ANALELE ȘTIINȚIFICE ALE UNIVERSITĂȚII „ALEXANDRU IOAN CUZA” DIN IAȘI (SERIE NOUĂ)

S E C Ț I U N E A I BIOLOGIE ANIMALĂ

TOMUL LX 2014

Editura Universității „Alexandru Ioan Cuza” din Iași

EDITORIAL BOARD

Editor-in-Chief: Prof. Dr. Gheorghe Mustață, Alexandru Ioan Cuza University of Iași,

Executive Editor: Reader Dr. Luminița Bejenaru, Alexandru Ioan Cuza University of Iași, Romania

Co-editors: Prof. Dr. Lotus Meșter, University of Bucharest, Romania Prof. Dr. Ion Moglan, Alexandru Ioan Cuza University of Iași, Romania Prof. Dr. Mircea Nicoară, Alexandru Ioan Cuza University of Iași, Romania Prof. Dr. Antonio Palanca-Soler, University of Vigo, Prof. Dr. Costică Misăilă, Alexandru Ioan Cuza University of Iași, Romania Assoc. Prof. Dr. Wietske Prummel, Royal University of Groningen, the Reader Dr. Ion Cojocaru, Alexandru Ioan Cuza University of Iași, Romania

Subject Editors: Reader Dr. Ștefan Zamfirescu, Alexandru Ioan Cuza University of Iași, Romania Lecturer Dr. Simina Stanc, Alexandru Ioan Cuza University of Iași, Romania Lecturer Dr. Mircea-Dan Mitroiu, Alexandru Ioan Cuza University of Iași, Romania

Scientific Board: Prof. Dr. Patrick Gillet, Western Catholic University of Angers, Prof. Dr. Vladimir Pešić, Universtity of Montenegro, Potgorica, Montenegro Prof. Dr. Ion Dediu, Institute of Ecology and Geography, Chișinau, Republic of Moldavia Prof. Dr. Iordache Ion, Alexandru Ioan Cuza University of Iași, Romania Prof. Dr. Mircea Varvara, Alexandru Ioan Cuza University of Iași, Romania Prof. Dr. Ion Andriescu, Alexandru Ioan Cuza University of Iași, Romania Prof. Dr. Ionel Miron, Alexandru Ioan Cuza University of Iași, Romania C.P.I. Dr. Dumitru Murariu, Grigore Antipa National Museum of Natural History, Bucharest, Romania Assoc. Prof. Dr. Christine Lefevre, National Museum of Natural History, Paris, France Reader Dr. Carmen Gache, Alexandru Ioan Cuza University of Iași, Romania Reader Dr. Ioan Coroiu, Babes-Bolyai University, Cluj-Napoca, Romania Lecturer Dr. Anca-Narcisa Neagu, Alexandru Ioan Cuza University of Iași, Romania Dr. Hab. Zbigniew Bocheński, Institute of Systematics and Evolution of , Polish Academy of Sciences, Krakow, Dr. Erika Gál, Archaeological Institute, Hungarian Academy of Sciences, Budapest,

Editorial address: Facultatea de Biologie Telephone: +40232201527 Universitatea „Alexandru Ioan Cuza” din Iași Fax: +40232201472 Bd. Carol I, Nr. 20A, 700505 Iași, România http://www.bio.uaic.ro/publicatii/anale_zoologie/anale_zoo_index.html

Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014

CONTENTS

RESEARCH ARTICLES ...... 5

Odette LOBIUC, Ioan MOGLAN & Andrei LOBIUC - DIVERSITY OF FAMILY () IN SOME PROTECTED AREAS FROM IASI COUNTY (ROMANIA) ...... 5

Emilian PRICOP - FIRST RECORD OF LITUS CAMPTOPTERUS NOVICKY, 1953 (HYMENOPTERA: MYMARIDAE) FROM PROVINCE (ROMANIA), WITH A KEY TO THE EUROPEAN SPECIES ...... 19

Oxana MUNJIU, Ion TODERAȘ, Elena ZUBCOV, Lucia BILETCHI & Igor SUBERNETKII - COMPOSITION AND DISTRIBUITION OF BENTHIC MACROINVERTEBRATES IN THE PRUTH RIVER (2012-2013) ...... 27

Marius Andrei RĂU, Gabriel PLĂVAN, Ștefan Adrian STRUNGARU & Mircea NICOARĂ - STUDY OF MACROINVERTEBRATE DIVERSITY AND SEASONAL DYNAMICS FROM BĂDĂRĂU LAKE – IAŞI, ROMANIA ...... 35

Elena-Daniela PRELIPCEAN (BOSOVICI), Mircea VARVARA & Ioan MOGLAN - EPIGENOUS FAUNA (ARTHROPODA: ARANEAE, INSECTA) IN THE RYE CROP FROM HORODNIC DE JOS (SUCEAVA COUNTY, ROMANIA) ...... 47

Mariana POPOVICI & Simina STANC - FREQUENCY OF SWINE (SUS DOMESTICUS AND SUS SCROFA) IN THE BRONZE AGE SETTLEMENTS ON THE TERRITORY OF ROMANIA ...... 53

Angela SIMALCSIK, Vasilica Monica GROZA & Robert Daniel SIMALCSIK - ANTHROPOLOGICAL ANALYSIS OF A SKELETON BELONGING TO MIDDLE BRONZE AGE, KOMARIV CULTURE, DISCOVERED IN SUCEAVA-CÂMPUL ŞANŢURILOR-STRADA PARCULUI (SUCEAVA COUNTY, ROMANIA) ...... 59

Robert-Daniel SIMALCSIK - DENTAL PATHOLOGIES IN POPULATIONS FROM A 4TH CENTURY A.D. NECROPOLIS AND A MEDIEVAL NECROPOLIS (14TH-17TH CENTURIES) FROM SĂBĂOANI, NEAMŢ DISTRICT ...... 77

Vasilica-Monica GROZA, Angela SIMALCSIK, Luminiţa BEJENARU & Robert-Daniel SIMALCSIK - OSTEOPATHIES IN THE POPULATION OF OLD IASI CITY (ROMANIA): THE NECROPOLIS OF THE “BANU” CHURCH, 16TH-19TH CENTURIES ...... 91

Vasilica-Monica GROZA, Angela SIMALCSIK & Luminiţa BEJENARU - ODONTOPATHIES IN THE POPULATION OF OLD IAȘI CITY (ROMANIA): THE NECROPOLIS OF THE “BANU” CHURCH, 16TH-19TH CENTURIES ...... 105

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SHORT COMMUNICATIONS ...... 115

Ionuț Ștefan IORGU - FIRST RECORD OF THE BUSH-CRICKET ISOPHYA CIUCASI (INSECTA: ORTHOPTERA: PHANEROPTERIDAE) IN MOLDAVIA (ROMANIA)...... 115

COMMENTARIES ...... 121

Ștefan-Adrian STRUNGARU, Gabriel PLĂVAN, Marius Andrei RĂU & Mircea NICOARĂ - A SHORT OVERVIEW FOCUSED ON INVERTEBRATE AND VERTEBRATE BIOINDICATORS FOR FRESHWATER ENVIRONMENTS CONTAMINATED WITH HEAVY METALS ...... 121

Monica LUCA & Gogu GHIORGHIȚĂ - THE INVASIVE SPECIES PERCCOTTUS GLENII – A THREAT FOR THE FRESH WATER ECOSYSTEMS ...... 129

Ion COJOCARU - THE THEORY OF SPONTANEOUS GENERATION, BETWEEN IDEOLOGY AND SCIENCE ...... 139

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DIVERSITY OF FAMILY LYCAENIDAE (LEPIDOPTERA) IN SOME PROTECTED AREAS FROM IASI COUNTY (ROMANIA)

Odette LOBIUC*, Ioan MOGLAN and Andrei LOBIUC Faculty of Biology, Alexandru Ioan Cuza University, Bd. Carol I 11, 700506 Iaşi, Romania. *[email protected]

Abstract. The Lycaenidae family is represented in Romania by 59 species, with the Polyommatinae subfamily comprising most of these species. Many taxa in Lycaenidae are vulnerable or critically endangered due to anthropic activities. To assess the diversity of species in this family, studies were conducted in five natural protected areas in Iași County: Fâneţele Seculare de la Valea lui David, Sărăturile de la Valea Ilenei, Poiana cu Schit, Pădurea Uricani and Pădurea Pietrosu. A total of 21 species were recorded in these areas. Of these species, one is endangered, three are near threatened and five are vulnerable. Our paper presents the list of recorded species, their frequency, synecological data and diversity indexes for the lycaenids communities in investigated areas, meanwhile offering new faunal data in these areas.

Keywords: Lycaenidae, biodiversity, protected areas

Rezumat. Diversitatea familiei Lycaenidae (Lepidoptera) din unele arii protejate din judeţul Iaşi (România). Familia Lycaenidae este reprezentată în România de 59 de specii, cu subfamilia Polyommatinae cuprinzând cele mai multe din specii. Numeroşi taxoni ai familiei Lycaenidae sunt vulnerabili sau critic periclitaţi datorită activităţilor antropice. Pentru a evalua diversitatea speciilor din această familie, s-au realizat cercetări în cinci arii protejate din judeţul Iaşi: Fâneţele Seculare de la Valea lui David, Sărăturile de la Valea Ilenei, Poiana cu Schit, Pădurea Uricani şi Pădurea Pietrosu. În total, s-au identificat 21 de specii în aceste zone. Dintre aceste specii, una este periclitată, trei sunt potenţial ameninţate şi cinci sunt vulnerabile. Studiul nostru prezintă lista speciilor înregistrate, frecvenţa lor, date sinecologice şi indici de diversitate ai comunităţii de licenide în zonele cercetate, în acelaşi timp oferind noi date faunistice în ariile respective.

Cuvinte-cheie: Lycaenidae, biodiversitate, arii protejate

Introduction The Lycaenidae family is one of the largest groups in Lepidoptera, with approximate 6000 species. Its representatives are found in all major biogeographical regions in the temperate and tropical areas. Although many species present strict habitat requirements, lycaenids can be met in various biomes and vegetal associations, from tropical forests to bush-type vegetation, meadows, wet areas or semidesertified areas (New, 1993). From the diversity point of view, the lycaenids are more frequent in the tropical region, especially in S America and S-E Asia, followed by Africa. In the neotropical ecozone, the Theclinae subfamily is the most abundent, followed by the Polyommatinae and Lycaeninae subfamilies. In the nearctic zone, the number of species is relatively small, representing half the number of species in palearctic region (New, 1993). In , the lycaenids are distributed rather heterogenous, with less than 10 species in some countries and up to more than 65 species in others ( 68, Hungary 55, Yugoslavia 62, 62, 52, Poland 53, France 68, 37, Great Britain 22, Ireland 8) (according to Fauna Europea). In Romania, 59 taxa represent the

- 5 - Odette Lobiuc et al. family, with the Polyommatinae subfamily comprising most of these species (Rákosy et al., 2003). Regarding the host plants of the larvae, the range is relatively large, including species from the Rosaceae, , Polygonaceae, Fagaceae, Ericaceae, Cornaceae, Geraniaceae etc. families. Larvae of certain species are euryphagous (Satyrium acaciae, S. w-album, Callophrys rubi, Lycaena phlaeas, minimus, C. argiades, C. osiris, Celastrina argiolus, Glaucopsyche alexis, Plebejus argus, P. argyrognomon, Aricia agestis, Polyommatus bellargus, P. semiargus, P. icarus, P. daphnis), others are stenophagous (Lycaena dispar, L. thersamon, C. decoloratus, Cupido alcetas, Polyommatus thersites) (Higgins & Riley, 1988; Ciochia & Stancă, 2000). Due to particular requirements of many species in the life cycle (myrmecophily in the larval stage) and to activities such as the change of agricultural practices, tourism, pollution, pesticide use, deforestation, urbanisation, climatic changes etc., numerous taxa in this group have become vulnerable or threatened (Munguira, 1993; Van Swaay et al., 2010). The diversity and ecology of many species was thus investigated in European countries, identifying potential threats of these species (Dennis, 2004; Bourn & Thomas, 1993; Brereton et al., 2008). In Romania, studies regarding the diversity of lepidopterans, including lycaenids, were conducted in Dobrogea region (Székely, 1994; Dincă et al., 2009; Dincă & Cuvelier, 2011), Transylvania (Rákosy, 1999; Cremene et al., 2002; Székely, 2004; Goia & Dincă, 2006; Rákosy, 2010), Moldavia (Corduneanu, 2011), Muntenia (Dincă, 2005). Potential threats as well as legislation issues are discussed for lepidopterans taxa in Romania by Rákosy (2005) and by Schmitt and Rákosy (2007). The current study investigates the richness of Lycaenidae taxa in some natural protected areas in Iași County, for which the data regarding the biodiversity of this group are relatively few. Corduneanu et al. (2011), however, identifies 17 Lycaenidae taxa in Valea lui David area. Protected natural areas were chosen due to a reduction in anthropic activities level and, possibly, a higher number of species present.

Material and Methods To assess the diversity of Lycaenidae taxa in natural protected areas in Iași County, five such areas were chosen: Fâneţele Seculare de la Valea lui David, Sărăturile de la Valea Ilenei, Poiana cu Schit, Pădurea Uricani and Pădurea Pietrosu. The protected areas under study are classified in I.U.C.N. IV category, some characteristics and vulnerabilities being given in Table 1, according to Nicoară & Bomher (2004). The chosen areas were selected for the different habitat types present and due to the lack of data regarding the diversity of lepidopterans for some of them. The material was collected between April and September 2012. were captured using a entomological net. Identification of taxa was done on the basis of external morphology and of male genitalia morphology. In forest-type habitats, specimens were observed and captured at woodside of the area under study. To evaluate the frequency of species, a number of 1-2 specimens observed per day was considered to designate a very rare (VR) species, 2-5 specimens per day was assigned to rare (R) species, 2-5 individuals per hour indicate a relatively frequent (RF) species and 6-15 individuals per hour signify a frequent (F) species.

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The ecological parameters for evaluated species were adapted from Dincă (2005). The categories of threat were taken from the Red List of lepidopterans of Rákosy et al. (2003). The characterization of investigated areas using the lycaenids populations in these reservations was realised through calculation of some ecological indexes that indicate the number of species and the structure of the lycaenids communities. The calculated parameters are: Abundance (A): the absolute number of individuals of one species that are present in an area (Simionescu, 1983). Constancy (C): the continuity of existence of one species in a given biotope (Simionescu, 1983).

Table 1. Protected areas under study.

Protected Area Geographical Altitude Habitat type Vulnerabilities areas (ha) coordinates - meadows and pannonic and - mowing and ponto-sarmatic salt marshes partial grazing Fâneţele - deciduous bushes - chaotic tourism Seculare de 47°12' N, 46.36 60.23-170.37 - ponto-sarmatic steppes - erosion la Valea lui 27°28' E phenomena in David percentages up to 70% - herbs and shrubs - grazing Poiana cu 46°59' N, 9.5 333.21-374.83 - hunting Schit 27°35' E

- Salicornia communities and - mowing and other annual species which partial grazing Sărăturile de colonise wet and sandy terrains 47°13' N, la Valea 5.9 55.31-71.19 - meadows and pannonic and 27°22' E Ilenei ponto-sarmatic salt marshes - woodside communities with tall higrophilous herbs - eurosiberian silvoseppe - forestry Pădurea 47°08' N, 68 76.46-163 vegetation with Quercus ssp. management Uricani 27°29' E - chaotic tourism - beech (Fagus taurica şi Fagus - grazing Pădurea 47°00' N, orientalis), hornbeam and silver - hunting 83 226.07-332.7 Pietrosu 27°40' E linden (Tilia tomentosa) forests - chemical 100-150 years of age treatments

CA = (npA/Np)*100 CA = the constancy of species A; npA = the number of samples in which the species A is present; Np = the total number of investigated samples. The value of this index classifies the species in one of the following classes: C1 - accidental, present in 1-25% of samples; C2 – accesory, present in 25.1-50% of samples; C3 - constant, present in 50.1-75% of samples; C4 – euconstant, present in 75.1-100% of samples. Dominance (D) expresses the relative abundance, indicating the percentage of participation of each species in the relationship (Simionescu, 1983): DA = (nA/N)*100

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DA = dominance of species A; nA = total number of individuals of species A, in investigated samples; N = total number of individuals of all species present in investigated samples. The percentage of individual dominance classifies the species in one of the following categories: D1 – subrecedent, below 1.1%; D2 – recedent, between 1.1-2%; D3 – subdominant, between 2.1-5%; D4 – dominant, between 5.1-10%; D5 – eudominant, above 10%. The ecological significance index (W) presents the relationship between the structural index (C) and the productivity index (D), indicating the position of a species in the biocenosis (Simionescu, 1983). WA = (CA*DA*100)/10000 The calculated value classifies the species as following: W1 – values below 0.1%; W2 – values between 0.1-1%; W3 – values between 1.1-5%; W4 – values between 5.1-10%; W5 – values above 10%. The W1 class corresponds to accidental species; classes W2 and W3 – to accessory species; classes W4 and W5 – to characteristic species. The Shannon-Wiener (H) diversity index varies directly with the number of species (Stan, 1995). The index value estimates the real value of diversity of the biocenosis (Botnariuc & Vădineanu, 1982). H = - ∑(i=1,S) Pi ln Pi S = the number of species; Pi = the proportion of each species. The degree of equitability of species on the basis of Shannon-Wiener index was calculated using the formula: EH = H/Hmax, Hmax = lnS (Magurran, 2004) Hmax = maximal diversity The Simpson (1/D) index is a diversity index illustrating the probability that two individuals in a sample to belong to the same species (Botnariuc & Vădineanu, 1982). 2 1/D = 1/ ∑(i=1,S) (Pi) Pi = the proportion of individuals of i species; S = the number of species. The degree of equitability of species on the basis of Simpson index was calculated using the formula: ED = (1/D)/S (Magurran, 2004)

Results and Discussion In the investigated areas, a total of 21 taxa were identified, belonging to 9 genera (Table 2). Out of these, three are a part of the Theclinae subfamily, three belong to Lycaeninae and fifteen to Polyommatinae. Most taxa were recorded at Fâneţele Seculare (15 species) and Poiana cu Schit (13 species), while the least taxa were identified at Pădurea Pietrosu (6 species). Within Sărăturile de la Valea Ilenei and Pădurea Uricani protected areas a number of 8 and respectively 10 species were identified. Among recorded taxa, three species are near threatened (Cupido minimus, Polyommatus daphnis and Polyommatus bellargus), five species are vulnerable (Satyrium acaciae, Satyrium w-album, Lycaena thersamon, Cupido decoloratus, Cupido osiris) and one species is endangered (Cupido alcetas) (Rákosy et al., 2003).

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Table 2. Systematic list of the Lycaenidae species identified in protected areas. Abbreviations: head of the table: L. t. s. = larval trophic source; Z. el. = zoogeographical element; Ec. ch. = ecological character; R. L. = red list. L. t. s.: 1 = consumers of herbaceous plants, excluding Poaceae; 2 = consumers of bushes and Ericaceae; 3 = consumers of decidous trees; 4 = myrmecophilous. Z. el.: Eua = Eurasiatic; Vam = West-Asiatic Mediterranean; E = European; Hol = Holarctic. Ec. ch.: M = mezophilous; Mxt = mezoxerothermophilous; Xt = xerothermophilous; Mt = mezothermophilous; Mh = mezohygrophilous; Hg = hygrophilous; Mg = migratory. R. L.: EN = endangered, VU = vulnerable, NT = near threatened, DD = data deficient.

No. of Frequency Ec. Species captured Data labels of observed L. t. s. Z. el. R. L. ch. individuals individuals Theclinae Pădurea Uricani: Satyrium acaciae 27.VI.2012; 7 RF 2; 3 Eua Mt VU (Fabricius, 1787) 23.VI.2012; 1.VII.2012 Poiana cu Schit: 4 F 8.VII.2012 Satyrium w-album Pădurea Uricani: Mt 1 R 3 Eua VU (Knoch, 1782) 27.VI.2012 Pădurea Pietrosu: 3 RF 3.VII.2012 Pădurea Uricani: Callophrys rubi 4 14.V. 2012; VR 1; 2 Eua Mt (Linnaeus, 1758) 27.VI.2012 Lycaeninae Valea lui David: 1 R 20.VI.2012 Poiana cu Schit: 1 VR Lycaena phlaeas 3.VII.2012 1 Hol Mxt (Linnaeus, 1761) Pădurea Uricani: 1 VR 27.VI.2012 Pădurea Pietrosu: 2 R 3.VII.2012 Valea lui David: 22.VI.2012; 9 RF 10.VII.2012; 15.VII.2012 Lycaena dispar Poiana cu Schit: 1 Vam Hg (Haworth, 1802) 5 RF 13.VI.2012 Pădurea Pietrosu: 5 20.VI.2012; RF 8.VIII.2012 Valea lui David: 24.IV.2012, 3 VR 20.VI.2012; 1.VIII.2012 Lycaena thersamon Sărăturile de la Valea 1 Vam Mt VU (Esper, 1784) 5 Ilenei: 27.IV.2012, RF 10.VII.2012 Pădurea Uricani: 2 R 27.VII. 2012

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No. of Frequency Ec. Species captured Data labels of observed L. t. s. Z. el. R. L. ch. individuals individuals Polyommatinae Valea lui David: 23.V.2012; 10 22.VI.2012; RF 10.VII.2012; 3.IX.2012 Sărăturile de la Valea Cupido argiades 3 F Ilenei: 10.VIII.2012 1 Eua M (Pallas, 1771) Poiana cu Schit: 3 F 16.VI.2012 Pădurea Uricani: 2 RF 4.VIII.2012 Pădurea Pietrosu: 3 F 18.VI.2012 Valea lui David: 3 23.V.2012, VR 10.VII.2012 Sărăturile de la Valea 1 RF Ilenei: 1.VIII. 2012 Poiana cu Schit: Cupido decoloratus Xt, 6 8.VII.2012; F 1 Vam VU (Staudinger, 1886) Mxt 15.VII.2012 Pădurea Uricani: 5 14.V.2012; F 27.VI.2012 Pădurea Pietrosu: 3 F 3.VII.2012 Sărăturile de la Valea 1 VR Cupido alcetas Ilenei: 27.VI.2012 1 Eua Mh EN (Hoffmannsegg, 1804) Pădurea Pietrosu: 2 VR 3.VII.2012 Pădurea Uricani: Cupido minimus 2 23.VI.2012; VR 1 Eua Xt NT (Fuessly, 1775) 4.VIII.2012 Cupido osiris Valea lui David: 2 VR 1 Eua Mt VU (Meigen, 1829) 10.VII.2012 Valea lui David: 24.IV.2012; 5 R 22.VI.2012; Celastrina argiolus 10.VII.2012 1; 2; Hol M (Linnaeus, 1758) Poiana cu Schit: 3; 4 2 R 15.VII.2012 Pădurea Uricani: 6 RF 23.VI.2012 Valea lui David: 2 R Glaucopsyche alexis 20.VI.2012 1; 4 Eua Mh (Poda, 1761) Sărăturile de la Valea 2 R Ilenei: 27.VI. 2012 Valea lui David: Plebejus argus 6 10.VII.2012; F 1; 4 Eua M (Linnaeus, 1758) 1.VIII.2012

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No. of Frequency Ec. Species captured Data labels of observed L. t. s. Z. el. R. L. ch. individuals individuals Sărăturile de la Valea Ilenei: 27.VI.2012; 12 F 10.VII.2012; 1.VIII.2012 Poiana cu Schit: 5 16.VI.2012; F 15.VIII.2012 Pădurea Uricani: 1 R 27.VI.2012 Valea lui David: 5 20.VI.2012, RF 1.VIII.2012 Plebejus Sărăturile de la Valea argyrognomon 1; 4 E Mt 2 Ilenei: 27.VI.2012; R (Bergsträsser, 1779) 10.VII.2012 Poiana cu Schit: 2 R 15.VII.2012 Valea lui David: 1 10.VII.2012; VR Aricia agestis Denis & 1.VIII.2012 1; 4 Eua Mxt (Schiffermüller, 1775) Poiana cu Schit: 2 3.VII.2012; VR 15.VII.2012 Valea lui David: Polyommatus 4 23.V.2012; R semiargus 22.VI.2012 1; 4 Eua M, Mg (Rottemburg, 1775) Poiana cu Schit: 1 VR 3.VII.2012 Valea lui David: 23.V.2012; 6 RF 20.VI.2012; 1.VIII.2012 Polyommatus icarus Sărăturile de la Valea 1; 4 Eua M (Rottemburg, 1775) 5 Ilenei: 10.VIII.2012; RF 3.IX.2012 Poiana cu Schit: 7 13.VI.2012; RF 15.VIII.2012 Valea lui David: Polyommatus thersites 6 10.VII.2012; RF 1; 4 Eua Mxt DD (Cantener, 1835) 3.IX.2012 Polyommatus daphnis Poiana cu Schit: 3. (Denis & 9 VII.2012; 8.VII.2012; RF 1; 4 Vam Mxt NT Schiffermüller, 1775) 15.VII.2012 Valea lui David: Polyommatus 4 20.VI.2012; R bellargus 3.IX.2012 1; 4 Eua Mt NT (Rottemburg, 1775) Poiana cu Schit: 1 VR 16.VI.20112. (1 ♂).

The number of taxa varies with the type of habitat, the most species being present in meadows and the least species present in forest-type habitats. In Europe, such distribution is mentioned in Munguira et al. (1993), especially due to the fact that forests

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(woodlands) are populated mostly by Theclinae species. These are met mainly at borders of clearings, situation encountered in our study. Preferences towards meadows are also recorded in mediterranean regions by Ştefănescu et al. (2005). Dry grasslands and steppes are inhabited by a larger number of species, among which Lycaena sp., Polyommatus sp., Plebejus sp., while shrublands are the habitat for Plebejus argus (Munguira et al., 1993). Similar accounts of taxa in different habitat types are enumerated by Rákosy (2010) for Romania. In steppe regions around Cluj (Transylvania), Cremene et al. (2002) identifies nine Lycaenidae taxa, among which Satyrium acaciae, Cupido osiris, Glaucopsyche alexis, Polyommatus bellargus, P. icarus and Plebejus argus the latter species being the most abundent. Dincă (2005) identifies 24 species of Lycaenidae in the steppe and forest habitats in Muntenia (Istriţa Hill), including Lycaena phlaeas, L. dispar, L. thersamon, Callophris rubi, Cupido argiades, C. alcetas, Celastrina argiolus, Glaucopsyche alexis, Plebejus argus, P. argyrognomon, Aricia agestis, Polyommatus thersites, P. icarus, P. daphnis and P. bellargus. Goia & Dincă (2006), in a study around the city of Cluj-Napoca, observes 45 Lycaenidae species in different habitats, with larger numbers in areas with lower degrees of forestation and in protected areas. The highest abundance of lepidopterans species in areas of Braşov County was recorded in meadows and hay meadows by Rákosy (2009). Of identified taxa, 28 lycaenid species are present, including protected ones. Considering the feeding preferences in the larval stage described in the literature, most (seventeen) species identified are consuming herbaceous plants (80.9%), three species have mixed preferences (14.3%), consuming herbaceous plants, shrubs and deciduous tree leaves, while one species prefers tree leaves only (4.8%). Proportions of preferences similar to those we found are reported in other steppe and forest habitats as well (Dincă, 2005). Regarding the zoogeographical element, the vast majority of species are Eurasiatic (66.7%), 19% are West-Asiatic Mediterranean, 9.5% are Holarctic and 4.8% are European. The ecological characters most present within observed species are mezothermophilous (7 species), mezoxerothermophilous (4 species) and mezophilous (5 species). The xerothermophilous character is found in two species while two species is mezohygrophilous and one species is hygrophilous. Among observed species, one is migratory (Polyommatus semiargus). Out of identified species, three are included in different lists for protection and conservation of fauna. Lycaena dispar is included in Annex 2 and Annex 4 of EU Habitats Directive as well as in Annex 3 and 4A of OUG 57/2007. Cupido alcetas and Cupido osiris are listed in Annex 4B of OUG 57/2007 (Goia & Dincă, 2006; Corduneanu, 2011). The presence of these species in the investigated areas emphasizes the need to protect and conserve these habitats. Conservation of such areas is further required by the fact that similar habitats are often exposed to threats such as urbanisation or intensification of agriculture or deforestations (Cremene et al., 2002; Dincă, 2005; Goia & Dincă, 2006). Furthermore, the current data and legislation on lepidopteran diversity appears insuficient in Romania (Rákosy, 2005). The frequency of observed species is variable, some taxa being however very rare or rare. It is the case of Callophrys rubi, Lycaena phlaeas, Cupido alcetas, C. minimus, C. osiris, Glaucopsyche alexis, Aricia agestis, Polyommatus semiargus, P. bellargus. The other taxa are rare to relatively frequent, depending on the area. The current study investigates lycaenid richness in five protected areas, for which only at Fâneţele Seculare, Corduneanu et al. (2011) evaluates the lepidopteran fauna. Our

- 12 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 study confirms most taxa identified in the reported paper, with the exception of Satyrium acaciae and S. spini. Further studies, however, are required to confirm the data we recorded in the investigated areas under climatic and floristic conditions of the subsequent flight seasons. By analysing the components and distribution of the communities of lycaenids in some protected areas, a set of characteristics can be observed for these areas.

Table 3. Sinecological analysis of lycaenids communities in Fâneţele Seculare de la Valea lui David area.

D C W No. Species A % Class % Class % Class

1 Plebejus argus 47 24.3 D5 100 C4 24.3 W5

2 Plebejus argyrognomon 29 15.0 D5 100 C4 15.0 W5

3 Polyommatus icarus 24 12.4 D5 100 C4 12.4 W5

4 Lycaena dispar 21 10.8 D5 100 C4 10.8 W5

5 Polyommatus thersites 21 10.8 D5 100 C4 10.8 W5

6 Cupido argiades 20 10.3 D5 100 C4 10.3 W5

7 Lycaena phlaeas 5 2.5 D3 100 C4 2.5 W3

8 Glaucopsyche alexis 5 2.5 D3 80 C4 2.07 W3

9 Aricia agestis 5 2.5 D3 80 C4 2.0 W3

10 Celastrina argiolus 4 2.0 D2 80 C4 1.6 W3

11 Polyommatus semiargus 3 1.5 D2 60 C3 0.9 W2

12 Polyommatus bellargus 3 1.5 D2 60 C3 0.9 W2

13 Lycaena thersamon 2 1.0 D1 60 C3 0.6 W2

14 Cupido decoloratus 2 1.0 D1 60 C3 0.6 W2

15 Cupido osiris 2 1.0 D1 60 C3 0.6 W2

In Fâneţele Seculare de la Valea lui David protected area, 15 lycaenids species were identified (Table 3). Of these, 6 present a higher abundance, being classified as eudominant (Plebejus argus, P. argyrognomon, Polyommatus icarus, Lycaena dispar, Polyommatus thersites, Cupido argiades). The other species are less abundant, with 3 subdominant species, 3 recedent species and 3 subrecedent species. Analysing the ecological significance index (W), 6 species can be considered characteristic, while the other species are accessory. In Poiana cu Schit protected area, 13 lycaenid species were identified, 7 of which having a higher abundance (Table 4). Of these, 6 species are eudominant and one species (Polyommatus daphnis) is dominant. Among species with a lower abundance, one species is subdominant (Plebejus argyrognomon), 2 are recedent and 3 are subrecedent. According to the ecological significance index (W), 7 species are characteristic, the rest are accessory. In Sărăturile de la Valea Ilenei protected area, of 8 lycaenid species identified, 5 are eudominant, characteristic for the area, also having a higher abundance (Table 5). The other 3 species are accessory. Of them, one is subdominant (Plebejus argyrognomon), and the other two are recedent (Glaucopsyche alexis, Cupido alcetas).

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Table 4. Sinecological analysis of lycaenids communities in Poiana cu Schit area.

D C W No. Species A % Class % Class % Class 1 Plebejus argus 41 18.3 D5 100 C4 18.3 W5

2 Cupido decoloratus 32 14.3 D5 100 C4 14.3 W5

3 Satyrium w-album 30 13.4 D5 100 C4 13.4 W5

4 Cupido argiades 30 13.4 D5 100 C4 13.4 W5

5 Lycaena dispar 27 12.1 D5 100 C4 12.1 W5

6 Polyommatus icarus 25 11.2 D5 80 C4 8.9 W4

7 Polyommatus daphnis 22 9.8 D4 100 C4 9.8 W4

8 Plebejus argyrognomon 5 2.2 D3 80 C4 1.7 W3

9 Aricia agestis 4 1.7 D2 60 C3 1.0 W2

10 Celastrina argiolus 3 1.3 D2 80 C4 1.0 W2

11 Lycaena phlaeas 2 0.8 D1 60 C3 0.5 W2

12 Polyommatus semiargus 1 0.4 D1 40 C3 0.1 W2

13 Polyommatus bellargus 1 0.4 D1 40 C2 0.1 W2

Table 5. Sinecological analysis of lycaenids communities in Sărăturile de la Valea Ilenei area.

D C W No. Species A % Class % Class % Class 1 Plebejus argus 43 26.5 D5 100 C4 26.5 W5

2 Cupido argiades 34 20.9 D5 100 C4 20.9 W5

3 Polyommatus icarus 28 17.2 D5 100 C4 17.2 W5

4 Cupido decoloratus 27 16.6 D5 100 C4 16.6 W5

5 Lycaena thersamon 21 12.9 D5 100 C4 12.9 W5

6 Plebejus argyrognomon 4 2.4 D3 80 C4 1.9 W3

7 Glaucopsyche alexis 3 1.8 D2 60 C3 1.1 W3

8 Cupido alcetas 2 1.2 D2 60 C3 0.7 W2

In Pădurea Uricani protected area, 10 species of lycaenids were identified (Table 6): 4 eudominant (Cupido decoloratus, Celastrina argiolus, Cupido argiades, Satyrium acaciae), 3 dominant, 2 recedent and one subrecedent species (Lycaena phlaeas). Of these, the eudominant species present the highest abundance in the area, also being characteristic species. The other species have a lower abundance, being considered accessory species according to the ecological significance index (W). In Pădurea Pietrosu protected area, 6 species of lycaenids were identified (Table 7), of which 4 are eudominant (Cupido decoloratus, Cupido argiades, Satyrium w-album, Lycaena dispar), one is subdominant (Lycaena phlaeas) and one is recedent (Cupido alcetas). The ecological significance factor (W) indicates the presence of 4 characteristic species and of 2 accessory species.

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Table 6. Sinecological analysis of lycaenids communities in Pădurea Uricani area.

D C W No. Species A % Class % Class % Class 1 Cupido decoloratus 34 26.5 D5 100 C4 26.5 W5 2 Celastrina argiolus 30 23.4 D5 100 C4 23.4 W5 3 Cupido argiades 26 20.3 D5 100 C4 20.3 W5 4 Satyrium acaciae 21 16.4 D5 80 C4 13.1 W5 5 Plebejus argus 5 3.9 D3 60 C3 2.3 W3 6 Satyrium w-album 4 3.1 D3 60 C3 1.8 W3 7 Lycaena thersamon 3 2.3 D3 60 C3 1.4 W3 8 Callophrys rubi 2 1.5 D2 60 C3 0.9 W2 9 Cupido minimus 2 1.5 D2 60 C3 0.9 W2 10 Lycaena phlaeas 1 0.7 D1 40 C2 0.3 W2

Table 7. Sinecological analysis of lycaenids communities in Pădurea Pietrosu area.

D C W No. Species A % Class % Class % Class 1 Cupido decoloratus 40 31.2 D5 100 C4 31.2 W5 2 Cupido argiades 33 25.7 D5 100 C4 25.7 W5 3 Satyrium w-album 29 22.6 D5 100 C4 22.6 W5 4 Lycaena dispar 21 16.4 D5 100 C4 16.4 W5 5 Lycaena phlaeas 3 2.3 D3 80 C4 1.8 W3 6 Cupido alcetas 2 1.5 D2 40 C2 0.6 W2

The Shannon-Wiener indices of investigated areas show that the highest diversity is present in Fâneţele Seculare de la Valea lui David and Poiana cu Schit areas, which are floristic reservation. The lowest diversity is present in Pădurea Pietrosu forest type reservation, the other two areas having similar intermediate values. Of these two, Sărăturile de la Valea Ilenei is a floristic reservation, while Pădurea Uricani is a forest area. The species equitability is lowest in Pădurea Pietrosu and highest in Fâneţele Seculare de la Valea lui David. The Simpson indices of the investigated areas is highest in Poiana cu Schit and Fâneţele Seculare de la Valea lui David floristic areas indicating the highest diversity. The lowest values are present in the Pădurea Pietrosu forest reservation. The values of this index present a different pattern compared to values of the Shannon-Wiener index. The Poiana cu Schit reservation in this case, presents a slightly higher diversity compared to Fâneţele Seculare de la Valea lui David and the Sărăturile de la Valea Ilenei area also has a higher diversity value compared to Pădurea Uricani area. The equitability of the species shows the highest values for Pădurea Uricani area and the lowest values in Sărăturile de la Valea Ilenei and Pădurea Pietrosu areas.

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2.50 0.450 0.426 0.400 0.400 0.372 2.00 0.347 0.350 0.309 0.300 1.50 0.250 H index 2.24 2.17 0.200 Equitability 1.00 1.77 1.80 0.150 1.50 0.50 0.100 0.050 0.00 0.000 Fâneţele Seculare Poiana cu Schit Sărăturile de la Pădurea Uricani Pădurea Pietrosu de la Valea lui David Valea Ilenei

Figure 1. Shannon-Wiener index and equitability of investigated areas lycaenids communities.

9.00 0.045 8.00 0.040 0.038 0.040 7.00 0.035 0.035 0.032 0.032 6.00 0.030 5.00 0.025 1/D index 4.00 0.020 Equitability 7.41 7.77 3.00 0.015 5.26 5.07 2.00 4.11 0.010 1.00 0.005 0.00 0.000 Fâneţele Seculare Poiana cu Schit Sărăturile de la Pădurea Uricani Pădurea Pietrosu de la Valea lui David Valea Ilenei

Figure 2. Simpson index and equitability of investigated areas lycaenids communities.

Conclusions The current study identifies 21 taxa of Lycaenidae family from 5 natural protected areas in Iași County. The number of taxa is variable according to the type of habitat. The distribution of recorded taxa in investigated areas is similar to those found in other regions of the country with the same types of habitat. Out of identified species, three are included in faunal protection and conservation lists, augmenting the protected regime of evaluated areas. In the same time, we present new data for four protected areas in Iași County regarding the biodiversity of Lycaenidae family. In the investigated areas, the highest diversity is present in two floristic areas (Fâneţele Seculare de la Valea lui David and Poiana cu Schit) as visible from the values of diversity indices. Compared to other floristic reservations, a high diversity can also be seen in Pădurea Uricani reservation.

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References Botnariuc, N., Vădineanu, A., 1982. Ecologie. Editura Didactică şi Pedagogică, Bucureşti. Bourn, N.A.D., Thomas, J.A., 1993. The ecology and conservation of the brown argus Aricia agestis in Britain. Biological Conservation, 63(1): 67-74. Brereton, T.M., Warren, M.S., Roy, D.B., Stewart, K., 2008. The changing status of the Chalkhill Blue butterfly Polyommatus coridon in the UK: the impacts of conservation policies and environmental factors. Journal of Conservation, 12(6): 629-638. Ciochia, V., Stancă, C., 2000. Catalogul colecţiei didactice de lepidoptere cu elemente de biologie a Universităţii „Lucian Blaga”- Sibiu. Ed. „Pentru Viaţă”, Braşov. Corduneanu, C., Balan, C., Surugiu, I., 2011. Diurnal observations over the lepidopteran fauna (Insecta: Lepidoptera) from “Valea lui David” Nature Reserve, Iasi County. Mnemosyne, 2: 39-49. Corduneanu, C., 2011. Protected Lepidopteran species (Insecta: Lepidoptera) in north-east of Romania. Travaux du Muséum National d’Histoire Naturelle «Grigore Antipa», LIV(1): 115-123. Cremene, C., Rákosy, L., Erhardt, A., 2002. Diversity of Macrolepidoptera in steppe habitats of Căianu Mic (Cluj). Entomologica Romanica, 7: 5-14. Dennis, R.L.H., 2004. Just how important are structural elements as habitat components? Indications from a declining lycaenid butterfly with priority conservation status. Journal of Insect Conservation, 8(1): 37- 45. Dincă, V., 2005. The Macrolepidoptera (Insecta: Lepidoptera) from Istriţa Hill (Buzău Country, Romania). Entomologica Romanica, 10: 5-24. Dincă, V., Cuvelier, S., Székely, L., Vila, R., 2009. New data on the Rhopalocera (Lepidoptera) of Dobrogea (south-eastern Romania). Phegea, 37(1): 1-21. Dincă, V., Cuvelier, S., 2011. Distribution and conservation status of Pseudophilotes bavius (Lepidoptera: Lycaenidae) in Dobrogea (south-eastern Romania). Phegea, 39(2): 59-67. Goia, M., Dincă, V., 2006. Structura şi răspândirea faunei de lepidoptere diurne (Hesperioidea & Papilionoidea) în împrejurimile municipiului Cluj-Napoca şi aspecte actuale ale influenţei antropozoogene asupra mediului de viaţă al acestora. Buletinul de informare al Societăţii Lepidopterologice Române, 17: 139- 197. Higgins, L.G., Riley, N.D.,1988. Guide des papillons d'Europe. Delachaux & Niestlé, Neuchâtel-Paris. Magurran, A.E., 2004. Measuring biological diversity. Blackwell Publishing. Munguira, M.L., Martin, J., Balletto, E., 1993. Conservation biology of Lycaenidae: a European overview. In New, T.R. (ed.). Conservation Biology of Lycaenidae (Butterflies). International Union for Conservation of Nature and Natural Resources, Gland, , 23-34. New, T.R., 1993. Introduction to the biology and conservation of the Lycaenidae. In New, T.R. (ed.), Conservation Biology of Lycaenidae (Butterflies). International Union for Conservation of Nature and Natural Resources, Gland, Switzerland, 1-20. Nicoară, M., Bomher, E., 2004. Ghidul ariilor protejate din Judeţul Iaşi. Editura Tipo Moldova, Iaşi. Rákosy, L., 1999. Lepidopterologische Biodiversität eines kleinräumigen steppenartigen Naturschutzgeibietes in Siebenbürgen (Suatu, Transsylvanien, Rumänien). Entomologica Romanica, 4: 49-68. Rákosy, L., Goia, M., Kovács, Z., 2003. Catalogul Lepidopterelor României / Verzeichnis der Schmetterlinge Rumäniens. Societatea Lepidopterologică Română, Cluj-Napoca. Rákosy, L., 2005. U.E. şi legislaţia pentru protecţia lepidopterelor din România. Buletinul de informare Entomologică, 16: 89-96. Rákosy, L., 2009. Macrolepidoptere din perimetrul comunei Şinca Nouă (jud. Braşov). Buletinul de informare Entomologică, 20: 5-26. Rákosy, L., 2010. Micul ghid al fluturilor din Situl natura 2000 Sighişoara-Târnava Mare. Ed. Fundaţia ADEPT Transilvania, Saschiz, Mureş. Schmitt, T., Rákosy, L., 2007. Changes of traditional agrarian landscapes and their conservation implications: a case study of butterflies in Romania. Diversity and Distributions, 13: 855-862. Simionescu, V., 1983. Lucrări practice de ecologie. Editura Universității “Al. I. Cuza”, Iaşi. Stan, Gh., 1995. Metode statistice cu aplicaţii în cercetări entomologice (VII). Buletinul de Informare al Societății Lepidopterologice Române, 6(1-2):67-96. Székely, L., 1994. Des nouveautés concernant la faune de lépidoptères de sud-ouest de la Doubroudja. Sargetia, Series Scientia Naturae Deva, 14: 139-144. Székely, L., 2004. Noutăţi lepidopterologice din sud-estul Transilvaniei (Judeţul Braşov, România). Buletinul de informare Entomologică, 14-15: 41-56.

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Stefanescu, C., Peñuelas, J., Filella, I., 2005. Butterflies highlight the conservation value of hay meadows highly threatened by land-use changes in a protected Mediterranean area. Biological Conservation, 126: 234- 246. Van Swaay, C., Cuttelod, A., Collins, S., Maes, D., Munguira, M.L., Šašić, M., Settele, J., Verovnik, R., Verstrael, T., Warren, M., Wiemers, M., Wynhoff, I., 2010. European Red List of Butterflies. : Publications Office of the .

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FIRST RECORD OF LITUS CAMPTOPTERUS NOVICKY, 1953 (HYMENOPTERA: MYMARIDAE) FROM MOLDOVA PROVINCE (ROMANIA), WITH A KEY TO THE EUROPEAN SPECIES

Emilian PRICOP “Alexandru Ioan Cuza” University of Iaşi, B-dul Carol I, 20A, 700505, Iaşi, Romania, [email protected]

Abstract. The present paper represents a contribution to the knowledge regarding the current and distribution of the rare species Litus camptopterus Novicky, 1953 and also the distribution of the common species Litus cynipseus Haliday, 1833 from two poorly surveyed Romanian regions – Moldova and Dobrogea. Litus camptopterus is recorded for the first time from Moldova province (Romania). An updated identification key to the European species is also given. In our paper we provide clear illustrations that will help species identification. Regarding species habitat some notes are also given.

Keywords: Litus camptopterus Novicky, Mymaridae, fauna, taxonomy, first record, Moldova province (Romania).

Rezumat. Prima semnalare a specie Litus camptopterus Novicky, 1953 (Hymenoptera: Mymaridae) din provincia istorică Moldova (România), cu o cheie de identificare pentru speciile europene. Aceasta lucrare reprezintă o contribuție la cunoașterea taxonomiei și distribuției speciei Litus camptopterus Novicky, 1953, dar și a distribuției speciei Litus cynipseus Haliday, 1833 din două regiuni istorice ale României – Moldova și Dobrogea. Specia Litus camptopterus este semnalată pentru prima dată din provincia istorică Moldova (România). Este întocmită și cheia pentru speciile europene ale genului. În lucrarea noastră oferim ilustrații clare, care vor ajuta la o identificare facilă a acestor specii. În ceea ce privește habitatul speciilor, sunt oferite note.

Cuvinte cheie: Litus camptopterus Novicky, Mymaridae, fauna, taxonomie, prima semnalare, provincia Moldova (România).

Introduction The species that belong to Litus Haliday, a genus probably cosmopolitan, remain poorly known in Europe. Only two valid species are described from this region: Litus cynipseus Haliday and L. camptopterus Novicky. Triapitsyn & Berezovskiy (2004) published the most recent monograph of this genus. In Romania, Botoc (1964) described the species Litus distinctus, later synonymised under Litus camptopterus Novicky, 1953 (Triapitsyn & Berezovskiy, 2004). Botoc (1974) recorded also L. cynipseus from Transilvania and Oltenia regions. Unfortunately Botoc’s chalcidoid collection is lost. Females of Litus Haliday are common, males are very rare (Viggiani, 1973). Pricop (2009a, b; 2013), recorded L. cynipseus from Moldova and later from Dobrogea. For our study, almost all specimens which belonging to Litus Haliday were collected during 2006-2014 from different locations in Eastern Romania. All specimens are deposited in the insect collection of the Department of Biology, “Alexandru Ioan Cuza” University of Iași, Romania. We also search for new characters that will define the European species. For species distribution: all countries are arranged in alphabetical order (Noyes, 2003). An updated identification key to the European species of Litus Haliday is also presented.

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Material and Methods The species have been collected with an entomological sweep-net and with yellow pan-traps from forest and grass-land vegetation. The material was collected from some areas of Moldova and Dobrogea (Romania). The examined specimens were mounted in Faure’s medium or in Canada balm. We have illustrated the morphology of the specimens utilizing microphotographs obtained with a “Canon” digital camera attached to the IOR optical microscope. For the scanning electron microscopy we used Vega Tescan electron microscope. We have illustrated the most discussed characters in this paper. We have illustrated female antennae, wings and habitus of some specimens. For accurate examination and measurements the appendages from one side of the body of some specimens were removed and slide mounted. We search for new taxonomic characters, to establish the limits of the intra-specific variation. Abbreviations: elev. = elevation; F1-F6 = funicle segments (articles); Leg. = the collector; UAIC = “Alexandru Ioan Cuza” University of Iasi; NHMW = Natural History Museum of Vienna; NT = Neamț; CT = Constanța; IS = Iași; SV = Suceava.

Results and Discussion Genus Litus Haliday, 1833 [= Neolitus Ogloblin, 1935] Short diagnosis: Body stocky (Fig. 1) and dark colored; 5-segmented tarsi; head, mesosoma and coxae strongly sculptured – Figs. 1; 3g, i (the sculpture is reticulated); mandibles bidentate; female funicle 6-segmented (Figs. 1; 2c; 3d, e, f), all funicle articles without longitudinal sensory ridges; clava compact (Fig. 1), with 4 longitudinal sensory ridges; male antenna 11-segmented (Viggiani, 1973); mesoscutum with distinct notauli; mesophragma projecting into gaster; forewings long and narrow; metasoma almost sessile, ovipositor relatively short (Triapitsyn & Berezovskiy, 2004; Pricop, 2013). The identification key was adapted from Triapitsyn & Berezovskiy (2004).

Figure 1. Habitus of Litus cynipseus (original).

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Identification key to the species of Litus Haliday from Europe, females: 1 (2). Scape about as long as the combined length of F3 to F6 (Figs. 1; 3d, e, f); F2 usually shorter than the pedicel; clava broad, about 2.4 times as long as wide in lateral view (Fig. 3d, e, f) and about as long as the combined length of F3 to F6 (Figs. 1; 3d, e, f); forewing with two rows of microtrichia along the margins and many more irregularly arranged microtrichia in the apical half (Fig. 3b) ...... L. cynipseus

2 (1). Scape evidently shorter than the combined length of F3 to F6 (Fig. 2c, d); F2 about as long (or longer) as the pedicel; clava narrower, about 3 times as long as wide in lateral view (Fig. 2c, e) and not as long as the combined length of F3 to F6 (clava much shorter in comparison to F3-F6 length combined); forewing blade bare except for the two rows of microtrichia along margins (Fig. 2b, d) ...... L. camptopterus

1. Litus camptopterus Novicky, 1953 (Fig. 2a, b, c, d, e) = Litus distinctus Boţoc, 1964 Material examined: 1♀, 8.05.2010, Cozla hill from Piatra Neamţ, NT (Leg. Pricop E.) collected with the sweep net from herbaceous vegetation in a meadow at 500 m elev.; 1♀, 8.05.2010, Piatra Neamţ, from a private garden with vegetables, flowers and Vitis vinifera, NT (Leg. Pricop E.), collected with yellow pan traps; 1♀, 14.08.2010, Piatra Neamţ, private garden, NT (Leg. Pricop E.), collected with yellow pan traps a 350 m elev.; 1♀, 24.08.2014 from Salix sp. near Bicaz river alluvial deposits in Hamzoaia, Tasca, NT (Leg. Pricop E.), collected with the sweep net. Hosts: Unknown. Distribution: Holarctic Region – Austria, Canada, China, , Kyrgyzstan, Republic of Korea, Romania, , USA (Noyes, 2003; Triapitsyn & Berezovskiy, 2004). L. camptopterus is recorded for the first time from Moldova province (Romania). Taxonomic notes: L. camptopterus is a species with long funicle segments (except F1) and a slender clava, in comparison with L. cynipseus a species with short funicle segments and a broad clava. In L. camptopterus the forewing blade is bare except for the two rows of microtrichia along margins (Fig. 2b). The fore wings of L. camptopterus are narrower in comparison to the forewings of L. cynipseus. Ovipositor is short. In Litus distinctus, described by Botoc (1964), the funicle segments seem to be a little longer in comparison to the funicle segments from the specimens redescribed as L. camptopterus by Triapitsyn & Berezovskiy (2004) and partially from our specimens also. Regarding L. camptopterus, the lack of any illustrations in Novicky’s (1953) paper has made this species difficult to identify. L. camptopterus it is a rare species recorded from Neamt County, Bistrița River Basin – Moldova Province (Romania). L. camptopterus is present in natural and anthropic ecosystems.

2. Litus cynipseus (Haliday, 1833) (Figs. 1; 3a, b, c, d, e, f, g, h, i) = Litus krygeri Kieffer, 1913 Material examined: Beside Botoc (1974), Pricop (2009a, b) recorded L. cynipseus from Cozla hill, Piatra Neamț area – Moldova, species collected with the sweep net from a meadow near the forest and later, Pricop (2013) recorded this species also from the Marine Biological Station Agigea, CT, collected with yellow pan traps from the sand dune reservation area – Dobrogea. In general, the illustrations from Figs. 1; 2 and 3 belong to

- 21 - Emilian Pricop specimens collected from Romania, except the illustrations from Fig. 3c, d which belongs to a specimen collected from and the illustration from Fig. 3e that belongs to a specimen from Austria.

b

a c

d

e Figure 2. Litus camptopterus Novicky: a – 1, species distribution in Piatra Neamț and surrounding areas (Moldova province map); b – forewing; c, e – variation in female antennae; d – female habitus (original).

L. cynipseus – additional material: 1♀, 11.05.2011, Cozla hill – Piatra Neamţ, NT (500 m elev.), (Leg. Pricop E.), collected with yellow pan traps from broadleaf forest dominated by Fagus sylvatica; 2♀, 2.08.2010, Rarău, SV (Leg. Pricop E.), collected with sticky traps from spruce forest (1400 m elev.); 1♀, 3.10.2011, Uricani forest, reservation area, IS (Leg. Pricop E.), collected with the sweep net and the aspirator (pooter) from mixed vegetation in the broadleaf forest area; 1♀, 26.09.2010, Codru Secular Giumalău in the reservation area (1300 m elev.) SV, (Leg. Pricop E.), collected with the sweep net and a pooter from mixed vegetation in the spruce forest; 1♀, 17.07.2010, Copou – Iaşi, UAIC area, (Leg. Pricop E.),

- 22 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 captured from a window in the Faculty of Biology, Laboratory of Zoology, UAIC; 2♀, 14.08.2010, Piatra Neamţ, from a small private garden with vegetables, flowers and Vitis vinifera, NT (Leg. Pricop E.), collected with yellow pan traps (330 m elev.); 1♀, 21.05.2006, The Botanical Garden from Iaşi (Leg. Pricop E.), collected with the sweep net and the pooter from a dry meadow; 1♀, 28.05.2014, Cernegura hill near Piatra Neamț (500 m elev.) NT (Leg. Pricop E.), collected with the sweep net and the aspirator from mixed vegetation in the broadleaf forest area dominated by Fagus sylvatica and Picea abies; 1♀, 25.07.2014, Dealul Vulpii – Botoaia stepic reservation, near Ciritei, NT (Leg. Pricop E.), collected with the sweep net from the stepic meadow area.

a b

d

c e

f

g

h i Figure 3. Litus cynipseus Haliday, 1833: a – head; b – forewing; c - body; d, e, f – small variations in female antennae; g – reticulated part of propodeum and mesopleuron; h – metasoma; i – reticulated coxae and ventral mesosoma (original).

Hosts: L. cynipseus was reared from Ocypus sp., Staphylinus sp., Staphylinus olens (Col., Staphylinidae); Limnobates sp. (Col., Hydrometridae) (Noyes, 2003).

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Distribution: Holarctic Region – Austria, Belgium, Bulgaria, Canada, Czech Republic, , , France, , Hungary, , Japan, Kyrgyzstan, Mexico, Morocco, Nepal, Netherlands, Romania, Serbia and Montenegro, Moldova, Republic of Korea, Russia, , Spain, Switzerland, , United Kingdom, USA (Noyes, 2003; Triapitsyn & Berezovskiy, 2004). In Eastern Romania, L. cynipseus is widely distributed. Taxonomic notes: In this region L. cynipseus is the most common species of this genus. Females are easy to identify from the narrow wings with almost parallel margins (Fig. 3b), F2 shorter than the pedicel, clava ovoid and compact at max 2.6 times as long as broad (Fig. 3f). Clava about 3 times more broad than the funicle articles. The ovipositor is short. The specimens collected from Agigea sand dune reserve (Constanța County) are smaller compared to the rest of the specimens. L. cynipseus is a common species, present in natural and anthropic ecosystems, distributed from sea level to hills and mountain areas of Dobrogea and Moldova provinces (Romania). Ulrich (1999) recorded L. cynipseus from a dry meadow on limestone. I had the good opportunity to examine a few specimens belonging to L. cynipseus (Fig. 4a, b, c) from the Natural History Museum of Vienna, specimens collected from different countries like England, Austria etc.

a

b

c Figure 4. Slide-mounted specimens belonging to L. cynipseus from the Museum of Natural History of Vienna (original).

Conclusions Litus camptopterus is here reported for the first time from Moldova province (Romania). In this region L. camptopterus is rarely collected and is recorded only from some areas of Bistrița River Basin (Neamț County). The species distribution sites are

- 24 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 situated above the 44°N. L. camptopterus belongs to different ecosystems from Bistrița River Valley. Species distribution and habitat is situated from about 330 to 500 m elevation. For L. camptopterus the specific habitat seems to be difficult to identify, its habitat is related to meadow vegetation, to Salix sp. and to the urban environment. L. cynipseus is a more common and widely distributed species. For L. cynipseus, we have been able to identify the natural habitat acording to NATURA 2000 and/or EUNIS, from Donita et al. (2005): 9410 Acidophilous Picea forests of the montane to alpine levels (Vaccinio-Piceetea), 91V0 Dacian beech forest (Symphylo-Fagion), 6240 Subpannonic steppic Grasslands and EUNIS: B1.4B1 Western Pontic fixed dunes. In our specimens of Litus camptopterus, although we found some variations in the length of the first four funicle segments (F1-F4), we also found that the ovipositor length/mid-tibia length ratio slightly varies from 0.72 to 0.78 and the forewings are very similar in shape. In the case of L. cynipseus, the ovipositor length/mid-tibia length ratio vary from 0.9 to 1.2 and length of the first three funicle segments (F1-F3) and length of all legs also vary, probably depending on body size variation. Regarding the female antennae of the two species (Litus camptopterus and L. cynipseus), the spindle-like sensillae are present and distributed in each species on F3, F4, F5 and F6. In Litus camptopterus the sensory ridges of clava are long, reaching (from the apex) the basal part of this segment. In L. cynipseus the sensory ridges of clava are shorter and are grouped in the middle and upper part of this segment, not reaching the base of clava. Numerically, seems to be more sensilla basiconica on the fore tibia of L. camptopterus than in the foretibia of L. cynipseus.

Acknowledgements This work was supported by the strategic grant POSDRU/159/1.5/S/133391, Project “Doctoral and Post-doctoral programs of excellence for highly qualified human resources training for research in the field of Life sciences, Environment and Earth Science” cofinanced by the European Social Fund within the Sectorial Operational Program Human Resources Development 2007-2013”. I am grateful to PhD Prof. Ionel Andriescu (UAIC) and PhD Prof. Ioan Moglan (UAIC) for references, advices, laboratory conditions and scientific support. I am also grateful to physicist Dumitru Raileanu (UAIC) for technical assistance regarding all SEM pictures. I thank also to Curator Dominique Zimmermann (NHMW) for providing me facilities for specimens examination.

References Botoc, M., 1964. Nouvelle contributions à l'étude des chalcidoidés de Roumanie (X). Studia Universitatis Babes- Bolyai, Cluj (Ser. Biol.), 1964(1): 79-85. Botoc, M., 1974. Noi contributii la studiul Mimaridelor (Hymenoptera, Chalcidoidea) din Romania. Studia Universitatis Babes-Bolyai, Cluj (Ser. Biol.), 19(1): 103-106. Donita, N., Pauca-Comanescu, M., Popescu, A., Mihailescu, S., Biria, I., 2005. Habitatele din Romania. Editura Tehnica Silvica, Bucureşti. Novicky, S., 1953. Vorläufige Beschreibung einiger Mymariden (Hymenoptera, Chalcidoidea), I. Teil. Entomologisches Nachrichtenblatt Österreichischer und Schweizer Entomologen, 5(1/2): 14. Noyes, J.S., 2003. Universal Chalcidoidea Database. World Wide Web electronic publication. (accessed 7 September 2014). Pricop, E., 2009a. A faunistic review of the Romanian Mymaridae and Mymarommatidae (Hymenoptera Chalcidoidea and Mymarommatoidea). In Lucrările Simpozionului Entomofagii şi rolul lor în

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păstrarea echilibrului natural, Analele Ştiinţifice ale Universităţii „Al. I. Cuza” din Iaşi, s. Biologie , Supliment, 121-128. Pricop, E., 2009b. Preliminary studies of the Mymaridae (Hym., Chalcidoidea) from Neamţ county, Romania, species distribution, vascular flora/vegetation, an ecological approach. AES Bioflux, 1(1): 13-29. Pricop, E., 2013. Identification key to European genera of the Mymaridae (Hymenoptera: Chalcidoidea), with additional notes. ELBA Bioflux, 5(1): 69-81. Triapitsyn, S.V., Berezovskiy, V.V., 2004. Review of the genus Litus Haliday, 1833 in the holarctic and oriental regions, with notes on the Palaearctic species of Arescon Walker, 1846 (Hymenoptera: Mymaridae). Far Eastern Entomologist, 141: 1-24. Viggiani, G., 1973. Ricerche sugli Hymenoptera Chalcidoidea. XXXVIII. Descrizione de maschio di Litus Haliday (1833) (Mymaridae). Bollettino del Laboratorio di Entomologia Agraria 'Filippo Silvestri', Portici, 30: 231-234. Ulrich, W., 1999. Phenology and life cycles of the parasitic Hymenoptera of a dry meadow on limestone. Polskie Pismo Entomologiczne, 68(4): 384-405.

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COMPOSITION AND DISTRIBUITION OF BENTHIC MACROINVERTEBRATES IN THE PRUTH RIVER (2012-2013)

Oxana MUNJIU*, Ion TODERAȘ, Elena ZUBCOV, Lucia BILETCHI and Igor SUBERNETKII Institute of Zoology, Academy of Sciences of Moldova, 1 Academiei Street, MD 2028, Chişinău, Republic of Moldova, *[email protected]

Abstract. The diversity and state of the benthic invertebrate communities from the Pruth River (within the borders of the Republic of Moldova) has been investigated during May of 2012-August of 2013. Twelve sampling stations have been chosen along the river, including Costeşti-Stânca reservoir. During the given period 159 taxa of the benthic macroinvertebrates have been identified. The alive individuals of Theodoxus transversalis (Pfeiffer, 1828) - rare species included in the IUCN Red List - have been registered. Also, in 2012 for the first time for Moldova was reported Pisidium moitesserianum (Paladilhe, 1866). The Pruth ecosystem supports a higher human impact in those river sectors, which correspond to Duruitoarea Nouă, Costeşti-Stânca and Giurgiuleşti sampling stations, thus reflecting the impact of environmental conditions upstream the Costeşti-Stânca dam and in area of Giurgiuleşti port, correspondingly. The most favourable conditions for development of benthic invertebrate communities are in the area of Branişte station, which is proved by biodiversity, the number of pollution-sensitive species and saprobity indexes.

Keywords: benthic macroinvertebrates, species diversity, density, biomass, molluscs, Ephemeroptera, Trichoptera, Pruth River

Rezumat. Structura şi distribuţia macronevertebratelor bentonice în râul Prut (2012-2013). Diversitatea şi starea comunităţilor nevertebratelor bentonice din râul Prut în limitele Republicii Moldova a fost investigată în perioada mai 2012-august 2013. Pe cursul râului, inclusiv lacul de acumulare Costeşti-Stânca, au fost selectate 12 staţii de colectare a eşantioanelor. Pe durata cercetărilor au fost identificaţi 159 de taxoni de macronevertebrate bentonice. Au fost depistaţi indivizi vii de Theodoxus transversalis (Pfeiffer, 1828) - specie rară, inclusă în Lista Roşie a IUCN. De asemenea, în anul 2012 în premieră pentru Moldova a fost înregistrat Pisidium moitesserianum (Paladilhe, 1866). Ecosistemul Prutului este supus unui impact antropic mai puternic în acele porţiuni de râu care corespund staţiunilor de colectare Duruitoarea Nouă, Costeşti-Stânca şi Giurgiuleşti, ceea ce reflectă influenţa condiţiilor de mediu în amontele barajului Costeşti-Stânca şi în regiunea portului Giurgiuleşti. Cele mai favorabile condiţii pentru dezvoltarea nevertebratelor bentonice sunt la staţiunea Branişte, fapt confirmat de biodiversitatea, numărul speciilor stenobionte şi valorile indicilor de saprobitate.

Cuvinte cheie: macronevertebrate bentonice, biodiversitate, efectiv numeric, biomasă, moluşte, Ephemeroptera, Trichoptera, râul Prut

Introduction Study of the composition and distribution of benthic invertebrates is one of the mandatory elements of the monitoring of freshwater ecosystems. Benthic invertebrates are a group of animals, which meets several requirements for the monitoring and indicator organisms: wide distribution, enough high density, relatively large size of the body, combination of populating certain biotopes and a certain degree of mobility. The Pruth River is the second largest river of the Republic of Moldova, originating in Ukraine and draining territories of Romania and Moldova. The total length of the river is 898 km, of which 695 km are within the borders of Moldova. As many other water courses, the Pruth River supported human intervention – in 1978, at 560 km from the mouth of the

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Pruth River, the Costeşti-Stânca reservoir was built. The Pruth River represents the last largest left tributary of Danube, and since it belongs to the Wetland Danube River Area, the investigation of its ecological state, including state of fauna, gets an international importance.

Material and Methods There have been collected more than 220 samples of the benthic macroinvertebrates from the Pruth River during May of 2012-August of 2013. The common sampling stations were Costeşti-Stânca, Branişte, Sculeni, Leuşeni, Leova, Cahul, Cîşliţa- Prut, and Giurgiuleşti, and only in July 2013 four more stations were investigated: Criva, Teţcani, Bădragii Noi, and Duruitoarea Nouă (Fig. 1).

Figure 1. Location of sampling sites along the Pruth River: 1) Criva, 2) Teţcani, 3) Bădragii Noi, 4) Duruitoarea Nouă, 5) Costeşti-Stânca, 6) Branişte, 7) Sculeni, 8) Leuşeni, 9) Leova, 10) Cahul, 11) Cîşliţa-Prut, and 12) Giurgiuleşti.

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Benthic invertebrates have been sampled in shallow zones of the Pruth River, including the Costeşti-Stânca reservoir, at depths up to 1.2 m, applying standardized methods (Rukovodstvo po metodam gidrobiologicheskogo analiza poverhnostnyh vod i donnyh otlojenii, 1983). The Petersen grab with area of capture of 1/40 m2 and dredge with area of capture of 8 m2 have been used for quantitative sampling and also a hand net for different substrates, including macrophytes, in the case of qualitative sampling. The preservation of the samples has been made by adding of 4% formaldehyde and 70% alcohol. All individuals have been sorted as much as possible by groups or species afterwards in the laboratory and identified by using identification keys (Jadin, 1952; Opredeliteli fauny Chiornogo i Azovskogo morei. Svobodnojivuscie bespozvonocinie, 1968, 1969, 1972; Opredeliteli presnovodnyh bespozvonochnyh Evropeiskoi chasti SSSR (plankton i bentos), 1977; Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 1994, 1995, 1997, 2000, 2001, 2004). The identification of species has been carried out by the stereomicroscope МБC-9 and upright microscope Jenaval (Zeiss), but since April 2013 the laboratory investigations have been performed by the aid of the stereomicroscope SteREO Discovery.V8 (Zeiss) and upright microscope Axio Imager А.2 (Zeiss). The hydrobiont biomass has been determined via their weighting on analytical balance ABS 80-4 Kern, being previously dried up on a paper filter till the disappearance of wet spots. The density and biomass of hydrobionts has been transferred into ind./m2 and g/m2, respectively. Saprobity indexes have been calculated accordingly to Pantle & Buck (1955), Sladecek (1973), and standardized methods of water quality investigation (Unifizhirovanye metody issledovaniia kachestva vod, 1977).

Results and Discussion The total number of invertebrate taxa has reached 159, including 20 taxa of Annelida, 22 of Chironomidae, 17 of Crustacea, 12 of Ephemeroptera, 11 of Trichoptera, 21 of Gastropoda, 17 of Bivalvia and 39 taxa of other groups. The most spread taxa were: Gordiacea – Gordius aquaticus (Linnaeus, 1758); Gastropoda – Lithoglyphus naticoides (C. Pfeiffer, 1828), Fagotia acicularis (Ferussac, 1823), Theodoxus fluviatilis (Linne, 1758), Theodoxus danubialis (C. Pfeiffer, 1828), Viviparus viviparus (Linne, 1758), Viviparus contectus (Millet, 1813); Bivalvia – Anodonta piscinalis (Nilsson, 1822), Unio tumidus (Philipsson, 1788), Crassiana crassa (Philipsson, 1788), Pisidium amnicum (Müller, 1774), Dreissena polymorpha (Pallas, 1771), Dreissena bugensis (Andrusov, 1897); Tibificidae – Tubifex tubifex (Müller, 1774), Branchiura sowerby (Beddard, 1892), Limnodrilus hoffmeisteri (Claparède, 1862); Lumbriculidae – Lumbriculus variegatus (Müller, 1773); Naididae – Ophidonais serpentina (Müller, 1773), Stylaria lacustris (Linnaeus, 1767); Myzidae – Limnomysis benedeni (Czerniavsky, 1882), Paramysis lacustris (Czerniavsky, 1882); Gammaridae – Dikerogammarus haemobaphes (Eichwald, 1841), Pontogammarus crassus (Sars, 1894), Chaetogammarus warpachowskyi (Sars, 1894), Chaetogammarus ischnus (Stebbing, 1899); Corophiidae – Corophium robustrum (Sars, 1895), Corophium curvispinum (Sars, 1895);

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Isopoda – Jaera sarsi (Valkanov, 1936); Ephemeroptera – Baetidae, Baetis rhodani (Pictet, 1843), Heptagenia flava (Rostock, 1877), Heptagenia coerulans (Rostock, 1877), Palingenia longicauda (Olivier, 1791), Potamanthus luteus (Linne, 1767); Odonata – Gomphus flavipes (Charpentier, 1825), Gomphus vulgatissimus (Linnaeus, 1758), Platycnemis pennipes (Pallas, 1771), Agrion splendens (Harris, 1782), Agrion virgo (Linnaeus, 1758); Heteroptera – Plea minutissima (Leach, 1817), Mesovelia sp., Nepa cinerea (Linnaeus, 1758), Aphelocheirus aestivalis (Fabricius, 1794); Coleoptera – Dytiscus sp., Haliplus sp.; Trichoptera – Hydropsychidae, Hydropsyche ornatula (McLachlan, 1878), Polycentropus sp., Ecnomus tenellus (Rambur, 1842), Triaenodes bicolor (Curtis, 1834), Hydroptila tineoides (Dalman, 1819), Limnephilidae; Chironomidae – Chironominae Polypedium gr.nubeculosum (Meigen, 1804), Microtendipes gr. chloris (Meigen, 1818), Chironomus plumosus (Linnaeus, 1758), Tanypodinae Tanypus punctipennis (Meigen, 1818), Tanypus vilipennis (Kieffer, 1918), Orthocladiinae Cricotopus gr. algarum (Kieffer, 1911), Diamesa insignipes (Kieffer, 1908); Ceratopogonidae – Bezzia hydrophila (Kieffer, 1909); Culicidae – Culicoides setosinervis (Kieffer, 1913). The taxa with the lowest occurrence were: Bryozoa, Collembola, Theodoxus transversalis (Pfeiffer, 1828), Pisidium moitesserianum (Paladilhe, 1866), Conchostraca, Notostraca, Ephemera vulgata (Linnaeus, 1758), Polymitarsis virgo (Oliver, 1791), Phryganeidae, Anabolia furcata (Brauer, 1857), Mystacides sp., Simuliidae, and Sialidae. Theodoxus transversalis (Pfeiffer, 1828) is a rare species included in The International Union for Conservation of Nature (IUCN) Red List. Only empty shells of this species were occasionally registered in the previous decade, but in summer of 2013 ten alive individuals were met. This is an important fact for monitoring and preservation of T. transversalis, since each population of this rare species and, respectively, their habitats need protection. Pisidium moitesserianum (Paladilhe, 1866) is a new species for Moldova, which was registered for the first time in 2012 at Branişte station. The highest number of rare species has been remarked at the Teţcani and Branişte stations, and among them are Theodoxus transversalis (Pfeiffer, 1828), Pisidium moitesserianum (Paladilhe, 1866), Ephemera vulgata (Linnaeus, 1758), Polymitarsis virgo (Oliver, 1791), Anabolia furcata (Brauer, 1857), species of Simuliidae, Sialidae. Also, there have been registered species, which are characteristic for clean zones of aquatic ecosystems – seven species of Ephemeroptera and nine species of Trichoptera. These stations are remarkable by their highest diversity of substrates: stones, gravel, sand, silty sand, silt, macrophytes, sunken trees, which, together with the absence of adverse effects of Costeşti- Stânca reservoir and waste water discharges, contribute to the creation of most favourable environmental conditions for lots species of benthic animals. Conchostraca and Notostraca have been identified only at Cîşliţa-Prut station. Along the river stream, the diversity of benthic invertebrates has differed significantly. The Branişte station has distinguished by the highest values of density and species diversity – there have been registered up to 85 species (Fig. 2).

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90 85 80 70 60 50 50 50 53 40 43 39 30 29 20 10 11 11 8 4 6 0 Cri Tet BaN DuN C-S Bra Scu Leu Leo Cah C-P Giu

Figure 2. Number of taxa along the Pruth River (Cri – Criva, Tet – Teţcani, BaN – Bădragii Noi, DuN – Duruitoarea Nouă, C-S – Costeşti-Stânca, Bra – Branişte, Scu – Sculeni, Leu – Leuşeni, Leo – Leova, Cah – Cahul, C-P – Cîşliţa-Prut, Giu – Giurgiuleşti).

The differences may occur because of various ecological conditions: hydromorphological, hydrochemical, type of substrate, and level of anthropogenic load. Taking in account the number of registered species, it shall be concluded the best environmental conditions have preserved in at Branişte and Cîşliţa-Prut. A relatively low number of species have been identified at Teţcani station, but it should be emphasised that the sampling in this river sector was performed only in July of 2013, as was mentioned above. Consequently, high number of rare species gives the ground to wait in the case of a regular sampling an extension of species list at Teţcani station. The least favourable for biodiversity sites are located near the dam of Costeşti- Stânca reservoir, downstream the confluence of the Jijia River and at the mouth of the Pruth River. In the last case the impact of Giurgiuleşti port is a strong one: beside the presence of unfavourable hydrochemical conditions, there is another factor, which affects the development of benthic invertebrates and namely the regular clearance of the river fairway. It has been observed that if the sampling is conducted after such works, the macroinvertebrates are missing in the benthic samples or their number and species composition are very limited, e.g. during the entire period of investigations the highest density was equal to 480 ind./m2, with a biomass of 0.8 g/m2 and it was composed of representatives of Tibificidae. There has been observed a reduction of the density without molluscs and total zoobenthos and biomass without molluscs (Fig. 3.1; 3.3; 3.4) downstream the Pruth River and an increase of the biomass of total zoobenthos, excepting the Giurgiuleşti station, where it is the lowest due to the most severe anthropogenic pressure. The increase of the biomass of total zoobenthos was accomplished with the contribution of large bivalve molluscs (Fig. 3.2). The total biomass has varied from 0.006 g/m2 to 2971.764g/ m2, the biomass without molluscs – from 0.006 g/m2 to 58.4 g/m2, the density of total zoobenthos – from 6 ind./m2 to 39000 ind./m2, and the density of zoobenthos without molluscs – from 4 ind./m2 to 32200 ind./m2 The density and biomass of benthic organisms have had the lowest values at Giurgiuleşti station (Fig. 3.2).

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1 2

3 4 Figure 3. Density and biomass of macrozoobenthos along the Pruth River in 2012-2013 (sampling sites: 1 – Branişte, 2 – Sculeni, 3 – Leuşeni, 4 – Leova, 5 – Cahul, 6 – Cîşliţa-Prut, 7 – Giurgiuleşti). The box represents the Mean, and the line on the top of the box shows the Standard Error of Mean.

Under the impact of anthropic factors primary the sensitive groups as Plecoptera, Ephemeroptera and Trichoptera get lost from the composition of macrozoobenthos. Representatives of Plecoptera have been identified at Branişte station during 2010-2011, but no one of them has been registered during 2012-2013. In samples collected at Giurgiuleşti station representatives of Plecopterа, Ephemeroptera and Trichoptera were absent (Fig. 4). The ecosystem of the Pruth River at the Cîşliţa-Prut station supports both a chemical and biological pollution. The presence of last one is demonstrated by the fact that in a range of samples such species as Corbicula fluminea (Müller, 1774) and Sinanodonta woodiana (Lea, 1834) constitute a significant part of macrozoobenthos according to their density, as well as biomass. As example, C. fluminea forms about 40% (200 ind./m2) of total density of molluscs and S. woodiana – more than 70% (260 g/m2) of total biomass of malacofauna at the Cîşliţa-Prut station. The state of benthic invertebrate communities and the density of invertebrate taxa are extremely important for the determination of water quality, and assessment of aquatic ecosystem state, in general. Thus, during the investigation period the saprobity indexes – SI (P&B) have varied from 1.709 (Branişte, February of 2013), which corresponds to the 1st

- 32 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 class of water quality, to 3.53 (Leuşeni, March of 2013) and 3.7 (Cîşliţa-Prut, February 2013), which corresponds to 5th class of water quality.

Criva Tetcani Badragii Noi Duruitoarea Noua Costesti-Stinca Braniste Sculeni Leuseni Leova Cahul Cislita-Prut Giurgiulesti

0 10 20 30 40 50 60 70 80 90 100 Mollusca Oligochaeta Chironomidae Crustacea Ephemeroptera Trichoptera Other groups

Figure 4. Taxonomic structure of macrozoobenthos of the Pruth River sampling sites in 2012-2013.

In this way, the saprobity zones calculated based on macrozoobenthos from the Pruth River have varied within β-mesosaprobic and α-mesosaprobic, and the water quality class within the moderately polluted and critically polluted.

Conclusions The results of carried out investigations have revealed that the sectors of the Pruth River, which are located in the area of Duruitoarea Nouă, Costeşti-Stânca and Giurgiuleşti sampling stations bear the highest pressure of the human factor and reflect the impact of environmental conditions upstream the Costeşti-Stânca dam and in area of Giurgiuleşti port, correspondingly. The most favourable conditions are in the area of Branişte sampling station, which is proved by high values of biological diversity, the presence of a large number of pollution-sensitive species and low values of saprobity indexes.

Acknowledgements Funding for this research came from the European Commission in the frame of the Cross-Border Cooperation Program Romania-Ukraine-Republic of Moldova – project MIS ETC-1150 RESOURCES PILOT CENTRE FOR CROSS-BORDER PRESERVATION OF THE AQUATIC BIODIVERSITY OF PRUT RIVER. Jechiu I., from the Institute of Ecology and Geography of the Academy of Sciences of Moldova, is thanked for helping with the preparation of map of sampling stations.

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References Jadin, V.I., 1952. Molliuski presnyh i solonovatyh vod SSSR. Moskwa-Leningrad: AN SSSR, 346 p. Opredeliteli fauny Chiornogo i Azovskogo morei. Svobodnojivuscie bespozvonocinie, 1968. Pod red. Morduhaia- Boltovskogo F.D. Tom 1. Prosteishie, gubki, kishecinopolostnye, cervi, sciupaltsevye. Kiev: Naukova dumka. Opredeliteli fauny Chiornogo i Azovskogo morei. Svobodnojivuscie bespozvonocinie, 1969. Pod red. Morduhaia- Boltovskogo F.D. Tom 2. Rakoobraznye. Kiev: Naukova dumka. Opredeliteli fauny Chiornogo i Azovskogo morei. Svobodnojivuscie bespozvonocinie, 1972. Pod red. Morduhaia- Boltovskogo F.D. Tom 3. Chlenistonogie (krome rakoobraznyh), molliuski, iglokozhie, shhetinkocheliustnye, hordovye. Kiev: Naukova dumka. Opredeliteli presnovodnyh bespozvonochnyh Evropeiskoi chasti SSSR (plankton i bentos), 1977. Pod red. Kutikovoi L.A., Starobogatova E.I. Leningrad: Gidrometeoizdat. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 1994. Pod red. Tsalolihina S.Ia. Tom 1. Nizshie bespozvonochnye. Sankt-Peterburg: Nauka. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 1995. Pod red. Tsalolihina S.Ia. Тom 2. Rakoobraznye. Sankt-Peterburg: Nauka. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 1997. Pod red. Tsalolihina S.Ia. Тom 3. Paukoobraznyeju. Nizshie nasekomye. Sankt-Peterburg: Nauka. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 2000. Pod red. Tsalolihina S.Ia. Тom 4. Dvukrylye nasekomye. Sankt-Peterburg: Nauka. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 2001. Pod red. Tsalolihina S.Ia. Тom 5. Vysshie nasekomye. Sankt-Peterburg: Nauka. Opredeliteli presnovodnyh bespozvonochnyh Rossii i sopredelinyh territorii, 2004. Pod red. Tsalolihina S.Ia. Тom 6. Molluski. Policheti. Nemertini. Sankt-Peterburg: Nauka. Pantle, R., Buck, H., 1955. Die biologische Überwachung der Gewässer und die Darstellung der Ergebnisse. Gas und Wasserfach, 96: 604-620. Rukovodstvo po metodam gidrobiologicheskogo analiza poverhnostnyh vod i donnyh otlojenii, 1983. Pod red. Abacumova V.A. Leningrad: Gidrometeoizdat. Sladecek, V., 1973. System of water quality from the biological point of view. Archiv für Hydrobiologie Ergebnisse der Limnologie, 7: 1-218. Unifizhirovanye metody issledovaniia kachestva vod, 1977. Chiasti 3. Metody biologicheskogo analiza. M.: SEV.

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STUDY OF MACROINVERTEBRATE DIVERSITY AND SEASONAL DYNAMICS FROM BĂDĂRĂU LAKE – IAŞI, ROMANIA

Marius Andrei RĂU*, Gabriel PLĂVAN, Ștefan Adrian STRUNGARU and Mircea NICOARĂ Faculty of Biology, Alexandru Ioan Cuza University, Bd. Carol I 11, 700506 Iaşi, Romania. *[email protected]

Abstract. In the investigations carried out on Bădărău Lake in 2011-2013 period during all the seasons on three established sites, the total number of taxa inventory was 61. There were calculated the indices of abundance, dominance, ecological significance (Dzuba) and diversity (Shannon and evenness or equitability index). The main points that we covered within the study was the evaluation and monitoring of the benthic macroinvertebrates as well as the analysis of the seasonal dynamics regarding this organisms.

Keywords: benthic macroinvertebrates, lake, seasonal dynamics, ecological index

Rezumat. Studii privind diversitatea și dinamica sezonieră a macronevertebratelor din lacul Bădărău – Iași, România. În studiul efectuat pe lacul Bădărău, în anii 2011-2013 de-a lungul tuturor anotimpurilor, pe trei stații stabilite, numărul total de taxoni a fost de 61. Au fost calculați indicii de abundență, dominanță, semnificație ecologică (Dzuba) și ai diversității (Shannon și indicele de echitabilitate). Principalele puncte pe care le-am acoperit în acest studiu au fost evaluarea și monitorizarea macronevertebratelor bentonice cât și analiza dinamicii sezoniere în ceea ce privește aceste organisme.

Cuvinte cheie: macronevertebrate bentonice, lac, dinamică sezonieră, indice ecologic

Introduction Benthic macroinvertebrates associations are unified into the structure of the great majority of continental aquatic ecosystems (streams, rivers, lakes and puddles), generally representing the numerical and/or biomass dominant components and the major links of the channels of matter and energy transfer (Plăvan & Nicoară, 2012). In many parts of the world, water is a limiting factor of the economy, food production and the spread of populations. The expanding of pollution is increasing pressures by depletion of underground springs, lowering groundwater levels and deterioration of ecological systems (Mustață, 2000). Bădărău Lake from Botanical Garden of Iași, Romania, in addition of the role of recreational area, which was originally designed, has the function of a complex ecosystem which tends to mature. The lake is characterized by a high specific diversity, including species belonging to all domains that compose a pool of stagnant water: neuston, pleuston, plankton, nekton and benthos. Specific composition, numeric and percentage abundances and density of each species were calculated to identify the benthic macroinvertebrates communities. The main objective of this study is the real and complex evaluation of the benthic macroinvebrates diversity as well as the analysis of ecosystem changes within the seasons regarding these organisms.

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Material and Methods The study was carried out on Bădărău Lake which it is situated in the Botanical Garden of Iași City, at an altitude of 54-60 m. The lake is located on Bădărău River, a tributary of Bahlui River. It was built between 1980-1982 and has an area of aproximately 25000 m2. It has a length of 400 m, a maximum depth of 2 m and a maximum volume of 7000 m3. The main influence on the climate is dry continental with frequent cold north winds in the winter. To establish an appropriate set of sampling sites, it is often useful to carry out preliminary samplings. In the case of qualitative analysis, samples will be collected from various habitats and it should have take into account the life cycles of various species of benthic organisms (Surugiu, 2008).

Figure 1. Location of the sampling sites on Bădărău Lake (Google Earth).

Sampling was performed during 2011-2013, in all the seasons. There were established three stations: Sampling Site 1 (Inflow), Sampling Site 2 (Forest) and Sampling Site 3 (Outflow). Three sets of benthic samples were collected from the three sampling sites during the months of October 2011, December 2011, April 2012, June 2012, November 2012, June 2013 and November 2013. The samples were taken at each sampling site using benthic net and a modified Petersen grab (170.5 cm2 covered area), after that being washed through a 0.25 mm sieve. GPS coordinates and the physico-chemical parameters (altitude, water and air temperature, pH, conductivity) were measured for every sampling site. The sampling sites have been established to intercept as accurate as possible the hydrobiological aspects, the aquatic vegetation (periphyton, macrophytes), riparian vegetation (shrubs, reed, herbaceous plants) and the substrate structure (rockfill, gravel, sand, mud). The macroinvertebrate diversity was estimated by the means of the Shannon- Wiener index (expressing community structure and evolution in time and space): H = - ∑n/N log2 n/N. Evenness (J) was calculated using Pielou index (expressing relative

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information within the ecosystem): J = H/Hmax, where Hmax is the maximal diversity calculated from: H(S)max = log2S (Magurran, 1988). Constancy (C) expresses the continuity of the species occurrence within the given biotope. This characteristic is considered to be a structural indicator, because it shows in what proportion one species participates to the biocoenosis structure edification. As high the indicator value is, as better adapted to the biotope conditions the given species is considered to be. Constancy was expressed by the relation: CA= nA/N x 100, in which: CA – species constancy; nA – number of samples with the species A; N – total number of samples examined. Dominance (D) shows the relationship between the contingents of a given species against the sum of the individuals belonging to other species with which it associates, expressing the relative abundance. Dominance is considered to be a productivity indicator, because it shows the percentage of species participation to the achievement of the biomass production within the biocoenosis. Dominance (D) was calculated using the relation: DA= nA/N x 100, in which: DA= species dominance; nA = total number of individuals belonging to species A, present in the samples researched; N= total number of individuals belonging to all the species present in the samples researched. The ecological significance index (W) represents the relationship between the structural indicator (C) and the productivity indicator (D), reflecting more eloquent one species position within the biocoenosis. The relation used was: WA = CAxDAx100/10,000, in which: WA - ecological significance index of the species A; CA - species constancy; DA species dominance. To enable interpretation of evidence from qualitative and quantitative point of view it was necessary individual screening and identification. Qualitative analysis of populations of macroinvertebrates was done through species inventory, quantitative analysis being done by statistical processing data obtained by taxa counting and determination (Nicoară, 2008).

Results and Discussion According to Bertrand (1954), the taxonomic-ecological spectrum of one biocoenosis only indicates the first step to the knowledge of the biocoenosis structure, and the knowledge of the organizational structure of a biocoenosis requires the understanding of the substance and energy manner of distribution to the community members. A series of ecologic indicators were calculated to characterize the biocoenosis: analytic (abundance, constancy, and dominance) and synthetic (ecological significance index). Benthic macroinvertebrates offers the best possibilities for the monitoring and evaluation of biological water quality. It is estimated that from the 100 different biological assessment methods currently existing, two-thirds rely on macroinvertebrates (Surugiu, 2008). A total number of 61 macroinvertebrates taxa were identified, including such animal groups as crustaceans, and mollusks summing a total of 4911 individuals. In the autumn of 2011, 17 taxa of benthic macroinvertebrates were identified following the sampling (Fig. 2). The most abundent taxa were Chironomidae family and Gammarus pulex, this two taxa being also the most eudominant in the lake. The most frequent taxa found in this season were Baetis spp. and Chironomidae family. Based on the index of ecological significance (Dzuba), Asellus aquaticus and Corixa spp. were

- 37 - Marius Andrei Rău et al. categorized as accessory taxa, Dytiscus spp. and Limnephilus spp. as accidental species, Baetis spp., Chironomidae family, Coenagrion puella and Gammarus pulex being the characteristic taxa.

Figure 2. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

Following the sampling in the winter of 2011, a total of 12 taxa were identified (Fig. 3). Chironomids, Gammarus pulex and Baetis spp. were the most abundent taxa in the lake. Regarding the frequency of the benthic macroinvertebrates found in the lake, Baetis spp. and Chironomidae family were the only constant organisms, the other ones being accidental. The dominance index indicated Chironomidae family and Gammarus pulex as eudominant, Baetis spp. as dominant, the rest of them being classified as subrecedent species. Based on the index of ecological significance, Chironomids, Gammarus pulex and Baetis spp. were classified as characteristic taxa. This analysis indicated also that the remaining taxa (9) were marked as accesory organisms. A total of 11 taxa were found in the spring of 2012 (Fig. 4). As in the past two seasons, Chironomidae family and Gammarus pulex were the most abundent taxa. Nemoura spp. and Limnephilus spp. had also an increased abundance value. Chironomids were also the most frequent organisms found in the lake as well as Baetis spp. genus. Oligochaetes were identified as accesory organisms. Chironomidae family was the only eudominant group (dominance index) and characteristic (Dzuba). The dominant taxa were Asellus aquaticus, Baetis spp., Limnephilus spp. and Nemoura spp. genus. Valvata naticina was indicated as accidental species in the lake.

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Figure 3. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

The summer sampling of 2012 resulted a total number of 15 taxa (Fig. 5). Gammarus pulex and Chironomidae family were eudominant and the most abundent in the lake. There is an observation in this season regarding the frequency of the species, indicating that no constant taxa were found in the lake. Following that we identified as accesory species Baetis spp., Chironomids, Gammarus pulex, Limnephilus spp., Notonecta spp. and Physa acuta, the rest being characterized as accidental organisms. Regarding the dominance index, Baetis spp. and Elodes marginata were categorized as dominant taxa, Asellus aquaticus, Limnephilus spp. and Naucoris spp. as subdominant, Corixa spp., Caenis spp., Coenagrion puella, Notonecta spp. and Physa acuta as recedent taxa. The only characteristic species was Gammarus pulex, the other remaining taxa being categorized as accessory organisms. A total of 34 taxa were identified in the sampling performed in the autumn of 2012 (Fig. 6). The most abundant benthic macroinvertebrates were Chironomidae family and Gammarus pulex. Asellus aquaticus, Baetis spp., Chironomids and Oligochaetes were the most frequent organisms. Following the dominance index, resulted that Chironomidae family and Gammarus pulex were categorised as eudominant taxa and Baetis spp. and Asellus aquaticus as dominant, the rest falling into subrecedent taxa. Based on the index of ecological significance, we identified as characteristic taxa Gammarus pulex, Asellus aquaticus, Baetis spp. and Chironomidae family, Ecnomus spp., Lymnaea spp., Orthetrum spp., Physa acuta, Platycnemis spp., Polycentropus spp., Coenagrion puella as accessory organisms and Aeshna spp., Cybister spp., Erpobdella spp. as accidental benthic macroinvertebrates.

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Figure 4. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

Figure 5. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

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Figure 6. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

Following the sampling in the summer of 2013, 26 taxa of benthic macroinvertebrates were identified (Fig. 7). Chironomidae family and Asellus aquaticus were the most abundant and frequent, being found in all the three sampling sites. Chironomidae family and Asellus aquaticus, as long as Gammarus pulex, were eudominant in the lake. As dominant species, only Baetis spp. was classified into this category, the rest of the taxa being indicated as subrecedent organisms. The characteristic species of this season, regarding the index of ecological significance (Dzuba), were Gammarus pulex, Asellus aquaticus and the individuals of Chironomidae family. Eiseniella tetraedra, Helobdella stagnalis and Naucoris spp. were designated as accessory taxa and Corixa spp., Dytiscus spp., Ecnomus spp., Erpobdella octoculata, Gerris spp., Hydrochara spp., Hydrophilus spp., Hydrovatus spp., Limnephilus spp., Microvelia spp., Physa acuta, Valvata cristata as accidental ones. A total of 27 taxa were identified through the last sampling, in the autumn of 2013, indicated, as well as in the most of the past samplings, Gammarus pulex and Chironomidae family, the most abundant benthic macroinvertebrates (Fig. 8). These two taxa indicated also high frequency in the lake and based on the dominance index, there were the only eudominant organisms. Asellus aquaticus and Chaoborus flavicans were indicated as

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Figure 7. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

Figure 8. Benthic macroinvertebrates dinamics in the samples taken from Bădărău Lake.

- 42 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 dominant species, the rest of the organisms identified in the sampling falling into the recedent and subrecedent categories. According to Dzuba index of ecological significance, Asellus aquaticus, Gammarus pulex and Chironomidae family were the characteristic taxa in the lake, Baetis spp., Caenis spp., Ecnomus spp., accessory taxa and Notonecta spp., Onychogomphus spp., Simpetrum pro-parte, accidental benthic macroinvertebrates. The highest value of evenness index was recorded at Sampling Site 3 (Outflow) in the summer of 2012, the lowest values being recorded in Sampling Site 2 (Forest) in the winter of 2011 and spring of 2012 (Fig. 9). The highest values of the Shannon diversity index during the entire study period were recorded in the summer of 2012 at Sampling Site 3 (Outflow), in the spring and autumn of 2012 at Sampling Site 1 (Inflow) and in the autumn of 2013 at Sampling Site 2 (Forest). Lowest values were recorded in Sampling Site 2 (Forest) in the winter of 2011 and spring of 2012 (Fig. 10). According to physico-chemical analysis and the eight bioindicator species identified, water quality of Bădărău Lake is β- mesosaprobic to α-mesosaprobic (relatively clear, with an organic loading) (Table 1). Seasonal dinamics of benthic macroinvertebrates showed that Sampling Site 2 (Forest) tend to remain the same regarding the total abundance, indicating only insignificant changes and a minor increase in individuals total abundance through the seasonal sampling, from 2011 to 2013 (Fig. 11). In the case of Sampling Site 1 (Inflow) we observed a major increasing of total abundance from 2012 to 2013, being explained by the high number of Gammarus pulex individuals found in this sampling site at the end of the study.

Figure 9. Benthic macroinvertebrates seasonal dinamics regarding evenness index.

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Figure 10. Benthic macroinvertebrates seasonal dinamics regarding Shannon diversity index.

Table 1. Macroinvertebrates as water quality bioindicators in Bădărău Lake.

No. Species Water quality bioindicator 1 Asellus aquaticus β- and α-mesosaprobic 2 Baetis spp. oligo- and β- mesosaprobic 3 Coenagrion puella oligo- and β- mesosaprobic 4 Erpobdella octoculata β- and α-mesosaprobic 5 Gammarus pulex oligo- and β- mesosaprobic 6 Limnephilus spp. oligo- and β- mesosaprobic 7 Lymnaea spp. β- mesosaprobic 8 Tubifex tubifex poly- and α-mesosaprobic

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Figure 11. Benthic macroinvertebrates seasonal dinamics regarding the total abundance.

Conclusions This paper gives an overview of the seasonal dynamics of benthic macroinvertebrates in Lake Bădărău – Botanical Garden of Iasi. After the analysis of samples, we have reached the conclusion that a total number of 4911 specimens belonging to 61 species and groups of macroinvertebrates were found during the period of the biological survey. Gammarus pulex (1684 individuals), Chironomidae family (1471 individuals), Asellus aquaticus (225 individuals) and Baetis spp. (209 individuals) were the most abundant benthic macroinvertebrates. Correlating the abundance of macroinvertebrates with the sampling points, it is clear that Chironomidae family was the most abundant taxa in Sampling Site 1 (Outflow) and Gammarus pulex in Sampling Site 3 (Inflow). Sampling Site 2 had the higher biodiversity in comparison with the other two sampling sites. High values of the Shannon diversity index during the entire study period were recorded in the summer of 2012 at Sampling Site 3 (Outflow), in the spring and autumn of 2012 at Sampling Site 1 (Inflow) and in the autumn of 2013 at Sampling Site 2 (Forest). The highest value of evenness index was recorded at Sampling Site 3 (Outflow) in the summer of 2012. The lowest values of both Shannon diversity index and evenness index were recorded in Sampling Site 2 (Forest) in the winter of 2011 and spring of 2012.

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Sampling Site 2 (Forest) tend to remain the same regarding the total abundance, indicating only insignificant changes and a minor increase in individuals total abundance through the seasonal sampling, from 2011 to 2013. We observed a major increasing of total abundance at Sampling Site 1 (Inflow) from 2012 to 2013, being explained by the high number of Gammarus pulex individuals found in this sampling site at the end of the study. Overall, Bădărău Lake is an ecosystem with high diversity in terms of taxa, as well as expressed by the number of specimens collected.

Acknowledgments This work was supported by the strategic grant POSDRU/159/1.5/S/133652, co- financed by the European Social Fund within the Sectorial Operational Program Human Resources Development 2007 – 2013.

References Magurran, A. E., 1988. Ecological Diversity and its Measurement. London, Croom Helm. Mustaţă, G., 2000. Hidrobiologie, Editura Universității „Al.I. Cuza” Iaşi. Nicoară M., 2008. Biodiversitatea mediilor acvatice. Editura Pim, Iași. Plavan, G., Nicoară, M., 2012. Studies on the distribution and biomass of the Oligochaetes (Annelida) from the Izvoru Muntelui Bicaz reservoir. Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, Biologie animală, LVIII: 7-12. Surugiu, V., 2008. Limnologie și saprobiologie. Compendiu de lucrări practice. Editura Tehnopress, Iași.

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EPIGENOUS FAUNA (ARTHROPODA: ARANEAE, INSECTA) IN THE RYE CROP FROM HORODNIC DE JOS (SUCEAVA COUNTY, ROMANIA)

Elena-Daniela PRELIPCEAN (BOSOVICI), Mircea VARVARA and Ioan MOGLAN Faculty of Biology, “Alexandru Ioan Cuza” University of Iași, Carol I Bvd., no. 20A, 700505 Iași, Romania, [email protected], [email protected], [email protected]

Abstract. The enthomofaunal material samples were made with the help of some soil pitfalls in the period April- August 2014 in a rye crop from Horodnic de Jos village, Suceava County, Romania. In total, there were collected 504 arthropoda specimens that belonged to the following classes: Arachnida, Miriapoda and Insecta. The Insects were net dominant that possessed 98% from the total of the collected individuals. Regarding to what concerns the insects, there were identified six orders, the dominant ones Coleoptera (59.9%), followed by Hymenoptera (34.0%). There were identified 11 families of Coleoptera, the most individuals belonging to the family of Carabidae (51.7%), followed by . Referring to Carabidae, there were identified 12 species, the most abundance being observed in Pseudoophonus rufipes species (41.3%).

Keywords: rye crop, soil pitfalls, epigenous enthomofauna, carabidae, Suceava County, Romania.

Rezumat. Fauna epigee (Arthropoda:Araneae, Insecta) din cultura de secară din localitatea Horodnic de Jos, Județul Suceava, România. Colectările de material entomofaunistic s-au făcut cu capcane de sol în perioada aprilie-august 2014 dintr-o cultură de secară din localitatea Horodnic de Jos, județul Suceava, România. În total s- au colectat 504 exemplare de artropode, din clasele Arachnida, Miriapoda și Insecta. Net dominante au fost Insectele care au deținut 98% din totalul indivizilor colectați. În ce privește insectele au fost identificate șase ordine, dominante fiind Coleopterele (59,9%), urmate de Hymenoptere (34,0%). Au fost identificate 11 familii de Coleoptere, cei mai mulți indivizi au aparținut familiei Carabidae (51,7%), urmată de Dermestidae. Referitor la Carabidae, au fost identificate 12 specii, abundența cea mai mare s-a constatat la specia Pseudoophonus rufipes (41,3%).

Cuvinte cheie: cultura de secară, capcane de sol, entomofaună epigee, carabide, județul Suceava, România.

Introduction The rye represents a straw cereal species that in Romania it’s produced mainly in the North area of the country. According to its importance for Suceava County economy, we set up to investigate the pests related to this crop but also the auxiliary enthomofauna. In the current paper we present the epigenous enthomofauna observed from a rye crop from Horodnic de Jos village, in Suceava County.

Material and Methods The epigenous fauna’s collecting was made with the help of 12 soil traps (Barber pitfalls), placed on a 5 m line distance between one another. The pitfalls were placed in the soil on 16 of April 2014, the material was collected from two in two weeks, until the very of August, including when the crop was collected. In all, there were made 8 samples. There was 4% formol in the pitfalls; the collecting was made with a mesh strainer. The collected material was placed in glass vessels, tagged and identified at the level of class, order, family and species (Radu & Radu, 1967; Lăcătușu & Ionescu, 1971; Lăcătușu et al., 1974;

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Pisică et al., 2002). The primary data of Carabidae were registered and statistically analysed, calculating the abundance, the constancy, the dominance and the ecological significance index (Turculeț & Varvara, 2006).

Results and Discussion In total, in the period of April-August there were collected 504 individuals of Arthropoda that belonged to 3 classes: Arachnida, Miriapoda and Insecta. The insects were net dominant that possessed 98.0% from the total individuals of the collected Arthropoda (Table 1). Referring to insects, in total, there were collected 494 individuals and were identified six orders: Orthoptera, Dermaptera, Heteroptera, Hymenoptera, Coleoptera and Diptera. The most insect specimens belonged to the order Coleoptera which possessed 59.9% from the total of insect individuals, followed by Hymenoptera (34%). The other orders were less numbering represented (Table 2). Hymenoptera were represented only by the family Formicidae and Orthoptera only by Gryllus campestris. Regarding Coleoptera there were collected 296 individuals which belonged to 11 families: Carabidae, Dermestidae, Elateridae, Meloidae, Tenebrionidae, Curculionidae, Silphidae, Scarabaeidae, Coccinellidae, Staphylinidae, Chrysomelidae. The most individuals belonged to the family of Carabidae, which possessed 51.7% from the total collected individuals of Coleoptera, followed by the families Dermestidae (31.4%) and Tenebrionidae (6.4%). The rest of Coleoptera families were less numbering represented (Table 3). Although the abundance of Dermestides has a high value, the diversity of this family is low, being represented by only one species ( laniarius Illiger). Referring to Carabidae, there were collected 160 individuals and were identified 12 species: Carabus cancellatus Illiger, Carabus scabriusculus Ol., Anisodactylus signatus Panzer, Pseudoophonus rufipes De Geer, Harpalus distinguendus Duftschmid, Pterostichus melanarius Illiger, Pterostichus strenuus Linne, Poecilus cupreus Linne, Metophonus punctatulus Duftschmid, Bembidion lampros Hrbst., Brachinus explodens Duftschmid and Brachinus crepitans. The biggest abundance has been observed in Pseudophonus rufipes which possessed 41.3% from the total Carabidae followed by Poecilus cupreus (22.5%), Carabus cancelathus (17.8%), Pterostichus melanarius (6.3%) and Anisodactylus signatus (5.6%). The rest of the species are represented by fewer individuals (Table 4). The Carabidae have presented the maximum of activity in the month of June and at the beginning of July. The rest of the Coleoptera families were represented only by one species, excepting the family of Silphidae which is represented by two species. In the matter of numbers, only Dermestidele and Tenebrionidae possess more individuals (Table 5). The statistic calculation applied to Carabidae, regarding the constancy of the obtained data it reveals that species Pseudoophonus rufipes and Poecilus cupreus are euconstant, Pterostichus melanarius is constant and the rest of the species are accessories and accidental. Regarding dominance, the species Pseudoophonus rufipes, Poecilus cupreus and Carabus cancellatus are eudominant, Pterostichus melanarius and Anisodactylus signatus are dominant and the rest are recedent and subrecedent (Table 6). The ecological significance index reveals that the species Pseudoophonus rufipes and Poecilus cupres are the best adapted to the ecological conditions from this place and are characteristic.

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The data from the specific literature referring to this type of enthomofauna for this particular cultured plant species are few. We’ve compared our results with the ones that Alexandru Dascalu obtained (in manuscript), from Zvoriștea village, Suceava County, in the year 1993, observing that there aren’t significant differences regarding the quality aspects, but they differ from the quantity point of view. So much so, referring to families of Coleoptera, the author has identified 12 families, the most abundance presenting the Carabidae family, but the number of Coleoptera individuals were a lot higher that in our case (2817 individuals among which 2698 individuals belonged to Carabidae). We consider that this big difference it owns in the first place to the fact that at Horodnic de Jos in the summer of 2014 and the beginning of 2014’s summer there was frequently raining, low temperatures and soil differences and agricultural techniques.

Table 1. Abundance (A) and dominance (D) of Arthropoda collected using soil pitfalls, in the rye crop from Horodnic de Jos, Suceava County.

Collecting dates in 2014

29.04 12.05 30.05 12.06 30.06 12.07 29.07 14.08 Total Taxon A D A D A D A D A D A D A D A D A D Order Araneae 1 3.1 2 4.2 ------1 0.9 3 4.8 - - 7 1.4 Class Miriapoda ------2 3.1 1 2.4 3 0.6 Class Insecta 31 96.9 46 95.8 25 100 91 100 87 100 112 99.1 59 92.2 43 97.7 494 98 Total 32 - 48 - 25 - 91 - 87 - 113 - 64 - 44 - 504 -

Table 2. Abundance (A) and dominance (D) of collected insects using soil pitfalls, in the rye crop from Horodnic de Jos village, Suceava County.

Collecting dates in 2014

29.04 12.05 30.05 12.06 30.06 12.07 29.07 14.08 Total Order A D A D A D A D A D A D A D A D A D Orthoptera 4 12.9 8 - 2 8.0 1 1.1 1 1.2 3 2.7 1 1.7 2 4.7 22 4.5 Dermaptera ------2 4.7 2 0.4 Heteroptera ------1 1.1 - - - - 2 3.4 1 2.3 4 0.8 Hymenoptera ------42 46.2 49 56.3 32 28.6 28 47.5 17 39.5 168 34.0 Coleoptera 27 87.1 38 - 23 92.0 47 51.1 37 42.5 76 67.9 27 45.8 21 48.8 296 59.9 Diptera ------1 - 1 - - - 2 0.4 Total 31 - 46 - 25 - 91 - 87 - 112 - 59 - 43 - 494 -

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Table 3. Abundance (A) and dominance (D) of Coleoptera collected using soil pitfalls, in the rye crop from Horodnic de Jos village, Suceava County.

Collecting dates in 2014 Total 29.04 12.05 30.05 13.06 30.06 12.07 29.07 19.08 Family A D A D A D A D A D A D A D A D A D Carabidae 8 29.6 7 18.4 2 8.7 22 46.8 29 78.4 60 78.9 12 44. 13 60.9 153 51.7 Dermestidae 16 59.3 15 39.5 18 78.3 24 51.1 4 10.8 8 10.5 5 18.5 3 14.3 93 31.4 Elateridae 3 11.1 4 10.5 ------3 4.0 - - 1 4.8 11 3.7 Meloidae - - 1 2.6 ------1 0.3 Tenebrionidae - - 10 26.3 3 13.0 - - 2 5.4 3 4.0 - - 1 4.8 19 6.4 Curculionidae - - 1 2.6 ------2 7.4 - - 3 1.0 Silphidae ------1 2.1 1 2.7 - - 1 3.7 - - 3 1.0 Scarabaeidae ------1 2.7 1 1.3 3 11.1 - - 5 1.7 Coccinellidae ------2 7.4 1 4.8 3 1.0 Staphylinidae ------1 1.3 - - 1 4.8 2 0.7 Chrysomelidae ------2 - 1 4.8 3 1.0 Total 27 - 38 - 23 - 47 - 37 - 76 - 27 - 21 - 296 -

Table 4. Abundance (A) and dominance (D) of Carabidae species collected using soil pitfalls, in the rye crop from Horodnic de Jos village, Suceava County.

Collecting dates in 2014 Total Species 29.04 12.05 30.05 12.06 30.06 12.07 29.07 14.08 A D A D A D A D A D A D A D A D A D Pseudoophonus rufipes 4 50.0 1 14.3 1 50.0 7 24.1 14 48.3 31 51.7 5 41.7 3 23.1 66 41.3 Poecilus cupreus 1 12.5 6 85.7 1 50 3 10.4 - - 18 30 3 25.1 4 30.8 36 22.5 Carabus cancelathus 1 12.5 - - - - 14 48.3 11 37.9 - - 1 8.3 - - 27 17.8 Pterostichus melanarius ------3 10.4 2 6.9 3 5.0 1 8.3 1 7.7 10 6.3 Anisodactylus signatus ------1 3.5 - - 6 10.0 - - 2 15.4 9 5.6 Brachinus crepitans ------1 3.5 - - - - 2 15.4 3 1.9 Bembidion lampros ------2 16.6 1 7.7 3 1.9 Carabus scabriusculus ------2 3.3 - - - - 2 1.3 Metophonus punctatulus ------1 3.5 ------1 0.6 Harpalus distinguendus 1 12.5 ------1 0.6 Pterostichus strenuus 1 12.5 ------1 0.6 Brachinus explodens ------1 3.5 ------1 0.6 Total 8 - 7 - 2 - 29 - 29 - 60 - 12 - 13 - 160

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Table 5. Silphidae, Scarabeidae, Dermestidae, Elateridae, Tenebrionidae and Meloidae collected with soil pitfalls, in the rye crop from Horodnic de Jos, Suceava County.

Collecting dates in 2014 Family: species 29.04 12.05 30.05 12.06 30.06 12.07 29.07 14.08 Total Silphidae: Necrophorus vespillo - - - - 1 2 - - 3 Silphidae: Phosphuga atrata - - - 1 - - - - 1 Scarabaeidae: Onthophagus sp. - - - - 1 1 3 - 5 Dermestidae: Dermestes laniarius 16 15 18 24 4 8 5 3 93 Elateridae: Agriostes lineatus 3 4 - - - - - 2 9 Tenebrionidae: Opatrum sabulosum - 10 3 - 2 3 - 1 19 Meloidae: Meloe proscarabaeus - 1 ------1 Coccinellidae: Coccinella 7.punctata ------2 1 3 Total 19 30 21 25 8 14 10 7 134

Table 6. Abundance (A), constancy (C), dominance (D) and ecological significance index (W) of Carabidae species, in the rye crop from Horodnic de Jos, Suceava County.

C D W A Species % % % Pseudoophonus rufipes 66 100 41.3 41.3 Poecilus cupreus 36 87.5 22.5 19.6 Carabus cancellatus 27 50.0 17.8 8.9 Pterostichus melanarius 10 62.5 6.3 3.9 Anisodactylus signatus 9 37.5 5.6 2.1 Brachinus crepitans 3 25.0 1.9 0.5 Bembidion lampros 3 25.0 1.9 0.5 Carabus scabriusculus 2 12.5 1.3 0.2 Harpalus distinguendus 1 12.5 0.6 0.08 Pterostichus strenuus 1 12.5 0.6 0.08 Metophonus punctatulus 1 12.5 0.6 0.08 Brachinus explondens 1 12.5 0.6 0.08 Total 160 - - -

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Conclusions From the rye crop from Horodnic de Jos village, in 2014’s year conditions, with the help of soil pitfalls there were collected in total 504 Arthropoda specimens, who belonged to three classes: Arachnida, Miriapoda and Insecta. Insects were net dominant. At the level of order, the collected insects belonged to six orders, dominant being Coleoptera. Although, Hymenoptera were also well numbering represented, the diversity was low, only the family of Formicidae. Coleoptera were represented by 11 families. Most individuals belonged to family Carabidae followed by the family Dermestidae, but, the last one, even with high abundance, was represented only by a single species, Dermestes laniarius. Referring to Carabidae, there were identified 12 species: Carabus cancellatus Illiger, Carabus scabriusculus Ol., Anisodactylus signatus Panzer, Pseudoophonus rufipes De Geer, Harpalus distinguendus Duftschmid, Pterostichus melanarius Illiger, Pterostichus strenuus Linne, Poecilus cupreus Linne, Metophonus punctatulus Duftschmid, Bembidion lampros Hrbst., Brachinus explodens Duftschmid and Brachinus crepitans Linne. The most individuals belonged only to 4 species: Pseudoophonus rufipes De Geer, Poecilus cupreus Linne, Carabus cancellatus Illiger, Pterostichus melanarius Illiger. The statistic calculation applied to Carabidae reveals that Pseudoophonus rufipes and Poecilus cupreus are characteristic for the investigated crop from Horodnic de Jos and the most well adapted to this place’s ecological conditions.

References Lăcătușu, M., Ionescu, A., 1971. Entomologie. Editura Didactică și Pedagogică, București. Lăcătușu, M., Tudor, C., Teodorescu, I., 1974. Lucrări practice de entomologie. Editura Universității București. Radu, G., Radu, V., 1967. Zoologia nevertebratelor, 2. Editura Didactică și Pedagogică, București. Pisică, C., Moglan, I., Cojocaru, I., 2002. Zoologia nevertebratelor (Lucrări practice de laborator, vol. II), Editura Universității „Alexandru Ioan Cuza” Iași. Turculeț, A., Varvara, M., 2006. Diversity and some Ecological Aspects of the Species of Carabidae (Coleoptera, Carabidae) in the Crop Ecosystem from the North of Moldavia (Suceava and Botoșani Counties). Studii și Cercetări, Universitatea Bacău: 257-267.

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FREQUENCY OF SWINE (SUS DOMESTICUS AND SUS SCROFA) IN THE BRONZE AGE SETTLEMENTS ON THE TERRITORY OF ROMANIA

Mariana POPOVICI* and Simina STANC “Alexandru Ioan Cuza” University, Faculty of Biology, Arheoinvest, Bd. Carol I, 20A, 700505, Iaşi, Romania, *[email protected]

Abstract. This study is a summary of swine records in Bronze Age archaeozoological samples from Romania, underling the importance of pig (Sus domesticus) and wild boar (Sus scrofa) in the diet of human populations in this period. The synchronic and diachronic relationships of pig and wild boar frequencies (reported to the total identified mammals) in the analyzed archaeozoological assemblages were established using the Multiple Correspondence Analysis. The mammal remains are predominant in the archaeozoological samples (compared with other faunal groups, as bird and fish); the importance of pig in the diet of human population increases in Bronze Age time, compared with previous periods. In the Chalcolithic-Bronze Age Transition pig represent in average 8.8% of all identified mammals, while in the Late Bronze Age an increase up to 24% was observed. In the case of wild boar, the tendency is a decrease in frequency from the Chalcolithic-Bronze Age Transition (8.7% of the identified mammals) to the Late Bronze Age (1%).

Keywords: Bronze Age, swine, economic importance, Romania.

Rezumat. Frecvența suinelor (Sus domesticus și Sus scrofa) în așezările de Epoca Bronzului de pe teritoriul României. Lucrarea reprezintă o sinteză a datelor privind suinele identificate în eșantioane arheozoologice, aparținând Epocii Bronzului de pe teritoriul României, și evidențiată importanța porcului domestic (Sus domesticus) și a mistrețului (Sus scrofa) în dieta populațiilor umane din această perioadă. Relațiile sincronice și diacronice privind frecvențele porcului domestic și ale mistrețului (raportate la numărul total al resturilor de mamifere identificate), în eșantioanele arheozoologice luate în studiu, au fost stabilite utilizând Analiza Corespondenței Multiple. Resturile provenind de la mamifere sunt predominante în toate eșantioanele arheozoologice (comparative cu celelalte grupe faunistice, cum sunt păsările și peștii); importanța porcului în dieta populațiilor umane crește de-a lungul Epocii Bronzului, comparative cu perioada anterioară. În perioada de tranziție Calcolitic-Epoca Bronzului, resturile de porc domestic au o pondere în medie de 8,8% din totalul resturilor de mamifere identificate, în timp ce în Epoca Bronzului s-a observat o creștere până la 24%. În cazul mistrețului a fost evidențiată o reducere a ponderii acestei specii – în Epoca Bronzului (1%) față de perioada de tranziție Calcolitic-Epoca Bronzului (8,7% din totalul resturilor de mamifere identificate).

Cuvinte cheie: Epoca Bonzului, suine, importanță economică, România.

Introduction This study concerns the Bronze Age cultures (including also the transition Neolithic-Bronze Age), which occupied the territory of present day Romania. The study is focused on subsistence as reflected by archaeozoological analyses. Previous archaeozoological papers concerning the Bronze Age period discussed the fauna discovered in different parts of the country (Haimovici, 1965; El Susi, 1996; Bindea, 2008). Our study is focused on the importance of pig and wild boar in the economy of human population in Bronze Age settlements on the Romanian territory. The studied assemblages were grouped according to following periods: Chalcolithic-Bronze Age Transition (3800-3500 BC), Early Bronze Age (3500-2200 BC), Middle Bronze Age

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(2200-1600/1500 BC) and Late Bronze Age (1600/1500-1100 BC). Also the settlements were grouped according to the regions of Romania: Moldavia, Banat, Wallachia, Transylvania and Dobrudja. Among the animal resources, domestic mammals constitute the majority. Animal husbandry was an important subsistence activity during Bronze Age in Romania, but wild mammal remains are still present, even if only in smaller amounts. The proportions of pig and wild boar remains in the archaeozoological samples have both regional and diachronical variations.

Material and Methods The data used in this synthesis comes from previous studies, as follows: Haimovici, 1962; 1965; 1966; 1970; 1972; 1974; 1980; 1991; 1994; 1997; 2006; El Susi, 1996; Perianu & Udrescu, 1990; Bindea, 2008. These papers had analyzed bone remains from following settlements: Chalcolithic-Bronze Age Transition: Banat (Bocşa-Colţan assemblage), Dobrudja (Cernavodă assemblage), Moldavia (assemblages: Folteşti, Stoicani, Horodiştea, Erbiceni); Early Bronze Age: Banat (assemblages: Foeni “Cimitirul Ortodox”, Moldova Veche Ostrov), Wallachia (assemblages: Glina, Militari-Câmpul Boja, Căscioarele), Transylvania (assemblages: Zoltan, Livezile, Hîrman, Şincai, Tărtăria, Ghida, Cicău, Ţebea, Boiu); Middle Bronze Age: Banat (assemblages: Ostrovu Corbului, Gornea- Păzărişte, Rogova, Foeni), Moldavia (assemblages: Sărata Monteoru, Bogdăneşti, Mîndrişca, Monteoru), Wallachia (assemblages: Popeşti, Militari-Câmpul Boja, Verbiţa), Transylvania (assemblages: Sîntion, Pecica, Derşida, Mintiu Gherlei, Carei, Sălacea, Săcueni, Medieşu Aurit Potău, Otomani), Late Bronze Age: Moldavia (assemblages: Valea Lupului, Truşeşti, Cavadineşti, Poşta Elan, Bîrlad, Piatra Neamţ, Gîrbovăţ), Transylvania (assemblages: Iclod, Zoltan). Swine remains are described in terms of frequency – number of identified specimens (NISP) and minimum number of individuals (MNI). The synchronic and diachronic relationships of pig and wild boar frequencies were established using the Multiple Correspondence Analysis (Greenacre, 2002). For statistical analysis, the software XLStat vers. 2012 were used.

Results and Discussion The faunal material collected during the archaeological excavations included 48919 mammal skeletal remains. Remains of cattle, sheep and goats dominate in samples numerically, with values over 70% of all identified mammal bones. The highest frequencies of swine bones are recorded in settlements of Middle Bronze Age (22.59% of all mammalian identified fragments) and Early Bronze Age (16.19%) (Fig. 1). In the swine group, pig was recorded in large proportion in Middle Bronze Age assemblages (Banat: 18.94%; Wallachia: 22.25%; Transylvania: 19.33%) and Late Bronze Age assemblages (Transylvania: 24.02%) (Fig. 2). This aspect is underlined in graphical illustration of ratio index (RI); differences between pig and wild boar frequencies of remains were observed in the assemblages from Moldavia (in Late Bronze Age, RI=17.62), Wallachia (in Early Bronze Age, RI=21.88), Transylvania (in Late Bronze Age, RI=24.05 (Fig. 3). The highest proportions for wild boar are in the samples of Banat region: 8.11% (in Chalcolithic-Bronze Age Transition assemblages), 8.12% (in Middle Bronze Age) and

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4.75% (in Early Bronze Age assemblages) (Fig. 2).

84.43% Late Bronze Age 15.57%

77.41% Middle Bronze Age 22.59%

83.81% Early Bronze Age 16.19%

Chalcolithic - Bronze 88.21% Age Transition 11.79%

other mammals % swine

Figure 1. Swine proportions (% NISP) in Bronze Age, estimated from the total mammal remains.

30.00 25.00 % pig %wild boar Linear (% pig) 20.00 15.00 10.00 5.00

0.00

Late BronzeAge

Late BronzeAge

Early BronzeAge Early BronzeAge Early BronzeAge

Transition Transition Transition

Middle BronzeAge Middle BronzeAge Middle BronzeAge Middle BronzeAge

Chalcolithic - BronzeAge Chalcolithic - BronzeAge Chalcolithic - BronzeAge Banat Dobrudja Moldavia Wallachia Transylvania

Figure 2. Swine proportions (% NISP) in the Bronze Age assemblages from Romanian territory, estimated from the total mammal remains.

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25.00 24.05 21.88 20.00 17.62 15.00 10.00 10.30 8.33 9.18 5.00 3.54 2.33 4.00 5.20 2.53

0.00 1.07

Late BronzeAge Late BronzeAge

Early BronzeAge Early BronzeAge Early BronzeAge

AgeTransition AgeTransition AgeTransition

Middle BronzeAge Middle BronzeAge Middle BronzeAge Middle BronzeAge

Chalcolithic - Bronze Chalcolithic - Bronze Chalcolithic - Bronze Banat Dobrudja Moldavia Wallachia Transylvania

Figure 3. Ratio Index of swine in the Bronze Age settlements, in Romania.

Multiple correspondence analysis based on the ratio pig/wild boar in archaeozoological assemblages reveals various associations between the studied variables: regions of Romania, ratio index and periods of Bronze Age. The results are shown in Figure 4, and the explanation is based on their position along the two main axes which represent

2 Asymmetric variable plot (axes F2 and F3: 28.18 %)

Banat EBA

1 Wallachia MBA Wallachia EBA Transylvania EBA Banat MBA Moldavia C-BA Tr Transylvania LBA

0

Transylvania MBA Moldavia MBA Moldavia LBA

Banat C-BA Tr F3(9.09 %) -1

-2

Dobrudja C-BA Tr

-3 F2 (19.09 %) -3 -2 -1 0 1 2 3

Figure 4. Multiple Corespondence Analysis (C-BA Tr – Chalcolithic-Bronze Age Transition, EBA – Early Bonze Age, MBA – Middle Bronze Age, LBA – Late Bronze Age).

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28.9% of the total variance (Axis 2-19.9% and axis 3-9%). The best dispersion of variables on the map were obtained for the axes 2 and 3. Axis 2 marks out the relationship between variables: Wallachia, Moldavia and Transylvania, characterized by a much higher pig proportion in oposite with the other regions and periods of the Bronze Age (Fig. 4). Axis 3 highlights the significant proportion of wild boar in Chalcolithic-Bronze Age Transition assemblages in opposition with the other superiods of Bronze Age.

Conclusions Among the animal resources, the mammals are predominant in archaeological samples. Besides cattle and sheep/goat, pig had an important role in the diet of human population in Bronze Age, recording an increase from the beginning toward the late Bronze Age. In the Chalcolithic-Bronze Age Transition pig represent about 8.86% of all identified mammals, while in the Late Bronze Age an increase up to 24.02% was observed. In the case of wild boar, the tendency is a decrease in frequency from the Chalcolithic-Bronze Age transition (8.7% of the identified mammals) to the Late Bronze Age (1%).

Acknowledgements This study was supported by the Romanian research program CNCS - UEFISCDI PN-II-RU-TE-2011-3-0146.

References Bindea, D., 2008. Arheozoologia Transilvaniei în pre- și protoistorie. Ed. Teognost, Cluj Napoca. El Susi, G., 1996. Vânători, pescari și crescători de animale în Banatul mileniilor VI i.Ch.-I d.Ch.. Ed. Mirton, Timișoara. Greenacre, M., 2002. The use Correspondence Analysis in the Exploration of Health Survey Data. Fundacion BBVA. Haimovici, S., 1962. Studiul comparativ al resturilor faunistice din epoca neolitică și cea a bronzului de la Valea Lupului. Analele Știintifice ale Universității „Alexandru Ioan Cuza” Iași, Științele naturii, 8(2): 291- 326. Haimovici, S., 1965. Studiul particularităților morfologice ale scheletului unor animale domestice și sălbatice descoperite în stațiunile Epocii Bronzului din România (Studiul paleofaunei din Epoca Bronzului). Teza de doctorat, Universitatea "Al. I. Cuza" Iași. Haimovici, S., 1966. Studiul materialului descoperit în așezarea din Epoca Bronzului (cultura Monteoru) de la Bogdanești. Arheologia Moldovei, IV: 119-136. Haimovici, S., 1970. Studiul faunei subfosile descoperită în așezarea de la Erbiceni (perioada de trecere de la Neolitic la Epoca Bronzului). Analele Știintifice ale Universității “Alexandru Ioan Cuza” Iași, s. Biologie, 16(1): 169-179. Haimovici, S., 1972. Studiul resturilor faunistice provenite din așezarea aparținând perioadei de trecere de la Neolitic la Epoca Bronzului de la Foltești. Arheologia Moldovei, 7: 97-102. Haimovici, S., 1974. La faune sous-fossile découverte dans la station éponyme de la civilisation Foltesti. Dacia, 18: 73-77. Haimovici, S., 1980. Studiul materialului faunistic din asezarea de la Mîndrișca (Valea Seaca) aparținând culturii Monteoru. Carpica, XII: 191-201. Haimovici, S., 1991. Materialul faunistic de la Gîrbovaț. Studiu arheozoologic. Arheologia Moldovei, 14: 153-166. Haimovici, S., 1994. Studiul unui lot de paleofauna provenit din așezarea eponimă a culturii Monteoru. Arheologia Moldovei, 17: 309-319. Haimovici, S., 1997. Studiul arheozoologic al unui lot de fauna descoperit în așezarea eponimă de la Glina. Thraco-Dacica, 18(1-2): 231-238. Haimovici, S., 2006. Studiul materialului faunistic descoperit în situl de cultură Noua de la Poșta Elan (jud.Vaslui), corelat cu cel găsit în alte situri. Arheologia Moldovei, XXIX: 223-234.

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Perianu, M., Udrescu, M., 1990. Studiu antropologic și arheozoologic al materialului osteologic din mormântul de la Căscioarele (jud. Călărași), Epoca Bronzului (Cultura Glina III). Cultură și civilizație la Dunărea de Jos, 5-7: 55-64.

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ANTHROPOLOGICAL ANALYSIS OF A SKELETON BELONGING TO MIDDLE BRONZE AGE, KOMARIV CULTURE, DISCOVERED IN SUCEAVA-CÂMPUL ŞANŢURILOR-STRADA PARCULUI (SUCEAVA COUNTY, ROMANIA)

Angela SIMALCSIK, Vasilica Monica GROZA and Robert Daniel SIMALCSIK Romanian Academy – Iași Branch, Department of Anthropological Research, Th. Codrescu Street, No. 2, Iași, Romania, [email protected]

Abstract. This paper concerns the anthropological analysis of the skeleton discovered in 2007 in the stone tomb (cista) in Suceava-Câmpul Şanţurilor-strada Parcului (Suceava County, Romania). The funerary monument was chronologically ranked in the Middle Bronze Age, the Komariv culture. The osteological remains discovered in Suceava-Câmpul Şanţurilor-strada Parcului belonged to a male, aged approximately 20-25 at death. The cranial index is dolichocranic. The frontal-parietal index is stenometopic. The occipital-parietal index is large-sized. The cranial bone relief is marked. The nasal region seems to have been narrow and quite high. The mandible is moderately robust, with medium robustness index. The postcranial skeleton is robust, clearly indented, with pronounced muscle insertions. The humeri record a euribrachic cross-section index. The femora are hyperplatymeric, with a prominent linea aspera and moderate pilaster. The tibiae show a mesocnemic cross- section index. The stature was appreciated only using the morphoscopic method and seems to be upper-medium to high-sized. The typological analysis shows predominantly Nordic characteristics in admixture with some Dinaric elements. The dentition suggests an excellent status of the dental health. This skeleton does not present any severe bone pathology. Some postcranial bones have specific traits which could be suggest the muscular massiveness and a series of not very stressful daily activities. The skeleton remains from the inventory do not present any signs of any possible ante mortem or peri mortem traumas.

Keywords: Suceava-Câmpul Şanţurilor-strada Parcului, Middle Bronze Age, Komariv (Komarow) culture, anthropological analysis

Rezumat. Analiza antropologică a unui schelet aparținând Epocii Bronzului Mijlociu, cultura Komariv), descoperit la Suceava-Câmpul Şanţurilor-strada Parcului (județul Suceava, România). Lucrarea de faţă prezintă analiza antropologică a scheletului descoperit în 2007 în cutia de piatră (cista) în punctul Suceava-Câmpul Şanţurilor-strada Parcului (judeţul Suceava, România). Monumentul funerar a fost încadrat chronologic în perioada mijlocie a Epocii Bronzului, cultura Komariv. Scheletul descoperit în Suceava-Câmpul Şanţurilor-strada Parcului aparţin unui individ de sex masculin cu vârsta la deces de circa 20-25 ani. Indicele cefalic este dolicocran. Indicele fronto-parietal este stenometop. Indicele occipito-parietal indică un occipital larg. Relieful cranian este evidenţiat. Regiunea nazală pare să fi fost îngustă şi destul de înaltă. Mandibula este moderat de robustă, indicele de robusticitate fiind unul de tip mijlociu. Scheletul postcranian este robust, reliefat, cu inserţii musculare accentuate. Humerusurile înregistrează un indice de secţiune euribrahic. Femurele sunt hiperplatimere, cu linea aspera proeminentă şi cu pilastru moderat. Tibiile prezintă un indice de secţiune mesocnemic. Statura, apreciată doar prin metodele morfoscopice, pare să fi fost supramijlocie spre mare. Analiza tipologică evidenţiază predominanţa caracterelor nordice în amestec cu unele elemente dinarice. Analiza dentiţiei ne sugerează o stare de sănătate dentară excelentă. Acest schelet nu prezintă nici o patologie osoasă severă. Unele oase postcraniene prezintă anumite caracteristici care ne pot sugera masivitatea musculară a acestui individ şi o serie de activităţi cotidiene nu prea solicitante. Pe resturile scheletice prezente în inventar nu am semnalat semne ale unor posibile traumatisme produse ante mortem sau peri mortem.

Cuvinte Cheie: Suceava-Câmpul Şanţurilor-strada Parcului, perioada mijlocie a Epocii Bronzului, cultura Komariv (Komarow), analiza antropologică

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Introduction During the diggings undertaken for the set out of a new dwelling foundation in 2007, in the town of Suceava, on the Eastern side of the archaeological site perimeter called Câmpul Şanţurilor, there was identified an anthropic deposit (construction) of stones which proved to be a funerary monument. The authors of the archaeological excavation were chronologically ranked in the Middle Bronze Age, Komariv (Komarow) culture (approximately 2200/2000-1600/1500 B.C.). “Komarow” is the old name for this culture in Polish. The term “Komariw” is used in this study according to the practice nowadays, this village was included in the Ukrain territory after the second world war. The place where the funerary monument was discovered is situated at about 300 m south-east of the Cetatea de Scaun, on the edge of the archaeological site called Câmpul Şanţurilor, 30 m away from the foundation of a church from the 15th century and its graveyard, at approximately 300 m to the west of the high terrace of Suceava River. The depth at which the stone building was discovered is 0.30-0.35 m. On the eastern side of the funerary building, at 0.40-0.45 m deep, but also in the tomb filling, inside the stone building there were discovered pottery shards from the Middle Bronze Age, namely the Komariv (Komarow) culture, which actually helped to make the chronological ranking of the discovery (Mareş, 2010). The funerary building (stone box or cista) is heading north-east – south-west, it is rectangular, rounded in the edges as it is made of massive Sarmatian lime, built horizontally and vertically, while the north-eastern edge is rounded (Fig. 1) (Mareş, 2010). Inside the stone building, 0.75 m deep from the present-day level, stretching on fertile soil, there was discovered a human skeleton, in crouching position on the right, in a south-west – north-east line, head to south-west and feet to north-east, looking to the east, left hand bent from the elbow lying on the trunk and the right hand straight next to the body (Fig. 2). Next to the skull (in the orbit area), at 10 cm distance there was found a hammer axe made of stone, ritually broken, still showing a small part of the hole for the handle (Mareş, 2010).

Figure 1. Suceava-Câmpul Şanţurilor-strada Parcului: the funerary building (stone box or cista) (Mareş, 2010).

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Figure 2. Suceava-Câmpul Şanţurilor-strada Parcului: the funerary building and the skeleton in situ (Mareş, 2010).

Material and Methods The preservation status of the skeleton found in Suceava-Câmpul Şanţurilor- strada Parcului is satisfactory, which made it possible to a detailed anthropological analysis, even if sometimes incomplete. The anthropological study began with the cleaning (by dry method) and restoration of the osteological remains, after which morphoscopic examination and collection of biometric data followed. Determination of sex and age at death was followed by conformative and morphoscopic analysis, typological analysis and investigation of the possible pathologies, anomalies and epigenetic characters. The determination of the sex and the estimation of the age at the time of death was done using the methods and techniques recommended by Périer (1935, 1949), Nemeskéri et al. (1960), Iordanidis (1961), Stradalova (1975), Ubelaker (1979), Ferembach et al. (1979), Brothwell (1981), Smith & Knight (1984), Buikstra & Ubelaker (1994), Mays (1998), Bruzek (2002), Walrate et al. (2004), White & Folkens (2005), Schmitt (2005), Latham & Finnegan (2010), and Blanchard (2010). In the anthropological analysis we used the main anthropometric measurements and some conformative and morphoscopic characteristics established from the values of direct measurements, and also from the conformation indices, by classical techniques recommended by Martin & Saller (1956-1966). Evaluation and categorization of the absolute and relative values made use of the dimorphic scales of Alexeev & Debetz (1964). The morphological observations were registered and analyzed with the methods

- 61 - Angela Simalcsik et al. recommended by Broca (1875), Eickstedt (1934) and Olivier (1969). The stature was estimated from the dimensions of the long bones of the upper (humerus, radius, and ulna) and lower (femur, tibia, and fibula) limbs. The dimensional scales proposed by Manouvrier (1892), Breitinger (1938), Bach (1965), Trotter & Gleser (1951; 1952; 1958) were also employed. The framing of stature in the appropriate sex category was made by Martin & Saler (1956-1959). For the typological characterization we have used the methods and scales of Eickstedt (1934), Vallois (1934; 1944; 1953; 1965), Coon (1939), Bunak et al. (1941), Biasutti (1959), Comas (1960), Boev (1972), Baker (1974), and King (1981). Identification of the skeletal pathologies and abnormalities, including non-metric cranial, dental and postcranial traits, also assessing their degree of development/severity were made by methods recommended by Wells (1964), Rohlin (1965), Brothwell & Sandison (1967), Janssens (1970), Buikstra & Cook (1980), Ortner & Aufderheide (1991), Buikstra & Ubelaker (1994), Campillo (1994), Mays (1998), Aufderheide & Rodriguez- Martin (1998), Cox & Mays (2000), Ortner (2003), Matshes et al. (2004), McCoy (2004), Mann & Hunt (2005), Bailey (2006), Roberts & Manchester (2007), Slootweg (2007), Katzenberg & Saunders (2008), Kimmerle & Baraybar (2008), Richard Scott (2008), Brickley & Ives (2008), Waldron (2009), Barnes (2012), and Leroux (2012). There were also analyzed skeletal particularities viewed in the literature as functional adaptations and occupational or life style markers, or mechanical enthesopathies.

Results and Discussion Preservation status. The skeleton is incomplete and fragmented. The fractures and fissures that led to fragmentation are produced post mortem. The cranial skeleton appears slightly better preserved than the postcranial one. The skull was fragmented during the cleaning process of the skeleton, and it was restored to calvaria. The cranium is represented by frontal, parietals, occipital, left temporal, the part of the left zygomatic, left maxilla and left half of the mandible. With many absent segments, the postcranial skeleton is represented only by incomplete bones from the limbs (femurs, tibias, humeri, radii and ulnae). We add to this inventory a fragment derived from the left hip bone, from the cotyloid cavity. The bone sample contains, also, parts of the long bones’ epiphyses, but they are highly degraded and fragile. Reattaching them to the shaft bone was impossible. The postcranial skeleton, in contrast to the skull, is highly affected by taphonomic processes. The external bony layer has got a consistent calcareous (limestone) deposits. Besides the petrified deposits, diaphyses of the femora, tibiae and humeri shows some black islands (?) derived, probably, from the pedological layer in which deceased has been submitted. Note that on any bone present in the sample (complete, restored or fragmented) we have not identifies any burn marks. Some diaphyseal fragments show tooth traces of the rodents produces post mortem. The bone inventory of the analyzed skeleton can be seen in figure 3. Sex determination. The quite robust appearance of the skeleton, the shape of the cranium, the quite smooth surface of the frontal bosses, the pronounced supraorbital relief, the appearance of the forehead (relatively narrow and slightly sloping), the rounded orbital upper margins, the pronounced curvature of the occipital, the big mastoid process, the characteristics of the mandible (medium-sized robusticity, square chin), the teeth size (moderate to large), the quite large cotyloid cavity of the preserved hip fragment, and the

- 62 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 pronounced postcranial bone relief (joints and muscle insertions), all these lead us to define this skeleton, definitely, that being male. Age at death estimation. Even though the bone inventory is incomplete, we can say that the skeleton shows no signs of involution, of degenerative bone conditions, neither the pathological processes which may be related to the advanced age. The tissue from the femoral and humeral meta-epiphyseal region is compact (1st stage by Nemeskéri et al., 1960). The epiphyses of the long bones are welded to the diaphyses. The IIIrd molar is present, with closed root apexes. The occlusal surface of the IIIrd molar crown is physiologically functional. The dental wear, estimated by Brotwell (1981), has got the 3rd degree, indulgently 3+. The molars cusps are slightly worn. The occlusal surface shows some small dentin islands, this situation is valid only for the Ist molar and for the canine teeth. The incisors show visible linear islands of the dentine. The cranial sutures are completely open (by Buikstra & Ubelaker, 1994). According to all these features, the age at death of this man is between 20-25 years old (adultus category).

complete

partial

absent

Figure 3. Bone inventory of the skeleton discovered in Suceava-Câmpul Şanţurilor-strada Parcului.

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Biometrical data and morphological features. The cranial anthropometric value regarding the main dimensions and indices analyzed are listed in Table 1. The longitudinal diameter (eu-eu) of the neurocranium (Figs. 4-7) offers a very long size; the transversal one (eu-eu) presents a large size, their report giving a cranial index of dolichocranic type. The forehead is moderately blunt. The minimum diameter of the forehead (ft-ft) is middle-sized. The maximum frontal width (co-co) offer a large size, meaning a stenometopic frontal- parietal index, therefore indicating a spherical contour of the forehead, with diverging margins from the parietals. Regarding the degree of occipital curvature, the skull presents a bulgy and short occipital. The width of the occipital (ast-ast) belongs to the very big-sized category. The occipital-parietal index is large-sized. The shape of the neurocranium in norma verticalis is ovoid (Fig. 7), in norma occipitalis is the one of “house” (Fig. 6). The cranial bone relief is marked. Regarding the development of the glabellar relief, it indicates 4th degree, the supraorbital – 1st-2nd degree, the mastoid – the maximum degree. The external occipital protuberance indicates 1st-2nd degree. The nuchal muscles impressions are very obvious. The development of the mastoid apophysis indicates 5th degree. The extremely poor preservation status of the facial skeleton makes him immeasurable, with some exceptions. Some morphoscopic observations can be made. The zygomatic bones are quite high and moderately revealed. Their orientation/position cannot be determined. The palate presents a divergent parabolic shape, a moderate to large depth and a medium width (enm2-enm2). The torus palatinus is missing. The nose seems to have been narrow and quite high. The maximum width of the nasal aperture (al-al) is very small. The shape of the pyriform aperture belongs to the “trench” type. The canine fossa is slightly outlined (1st degree). The nasal spine was, probably, medium-sized. It was broken during cleaning process. The mandible, of which was preserved only the left half (Fig. 8), is moderately robust. Its depth is small. The height of the horizontal ramus is medium to high. The vertical ramus is short, wide and gently sloping. The mandibular mental protuberance is quite marked, with the pyramidal aspect. The gonial relief appears quite pronounced, easily designed out of horizontal ramus plane. The robustness (section) index of the mandible is medium. The torus mandibularis is missing. The general statement of the dentition supports the age at death of this man (20-25 years old) and the excellent status of the dental health. In the bone inventory are present only the left upper and lower dental arcades (Figs. 8-9). This individual has not suffered ante mortem tooth loss. The left central upper incisor has been lost post mortem. In the alveoli are present all the teeth from the two left arcades, as follows: central and lateral incisors, canines, Ist and IInd premolars, Ist, IInd and IIIrd molars. In total, 15 teeth are present in the alveoli, seven on the left upper arcade and eight on the left one. The dental wear (incisal and occlusal) is very low. The loss of tooth structure is physiological, attrition-type, produced during the functionality of the stomatognathic system (chewing movements). None of the teeth from the sample don’t have dental calculus or tooth decay. The dental hypoplastic defects are missing. There are no signs of the alveolar resorption. The postcranial skeleton (extremely incomplete and fragmented) has got pronounced muscle insertions. Because of the failure of the restoration process, we took from the limb bones (humerus, radius, ulna, femur and tibia) only the diameters and the circumferences (Table 2). The humerus (Fig. 10) shows pronounced deltoid muscle

- 64 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 insertions. This bone doesn’t have a supratrochlear foramen, and records a euribrachic diaphyseal section index. The section indices for radiuses and ulnae show the same situation. The femur (Fig. 11) is hyperplatymeric in the subtrochanteric region, with pronounced muscles insertions, forming pits and ridges. Linea aspera is prominent. The pilasters are presents in both femurs. The pilasteric index is slightly higher at the left femur. The low mesocnemic index indicates a tibial flattening. Tibias don’t have the hyperdorsiflexion signs. The high degree of fragmentation and the absence of the limb bone lengths led to the impossibility of the stature’s estimation. Using only the morphoscopic method, we appreciate for this man an upper-medium or a high-sized stature. The typological analysis was made, mainly, based on cranial measurements. In addition, we took the cranial and postcranial morphoscopic characteristics. The skeleton shows predominantly Nordic characteristics (namely: the cranial indices, the shape of the cranial vault, the mandible features, the skeletal robustness), to which it joins, to form a mix, some Dinaric elements.

Figure 4. Norma frontalis of the skull. Figure 5. Norma lateralis of the skull.

Figure 6. Norma occipitalis of the skull. Figure 7. Norma verticalis of the skull.

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Table 1. Cranial anthropometric values regarding the main dimensions (mm) and indices analyzed, and their classifications into appropriate categories (↓ = at the lower end of the category; ↑ = at the upper end of the category).

Martin Measured Appropriate Characters No. value (mm) category 1 g-op (maximum cranial length) 194 very long 8 eu-eu (maximum cranial breadth) 145 large ↓ 9 ft-ft (minimum frontal breadth) 95 middle 10 co-co (maximum frontal breadth) 122? large ↓ 12 ast-ast (maximum occipital breadth) 120? very large 43 fmt-fmt (upper facial breadth) 102 43(1) fmo-fmo (internal biorbital breadth) 94 54 al-al (nasal breadth) 22? very narrow 63 enm2-enm2 (internal palatal breadth) 40? middle 68 mandibular length 64? very short 69 id-gn (chin height) 33 69(1) Height of the mandibular body 32 middle ↑ 69(3) Breadth of the mandibular body 13 middle ↑ 70 Maximum ramus height 64 71 Minimum ramus breadth 33

Measured Appropriate Indices value category 8/1 Cranial index 74.74 dolichocranic ↑ spherical forehead, 9/10 Frontal-transversal index 77.9 diverging margins 9/8 Frontal-parietal index 65.5 stenometopic ↑ 12/8 Parietal-occipital index 82.7 large 9/43 Frontal-parietal index 93.1 middle 71/70 Mandible branch index 51.6 69(3)/69(1) Mandible robustness index 40.6 middle

Figure 8. Mandible (the left half). Figure 9. The left half of the maxilla.

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Table 2. Postcranial anthropometric values regarding the main dimensions (mm) and indices analyzed, and their classifications into appropriate categories (↓ = at the lower end of the category; ↑ = at the upper end of the category).

Measured (mm)/calculated Martin value Characters and indices No. and appropriate category left right

5 Maximum diameter at midshaft - 26 6 Minimum diameter at midshaft - 20 7 Minimum circumference of the diaphysis - 65 76.92 Humerus 6/5 Diaphyseal cross-section index - euribrachic

3 Minimum circumference of the diaphysis 47 - 4 Transverse diameter at midshaft 16 - 5 Anterior-posterior diameter at midshaft 12 - Radius 5/4 Diaphyseal cross-section index 75.0 -

11 Anterior-posterior diameter - 15 12 Medial-lateral diameter - 18 Ulna 11/12 Diaphyseal cross-section index - 83.0 6 Anterior-posterior midshaft diameter 33 31 7 Medial-lateral midshaft diameter 31 30 8 Midshaft circumference 97 95

9 Medial-lateral subtrochanteric diameter - 38 Anterior-posterior subtrochanteric 10 - 27 diameter Femur 106 103 6/7 Pilasteric index with pilaster ↓ with pilaster ↓ 71.0 10/9 Platymeric index - hyperplatymeric 8 Anterior-posterior midshaft diameter 36 - 9 Medial-lateral midshaft diameter 24 - 8a Maximum diameter at the nutrient foramen 42 -

Medial-lateral diameter at the nutrient 9a 29 - foramen Tibia 10b Minimum circumference of the diaphysis 84 - 9/8 Midshaft cross-section index 66.7 - 69.0 9a/8a Platycnemic index - mesocnemic

Pathologies, nutritional stress markers. The bone anomalies/pathologies show how the bone structure did not follow the normal structure under the influence of several genetic, exogenous or teratogenic factors. Unlike the major ones, which most of the times are lethal, less serious anomalies/pathologies only leave marks on the skeleton and cause slight health problems (Barnes, 2012). On tabula externa ossis cranii, in the area next to the lambda point, both the occipital and the parietals of the skeleton under study present foramina of cribra cranii externa type, also called porotic hyperostosis. Porotic hyperostosis is a pathological condition which affects the external part of the skull vault, seen in the appearance of a

- 67 - Angela Simalcsik et al. pointed network/structure which can be irregular (less serious) or uniform (very serious) (Walker et al., 2009). In this case, it is a first degree condition (by Stuart-Macadam, 1991). The porosity seems to have been active at the time of death (there are no signs of regeneration).

Figure 10. Humeri, Figure 11. Femora, posterior view. posterior view.

The skull porosity is a meaningful-suggestive indicator of some nutrition deficiencies and chronic illnesses, thus becoming an indirect and non-specific marker of the life quality and conditions, more precisely the health state and the nutrition habits (Walker et al., 2009; Piontek & Kozlowski, 2002). Generally speaking, the porosities seen on tabula externa ossis cranii develop during early childhood. They are less frequent among teenagers and much rarer among adults (Stuart-Macadam, 1985; Mays, 1998). In case it is not spread on the whole bone system, the first cause for the appearance of skull porosity, there should be considered sideropenic anemia (Ortner, 2003), namely iron deficiency, hence the risk to become ill is higher, especially for catching diseases (due to bacteria, viral illnesses, mycotic or parasitic diseases). The iron deficiency is given by insufficient quantities of iron in food or there are deficiencies in iron absorption or iron metabolism. There is a direct link between the acute gastroenteritis or parasites infections and the iron quantity, as they influence each other. Other causes which might lead to the appearance of porotic hyperostosis are vitamin C, D, B12, B6, B9 deficiencies as well as other inflammatory processes in the skull, osteomyelitis, and traumas (Walker et al., 2009).

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The non-metrical traits also called discontinuous, epigenetic or discrete traits/features are expressions of the variations noticed on the skull bones (including dental structures) and postcranial bones. Their importance lies on the hypothesis that they are more or less hereditary and might be used in relation to the ancestors, proving to be useful in estimating the biological affinity of the disappeared populations (Carson, 2006). As regards the non-metrical features, generally speaking, and the dental characteristics in particular, the analysis gains relevance only in the case of a population study, nevertheless this does not apply for this study. Still, we feel obliged to mention that the skeleton under study presents such discontinuous features. Thus, the analyzed skeleton which belonged to an adult male (20-25 years old), discovered in Suceava-Câmpul Şanţurilor-strada Parcului, chronologically ranked in the Middle Bronze Age, the Komariv (Komarow) culture presents two non-metrical features on the top skull and two on the dental structures. On the labdoid suture of the skull, in its right part, we notice the presence of a supplementary bone of small dimensions, called Wormian bone or intra sutural bone. The Wormian bones are anomalies of the normal fusion model of the ossification centres (Jeanty et al., 2000). Some authors correlate their appearance to congenital abnormalities or abnormalities of the central nervous system. Most authors consider that the appearance of intra sutural bones is controlled by genetic factors and it represents a variant of normality included in the group of non-metrical traits (El-Najar & Dawson, 1977). The left supraorbital region of the frontal bone presents an indentation also called the supraorbital notch – a non-metrical feature of the skull which appears as a reaction of the body to adapt by thermoregulation to the low temperatures in the environment, so as not to lose heat through the neuro-vascular system. As a consequence, the nerves and the blood vessels become thicker and deeply settled in the bone structures. The presence of such an indentation might suggest, in an indirect way, the cold and humid climate in the area where the individual lived (Tomaszewska et al., 2013). For the dental non-metrical features, the authors recommend they should be registered separately for the two arcades: 20 traits for the upper arcade and 16 for the lower arcade (McCoy, 2004; Bailey, 2006; Richard Scott, 2008; Leroux, 2012). For this particular study we mention only two such features in the dentition of the skeleton under study. The mandibular first molar presents an additional cusp, the occlusal surface presenting five cusps in all. The mandibular third molar has three extra cusps, showing on the occlusal surface a total of seven cusps. Activity-induced musculoskeletal stress markers, enthesopathies. At the level of the postcranial skeleton, on certain bones of the upper and lower limbs of this skeleton, we have identified a series of special characteristics which have been appreciated by the literature in the field as functional adjustments or adaptations, indicators of the physical activity and life-style (Larsen, 1997; Molleson, 2007). The humeri present highlighted insertions of the deltoid muscles (having a role in moving the arm forward, backward and on the side), the brachial biceps (playing a role in the flexion and extension of the forearm and the supination movement) and the pectoral (playing the role of bringing the arm upwards). This muscles group has left deep and rough signs on the humerus diaphyseal surface, which suggests that this person did activities which intensely used the arm muscles (such as repeatedly lifting heavy objects) (Molleson, 2007). The Euribrachic category of the humeral diaphyses comes to support this statement.

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The femurs, in the region of the gluteal tuberosity present a moderate to pronounced subtrochanteric prominence, seen in the crest and cavity/fossa. Even more, the femoral diaphysis presents a pilaster and the linea aspera shows enthesophytes – bony projections which are actually ossified entheses (which are, in fact, the connective tissue between tendon or muscle ligament and bone). These characteristics are strictly related to the intense activity of the muscle groups inserted on the back side of the femur: vastus lateralis and vastus medialis (playing a role in the extension of the lower limb), adductors muscle (contributing to the adduction, flexion and external rotation of the hip), the short head of the biceps femoris (influencing the hip extension and the knee flexion) and the gluteus maximus (influencing the abduction and the lateral rotation of the lower limb) (Teodorescu, 1982). This group of muscles left prominent and rugged signs, together with enthesophytes, which suggest an overstressing due to repeated activities such as walking or running on long distances, sustained walking on uneven ground, climbing, jumping (Molleson, 2007). The femoral hyperplatymery noticed in both hip bones comes to enforce these suppositions. We have to underline the fact that the skeleton remains from the inventory do not present any signs of any possible traumas, all the fractures or fissures which led to the fragmentation of this skeleton appeared post mortem. We should not overlook the fact that the bone inventory does not include the back spine, ribs, the pectoral arch and the pelvic arch, and the feet and hand bones. Considering the bone inventory (unfortunately incomplete) of the skeleton under study, we support up to a certain point the statement made by Ion Mareş (2010), according to which the skeleton might belong to a person who held an important position in the community. The social status is revealed by the three important elements exposed by the archaeologists who studied the funerary monument: the presence of the hammer-axe, the structure of the stone building and the considerable effort made by community members to turn the funeral into a complex event, to sacrifice time and make physical efforts to expose the passing away of a young man. In Ion Mareş’ opinion (2010), the funerary monument discovered at Suceava-Câmpul Şanţurilor required the use of approximately two tons of gritstone, most probably taken out from the bank of Suceava river. From the tomb to the edge of the Suceava river terrace there are approximately 300 meters. The effort made to transport the stone and build the funerary monument came from a large number of community members and the workload is huge, undoubtedly group-work. It is clear that not every community member received such a “treatment” when they passed away, only those who held an important position or, maybe, were high society. The stone tomb discovered at Suceava-Câmpul Şanţurilor-strada Parcului, chronologically ranked in the Middle Bronze Age, the Komariv (Komarow) culture is the only one ever discovered in the town of Suceava (Mareş, 2010). In the Suceava County there have been discovered and investigated from the archaeological point of view several funerary monuments belonging to Komariv (Komarow) culture, out of which we mention here only a few: the tumular necropolises from Horodnic de Jos, in Vârfu Colnicului point (Kaindl, 1903; Ignat, 1981; Niculică et al., 2014; Niculică, 2010) and Brădet point (Kaindl, 1903; Burtănescu, 2002); the tumular necropolis from Adâncata-Imaş (Niculică et al., 2005; Budui & Niculică, 2012, 2013; Niculică et al., 2013); the necropolis from Hârtop-Sub Plopi (Ursulescu & Popovici, 1987); the stone box (cista) tomb from Şerbăneşti (Ignat & Popovici, 1980). Other discoveries made in the area, included in the Costişa-Komarow

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(Komariv) culture are the ones from Prăjeni-Lutărie (Ursulescu & Şadurschi, 1988) and from Cotârgaci, both in the Botoşani County (Moscalu, 1989; Perianu, 1989; Dumitroaia, 2001). Unfortunately, out of the monuments mentioned above, only the tumular necropolis from Adâncata-Imaş (Suceava County) benefited from a proper anthropological analysis (Simalcsik & Niculică 2012). The study of the osteological series from Adâncata- Imaş highlighted the presence of a total of 18 individuals (14 inhumed and 4 incinerated) in the eight tumuli where there were discovered human bone remains. Male skeletons are predominant. As regards the age at death, out of the 18 deceased, 12 died right after the age of 20 (adults), hence having gone over the critical childhood age. The other 6 people are distributed as follows: in a single case the age cannot be estimated and in the other 5 cases, the people did not reach the age of 20, which can be rendered in a percentage of 28% out of the total population which ”succumbed” in childhood or teenage years. Unfortunately, the conservation state of the skeleton material from Adâncata-Imaş made it impossible to undertake some detailed typological and paleopathological analyses.

Conclusions The disinterred skeleton from the stone tomb, discovered in Suceava-Câmpul Şanţurilor-strada Parcului, chronologically ranked in the Middle Bronze Age, the Komariv (Komarow) culture, belonged to a male, aged approximately 20-25 at death (in the adultus category). Even if the skeleton is not complete, the conservation state of the remaining bones in the inventory is satisfactory. The bones are robust, clearly indented, both in the skull segment, as well as in the postcranium. The humerus is euribrachic, the femur is hyperplatymeric (with pilaster), and the tibia is mesocnemic. The stature, assessed only in morphoscopic traits seems to have been upper-medium or even high. From the typological point of view, this skeleton presents mostly Nordic features, in a combination with certain Dinaric traits. The teeth present a slightly worn occlusal / masticatory surface. None of the teeth is affected by calculus deposits, decay or any other infectious processes, a situation which proves an excellent oral health. This man did not lose any teeth ante mortem. Enamel hypoplasia is not present in any way, which suggests that during childhood (the period when the dental crowns are formed and the teeth crowns are calcified) the man did not experience any severe physiological stress (serious illness, malnutrition). The slight teeth wear of physiological type indicates the man’s preference for soft food. We should not overlook the young age at the moment of death (20-25 years old). Mention should be made that the skeleton does not present any severe bone pathology. The only pathological disease that could be identified is porotic hyperostosis (cribra cranii externa) – an indirect clue for nutritional stress and the health state. The exocranial porosity, active at the moment of death is localized in the area of the lambda point (on the occipital and parietals). The presence of porotic hyperostosis in itself, even if not very serious in this case, indirectly indicates a nutritional deficiency, hence slight health problems. In the absence of generalized bone porosity, as the first cause for the appearance of exocranial porosity, there should be mentioned the iron deficiency which entails an increased risk of illnesses, especially catching diseases. Other possible causes which might be mentioned here are C, D, B12, B6, B9 vitamins deficiency.

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The analyzed skeleton also presents a few non-metrical traits (two cranial and two dental). We mention the presence of an additional/accessory ossicle (Wormian bone) of small dimensions on the right part of the lambdoid suture – as a variant of normality. In the supraorbital area of the frontal bone there is noticed the presence of the supraorbital notch, which is an indirect indicator of the cold and humid climate where the person lived. Out of the dental abnormalities we underline the presence of additional cusps (1st molar has five cusps and the 3rd molar has seven cusps). Some postcranial bones have specific traits which could suggest the muscular massiveness of this man, but also a series of not very stressful daily activities. The characteristics of the humerus somehow betray the physical efforts made especially using the arms, such as, maybe, lifting of some burdens. The structural features of the femur and tibia indicate activities such as walking and running on long distances, fast walking on rugged/uneven land, maybe climbing or jumping. In conclusion, except nutritional deficiency but not very serious (most probably a slight sideropenic anemia) this men did not have any other health problems. The teeth were in excellent health. The abnormalities noticed on the skeleton remaining bones are actually variants of normality. Even more, the skeleton does not present any feature that would indicate exaggerated physical effort, the occupational indicators show only moderate physical labour. There are no signs of possible traumas which might have occurred ante mortem or peri mortem, all the fractures/fissures which led to the skeleton fragmentation appeared post mortem. The results of the anthropological study on the skeleton discovered in the stone box (cista) from Suceava-Câmpul Şanţurilor-strada Parcului should be considered a positive element in the overall knowledge on the behaviour and funerary customs of the Komariv (Komarow) population within the borders of our country. We plan on doing a comparative study in the near future, in the fortunate situation in which the archaeological research from Suceava region will bring to light the necropolis of Komariv (Komarow) culture, the same one mentioned as possible to exist by the Ioan Mareş – coordinator of the team which discovered the funerary monument from Suceava- Câmpul Şanţurilor-strada Parcului.

Acknowledgements The team responsible for uncovering and studying the funerary monument from Suceava-Câmpul Şanţurilor-strada Parcului consists of archaeologists from Bucovina Museum from Suceava, as follows: PhD Ion Mareş (responsible), PhD Bogdan-Petru Niculică, Ticu Dolenschi, Paul Ciurari, Sânziana Bedreagă, and Cătălina Ungureanu. We thank them for our involvement in the analysis of this material. We thank especially Mr. Ioan Mareş for the osteological material made available for the anthropological study and for information provided on the archaeological context. Also, we thank Mr. Bogdan-Petru Niculică for the references on archaeological research on the Komariv (Komarow) culture from Romania.

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DENTAL PATHOLOGIES IN POPULATIONS FROM A 4TH CENTURY A.D. NECROPOLIS AND A MEDIEVAL NECROPOLIS (14TH-17TH CENTURIES) FROM SĂBĂOANI, NEAMŢ DISTRICT

Robert-Daniel SIMALCSIK Romanian Academy – Iași Branch, Department of Anthropological Research, Str. Th. Codrescu, no. 2, Iași, Romania, [email protected]

Abstract. This paper concerns the paleopathological dental analysis of the skeletal series coming from two necropolises found on the territory of Săbăoani village (Neamţ County, Romania). The first necropolis is chronologically ranked in the 4th century A.D., Sântana de Mureş culture. The anthropological analysis in this case was made on 57 skeletons only from inhumation graves (16 men, 19 women and 22 of indeterminate gender). The second necropolis is chronologically included in the late medieval period, the period between the middle of the 14th century and the 17th century. The anthropological analysis in this second case was made on 937 skeletons (405 men, 290 women and 242 of indeterminate gender). Dental wear appears as a physiological result (attrition), produced during the functioning of the mastication process. Among the dental pathologies, we have identified in the two necropolises the following: the dental caries (decay), intra vitam tooth loss, paradontosis, partial or total edentation, apical granuloma and dental calculus. The presence of dental pathologies is in between low and moderate for the skeleton series coming from the necropolis in the 4th century and from moderate to increase for the late medieval necropolis. The highest values are recorded by intra vitam tooth loss and by dental caries. Paradontosis is present only in the medieval series. The apical granuloma exhibits lower frequencies in both necropolises. The dental calculus (supragingival) records lower values compared with caries. It is deposited on the mastication as well as the frontal teeth area. All this observations suggest a preference for cariogenic nutrients (rich in carbohydrates) to the disadvantage of animal nutrients (diary products, meat), and the possible nutrient deficiencies.

Keywords: Săbăoani, 4th century A.D., 14th-17th centuries, paleopathological dental analysis

Rezumat. Patologii dentare în populaţii vechi din Săbăoani (judeţul Neamţ, România): date paleoantropologice privind două necropole de secol IV d. Hr. şi de secole XIV-XVII. Studiul de faţă analizează paleopatologic dentiţia a două serii scheletice provenite din necropolele descoperite la Săbăoani (judeţul Neamţ, România). Prima necropolă este încadrată cronologic în secolul IV d. Hr. (cultura Sântana de Mureş). Analiza antropologică în acest caz s-a realizat pe 57 schelete provenite din morminte de inhumaţie (16 bărbaţi, 19 femei şi 22 indeterminabile ca sex). A doua necropolă este încadrată cronologic între jumătatea secolului XIV şi secolul XVII. Analiza antropologică în acest de-al doilea caz s-a realizat pe 937 schelete provenite din morminte de inhumaţie (405 bărbaţi, 290 femei şi 242 indeterminabile ca sex). Uzura dentară în cele două serii analizate este fiziologică, de tip atriţie. Patologiile identificate sunt: caria dentară, căderea/pierderea dentară in vivo, paradontoza, edentaţia parţială şi/sau totală, granulomul apical şi tartrul supragingival. Prezenţa patologiilor dentare este slabă spre moderată în seria scheletică din secolul IV d. Hr. şi moderată spre ridicată în cea medievală. Cele mai ridicate valori sunt înregistrate de căderile/pierderile dentare in vivo şi de carii. Paradontoza este prezentă doar în seria scheletică medievală. Granulomul apical înregistrează frecvenţe scăzute în ambele necropole. Tartrul dentar (supragingival) înscrie valori mai scăzute comparativ cu cariile, stratul depus fiind localizat atât în regiunile masticatoare, cât şi în cele jugale. Toate aceste constatări ne sugerează preferinţa pentru alimentele cariogene (bogate în carbohidtraţi), în detrimentul celor de origine animală (produse lactate şi carne), dar şi posibile carenţe alimentare.

Cuvinte Cheie: Săbăoani, secolul IV d. Hr., secolele XIV-XVII, analiza paleopatologică dentară

Introduction Săbăoani village is part of Neamţ County (Fig. 1) and it is considered the biggest as well as one of the oldest Moldavian villages in Romania.

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The climate in this area is transitional-continental, as in the whole Central Moldavian Plateau. The altitude and the neighbourhood to Siret and Moldova Rivers include the area in the high plateau category, with average temperatures of 8-6°C and rainfall of 700-1000 mm/year (Bojoi & Ioniță, 1974). The Scandinavian climate influence is rarely felt, during the winter, depending on the circulation of the polar air masses (Amăriucăi, 2000).

Figure 1. Location of the settlement of Săbăoani, Neamţ County (https://www.google.com/maps/place/Săbăoani).

The soil in the confluence terraces of Moldavia with Siret River and the high terraces of Siret are fertile and generally known as Chernozem soils. The alluvial plains and inferior terraces are covered with alluvial and soils and protosoils of south-western confluence and only on the right side of Siret River there are present bogs on small areas. The higher hills of the Central Plateau present brown luvic soils (Andone & David, 1992). Săbăoani village is among the oldest communities of Catholics in Moldavia. There are two theories related to the oldest core of the village. The first says that it is approximately 3 kilometres away to the east from the present day position, on the edge of the upper terrace, on the side of the hill, on the way to Răchiteni nowadays, the place being called Berindeşti. The second theory disagrees with the first, as it states that part of Berindeşti later became Săbăoani, a new village (Doboș, 2002).

Material and Methods The osteological material which was the basis for this study is represented by the human skeleton series coming from the two necropolises found on the territory of Săbăoani village (Neamţ County). The first necropolis, chronologically ranked in the 4th century

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A.D., Sântana de Mureş culture (Ursachi, 2010), was studied during 10 campaigns of systematic diggings coordinated by Vasile Ursachi, an archaeologist from the History Museum in Roman, Neamţ County. The second necropolis from Săbăoani, chronologically included in the late medieval period, the period between the middle of the 14th century and the 17th century (Hordilă, 2000), was studied during 1986-2000. The diggings were coordinated by archaeologist Domniţa Hordilă, from the History Museum in Roman (Neamţ County). In the biritual necropolis from the 4th century A.D. (Sântana de Mureş culture) from Săbăoani, there were discovered a total of 102 graves, out of which 88 inhumation and 14 incineration graves (Ursachi, 2010). The anthropological analysis was made on 57 skeletons only from inhumation graves (16 men, 19 women and 22 of indeterminate gender), complete or incomplete. In the late medieval necropolis from Săbăoani (14th-17th centuries) there were discovered 1455 inhumation graves in all (Hordilă, 2000), out of which we have received 937 for analysis, complete or incomplete. The conservation state of the osteological material is satisfactory, generally speaking; still there are some cases of advanced fragmentation or lack of bone regions which made it quite difficult to strictly follow the analysis stages. The latter situation applies mostly to the osteological material from the necropolis from the 4th century A.D. We mention that we have excluded the skeletons from which we could not gather data on the health state of the masticatory system. For the estimation of the age at death for those who died under 20 (infans I, infans II and juvenis) we have considered the following aspects: evolution of the temporary and permanent dentition, the stages in development of the tooth buds, degrees of intergrowth of the long bone epiphysis in their diaphyses and the epiphyseal disks of the vertebrae in their bodies. We have used the methodology proposed by Maresh (1955, 1970), Moorrees et al. (1963), Trotter & Peterson (1969), Fazekas & Kosa (1978), Ubelaker (1978), Jeanty (1983), Scheuer & Black (2000), Schaefer et al. (2009). For the subjects who overpassed the growth period, namely those who had more than 20 years (adultus, maturus and senilis), the traits which acted as instruments for the estimation of the age at death were the following: evolution of the symphyseal surface of pubis and the surface of the sacroiliac joints, the changes in the spongiosal tissue of the humerus and thigh bone, the degree of tooth wear (in the mastication surface of the teeth), some phenomena of skeletal degeneration due to aging and cranial suture obliteration. We used as guide the methodology recommended by Nemeskéri et al. (1960), Ferembach et al. (1979), Ubelaker (1978), Brothwell (1981), Buikstra & Ubelaker (1994), Mays (1998), White & Folkens (2005), Schmitt (2005). Gender was established considering a complex of morphoscopic traits, as follows: shape and width of the pelvis, the aspect of the greater sciatic notch, the curving degree of the sacrum, the massiveness and the robustness of the skeleton, the development of the joints and muscle insertions, the development of the skull bones relief, the forehead shape and angle, the aspect of the orbital margin, the dimension and strength of the mandible, the shape and type of mandibular chin, the shape and dimension of the teeth, the dimension of the mastoid process. For this purpose we have used the methodology recommended by Iordanidis (1961), Stradalova (1975), Ubelaker (1978), Buikstra & Ubelaker (1994), Bruzek (2002), Walrate et al. (2004), Blanchard (2010). The tooth wear was estimated under the limits proposed by Périer (1935, 1949), Smith & Knight (1984), Brothwell (1981). The dental pathologies and their severity were

- 79 - Robert-Daniel Simalcsik analysed considering the methods recommended by Janssens (1970), Ortner & Aufderheide (1991), Buikstra & Ubelaker (1994), Mays (1998), Aufderheide & Rodriguez-Martin (1998), Ortner (2003), Slootweg (2007), Katzenberg & Saunders (2008), Richard Scott (2008), Waldron (2009).

Results and Discussion The dental paleopathological study of the skeletons discovered in the two necropolises from Săbăoani, chronologically belonging to the 4th century A.D. (Sântana de Mureş culture) and to the 14th-17th centuries (late medieval period) has brought to light several interesting cases. Among the dental pathologies known until now, we have identified in the two necropolises the following: dental caries (decay), in vivo tooth loss, paradontosis, partial or total edentation, apical granuloma and dental calculus. We will present each of them in what follows. Dental caries. The caries (decay, cavity) is a condition given by an infection, bacterial in origin, which causes demineralization and destruction of the hard tissues of the teeth. It is considered that the decayed lesions are signs of the disequilibrium between the calcium ions and the phosphates in the dental tissue and saliva, whereas the bacteria act as mediator. The main factors are precarious oral hygiene, a diet rich in carbohydrates, the life style and the genetic predisposition. Even if it is considered an illness of the modern civilisation, the dental caries has been described even since antiquity, its definition being limited to a few symptoms and its causes remain unclear. The human beings have presented dental cavities from the oldest times, but with a smaller prevalence rate in comparison to the one registered nowadays. The increase in prevalence might be explained by the change in the food components experienced throughout the centuries, especially when changing food coming from hunting and picking (meat and fruit) to food rich in carbohydrates (cereals), made available once the first agricultural activities appeared. The link between the growth in caries prevalence and the high consumption of carbohydrates changes the cavities into an indicator, even if it might be less specific, of the dietary habits. As for the relationship between the dietary habits and caries, we should mention that certain nutritional deficiencies (inappropriate intake of A, C and D vitamins), lack of oligoelements or essential amino acids in the dental growth can lead to an increased vulnerability of the teeth to the cariogenic agents, seen in the decrease of dental stamina (Iovan, 2011; Cucina et al., 2011). In the two necropolises from Săbăoani analysed in this paper, the dental decay, as we would expect, ranges between different values. The skeleton from the necropolis from the 4th century A.D. is less affected by this infectious process in comparison to the medieval series in the 14th-17th centuries. The presence of the dental caries is approximately 10% in the first case and between 30-40% in the latter. As for the sexual dimorphism, it can be clearly seen in the skeleton series from the 4th century A.D. in which men are more affected by the cariogenic agents than women. In case of the medieval skeleton series, the sexual dimorphism is also clearly seen, the cavity being present in the dental arcades of 295 men and 156 women. Table 1 presents the numbers and the percentages of dental caries incidence in the skeleton series from the medieval necropolis (14th-17th centuries) from Săbăoani – a sample with a representative number of people.

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The higher percentages in the whole medieval population are met for the arcades with 1-2 cavities which were probably easier to be endured than any other greater number. The latter situation determined the sufferer to take out teeth (a phenomenon which is described separately in this paper, together with the paradontosis cases).

Table 1. Dental caries frequency in the skeleton series from the medieval necropolis (14th-17th centuries) from Săbăoani.

Males Females Frequency of dental caries N % N % Without caries 94 31.86 55 35.25 1-2 caries 120 40.69 64 41.03 3-4 caries 40 13.56 10 6.41 5-6 caries 41 13.89 9 5.77 6 and more caries - - 18 11.54

Regarding the seriousness of the cavity, it varies from the first to the fifth degree (Fig. 2), when the entire dental crown is decayed and the bacterial infection reaches the pulp chamber. For this study, we have most commonly met the 2 and 3 degree cavities (Fig. 3), followed by the 1, 4 and 5 degree cavities. As for the location, we have noticed both occlusal cavities (Fig. 4) and vestibular (Fig. 3). The teeth most seriously affected by decay are molars I, followed by molars II (Fig. 4), premolars I and II, canines and incisor I and II. Should we make a comparison between the arcades, the lower one (mandibular) is much more affected by caries in comparison to the upper one. Considering the process of decay in the temporary dentition, in the group of children named infans I (0-7 years old) we have noticed the presence of a pathological process in a number of 11 subjects out of a total of 98 under study, which means approximately 11% of the infans population (a small percentage for the medieval times). Most cavities in children are seen on the permanent molar I.

Figure 2. Male, 30-35 years old. Figure 3. Male, 35-40 years old. Săbăoani, medieval necropolis. Săbăoani, medieval necropolis. Tooth Severe tooth caries (5th degree) and vestibular caries (2-3 degree) on the II and ante mortem tooth loss. the III mandibular left molars.

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Figure 4. Female, 25-30 years old. Săbăoani, medieval necropolis. 4 degree tooth caries on the II mandibular right molar.

Intra vitam tooth loss. The tooth loss during the lifetime might be grouped considering the aspect of the mandibular alveoli. The alveoli of the teeth lost only a short time before death are porous, while those lost a long time before death (at least 6 months) are completely closed and resorbed. Tooth loss falls into two categories, physiological and demanded. The physiological tooth loss appears when the alveoli eliminates itself the affected tooth, as well as its root. The demanded tooth loss (extractions) appears only when the sufferer chooses to take out the painful teeth out of the alveolus (Iovan, 2011; Cucina et al., 2011). The demanded tooth loss was widely seen during the medieval times. The first cause of intra vitam tooth loss is the cavity. Actually, it is unanimously admitted that the teeth which fall or are extracted from the alveolus necessarily go through a pathological process, mostly cavities and their complications. Paradontosis is another cause of dental loss, as it is an acute degenerative illness of the tissues which keep the teeth fixed in the alveoli. Paradontosis derives from untreated gingivitis and it means the gum tissue is destroyed, leading to its drawn-back aspect and the appearance of lesions (Firu, 1967). During this study we have noticed a great number of dental losses during the lifetime, in the series come from the necropolis from the 4th century A.D., as well as from the series from the 14th-17th centuries. In the sample from the 4th century A.D., the intra vitam tooth loss had a frequency of approximately 25%, and for the medieval period (the 14th-17th centuries) the frequency was of approximately 40%. Related to paradontosis, this study mentions this phenomenon only for the skeleton series from Săbăoani, registering a presence of approximately 26-27% in both genders. If we analyse the data from the point of view of the sexual dimorphism, in the skeleton series from the 4th century, the tooth loss presents a higher percentage in women than in men. As we have mentioned before, men registered a greater percentage of cavities than women. The intra vitam dental loss is more frequent in the mastication region, which means it mainly affected molars (Fig. 5). We have identified this pathological phenomenon in the region of the premolars as well (Fig. 6), as in the front region (of the incisors and canines). Most arcades with losses present healed alveoli, fully unswollen, which proves that the phenomenon of physiological rejection or demanded extraction of the decaying

- 82 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 teeth happened a long time before death.

Figure 6. Female, 45-50 years old. Figure 5. Female, 45-50 years old. Săbăoani, Săbăoani, the 4th century A.D. necropolis. the 4th century A.D. necropolis. Ante mortem Ante mortem tooth loss (mandibular tooth loss in the masticatory areas. premolars and molars).

In the skeletal series from the medieval necropolis (14th-17th centuries) from Săbăoani, in both genders, the percentage of individuals without any teeth loss is rather low (of approximately 45%). The phenomenon grows especially if we make a comparison with other medieval synchronous necropolises. The percentage of 45% for complete arcades, namely no dental loss is a very low one, and this fact is correlated to the analysis on the decaying process. The presence of a dental loss of 40% in the medieval skeletal series under study (together with the increased frequency of cavities and paradontosis) suggests that this population lived in precarious hygiene. The dental loss during their lifetime affected mainly the mastication areas (Figs. 7-8), as regards the skeleton series from the 4th century. As a preference, mostly affected were the individuals older than 45-50.

Figure 7. Female, 60-65 years old. Figure 8. Male, 60-65 years old. Săbăoani, Săbăoani, medieval necropolis. Partial medieval necropolis. Ante mortem tooth loss of extended edentation of the maxillary the mandibular masticatory areas. Dental abscess masticatory areas. of the anterior teeth.

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We have included in Table 2 the frequency of the dental loss and paradontosis for the skeleton series from the medieval necropolis from Săbăoani – a sample with a representative number of individuals. It is noticed that the arcades without any tooth loss present similar percentages for both genders and the same applies for the category of the arcades with 5-6 teeth loss, even for the paradontosis condition. For the other two categories (arcades with 1-2 and 3-4 tooth loss) the sexual dimorphism is better seen, in favour of one gender or the other. Thus, arcades with 1-2 tooth loss are more frequent in women and those with 3-4 teeth loss in men. We need to take into account the fact that part of the decayed teeth (previously analysed) fell or were extracted intra vitam and this fact changes somehow the frequency of cavities. We believe that the members of the community from Săbăoani were concerned about the oral hygiene in the sense that they extracted the painful teeth with cavities, especially if they had more than two cavities.

Table 2. Frequency of tooth loss and paradontosis for the skeleton series from the medieval necropolis from Săbăoani, 14th-17th centuries. Male Female Frequency of tooth loss N % N % Without tooth loss 115 38.99 67 42.96 1-2 missing teeth 25 8.47 24 15.38 3-4 missing teeth 38 12.88 8 5.12 5-6 missing teeth 38 12.88 16 10.26 Paradontosis 79 26.78 41 26.28

Total/partial edentation. Edentation is a pathological condition characterised by the absence of one or several teeth from the arcades. Partial edentation (intra vitam tooth loss discussed in the above paragraph here included) refers to the lack of more than 3 dental units in a row, on a half-arcade. Total edentation means the lack of all teeth in the oral cavity due to their loss after the natural eruption and growth. Total edentation might be caused by several factors. The dental decay and its complications is the main cause. The second cause, as frequent as the first one is parodontopathy which refers to the gradual withdrawing of the tissues which support the teeth, this phenomenon being followed by the spontaneous loss or tooth extraction. One of the evolutive forms responsible for the tooth loss is deep chronic marginal periodontitis. In this illness, the alveolar bone is affected and there is observed serious bone resorption and atrophy. Other conditions which could lead to total edentation are the infectious processes in bone or soft tissues, tumours or facial traumas (Firu, 1967; Slootweg, 2007). The side effect of edentation is the bone remodelling of the facial area, meaning that the bone mass becomes resorptive and atrophic. This phenomenon leads to the fall of the lower part of the face bones as the upper jaw and the mandible decrease in height, but also due to the decrease of the bone tissue. The alveolar bone resorption means this part of the body decreases until it no longer exists, thus being useless to the body, as a consequence of no longer fulfilling the functions he had been created for. Atrophy is the decrease in size of an organ due to deficient nutrition. In case of the upper jaw, resorption and atrophy take place in a centripetal (concentric) manner whereas for the mandible the process is reversed, it is centrifugal. Once age advances, the bone atrophy becomes deeper as a consequence of slowing down the metabolic processes, especially due to the

- 84 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 appearance of senile osteoporosis. It has been noticed that the female gender is more vulnerable to a negative evolution for edentation, as it comes mingled to endocrine disorders and osteoporosis present at menopause (Waldron, 2009). This study proves the presence of edentation is higher in the sample from the medieval necropolis (14th-17th centuries) in comparison to that from the 4th century A.D. Generally speaking, partial edentation, in other words the lack of three teeth in a row (Figs. 6-8) is much more frequent in comparison to total edentation (Figs. 9-10). When we consider sexual dimorphism, total edentation affects mainly the female gender, especially after the age of 50. The phenomena of resorption and atrophy of the jaw regions were highlighted only in women (Fig. 10).

Figure 9. Male, 55-60 years old. Săbăoani, Figure 10. Female, 55-60 years old. medieval necropolis. Total maxillary Săbăoani, medieval necropolis. Total edentation, without atrophy or resorption. maxillary edentation, with atrophy and resorption of the bone tissue.

Granuloma. Granuloma, also called the periapical or radicular cyst or abscess, refers to an infection in the radicular apex area, which is chronic as it had not been treated. This formation has degenerative effects on the alveolar edge as well as on the dental arcades as they are formed due to a defensive reaction of the apical tissues to neutralize and isolate the toxic products (sent out by the bacterial infections) from the radicular canal. The cyst entails large cavities in the bone, having regular walls around the dental apex. Granuloma is more frequent in the upper jaw than in the mandible (Slootweg, 2007; Ştefănescu, 2007). In the skeletal series from Săbăoani, the dental granuloma was noticed more frequently in the medieval necropolis (14th-17th centuries), registering a presence of approximately 2%. In the skeleton series from the 4th century A.D., this infectious process was noticed on the alveoli of two individuals, males. Sexual dimorphism related to this pathological phenomenon is noticeable, as the male gender is more affected than the female. In the skeleton series from the medieval necropolis, the apical granuloma was noticed on 49 skeletons (37 males and 12 females). As for the positioning, it appears mostly in the molars (29 cases) (Figs. 11; 13), but also in the incisors (11 cases) (Fig. 12) and canines (7 cases) (Fig. 8) as well as premolars (2

- 85 - Robert-Daniel Simalcsik cases). We mention that the 7 cases of apical granuloma on the canine belong to the male gender, without exception (Fig. 8).

Figure 12. Female, 50-55 years old. Figure 11. Male, 45-50 years old. Săbăoani, Săbăoani, medieval necropolis. medieval necropolis. Dental abscess of the right Dental abscesses of the mandibular mandibular I molar. I and II incisors.

Figure 13. Female, 25-30 years old. Săbăoani, medieval necropolis. Dental abscesses of the right I and left II mandibular molars.

Dental calculus. Dental calculus, also called dental tartar is a concretion of mineral salts on the teeth surface, as a result of the mineralization of the bacterial plaque in an alkaline environment. Tartar actually represents the calcified (hardened) plaque due to its presence on the teeth surface. There are two types of dental calculus: above gum calculus deposits (supragingival) and below the gingival margin (subgingival). Supragingival tartar (salivary duct adjacent) is placed mainly on the inferior front teeth and it appears as yellowish deposits, at first reduced in consistency, soft, breakable and easily to be removed. In time, its consistence and adherence increase and its colour becomes darker,

- 86 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 influenced by food pigments or other harmful substances pigments. The subgingival tartar is brownish, it is hard and it presents high density and it is strongly built-up and it is harder to remove than the supragingival one (Aufderheide & Rodriguez-Martin, 1998). Dental calculus is the origin of most periodontal illnesses and it is also the main cause for cavities. The mineralization degree is tightly connected to the age of the dental plaque, related to its width, chemical composition and the type of bacteria which are present. Several microscopic studies on the dental calculus from the prehistoric skeletons have shown a wide range of components, apart from bacteria: pollen grains, plant pieces, fur filaments (Aufderheide & Rodriguez-Martin, 1998). Increased prevalence of dental calculus was noticed in the medieval population involved in hunting, the included factors being the lack of oral hygiene, cooking techniques, a diet rich in animal proteins (meat and diary products) or cultural activities (Lukacs, 1989). It is important to note down that there is a negative cause-effect link between the appearance of tartar and cavities. Tartar develops in an alkaline environment (in demineralization) whereas cavities appear in an acid environment (by demineralization), hence the two processes are simultaneously incompatible. Nevertheless, in most cases, the dental arcades exhibit both calculus and cavities. It has been proven that the tartar formed on a tooth with a cavity actually blocks its development (Aufderheide & Rodriguez-Martin, 1998).

Figure 14. Male, 50-55 years old. Figure 15. Male, 35-40 years old. Săbăoani, Săbăoani, medieval necropolis. medieval necropolis. Dental supragingival calculus Dental supragingival calculus accumulation accumulation on the right on the mandibular frontal teeth. mandibular masticatory area.

Figure 16. Male, 35-40 years old. Săbăoani, medieval necropolis. Dental supragingival calculus accumulation on the mandibular frontal teeth.

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In this study, tartar does not register a significant presence, the frequency being approximately 5% in the skeleton series from the 4th century A.D. and around 9% in that coming from the medieval necropolis. Sexual dimorphism is clearly expressed, by this proving that all the cases of tartar discovered in this study belong to the male gender, hence, again, we might deduce the preference expressed by men for nutrients of animal origin. Related to the positioning, this study highlights that tartar deposits are seen especially on the surfaces of mastication (Fig. 15), but they also appear frequently in the frontal area (Figs. 14; 16). In all cases identified as having been affected by a pathological process, the dental calculus is of supragingival type (salivary duct adjacent). In some cases, the tartar layer is thin, of reduced consistency, not too tightly built-up on the teeth and it can be easily removed. Still, more frequent are the cases in which the tartar is hard, dark and strongly stuck on the crown, thus being very difficult to remove.

Conclusions The paleopathological study on the dentition of the skeletons discovered in the necropolises from the region of Săbăoani village led to interesting results. Interpreted in a comparative manner, they generated important conclusions as regards the health condition of the teeth. The two necropolises discovered on the territory of Săbăoani village belong to an important historical period, as archaeologists Vasile Ursachi and Domniţa Hordilă state. The first and oldest necropolis is included in the 4th century and it belongs to Santana de Mureş culture and it is represented in this study by 57 skeletons from inhumation graves. The second necropolis was included in the period between the 14th-17th centuries, part of the late medieval period, here represented by 937 skeletons coming from inhumation graves. The conservation status of the osteological material is satisfactory, in general terms. The presence of dental pathologies is in between low and moderate for the skeleton series coming from the necropolis in the 4th century and from moderate to increase for the late medieval necropolis. Considering the whole osteological sample, we have registered the following deviations from the normal structures: cavities, intra vitam dental loss, paradontosis, partial or total edentation, apical granuloma and dental calculus. Dental wear appears as a physiological result (attrition), produced during the functioning of the mastication process. We have not noticed the presence of other types of wear, such as erosion or abrasion, a situation which applies to both skeletal series. The dental decay exhibits a frequency of 10% in the necropolis from the 4th century A.D. and approximately 40% in the medieval skeleton series, in both men and women. Intra vitam tooth loss most of the times appeared as a consequence of cavities and sometimes of paradontosis, registers a value of 25% in the necropolis from the 4th century A.D. and approximately 40% in that from the 14th-17th centuries. Paradontosis is not present in the series from the 4th century A.D. while in the medieval series it presents a percentage of approximately 26-27% in both genders. Partial or total edentation appears much more frequently in the medieval necropolis in comparison to that from the 4th century A.D.

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The apical granuloma exhibits lower frequencies in both necropolises, of only 0.3% in the skeleton series from the 4th century and approximately 2% in the medieval one. Supragingival tartar presents a frequency of 5% in the series from the 4th century A.D. and 9% in the medieval series, as it is deposited on the mastication as well as the frontal teeth area. The data regarding the prevalence of the dental pathologies in the two necropolises under study suggest valuable information on the nutrition habits of the community members. Even more, they slightly insinuate the part of the living conditions for the two historical periods. In both skeletal series coming from the two necropolises from Săbăoani, mostly the medieval one, the dental pathologies prove to have a high percentage. The preference for cariogenic nutrients (rich in carbohydrates) to the disadvantage of animal nutrients (diary products, meat), together with the possible nutrient deficiencies, such as some avitaminosis, the lack of oligoelements or scarcity in certain essential amioacids required for the teeth growth are part of the factors which entailed increased vulnerability of the stomatognathic apparatus, seen in the decrease of dental strength.

Acknowledgements We kindly thank the researchers who coordinated the archaeological diggings conducted on the territory of Săbăoani village (Neamţ County) – PhD Vasile Ursachi and PhD Domnița Hordilă from the History Museum in Roman, Neamţ County. They provided us with the osteological material from the necropolises in Săbăoani in order to conduct our study.

References Amăriucăi, M., 2000. Şesul Moldovei Extracarpatice dintre Păltinoasa şi Roman. Editura Corson, Iaşi. Andone, M., David, G., 1992. Istoria oraşului Roman: (1392-1992). Societatea Culturală “Roman-600”, Roman. Aufderheide, A.C., Rodriguez-Martin, C., 1998. Cambridge Encyclopedia of Human Paleopathology. Cambridge. Blanchard, B.K., 2010. A study of the accuracy and reliability of sex estimation methods of the human pelvis. A Thesis presented to the Faculty of California State University, Chico. Bojoi I. Ioniţă I., 1974, Judeţul Neamţ. Editura Academiei Române, Bucureşti. Brothwell, D.R., 1981. Digging up bones. British Museum of Natural History. London. Bruzek, J., 2002. A method for visual determination of sex, using the human hip bone. American Journal of Physical Anthropology, 117: 157-168. Buikstra, J.E., Ubelaker, D.H., 1994. Standards for Data Collection from Human Skeletal Remains. Arkansas Archaeological Survey Research Series, 44, Fayetteville. Cucina, A., Cantillo, C.P., Sosa, T.S., Tiesler, V., 2011. Carious Lesions and Laize Consumption Among the Prehispanic Maya: An Analisys of a Coastal Community in Northern Yucatan. American Journal of Physical Anthropology, 145(4): 560-567. Doboş, F., 2002. Săbăoani, file de istorie. Editura Presa Bună. Fazekas, I.Gy., Kosa, F., 1978. Forensic Fetal Osteology. Budapest Akademiai Kiado. Ferembach, D., Schwidetzky, I., Stloukal, M., 1979. Recommandations pour determiner l’age et le sexe sur le squelette. Bulletins et Memoires de la Societe d’Anthropologie de Paris, XIII(6, 1): 7-45. Firu, P., 1967. Stomatologia infantilă. Editura Pedagogică, Bucureşti. Hordilă, D., 2000. Câteva consideraţii de ordin arheologic privind populaţia catolică din zona Romanului în secolele XIV-XIX. Buletin Istoric, 1: 59-82. Iordanidis, P., 1961. Determination du sexe par les os du squelletts. In Annales de Medecine Legale, Criminologie, Police Scientifique et Toxicologie, 41: 280-291. Iovan, G., 2011. Caria dentară. Repere etiologice şi patogenice. Editura „Gr. T. Popa” Iaşi. Janssens, P.A., 1970. Paleopathology. London. Jeanty, P., 1983. Fetal limb biometry. Radiology, 147: 601-602. Katzenberg, M.A., Saunders, R.S., 2008. Biological Anthropology of the Human Skeleton. Hoboken.

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Lukacs, J.R., 1989. Dental Palaeopathology. Methods for reconstructing Dietary Patterns. In Iscan M.Y., Kennedy K.A.R. (eds), Reconstruction of Life from the Skeleton, New York, 261-286. Maresh, M.M., 1955. Linear growth of long bones of extremities from infancy through adolescence. American Journal of Diseases of Children, 89: 725-742. Maresh, M.M., 1970. Measurements from roentgenograms. In McCammon R.W. (Ed.), Human Growth and Development, Springfield, IL: C.C.Thomas. Mays, S., 1998. The archaeology of human bones. Ed. Routledge. Moorrees, C.F.A., Fanning E.A., Hunt E.E., 1963. Age variation of formation stages for ten permanent teeth. Journal of Dental Research, 42: 1490-1502. Nemeskéri, J., Harsányi, L., Acsády, Gy., 1960. Methoden zur Diagnose des Lebensalters von Skelettfunden. Anthropologhischer Anzeiger, 24: 70-95. Ortner, D.J., 2003. Identification of Pathological Conditions in Human Skeletal Remains. Oxford. Ortner, D.J., Aufderheide, A.C., 1991. Human paleopathology. Current Syntheses and Future Options. Washington-London. Périer, A.L., 1935. Observation sur le phénomène de l’abrasion dentaire fonctionnelle chez un groupe ethnique inférieur. L’Odontologie, 28(10): 687-697. Périer, A.L., 1949. Usure, abrasion, érosion. Pratique Odonto-Stomatologique, 140: 1-7. Richard Scott, G., 2008. Dental Morphology. In (Eds.) M.A. Katzenberg, R.S. Saunders, Biological Anthropology of the Human Skeleton, Hoboken, 265-298. Schaefer, M., Black, S., Scheuer, L., 2009. Juvenile osteology. Elsevier Academic Press. Scheuer, L., Black, S., 2000. Developmental Juvenile Osteology. Elsevier Academic Press. Schmitt, A., 2005. Une nouvelle methode pour estimer l’age au deces des adultes a partir de la surface sacro- pelvienne iliaque. Bulletine et Memoire de la Societe d’Anthropologie de Paris, 17(1-2): 1-13. Slootweg, P. J., 2007. Dental pathology. A practical introduction. Berlin. Smith, B.G., Knight, J.K., 1984. An index for measuring the wear of teeth. British Dental Journal, 156: 435-438. Stradalova, V., 1975. Sex differences and sex determination on the sacrum. Anthropos, 13(3): 237-244. Ştefănescu, D.G., 2007. Studiul cariei dentare la populaţia din Moldova aparţinând culturii Sântana de Mureş, sec. III-IV. Teză de doctorat (rezumat), Universitatea de Medicină şi Farmacie „Gr. T. Popa”, Iaşi. Trotter, M., Peterson, R.R., 1969. Weight of bones during the fetal period. Growth, 33: 167-184. Ubelaker, D.H., 1978. Human Skeletal Remains: Excavation, Analysis and Interpretation. Smithsonian Institute Press, Taraxacum, Washington. Ursachi, V., 2010. Săbăoani. Monografie arheologică, I-II. Casa Editorială Demiurg, Iaşi. Waldron, T., 2009. Palaeopathology. Cambridge. Walrath, D. E., Turner, P., Bruzek, J., 2004. Reliability test of the visual assessment of cranial traits for sex determination. American Journal of Physycal Anthropology, 125: 132-137. White, T.D., Folkens, P.A., 2005. Human bone manual. Elsevier Acadmic Press.

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OSTEOPATHIES IN THE POPULATION OF OLD IASI CITY (ROMANIA): THE NECROPOLIS OF THE “BANU” CHURCH, 16TH-19TH CENTURIES

Vasilica-Monica GROZA1, Angela SIMALCSIK1, Luminiţa BEJENARU1, 2 and Robert- Daniel SIMALCSIK1 1 Romanian Academy – Iași Branch, Department of Anthropological Research, Str. Th. Codrescu, no. 2, Iaşi, Romania, [email protected], [email protected] 2 “Alexandru Ioan Cuza” University, Faculty of Biology, Bd. Carol I, 20A, Iaşi, 700505, Romania

Abstract. This study describes the incidence and the analysis of a series of osteopathies observed in the skeletal series found in the necropolis of the ancient “Banu” Church of Iași City (Romania). According to the authors of the excavations, the necropolis was used between the first half of the 16th century and the beginning of the 19th century. The osteological material consisting of 67 skeletons (children, teenagers, adults, mature and senile) derives from both individual tombs and reinhumation tombs. The distribution by gender and by age categories indicates a higher male mortality rate as opposed to females, resulting in a higher masculinity index. Most deaths were recorded in the maturus category (59.70%), followed by infans I and II (26.15%), senilis (5.97%), juvenis (4.48%) and adultus (2.99%). The incidence of pathologies, anomalies and non-metric features was estimated both separately by gender and for the entire osteological sample. In the cranial segment (for the entire osteological series), wormian bones and metopic suture have equal incidence – 2.98%, followed by porotic hyperostosis, cribra orbitalia, and premature synostosis of the sagittal suture (with equal incidence – 1.49%). In the post-cranial segment, extra facets of the tibia recorded 8.98%, followed by osteoarthritis (7.46%) and supratrochlear foramen of the humerus (5.97%). Lower incidences were recorded for spina bifida occulta and spondylolysis (2.98% each).

Keywords: Iași City, 16th-19th centuries, necropolis, skeletal series, osteopathies

Rezumat. Osteopatii la populaţia vechiului oraș Iaşi (România): necropola Bisericii „Banu”, secolele XVI- XIX. Lucrarea prezintă incidenţa şi studiul unor osteopatii semnalate la seria osteologică deshumată din necropola situată în incinta Bisericii „Banu” din orașul Iaşi (România). Conform autorilor săpăturii arheologice, limitele de folosire în timp a necropolei se situează între prima jumătate a secolului al XVI-lea şi începutul secolului al XIX- lea. Materialul osteologic reprezentat prin 67 de schelete (copii, adolescenţi, adulţi, maturi şi senili) provine atât din morminte individuale cât şi din morminte cu reînhumări. În urma repartiţiei pe sexe şi pe categorii de vârstă s- a constatat o pondere mai mare a bărbaţilor comparativ cu femeile, ceea ce indică un indice de masculinitate mai crescut. Semnalăm o predominanţă a deceselor în intervalul maturus (59,70%), urmate de categoriile, infans I şi II (26,15%), senilis (5,97%), juvenis (4,48%) şi adultus (2,99%). Prezenţa patologiilor, anomaliilor şi a caracterelor non-metrice a fost calculată atât separat pe sexe, cât şi pentru întregul eşantion osteologic. La nivelul segmentului cranian (pentru întreaga serie scheletică), oasele wormiene şi sutura metopică prezintă procente egale – 2,98% urmate de hiperostoza porotică, cribra orbitalia şi sinostozarea prematură a suturii sagitale (cu procente egale – 1,49%). În segmentul postcranian, faţetele suplimentare pe articulaţia tibio-talară înregistrează un procent de 8,98%, urmate de osteoartrită (7,46%) şi foramenul humeral supratrohlear (5,97%). Frecvenţe reduse au înregistrat spina bifida occulta şi spondiloliza (procente egale – 2,98%).

Cuvinte cheie: orașul Iași, secolele XVI-XIX, necropolă, serie de schelete, osteopatii

Introduction It is a well known fact that bone diseases (the incidence and gravity of osteopathies) are used as “markers” for advanced ages, but they can also be associated with hygienico- sanitary conditions (infections), diet, hard labour, carrying burdens, etc.

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Paleopathology analyses the origins of diseases, their spreading, dynamics, evolution and progress for long periods of time, as well as the way people adapted to changes in their environment (Gładykowska-Rzeczycka, 2004-2006; Roberts & Manchester, 2005). It is also used to evaluate the ancient populations’ biological structure, when historical sources provide very little information (Gładykowska-Rzeczycka, 2004-2006). Pathological aspects are induced by the interaction between genetic and non-genetic factors (the latter deriving from the environment). Bone anomalies are changes in the normal bone structure induced by disruptions of the chemical or metabolic functions under the influence of exogenous, genetic or teratogenic factors (Gregg & Gregg, 1987). Bone anomalies can appear before or after birth and they range from less serious forms to major and sometimes lethal anomalies (Barnes, 1994; Masnicová & Beňuš, 2003). Most frequently encountered in the human skeleton are bone fusion, ossification and underdevelopment anomalies and ageneses, all of these identified in numerous prehistoric and historic osteological series around the world (Zimmerman & Kelley, 1982). Non-metric features (also called epigenetic, discontinuous morphological or discrete traits) are forms of variations observed in the bone structure. Non-metric features are hereditary to a certain extent and for that matter they can be used in research concerning ancestral connections; however their genetic foundation is unclear (Carson, 2006). Archaeological research conducted on the premises of the current “Banu” Church of Iași City within an edifice rehabilitation project brought to light a piece of the old church necropolis. The necropolis was discovered and excavated in 2011 by Mrs. Stela Cheptea, PhD and C.S. I archaeologist, and her collaborators at the Centre for European History and Civilization within the Iași Branch of the Romanian Academy. The old church was dated from 1705 (Bogdan, 1997-2004), and it was known at the time under the name of “The Falling Asleep of the Virgin Mary” Church. In his monograph on the city of Iași, N.A. Bogdan (1997-2004) mentions that during the 1650 Kazakh invasion, the wooden church was burnt down and a new majestic Baroque edifice made of stone was built in its stead in 1800 – the current “Banu” Church. According to the data provided by the authors of the excavations, the necropolis was used between the first half of the 16th century and the beginning of the 19th century. This study intends to analyze the incidence and characteristics of specific pathologies, anomalies and non-metric features observed in the osteological series exhumed from the necropolis located on the premises of the “Banu” Church of Iași.

Material and Methods The osteological material consisting of 67 skeletons (children, teenagers, adults, mature and senile) comes from both numbered tombs (individual for the most part – skeletons noted with “M”), but mostly from reinhumation tombs (skeletons noted with “R”). In the total of 67 skeletons, there were 18 children, three teenagers, two adults, 40 mature and only four senile people. In most cases, the precarious state of preservation, as well as the skeleton deterioration during excavations, impeded us to abide strictly by the analysis stages. The bone fragments were subjected to a thorough morphoscopic analysis, which determined each skeleton’s age and gender, anomalies, pathologies and the epigenetic features of the cranial and post-cranial bones. In order to determine the gender, we considered a set of features, using all the bone pieces which provided us with metric and morphological data. Thus, we analyzed the shape of the pelvis, the opening of the greater

- 92 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 sciatic foramen, the sacral curvature, the robustness of the long bones, the development of the joints and muscle insertions, the development of the cranial relief, the forehead’s shape and degree of inclination, the mandible robustness, the shape and type of the mandibular mentonian region and the teeth shape and size. So as to establish the age at death, we used all the criteria recommended by classical methodology. For the subjects under 20, we examined: the primary teeth eruption, the permanent teeth eruption, the dental buds stages of development, the long bones epiphyses fusion with the respective dyaphises and the fusion of the epiphyseal discs with the vertebrae. For the subjects under 20 (infans I, infans II and juvenis), the age at death was established according to the methodology suggested by Maresh (1955, 1970), Moorrees et al., (1963), Trotter & Peterson (1969), Fazekas & Kosa (1978), Ubelaker (1979), Jeanty (1983) and Schaefer et al., (2009). For the subjects past 20, the features we used to determine the age at death were: changes in the symphyseal surface and the sacropelvic surface of ilium, changes in the spongy tissue found in the epiphyses of the humerus and femur, certain skeletal involution phenomena and the existence of pathological processes that can be associated with aging, cranial suture obliteration and masticatory surface abrasion. The gender and age at death for the subjects past 20 (adultus, maturus şi senilis) was determined using the methods, criteria and techniques recommended by Ferembach et al., (1979), Ubelaker (1979), Brothwell (1981), Buikstra & Ubelaker (1994), Bruzek (2002), Walrath et al., (2004) and Schmitt (2005). Pathologies, anomalies, and cranial and post-cranial non-metric features were subjected to a thorough analysis. In order to identify them, we used the methods, criteria and techniques recommended by Mays (1998), Waldron (2009), Ortner (2003), Aufderheide & Rodriguez-Martin (1998) and Barnes (1994).

Results and Discussion In the total of 67 skeletons subjected to the study, we identified 21 subadults (infans I, infans II and juvenis) and 46 subjects past the age of 20 (adultus, maturus and senilis). Subadult mortality rate, including the subjects between 0 and 20 years, has significant values (17.19% for infans I, 8.96% for infans II and 4.48% for juvenis), which means that more than one third of the population (31.35%) didn’t reach adulthood. In the interval 20-x years, the highest number of deaths was recorded in the maturus category (59.70-32.84% for men and 26.87% for women), whereas in the adultus and senilis categories, the mortality rate is considerably lower (adultus: 2.99% for men; senilis: 5.97-4.48% for men and 1.49% for women). The analysis of mortality by gender revealed a higher male mortality rate (29 men as opposed to 20 women, with a masculinity index of 1.45). This disparity was observed in all the age groups (juvenile, adult, mature and senile). The incidence of osteopathies in the osteological series discovered on the premises of the “Banu” Church of Iași (16th-19th centuries) is presented in Table 1. Values were calculated both for the two genders (29 men and 20 women) and for the entire lot (67 subjects). Due to the precarious preservation of the subadults and the fact that certain osteopathies become obvious later in life, we only identified a child supratrochlear foramen of the humerus.

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In the cranial segment (for the entire osteological series), we identified wormian bones and metopic suture (with equal incidence – 2.98%), followed by porotic hyperostosis, cribra orbitalia and premature sagittal suture closure (also with equal incidence – 1.49%) (Table 1). Porotic hyperostosis, also known as exocranial porosity or external cribra cranii, appears due to abnormal increase in the bone tissue on the exocranium. The presence of porotic hyperostosis can allude to various diseases, such as: infantile scurvy, rickets, iron deficiency anemia, osteitis, osteomyelitis, periostitis, inflammatory processes of the cranial bones and skull or post-mortem osseous changes (Ortner, 2003; Walker et al., 2009). Porotic hyperostosis is one of the most frequently studied indicators of the subjects (skeletons) state of health, also providing valuable information about the environmental conditions during growth and development (Vercellotti et al., 2009). In the skeletal series discovered on the premises of the “Banu” Church of Iași, porotic hyperostosis was identified in a single skull (1.49% of the total population), which belonged to a female aged circa 30-35 years (Fig. 1).

Table 1. Incidence of pathologies, anomalies and non-metric bone features in the skeletal serie exhumed from the necropolis of the “Banu” Church of Iași (16th-19th centuries). Men Women Children Total Pathologies/anomalies/ (14-x years) (14-x years) (0-14 years) non-metric features N (%) N (%) N (%) N (%) porotic hyperostosis* - - 1/20 5 - - 1/67 1.49 cribra orbitalia* 1/29 3.44 - - - - 1/67 1.49

premature sagittal suture closure** 1/29 3.44 - - - - 1/67 1.49

metopic suture*** 1/29 3.44 1/20 5 - - 2/67 2.98 segment Cranial Cranial wormian bones*** 2/29 6.89 - - - - 2/67 2.98 osteoarthritis (Schmorl’s nodules, 5/29 17.24 - - - - 5/67 7.46 osteophytes, bone erosion)* spina bifida occulta** 2/29 6.89 - - - - 2/67 2.98

cranial cranial vertebral spondylolysis** 1/29 3.44 - - - - 1/67 2.98 -

segment supratrochlear foramen of 2/29 6.89 1/20 5 1/18 5.55 4/67 5.97 Post humerus*** extra facets on tibiotalar joint*** 5/29 17.24 1/20 5 - - 6/67 8.98 * Pathologies; ** anomalies; *** non-metric features.

Figure 1. Porotic hyperostosis in parietal bones (R12, ♀, 30-35 years).

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Another pathology we identified in the cranial segment is cribra orbitalia (exocanian orbital porosity) which appears as a point-like corrosion of the external compact layer of the orbital roof and the thickening of the spongy bone layer (Kozak & Krenz-Niebala, 2002). The presence of this porosity alludes to an iron deficiency in the organism, due to insufficient dietary intake of iron or an environment laden with pathogenic agents (Stuart-Macadam, 1992). In the case in which there is no additional proof to support this deficiency (such as a histological survey), experts recommend caution when establishing a final diagnosis of iron deficiency anemia. Thus, in the absence of exact diagnosis methods, other than simple morphoscopic examination, cribra orbitalia is a non- specific indicator of diseases (Aufderheide & Rodriguez-Martin, 1998; Wapler et al., 2004). In some cases, we encountered both cribra orbitalia and porotic hyperostosis in the same subject, which is a solid argument supporting the theory that the two affections are consequences of the same systemic problem. Cribra orbitalia is more frequently encountered than porotic hyperostosis. If cribra orbitalia represented an earlier and less serious expression of the pathological process which also determines porotic hyperostosis, it could explain the much higher incidence of this pathology (Walker et al., 2009). In the present study, cribra orbitalia was identified in a single subject (1.49% of the total population) – a teenager aged 16-18 (Fig. 2).

Figure 2. Cribra orbitalia in the right orbit (M 37-A, ♂, 16-18 years).

Premature fusion of the cranial sutures (agenesis). Craniosynostosis appears between the ages of 30 and 40 years on the internal surface and 10 years later on the external surface. The fusion normally begins in the bregma point and then expands successively to the sagittal, coronal and lambdoid suture. Thus, craniosynostosis (premature fusion of the cranial sutures) can be considered a normal process which appears at an abnormally early age. Craniosynostosis or suture agenesis can appear as an isolated case or as part of polytropic syndromes (Aufderheide & Rodriguez-Martin, 1998). The connection between cranial malformations and craniosynostoses was suspected from the beginning of the 19th century and throughout the next decades and it became obvious that some of these craniosynostoses were associated with other congenital anomalies (David et al., 1982). In the total population we analyzed, premature fusion of the sagittal suture was identified in a single case (1.49%), in a male aged 18-20 years (Fig. 3). Metopic suture is located between the frontal bones, from the anterior point of the anterior fontanelle to the glabella. The fusion of the metopic suture begins from the glabella, advances progressively to the upper area and ends at the anterior fontanelle

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(Weinzweig et al., 2003). Metopic suture normally closes between the first and the second year of life and is usually completely fused by the age of three, but it can sometimes remain open until the age of seven. Also, in some cases metopic suture remains open throughout life and it can be identified even in elders (Ide et al., 2003; Nakatani et al., 1998). In the studied osteological series, metopic suture was identified in two subjects (2.98% of the total population), specifically a male aged 40-45 years (Fig. 4) and a mature female (50-55 years) (Fig. 5).

Figure 3. Skull, vertical norm, premature fusion of the sagittal suture (R 5, ♂, 18-20 years).

Figure 4. Frontal bone, metopic suture Figure 5. Frontal bone (incomplete), metopic (R 9, ♂, 40-45 years). suture (R 17, ♀, 50-55 years).

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Wormian bones, also called intersutural bones or Inca bones, are small bones that appear within cranial sutures. Their number and shape can vary from one person to another. They are present in the frontal and occipital bones and in some cases they lead to erroneous diagnoses of the cranial fractures (Nayak, 2008). Wormian bones can be a common occurrence and they seem to be genetically conditioned in some populations (Kaplan et al., 1991). Although the mechanisms responsible for the appearance of wormian bones are unknown, some studies indicated that their presence can be useful in identifying anomalies of the skull and central nervous system (Pryles & Khan, 1979; Das et al., 2005). In the total number of subjects we analyzed, wormian bones (within the lambdoid suture) were identified in two males (2.98% of the population) aged between 35 and 50 years (Figs 6; 7).

Figure 6. Two wormian bones within the Figure 7. Two wormian bones within the lambdoid lambdoid suture (one bone 23.5 mm long and suture (M 29, ♂, 50 years). 12.5 mm wide) (R 8, ♂, 35-40 years).

In the post-cranial segment (for the entire osteological series), the highest incidence was recorded by extra facets on the tibiotalar joint (8.98%), osteoarthritis (7.46%) and supratrochlear foramen of the humerus (5.97%). Spina bifida occulta and spondylolysis had lower incidences (2.98% each) (Table 1). An important observation is that the female gender appears less pathologically conditioned in comparison with the male gender, regardless of the skeletal segment (cranial or post-cranial). This claim has some reservations due to the smaller number of female subjects (the studied series consists of 67 subjects and the sex ratio is 29/20). Osteoarthritis is a degenerative joint disease caused by cartilage loss in a joint. This leads to lesions due to direct contact between the bones and in the attempt to repair the lesion, the joint’s bone reacts by producing another bone (Aufderheide & Rodriguez- Martin, 1998). Osteoarthritis generally affects the hip, hand, knee, lumbar and neck joints. We noticed that after the age of 50, women are much more frequently affected by osteoarthritis in comparison with men (Lawrence et al., 2008). The cause determining osteoarthritis is yet unknown, however there are certain factors which amplify the risk of developing this disease, such as: heredity, overweight, joint lesions, repeated overstrain in certain joints, lack of physical activity, nervous lesions and aging. The study of a genealogical database concerning the population of Iceland revealed that hand osteoarthritis and hip osteoarthritis

- 97 - Vasilica-Monica Groza et al. are strongly influenced by genetic inheritance, spreading over several generations (Jónsson et al., 2003). In the analyzed skeletal series, osteoarthritis was identified in five male subjects (aged 40-65 years) (7.46% of the total population), being mainly represented by osteophytes (Figs 8; 9), Schmorl’s nodules (Fig. 10) and bone porosity (corrosion) (Figs. 11; 12).

Figure 8. Dorsal vertebrae, Figure 9. Left cubitus, osteophytes Figure 10. Dorsal vertebrae, marginal osteophytes in the uperior epiphysis (R 1(B), Schmorl’s nodules (M 29, ♂, (M 11, ♂, 45-50 years). ♂, 40-45 years). 50 years).

Figure 11. Left tibia, bone porosity Figure 12. Humeri, porosity (corrosion) (corrosion) (R 10, ♂, 55-60 years). in the shoulder joint area (M 43, ♂, 60-65 years).

Spina bifida occulta (occult spinal dysraphism) Spina bifida (spinal dysraphism, spinal defect, neural tube defect, open spine) includes all forms of congenital spine abnormalities resulting in a faulty neural arch which allows the meninx or neural elements to herniate (Dawson, 2006). Spina bifida has multifactor origins, including a genetic predisposition and environmental factors; nevertheless, the exact cause remains unknown (Windham & Bjerkedal, 1982). In the past decade, scientists noticed that folic acid, zinc and selenium deficiencies in the maternal metabolism can lead to neural tube defects in the embryo. The three nutritional substances

- 98 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 are necessary for the genetic control of cell growth during morphogenesis (Barnes, 1994). Consequently, maternal diet, exposure to teratogenic factors and genetic predisposition acting together can lead to spinal dysraphism. Occult spinal dysraphism can be located on any spinal segment, starting at the atlas and ending at the sacrum; however, it is most frequently encountered in the lumbosacral region (L5-S1), recording incidences up to 25% (Barnes, 1994). Epidemiologic studies have revealed a wide variation in the prevalence of spina bifida based on ethnicity, race, geography and temporal trends (Josan et al., 2008). Paleoanthropological research concerning spina bifida was centered on the sacrum bone. Sacral spina bifida occulta was frequently identified and reported by paleopathologists as a congenital anomaly (Kumar & Shane Tubbs, 2011). Sacral spina bifida occulta (sacral occult spinal dysraphism) was recorded in two male subjects (2.98% of the total population) aged between 18-55 years, in the segments S4-S5 and S3-S5, respectively (Figs 13; 14).

Figure 13. Sacrum, sacral spina bifida occulta Figure 14. Sacrum, spina bifida occulta (segment S4-S5) (R 5,♂, 18-20 years). (segment S3-S5) (M 45, ♂, 50-55 years).

Vertebral spondylolysis Spondylolysis can be described as an ossification failure in the pars interarticularis area of the vertebra, leading to its separation in two parts: a ventral part consisting of the vertebral body and a dorsal part represented by the spinous process and the inferior articular process (Aufderheide & Rodriguez-Martin, 1998). L3-L5 lumbar vertebrae, respectively the first sacral vertebra (S1) are the most frequently affected. Bilateral spondylolysis is most frequently encountered in males (Barnes, 1994). In the studied series, lumbar spondylolysis in the L4 vertebra was identified in a single case (2.98% of the total population), specifically a male aged 45-50 years (Fig. 15). Supratrochlear foramen of the humerus is considered by some authors an atavism (Mahajan, 2011), whereas others (Singhal & Rao, 2007) see it as a result of bone atrophy after ossification, induced by the mechanical pressure of the olecranon process over the supratrochlear area of the humerus, resulting in the atrophy of the olecranon process. In the studied series, supratrochlear foramen of the humerus (olecranian

- 99 - Vasilica-Monica Groza et al. perforation) was identified in four subjects (5.97% of the total population), specifically two men (aged 18-40 years) (Figs 16; 17), a woman (aged around 60) (Fig. 18) and a child (13- 14 years) (Fig. 19).

Figure 15. Lumbar vertebra (L4), bilateral spondylolysis (R 3, ♂, 45-50 years).

Figure 16. Humeri, supratrochlear foramen Figure 17. Left humerus, supratrochlear (R 5, ♂, 18-20 years). foramen (M 23, ♂, 35-40 years).

Figure 18. Left humerus, supratrochlear foramen (M Figure 19. Right humerus, supratrochlear 33, ♀, 60 years). foramen (R23, 13-14 years).

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Extra facets on the tibiotalar joint are caused, according to some authors (Tulsi & Prasada Rao, 1968) by mechanical stress (the habit of squatting), whereas others (Satinoff, 1972) see it as a post-cranial epigenetic trait. The study of heredity revealed that the squatting facet can be seen on the tibia and the talus during the fetal period and during childhood. With age, if squatting is no longer part of the daily routine, the associated facet disappears. Consequently, the main reason which determines the appearance of this facet is represented by the frequent squatting movements (Baykara et al., 2010). Extra facets on the inferior tibial epiphyses were identified in five men aged 35-60 years (Figs. 20-24) and a woman aged 55-60 years (8.98% of the total population) (Fig. 25).

Figure 20. Tibiae, extra facets on the Figure 21. Right tibia, Figure 22. Tibiae, extra facets on joint (M 23, ♂, 35-40 years). extra facet on the joint the joint (M 29, ♂, 50 years). (M 39, ♂, 40 years).

Figure 23. Left tibia, extra Figure 24. Left tibia, extra Figure 25. Tibiae, extra facets on the facets on the joint (R 16 facets on the joint joint (R 10, ♂, 55-60 years). (A), ♂, 50-55 years). (R 10, ♂, 55-60 years).

Conclusions The osteological material analyzed in this study was exhumed in 2011 under the guidance of the archaeological team from the Centre for European History and Civilization of Iasi. Archaeological excavations brought to light 67 human skeletons found in inhumation tombs (individual or reinhumations). This skeletal series represents a small segment of the population inhabiting the city of Iași during the 16th-19th centuries.

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The distribution by gender of the 67 skeletons revealed a significant child mortality rate (0-14 years: about 27%), a maximum incidence of death in the mature range (30-60 years: about 60%) and a relatively low mortality rate after the age of 60. We observed a higher male mortality rate (43.28% for men as opposed to 29.85% for women). Paleopathological aspects, linked to the main paleodemographic indicators, can provide valuable information about the environmental and socioeconomic conditions in which this population performed its daily activities. If we refer to the entire skeletal series, the main indicators of the state of health in the cranial segment are cribra orbitalia and porotic hyperostosis, with equal incidence (1.49%). The presence of these exocranial porosities is frequently used as an instrument to evaluate the state of health and the nutritional status of the past populations; at present, they are also regarded as potential indicators of the environmental conditions. Among the types of anomalies, we identified premature fusion of the cranial sutures (craniosynostosis produced at an abnormally early age), with an incidence of 1.49%. Non-metric (epigenetic) features such as wormian bones and metopic suture identified in the cranial segment (with equal incidence – 2.98%) can be inherited and for that matter they can be used in studies concerning the biological affinity of the human populations from the past. In the post-cranial segment of the analyzed series, osteoarthritis related affections (Schmorl’s nodules, osteophytes, bone corrosion – 7.46%) were identified exclusively in males aged between 40 and 65 years, being mainly located in the spine and the joints of appendicular skeleton. The main factors responsible for this affection are age and genetic predisposition; however certain physical activities also play an important role in the matter. The presence of non-metric features such as the supratrochlear foramen of the humerus (5.97%) and extra facets on the tibiotalar joint (8.98%), indicates that the subjects practiced intense physical activities which strained their arms (repetitive flexion and extension movements) and that they spent a lot of time in the squatting position or doing many repetitive genuflections. The only anomalies identified in the post-cranial segment for this population were spina bifida occulta and vertebral spondylolysis, with equal incidence (2.98%). An important observation is that the male gender appears more pathologically conditioned in comparison with the female gender, regardless of the skeletal segment (cranial or post-cranial).

Acknowledgements We thank Mrs. Stela Cheptea, PhD (C.S. I archaeologist at the Centre for European History and Civilization, Iasi) for the osteological material made available for the anthropological study.

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Bruzek, J., 2002. A method for visual determination of sex, using the human hip bone. American Journal of Physical Anthropology, 117: 157-168. Buikstra, J.E., Ubelaker, D.H., 1994. Standards for Data Collection from Human Skeletal Remains. Arkansas Archaeological Survey Research Series No 44, Fayetteville. Carson, E.A., 2006. Maximum – likelihood variance components analysis of heritabilities of cranial nonmetric traits. Human Biology, 78 (4): 383-402. Das, S., Suri, R., Kapur, V., 2005. Anatomical observations on os inca and associated cranial deformities. Folia Morphologica, 64: 118-121. David, J.D., Poswillo, D., Simpson, D., 1982. The Craniosynostoses. Causes, Natural History and Management. New York, Heidelberg, Berlin: Springer-Verlag. Dawson, F., 2006. Spinal Dysraphism. Medical Bulletin, 11 (12): 7-11. Fazekas, I.Gy., Kosa, F., 1978. Forensic Fetal Osteology. Budapest Akademiai Kiado. Ferembach, D., Schwidetzky, I., Stloukal, M., 1979. Recommandations pour determiner l’age et le sexe sur le squelette. Bulletins et Memoires de la Societe d’Anthropologie de Paris, XIII, 6, 1: 7-45. Gładykowska-Rzeczycka, J.J., 2004-2006. Palaeopathology in Poland at the beginning of the 21st century. Studies in Historical Anthropology, 4: 25-48. Gregg, J.B., Gregg, P.S., 1987. Dry Bone. Dakota Territory Reflected. Sioux Printing. Ide, Y., Inukai, Y., Yoshida, S., Sato, I., 2003. The internal structure of bony tissue of a human metopic suture by Soft X-ray. Okajimas Folia Anatomica Japonica, 79: 169-173. Jeanty, P., 1983. Fetal limb biometry. Radiology, 147: 601-602. Jónsson, H., Manolescu, I., Stefansson, S.E., Ingvarsson, TH., Jonsson, H.H., Manolescu, A., Gulcher, J., Stefansson, K., 2003. The inheritance of hand osteoarthritis in Iceland. Arthritis Rheumatism, 48: 391- 395. Josan, V., Morokoff, A., Maixner, W.J., 2008. Epidemiology and aetiological factors. In: Memet Ö.M., Cinalli G., Maixner W.J. (coord.): The Spina Bifida. Management and Outcome. Springer-Verlag Press, 59-65. Kaplan, S.B., Kemp, S.S., Oh, K. S., 1991. Radiography manifestations of congenital anomalies of the skull. Radiology Clinics of North America, 29(2): 195-218. Kozak, J., Krenz-Niebała, M., 2002. The occurrence of cribra orbitalia and its association with enamel hypoplasia in a medieval population from Kolobrzeg, Poland. Variability and Evolution, 10: 75-82. Kumar, A., Shane Tubbs, R., 2011. Spina bifida: a diagnostic dilemma in paleopathology. Clinical Anatomy, 24: 19-33. Lawrence, R.C., Felson, D.T., Helmick, C.G., Arnold, L.M., Choi, H., Deyo, R.A., Gabriel, S., Hirsch, R., Hochberg, M.C., Hunder, G.G., Jordan, J.M., Katz, J.N., Maradit Kremers, H., Wolfe, F., 2008. National Arthritis Data Workgroup. Estimates of the prevalence of arthritis and other rheumatic conditions in the United States. Part II, Arthritis and Rheumatism, 58 (1): 26-35. Mahajan, A., 2011. Supratrochlear foramen; Study of humerus in North Indians. Professional Medical Journal, 18 (1): 128-132. Mays, S., 1998. The archaeology of human bones. Ed. Routledge. Maresh, M.M., 1955. Linear growth of long bones of extremities from infancy through adolescence. American Journal of Diseases of Children, 89: 725-742. Maresh, M.M., 1970. Measurements from roentgenograms. In: McCammon, R.W. (ed.): Human Growth and Development. Springfield, IL: C.C.Thomas. Masnicová, S., Beňuš R., 2003. Developmental Anomalies in Skeletal Remains from the Great Moravia and Middle Ages Cemeteries at Devin (). International Journal of Osteoarchaeology, 13: 266-274. Moorrees, C.F.A., Fanning, E.A., Hunt, E.E., 1963. Age variation of formation stages for ten permanent teeth. Journal of Dental Research, 42: 1490-1502. Nayak, S.B., 2008. Multiple Wormian bones at the lambdoid suture in an Indian skull. Neuroanatomy, 7: 52-53. Nakatani, T., Tanaka, S., Mizukami, S., 1998. A metopic suture observed in a 91-year-old Japanese mal. Kaibogaku Zasshi, 73: 265-267. Ortner, D.J., 2003. Identification of Pathological Conditions in Human Skeletal Remains. Elsevier Academic Press. Pryles, C.V., Khan, A.J., 1979. Wormian bones. A marker of CNS abnormality?, American Journal of Diseases of Children, 133: 380-382. Roberts C., Manchester, K., 2005. The Archaeology of Disease. Third Edition, Sutton Publishing Limited, United Kingdom. Satinoff, M.I., 1972. Study of the squatting facets of the talus and tibia in ancient Egyptians. Journal of Human Evolution, 1 (2): 209-212. Schaefer, M., Black, S., Scheuer, L., 2009. Juvenile osteology, Elsevier Academic Press.

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ODONTOPATHIES IN THE POPULATION OF OLD IAȘI CITY (ROMANIA): THE NECROPOLIS OF THE “BANU” CHURCH, 16TH-19TH CENTURIES

Vasilica-Monica GROZA1, Angela SIMALCSIK1 and Luminiţa BEJENARU1, 2 1 Romanian Academy-Iasi Branch, Department of Anthropological Research, Str. Th. Codrescu, no. 2, Iaşi, Romania, [email protected], [email protected] 2 “Alexandru Ioan Cuza” University, Faculty of Biology, Bd. Carol I, 20A, Iaşi, 700505, Romania

Abstract. This study describes the incidence and the analysis of a series of odontopathies observed in a skeletal series dating from the 16th-19th centuries, found in the necropolis of the ancient “Banu” Church of Iași City (Romania). The unearthed material is mostly derived from reinhumation tombs and includes 67 skeletons, out of which 18 children, three adolescents 2 adults, 40 mature and 4 senile. The distribution by gender indicates a higher male mortality rate (29 men as opposed to 20 women). Among the odontopathies observed in the 67 subjects, dental caries, radicular remains, supragingival dental calculus and edentia have equal incidence – 5.97%, whereas microdontia recorded 1.49%.

Keywords: Iași City, necropolis, 16th-19th centuries, “Banu” Church, odontopathies

Rezumat. Odontopatii la populaţia vechiului oraș Iaşi (România): necropola Bisericii „Banu”, secolele XVI- XIX. În studiul de faţă autorii prezintă incidenţa şi analiza unor odontopatii semnalate la o serie osteologică care datează din secolele XVI-XIX, deshumată din necropola vechii Biserici „Banu” din Iaşi. Materialul deshumat, provenit în mare parte din morminte cu reînhumări, este reprezentat prin 67 de schelete, dintre care 18 aparţin copiilor, trei adolescenţilor, două adulţilor, 40 maturilor şi patru senililor. Repartiţia pe sexe, indică o rată de mortalitate mai ridicată a bărbaţilor faţă de femei (29 bărbaţi faţă de 20 de femei). Dintre odontopatiile semnalate la cei 67 de subiecţi, cariile dentare, resturile radiculare, calculusul dentar supragingival şi edentaţia prezintă procente egale – 5,97%, iar microdonţia înregistrează un procent de 1,49%.

Cuvinte cheie: orașul Iași, Biserica Banu, necropolă, secolele XVI-XIX, odontopatii

Introduction Dental remains, the hardest and most chemically stable tissues in the organism (Roberts & Manchester, 2005), provide valuable information about a population’s state of health and nutrition (Turner 1979; Lukacs, 1992). In an anthropological study, having knowledge about the tooth disorders, as well as the dental pathology in general, has great significance in establishing a connection between these disorders and the culture of the age. Dental paleopathology identifies and explains tooth and jaw anomalies in the past populations. The analysis of the dental pathologies and anomalies is based on two research traditions. The first one refers to the relation between tooth diseases and cultural factors, such as nutrition and subsistence diet, whereas the second targets developmental dental anomalies, which are more significantly influenced by genetic factors. Ethnographic documentation and information concerning diet and food preparation are requisite in order to better understand the incidence of the various tooth pathologies (Lukacs, 2012). The use of teeth as instruments in a wide area of activities non- related to diet is improved and amplified by ethnographic research (Larsen, 1985; Brown & Molnar, 1990).

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Infectious diseases, such as caries, are some of the most common affections observed in archeological populations, whereas the jaw degenerative diseases include ante- mortem tooth loss, usually in elders. Development problems include enamel hypoplasia and genetic anomalies (Roberts & Manchester, 2005). We remarked that tooth disorders don’t develop separately, but there is a complex relation between them (Lukacs, 1989). Archeological excavations conducted in 2011 by the Centre for European History and Civilization, Iasi Branch of the Romanian Academy, under the guidance of C.S. I archaeologist PhD Mrs. Stela Cheptea, on the premises of the current “Banu” Church of Iași City (Romania), brought to light a part of the necropolis of the ancient church, dating from 1705 (Bogdan, 1997-2004). The authors of the diggings indicate that the necropolis was used between the first half of the 16th century and the beginning of the 19th century. This study intends to analyze the prevalence and characteristics of specific odontopathies (dental caries, radicular remains, dental calculus/scale, edentia and microdontia) observed in the osteological series exhumed from the necropolis located on the premises of the “Banu” Church of Iași City.

Material and Methods The osteological material consists of 67 skeletons (children, teenagers, adults, mature and senile) coming from both numbered tombs (individual for the most part – skeletons noted with “M”), but mostly from reinhumation tombs (skeletons noted with “R”). Of the total of 67 skeletons, 46 belonged to adults, 18 to children and only three belonged to teenagers. The precarious state of preservation, as well as the skeleton deterioration during excavations, impeded us to abide strictly by the analysis stages. The study of the osteological material was preceded by a process marking and restoration of the main segments of each skeleton using the available fragments, as well as a process of filling up the missing sections with a moldable mass. This allowed us to proceed afterwards to the bio-morphoscopic analysis in order to determine each subject’s age and gender. So as to establish the gender and age at the time of death for the subjects over 18 years, we used the methods, criteria and techniques recommended by Ferembach et al., (1979), Ubelaker (1979), Brothwell (1981), Buikstra & Ubelaker (1994), Bruzek (2002), Walrath et al., (2004) and Schmitt (2005). As concerns the subadults, the age at death was determined based on the primary teeth eruption, the permanent teeth eruption, the dental buds stages of development, the epiphyses fusion with the respective dyaphises and the fusion of the epiphyseal discs with the vertebrae. For the subjects under 20 years (infans I, infans II and juvenis), the age at death was established according to the methodology suggested by Maresh (1955, 1970), Moorrees et al., (1963), Trotter & Peterson (1969), Fazekas & Kosa (1978), Ubelaker (1979), Jeanty (1983) and Schaefer et al., (2009). Since teeth are considered human remains which are very well preserved over time, they can provide information about the health state or the lifestyle of a past population. In this study we confined ourselves to macroscopic observations. In order to identify dental pathologies, we cleaned the teeth (found in the alveoli or lost post mortem) using a soft brush, to eliminate the particles that would have impeded visual inspection. Caries (when perforations affected the dentin) were recorded based on their position and gravity. We identified the cases of edentia (partial or total absence of

- 106 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 teeth in the oral cavity due to falling after eruption caused by several factors) (Ionescu, 2005). We also observed the presence of radicular remains in the upper and lower jaw. We identified dental calculus or scale (grey-white mineralized plaque composed primarily of calcium phosphate), firmly attached to the dental surface. Depending on the positioning on the tooth crowns or exposed roots, dental calculus can be supragingival or subgingival (Waldron, 2009). A tooth anomaly observed in this population was microdontia, a condition in which one or several teeth appear smaller than normal (for example, short and narrow roots and crowns) (Lyngstadaas et al., 1996).

Results and Discussion Considering the total 67 skeletons subjected to study, we are first and foremost struck by the relatively high child mortality rate (0-14 years: 26.87%), which shows that approximately a quarter of the population died before reaching adolescence. If we add to this the teen mortality rate (14-18 years: 4.48%), we observe that more than one third of the subjects (31.35%) didn’t reach adulthood. The highest mortality rate is associated with the age interval 0-7 years (17.91%), whereof half of the skeletons belonged to the interval 0-1 years (6 out of 12 children). Among the population that reached at least adulthood, (20-x years: approximately 68.66%), 59.70% of the subjects died in full maturity (32.84% males and 26.87% females) and a were 5.97% (4.48% males and 1.49 females) reached the old age (60-x years); only 2 deaths were recorded in the adult stage (2.99%, both of them males). The analysis of mortality by gender revealed a higher male mortality rate (29 men as opposed to 20 women, with a masculinity index of 1.45). This disparity was observed in all the age groups (juvenile, adult, mature and senile). The presence of odontopathies in the osteological series discovered on the premises of the “Banu” Church of Iași (16th-19th centuries) is rendered in Table 1. The incidence was calculated for the two genders (29 men and 20 women), as well as for the entire lot of subjects (67). Among the odontopathies observed, edentia, dental caries, dental calculus and radicular remains had equal incidence – 5.97%, whereas microdontia had the lowest incidence at 1.49% (Table 1).

Table 1. Incidence of odontopathies in the osteological series exhumed from the necropolis of the “Banu” Church of Iași (16th-19th centuries). Men Women Children Total (14-x years) (14-x years) (0-14 years) Odontopathies N (%) N (%) N (%) N (%) Dental caries 3/29 10.34 1/20 5 - - 4/67 5.97 Radicular remains 3/29 10.34 1/20 5 - - 4/67 5.97 Dental calculus/scale 3/29 10.34 1/20 5 - - 4/67 5.97 Edentia 3/29 10.34 1/20 5 - - 4/67 5.97 Microdontia 1/29 3.44 - - - - 1/67 1.49

Dental caries has a multifactor etiology, presenting various degrees of gravity, from opaque stains to large cavities affecting the teeth (Roberts & Manchester, 1995). The spreading of this disease in historical populations is undoubtedly associated with a carbohydrate-rich diet (Cucina et al., 2011). This suggests that carbohydrate-rich foods are associated with a wider spreading of dental caries, the latter leading to physiological

- 107 - Vasilica-Monica Groza et al. changes of the oral cavity and changes of the bacterial flora in the oral cavity. Consequently, bioarchaeological studies use the incidence of dental caries as a nonspecific indicator of the eating behavior (Temple, 2011). Powell (1985) indicates that the main factors influencing dental caries are: environmental factors (oligoelements present in food and water), pathogenic agents (bacteria causing the disease), exogenous factors (diet, oral hygiene) and endogenous factors (teeth shape and structure). As regards the incidence of dental caries, some researchers noticed that it is much higher in female subjects as opposed to male subjects (Fujita et al., 2007; Temple & Larsen, 2007), due to the fact that men eat bigger amounts of non-cariogenic foods, whereas women eat larger quantities of cariogenic carbohydrates (Temple, 2011). There are authors who suggest that people belonging to a higher social class have access more easily to cariogenic foods, including exotic products containing high levels of sucrose (Cucina & Tiesler, 2003). Alternatively, there is the belief that the upper class can afford to eat more meat proteins, which would consequently lead to less caries (Larsen, 1997). As a specific trait of the ancient populations we studied, mandibular teeth – mainly lateral teeth – are generally less resistant than maxillary teeth. In the studied series, dental caries was observed in four subjects, more specifically three men aged between 22 and 50 (Figs. 1-3) and one women aged 35-40 years (Fig. 4). In three of the cases, dental caries affected the mandibular first molars, whereas in one case it affected the mandibular second molars.

Figure 1. Mandible, occlusal caries, gr. III-IV (left Figure 2. Mandible, approximal caries, gr. II and right second molar) (R 2, ♂, 22-25 years). (right first molar) (M 36, ♂, 40 years).

Figure 3. Mandible, occluso-approximal caries, gr. Figure 4. Mandible, occlusal caries, gr. II II (left first molar) (M 28, ♂, 45-50 years). (left first molar) (R 18, ♀, 35-40 years).

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Radicular remains. Dental abrasion can be induced both by the use of teeth in the mastication process (which leads to the wearing of the contact surface, due to the disappearance of the tough substance from the teeth) and by tooth pathology (Ştefănescu, 2007). Five degrees of dental abrasion established by Périer are used to highlight the disappearance of the enamel form the dentin (Chira, 1981). The fifth degree is represented by pronounced abrasion, leading to the disappearance of the crown, which makes visible the pulp chamber. This is how radicular remains result. Radicular remains were identified in three male subjects (aged between 22 and 65, Figs. 5-7) and one female subject (40-45 years), affecting the mandibular and maxillary teeth (Fig. 8).

Figure 5. Upper jaw, radicular remain (second Figure 6. Mandibular fragment, radicular premolar and first molar on the left). remain (right first molar) (R 2, ♂, 22-25 years). (R 8, ♂, 35-40 years).

Figure 7. Upper jaw fragment, radicular remain Figure 8. Upper jaw fragment, radicular (right first premolar) remain (left second premolar) (R 16(B), ♀, (R 15(A), ♂, 60-65 years). 40-45 years).

Dental calculus/scale. Dental calculus (scale) appears in the form of a mineralized plaque composed primarily of calcium phosphate. Depending on its localization, either on

- 109 - Vasilica-Monica Groza et al. the tooth crown or the exposed roots, there are two forms of calculus: supragingival and subgingival (Waldron, 2009). According to Waldron (2009), there is an inverse relationship between calculus and caries, since calculus needs an alkaline environment to develop, whereas caries develop in an acidic environment, which leads to the logical conclusion that the two processes are incompatible. Dental calculus appears most frequently on the teeth located closest to the salivary glands (especially mandibular incisors and maxillary molars) (Roberts & Manchester, 2005). In the analyzed osteological series, supragingival dental calculus was identified in five cases, more specifically four men aged between 18 and 50 (Figs. 9-11) and a mature woman (35-40 years) (Fig. 12).

Figure 9. Mandible fragment, supragingival Figure 10. Mandible, supragingival dental calculus dental calculus (C, P1, P2, M1 – right (I1, I2, C – right hemiarcade; I2, C, P1 – left hemiarcade) (R 3, ♂, 45-50 years). hemiarcade) (R 5, ♂, 18-20 years).

Figure 11. Mandible, supragingival dental Figure 12. Mandible fragment, supragingival dental calculus (C – left hemiarcade) calculus (I1, I2, C, P1, P2, M1, M2 – right (R 2, ♂, 22-25 years). hemiarcade) (M 26 (B), ♀, 35-40 years).

Edentia refers to the partial or total absence of teeth in the oral cavity, caused by their falling after eruption due to several factors. The main cause for edentia is dental caries

- 110 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 and its complications. There are also other disorders, such as infections of the soft tissues or bone tissues (osteomyelitis), tumors or facial traumas which can lead to edentia (Ionescu, 2005). Edentia can be: frontal (missing incisors and canines, with interlaced spaces), lateral (missing premolars and molars, where the spaces can be unilateral or bilateral), terminal (uniterminal or biterminal, located exclusively anteriorly), mixed (interlaced and terminal spaces), subtotal (with 1-2 remaining teeth), or complete including both the upper and the lower jaw (Ştefănescu, 2007). A secondary effect of edentia is the remodeling of the affected portion of facial skeleton, defined by the bone mass reduction through resorption and atrophy; this causes the lower side of the face to fall, due to the upper jaw and mandible decrease in height and the reduction of the mandibular thickness (Firu, 1967). Edentia was identified in three male subjects aged between 50 and 65 (Figs. 13-15) and a female subject aged 60-65 years (Fig. 16). Edentia is total in all cases, which is the most frequently encountered form after the age of 50.

Figure 13. Mandible, edentia Figure 14. Mandible fragment, edentia (R 15 (A), ♂, 60-65 years). (R 15 (B), ♂, 50-55 years).

Figure 15. Incomplete mandible, edentia Figure 16. Mandible, edentia (M 43, ♂, 60-65 years). (M 32, ♀, 60-65 years).

Microdontia is a tooth anomaly defined by the appearance of one ore several teeth with smaller dimensions than normal (for example: short and narrow roots and crowns).

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This anomaly can affect a single tooth or can be partially or fully generalized, which is rarely encountered in specialized literature. Microdontia can frequently appear in association with hypodontia. Microdontia involving a single tooth is quite common. The most frequently affected teeth are maxillary lateral incisors and third molars (Lyngstadaas et al., 1996). Microdontia is an inherited developmental anomaly, with dominant traits, handed down through crisscrossed inheritance, which means that the micro-dimensioned teeth are inherited from one parent, whereas the over-dimensioned teeth are inherited from the other parent. The incidence of microdontia in a population records values between 1% and 8%. It can appear as the result of a normal eruption or it can be caused by specific genetic disorders, case in which all teeth are micro-dimensioned (Down syndrome or hypophyseal nanism) (Lyngstadaas et al., 1996). In the series we studied, microdontia was identified in the upper jaw (third molar on the right hemiarcade) belonging to a male aged 18-20 (Fig. 17).

Figure 17. Upper jaw, microdontia (third molar, right hemiarcade) (R 5, ♂, 18-20 years).

Conclusions The 67 human skeletons analyzed in this study were discovered in the necropolis exhumed in 2011 on the premises of the “Banu” Church of Iași City. The authors of the excavations date the necropolis between the first half of the 16th century and the beginning of the 19th century. The 67 skeletons belong to 18 children (0-14 years: 26.15%), three teenagers (2 ♂ and 1 ♀: 4.48%), two adults (2 ♂: 2.99%), 40 mature (22 ♂ and 18 ♀: 59.70%) and four senile (3 ♂ and 1 ♀: 5.97%). It is well known that during the Middle Ages there were many highly cariogenic foods which favored the disappearance of the thin enamel coating, leaving the dentin exposed. We also need to consider that the absence of some oligoelements or certain amino acids that are essential in teeth development can decrease teeth resistance, increasing their vulnerability to cariogenic agents. The odontopathies observed in this series mainly affect the male gender; thus, we identified dental caries, radicular remains, supragingival dental calculus and edentia (with equal incidence – 5.97%). As regards dental anomalies, we observed microdontia (defined by the

- 112 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 appearance of one ore several teeth with smaller dimensions than normal) in a single male subject (1.49%).

Acknowledgements We thank Mrs. Stela Cheptea, PhD (C.S. I archaeologist at the Centre for European History and Civilization, Iasi) for the osteological material made available for the anthropological study.

References Bogdan, N.A., 1997-2004. Oraşul Iaşi – Monografie istorică şi socială ilustrată (ediţia a III-a). Editura Tehnopress, Iaşi, 228-229. Brothwell, D.R., 1981. Digging up Bones. Cornell University Press, London. Brown, T., Molnar, S., 1990. Interproximal Grooving, and Task Activity in Australia. American Journal of Physical Anthropology, 81: 545-554. Bruzek, J., 2002. A method for visual determination of sex, using the human hip bone. American Journal of Physical Anthropology, 117: 157-168. Buikstra, J.E., Ubelaker, D.H., 1994. Standards for Data Collection from Human Skeletal Remains. Arkansas Archaeological Survey Research Series No 44, Fayetteville. Chira, I., 1981. Morfopatologia funcțională a aparatului dento-maxilar. Editura Didactică şi Pedagogică, București, 160-161. Cucina, A., Tiesler, V., 2003. Dental caries and ante-marten1 tooth loss in the Northern Peten area, Mexico: a biocultural perspective on social status differences among the Classic Maya. American Journal of Physical Anthropology, 122: 1-10. Cucina, A., Cantillo, C.P., Sosa, T.S., Tiesler, V., 2011. Carious Lesions and Maize Consumption among the Prehispanic Maya: An Analysis of a Coastal Community in Northern Yucatan. American Journal of Physical Anthropology, 145 (4): 560-567. Fazekas, I.Gy., Kosa, F., 1978. Forensic Fetal Osteology. Budapest Akademiai Kiado. Ferembach, D., Schwidetzky, I., Stloukal, M., 1979. Recommandations pour determiner l’age et le sexe sur le squelette. Bulletins et Memoires de la Societe d’Anthropologie de Paris, XIII, 6, 1: 7-45. Firu, P., 1967. Stomatologia infantilă. Ed. Pedagogică, Bucureşti. Fujita, H., Asakura, K., Ogura, M., 2007. Age-and sex-related dental caries prevalence in Japanese from the Jomon period. Journal of Oral Biosciences, 49: 198-204. Ionescu, E., 2005. Anomaliile dentare, Cartea Universitară, Bucureşti. Jeanty, P., 1983. Fetal limb biometry. Radiology, 147: 601-602. Larsen, C. S., 1985. Dental Modification and Tool Use in the Western Great Basin. American Journal of Physical Anthropology, 67(4): 393-402. Larsen, C.S. 1997. Bioarchaeology: interpreting behavior from the human skeleton. Cambridge, Cambridge University Press, 76-77. Lingstadaas, S.P., Nordbo, H., Gedde-Dahl, T., Thrane, P.S., 1996. On the genetics of hypodontia and microdontia: synergism or allelism of major genes in a family with six affected members. Journal of Medical Genetics, 33: 137-142. Lukacs, J.R., 1989. Dental paleopathology: methods for reconstructing dietary patterns. In: Iscan M.Y. and Kennedy K.A.R. (Eds): Reconstruction of life from the skeleton. New York, Alan Liss, 261-286. Lukacs, J.R., 1992. Dental Paleopathology and Agricultural Intensification in South Asia: New Evidence from Bronze Age Harappa. American Journal of Physical Anthropology, 87 (2): 133-150. Lukacs, J.R., 2012. Oral Health in Past Populations: Context, Concepts and Controversies. In: Grauer A.L., (Ed.): A Companion to Paleopathology, First Edition, Blackwell Publishing Ltd., Oxford, United Kingdom, 553-581. Maresh, M.M., 1955. Linear growth of long bones of extremities from infancy through adolescence. American Journal of Diseases of Children, 89: 725-742. Maresh, M.M., 1970. Measurements from roentgenograms. In: McCammon, R.W. (Ed.), Human Growth and Development, Springfield, IL: C.C.Thomas. Moorrees, C.F.A., Fanning, E.A., Hunt, E.E., 1963. Age variation of formation stages for ten permanent teeth. Journal of Dental Research, 42: 1490-1502. Piperno, D., 1988. Phytolith analysis: archaeological and geological perspective. London, Academic Press. Powell, M.L. 1985. The analysis of dental wear and caries for dietary reconstruction. In: Gilbert R.I. and Mielke J.H. (Eds.): Analysis of prehistoric diets. Academic Press, London, 307-338.

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Roberts Ch., Manchester, K., 1995. The Archaeology of Disease. Cornell Publishing Limited, Cornell University Press, Ithaca, New York, 45-47. Roberts Ch., Manchester, K., 2005. The Archaeology of Disease. Third Edition, Sutton Publishing Limited, United Kingdom, 63-84. Schaefer, M., Black, S., Scheuer, L., 2009. Juvenile osteology, Elsevier Academic Press. Schmitt, A., 2005. Une nouvelle methode pour estimer l’age au deces des adultes a partir de la surface sacro- pelvienne iliaque. Bulletine et Memoire de la Societe d’Anthropologie de Paris, 17(1-2): 1-13. Ştefănescu, D.G., 2007. Studiul cariei dentare la populaţia din Moldova aparţinând culturii Sântana de Mureş, sec. III-IV e.n., Teză de doctorat (Rezumat). Universitatea de Medicină şi Farmacie ,,Gr. T. Popa” Iaşi. Temple, D.H., Larsen, C.S., 2007. Dental caries as evidence for agriculture and subsistence variation during the Yayoi period in prehistoric Japan: biocultural interpretations of an economy in transition. American Journal of Physical Anthropology, 134: 501-512. Temple, D.H., 2011. Variability in Dental Caries Prevalence between Male and Female Foragers from the Late/Final Jomon Period: Implications for Dietary Behavior and Reproductive Ecology. American Journal of Human Biology, 23: 107-117. Trotter, M., Peterson, R.R., 1969. Weight of bones during the fetal period. Growth, 33: 167-184. Turner, C.G., 1979. Dental Anthropological Indications of Agriculture among the Jomon People in Central Japan: X. Peopling of the Pacific. American Journal of Physical Anthropology, 51: 619-636. Ubelaker, D.H., 1979. Human Skeletal Remains: Excavation, Analysis and Interpretation. Smithsonian Institute Press. Waldron, T., 2009. Palaeopathology. Cambridge University Press. Walrath, D.E., Turner, P., Bruzek, J., 2004. Reliability test of the visual assessment of cranial traits for sex determination. American Journal of Physycal Anthropology, 125: 132-137.

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FIRST RECORD OF THE BUSH-CRICKET ISOPHYA CIUCASI (INSECTA: ORTHOPTERA: PHANEROPTERIDAE) IN MOLDAVIA (ROMANIA)

Ionuț Ștefan IORGU "Grigore Antipa" National Museum of Natural History, Kiseleff blvd., no. 1, 011341, Bucharest, Romania, [email protected]

Abstract. The bush-cricket Isophya ciucasi is recorded for the first time outside its locus typicus. Recently described from Ciucaş Mountains, the species was found in Vrancea Mountains in the summer of 2013. As the species was described as cryptic morphologically, the acoustic signals of males from the two known populations were compared.

Keywords: bush-cricket, Isophya ciucasi, Moldavia, bioacoustics.

Rezumat. Prima semnalare a cosaşului Isophya ciucasi (Insecta: Orthoptera: Phaneropteridae) în Moldova (România). Cosaşul Isophya ciucasi este semnalat pentru prima dată în afara locus typicus. Recent descrisă din Munţii Ciucaş, specia a fost găsită în vara anului 2013, în Munţii Vrancei. Deoarece specia a fost descrisă ca fiind criptică morfologic, am comparat semnalele acustice ale masculilor din cele două populaţii cunoscute.

Cuvinte cheie: cosaş, Isophya ciucasi, Moldova, bioacoustică.

Comprising more than 45 species, Isophya is the second largest Phaneropteridae genus in Europe. From the taxonomical point of view, genus Isophya is one of the most problematic of European Orthoptera, their morphological similarity making identification of species very difficult (Heller et al., 2004). As Isophya are stridulating species, male calling song analysis is usually regarded as the most important instrument for correct identification (Heller et al., 2004; Orci et al., 2010; Iorgu, 2012; Szövényi et al., 2012). The first individuals of Isophya ciucasi Iorgu I.Ş. & Iorgu E.I. were audio recorded in the summer of 2008 in Ciucaş Mountains and the species was later described as new, in 2010. Isophya ciucasi is very similar in morphology with Isophya camptoxypha, but the syllables produced during the calling song are different in structure. Until present study, the species was known only from the subalpine plateau of the Ciucaş Mountains, Eastern Carpathians. The bush-crickets were collected during six expeditions in Ciucaş Mountains and one in Vrancea Mountains. Adults and last instar nymphs caught in the wild were transported in aerated plastic containers and audio recorded indoors. The digital recorders EDIROL R-09HR was used. Ultramic200K external microphone was used for obtaining the full spectrum of song. Sound analysis was performed with the software Audacity 2.0.5 and Batsound 4. Oscillographic analysis terminology was used according to Heller et al. (2004), Orci et al. (2010) and Iorgu (2012): SD – period from the beginning of first impulse to the end of last impulse of the main impulse series of a syllable; NI – number of impulses of the main impulse series of the syllable; GS – period of time elapsed from the end of a syllable to the beginning of the next syllable; DAC – time elapsed from the last impulse of the main

- 115 - Ionuț Ștefan Iorgu impulse series of the syllable to the first after-click (-s); NAC – number of after-clicks produced after a syllable; STD – period from the beginning of first impulse to the end of last impulse of the main impulse series of a syllable, or end of its last after-click, if present. Photos were taken with a Canon DSLR camera, equipped with 100 mm 1:1 macro lens and 18-135 mm lens.

Order Orthoptera Suborder Ensifera Family Phaneropteridae Isophya ciucasi Iorgu I.Ş. & Iorgu E.I., 2010 (Fig. 1)

Material examined. 2 ♂♂ 1 ♀, Ciucaş cabin, Ciucaş Mountains, 1585 m a.s.l., 27.06.2008, leg. I.Ş. Iorgu; 3 ♂♂ 3 ♀♀, Ciucaş cabin, Ciucaş Mountains, 1600 m a.s.l., 17.07.2009, leg. I.Ş. Iorgu; 23 ♂♂ 9 ♀♀, Tigăile Mari, Ciucaş Mountains, 17.07.2010, leg. I.Ş. Iorgu; 10 ♂♂ 4 ♀♀, Ciucaş cabin, Ciucaş Mountains, 14.08.2010, leg. I.Ş. Iorgu; 7 ♂♂ 5 ♀♀, Tigăile Mari, Ciucaş Mountains, 29.07.2011, leg. I.Ş. Iorgu; 2 ♂♂ 1 ♀♀, Ciucaş cabin, Ciucaş Mountains, 09.08.2012, leg. I.Ş. Iorgu; 8 ♂♂ 3 ♀♀, Goru, Vrancea Mountains, 29.06.2013, leg. I.Ş. Iorgu. Audio recordings. 2 ♂♂, Ciucaş cabin, Ciucaş Mountains, 27.06.2008 (24ºC); 3 ♂♂ 3 ♀♀, Ciucaş cabin, Ciucaş Mountains, 17.07.2009 (27ºC); 19 ♂♂ 6 ♀♀, Tigăile Mari, Ciucaş Mountains, 17.07.2010 (24ºC); 7 ♂♂, Ciucaş cabin, Ciucaş Mountains, 14.08.2010 (26ºC); 3 ♂♂, Tigăile Mari, Ciucaş Mountains, 29.07.2011 (25ºC); 2 ♂♂, Ciucaş cabin, Ciucaş Mountains, 09.08.2012 (24ºC); 4 ♂♂, Goru, Vrancea Mountains, 29.06.2013 (20ºC). Remarks. Males sing at dusk and during the night. The song consists of a long series of syllables, sometimes lasting for more than 20 minutes. Each syllable is formed of a compact series of impulses, always followed by several after-clicks. All sounds are produced during the closing stroke of tegminae. Spectrographic analysis indicates frequencies between 15-60 kHz (Fig. 2). First data on the song (8 ♂♂ recorded in 2008 and 7 ♂♂ in 2009) showed that the compact series of 10-22 impulses (mean ± SD: 18.4 ± 4.82) lasts for 11-22 ms (mean ± SD: 18.2 ± 4.32) and the following after-clicks (13-23 impulses, mean ± SD: 17.6 ± 3.71) last for 200-314 ms (mean ± SD: 228 ± 25.31) (Iorgu & Iorgu, 2010). New recordings in the type locality (33 ♂♂ recorded in 2010, 7 ♂♂ in 2011 and 2 ♂♂ in 2012) showed a higher amplitude in song characteristics (Fig. 2, Table 1). Compared with males from Ciucaş, specimens from Vrancea showed insignificant differences in song production: relative longer syllable duration (mean 21.81 ms, respectively 15.56 ms) and delay of after-clicks (mean 117.45 ms, respectively 101.2 ms).

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Figure 1. Habitus and habitat photos of Isophya ciucasi: a - female, Ciucaş Mountains, 28.07.2011; b - male, Ciucaş Mountains, 28.07.2011; c - male, Vrancea Mountains, 29.06.2013; d - female, Vrancea Mountains, 29.06.2013; e - Ciucaş Mountains, 17.07.2010; f - Vrancea Mountains, Goru Peak, 29.06.2013.

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Figure 2. Sound analysis in Isophya ciucasi males: a, b - oscillograms; c, d - detailed syllable; e - spectrum; f - spectrogram (a, c, e, f - Vrancea Mountains; b, d - Ciucaş Mountains).

Table 1. Examined male song characters in Isophya ciucasi SD (ms) NI GS (ms) DAC (ms) NAC STD (ms) Min. 7 3 134 52 10 209 Ciucaş Max. 24 26 600 167 30 438 (n = 34, Mean 15.56 15.32 378.08 101.2 - 273.76 24-28ºC) S.D. 5.12 6.74 116.95 38.31 - 56.76 Min. 17 7 167 81 8 302 Vrancea Max. 26 20 289 161 21 459 (n = 4, Mean 21.81 12.81 212.09 117.45 - 386.63 20ºC) S.D. 3.02 4.02 37.1 30.39 - 50.57

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Acknowledgements The present study was supported by project “Completing and improving the information on wildlife species from Putna Vrancea Natural Park in accordance with national standards”. We acknowledge the anonymous reviewers for their advices on improving the paper.

References Heller, K.-G., Orci, K.M., Grein, G., Ingrisch, S., 2004. The Isophya species of central and western Europe (Orthoptera: Tettigonioidea, Phaneropteridae). Tijdschrift voor Entomologie, 147: 237-258. Iorgu, I.Ş., 2012. Acoustic analysis reveals a new cryptic bush-cricket in the Carpathian Mountains (Orthoptera, Phaneropteridae). ZooKeys, 254: 1-22. Iorgu, I.Ş., Iorgu, E.I., 2010. A new species of Isophya (Orthoptera: Phaneropteridae) from the Romanian Carpathian Mountains. Travaux du Muséum d’Histoire Naturelle “Grigore Antipa”, 53: 161-170. Iorgu, I.Ş., Iorgu, E.I., 2011. Bioacoustics in bush-crickets, crickets and grasshoppers (Insecta: Orthoptera) from Ciucaș Mountains (Eastern Carpathians, Romania). Brukenthal. Acta Musei, VI.3: 427-446. Orci, K.M., Szövényi, G., Nagy, B., 2010. A characterisation of the pair forming acoustic signals of Isophya harzi (Orthoptera, Tettigonioidea, Phaneropteridae). Acta Zoologica Academiae Scientiarum Hungaricae, 56(1): 43-53. Szövényi, G., Puskás, G., Orci, K.M., 2012. Isophya nagyi, a new phaneropterid bush-cricket (Orthoptera: Tettigonioidea) from the Eastern Carpathians (Caliman Mountains, North Romania). Zootaxa 3521: 67- 79.

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A SHORT OVERVIEW FOCUSED ON INVERTEBRATE AND VERTEBRATE BIOINDICATORS FOR FRESHWATER ENVIRONMENTS CONTAMINATED WITH HEAVY METALS

Ștefan-Adrian STRUNGARU*, Gabriel PLĂVAN, Marius Andrei RĂU and Mircea NICOARĂ Faculty of Biology, Alexandru Ioan Cuza University, Bd. Carol I 11, 700506 Iaşi, Romania. * [email protected]

Abstract. The invertebrate and vertebrate bioindicators are known as biomonitors which are used to indicate the negative effects caused in time by different pollutants from water, air and soil. These species are living in natural environments and are not the same model species/organisms which are used in toxicological studies conducted in laboratory conditions. Can the bioindicator species adapt to the negative effects of the pollutants? Each aquatic environment is unique and each species on this planet has the capacity to adapt itself to the environmental conditions. The aim of present study was to investigate the main bioindicators of freshwater environments contaminated with heavy metals, in order to be used in future studies.

Keywords: analysis methods, bioindicators, heavy metals, sampling.

Rezumat. Un scurt comentariu axat pe nevertebratele si vertebratele bioindicatori ai mediilor acvatice dulcicole contaminate cu metale grele. Vertebratele şi nevertebratele bioindicatoare mai sunt cunoscute ca biomonitori care indică efectele negative cauzate în timp de către diferiţi poluanţi din apă, aer şi sol. Aceste specii care sunt întâlnite în natură nu sunt aceleaşi cu organismele model utilizate în studiile toxicologice în condiții de laborator. Sunt capabile organismele bioindicatoare să se adapteze la efectele negative ale poluanţilor? Fiecare mediu acvatic este unic şi fiecare specie de pe această planetă are capacitatea să se adapteze la condiţiile de mediu. Scopul acestui studiu a fost investigarea principalilor bioindicatori ai mediilor acvatice dulcicole contaminate cu metale grele, care ar putea fi utilizați în studiile viitoare.

Cuvinte cheie: metode de analiză, bioindicatori, metale grele, prelevare probe.

Introduction The invertebrate and vertebrate bioindicators are known as biomonitors which are used to indicate the negative effects caused in time by different pollutants from water, air and soil. These species are living in natural environments and are not the same model species/organisms which are used in toxicological studies conducted in laboratory conditions. A certain species can be classified as a bioindicator of the environmental health after the analysis of several specific criteria: is it common in many ecosystems? Can it tolerate a large group of toxicants, those which cannot be tolerated creating disturbances of the populations by decreasing the number of the individuals and triggering migrations in order to avoid the pollution sources? One possibility to measure the pollutant effect upon the bioindicator is the decrease of population density or the absence of the species from an area where it was previously present in high abundance. Many studies proposed databases with bioindicator species and many of them were focused on water quality analysis. For instance, Neumann et al., 2003 presented in their study the data base of LIMPACT with 39 bioindicators that were observed in the studied stream (Taxa: Turbellaria, Oligochaeta, Gastropoda, Amphipoda, Isopoda, Plecoptera, Coleoptera, Diptera, Ephemeroptera, Megaloptera and Trichoptera). The results

- 121 - Ștefan-Adrian Strungaru et al. were based on the interpretation of the abundances and rule syntax. These showed the level of contamination with pesticides correlated with nine water-quality and morphological parameters. Other authors proposed benthic larvae of insects for pesticides and heavy metals after complex chemical analyses. The freshwater mollusks are considered to be the best bioindicators for the contamination with pesticides and heavy metals in freshwater environments, but the interactions with environmental conditions release questions about the bioindicators. Can the bioindicator species adapt to the negative effects of the pollutants? Each aquatic environment is unique and each species on this planet has the capacity to adapt to environmental conditions. The aim of present study was to investigate the main bioindicators of freshwater environments contaminated with heavy metals, in order to be used in future studies.

Materials and methods required for heavy metals analysis in freshwater ecosystems The first step in this investigation is the choice of the analyzed environment that is important for humans and is rich in biodiversity. All the freshwater environments are important to sustainability of the life forms on this planet. There is a hierarchy of the importance of freshwater ecosystems based on: biodiversity richness, endangered species, economical activities, freshwater usage for drinking. The second step is based on the identification of the possible heavy metals pollution sources based on anthropogenic activities e.g.: heavy industry, mining, agriculture, car traffic, waste waters and wastes disposal. Next step is the choice of the bioindicators. The scientific literature provides valuable information about bioindicators; if not, we must find the species which respond to the specific questions. The indicators for heavy metals in fresh water ecosystems are: water, sediments and biota. In case of biota there are several ways of finding the indicators of heavy metals pollution: analysis of the food web (it is very complex, there are necessary many resources; the analysis should begin with producers-planktonic algae and end with highest consumer/predator-fish, birds, mammals and humans), organisms with high capacity of metal absorption from environment (benthic organisms like gastropods, bivalves, macrophytes) or species with a short life cycle (their population can be damaged very fast by the activity of the pollutant). Very important are the sampling period (season of the year) and the number of samples (for sediments, water and biota) to prove the heavy metals pollution in the studied environment. The main problem that may appear is the accidental contamination of the samples. That is why the person that runs the sampling must use uncontaminated bottles and bags metal free made from PET. The measurement of water parameters (pH, conductivity, ORP, TDS, water temperature, salinity, dissolved oxygen, oxygen saturation and others) is useful because the correlation between them and metal content (water, sediment, biota) may offer a complex image about metal absorption and toxicity. Once the samples are collected they must be preserved for metals analysis in laboratory. The biota and sediment samples can be preserved by freezing at -20⁰C in PE bags. Water samples are preserved by acidification with nitric acid (Marcovecchio et al., 2007; Biziuk et al., 2010). The equipment and technique used may be different. One of the most frequent technique used for these types of analyzes is AAS (Atomic Absorption Spectrometry). The AAS heavy metals analysis is a modern technique for measuring the

- 122 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 metals concentration in life forms and environment, that was used since 1955 (L’vov, 2005). It is a technique that is still developing nowadays. It is split in two main methods of analyses: GF-AAS (on graphite furnace with inert gas as transporter) for low concentrations of metal in samples (µg g-1) and Fl-AAS (flame-AAS with air-acetylene mixture) for high concentrations of metal in samples (mg g-1). Before the analysis, the samples must be weighted and digested with acid mixtures; this is different for each sample type. Microwave digestion system with pressure teflon vessels is often used. The biometric measurements and biological age of the organisms are important variables for correlations. The AAS must be calibrated with certificated standard solutions that are not expired and are kept in conditions recommended by the producer. The blank sample and water for the dilution must be ultrapure with no content of the elements that will be analyzed, in order to avoid possible errors. All the bottles and pipettes must be uncontaminated; this can be proved by absorbance measurements. The metal element with its specific wave length does not have any absorbance if not present in the sample. The reference materials for sediments and biota are required to test the accuracy of the device, and QC samples are also required. It is recommended to firstly test the presence of the metal in samples, to do the calibration curves and to dilute the sample if the concentration exceeded (Fig. 1).

Figure 1. The absorbance and spectrum number that prove the existence of cadmium in sample.

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The calibration (Fig. 2) of the device is most important for the measurements and data validation. If the metal concentrations in samples exceed the modern devices capacity, the samples must be diluted. All the results and data must be statistical validated and interpreted.

Figure 2. Examples of calibration for nickel and chromium, using GF-HR-CS-AAS ContrAA 600 AnalitikJena.

Groups of invertebrates and vertebrates with potential in the monitoring of heavy metal pollution Macroinvertebrates are the most important bioindicators in heavy metal pollution because they can provide valuable information about pollution sources. The groups with potential in heavy metals monitoring are: Plecoptera, Ephemeroptera, Trichoptera, Hemiptera, Coleoptera, Odonata, Diptera, Crustacea, Gastropoda, Bivalvia, Tricladi, Hirudinea, Oligochaeta (Arimoro et al., 2009; Testi et al., 2012; Moldovan et al., 2013). The disturbance in their populations can be caused by the metal toxicity. It is very important that the metal concentration from their bodies to be correlated with population density. There are disadvantages because many of them are relatively small and more effort is required to obtain the biomass for metal analysis. Another disadvantage is the locomotion capacity and the capacity to tolerate in time the toxic effects of the pollutant (Arimoro et al., 2009; Testi et al., 2012; Moldovan et al., 2013). Bivalves and other species with low

- 124 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 capacity of locomotion are the best candidates; if they are big enough we can do analyzes on different organs. The metals accumulate in organisms by time for the chronic pollution. Another variable is the biological life cycle. In the acute pollution, the species with a faster biological cycle are the best because pollutants can enter very fast in the cells and create damages. In the case of vertebrates, the juveniles are the best indicators in the acute poisoning with heavy metals. For instance, the tadpoles that absorb very fast the lead into their bodies (Strungaru et al., 2012) and the mortality of the young fish which is the first indicator of the high contamination and disturbance of the environment (Hoffman et al., 2002). The abnormal developing and the behavior of the young fish are symptoms for heavy metals contamination. The accumulation of the metals must be studied for different organs in order to set the target organ of the toxicant from environment (Fig. 3) because there is the possibility for the toxic metal to be localized inside an organ only. The water birds are indicators in chronic exposure for heavy metals in freshwater environments. The high juvenile’s mortality and food source contamination are huge problems. The main in vivo analyses for water birds contamination with heavy metals are for blood, feather and excrement. This method does not provide significant results all the time and there is the possibility that the birds were accidentally contaminated during migration. The analysis of the internal organs (liver, kidneys, spleen and bones) are the most important to prove the contamination (Binkowski et al., 2013).

Figure 3. The metal pathway from environment in a predator and to the target organs.

In case of mammals, the blood samples and urine are used to observe the level of contamination with heavy metals. The otter (Lutra lutra B.) can be a good indicator because is a predator species. The behavior is a preliminary indicator in case of metal contamination. In humans, especially in the mining areas, there were recorded many cases of exposure to cadmium and lead that caused new types of diseases, damages to the body

- 125 - Ștefan-Adrian Strungaru et al. and abnormal child development (Andujar et al., 2010; Menai et al., 2012; Faramawi et al., 2012; Sun et al., 2014; Kim et al., 2013; Inaba et al., 2005). All the results and conclusions should be integrated with education, health, strategies and future investments to reduce the anthropogenic pressure caused by pollution (Fig. 4) on freshwater environments.

Figure 4. Results from freshwater monitoring studies of the heavy metals pollution integrated to reduce the anthropogenic pressure.

Conclusions This short overview presented a few groups of invertebrates and vertebrates capable to react in the process of heavy metals pollution of the freshwater ecosystems. In case of invertebrates, population densities must be correlated with the metal from environment and from their bodies. For vertebrates must be identified the target organs where each studied metal proves its negative effects. It will be a problem in the future with freshwater resource if the people will not react with strategies for reducing the anthropogenic pressure. The education, health, investments and resources management integrated with the data provided by indicators is the key in the protection of the freshwater ecosystems.

Acknowledgments This work was supported by the strategic grant POSDRU/159/1.5/S/133391, Project “Doctoral and Post-doctoral programs of excellence for highly qualified human resources training for research in the field of Life sciences, Environment and Earth Science” cofinanced by the European Social Fund within the Sectorial Operational Program Human Resources Development 2007-2013.

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References Andujar, P., Bensefa-Colas, L, Descatha, A., 2010. Acute and chronic cadmium poisoning. Revue de Medecine Interne, 31: 107-115. Arimoro, F.O., Ikomi, R.B., 2009. Ecological integrity of upper Warri River, Niger Delta using aquatic insects as bioindicators. Ecological Indicators, 9: 455-461. Binkowski, Ł.J., Sawick-Kapusta, K., Szarek, J., Strzyżewska, Felsmann, M., 2013. Histopathology of liver and kineys of wild living Mallards Anas platyrhynchos and Coots Fulica atra with considerable concentrations of lead and cadmium. Science of Total Environment, 450-451: 326-333. Biziuk, M., Beyer, A., Zukowska, J., 2010. Preservation and storage of water samples. Analytical Measurements in

Aquatic Environments, 19-39. Faramawi, M.F., Liu, Y. , Caffrey, J.L., Lin, Y.-S., Gandhi, S., Singh, K.P., 2012. The association between urinary cadmium and frontal T wave axis deviation in the US adults. International Journal of Hygiene and Environmental Health, 215: 406-410. Hoffman, J.D., Rattner, A.B., Burton, G.A., Jr., Cairns, J., 2002. Handbook of ecotoxicology, Second edition. CRC Press. Inaba, T., Kobayashi, E., Suwazono, Y., Uetani, M., Oishi, M.N., Nogawa, K., 2005. Estimation of cumulative cadmium intake causing Itai-itai disease. Toxicology Letters, 159: 192-201. Kim, S., Arora, M., Fernandez, C., Landero, J., Caruso, J., 2013. Lead, mercury, and cadmium exposure and attention deficit hyperactivity disorder in children. Environmental Research, 126: 105-110. L’vov, B.V., 2005. Fifty Years of Atomic Absorption Spectrometry. Journal of Analytical Chemistry, 60(4): 382- 392. Marcovecchio, J.E., Botté, S.E., Freije, R.H., 2007. Heavy metals, major metals, trace elements. In Handbook of water analysis, second edition, 275-311. Menai, M., Heude, B., Slama, R., Forhan, A., Sahuquillo, J., Charles, M.-A., Yazbeck, C., 2012. Association between maternal blood cadmium during pregnancy and birth weight and the risk of fetal growth restriction: The EDEN mother-child cohort study. Reproductive Toxicology, 34: 622-627. Moldovan, O.T., Meleg, I.N., Levei, E., Terente M., 2013. A simple method for assessing biotic indicators and predicting biodiversity in the hyporheic zone of a river polluted with metals. Ecological Indicators, 24: 412-420. Neumann, M., Baumeister, J., Liess, M., Schulz, R., 2003. An expert system to estimate the pesticide contamination of small streams using benthic macroinvertebrates as bioindicators. II. The knowledge base of LIMPACT. Ecological Indicators, 2: 391-401. Strungaru, Ş.-A., Jitar, O., Plăvan, G., Nicoară, M., 2012. Lead accumulation in the bodies of Rana tadpoles (Anura: Ranidae). Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, LVIII: 93-98. Sun, H., Chen, W., Wang, D., Jin, Y., Chen, X., Xu, Y., 2014. The effects of prenatal exposure to low-level cadmium, lead and selenium on birth outcomes. Chemosphere,108: 33-39. Testi A., Fanelli G., Crosti R., Castigliani V., D’Angeli D., 2012. Characterizing river habitat quality using plant and animal bioindicators: A case study of Tirino River (Abruzzo Region, Central Italy). Ecological Indicators, 20: 24-33.

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THE INVASIVE SPECIES PERCCOTTUS GLENII – A THREAT FOR THE FRESH WATER ECOSYSTEMS

Monica LUCA1 and Gogu GHIORGHIȚĂ1, 2 1 “Alexandru Ioan Cuza” University, Faculty of Biology, Bd. Carol I, 20A, Iași, 700505, Romania, [email protected], [email protected] 2 Academy of Romanian Scientists, Piatra Neamț Branch, Romania, [email protected]

Abstract. The introduction of non-native species in new habitats had, in several occasions, damaging effects on the local biodiversity, when the introduced species managed to establish there and later became invasive. An example was the fresh-water fish species Perccottus glenii, which was introduced in Russia, near at the beginning of the 20th century, and only later, during the past two decades, its dissemination covered a large area, being pinpointed from Eurasia to the west of Europe. Because of its high resistance to extreme environmental conditions and of the economic loss caused in the fishing ponds, the Amur sleeper – as also called – became a real threat for the fresh-water ecosystems in Asia both and Europe. Many studies developed an intensive geographic survey of Perccottus glenii and they pointed out its morphologic and physiologic features in order to build a comprehensive profile of this fast invader; however, they did not include the genetic analysis, which represented a key-factor for assessing its invasion potential. This later data would complete the species profile, with the final purpose of planning certain programs for the conservation of native fresh-water species from the invaded areas.

Keywords: Perccottus glenii, invasion, dissemination, loss of biodiversity

Rezumat. Specia invazivă Perccottus glenii – o amenințare pentru ecosistemele de apă dulce. Introducerea speciilor non-native în habitate noi a avut, în repetate rânduri, efecte devastatoare asupra biodiversității locale, atunci când speciile introduse au reușit să dobândească stabilitatea și să devină ulterior invazive. Un exemplu l-a constituit și peștele de apă dulce Perccottus glenii, care a fost introdus în Rusia, lângă Sankt Petersburg, la începutul secolului XX, și abia mai târziu, pe parcursul ultimelor două decenii, răspândirea sa a acoperit o arie largă, fiind semnalat din Eurasia până în vestul Europei. Datorită rezistenței sale la condiții de mediu extreme și din cauza daunelor economice pe care le provoacă în iazurile de pește, guvidul adormit – după cum mai este denumit în limbaj popular – a devenit o adevărată amenințare pentru ecosistemele de apă dulce atât în Asia cât și în Europa. Numeroase studii s-au axat pe o monitorizare geografică intensivă a speciei Perccottus glenii și au pus în lumină, de asemenea, trăsăturile sale morfologice și fiziologice, pentru a construi o imagine cât mai cuprinzătoare a acestei specii invazive; totuși, aceste studii nu au inclus analize genetice, care reprezintă un factor cheie în cuantificarea potențialului invaziv. Analizele genetice ar completa profilul speciei, cu scopul de a planifica în final anumite programe de conservare pentru speciile native de pești de apă dulce din arealele invadate.

Cuvinte cheie: Perccottus glenii, invazie, răspândire, pierderea biodiversității

Introduction The millennial development of human society has had – in several occasions – a direct impact on the environment, bringing radical changes, whether they were deliberate or not, and ichthyofauna represents one of the most affected elements. Native fish species on the entire planet faced, on one hand, the immediate effect of human activity (by the deliberate introduction of other alien fish species) and their subsequent spread beyond human control, and, on the other hand, the downsides of pollution and greenhouse effect. They caused climate transformations followed by the expansion of invasive species to the detriment of native ones.

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The reasons that triggered animals’ movement by humans outline significant differences in the timing of their introductions in new habitats. One of the main elements that facilitated the spatial displacement of different species on Earth and that intensified the extent of the natural movement of species between neighbouring areas was the emergence and development of trade and exchange networks in ancient times, but also, later, of human cultural interests or hobbies. Unlike the introduction of different mammals, which goes back in time for thousands of years (the late Mesolithic and Neolithic period), when many domestic animals were introduced in new continents (Anderung, 2006; Albarella et al., 2007; Larson et al., 2007; Zeder, 2008; Ottoni et al., 2013) – the main introductions of exotic fish species in countries outside their natural habitat are more recent on the European continent – only to a few centuries ago – and they also follow the commercial routes (Keller et al., 2011). Most alien fresh-water fish species started to be introduced in Europe in the 19th century (Benigno, 2001), even if other European species might have been introduced in other continents, like eastern Asia (Japan), a few centuries earlier – for instance, the common carp, Cyprinus carpio (Balon, 2004). The history of fishery is much older than the one of fresh-water fish introductions, though it did develop later than the process of mammals’ domestication. Archaeological evidence describes a peak of fishing processing facilities in the 1st-2nd centuries AD (Scearce, 2009). The temporal distance between “fish domestication” and its introduction to entirely new habitats was influenced mainly by humans’ requirements and needs. In ancient times, the management of food resources represented, on one hand, a basis for subsistence, and, on the other, a key-factor in the process of social and economic evolution of humans, which continued later, in medieval and modern times. Starting with the past few centuries, food resources also became the subject of scientific research, of different hobbies, or they were used just for decoration purposes. Thus, from the 19th century, fish species started to be introduced to new habitats for the following: sports, improvement of wild stocks, aquaculture, decorations and biological control or by simple accident (Benigno, 2001). Nevertheless, irrespective of the purpose of their introduction, this process can have severe consequences on the environment if the introduced species pass through all stages which lead to the status of invasive species. In this sense, a very good example is the fresh-water species Perccottus glenii, also known as the Amur sleeper. This species is one of the many exotic ones introduced in European countries in the 20th century, mostly in the first half, before the Second World War (Holčík, 1991). From its Eurasian location, where its introduction was reported for the first time, during the past decades, it has managed to spread massively in many European countries, from the east to the west of Europe (Reshetnikov, 2010). Its high capacity to adapt to new water environments with different hydro-climatic conditions, as well as its velocity of spread were a big sign of warning for ecology and other related scientific branches. Specialists have started analyzing its features as invasive species and foreshadowing its spread with the purpose of conceiving conservation programs extremely useful for the protection and ecological improvement of biocenosis in the invaded areas (Lawson Handley et al., 2011).

Perccottus glenii – a fast colonizing species in a large area The main native area where the Amur sleeper was introduced from – at the

- 130 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 beginning of the 20th century– is the basin of the Amur River, in the Far East of Asia, covering a part of the Russian, Chinese and North-Korean territories, where it was found mostly in the floodplain waters of smaller or even larger Amur tributaries, like Zeya, Sungari and Ussuri (Bogutskaya & Naseka, 2002). Its native range also includes the water bodies of the tributaries of the gulfs of the Yellow Sea: Gou, Liao, Never, Yalu Rivers from China and North Korea. In the east, the native range reaches the north-west of the Sakhalin Island, in Russia (Reshetnikov, 2013); hence, it includes the tributaries of the Sea of Japan: Suifun, Tumen and the Liushung region, previously known as Port Arthur (Bogutskaya & Naseka, 2002). The distribution range of this species increased significantly with the introductions that took place during the 20th century. The oldest reported introduction of Perccottus glenii dates to 1912, when the species was introduced from the Zeya River, tributary of the Amur River, into an area close to Saint Petersburg (where it was kept for a few years as an aquarium fish and released later, in 1916, in a garden pond). The second earliest introduction is reported from 1948, when the Amur sleeper was brought to also from its native range, after an expedition to the Amur River (Bogutskaya & Naseka, 2002). The occurrence of Perccottus glenii in other countries is recorded after the middle of the 20th century: the earliest appearance of this species in Asia (Uzbekistan) dates from 1961, (Reshetnikov, 2010) and in Europe, Ukraine, from the beginning of the ‘70s (Kvach, 2012). A more intense survey of Perccottus glenii’s geographical spread began only after 1990; in fact, the past two decades have recorded the peaks in velocity and space for this species, to such an extent that it has become a possible threat to many fresh-water ecosystems. Thus, two invasion areas are described: the first one near its native range, and other one in Eurasia and Europe, where its spread starts with the main points of its introduction: Saint Petersburg, Moscow and Ukraine. A particular case is Germany, where the Amur sleeper might also have been introduced by mistake, for aquaculture purposes, alongside other fresh-water fish species with economic value (Reshetnikov & Schliewen, 2013). Thus, by 2000, the species Perccottus glenii was already identified in the West of Ukraine, in the Latorica and Borzahava rivers (Reshetnikov, 2010), in the Tisza River in Hungary, (Covaciu-Marcov et al., 2011) in the basins of Latorica and Bodrog rivers in Slovakia, (Koščo et al., 2003) as well as in Germany. However, in the last country, the individuals were collected at least 10 years later (Reshetnikov & Schliewen, 2013). Since 2001, the Amur sleeper has been recorded in even more countries, starting with Serbia – where it gets out easily from the upper stream of the Tisza River – in Jazovo fishpond, in the basin of the Tisza River (Simonović et al., 2006). In 2001, it was identified for the first time in Romania, in the Suceava River (Nalbant et al., 2004), with the only difference that it must have got here from Ukraine (Reshetnikov, 2010). In 2005, new recordings of this species appeared in the Romanian sector of the Danube, both on the middle stream (at the Iron Gates) and in the Danube Delta (Popa et al., 2006; Reshetnikov, 2010; Năstase, 2008). It has also been pinpointed in the Bulgarian sector of the Danube, corresponding to the Romanian one (Jurada et al., 2006). After 2005, the presence of this fish species was further mentioned in Romania in the Mureş River basin (Copilaș-Ciocianu & Pârvulescu, 2011), as well as in other European countries, like Moldova (Mosu, 2007), (Caleta et al., 2010), (Pupiņa & Pupiņš, 2012), , Finland, and Slovenia

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(Reshetnikov, 2010). It has also been spotted in many other provinces of the Russian Federation. By 2008, it had already spread in 36 Russian provinces, 12 European countries (Latvia, , Estonia, Byelorussia, Ukraine, Moldova, Poland, Slovakia, Hungary, Serbia, Bulgaria, Romania) and three Asian countries (, Uzbekistan and Mongolia), (Reshetnikov, 2010) (Figs. 1-2).

Figure 1. First introductions of Perccottus glenii in the East of Europe (after Reshetnikov, 2010, 2013).

Figure 2. Current range of Perccottus glenii in Eurasia (http://en.wikipedia.org/wiki/Chinese_sleeper#mediaviewer/File:Perccottus_range_map.png).

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Morpho-physiological traits favouring the establishment and spread of Perccottus glenii in non-native habitats To explain the success of invasive fish species in gaining stability in a new habitat, many conditions have been described. Some of them are related strictly to exogenous traits, like nutrients quantity in the invaded area, (directly connected to their trophic status) the size of the native range or the distance from the nearest native source. However, it is also worth underlining the traits regarding the species itself, like the morphology, (including the maximum adult size) the parental care, the physiological tolerance to extreme environmental factors, the prior invasion success or the propagule pressure (Marchetti et al., 2004). For this matter, the reduced size of Perccottus glenii makes it easy to be transported, by mistake, together with other species, for aquaculture purposes (Reshetnikov & Schliewen, 2013). Perccottus glenii reaches between 14-25 cm in length and a maximum weight of 260-270 g, with a faster growth in the non-native range than in the native one (Bogutskaya & Naseka, 2002). The exceptions are a consequence of factors like climate differences, food resources or zooplankton density (Grabowska et al., 2011). For a species to become stabile in a new habitat and invasive later, traits connected to growth and development are very important, such as early sexual maturity, short generation time or rapid growth (Haynes, 2009). Perccottus glenii reaches sexual maturity very early, after the first year of life, and it presents sexual dimorphism: males are larger than females (explained precisely by the fact that females spend the energy for somatic growth on gonad production). In exchange, males offer parental care to their offspring, which, according to some scientists, is another strategic behaviour for better survival of an invasive species (Grabowska et al., 2011). Alongside its reduced size and fast growth, there are other physiological traits which bring more advantages to the species Perccottus glenii in the fight for survival and spread. These traits mainly refer to the resistance to quite high and very low temperatures, to the rate of warming and to the different concentrations of chemical components in the invaded fresh water ecosystems. Thus, the dissemination area of Perccottus glenii is much influenced by different climate parameters. The maximum temperature ensuring survival is strictly connected to the velocity of warming, and from this point of view, it was shown that, under a low warming rate, the temperature can rise up to 38 degrees, while under a high warming rate, the fish does not survive to more than 35 degrees (Golovanov & Ruchin, 2011). In addition, considering the high temperatures in the south of the European countries, this factor can be a limiting one for the dissemination of the Amur sleeper in this area, and also in the extreme north of Europe, where the low temperatures reached in the warmest months of the year can slow down the reproduction rate (Reshetnikov & Ficetola, 2011). The proportion of certain natural phenomena like precipitation and evapotranspiration represents another important factor because, through these processes, the concentration of chemical compounds in the water – as the sodium salts, chlorides and oxygen – rises. Nonetheless, Perccottus glenii presents such a resistance to these chemical changes that it can survive the complete drying up of the water by hiding in a mucus shell (Bogutskaya & Naseka, 2002).

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Enemies of Perccottus glenii in the invaded area One of the biggest advantages of an invasive species is the lack of enemies in new habitats, which facilitates the survival, proliferation and subsequent dissemination of the species. Considering anthropogenic stress as a first type of enemy, often encountered for many species of fish and caused by the potential economic value of these species for humans, Perccottus glenii is free from this stress factor, as it has never been of interest for human nutrition or any kind of other use (Reshetnikov, 2004). Still, many introductions were made by humans, who transported the species by mistake together with other species, aiming to improve the breed stocks in different European ponds, or who used it as bait in fishing activities (Reshetnikov & Schliewen, 2013; Caleta et al. 2010). Many survey studies carried out so far on Perccottus glenii have showed that this species is not likely to be identified on the main course of big rivers. It inhabits only small water bodies, poor in fish species, thus avoiding as much as possible the presence of any predators or even of food competitors (Reshetnikov & Schliewen, 2013). This little number of predators is also due to the inappropriate biochemical conditions encountered generally in small and isolated water bodies. Here, the oxygen level is very low and the survival capacity of Perccottus glenii in such conditions is significantly higher than the one of other predator fish species, like the perch, bleak, roach, goldfish or carp (Caleta et al., 2010). This limiting environmental factor helps the species Perccottus glenii escape not only from the predator enemies, but also from their competitors for food, since the trophic spectrum of species like the goldfish or roach overlaps the one of Perccottus glenii by high proportions (81.2% and 49.4%, respectively), (Bogutskaya & Naseka, 2002). Besides the extreme environmental conditions, this species can eliminate many fish enemies by transmitting parasites to the native fish or other aquatic or semi-aquatic vertebrates (Reshetnikov & Schliewen, 2013), or through its large trophic spectrum, which inludes larvae of insects and amphibians, micro-crustaceans, worms, and also small fish, causing great economical loss for the fish ponds and also a decrease in the vertebrate species richness in many small water bodies (Reshetnikov, 2004; Koščo et al., 2008; Reshetnikov & Schliewen, 2013).

Beyond the geographical survey, anatomy and physiology of Perccottus glenii – the genetic perspective for the risk analysis of its invasion Generally, biological invasions represent fast evolving events which create genetically dynamic populations, in both space and time, and the genetic population traits have a great impact on both the dissemination capacity and the dispersal area of certain invasive species (Lee, 2002; Lindholm et al., 2005). Nevertheless, the genetic tools have been used so far for a quite reduced number of invasive species in Europe, most of them belonging to the Cyprinidae family (Alcaraz Cazorla, 2007; Gorgan, 2007; Mezhzherin & Lisetskii, 2004; Kalous et al., 2012; Kang et al. 2014). Many morpho-physiological studies and geographical surveys, but very few genetic analyses have been carried out so far for Perccottus glenii and, in order to get a more precise and complete image regarding its invasive status, a genetic approach is required, with the purpose of assessing the variability in and between populations. A first problem the scientists have dealt with for decades in the research carried out for Perccottus glenii is taxonomy, concerning the order of this species. The univocal fact regarding the

- 134 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 taxonomy of Perccottus glenii is that it belongs to the Perciformes order, suborder Gobioidei, but the families included in this suborder have been long time discussed. By the middle of the 20th century, different scientists included the species Perccottus glenii in the Eleotridae family which, alongside other two families – Gobiidae and Periophthalmidae – make up the Gobioidei suborder (Terlecki & Palka, 2012). The Amur sleeper continued to be included in the Eleotridae family even towards the end of the 20th century (Manteifel & Bastakov, 1986). Only later, in the last decade of the 20th century, Perccottus glenii was assigned to the Odontobutidae family, alongside the genus Micropercops and Odontobutis (Terlecki & Palka, 2012). However, the problem was only apparently solved, because later publications continued to include the Perccottus glenii species in the Eleotridae family (Kas’yanov & Goroshkova, 2003). The taxonomy based on phenotypic traits was finally clarified by the phylogenetic study carried out for the Gobioidei suborder first in 2005, when four mitochondrial complete genes were analysed and a phylogenetic tree was built. According to this genetic data, the species Perccottus glenii actually belongs to the Odontobutidae family, a monophyletic family that – together with the monophyletic Rhyacichthyidae family – forms a sister group which split for the first time in the Gobioidei suborder. These results were supported later, in 2013, by an even more accurate phylogenetic tree, based on five complete mitochondrial genes (Tacker & Hardman, 2005; Agoretta et al., 2013). Thus far, taxonomy represents the only issue where the genetic tools have been used for solving; however, further genetic analysis must be carried out to establish the precise origin of invasive populations of Perccottus glenii, their invasion routes, as well as their genetic variability both in and between populations. The purpose is to design and implement specific strategies for the conservation of the natural biocenosis. The high spread speed of the invasive populations of Perccottus glenii both in Asia and Europe (covering a significant number of contries in only few decades), the presence of certain morphologic and physiologic features like the reduced adult size, its fast growth and early sexual maturity (which is acieved within only one year of life), the resistance to extreme environmental conditions, like the wide range of temperature (which also enlarges considerably the invasion area from the South to the North of Europe) or the low level of oxigen (which favors the colonizing of small water bodies with a very low number of other fish species), the lack of enemies (as a consequence of the habitat requirements or simply for its no economical value), its trophic spectrum (which includes small fish) and its often reported status of parasites carrier, represent special features which transform the Amur sleeper into a real threat for many fresh-water ecosystems. And above all these, the lack of knowledge about any genetic traits which trigger the adaptabilty and spread of this species, makes the invasive fish Perccottus glenii to become a imminent danger for the fresh-water ecosystems.

Conclusions Thus far, the research activity for Perccottus glenii shed light on many details regarding the morphological and physiological traits of this species, as well as its geographical dissemination, and all these details describe the perfect portrait of an invasive species. Considering the recorded findings of Perccottus glenii outside its native range, this invasive species has spread remarkably fast on a very large area.

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The reduced size of this fish, alongside its high prolificacy and the resistance to extreme environmental conditions, represent another big advantage that enables Perccottus glenii to survive and become stable in completely new habitats, and then to disseminate to the surrounding areas. The resistance to extreme environmental conditions gives the Amur sleeper the capacity to exploit different ecological niches and thus to adapt more easily to a different trophic spectrum. This typical trait for an invasive species represents another advantage for survival and dissemination, as it helps the species escape its natural enemies. Considering that Perccottus glenii represents a threat for the biodiversity preservation in many fresh-water ecosystems, and that there is yet no genetic study regarding the variability both in and between populations, this type of analysis is almost mandatory for assessing the Amur sleeper’s capacity to adapt to different fresh-water environments and for predicting its future dissemination.

Acknowledgements This work was supported by a grant of the project POSDRU 159/1.5/S/133652.

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Hoese, D.F., Gill, A.C. 1993. Phylogenetic relationships ofeleotridid fishes (Perciformes: Gobioidei). Bull. Mar. Sci., 52(1): 415-440. Holčík, J., 1991. Fish Introductions inEurope with Particular Reference to its Central and Eastern Part. Can. J. Fish. Aquat. Sci., 48: 13-23. http://www.zin.ru/animalia/pisces/eng/taxbase_e/species_e/perccottus/perccottus_glenii_eng.pdf. Jurajda, P., Vassilev, M., Polačik, M., Trichkova, T., 2006. A First Record of Perccottus glenii (Perciformes: Odontobutidae) in the Danube River in Bulgaria. Acta Zoologica Bulgarica, 58(2): 279-282. Kalous, L., Bohlen, J., Rylková, K., Petrtýl, M., 2012. Hidden diversity within the Prussian carp and designation of a neotype for Carassius gibelio (Teleostei: Cyprinidae). Ichthyological Exploration of Freshwaters, 23(1): 11-18. Kang, J.-H., Noh, E.-S., Lim, J.-H., Han, H.-K., Kim, B.-S., Lim, S.-K., 2014. Genetic Differentiation of Carassius auratus and C. Cuvieri by the Cytochrome C Oxidase I Gene Analysis. Aquaculture Research & Development, 5(3): 1-4. Kas’yanov, A.N., Goroshkova, T.V., 2012. Morphological Features of the Amur Sleeper(Perccottus glenii, Perciformes, Eleotridae) Introduced into Water Bodies of European Russia. Contemporary Problems of Ecology, 5(1): 58-7. Keller, R.P., Geist, J., Jeschke, J.M., Kühn, I., 2011. Invasive species in Europe: ecology, status, and policy. Environmental Sciences Europe, 23(23): 1-17. Koščo, J., Lusk, S., Halačka, K., Lusková, V., 2003. The expansion and occurrence of the Amur sleeper (Perccottus glenii) in eastern Slovakia. Folia Zoologica, 52(3): 329-336. Koščo, J., Manko, P., Miklisova, D., Košuthova, L., 2008. Feeding ecology of invasive Perccottus glenii (Perciformes, Odontobutidae) in Slovakia. Czech Journals of Animal Science, 53(11): 479-486. Kvach, Y., 2012. First record of the Chinese sleeper Perccottus glenii Dybowski, 1877 in the Ukrainian part of the Danube delta. BioInvasions Records, 1(1): 25-28. Larson, G., Albarella, U., Dobney, K., Rowley-Conwy, P., Schibler, J., Tresset, A., Vigne, J-D, Edwards, C.J., Schlumbaum, A., Dinu, A., Bălășescu, A., Dolman, G., Tagliacozzo, A., Manaseryan, N., Miracle, P., Van Wijngaarden-Bakker, L., Masseti, M, Bradley, D. G., Cooper, A., 2007. Ancient DNA, pig domestication, and the spread of the Neolithic into Europe. PNAS, 104: 4834-4839. Lawson Handley, L.-J, Estoup, A., Evans, D.M., Thomas, C.E., Lombaert, E., Facon, B., Aebi, A., Roy, H.E, 2011. Ecological genetics of invasive alien species. International Organization for Biological Control, 56: 409-428. Manteifel, Y.B., Bastakov, V.A, 1986. Percottus glehni Dybowski - a new colonizer in the ichthyofauna of Lake Glubokoe. Hydrobiologia, 141: 133-134. Marchetti, M.P., Moyle, P.B., Levine, R., 2004. Invasive species profiling? Exploring the characteristics of non- native fishes across invasion stages in California. Freswater Biology, 49: 646-661. Mezhzherin, S.V., Lisetskii, I.L., 2004. The Genetic Structure of European Goldfish Carassius auratus s. lato (Cyprinidae) in Ukrainian Water Bodies: An Analysis of Bisexual Samples. Biology Bulletin, 31(6): 574-581. Mosu, A., 2007. Invazia in Unele Ecosisteme Acvatice ale Republicii Moldova a Peștelui Alogen – Perccottus glenii Dybowski, 1877 (Perciformes: Odontobutidae). In Problemele Actuale ale Protecției și Valorificării Durabile a Diversității Lumii Animale: Materialele Conferintei a VI-a a Zoologilor din Republica Moldova cu Participare Internațională (Chișinău, 18 -19 Octombrie 2007), 170-172 (in Romanian). Nalbant, T.T., Battes, K.W., Pricope, F., Ureche, D., 2004. First record of the Amur sleeper Perccottus glenii (Pisces: Perciformes: Odontobutidae) in Romania. Travaux du Museum National d'Histoire Naturelle “Grigore Antipa”, 47: 279-284. Năstase, A., 2008. First Record of Amur sleeper Perccottus glenii (Perciformes, Odontobutidae) in the Danube Delta (Dobrogea, Romania). Acta Ichthiologica Romanica, II, 167-175. Ottoni, C., Flink, L.G., Evin, A., Georg, C., De Cupere, B., Van Neer, W., Bartosiewicz, L., Linderholm, A., Barnett, R., Peters, J., Decorte, R., Waelkens, M., Vanderheyden, N., Ricaut, F.X., Cakırlar, C., Cevik, O., Hoelzel, A.R., Mashkour, M., Karimlu, A.F.M., Sheikhi, S.S., Daujat, J., Brock, F., Pinhasi, R., Hongo, H., Perez-Enciso, M., Rasmussen, M., Frantz, F., Megens, H.J., Crooijmans, R., Groenen, M., Arbuckle, B., Benecke, N., Strand Vidarsdottir, U., Burger, J., Cucchi, T., Dobney, K., Larson, G., 2013. Pig Domestication and Human-Mediated Dispersal in Western Eurasia Revealed through Ancient DNA and Geometric Morphometrics. Mol. Biol. Evol, 30(4): 824-832. Popa, L.O., Popa, O.P., Pisică, E.I., Iftime, A., Matacă, S., Diaconu F., Murariu, D., 2006. The first record of Perccottus glenii Dybowski, 1877 (Pisces: Odontobutidae) and Ameiurus melas rafinesque, 1820

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(Pisces: Ictaluridae) from the Romanian sector of the Danube. Travaux du Muséum National d’Histoire Naturelle «Grigore Antipa», 49: 323-329. Pupiņa, A., Pupiņš, M., 2012. Invasive fish Perccottus glenii in biotopes of Bombina bombina in Latvia on the north edge of the fire-bellied toad’s distribution. Acta Biologica Universitatis Dau Gavpiliensi, 3: 82- 90. Reshetnikov, A.N., 2004. The fish Perccottus glenii: history of introduction to western regions of Eurasia. Hydrobiologia, 522: 349-350. Reshetnikov, A.N., 2010. The Current Range of Amur sleeper Percottus glenii Dybowski, 1877 (Odontobutidae, Pisces) in Eurasia. Russian Journal of Biological Invasions, 1: 119-126. Reshetnikov, A.N., 2013. Spatio-temporal dynamics of the expansion of rotan Perccottus glenii from Wets- Ukrainian centre of distribution and consequences for European freshwater ecosystems. Aquatic Invasions, 8: 1-14. Reshetnikov, A.N., Ficetola, G.F., 2011. Potential range of the invasive fish rotan (Perccottus glenii) in the Holarctic. Biological Invasions, 13(12): 2967-2980. Reshetnikov, A.N., Schliewen, U.K., 2013. First record of the invasive alien fish rotan Perccottus glenii Dybowski, 1877 (Odontobutidae) in the Upper Danube drainage (Bavaria, Germany). Journal of Applied Ichthyology, 29: 1367-1369. Scearce, C., 2009. European Fisheries History: Pre-industrial Origins of Overfishing. ProQuest, Discovery Guides, 1-13, http://www.csa.com/discoveryguides/fish/review.php Simonović, P., Marić, S., Nikolić, V., 2006. Record of the Amur sleeper Perccottus glenii ( Odontobutidae) in Serbia and its recent status. Archives of Biological Science, Belgrade, 58(1): 7-8. Terlecki, J., Pałka, R., 2012. Occurence of Perccottus glenii Dynowski 1877 (Perciformes, Odontobutidae) in the middle stretch of the Vistula River, Poland. Archives of Polish Fisheries, 7(1): 141-150. Thacker, C.E., Hardman, M.A., 2005. Molecular phylogeny of basal gobioid Wshes: Rhyacichthyidae, Odontobutidae, Xenisthmidae, Eleotridae (Teleostei: Perciformes: Gobioidei). Molecular Phylogenetics and Evolution, 37: 858-871. Zeder, M.A., 2008. Domestication and early agriculture in the Mediterranean Basin: Origins, diffusion, and impact. Proceedings of the National Academy of Sciences of the USA, 105(33): 11597-11604.

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THE THEORY OF SPONTANEOUS GENERATION, BETWEEN IDEOLOGY AND SCIENCE

Ion COJOCARU Faculty of Biology, Alexandru Ioan Cuza University, Bd. Carol I 11, 700506 Iaşi, Romania. [email protected]

Abstract. Belief in spontaneous generation of beings had a millenial existence, right up to the end of the 19th century, and stayed in the debate of two camps: spontanists and anti-spontanists. Today the problem of spontaneous generation is a closed case in the history of science. What makes it unique but this dogma is her ideological interpretation extremely varied, being invoked by materialistic or idealistic thinkers, theologians or darwinist biologists. For demonstration of these traits, the present work is structured around an ideological theme: idealist spontanism, materialist spontanism, idealist anti-spontanism, materialist anti-spontanism. It is remarkable that an idea so easily dismissed today, just on the basis of elementary knowledge and logical arguments persisted so long in history. The theory of spontaneous generation was not unmasked but little by little, in successive stages: first to the large, highly organized organisms, then for small organisms and, finally, for the microscopic (Leeuwenhoek: animalculi). The disproving of the spontaneous generation dogma involved, in the ideological plan, the release of preconceived ideas, and in practical, eliminationthe sources of errors in experiments and perfecting experiments.

Keywords: spontaneous generation theory, materialist spontanism, idealist spontanism, idealist anti-spontanism, materialist anti-spontanism

Rezumat. Teoria generației spontane, între ideologie și știință. Credința în generația spontană a ființelor a avut o existență multimilenară, până spre sfârșitul secolului al XIX-lea, și a stat în dezbaterea a două tabere: spontaniștii și anti-spontaniștii. Astăzi problema generației spontane este un caz închis în istoria științei. Ceea ce face unică însă această dogmă este interpretarea ei ideologică extrem de variată, fiind invocată de gânditori materialiști sau idealiști, de teologi sau biologi darwiniști. Pentru evidențierea acestor trăsături, prezenta lucrare este structurată pe o tematică ideologică: spontanism idealist, spontanism materialist, respectiv anti-spontanism materialist, anti- spontanism idealist. Este remarcabil faptul că o idee atât de ușor de respins astăzi, doar pe baza unor cunoștințe elementare și argumente de logică, a persistat atât de mult în istorie. Eroarea teoriei generației spontane nu a fost demascată decât încetul cu încetul, în etape succesive: mai întâi pentru organisme mari, superioare, apoi pentru organisme mici și, în cele din urmă, microscopice (Leeuwenhoek: animalculi). Demontarea dogmei generației spontane a presupus, în plan ideologic, eliberarea de dogme preconcepute, iar în plan practic, eliminarea surselor de erori din experimente și perfecționarea experimentelor.

Cuvinte cheie: teoria generației spontane, spontanism idealist, spontanism materialist, anti-spontanism idealist, anti-spontanism materialist

Introduction Spontaneous generation of organisms represented a conception about the occurrence of organisms in different environments, not from their parents too, but even in the material or assumed immaterial elements, in a relatively short time. F. A. Pouchet a defender in the modern period of spontaneous generation, gives the following definition: "Spontaneous generation is occurence of a new organized beings lacking parents and whose main elements were entirely extracted from the surrounding substances" (Pouchet, 1859). The first ideas on spontaneous generation of organisms originate from ancient times. Aristotle asserted that some animals come from parents and others do not. There is

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Ion Cojocaru the belief that the complex organisms, small or large, previously known as fauna, may arise from mineral elements or from dead matter in the decomposition, at temporal scale (hours, days, seasons) (Aristoteles, 1860, 1868). This thing called spontaneous generation (generatio spontanea) can be repeated whenever shall meet the biogenetic conditions envisaged. Another situation is that of living organisms that occur in other living organisms (host organisms in current terms), which does not relate (intestinal worms, for example), called equivocal generation (generatio aequivoca), heterogenesis or xenogenesis. Emergence of insects in galls on plants fall into this category also. Compared to these mysterious origins of living beings, the natural occurrence of organisms through reproductive processes known, was called the univocal generation (generatio univoca). Biogenetics factors suspected to be responsible for spontaneous generation were very different: the Sun's rays (Diodore of Sicily); divine powers which have left since the days of creation, whether material, "divine seed" and awaiting favourable conditions for development (Augustine); Special creation of divinity, without the existence of \"divine seed\" (Thomas Aquinas); the mark of divine creation (Jan Swammerdam); \"ferments\" contained in matter (Van Helmont); animal spirits, or seminal release from corpses (Athanasius Kircher); the primordiums such eggs, which can be formed directly from inert matter (Harvey: primordium = egg appeared spontaneously) (Rostand, 1975; Bastian, 2001). "The theory" of spontaneous generation (which, in fact, did not meet the conditions of a scientific theory ever) was dismantled little by little, being removed permanently from Pasteur, in the second half of the 19th century. The idea of spontaneous generation was differently understood and presented. He spoke of spontaneous emergence of individuals of certain species (Aristotle, tomist dogma), or spontaneous emergence of life itself in the early times of the world (Buffon, Diderot, Maupertuis, etc.). On the other hand, the rejection of the theory of spontaneous generation was very different exploited: rejecting the possibility of occurece of forms of life observed (Redi, Leeuwenhoek, Pasteur, Spallanzani), the possibility of the creation of beings (Swammerdam) or denying the possibility of moving naturally from matter devoid of life to life (Pasteur's initial hesitation, clergymen). It is remarkable that an idea (also called Virchow, 1855 "heresy"), so easily dismissed today, just on the basis of elementary knowledge and logical arguments persisted so long in history. What makes it unique but this dogma is her extremely varied ideological interpretation, being operated as materialistic or idealistic, thinkers and theologians or darwinist biologists.

I. The dogma of spontanism and its disproving The disproving of spontaneous generation theory dogma was a gradual process, influenced by developments in scientific knowledge, improvement of methods of experimentation, the elimination of sources of errors in experiments and the release of preconceptions. In fact, removing the spontaneous generation dogma meant overcoming a capital impasse, conceptual and methodological, appeared between spontanists and anti- spontanists: how many times anti-spontanists will carry out experimental conditions such as microorganisms may no longer appear in the culture medium, spontanists will object that desiring to prevent the intervention of germs, they disturbed, in fact, the conditions which allow the formation of life. Thus, Needham will claim in 1748 that an excessive heating of

- 140 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 matter infused, destroy "vegetative force" necessary aggregations of organic molecules in living beings; torturing nature, "forces it to be a false witness" he said (Rostand, 1975). Instead, whenever spontanists will present facts that seem to advocate in favor of a spontaneous genesis, anti-spontanists they argued that they never managed to take all the necessary measures to exclude the germs. One can identify three stages/aims of abolition, experimental way, spontaneous generation theory: (1) the invalidation of spontaneous generation theory for macroscopic animals (F. Redi, M. Malpighi, A. Vallisneri); (2) invalidation of spontaneous generation theory for microorganisms, through isolation and heat the sample (L. Spallanzani); (3) the invalidation of the theory of spontaneous generation for microorganisms, in natural conditions (normal temperature, presence of air, especially oxygen) (Pasteur, Schwann, Tyndall). To achieve these objectives were required the invention of three categories of methods: (1) environmental organic putrescibil isolation from the external environment (Redi). (2) isolation and heated to boiling temperature (Needham, Spallanzani). (3) insulation, heating, cooling, constant contact with the external air, plus use of biological liquids placed in sterile air (Pasteur).

II. Spontanism and anti-spontanism Division of authors who have discussed the theory of spontaneous generation in spontanists and anti-spontanists is very general and relative. In general, spontanists are the supporters and defenders of the theory of spontaneous generation, and anti-spontanists those who do not support this conception. But there were all grades and shades in assigning these names for a specific author. For example, Redi was anti-spontanist with regard to the origin of insects from dead matter, but spontanist with regard to the occurrence of insects from the galls on plants. Swammerdam was anti-spontanist in that it does not accept the birth of living things in the mud, manure, dead bodies, etc., but did not say that the spontaneous emergence as such is impossible, it is put on the account Creator.

The ideological movement of the spontanism Theory of spontaneous generation was supported, along time with arguments mostly idealistic or predominantly materialistic. Over time, is observed an ideological movement of the concept of spontaneous generation, from a naive materialism (Anaximander), toward a philosophical idealism (Empedocles, Aristotle) and religious (Albertus Magnus, Augustine, Thomas Aquinas), then toward a materialism that was the framework of some scientific theories (Buffon, Haeckel). Doctrinal separation in idealistic spontanism materialistic spontanism is general and not exclusive. As the opera's great ancient thinkers, and later, non-scientific doctrines have incorporated in different proportions elements of idealistic thinking and materialistic thinking. Thus, the preformism and the pangenesis theory of antiquity had a materialistic character, trying to explain the phenomenon of heredity and embryonic development through living matter's properties. Then, this doctrine take the form of Idealist doctrine, being taken over by philosophers theologians, including the preformation in the divine plan Act (Augustine). The vitalism originated in idealist Aristotelian thinking about entelechia and psyche, but it represented a reaction to mecanicist materialism from the 17th century (analogy between the supreme force of inanimate bodies, viewed as a machine-the gravity,

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Ion Cojocaru and the supreme force of living beings - the vital force). Thus, Benoît de Maillet (1656- 1758) consider that germs, which included living beings in miniature, were spread everywhere in nature and are not created by the deity. This scenario, in which divinity is not involved was a materialist scenario on the origin of species. Maupertuis will imagine combining seminali germs (also Maillet's germs from nature), not just on the basis of chemical affinity, but also through an instinct, desire, or intelligenceto associate, opening the way vitalistic doctrine. Mechanistic materialist explanation of the origin of life will lead to an idealistic vision. Epigenesis theory as a theory of embryonic development have, in the conception of Aristotle, a materialistic character, but she served an idealistic ideology – the finalism of biological transformation (intrinsic goal of the motion of the matter) (Balme, 1962; Gotthelf, 1986). The theory of spontaneous generation, like other old dogmatic doctrines (pangenesis, preformism, vitalism), underwent radical, ideological changes over time, oscillating between naive materialism, creationist, vitalist and preformist idealism, and scientific materialism. Find it interesting that an institution which declares that recognises the dogmas "immutable", the Church, had with regard to the theory of spontaneous generation diametrically opposed attitudes: the incorporation of the theory of spontaneous generation in the dogma of the Church (12th-13th centuries), to outright rejection of this conception, which had become, over time, an assumption too materialistic and taken in laughter of the experimental science (18th-19th centuries). With the advancement of science, knowing more and more about the environmental chemistry and the way of life of organisms, numerous experimenters will pass, with regard to the theory of spontaneous generation, from a creationist, religious spontanism, toward a dualistic philosophy (specific of "natural theology"), in which Divinity is offered a role less important. As the theory of spontaneous generation assigned an increasingly materialist, idealist orientation the theory of spontaneous generation finds refuge in vitalism: in the 18th century (1748), John Needham was talking about a vegetative force which allows organic molecules to the living beings agrege. Gradually, is taking shape the idea of a vital force called exciting cause (J. B. Lamarck, 1744-1829), or vital energy (Pouchet, 1859). About this this direction were, from the beginning, also critical reactions. Charles Bonnet (1720-1793), opponent of the theory of spontaneous generation said: "When to explain the emergence of animalculi in a liquid are resorting to\" vegetative capacity", no place does the words instead of things?" and Claude Bernard, the great French physiologist (1813-1878) show that we can use the expression "vitality" as chemists use the word "affinity". At the end of the 17th century, the idea of spontaneous generation in other organisms of living beings, which they were not related (e.g. intestinal worms), have fewer supporters. The idea that Divinity would have imposed these creations were not sympathized; on the other hand, was more widely accepted belief that all organisms originate through the union of a male and a female. Result that the new living being was preformed miniature in the seed of one of the two parents. The discovery of the phenomenon of partenogenesis from Charles Bonnet in aphids, in 1745, has been interpreted as an argument in favor of preformism. The preformism fit perfectly with the doctrine of the Church in relation to a pre-set plan regarding the origin of living beings. A. The idealist spontanism. The idealist spontanism put at the origin of spontaneous generation a immaterial force of matter, or the Act of divinity. There is a

- 142 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 gradual movement of this conception, from the principles of Aristotelian philosophy, in direction of religious dogmas and vitalist science. As a result we will separate the idealistic spontanism into three categories: (1) the philosophical; (2) the theological and (3) the vitalist. 1. The philosophical spontanism. Ancient mythology offered explanations for almost all things otherwise inexplicable that they find the man. Thinkers such as Empedocles, Democritus, Epicurus, Theophrastus, Lucretius, Vergilius and Aristotle (but not Pythagoras) (Chiorcea, 2005) have agreed, in one form or another, the idea of spontaneous generation. Aristotle, dualist thinker, in his Historiaanimalium says several times that animals of different kinds appear directly in the material elements and pneuma-a vital force (heat) translated into Latin by anima (soul). He explains, in Generatione animalium, that living things appear from decaying matter, vegetable matter (some insects), the internal secretions of some animals and excrement (some parasites), or in any thing dry that gets wet. He believed that "everything is alive can be achieved not only as a result of mating, but also by the decomposition of soil". Aristotle rejected the idea that the universe or Earth had a beginning, as a result the process of spontaneous generation occurs all the time, not just at the beginnings, as believed by other Greek thinkers. In the 17th century, Descartes, mecanicist, materialist philosopher considered spontaneous generation as a process naturally knowable of complicated machinery construction – beings, from non-living matter, without the intervention of the vital forces, but after the creator's laws (Dorobanțu, 1980). 2. The theological spontanism. The theory of spontaneous generation will come to the attention of the Church, it will adapt and integrate religious dogma. The Church turn into dogma the idea of Aristotle, being in her support, transforming the conception of spontaneous generation in a official conception, fact which constituted a serious obstacle to the progress of scientific knowledge. Among the early fathers of the Church, Gregory of Nyssa (335-394) and Augustine (354-430) argued that not all kinds of plants and animals were created by God directly, rather some appeared in historical times, indirectly, from those created in the beginning. Augustine believed that spontaneous generation is due to the divine powers (rationes seminales). All beings that it produces have been created even from the fifth or sixth day of Creation. These "divine seed" will wait wait for a period more or less long, having the possibility to develop further, in favourable conditions, through natural processes. The idea that organisms might change through natural processes was considered as a possibility by a number of Christian theologians of the middle ages, including Albert the Great (Albertus Magnus, 1193/1206-1280) and his pupil Thomas Aquinas (1224-1274). Thomas Aquinas accepted the spontaneous generation as a form of God's special creation, without the involvement of "divine seed". He considered that the development of spontaneous living things, such as worms and flies, from a non-living matter, such as altered meat, is not incompatible with the Christian faith or philosophy. In the 12th century, the idea that a variety of geese can occur from a marin crustacean ciriped, had implications in the practice of lent when the fish was permitted, not the bird. Monks ate in the days of fasting meat of duck and goose, counting that they originate from fish, until Pope Innocent III himself was decided to intervene (1215), prohibiting this. Albert the Great, the great dominican theologian and scholastic philosopher takes in his books legends about the birth of some birds of the crustaceans from

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Ion Cojocaru the sea, or the birth of ducks, geese and even from certain fruit trees. The concept of spontaneous generation of creative force thanks to the beings of the deity is widely accepted. A representative of this was Van Helmont (1577-1644). 3. The vitalist spontanism. The vitalism was developed as a reaction to mecanicist materialism which does not deny existence of spirit, but give them a secondary role in relation to physical-chemical factors. The vitalism, in turn, does not deny the role of physical and chemical factors, but consider that life cannot be reduced to them. Biological processes are directed to specific, vital forces. Needham spoke of a "vegetative force", at 1748 (supported by Buffon), which allow aggregation of organic molecules in living beings. Later, rival of Pasteur's debates, F. A. Pouchet (1800-1872) (Hétérogénie ouTraité de la Génération spontanée, 1859), presents an idealised conception of the spontaneous generation of creationist-vitalist type, that wishes to support with "scientific" experiments (concession of materialism). He admits the primordial divine creation, spontaneous generation is a gift of divine beings explained in terms of vitalism: "each primary organic molecule possesses, without a doubt, two elements: one material and other vital ". For life to emerge-says Pouchet – are needed and sufficient the conditions: a body that can rot, water, air and a suitable temperature. The old claim that, for example, from dirty laundry moistened and cheese or wheat grains etc. may arise spontaneously mice (van Helmont) are considered by Pouchet "exaggerations" mentioned by some to compromising the theory of spontaneous generation. From the composition of some materials in the environment will not obtain an animal never completely formed, but only the egg from which it may subsequently develop adult beings (Eșanu, 1985). Even sexual reproduction was considered a spontaneous generation produced by the vital force of the elements coming together at mating. The vital force not acting randomly, but according to the will of the divine. B. The materialist spontanism. In this category falls the old interpretations, naive, of the genesis from matter of living beings, together with the erroneous results of some scientific experiments, or ideological response to the invocation of supernatural in explaining the origin of living beings. As a result we distinguish in the materialist spontanism, four categories: (1) the materialist, naïve spontanism; (2) thematerialist, philosophical spontanism (3) the anti-theological materialist spontanism; (4) the materialist spontanism incorporated into scientific theories. 1. The materialist, naïve spontanism. A number of Greek thinkers, Anaximander, Anaximenes, or Xenophanes believed that plants, animals and humans arose in the interaction of the elements of nature: a primordial mud (mixture of Earth and water) under direct solar heat. In this case, the spontaneous generation appears as an expression of the properties of matter - a form of naive materialism.The materialist, naïve spontanism was the consequenceof the lack of knowledgethe way of life of organisms and observation errors. 2. The materialist, philosophical spontanism. Democritus believed that life is the result of mechanical forces of nature (Mohan and Neacșu, 1992). In this case, the spontaneous generation appears as an expression of the properties of matter-a form of materialistic thinking. F. Bacon (1561-1626), author of the inductive method of the new times, no doubts of spontaneous generation, at least in terms of a lot of plants and lower animals. He defines the process of spontaneous generation as a path by which one can make the transition from non-living to the living world.

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3. The anti-theological materialist spontanism. Many scholars have interpreted the Pasteur's experiences as a proof of absolute impossibility to pass on naturally from non- living matter to living matter. As a result, biologists and materialist philosophers, darwinist evolutionists such as Ernst Haeckel,have tolerated the theory of spontaneous generation, in order not to admit the Act of divine creation. Haeckel declared: "to deny the spontaneous generation mean to admit a miracle, to admit a divine creation of life". In an era heavily influenced by fixism and creationism, the idea that beings arose spontaneously from inorganic matter was the only non-creationist scientific conception and even anti-creationist (Eșanu, 1985). In our country, materialist and darwinist philosophers have defended the point of view of Haeckel in spontaneous generation, in response to alternative Creationism: Vasile Conta in Teoria ondulațiunii universale (Conta, 1876, 1876a, 1876b, 1876c, 1876d, 1876e, 1876f, 1877, 1877a, 1877b, 1877c) or Nicolae Leon, in Generațiunea spontanee și darwinismul (Leon, 1903). 4. The materialist spontanism incorporated into scientific theories. Materialistic rationalism that characterize scientific research was born under the dualist philosophy. The historical development of the theory of spontaneous generation show a change to its content, moving from idealism to materialism as scientific knowledge progressed. Spontaneous generation is explained by the properties of matter showing, on this way, a link between the non-living and living. The dualistic philosophy allowed the progress of science, avoiding as far as possible, the conflict with religious dogma. Descartes (1596-1650), father of rationalistthinking, assert that all existing structures, however complex they may be, are the result of a natural process, but after the laws established by God. Even if dualist philosophers maintained, circumstantial, the sovereign role of the Divinity in nature, scientific explanation of specific phenomena became increasingly important. Redi, in concession to the spontaneous emergence of insects in galls on plants, talks about "live" juices of the plant that generates insects and the parasites of animals originating in "living juices" of the host animal. A materialist vision of the spontaneous generation has also W. Harvey (1578-1657), who believed that beings come from eggs that appeared spontaneously ("ex ovo omnia"). A precursor to evolutionary theory, Buffon (1707-1788) issues a materialist theory of spontaneous generation with with a vitalist component, which did not bring a progress in science – the theory of organic molecules. According to Buffon, all beings, from the simplest infusoria to man, are made up of some particles-"organic molecules"- qualitatively different from the particles from which inorganic matter is formed. "Organic molecules" are spread everywhere, are immortal, and the beings represent only certain combinations of these molecules. After being death, decompose only combination, without those molecules to perish. The number of organic molecules is fixed, but they can be combined in infinite ways, giving the infinite diversity of nature. Buffon resume, in fact, Athanasius Kircher’s conception, "organic molecules " being in fact "volatile animals spirits" of the cleric. The idea of combining by hazard of some molecules or living germs spread in nature, through which it can produce higher organisms, was widely accepted until the middle of the 18th century (Maupertuis, La Mettrie, Diderot and others). To point out that the formulation of such ideas in the 18th century, represents a considerable regress. Scientific thinking could not reject from the beginning the dogma of spontaneous generation. Explanatory gaps in certain areas, such as the cell theory or of evolution theory,

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Ion Cojocaru allowedkeeping the theory of spontaneous generation, especially for explanation of the origin of life or of the great differences between the groups of living things. Thus, Lamarck and Geoffroy, deists and evolutionists, have invoked often spontaneous generation to explain their discontinuities between groups of living things. The combination of the concept of spontaneous generation and the scala naturae led, in fact, to Lamarck's theory of evolution: the spontaneous emergence of simple beings from inanimate matter, which then transforms slowly into more complex forms (Mayr, 1989). Development of cell theory by Mathias Schleiden and Theodor Schwann in the early part of the 19th century paved the way for understanding the unicellular organisms reproduction through cell division. Question asked by Leeuvenhoek on the origin of animalculi can now find the answer. But, to understand the process of cell division, as a way of origin of new microorganisms from the preexisting microorganism, Schleiden believed that young cells occur inside the old cell from a fluid substance without structure called citoblastem. This explanation of the origin of living forms fits into the logical explanation of schema theory of spontaneous generation (Mayr, 1989).

The ideological movement of the anti-spontanism The historical rejection of the theory of spontaneous generation was made from two completely different considerations: (1) finding that this conception of materialistic type don't fall into the dogmatic claims of the Church (anti-spontanism idealistic, theological) and (2) the development of experimental science, able to dismiss baseless theories (scientific, materialist, anti- spontanism). 1. The idealist, theological anti-spontanism. The idea that living beings could take birth through the decomposition of corpses came at a time, in contradiction with the the biblical data. Why was designed Noah's Ark, if organisms can be born from the corpses of dead animals, asks Thomas Browne (1605-1682), English physician and writer. The dutch naturalist Jan Swammerdam (1637-1680) was a great opponent of the theory of spontaneous generation (Botnariuc, 1961). For Swammerdam, the concept that one animal could arise from another or from putrification by chance is impious and irreligious. He studied the development of many animals, metamorphosis in insects and frogs, and made meticulous observations on those smaller animals (mites, lice, efemerids, bees) (Historia insectorum generalis, 1669). Swammerdam is very impressed by the complex organization of these small living machines and considered crazy to admit that the blind hazard can composed of rotting, incidentally the complex organisms. His assessment is correct fair, but explains these wonders of nature as a result of divine creation. For Swammerdam, the anti- theological spontanism does not lead to discrediting the theory of spontaneous generation, but only replaced the natural causes of this pseudo-phenomenon with voluntary action of divinity. The theory of spontaneous generation, started from the naïve thinking, mythological, idealist vision of mankind about the various phenomena of nature, gradually acquired a character materialistic, even being imagined as a natural process that man, however, does not understand well this. Spontaneous generation was, of course, in contradiction with the dogma of a unique creations at the beginning of the world. Such a natural process of creation, that no longer involves the divinity, could no longer be accepted by the Church. Spontaneous generation seemed a faith for atheism. The same thing it has been argued, a century before, also by Voltaire (preformist in biology), an enemy of

- 146 - Analele Științifice ale Universității „Alexandru Ioan Cuza” din Iași, s. Biologie animală, Tom LX, 2014 atheism, which did not want to take the Creator the role of Planner and active force in the emergence of forms of life (Lepeltier, 2009). Pasteur proved through scientific experiments error of the spontaneous generation theory and showed that the environment around us is full of micro-organisms that come from some of the others, after a process of reproduction. But, during a Conference at the Sorbonne, Pasteur make the association between the spontaneous appearance of living beings, from matter, with the materialistic doctrine of non-believers: if there is spontaneous generation, "what good the idea of a creator God?" (Rostand, 1975). The clergy have adapted these words in their best interests, praising Pasteur which brings such an evidence of the involvement of the divine creative force. Matter itself has no creative power, only God. Abolition of the theory of spontaneous generation by Pasteur has left some evolutionists the impression that it denies the possibility of evolution from non-living to living, a fact which has negatively influenced the research about the origin of life. It appears, then, that the demolition of this "theory" has been differently exploited, both the evolutionists and creationists.. 2. The materialist, scientific anti-spontanism. The field of action to prove the theory of spontaneous generation was neither philosophy nor religion, but experimental science. It was necessary to be imagined experiments to eliminate the sources of error in formulating of the explanations. Leeuwenhoek, practitioner, discoverer of microorganisms, spontaneous generation denied preferring to adhere to the old pre-existence germs theory in the Universe or panspermia (as described by the theory of Anaxagoras, 500-428 BCE). The experimental contributions of anti-spontanists, regardless of their religious or philosophical orientation, resulted in the consolidation of the materialist vision about natural processes. The whole scientific contribution of scholars such as Redi, Vallisneri, Malpighi, Bonnet, Spallanzani, Pasteur, Schwann, Tindall had finally allowed the victory of biology as a science. Experimental science disassembled spontaneous generation theory, as pointed out above, gradually and in stages, from the higher forms of life, from the simplest ones. After Darwin proposed his theory of descendence, the concept of spontaneous generation had suffered violent attacks, followed by it rejection on the experimental wayby Pasteur and others. Darwin himself said that the conditions which preceded the emergence of life are still present and have always been present, but the actual living things would devour any more complex substance necessary of life (Mayr, 1989). This scenario suggests that the emergence of life was a unique and historic process, and no spontaneous generation of complex beings, previously known. Pasteur's experiments are simple; they derive directly from the technique inaugurated in the 18th century of Needham and Spallanzani; but to correctly execute them, it was necessary a clarity of mind, perseverance, a logical tactic. Pasteur introduced the sterilization of samples at a temperature of boiling water, 100 C°; subsequently, were found spores (Bacillus subtilis) that rezisted at that temperature, but which were destroyed at a temperature of 120 C° (Taton, 1972). John Tyndall (1877) publishes his method for fractional sterilization, showing the existence of heat-resistant bacterial spores. Pasteur could finally conclude with certainty: "spontaneous Generation is a chimera ... No, there is no circumstance known to be able to affirm that the microscopic beings have come into the world without germs, without parents like them. Those who

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Ion Cojocaru claim have been victims of the bad experiences made, flawed by errors, which they knew not to see them or could not avoid them" (Rostand, 1975). Today it is generally accepted that life on our planet is a unique phenomenon of occurrence of life from inorganic matter, through natural processes, following an long prebiotic chemical developments. Artificial life can be produced under laboratory conditions? This is another issue. Theoretically, in terms of chemistry and molecular biology, it would seem possible. Basically though, beyond methodological difficulties,expected and unexpected, remains to be seen if mankind can recreate at extremely small temporal scale a grand and unique process, produced by itself, at spatial- temporal scale extremely large.

Conclusions The spontaneous generation is a pseudo-phenomenon inherited from the ancient thinking, adapted to different ideological conjecture of the time. Spontaneous generation problem divided the authors into two camps: spontanists and anti-spontanists. It was not a camp to hold the truth always and another who always to go wrong. There was, however, a camp-the spontanists, situated on a runway entirely wrong, and another, that of antispontanists - directed correctly, even though more committed errors in method. These camps did not have a well-defined ideological identity and constant, ranging from idealism to materialism (Table 1). The spontanism presents itself as being appropriate, naïve, philosophical, mystical, idealistic, vitalist, materialistic, without scientific value. The Church, circumstantial, has accepted the theory of spontaneous generation, the spontaneous occurrence of beings, being regarded as a divine creation, or he rejected the spontanism, when the spontaneous generation was regarded as a natural phenomenon, material. The Spontanism was a refuge of materialist scientists, who did not want to turn to the Act of creation (E. Haeckel). The Anti-spontanism is presented as being rationalist, materialist: naive or scientifically accepted by some theologians as a reaction to the materialist spontanism (theological anti- spontanism). Spontaneous generation, as pseudo-phenomenon, was at first interpreted as an expression of the creative powers of matter, then as an expression of divinity, then, the divinity and nature have also been deprived of these powers. Ideas about spontaneous generation were influenced not only by the quality of observations and experiments, but the flow of cultural ideas (philosophical, religious, mythical), which circulated and were at one time dominant in society. The same pseudo-phenomenon, spontaneous generation, was materialist interpreted by the ancient naïve thinking, idealist creationist of scholastic thought of the Church, idealist vitalist (van Helmont, F. Pouchet) materialist and vitalist (Buffon), scientific materialist (Pasteur). The same pseudo-phenomenon, spontaneous generation, was accepted and criticized by the Church, was supported and criticized by scientists.

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Table 1. The ideological orientations of spontanism and anti-spontanism and the main representatives.

Spontanism Anti-spontanism Th. Browne Aristotle Swammerdam philosophical Descartes theological Voltaire

idealist Pasteur in clerical

vision Augustin theological Toma d’Aquino idealist A. Kircher Van Helmont J. Needham vitalist Buffon F.A. Pouchet Redi

Anaximander Leeuvenhoek

Anaximenes Malpighi naive Xenophanes scientific Spallanzani Avicenna Ch. Bonnet

J. Carda Cl. Bernard materialist philosophical Democritus Pasteur F. Bacon anti-theological E. Haeckel materialist W. Harvey incorporated in Buffon scientific theories De Maillet Lamarck Schleiden

The theory of spontaneous generation was an error supported by enlightened spirits-Descartes, F. Bacon, Buffon, Lamarck, Haeckel, which expresses the naivety of free thinking or, on the contrary, the power of preconceived ideas , the power of authority or tradition. This error was not unmasked but little by little, in successive stages: for large organisms, then small and microscopic. The spontanists will oscillate between spontaneous emergence of higher adult animals (van Helmont), eggs (Pouchet), from the miraculous apparition of life, at explanations based on vitalistic mechanisms. The dismantling of the spontaneous generation dogma involved, in the ideological plan, the release of preconceived ideas, and in practical, eliminationthe sources of errors in experiments and perfecting experiments.

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