Arthonia Parietinaria E a Common but Frequently Misunderstood Lichenicolous Fungus on Species of the Xanthoria Parietina-Group
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fungal biology 120 (2016) 1341e1353 journal homepage: www.elsevier.com/locate/funbio Arthonia parietinaria e A common but frequently misunderstood lichenicolous fungus on species of the Xanthoria parietina-group Antonia FLEISCHHACKERa, Martin GRUBEa,*, Andreas FRISCHb, Walter OBERMAYERa, Josef HAFELLNERa aInstitut fur€ Pflanzenwissenschaften, NaWi Graz, Karl-Franzens-Universitat,€ Holteigasse 6, 8010 Graz, Austria bDepartment of Ecology, Swedish University of Agricultural Sciences, P.O. Box 7044, 750 07 Uppsala, Sweden article info abstract Article history: Arthonia parietinaria is described as new to science. Host of the type and at the same time Received 31 December 2015 the only confirmed host species is the foliose macrolichen Xanthoria parietina. Sequence Received in revised form data of nucLSU rRNA genes reveal a close relationship to Arthonia molendoi. A. parietinaria 29 May 2016 is recorded for many countries in Europe, western Asia, and northern Africa. Accepted 6 June 2016 ª 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved. Corresponding Editor: Ana Crespo Keywords: Arthoniales Arthoniomycetes Ascomycetes Lichen parasites Introduction includes 38 species, while the current version of an online- synopsis lists 115 species (Lawrey & Diederich 2015). In a pri- According to the traditional concepts, Arthonia Ach. (1806) rep- vate database we accumulated about 200 heterotypic names resents one of the large genera of ascomycetes, comprising that would be relevant in connection with a generic revision both lichenized and lichenicolous species. The number of ac- (J.H., unpubl.). And the process of inventory is still going on cepted species is given with 491 in the latest edition of the with additional species described almost every year. ‘Dictionary of the fungi’ (Kirk et al. 2008), which seems to be Due to high values of nitrogen deposition in European a rather conservative approximation. A search in MycoBank landscapes, the eutrophic conspicuous orange Xanthoria parie- (http://www.mycobank.org) reveals more than 1200 infrage- tina is one of the few foliose macrolichens with populations neric names linked to the genus. Also the proportion of lichen- increasing both in number and size since about the 1990s icolous taxa in Arthonia is remarkable. The somewhat out- (e.g. Frahm et al. 2009), whereas even common acidophytic dated, but still most comprehensive key for lichenicolous species like Hypogymnia physodes responded by decreasing Arthonia species at a world wide scale (Clauzade et al. 1989) thallus population numbers (e.g. Herk et al. 2003). As * Corresponding author. E-mail address: [email protected] (M. Grube). http://dx.doi.org/10.1016/j.funbio.2016.06.009 1878-6146/ª 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved. 1342 A. Fleischhacker et al. a consequence lichenicolous fungi confined to X. parietina as agarose gel stained with GelRed (Biotium, Hayward, Califor- their host face a situation with much more substrate available nia, USA) under UV light. PCR products were purified either us- for colonization. The fungus flora of decaying thalli of X. pari- ing the QIAquick PCR Purification Kit (Qiagen) or gel extracted etina has already been studied by Etayo & Berger (2009).Itis and purified using the QIAquick Gel Extraction Kit (Qiagen). dominated by anamorphic states of both ascomycetes and ba- Sequencing of the amplicons was provided by Uppsala Ge- sidiomycetes. In contrast, vital thalli are colonized by a differ- nome Centre (Sweden). The sequences were assembled in Bio- ent set of species. The entire fungal flora of X. parietina as Edit v.7.1.3 (Hall 1999) and their identity checked with the Blast single host species was studied in a project resulting in the search in GenBank and the local Blast functionality imple- master thesis for one of us (Fleischhacker 2011). mented in the BioEdit v.7.1.3 package. The nLSU sequences One of the fungi we frequently came across during this sur- were aligned in RNAsalsa (Stocsits et al. 2009) using the sec- vey was an Arthonia species, commonly determined as Artho- ondary structure of Saccharomyces cerevisiae (nLSU; Kjer 1995) nia molendoi (e.g., Santesson et al. 2004). However, a closer as template. The final alignment comprised 676 sites and the analysis revealed fine phenotypic differences, and here, we phylogeny was reconstructed with a Maximum Likelihood also confirm the distinctiveness of this lichenicolous fungus (ML) approach using RAxML (Stamatakis 2014) with raxmlGUI by molecular data. We therefore describe the species below interface (v.1.5, Silvestro & Michalak 2012). Myriostigma candi- as new to science and provide a discussion of apparently re- dum was used as outgroup lineage. A GTR þ GAMMA model lated species. was applied and 1000 bootstrap replicates were run. The phy- logenetic tree was visualized in FigTree v.1.4. In addition to the specimens cited below together with the Material and methods treatment of Arthonia parietinaria, the following material was compared: Methods for phenotype analyses Arthonia anjutae S. Kondr. & Alstrup (on Teloschistes spinosus For the morpho-anatomical investigation air-dried herbarium (th.)) specimens have been analysed. External morphology was Note: Following ICN Art. 60.12, Note 4, the gender of the epi- e  studied with a dissecting microscope (WILD M3, 6.4 40 ). An- thet with the original spelling ‘anjutii’ is corrected because atomical studies of the thallus and the ascomata were carried the epithet is derived from the female first name Anyuta. e  out under the light microscope (LEICA DMRE, 100 1000 ). Sec- Australia: South Australia, Roopera station near Whyalla, tioning was performed with a freezing microtome (LEITZ, sec- eastern fence of Extension Paddock, 1969, R. Rogers, det. J. e m tions of 12 15 m) but squash preparations were also used, Hafellner (herb. Hafellner 19576). especially for ascus analysis. Preparations were mounted in water. When necessary, contrasting was performed by a pre- Arthonia clemens (Tul.)Th.Fr. (on Rhizoplaca chrysoleuca (ap.)) treatment with lactic acidecotton blue (MERCK 13741). Amy- Europe: Italy: Prov. Sondrio, Ortler Gruppe, Pian di Cembro € loid reactions in hymenia were observed both progressively uber dem Val di Cortena, an der Straße von San Pietro nach and regressively by the use of Lugol’s reagent (MERCK 9261). Trivigno, 1400e1450 m alt., 1975, H. Mayrhofer (herb. Hafellner Conidiogenesis of pycnoconidia was studied in erythrosin B 933 in GZU). e Asia: Russia: Siberia, Chukotka, on the upper (ALDRICH 19,826-9) in 10 % ammonia. Sections and squash reaches of the river Milkera, 1977, M. P. Andreev, det. J. Hafell- preparations were not pretreated with KOH unless otherwise ner (GZU). stated, namely for the detection of hemiamyloid reactions in Arthonia epiphyscia Nyl. the hymenium. Measurements refer to dimensions in tap wa- Hosts: Physcia caesia (th.) (1, T), Physcia dubia (th.) (2), Physcia ter. The terminology of pycnidial characters follows Vobis & phaea (th., ap.) (3). Hawksworth (1981). Europe: Austria: Burgenland, Sudburgenland,€ in Deutsch- Kaltenbrunn, ca. 265 m, GF 8962/2, (1), 1990, J. Hafellner 26075 Molecular analyses & W. Maurer (GZU). e Karnten€ (Carinthia), Nationalpark Hohe Tauern, Glockner-Gruppe, S Hange€ des Fuscherkar Kopfes, N Hymenia (about 20e50  50e100 mm in size) of four to five ober der Hofmannshutte€ [470501000N/124402000E], ca. 2500 m, ascomata of the lichenicolous species were carefully cut GF 8942/1, (3), 1988, J. Hafellner 32049, M. Walther, & A. Hafell- from the host tissue, placed in 1.5 ml microtubes using a ster- ner (GZU). e Steiermark (Styria), Niedere Tauern, Wolzer€ ilized forceps (Dupont Nr. 5), and used for a direct PCR ap- Tauern, Hochgroßen,€ nordostlicher€ Seitengipfel, im Gratber- proach (Grube 2005). Parts with either amorphous pigments eich [472705000N/141503500E], ca. 2050 m, GF 8551/2, (2), 1989, J. (incl. melanized portions) were removed from the cuttings Hafellner 26309 (GZU). e Norway:Oppland,Gem.Vaga, N- as far as possible as these pigments impair PCR efficiency. Ufer des Sees Vagavatn ca. 2 km W von Vagamo, ca. 450 m, PCR reactions of 20 ml (DNA extractions) and 30 ml (direct (2), 1984, J. Hafellner 11424 & A. Ochsenhofer (herb. Hafellner). PCR) were used. Each 10 ml of PCR mix contained 0.5e1 ml geno- Arthonia molendoi (Heufl. ex Frauenf.) R. Sant. (on Xanthoria mic DNA extraction (or the lichen sample), 0.4 ml of each À elegans (th., ap.)) primer (20 pmol ml 1), and 5 ml HotStarTaq Plus Mastermix Europe: Austria:Karnten€ (Carinthia), Hohe Tauern, Kreuzeck- Kit (Qiagen). Primers LIC24R and LR7 were used for PCR ampli- Gruppe, Schwarzsteinwande€ (Sudw€ ande€ des Schwarzsteins) E fication of nuclear ribosomal large subunit RNA gene frag- der Hochtristen, N der Turgger Alm [464705500N/130901000E], ments (nLSU) (Vilgalys & Hester 1990; Miadlikowska & 1950e2150 m, GF 9244/2, 1978, J. Hafellner 3875 (GZU). e Lutzoni 2000). The PCR products were visualized in a 0.5 % A. parietinaria e A common but frequently misunderstood lichenicolous fungus 1343 Steiermark (Styria), Nordalpen (Nordliche€ Kalkalpen), ‘Ragnitzbach’ at lower end of Dr Hanischweg, 470403500N/ Ennstaler Alpen, Gesauseberge€ E von Admont, Gr. Buchstein, 152805000E, ca. 380 m s. m., GF 8958/2, old orchard, on canopy NW-Abhange,€ am W-Fuß der Abbruche€ der Admonter Frauen- branches of recently felled Juglans regia,onXanthoria parietina, mauer, 473605500N/143503000E, ca. 1720 m, GF 8353/4, 2005, J. 1 July 2010, J. Hafellner 77067 & W. Obermayer (GZU e holo- Hafellner 67595 & A. Hafellner (herb. Hafellner). e Liechten- type, BCN, BR, CANB, E, GZU, LE, M, NY, PRM, UPS e isotypes). stein: Eastern Alps, Ratikon,€ mountain ridge between Aug- Isotypes to be distributed in Hafellner, Lichenicolous Biota no. stenberg and Nospitz, SSW above the village Malbun, S adhuc ined. above Vaduzer Tali,€ 470502000N/093601500E, ca. 2060 m, 2008, Etymology: parietinarius 3 (Lat.), belonging to [Xanthoria] J. Hafellner 72834 (GZU). e Spain: Prov. Gerona, Pyrenaen,€ parietina. Nuria N von Ribas de Freser, NE von der Bergstation der Zahn- Iconography: Vezda 1970: 223, Fig 1 (drawings of asci, asco- radbahn, 2100e2200 m, 1983, J.