Assessing Past Agrobiodiversity of Prunus Avium L. (Rosaceae): A

Veget Hist Archaeobot DOI 10.1007/s00334-011-0310-6

ORIGINAL ARTICLE

Assessing past agrobiodiversity of Prunus avium L. (Rosaceae): a morphometric approach focussed on the stones from the archaeological site Hoˆtel-Dieu (16th century, Tours, France)

Pauline Burger • Jean-Frederic Terral • Marie-Pierre Ruas • Sarah Ivorra • Sandrine Picq

Received: 18 October 2010 / Accepted: 20 July 2011 Springer-Verlag 2011

Abstract Abundant and diverse Prunus fruitstone remains century Hoˆtel-Dieu cesspit at Tours, France, revealed that from cherries, plums, sloes, peaches, etc. are frequently the morphological diversity is structured into two distinct recovered from archaeological waterlogged contexts such as morphotypes which diverge mainly according to geometri- wells, latrines, lake dwellings etc. in Europe. The distinction cal features. Finally, the comparison between morphological between most of the Prunus species, based on traditional features of these well-preserved archaeological stones and morphological characters of the fruit stones, is usually not modern reference material including P. avium, P. cerasus problematic. However the discrimination between P. avium and P. 9 gondouinii, suggests that these two morphotypes, L., P. cerasus L. and related cherry species, based on clas- which have been initially attributed to P. avium (long stones) sical criteria alone, often turns out to be ambiguous because and P. avium/cerasus (rounded stones) according to tradi- of the increasing number of varieties which have been bred tional morphological parameters, would correspond to two since Roman times. By combining geometric and traditional different cultivated varieties, both belonging to Prunus morphometrical approaches, the overall variation in shape avium. Results presented in this work constitute new and and size of stones from French and Swiss excavations dating preliminary data obtained during the development of this from the 1st century to the 16th century A.D. were assessed. project that throw light on morphological variability and Among these important archaeobotanical data, the detailed biosystematic aspects. examination of 100 waterlogged stones from the 16th Keywords Agrobiodiversity Á Archaeobiology Á Communicated by M. Latałowa. Cultivars Á Morphometrics Á Morphotypes Á Prunus avium

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P. Burger (&) Á J.-F. Terral Á S. Ivorra Á S. Picq Most Prunus species are deciduous and often spiny trees Centre de Bio-Archeólogie et d’Ecologie (CBAE) (UMR 5059 and shrubs with fruits as drupes, usually with a juicy fruit CNRS/Universite´ Montpellier 2/EPHE), Equipe Ressources flesh (Hanelt 1997; Rehder 1940). The Prunus genus which Biologiques, Socie´te´s, Biodiversite´, Institut de Botanique, 163 rue Auguste Broussonet, 34090 Montpellier, France comprises more than 400 species (Rosaceae family; Lin- e-mail: [email protected] naeus 1753), is widely distributed in the northern hemi- sphere with many wild and cultivated representatives in J.-F. Terral Europe. It is an economically and ecologically important Universite´ Montpellier 2, Place Euge`ne Bataillon, 34095 Montpellier cedex 5, France group with many cultivated species, notably P. dulcis (Mill) D.A.Webb (almond), P. armeniaca L. (apricot), M.-P. Ruas P. persica (L.) Batsch (peach), and P. domestica L. l.s. Archeózoologie, Archeóbotanique-Socie´te´s, Pratiques et (plums, in the wide sense). Despite the geographical range Environnements (AASPE) (UMR 7209 CNRS/MNHN), Museúm National d’Histoire Naturelle, 55 rue Buffon, 75231 and the popularity of this genus, the evolution history and Paris Cedex 05, France taxonomy of Prunus remain unclear. Progenitors of many 123 Veget Hist Archaeobot species and cultivars are often hypothesized, but not highly regarded. The establishment of the cherry orchard definitively identified. (‘‘Kirschgarten’’) tradition in northern and central Europe Prunus avium L. (sweet and wild cherries), P. cerasus would be linked to the increasing medieval taste for these L. (sour, dwarf or morello cherry), P. fruticosa (ground flavoursome fruits (Kroll 2007; Kroll and Willerding 2004). cherry) and P. 9 gondouinii (duke cherry), along with P. cerasus was well represented in these orchards, alongside related cherry species and their interspecific hybrids con- P. mahaleb, P. fruticosa and P. avium in the early Slavonic stitute the Eucerasus section, part of the Cerasus (Mill.) stronghold of Mikulcˇice in Moravia (Opravil 1998) and in Focke subgenus of the genus Prunus (Santi and Lemoine several medieval sites located in eastern central Europe 1990; Thorne 1992). This classification was defined based (Kroll 2007; Kroll and Willerding 2004; Medovic´ 2004). on morphological criteria by Rehder (1947) and later by The duke cherries from the fertile P. 9 gondouinii Rehd. Kru¨smann (1978), and has been further confirmed by species, previously known as P. acida Dum, Cerasus reg- chloroplast DNA variation analysis (Badenes and Parfitt alis or Prunus avium ssp. regalis, seem to be intermediate 1995). between P. avium and P. cerasus (Dirlewanger et al. 2009; The ancestors of the modern P. avium (sweet cherries) Faust and Suranyi 1997; Saunier and Claverie 2001). Based seem to have originated around the Caspian and Black on the analysis of 75 AFLP markers, P. avium and P. cer- Seas, from where they have slowly spread, a phenomenon asus were established as the progenitors of P. 9 gondouinii initially driven by birds, hence the species name P. avium Rehd. (Tavaud 2002; Tavaud et al. 2004): duke cherries (Dirlewanger et al. 2007). Requiring warm and dry sum- would result from the pollination of sour cherry by unre- mers, but adequate rainfall or irrigation during the growing duced gametes of sweet cherry (Iezzoni et al. 1990). season, the natural range of P. avium covers the European Since antiquity in Europe, archaeobotanical data have temperate regions from the southeastern part of Russia to recorded a significant increase in fruit species diversity the northern part of Spain (Hedrick 1915). According to (Ruas 1992, 1996; van Zeist 1991). In his Natural History, archaeobotanical data, the wild P. avium seems to have Pliny the Elder (1st c. A.D.) mentioned notably nine vari- been collected by Mesolithic hunter-gatherers in southern eties of sweet cherries planted in Rome (Andre´ 1981), France, but very few charred stones have been found suggesting that several cherry varieties had already been (Vaquer and Ruas 2009). Most of the other mentions from imported and probably cultivated at this time in the Roman Mesolithic sites in northern Europe seem doubtful (Bakels southern provinces, such as the Narbonnaise in France 1991). The fruit are more frequently recorded from the (Ruas et al. 2006). From this period, a morphological Neolithic and Bronze Ages 5500–4000 B.C. (Hedrick 1915; diversification of Prunus stones was noticed which is Marshall 1954; Bertsch and Bertsch 1949, Out 2009). It is related to, after written sources, an increasing number of suggested that sweet cherries may either be indigenous in fruit varieties (Amigues 2002; Andre´ 1981; Raspail 1838; southern Europe as small isolated populations, or may have Quellier 2003). In order to identify more precisely the fruit been introduced in these areas since the Neolithic. remains found in archaeological excavations and particu- The ground cherries, from the most cold hardy cherry larly to distinguish varieties among these taxa, various species P. fruticosa, are widespread over the major part of typological analyses based on macroscopic characters of central Europe, Siberia and northern Asia (Hedrick 1915). plum and sloe stones have been proposed over time by Based on fossil evidence, P. cerasus seems to have several authors (Baas 1974;Ro¨der 1940; Rybin 1936; originated around the Caspian Sea, from an area very sim- Werneck 1958). In 1978, Behre developed a typological ilar to that of sweet cherries, and it appears to be native to analysis of stones by defining a number of ‘Prunus northwest and central Europe (Dirlewanger et al. 2009; Formenkreise’ or types during his analysis of the Watkins 1995). The hybrid origin of P. cerasus was first P. domestica L. stones from Haithabu, Germany, 9th–11th suggested by Olden and Nybon (1968) on the basis of centuries A.D. These ‘Prunus Formenkreise’ were defined morphological and biochemical evidence. Continuous according to stone size, surface sculpture and shape variations between the P. fruticosa and P. avium charac- (finding expression in the indices), and were then related to teristics were observed in sour cherry throughout its geo- modern varieties. This method has been applied by various graphical range. P. cerasus seems to be closer to P. avium in archaeobotanists to stones from numerous European western Europe, whereas it is most closely related to excavations and used notably by van Zeist and Woldring P. fruticosa in eastern Europe (Hillig and Iezzoni 1988; (2000) to characterise P. domestica L. endocarps from the Krahl et al. 1991). Many genetic studies have confirmed the late- and post-medieval occupation deposits in the town hybrid origin of P. cerasus and have shown P. fruticosa and centre of Groningen, The Netherlands. Based on physical P. avium as its progenitors (Hancock and Iezzoni 1988; and morphological characteristics, the authors distin- Santi and Lemoine 1990; Schuster and Schreiber 2000). guished 13 different types of plum, some related to recent During the early Middle Ages, cherries and plums were varieties. 123 Veget Hist Archaeobot

At the same time, Kroll (1978) proposed criteria to dis- – Interpret these results in term of agrobiodiversity by tinguish P. avium and P. cerasus stones from the medieval comparison of archaeo-diversity with a collection of and modern waterlogged material from Lu¨beck, Germany. modern stones from P. avium, P. cerasus and The overlapping of quantitative size and shape features did P. 9 gondouinii varieties. not allow identification of all stones except those with – Open new perspectives in the research of the history of extreme phenotypes. More recently, Kroll (2007) showed the varietal inheritance of the cherry. that archaeological P. avium and P. cerasus could be easily differentiated on the basis of traditional morphological features such as surface sculpture, dimensions, etc., pro- Materials and methods vided that the morphological features of the fruitstones were observable, despite a long period of deposition. The study The cherry fruit is a drupe consisting of an exocarp, a thick was carried out with success for the Haithabu excavation and fleshy mesocarp and a hard and woody stone (endo- where the early Middle Age material was identified as carp) surrounding and protecting a single seed. The stone is P. cerasus. For later periods, the author considers that a structure with bilateral symmetry consisting of two identification of cherry stones becomes more problematic pseudo-valves deriving from a single carpel, sutured on the because of the disappearance of the morphological features. margin (Figs. 1, 2). Indeed, we have noticed that physical characteristics such as the cherry stone surface sculpture and hilum tend to Archaeological stones disappear due to erosion through time and because of the type of deposit, for example acidity in the cesspit. So the The material analyzed is based on 717 well-preserved efficiency of this method is often limited, particularly in stones originating from Roman and medieval archaeolog- the case of cherry stones, which are often classified under ical features dating from the 1st to the 16th century A.D., the generic term of P. avium/cerasus (Baas 1951). These and from ten French and three Swiss sites (Table 1). taphonomic constraints add to the problems related to inter- Among this collection of cherry stones from various specific hybridization and cultivation practices which tend excavations, we have particularly examined material from to increase the morphological diversity. In addition, the the cesspit of the 16th century site of the Hoˆtel-Dieu at very local aspect of varietal improvement, the probable Tours, France. It comprises 100 stones of Prunus spp. disappearance of ancient varieties, and the dissimilarity among many other waterlogged remains of food waste between these ancient varieties and the modern ones, all from a hospital. A substantial variability of stone mor- complicate cherry species identification even more. Diffi- phologies according to shape, size and hilum structure had culties in circumscribing species due to lack of diagnostic already been noticed during the archaeobotanical study, characters are also known from the genus Malus where suggesting the existence of two mixed populations, one widespread crossability, introgression and cultivation may blur taxonomic boundaries (Dickson et al. 1991). Recently, in their study of Prunus stones from the Roman vicus Tasgetium (Eschenz, Switzerland), Pollmann et al. (2005) obtained promising results by using ancient DNA to answer archaeobotanical issues. Such archaeobotanical research is supported by the development of several regional centres for conservation of ancient cultivars in Europe (Chauvet 1999;Ko¨rber-Grohne 1996). According to this scientific context, the present work aims to apply to Prunus avium/cerasus stones an analytical method previously successfully used in the study of Olea europaea L. (olive) stones (Terral et al. 2004; Newton et al. 2006) in order to: – Evaluate the range of stone shape variation shown by the whole of the bioarchaeological material at our disposal. – Distinguish morphotypes on the basis of the model of Fig. 1 Two different morphological types of cherry stones recog- the 16th century Hoˆtel-Dieu archaeobotanical material nized within the Hoˆtel-Dieu archaeological material, 16th century, from Tours, France. Tours, France; scale bar 5mm

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Fig. 2 Analytical processing for quantifying the morphological structure of a cherry stone photographed in ventral view and morphometric (shape and size) attributed possibly to P. avium (the longer ones), the other angles or ratios—but the overall object shape is not really possibly to P. cerasus or P. avium/cerasus (the shorter and measured (Rohlf and Marcus 1993). Considering only rounder ones) (Ruas, unpublished) (Fig. 1). simple measurements, this approach is not nearly as powerful as it could be, since it does not take into account the Comparative reference material geometrical relationships among these data. To solve this limitation, we have combined shape morphometry with To gain insight into the significant identification characters, traditional measurements, as has been done previously for the present study used reference material composed of 542 other fruits such as olive (Terral et al. 2004) and Prunus L. stones from various cherry varieties. Thus, we have col- section Prunus (Nielsen and Olrik 2001; Depypere et al. lected stones from both old and current varieties of 2007, 2009). Shape morphometrical methods are effective P. avium (N = 419), P. cerasus (N = 98) and P. 9 in capturing information about the three-dimensional shape gondouinii (N = 25) obtained from INRA (Institut of biological objects and in testing for differences in shapes National de Recherche Agronomique, Bourran, France) within and among samples of organisms. These powerful (Table 2). P. fruticosa stones were not included in the statistical procedures were applied in exploratory studies in study as they cannot be easily distinguished from wild taxonomy and evolution (Rohlf and Marcus 1993). types of P. avium (Olden and Nybom 1968). In this work, each stone was photographed in ventral view with a digital camera (60 mm f/2.8D lens) placed at Morphometrical analyses the fixed distance of 35 cm from the stones. Photographs of stones were first cut out, and then converted to black-and- Size analysis corresponds usually to discrete measure- white before being resized (Fig. 2). This procedure was ments—distances between defined points (landmarks), and carried out with R software (R Development Core Team

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Table 1 Archaeological material analysed Archaeological sites, town (department), context N Dating century Excavation supervisor Archaeobotanist A.D.

France Place d’Assas, Nıˆmes (Gard), well 42 1st F. Conche N. Rovira Rue Ste Catherine, Vannes (Morbihan), well 41 1st A. Triste M.-P. Ruas BK14050, Biesheim-Kunheim, Haut-Rhin, cesspit 32 1st–2nd M. Redde´ P. Vandorpe BK14064, Biesheim-Kunheim, Haut-Rhin, cesspit 44 1st–2nd M. Redde´ P. Vandorpe BK14104, Biesheim-Kunheim, Haut-Rhin, cesspit 31 1st–2nd M. Redde´ P. Vandorpe La Roquette, Cavillargues (Gard), well 29 4th–5th B. and H. Petitot, S. Alix L. Bouby Rue des veaux, Strasbourg (Bas-Rhin), Ill river banks 44 10th–12th M. Werle´ C. Schaal Charavines, Colletie`re (Ise`re), handcraft shops 41 11th E. Verdel, M. Colardelle K. Lundstro¨m-Baudais/C. Schaal Charavines, Colletie`re (Ise`re), extension area 50 11th E. Verdel, M. Colardelle K. Lundstro¨m-Baudais/C. Schaal Charavines, Colletie`re (Ise`re), stalling area 45 11th E. Verdel, M. Colardelle K. Lundstro¨m-Baudais/C. Schaal Place Me´te´zeau, Dreux (Eure-et-Loir), cesspit 21 12th P. Dupont M.-P. Ruas Rue du rempart Etampes (Essonne), unknown 24 12th X. Peixoto M.-F. Sellami Place Me´te´zeau, Dreux (Eure-et-Loir), cesspit 24 16th P. Dupont M.-P. Ruas Place de la cathe´drale, Tours (Indre-et-Loire), cesspit 100 16th A.-M. Jouquand M.-P. Ruas Switzerland Oberwinterthur, (Zu¨rich canton), cesspit OWKW76 23 1st P. Vandorpe Oberwinterthur, (Zu¨rich canton), cesspit OWKW78 35 1st P. Vandorpe Oberwinterthur, (Zu¨rich canton), cesspit OWKW78(1) 25 1st P. Vandorpe Schoffelgasse Zu¨rich, (Zurich canton), cesspit 34 13th M. Ku¨hn Hallwyl Castle, Seengen, (Argovie canton), ditch 32 14th–15th M. Ku¨hn

2005) using the functions proposed by Claude (2008)(R this study, a standardized Principal Component Analysis functions are indicated in italic). (PCA) was carried out on 717 stones and eight quantitative Geometrical analysis of the stones was carried out fol- parameters presented previously (four by polynomial equa- lowing the procedure developed by Terral et al. (2004). In tions/two equations by stone). ventral view, the two external half outlines between two Morphological variation in the archaeological material homologous landmarks, the base of stone and the apex, are shown by PCA was then analysed by testing correlation obtained with the Conte() function and were similarly between the size features measured and new coordinates of adjusted in an orthonormed basis and standardized by size cherry stones in the PCA space. The combination of results with the BooksteinM() function (Baseline superimposition) from shape and size analyses allowed us to test a possible (Bookstein 1991). Then, a least-squared third-degree allometric phenomenon between shape and size which are polynomial curve was fitted to each half outline defined by a priori two independent parameters. Especially for the 20 equally spaced points (Fig. 2). Finally, two third degree Hoˆtel-Dieu archaeological stones, PCA results have been polynomial equations described the external geometrical thought of as revealing the internal structure of the data. structure of the stone. The eight quantitative parameters Finally, a discriminant analysis (DA) was carried out on (four by half outline) obtained may be easily used as 542 stones from a non-exhaustive collection of material variables in multivariate statistical analyses. from modern varieties belonging to P. avium, P. cerasus and In addition, length (mm), thickness (mm) and area P. 9 gondouinii, eight quantitative variables (the eight (mm2) were measured in ventral view using computerized shape parameters from the polynomial morphometrical image analysis systems on digitized photographs (Fig. 2). analysis) and one qualitative parameter with three distinct modalities, corresponding to the allocation of stones to a Statistical analyses Prunus species. This multivariate statistical analysis aimed to test morphological differentiation between fruitstones of In order to evaluate the range of shape diversity in the modern Prunus species and finally, to provide tangible ele- P. avium/cerasus archaeological material at our disposal for ments for identifying the Hoˆtel-Dieu archaeological stones.

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Table 2 Prunus species, Species Cultivated variety N varietal denomination, origin and number of modern stones Prunus avium Alex III 10 analysed as reference material (in brackets, plant material— Argot 13 Prunus species or variety—used Black Star (Prunus mahaleb)19 as rootstock) Brooks (Edabriz) 17 Early Bigi 10 Early Star 18 Europepice 93-17 (MM14) 17 Europepice 94-04 (MM14) 29 Firmred 10 Giant Red (MM14) 28 Grace Star 10 Grace Star (Prunus mahaleb)21 Lalastar 10 Lodi 20 Masdel Kabel 10 Panaro 1 Sweet Early (Prunus mahaleb)21 Penny 10 Ruby 3 Sandra Rose 10 Santina 10 Simcoe Probla 7 Skeena (MM14) 9 Sumbigo 10 Sumcoro 10 Sumele 8 Sweet Early Panaro 1 20 Tieton 10 V3648 12 V3868 20 Vanda 7 Victor 10 Prunus cerasus Griotte du Nord 5 Haut-Rhin acide 9 Olivet Haˆtive 5 Olivet Tardive 6 Reine Hortense 2 Toulennea 71 Prunus 9 gondouinii Cerise Cure 4 All the accessions come from Griotte de Provence 7 INRA, Bordeaux, France Gros Guin de Cœur 7 a From different clonal Impe´ratrice Euge´nie 7 individuals

Results (x [ 0). The second axis (PCA2), orthogonal to the PCA1, explains 18.1% of the variability and contributes to dis- The 2-dimensional plot (PCA1-2) explaining 93.2% of the tinguish asymmetrical elliptic stones (y [ 0) and rather total inertia shows that archaeological stones are mainly symmetrical elliptic/oval stones (y \ 0) (Fig. 3). In any distributed along the ﬁrst factorial axis (PCA1), expressing case, as no clear structure in morphological diversity was 75.1% of the total morphological variability. This axis highlighted, we have hypothesized that the ﬁrst axis appears to differentiate ovate (x \ 0) from obovate stones expresses shape variation induced by size differences, and

123 Veget Hist Archaeobot independently the second axis allows discrimination of the Table 3 Tests of linear correlation between stone morphometrical stones according mainly to shape features. descriptors: size measurements (length, thickness and area) and geo- In order to test this hypothesis, a correlation analysis metrical features (new coordinates in the two first dimensions of PCA); R = Pearson correlation coefficient between each morphometric descriptor was carried out (Table 3). The results show that the existence of correlations Tested correlations RP-value Significance among size variables emphasizes the limits of traditional Length–thickness 0.34 \0.0001 *** morphometry to characterize stones and to discriminate Length–area 0.65 \0.0001 *** different statistical populations, or Prunus morphotypes. In PCA1–length 0.35 \0.0001 *** addition, size is generally influenced by environmental PCA1–thickness -0.23 \0.0001 *** factors such as climatic and edaphic parameters, or human PCA1–area 0.13 \0.001 *** practices in the case of cultivated plants. In addition, as PCA2–length -0.10 0.007 * the correlation between traditional characters and PCA1 is PCA2–thickness -0.05 0.21 ns significant, the first PCA axis reflects a pattern of size PCA2–area 0.06 0.10 ns variations, from small (x \ 0) to large stones (x [ 0). Independently, even if a slight correlation between PCA2 n.s. not significant and ‘length’ was noticed, the second axis revealed shape * Significant; *** highly significant differences (Table 3). Thus, these results underline an allometry phenomenon, thus a relationship between size and shape, however almost non-existent on the second axis of the these two morphotypes attributable to PCA2 was validated PCA, which explains mainly shape variations. by an analysis of variance (ANOVA) carried out on the new After evaluation of the overall range of morphological coordinates (PCA1 and PCA2) of stones in the morpholog- diversity in the archaeological material, geometrical vari- ical space (PCA biplot 1-2) (Table 4). ability of the archaeological stones from the Hoˆtel-Dieu site In order to find out the systematic status of these two (Tours, France) was precisely examined. Coordinates of Prunus forms, we have then performed a discriminant these stones in the first axis of PCA appeared relatively analysis (DA) on the reference material (542 modern homogeneous and normally distributed (Shapiro–Wilk stones) presented in Table 2. The overall discriminant normality test: W = 0.99, P-value = 0.87), while those power computed by the discriminant analysis is equal to concerning PCA2 were not (Fig. 4a). Their frequency dis- 86.5% in which 93.2% and 77.6% of stones belonging to tribution, whose bimodality was tested, implies two signif- P. avium and P. cerasus respectively are well-differenti- icant distinct populations of stones which may correspond to ated from their relatives. However, the discrimination rate two different shape morphotypes (Fig. 4b). The existence of of P. 9 gondouinii stones is weak, only equal to 8%.

Fig. 3 PCA analysis biplot 1-2 showing the overall morphological variation of 717 cherry stones from 13 archaeological sites

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Fig. 5 Discriminant analysis biplot 1-2 showing: a a pattern of shape c variation in Prunus spp.; b the location in morphological space, of the Hoˆtel-Dieu archaeological stones allocated to Prunus avium

presents shared or intermediate morphological features as shown by previous studies (Dirlewanger et al. 2009; Faust and Suranyi 1997; Saunier and Claverie 2001). When compared to this preliminary discriminant model as additional samples, the Hoˆtel-Dieu archaeological stones were assigned to a Prunus species. The probability (P) that archaeological stones belong to a species was calculated using the Mahalanobis distance between stones and each group centroid in the morphological space defined by the discriminant analysis. We have considered the allocation reliable and very accurate if P [ 0.80. In this case, 91 archaeological stones were attributed to P. avium. More- over, five stones were allocated to P. avium with a probability ranging between 0.7 and 0.8, and four were not classified (P \ 0.7) (Fig. 5). Finally, according to results from the Hoˆtel-Dieu archaeological material structured in two different morphotypes (Figs. 4, 5), we suggest that these distinct morphotypes correspond to two different cultivated varieties of P. avium.

Discussion

By comparing stone size, surface sculpture and shape of archaeological P. domestica L. stones with modern varieties, Behre (1978) began morphometric studies on archaeobotanical material. Traditional stone parameters (dimension, ratio and surface sculpture description) were used to characterise archaeological cherry stones. But for more recent historical periods, it appears that these characteristics are not sufficient to differentiate cherry species because of the overlapping ranges of stone size and shape parameters Fig. 4 a Bimodal distribution of the Hoˆtel-Dieu stones in relation to (Kroll 1978), probably resulting from centuries of cultiva- PCA2 scores. Results from the normality test (Shapiro–wilk test) are presented for each population/morphotype identified. b The two tion and hybridization. Most archaeobotanists consequently distinct morphotypes highlighted in the Hoˆtel-Dieu archaeological expressed reservation about this type of analysis and they material, emphasized within the overall morphological diversity continued to use the term of P. avium/cerasus for archaeo- shown by PCA1-2 biplot botanical fruitstones. In this context, the originality and the innovation of our study is to apply morphometrical methods The DA biplot 1-2 illustrates this pattern. It shows that combining geometric and traditional approaches to assess canonical scores of the first axis, explaining 94.7% of the the overall variation in shape and size of cherry stones, as total variance, contribute highly to discriminate the three previously achieved successfully for other fruit species of Prunus species (Fig. 5). The second axis (5.3% of the total interest such as Olea europaea L. (Terral et al. 2004) and variance) structures the morphological diversity according Vitis vinifera L. (Terral et al. 2010). to shape differences, following most probably an analogous The co-occurrence of two Prunus avium varieties in trend shown by the PCA carried out on archaeological Tours is not surprising given the technical improvement of stones. Morphological variability of P. 9 gondouinii fruit cultivation during the Renaissance. Indeed, in northern appears to be significantly higher than that of its ancestors. France, cherries became highly marketed products. They Compared to its parental relatives, P. 9 gondouinii were intensively cultivated in orchards and not only in

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Table 4 Results of testing the division of the Hoˆtel-Dieu Prunus stones into two distinct statistical populations (or morphotypes) using analysis of variance (ANOVA) applied to principal components (PCA1 and PCA2) of the PCA carried out on shape descriptors Tested principal component Statistical populations Analysis of variance (ANOVA) POP1 POP2 Wilk’s Lambda F-test DF P-value Mean SD Mean SD

PCA1 0.714 0.466 0.581 0.592 0.986 1.420 1, 103 0.236 n.s. PCA2 -0.3 0.124 0.062 0.146 0.380 167.990 1, 103 \0.001* POP statistical populations (see Fig. 4), SD Standard deviation at P = 0.05, DF degree of freedom, n.s. not significant * Highly significant gardens of the elite as a result of curiosity (Quellier 2003). In taxonomical, bio-archaeological and historical perspectives. the current state of research, results highlight the fact that the The combination of geometrical morphometry (baseline morphometrical method used in this work allows us to rule superimposition method) and traditional measurements out the doubt remaining about the characterization of the brings clues to define archaeological morphotypes and to stones in the cesspit. According to a rental dated from 1587, link them to current species or forms. In the future, we need the Hoˆtel-Dieu hospital was occupied both by religious to increase our data corpus of the available reference and personnel of the institution and people affected by various archaeological material. Study of a larger number of refer- diseases, notably leprosy (Jouquand et al. 1996). The social ence stones from wild forms of cherry, diverse varieties and position of these occupants could be related to the con- various origins and of archaeological stones from an sumption of two different cherry varieties. The mention of a increasing number of excavation sites would probably pro- garden in the hospital suggests that these fruits and other vide important results. We particularly need to include plants were probably produced locally (Jouquand et al. archaeological cherry stones from northwestern and eastern 1996). Nevertheless, these results should be interpreted with European sites and notably ancient P. cerasus stones. We caution, as the relevance of the use of stones from modern will also improve the methodological approach by inte- cherry cultivars as reference material for identification of grating analysis of stones performed in different orienta- ancient stones from archaeological contexts could be biased tions, using powerful methods such as the Elliptic Fourier by hybridisation and introgression. We cannot exclude that Transforms (EFT) and 3D analysis. Finally, such a study these stones may correspond to P. fruticosa as the mor- opens new and interesting perspectives into the under- phology of its stones overlaps with wild P. avium ones. standing of the biogeographical and evolution history of the Moreover, in the present state of research, it is impossible to cherries. In addition, it will be fascinating to reveal if dif- identify the origin of these cherries. Were these varieties ferent varieties were consumed by different social categories cultivated in the Tours region? Did the cherries come from of people since the Roman period in the provinces, and to more distant places to be finally marketed in Tours? discuss the role of these varieties in marking social dis- To be more accurate, this study should be completed by tinction. Indeed, the economic and social status of cherries the analysis of archaeological stones from different his- has changed in European societies between classical and torical periods and different geographical origins. We medieval times. These fruits played an important social role should also use our method to confirm or re-evaluate the in the medieval elite diet regime (Grieco 1996) before nature of archaeological fruitstones already attributed to a becoming a more common fruit during the later centuries particular species based on traditional parameters. These (Quellier 2003). promising preliminary results open new and interesting perspectives on the assessment of cherry agrobiodiversity Acknowledgments We wish to thank all the archaeobotanists at different taxonomic levels (species, subspecies and (Bouby L, Derreumaux M, Hallavant C, Ku¨hn M, Rovira N, Schaal C, Sellami M-F, Vandorpe P, Woldring H, Zech-Matterne V), the exca- variety) and on the understanding of its cultivation and vation supervisors (Demolon P, Dupont P, Jouquant A-M, Plumier J, consumption history in Europe. Re´dde´ M, Triste A, Verdel E and Colardelle M) and all the people who collected and sent us stones from archaeological excavation sites in France, Switzerland and Belgium. We also are grateful to Ste´phanie Mariette (UREF—Unite´ de recherche sur les espe`ces fruitie`res, Conclusions Villenave d’Ornon, France) and to Evelyne Leterme (Domaine de Barolle—Montesquieu, France) for their kind cooperation in the col- The morphometric study presented in this work constitutes lection of reference stones respectively from the INRA and the CVRA orchards. Finally, we would like to thank Emma Passmore who helped an innovative archaeobiological contribution intended to us to improve our English. We also would like to thank the two characterize past cherry agrobiodiversity in methodological, anonymous referees for their helpful comments and suggestions. This

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