Low Genetic Variation in Subpopulations of an Endangered Clonal Plant Iris Sibirica in Southern Poland

Ann. Bot. Fennici 45: 186–194 ISSN 0003-3847 (print) ISSN 1797-2442 (online) Helsinki 27 June 2008 © Finnish Zoological and Botanical Publishing Board 2008 Low genetic variation in subpopulations of an endangered clonal plant Iris sibirica in southern Poland Kinga Kostrakiewicz1,* & Ada Wróblewska2 1) Department of Plant Ecology, Institute of Botany, Jagiellonian University, Lubicz 46, PL-31-512 Kraków, Poland (corresponding author’s e-mail: [email protected]) 2) Institute of Biology, University of Białystok, Świerkowa 20B, PL-15-950 Białystok, Poland Received 2 May 2007, revised version received 5 June 2007, accepted 31 July 2007 Kostrakiewicz, K. & Wróblewska, A. 2008: Low genetic variation in subpopulations of an endangered clonal plant Iris sibirica in southern Poland. — Ann. Bot. Fennici 45: 186–194. The spatial genetic structure in three subpopulations of the endangered clonal plant Iris sibirica from southern Poland was investigated. The subpopulations occurred in different habitats, i.e. in a Molinietum caeruleae community, a Phragmites australis patch and in a willow brushwood. Using 13 enzymatic systems, sixteen loci were evaluated. The very low genetic diversity (P = 0%–18.7%, A = 1.0%–1.19%, HO = 0.000–0.009) observed within the subpopulations is probably due to lack of recruitment, habitat fragmentation and/or historical causes. Five distinct multilocus genotypes, detected from 148 collected samples in the subpopulations, supported this observation. This fact illustrated that only clonal growth could maintain the present low genetic variation through the domination of a single or a few clones within these sites. Moderate genetic differentiation (FST = 0.077, P < 0.001) that varies strongly between pairs of subpopulations, was observed, thereby suggesting substantial gene flow between populations. Key words: clonal growth, conservation, genotypic variation, Iridaceae, population subdivision Introduction well. Recent reviews of plant allozyme litera- ture have revealed the relationships between Knowledge of spatial genetic structure is of enor- life history traits and the patterns of genetic mous significance when concerning rare or endan- and genotypic variability (Loveless & Hamrick gered species. In these cases, determination of the 1984, Ellastrand & Roose 1987, Hamrick & genetic resources in the study group is essential Godt 1989, Hamrick et al. 1991). Since the early from the viewpoint of their conservation, both on 1960s, the usage of molecular markers (alloz- a local scale and throughout their whole geograph- ymes and DNA), particularly in the population ical range. Undoubtedly, the evolutionary poten- genetics of clonal plants has allowed not only to tial of a species depends upon the level of genetic assess the level of the population genetic vari- variability retained in its natural populations. ability, but also to determine the affiliations of The persistence of such populations depends ramets to specific genotypes (McLellan et al. on their genetic and genotypic variability as 1997, Brzosko et al. 2002). Molecular markers ANN. BOT. FENNICI Vol. 45 • Low genetic variation in subpopulations of Iris sibirica 187 have then provided answers to previously open Therefore, the aims of our investigations were: questions in the field of demography, concern- (1) to assess the intensity of generative and ing especially endangered or vulnerable clonal vegetative reproduction, (2) to determine the plants (Krenova & Lĕps 1996, Ayres & Ryan level of genetic and genotypic variability in the 1999, Fischer et al. 2000, Hannan & Orick 2000, subpopulations, and (3) to describe the effect of Xie et al. 2001, Brzosko et al. 2002, Brzosko & life history traits and habitats on the genetic and Wróblewska 2003). genotypic diversity among the subpopulations. One of the rare and strictly protected clonal plants in Poland is Iris sibirica. It is listed as a species of the Euro-Siberian subelement, whose Material and methods distribution ranges from central Europe to central Asia (Hultén & Fries 1986). The individuals Study sites and characteristics of of I. sibirica consist of numerous leaf rosettes subpopulations and flowering stems connected by permanent rhizomes with short internodes. The species is The field studies were carried out at the Kostrze characterised by an iterative type of growth site, situated in the western part of Kraków, resulting from the processes of senility, regener- southern Poland (Fig. 1 insert). The patches of ation and outgrowth, which proceed with differ- Molinietum caeruleae community are all that ent intensities in subsequent phases of develop- remain of the vast meadows, which once occu- ment (Harper 1977). Regeneration occurs at the pied an area along the Vistula river (Zarzycki margin of the clone, whereas the senility proc- 1956, 1958). The changes in water regime and esses at its centre lead to a gradual expansion the cease of grass cutting has enabled shrub of an ‘empty centre’ or ‘disintegration zone’. As vegetation to colonise the area and has after a result of advanced senility processes (dying), several decades led to a transformation of the the divisions occur among the oldest sections of herb layer (Dubiel 1991, 1996, Kostrakiewicz the rhizomes and the groups of their independent 2001). At present, the vast patches of Molinietum fragments emerge in the form of a single indi- caeruleae display partial or even significant dis- vidual in the genetic sense (Klimeš et al. 1997 turbance (Dubiel 2005). and http://www.butbn.cas.cz/klimes). Three neighbouring subpopulations of I. Iris sibirica also reproduces sexually. While sibirica were selected for this study (Fig. 1). seedlings rarely appear, they usually occur in Spaced some 200 m from one another, they the vicinity of the parental plants (Kostrakiewicz occurred in distinctly different habitats. The first 2007). The I. sibirica populations occur in a one, numbering 52 ramet clusters, occurred in a range of various communities from tall-herb to patch where Phragmites australis was the domi- shrub and deciduous woods or even coniferous nant species (side code PH). The second subpop- forests communities. However, populations of ulation, formed by 174 ramet clusters, occurred this species are also found in the community of in a willow brushwood (Salix rosmarinifolia and Molinietum caeruleae, characterised by the pres- S. cinerea), with a major proportion of sedges ence of other rare and protected species, such (e.g. Carex gracilis and C. nigra, side code SX). as Dianthus superbus, Gentiana pneumonanthe, The third subpopulation of 506 ramet clusters Gladiolus imbricatus, Trollius europaeus and was situated in a patch of Molinietum caeruleae, Orchis latifolia (Kostrakiewicz 2004). Unfortu- characterised by a considerable species diversity nately, the communities of this type are disap- (site code MO). In 2006, a single permanent plot pearing, and are listed by The World Conserva- was established in each subpopulation for the tion Union (IUCN) among the most endangered analyses of life history traits and spatial genetic ones in Europe (Denisiuk 1991). structure. The distribution of all ramet clusters We investigated genetic and genotypic diver- was determined with 10 of these being marked sity among I. sibirica subpopulations in the on a cartogram. The distance between the neigh- context of the above-mentioned problems in bouring ramet clusters was at least 2 m. In each three distinct communities in southern Poland. subpopulation, the average number of ramets in 188 Kostrakiewicz & Wróblewska • ANN. BOT. FENNICI Vol. 45 Fig. 1. Locations of the Iris sibirica subpopulations analysed in this study. a cluster was determined, as well as the aver- superoxide dismutase (Sod) and triose-phosphate age proportion of fruits sets in a cluster, and the isomerase (Tpi). The other allozymes such as number of juveniles. The maximum length and malic enzyme (Me), 6-phosphogluconate dehy- width of the disintegration zone was also meas- drogenase (6Pgd), phosphoglucomutase (Pgm), ured and the differences in the size of the disinte- and phosphoglucose isomerase (Pgi) were gration zones were tested using ANOVA (Tukey screened using cellulose acetate plates (Titan test). From each group of ramets in a cluster evi- III Zip Zone, Helena Laboratories). Enzyme dently growing from the same rhizome fragment, staining with minor modifications followed the a single leaf sample was collected for genetic protocols developed by Richardson et al. (1986), analyses. In total, 148 samples were obtained, Soltis and Soltis (1989), and Szweykowski and including 62 samples from the patch with a Odrzykowski (1990). predominance of Phragmites australis (PX), 34 samples from willow brushwood (SX), and 52 samples from the patch of Molinietum caeruleae Genetic and genotypic variation (MO) (Table 1). The proportion of polymorphic loci (P), the mean number of alleles per locus (A), the mean Allozyme analysis observed (HO) and expected (HE) heterozygos- ity were calculated using TFPGA (Miller 1997). For each ramet, a 3-cm-long leaf fragment was Deviations from the Hardy-Weinberg equilib- extracted in a 30 µl of PVP buffer (Mitton et al. rium (HWE) were tested using an exact test 1979). The majority of enzyme systems were of HWE with a Markov chain algorithm with separated on 11% horizontal starch gels. A lith- GENEPOP (Raymond & Rousset 1995). F-sta- ium-borate/Tris-citrate buffer system at pH 8.3 tistics according to Weir and Cockerham (1984) was used to separate aconitase

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