Can Soil Seed Banks Serve as Genetic Memory? A Study of Three Species with Contrasting Life History Strategies

We attempted to confirm that seed banks can be viewed as an important genetic reservoir by testing the hypothesis that standing (aboveground) plants represent a nonrandom sample of the seed bank. We sampled multilocus allozyme genotypes from three species with different life history strategies: Amaranthus retroflexus, Carduus acanthoides, Pastinaca sativa. In four populations of each species we analysed the extent to which allele and genotype frequencies vary in consecutive life history stages including the summer seed bank, which has been overlooked up to now. We compared the winter seed bank (i.e., seeds collected before the spring germination peak), seedlings, rosettes, the summer seed bank (i.e., seeds collected after the spring germination peak) and fruiting plants. We found that: (1) All three species partitioned most of their genetic diversity within life history stages and less among stages within populations and among populations. (2) All genetic diversity parameters, except for allele frequencies, were similar among all life history stages across all populations in different species. (3) There were differences in allele frequencies among life history stages at all localities in Amaranthus retroflexus and at three localities in both Carduus acanthoides and Pastinaca sativa. (4) Allele frequencies did not differ between the winter and summer seed bank in most Carduus acanthoides and Pastinaca sativa populations, but there was a marked difference in Amaranthus retroflexus. In conclusion, we have shown that the summer seed bank is not genetically depleted by spring germination and that a majority of genetic diversity remains in the soil through summer. We suggest that seed banks in the species investigated play an important role by maintaining genetic diversity sufficient for recovery rather than by accumulating new genetic diversity at each locality.     

Restoration of genetic diversity from soil seed banks in a threatened aquatic plant, <Emphasis Type="Italic">Nymphoides peltata</Emphasis>

Populations of a threatened aquatic plant, Nymphoides peltata, have rapidly degenerated under the influence of recent artificial changes in Lake Kasumigaura of Japan. To estimate the potential of soil seed banks for genetic restoration of the species, we used 10 microsatellite markers to analyze the genetic variation in adults and in seedlings that emerged from soil seed banks. About 187 leaf samples from the cultured stocks that were collected in 17 adult subpopulations in 1995 and 2000 and from three subpopulations that were newly discovered in 2002 were analyzed. As a result, only 18 genets could be identified, suggesting that clonal diversity of the adult population had already become extremely low. Genetic tests were performed on 430 seedlings from seed banks at six locations of natural lakeshores and three of the restoration sites that were artificially constructed in an attempt to assign them to the remnant adult population; many of the seedlings showed genetic variation different from the adults. Furthermore, the seedlings preserved seven alleles that had been lost from remnant adults. However, they had lower average numbers of alleles and heterozygosity levels (NA = 1.5–3.1, H _E = 0.146–0.487) than the remnant adults (NA = 3.5, H _E = 0.539) and showed high inbreeding coefficients, suggesting that the seed banks were produced by inbreeding. Thus, although the seed banks had a certain potential to restore genetic diversity, the fitness reduction in seed banks caused by inbreeding could affect the success of restoration based on seed banks.     

Diversity of Endophytic Fungal Community in Leaves of Artemisia argyi Based on High-throughput Amplicon Sequencing

To investigate the community structure and diversity of endophytic fungi in the leaves of Artemisia argyi, leaf samples were collected from five A. argyi varieties grown in different cultivation areas in China, namely, Tangyin Beiai in Henan (BA), Qichun Qiai in Hubei (QA), Wanai in Nanyang in Henan (WA), Haiai in Ningbo in Zhejiang (HA), and Anguo Qiai in Anguo in Hebei (AQA), and analyzed using Illumina high-throughput sequencing technology. A total of 365,919 pairs of reads were obtained, and the number of operational taxonomic units for each sample was between 165 and 285. The alpha diversity of the QA and BA samples was higher, and a total of two phyla, eight classes, 12 orders, 15 families, and 16 genera were detected. At the genus level, significant differences were noted in the dominant genera among the samples, with three genera being shared in all the samples. The dominant genus in QA was Erythrobasidium, while that in AQA, HA, and BA was Sporobolomyces, and that in WA was Alternaria, reaching a proportion of 16.50%. These results showed that the fungal community structure and diversity in QA and BA were high. The endophytes are of great importance to the plants, especially for protection, phytohormone and other phytochemical production, and nutrition. Therefore, this study may be significant with the industrial perspective of Artemisia species.     

The potential of soil seed banks of a eucalypt wetland forest to aid restoration

Soil seed banks can play an important role in the regeneration of wetland vegetation. However, their potential role in the restoration of degraded wetland forests is less certain. I surveyed the soil seed bank and extant floras of four sites across a eucalypt wetland forest of variable vegetation condition. At each site, the extant vegetation was surveyed within two 5 × 5 m² quadrats, each from which five composite soil seed bank samples were collected. Across the four sites, 57 (including 18 exotic) species were identified in the extant vegetation, while from the seed bank samples 6379 seedlings emerged from 80 taxa, 33 of which were exotic species. The soil seed bank was dominated by native and exotic monocots, and contained very few seeds of wetland tree or shrub species. Overall, the similarity between the extant and seed bank floras was very low (~24 %). Soil seed banks are likely to be of limited use in the restoration of degraded wetland forests, because the dominant species in such systems—woody and clonal plants—are typically absent from the soil seed bank. Wetland soil seed banks may contribute to the maintenance and diversity of understorey vegetation, however, they may also act as a source of exotic plant invasions, particularly when a wetland is degraded.     

Genetic characterization of cassava (Manihot esculenta) landraces in Brazil assessed with simple sequence repeats

Based on nine microsatellite loci, the aim of this study was to appraise the genetic diversity of 42 cassava (Manihot esculenta) landraces from selected regions in Brazil, and examine how this variety is distributed according to origin in several municipalities in the states of Minas Gerais, São Paulo, Mato Grosso do Sul, Amazonas and Mato Grosso. High diversity values were found among the five above-mentioned regions, with 3.3 alleles per locus on an average, a high percentage of polymorphic loci varying from 88.8% to 100%, an average of 0.265 for observed heterozygosity and 0.570 for gene diversity. Most genetic diversity was concentrated within the regions themselves (H_S = 0.52). Cluster analysis and principal component based scatter plotting showed greater similarity among landraces from São Paulo, Mato Grosso do Sul and Amazonas, whereas those from Minas Gerais were clustered into a sub-group within this group. The plants from Mato Grosso, mostly collected in the municipality of General Carneiro, provided the highest differentiation. The migration of human populations is one among the possible reasons for this closer resemblance or greater disparity among plants from the various regions.     

Genetic Structure,Diversity and Long Term Viability of a Medicinal Plant,Nothapodytes nimmoniana Graham. (Icacinaceae), in Protected and Non-Protected Areas in the Western Ghats Biodiversity Hotspot

Background and Question

The harvesting of medicinal plants from wild sources is escalating in many parts of the world, compromising the long-term survival of natural populations of medicinally important plants and sustainability of sources of raw material to meet pharmaceutical industry needs. Although protected areas are considered to play a central role in conservation of plant genetic resources, the effectiveness of protected areas for maintaining medicinal plant populations subject to intense harvesting pressure remain largely unknown. We conducted genetic and demographic studies of Nothapodytes nimmoniana Graham, one of the extensively harvested medicinal plant species in the Western Ghats biodiversity hotspot, India to assess the effectiveness of protected areas in long-term maintenance of economically important plant species.

Methodology/Principal Findings

The analysis of adults and seedlings of N. nimmoniana in four protected and four non-protected areas using 7 nuclear microsatellite loci revealed that populations that are distributed within protected areas are subject to lower levels of harvesting and maintain higher genetic diversity (H_e = 0.816, H_o = 0.607, A = 18.857) than populations in adjoining non-protected areas (H_e = 0.781, H_o = 0.511, A = 15.571). Furthermore, seedlings in protected areas had significantly higher observed heterozygosity (H_o = 0.630) and private alleles as compared to seedlings in adjoining non-protected areas (H_o = 0.426). Most populations revealed signatures of recent genetic bottleneck. The prediction of long-term maintenance of genetic diversity using BOTTLESIM indicated that current population sizes of the species are not sufficient to maintain 90% of present genetic diversity for next 100 years.

Conclusions/Significance

Overall, these results highlight the need for establishing more protected areas encompassing a large number of adult plants in the Western Ghats to conserve genetic diversity of economically and medicinally important plant species.     

Genetic diversity through life history of Dioon edule Lindley (Zamiaceae, Cycadales)

The distribution of genetic diversity and structure for three populations of Dioon edule Lindley (Zamiaceae) at Monte Oscuro (MO), El Farallón (EF) and Rancho del Niño (RN) in Veracruz, Mexico was studied using 20 allozyme loci, considering four life history classes: seeds, seedlings, juveniles and adults. The MO population is genetically less diverse than the EF and RN populations. Total and local inbreeding differ significantly between life history classes. An increment of inbreeding among all classes was observed, and genetic differentiation among populations was higher in seeds and seedlings than in juveniles and adults. In terms of percentage of polymorphic loci, the MO seeds showed least (80%), followed by RN (95%) and EF had the highest values (100%), probably because of a reduction in effective population size and habitat fragmentation processes. In this context, the mean effective population size was 23.2 ± 11.3 for all populations. We conclude that seed cohorts in EF and RN represent a reservoir of genetic diversity within these two populations. Also, preservation of adult plants is an essential aspect to consider in management and conservation efforts for populations of Dioon edule in natural conditions.     

Diversity and Species Relationship in Pearl Millet (Pennisetum typhoides) and Related Species

Thirteen accessions of pearl millet (Pennisetum typhoides (L) Leeke) collected from different states of India and eight wild species of the genus Pennisetum across the world were analyzed for genetic diversity using AFLP markers. A combined analysis of eight primer combinations showed 35% polymorphism among P. typhoides accessions while analysis with five primer combinations showed 99% polymorphism among the wild species. The dendrogram constructed for the P. typhoides accessions based on the UPGMA method revealed two major clusters with samples from Gujarat forming a separate cluster from the rest of the samples. Principal component analysis of the same data set revealed similar results with the first principal component accounting for 65% of the total variation. The percentage of rare and common alleles contributing to the diversity in the sample was analyzed using the Shannon Weiner diversity index. The SW index revealed that the samples from Gujarat contributed significantly to the overall diversity among the accessions. Among accessions of each geographical region, considerable variation was revealed by SW index with samples from Tamil Nadu being most polymorphic. The genetic diversity in the accessions could be utilized for future breeding work. The dendrogram constructed for the wild species revealed the extent of genetic diversity among them. Analysis with one primer combination showed P. typhoides being closer to P. mollissimum than to the other analyzed species.     

Taxonomic Status,Phylogenetic Affinities and Genetic Diversity of a Presumed Extinct Genus,Paraisometrum W.T. Wang (Gesneriaceae) from the Karst Regions of Southwest China

Background

The karst regions in South China have an abundance of endemic plants that face high extinction risks. The Chinese Gesneriaceae endemic Paraisometrum mileense ( = Oreocharis mileensis), was presumed extinct for 100 years. After its re-discovery, the species has become one of five key plants selected by the Chinese forestry government to establish a new conservation category for plants with extremely small populations. For conservation purposes, we studied the phylogenetic and population genetic status of P. mileense at the three only known localities in Guangxi, Guizhou and Yunnan.

Methodology/Principal Findings

We collected 64 samples (52 species) of Oreocharis and 8 samples from three provinces of P. mileense and generated molecular phylogenies, and inferred that P. mileense represents a relatively isolated and derived taxonomic unit within Oreocharis. Phylogeographic results of 104 samples of 12 populations of P. mileense indicated that the populations in Yunnan have derived from those in Guangxi and Guizhou. Based on AFLP data, the populations were found to harbor low levels of genetic diversity (He = 0.118), with no apparent gradient across the species’ range, a restricted gene flow and significant isolation-by-distance with limited genetic differentiation among the populations across the three provinces (F _ST = 0.207, P<0.001). The 10 populations in Yunnan were found to represent two distinct lineages residing at different altitudes and distances from villages.

Conclusion/Significance

The low levels of genetic diversity found in P. mileense are perhaps a consequence of severe bottlenecks in the recent past. The distribution of the genetic diversity suggests that all populations are significant for conservation. Current in situ and ex situ measures are discussed. Further conservation actions are apparently needed to fully safeguard this conservation flagship species. Our work provides a model of an integrated study for the numerous endemic species in the karst regions with extremely small populations.     

Genetic differentiation and diversity analysis of medicinal tree Syzygium cumini (Myrtaceae) from ecologically different regions of India

This study represents the agro-ecological zone wise surveys of molecular variation of important medicinal tree Syzygium cumini Linn. (Jamun) which is native to India. It is used world wide in treatment of diabetes. Despite of its diverse medicinal properties no molecular data is available about the pattern of variation in its natural range. Populations of S. cumini in India are located in different habitats which differ from each other with regard to ecological factors. In this study, random amplified polymorphic DNA (RAPD) markers were used to detect inter and intra levels of genetic variations of sixteen S. cumini genotypes collected from three major agro-ecological zones of India. A total of 220 amplification products were scored of which 87.50 % were polymorphic. The level of polymorphism ranged from 47.69 % to 74.87 % polymorphic bands per population and was correlated with population size. Different measures of diversity: Shannon’s index of phenotypic diversity (I) = 0.451 ± 0.230; Nei’s genetic diversity (h) = 0.300 ± 0.172; effective number of alleles per locus (Ne) = 1.51 ± 0.347; total species diversity (Hsp) = 0.315 ± 0.031 and within population diversity (Hpop) = 0.158 ± 0.104 showed high genetic diversity at species level. Coefficient of genetic differentiation (Gst =0.498; Nm = 0.503) revealed significant genetic differentiation among the populations. Most of the genetic variations are contained among the populations. The results of cluster analysis and principal component analysis (PCA) give only little evidence for an ecotypic differentiation of S. cumini populations. Present genetic structure of population suggests ex situ conservation in seed banks in which seeds from at least five populations need to collected and conserved. Secondly, our study provides practical information to herbal drugs manufactures who use Jamun as a raw material.     

Genetic diversity and structure among natural populations of Sindora glabra in Hainan Island,China as revealed by ISSR markers

Sindora glabra, one of the second-class national protective plants in China, has important ecological and commercial values. To understand the genetic diversity and structure of S. glabra, eleven natural populations from four areas of Hainan Island in China were analyzed by inter-simple sequence repeat (ISSR) markers. Thirteen primers were screened out and used to amplify 157 DNA samples. A total of 122 bands were obtained, among which 114 (93.4%) bands were polymorphic. Genetic parameters including average number of effective alleles (1.547), Nei's gene diversity (0.321), Shannon's information index (0.482), and gene differentiation coefficient (0.1944) revealed a high level of genetic diversity maintained in S. glabra populations. The variation within populations accounted for 85.6% of total variation based on analysis of molecular variance. Genetic distance analysis showed that 11 populations could be divided into three groups and populations from the same areas were classified as one group, suggesting that high genetic diversity of S. glabra was attributable to geographic isolation. Together, introduction of germplasm from distant natural distribution areas would be a sound strategy for germplasm resource conservation of S. glabra and selection of elite individuals from populations of far relationship for hybridization is of great importance for breeding improvement programs in future.     

Mitochondrial DNA Diversity of Honey Bees (Apis mellifera) from Unmanaged Colonies and Swarms in the United States

To study the genetic diversity of honey bees (Apis mellifera L.) from unmanaged colonies in the United States, we sequenced a portion of the mitochondrial DNA COI–COII region. From the 530 to 1,230 bp amplicon, we observed 23 haplotypes from 247 samples collected from 12 states, representing three of the four A. mellifera lineages known to have been imported into the United States (C, M, and O). Six of the 13 C lineage haplotypes were not found in previous queen breeder studies in the United States. The O lineage accounted for 9% of unmanaged colonies which have not yet been reported in queen breeder studies. The M lineage accounted for a larger portion of unmanaged samples (7%) than queen breeder samples (3%). Based on our mitochondrial DNA data, the genetic diversity of unmanaged honey bees in the United States differs significantly from that of queen breeder populations (p < 0.00001). The detection of genetically distinct maternal lineages of unmanaged honey bees suggests that these haplotypes may have existed outside the managed honey bee population for a long period.     

High genetic variation among closely related red oak (<Emphasis Type="Italic">Quercus rubra</Emphasis>) populations in an ecosystem under metal stress: analysis of gene regulation

Metal toxicity is a major abiotic stressor of plants. It has been established that changes in genetic variation occur very rapidly in plants in response to environmental stressors such as increased levels of metals. Quercus rubra (red oak) is a pioneer species in mining regions contaminated with metals in Northern Ontario (Canada). The objectives of the study were to (1) determine the level of genetic variation in Q. rubra populations from mining damaged ecosystems using RAPD marker system and (2) assess the level of gene expression of candidate genes for nickel resistance. Total gene diversity (H_T) and the mean gene diversity among populations (H_S) were 0.22 and 0.19, respectively. The percent of polymorphic loci within populations was high ranging from 61 % (Capreol) to 72 % (Daisy Lake) despite a high level of gene flow (2.4). The population differentiation (G_ST) value was low (0.17). No significant difference was found among the contaminated and reference sites for all the genetic parameters estimated. Hence, all the Q. rubra populations from the metal-contaminated and damaged ecosystems are genetically sustainable. Moreover, this study reveals that all populations were genetically closely related with genetic distance values varying from 0.17 to 0.35. A zinc finger protein of Arabidopsis thaliana (ZAT11) gene involved in nickel resistance was differentially expressed in samples analyzed. There was a 120 times higher of ZAT11 expression in samples from metal contaminated areas of Wahnapitae Dam compared to other metal contaminated and uncontaminated sites, but no association between soil metal levels and expression of ZAT11 was established.     

Plant genetic diversity and grazing management on the Qinghai-Tibetan Plateau: A case study of a dominant native wheatgrass (Elymus nutans)

Grazing management on the Qinghai-Tibetan Plateau in China concerns farmers, scientists and policy makers because of its geographical uniqueness. To investigate the effects of grazing on genetic diversity in native plants, the dominant perennial wheatgrass Elymus nutans was used as a model to examine genetic diversity and spatial population structure under different grazing intensities. Intra-population genetic diversity in three grazed areas was higher than un-grazed area; however, the genetic diversity decreased with increasing grazing intensity. Genotypes from four areas were separated into two groups (grazed and un-grazed) through a clustering analysis. Approximately 12% (G_st = 0.1487) of the DNA variation was apportioned among populations by AMOVA indicating a relatively low degree of inter-population genetic differentiation. Significant spatial auto-correlations and subpopulation structure were also observed within the four areas, and the genetic ‘patch’ size decreased with increasing grazing intensity. On the high and cold Qinghai-Tibetan Plateau, grazing is suitable for the maintenance of genetic diversity, and a light grazing intensity is optimal for the conservation of perennial grass germplasm and long-term grassland management.     

Genetic Diversity and Distribution of the Ciguatera-Causing Dinoflagellate Gambierdiscus spp. (Dinophyceae) in Coastal Areas of Japan

Background

The marine epiphytic dinoflagellate genus Gambierdiscus produce toxins that cause ciguatera fish poisoning (CFP): one of the most significant seafood-borne illnesses associated with fish consumption worldwide. So far, occurrences of CFP incidents in Japan have been mainly reported in subtropical areas. A previous phylogeographic study of Japanese Gambierdiscus revealed the existence of two distinct phylotypes: Gambierdiscus sp. type 1 from subtropical and Gambierdiscus sp. type 2 from temperate areas. However, details of the genetic diversity and distribution for Japanese Gambierdiscus are still unclear, because a comprehensive investigation has not been conducted yet.

Methods/Principal Finding

A total of 248 strains were examined from samples mainly collected from western and southern coastal areas of Japan during 2006–2011. The SSU rDNA, the LSU rDNA D8–D10 and the ITS region were selected as genetic markers and phylogenetic analyses were conducted. The genetic diversity of Japanese Gambierdiscus was high since five species/phylotypes were detected: including two reported phylotypes (Gambierdiscus sp. type 1 and Gambierdiscus sp. type 2), two species of Gambierdiscus (G. australes and G. cf. yasumotoi) and a hitherto unreported phylotype Gambierdiscus sp. type 3. The distributions of type 3 and G. cf. yasumotoi were restricted to the temperate and the subtropical area, respectively. On the other hand, type 1, type 2 and G. australes occurred from the subtropical to the temperate area, with a tendency that type 1 and G. australes were dominant in the subtropical area, whereas type 2 was dominant in the temperate area. By using mouse bioassay, type 1, type 3 and G. australes exhibited mouse toxicities.

Conclusions/Significance

This study revealed a surprising diversity of Japanese Gambierdiscus and the distribution of five species/phylotypes displayed clear geographical patterns in Japanese coastal areas. The SSU rDNA and the LSU rDNA D8–D10 as genetic markers are recommended for further use.     

Micro-geographic landscape features demarcate seedling genetic structure in the stream lily,Helmholtzia glaberrima

In this study, 169 stream lily (Helmholtzia glaberrima) seedlings from six micro-drainages were genotyped with AFLP markers to quantify the impact that topographic landscape features and altitude may have in shaping patterns of genetic diversity within individual populations. A global analysis of genetic diversity detected significant genetic differentiation among micro-drainages (F_ST = 0.22, P < 0.01). The observed genetic structure of sampled sites conformed to a hierarchical model of gene flow. Assignment tests also supported a hierarchical model of gene flow as only one dispersal event among the sampled micro-drainages was detected. This suggests that opportunities for seed dispersal in H. glaberrima are highly constrained by patterns of hydrographic networks even at a local scale. In contrast, altitude had little impact on partitioning of genetic diversity as no increase in genetic diversity was evident among individuals in the upper (0.18 ± 0.02), and lower (0.17 ± 0.02) areas of micro-drainages. Overall these results suggest that the influence of freshwater landscape features can vary widely the effect on the patterns of genetic diversity of seedlings in stream lily populations.     

Genetic diversity of wild and cultured swamp eel (Monopterus albus) populations from central China revealed by ISSR markers

Asian swamp eel is a highly commercial fish, primarily for China and other Asian countries. The aim of this paper is to evaluate the genetic diversity of wild and cultured samples of Asian swamp eel Monopterus albus using ISSR markers. A total of 129 individuals belonging to three wild samples, Xiantao (XT), Huanggang (HG), Xinyang (XY) and three cultured samples, Wuhan (WH), Jingzhou (JZ) and Nanjing (NJ) were randomly selected for genetic analysis. Twelve ISSR primers were used for screening the six populations and 110 loci were obtained. The polymorphic loci were estimated to be 54%, 56.3%, 58.2%, 60.6%, 69.5% and 71% in NJ,WH, JZ, XT, HG and XY samples, respectively. Average heterozygosity value varied from 0.1956 to 0.2449. The three wild samples showed higher genetic diversity than the cultured samples (P < 0.05), including polymorphic bands (PPB), observed number of alleles per locus (_to), effective number of alleles per locus (Ne), Nei’s gene diversity index (H) and Shannon’s information index (I).     

Assessing the diversity of arbuscular mycorrhizal fungi in semiarid shrublands dominated by Artemisia tridentata ssp. wyomingensis

Variation in the abiotic environment and host plant preferences can affect the composition of arbuscular mycorrhizal (AMF) assemblages. This study analyzed the AMF taxa present in soil and seedlings of Artemisia tridentata ssp. wyomingensis collected from sagebrush steppe communities in southwestern Idaho, USA. Our aims were to determine the AMF diversity within and among these communities and the extent to which preferential AMF–plant associations develop during seedling establishment. Mycorrhizae were identified using molecular methods following DNA extraction from field and pot culture samples. The extracted DNA was amplified using Glomeromycota specific primers, and identification of AMF was based on phylogenetic analysis of sequences from the large subunit-D2 rDNA region. The phylogenetic analyses revealed seven phylotypes, two within the Claroideoglomeraceae and five within the Glomeraceae. Four phylotypes clustered with known species including Claroideoglomus claroideum, Rhizophagus irregularis, Glomus microaggregatum, and Funneliformis mosseae. The other three phylotypes were similar to several published sequences not included in the phylogenetic analysis, but all of these were from uncultured and unnamed glomeromycetes. Pairwise distance analysis revealed some phylotypes with high genetic variation. The most diverse was the phylotype that included R. irregularis, which contained sequences showing pairwise differences up to 12 %. Most of the diversity in AMF sequences occurred within sites. The smaller genetic differentiation detected among sites was correlated with differences in soil texture. In addition, multiplication in pot cultures led to differentiation of AMF communities. Comparison of sequences obtained from the soil with those from A. tridentata roots revealed no significant differences between the AMF present in these samples. Overall, the sites sampled were dominated by cosmopolitan AMF taxa, and young seedlings of A. tridentata ssp. wyomingensis were colonized in relation to the abundance of these taxa in the soil.     

Patterns of genetic and taxonomic differentiation in three Melitaea (subg. Mellicta) species (Lepidoptera, Nymphalidae, Nymphalinae)

In conservation genetics the existence of Evolutionarily Significant Units (ESU) is a crucial question in threatened or vulnerable species. It is of particular concern to determine whether different subspecies or ecotypes of a species can be considered as separate ESUs. Some Melitaea subg. Mellicta species (e.g. Mellicta aurelia, Mellicta britomartis) are declining or scarce in Europe. Therefore, the level of genetic differentiation and pattern of genetic variation were surveyed in three Melitaea (subg. Mellicta) species. Their habitat requirements and food plants partly overlap; accordingly they often co-occur in the same habitat. M. britomartis and M. aurelia have one brood per year in Hungary, while Mellicta athalia has a monovoltine and a bivoltine ecotype. The purpose of the study was to estimate the number of genetically differentiated ESUs among these species in the Carpathian basin. Samples were taken from 5 Hungarian regions and a few samples were collected in Transylvania as well. Enzyme polymorphism was studied using polyacrylamide gel electrophoresis. The structure of genetic variation was analysed by F-statistics, AMOVA, PCA and Bayesian clustering method. UPGMA dendrogram was constructed on the basis of Cavalli-Sforza and Edwards chord distances. The three species are clearly differentiated from each other in all statistical analyses. They are evidently different Evolutionary Significant Units. The two ecotypes of M. athalia, however, do not show any genetic differentiation.