Effects of population size on reproductive success of the endangered and endemic species Primula merrilliana

The reproductive ability related to the population size of the endangered and endemic species Primula merrilliana Schltr.was investigated.In 26 populations observed,only four contain more than 500 flowering individuals,whereas most of them(53.8%) consist of less than 100 flowering individuals.Though the ratio of pin and thrum plants keeps its balance at 1:1 for all populations,the frequency of pin and thrum flowers was significantly biased in most small populations.Population size strongly affected reproductive success; plants in small populations produced significantly fewer flowers and fruits per plant and fewer seeds per fruit,and therefore fewer seeds per plant.The floral density was another important factor that influenced the reproductive success of P.merrilliana,because four main reproductive success parameters (fruits per plant,seeds per fruit,seeds per plant,and the proportion of flowers setting fruit) were all positively correlated with floral density.The size of plants and the number of leaves per plant (measure of habitat quality) were not influenced by the variation of population size,suggesting that the reduced fecundity in small populations may not be a consequence of lower habitat quality,inbreeding depression and pollen limitation as a result of less attractiveness in small populations are therefore likely explanations for these patterns.     

Effects of habitat fragmentation on the reproductive success of the nectar-producing orchid Gymnadenia conopsea and the nectarless Orchis mascula

Habitat fragmentation decreases plant population sizes and increases spatial isolation, which hampers the exchange of seeds and pollen between fragmented populations. This may result in decreased population viability. We compared the effects of population size and isolation on the reproductive success of two orchid species, Gymnadenia conopsea (nectar-producing) and Orchis mascula (nectarless) growing in highly fragmented calcareous grassland in southern Belgium. We expected that the nectar-producing species would be more susceptible to the negative reproductive consequences of habitat fragmentation compared to the nectarless species. Nectar production has been associated with increased geitonogamous pollination and, therefore, with lower seed viability. Our results show that seed viability increased with increasing population size in O. mascula, whereas it was always low in G. conopsea, even in large populations. In contrast, percentage fruit set was positively related to population size in the nectar-producing G. conopsea, but no such effect was observed in the nectarless O. mascula, where fruit set was low even in large populations. Population isolation was not related to reproductive success for either species. Our results suggest that even in large populations, where pollinators are expected to be more abundant, increased geitonogamous pollination reduces seed viability in the nectar-producing G. conopsea. In contrast, seed viability in O. mascula seems to benefit from increased pollinator availability in larger populations. For the latter species, however, fruit set remains low, even in large populations, compared to G. conopsea. This may be explained by the relatively low attractiveness of nectarless orchid species for pollinators. Our results indicate that small population size may negatively influence reproductive success in both nectarless and nectar-producing orchids by reducing seed viability and fruit set, respectively.     

Sample size for collecting germplasms — a polyploid model with mixed mating system

The present paper discusses a general expression for determining the minimum sample size (plants) for a given number of seeds or vice versa for capturing multiple allelic diversity. The model considers sampling from a large 2 k-ploid population under a broad range of mating systems. Numerous expressions/results developed for germplasm collection/regeneration for diploid populations by earlier workers can be directly deduced from our general expression by assigning appropriate values of the corresponding parameters. A seed factor which influences the plant sample size has also been isolated to aid the collectors in selecting the appropriate combination of number of plants and seeds per plant. When genotypic multiplicity of seeds is taken into consideration, a sample size of even less than 172 plants can conserve diversity of 20 alleles from 50,000 polymorphic loci with a very large probability of conservation (0.9999) in most of the cases.     

TEMPORAL VARIATION OF FLOWER AND FRUIT SIZE IN RELATION TO SEED YIELD IN CELANDINE POPPY (CHELIDONIUM MAJUS; PAPAVERACEAE)

Ecological and evolutionary studies typically consider variation in single reproductive characters in isolation, without considering how they might be correlated with other reproductive and vegetative characters. In our study, we examined temporal patterns of variation and correlation in flower diameter and fruit length during a reproductive phase in two Massachusetts populations of the herb, Chelidonium majus. We also examined the relationships of such variation to measurements of seed yield components (mean seed weight and number per fruit) and aspects of plant vegetative size. Most of the variation in the sizes of reproductive characters occurred within individual plants, instead of among plants or between populations. Flower and fruit sizes as well as seed number per fruit declined significantly during the season in both populations. Only mean seed size per fruit was relatively stable for individual plants in both populations. Conserving resources by a gradual reduction in the size of reproductive characters over the season may be a strategy for maternal plants to continue seed production. The strong, persistent patterns of correlation between certain characters, such as flower and fruit size, in spite of extensive phenotypic plasticity, was interpreted as indirect evidence for developmental correlation. Furthermore, vegetatively larger plants produced not only more flowers and fruits, but also consistently larger flowers and fruits. The results emphasize that variation in fitness characters, such as seed size and number, should not be viewed in isolation from vegetative characters, flower, and fruit sizes in ecological and evolutionary studies, if the goal is to understand the mechanisms of natural selection in wild populations.     

Sexual reproduction and variation in floral morphology in an ephemeral vernal lily,Eyythronium americanum

Summary In a riparian population of Erythronium americanum (Liliaceae) in central New Jersey, experimentally self-pollinated plant produced markedly fewer fruit and fewer seeds per fruit than hand-outcrossed and open pollinated plants, even though differences were not evident between pollen tubes that penetrated stigmas from self or foreign pollen. This weak self-compatibility and a positive relation between the percentage of seeds set by outcrossed plants and the distance between pollen donor and recipient plants indicate that this population could be susceptible to inbreeding depression.Limited resources for seed development apparently constrained maximal seed production, based on low seed set (40.6%) by hand-pollinated plants and positive correlations for these plants between plant size and the number and size of seeds set. In contrast, naturally-pollinated plants set a smaller proportion of their ovules, suggesting that limited pollinator service reduced the quantity of seeds produced in this population. Free-foraging bees usually removed more than half of the available pollen in a single visit, so that individual plants probably have few opportunities to disseminate their pollen.Even though sexually reproductive ramets produce only a single flower per year, less than a third of variation in floral morphology is associated with variation in plant size. Within the flower, the sizes of some closely associated structures, such as the style and ovary, and the anthers and filaments, vary essentially independently of one another. Production of nectar and pollen, the ultimate attractors of pollinating insects, was positively correlated with flower size.     

Reproductive biology and genetic structure in Lloydia serotina

Concern regarding the conservation status of small, isolated populations of the arctic-alpine plant species Lloydia serotina prompted research to establish the status and performance of this species in Wales, in comparison with large populations in its more typical alpine habitat. Relationships between reproductive strategies and genetic variation were investigated in a number of populations, representing a wide habitat, geographic and population size range. In all populations, vegetative reproduction predominates over sexual reproduction, but seed produced is viable and germinates readily under controlled conditions. Smaller, peripheral populations produced fewer flowers and seeds than the larger ones, but all populations studied supported significant percentages (>30%) of male plants, due to either the occurrence of androdioecy in this species or to a resource limited breeding system. Analysis of allozyme variation in sixteen populations from North America, the European Alps and Wales showed lower levels of genetic variation in smaller populations which averaged 1.1–1.2 alleles per locus and 10–20% of loci polymorphic, whereas larger populations averaged 1.4 alleles per locus and 30–40% polymorphic loci. This applied especially to the most northerly and southerly populations in North America, suggesting the occurrence of genetic drift in these small, peripheral populations. F-statistics suggest relatively high levels of differentiation among smaller populations, even among those closely related geographically, but genetic variation has been retained in all but one population, possibly due to infrequent sexual reproduction by long lived clones. RAPD analysis of four small populations in Wales provided further evidence of clonal growth and possible inbreeding dominating a mixed mating reproductive system with consequent genetic structuring in these populations.     

The role of the tolerance–fecundity trade‐off in maintaining intraspecific seed trait variation in a widespread dimorphic herb

Coexistence of species with different seed sizes is a long‐standing issue in community ecology, and a trade‐off between fecundity and stress tolerance has been proposed to explain co‐occurrence in heterogeneous environments. Here we tested an intraspecific extension of this model: whether such trade‐off also explains seed trait variation among populations of widespread plants under stress gradients. We collected seeds from 14 populations of Plantago coronopus along the Atlantic coast in North Africa and Europe. This herb presents seed dimorphism, producing large basal seeds with a mucilaginous coat that facilitates water absorption (more stress tolerant), and small apical seeds without coats (less stress tolerant). We analysed variation among populations in number, size and mucilage production of basal and apical seeds, and searched for relationships between local environment and plant size. Populations under higher stress (higher temperature, lower precipitation, lower soil organic matter) had fewer seeds per fruit, higher predominance of basal relative to apical seeds, and larger basal seeds with thicker mucilaginous coats. These results strongly suggest a trade‐off between tolerance and fecundity at the fruit level underpins variation in seed traits among P. coronopus populations. However, seed production per plant showed the opposite pattern to seed production per fruit, and seemed related to plant size and other life‐cycle components, as an additional strategy to cope with environmental variation across the range. The tolerance–fecundity model may constitute, under stress gradients, a broader ecological framework to explain trait variation than the classical seed size–number compromise, although several fecundity levels and traits should be considered to understand the diverse strategies of widespread plants to maximise fitness in each set of local conditions.     

Comparison of quantitative genetic parameters between two natural populations of a selfing plant species, Medicago truncatula Gaertn.

 In this paper we compare mean values, heritability estimates, coefficient of genetic variation, and genetic correlations among several fitness components of two natural populations of a selfing plant species, Medicago truncatula L. It is shown that the population that had been found most polymorphic for molecular markers in a previous study was also the most variable for quantitative characters. Depending on the traits, the larger heritabilities in this population were due to either larger coefficients of genetic variances or smaller coefficients of environmental variances. Whereas genetic and phenotypic correlation matrices were very similar within each population, they were quite different between populations. In particular, although a positive correlation between age and size at maturity was found in both populations, the correlation between age at maturity and reproductive success was negative in the more variable population (late flowering plant, with a larger size at flowering, produced fewer pods), whereas no correlation was observed in the less variable population. We suggest that while in the less variable population all individuals have a high reproductive effort, several strategies coexist in the more variable population, with some early-flowering genotypes showing a high reproductive effort and other late-flowering genotypes showing a larger competitive ability through increased vegetative growth. Received: 25 June 1996 / Accepted: 11 October 1996     

海拔对青藏高原东部毛茛科植物繁殖特征和资源分配的影响

通过野外采样对青藏高原东部高寒草甸上两个海拔间3种常见毛茛科植物条裂银莲花(Anemone trullifo-liavar.linearis)、粗距翠雀花(Delphinium pachycentrumHemsl.)和钝裂银莲花(Anemone obtusiloba)的繁殖性状和资源分配进行了研究,并对植物在极端环境下采取的繁殖策略及繁殖模式进行了探讨.结果显示:(1)3个物种的株高都随着海拔的升高而降低;同一物种的花大小在不同海拔间均无显著差异;条裂银莲花单个种子重随海拔的升高而增加,但虫食数/株、结籽率和种子数/株均随着海拔的升高而减少;钝裂银莲花的繁殖分配、虫食数/株、种子数/株和单个种子重均随着海拔的升高而减小.(2)不同海拔条裂银莲花的个体大小与单花重、雄蕊重、虫食数/株均呈显著正相关,且回归斜率在海拔间有显著差异.粗距翠雀花花期的雄蕊重与个体大小呈显著正相关,但这种异速关系不受海拔和个体大小的影响;钝裂银莲花的个体大小与花期的所有繁殖特征以及果期的种子重/果实均呈显著正相关,不同海拔间个体大小与种子重/果实的斜率差异显著,且与花期各繁殖性状异速关系的截距差异显著.研究表明海拔对植物的个体大小以及种群间的繁殖对策和繁殖成功率有着重要的影响.     

Fruit production in Sorbus aucuparia L. (Rosaceae) and pre-dispersal seed predation by the apple fruit moth (Argyresthia conjugella Zell.)

Variation in fruit production and pre-dispersal seed predation by Argyresthia conjugella was studied in␣four populations of Sorbus aucuparia in northern Sweden.␣The number of infructescences, fruits per infructescence, consumed seeds and developed unattacked seeds per fruit were scored in marked trees from 1984 to 1990. The results showed that the number of fruits produced in each population determined the number of seed predators occurring in the host population, as the yearly number of seed predators was significantly and positively correlated with yearly number of fruits, in all but one population. The seed predators showed a delay in response to variation in number of fruits produced. This lag in response resulted in a large proportion of fruits being attacked and seeds consumed in a bad fruiting year that followed a good fruiting year, and vice versa. The proportion of fruits attacked and seeds consumed was largest in the population showing the greatest between-year variation in fruit production and lowest in the population showing the lowest between-year variation in fruit production. Furthermore, the individuals within the former population were synchronised, while they were not in the latter population. These results contradict one of the possible explanations of mast-seeding, where large synchronised between-year variation is supposed to reduce pre-dispersal seed predation. Instead, differences in attraction of the seed predator to differences in fruit crop size could explain the observed difference in seed predation between the two populations with opposite fruiting patterns. Within each population, irrespective of year, the proportion of fruits attacked and seeds consumed was independent of a tree's fruiting display. Therefore, trees with high fruit production, despite harbouring the largest number of seed predators, produced the largest number of developed seeds in absolute numbers, compared to trees that produced few fruits. Received: 25 February 1996 / Accepted: 30 November 1996     

GENETIC DIVERSITY AND POPULATION STRUCTURE IN CAMELLIA JAPONICA L. (THEACEAE)

Camellia japonica is a widespread and morphologically diverse tree native to parts of Japan and adjacent islands. Starch gel electrophoresis was used to score allelic variation at 20 loci in seeds collected from 60 populations distributed throughout the species range. In comparison with other plant species, the level of genetic diversity within C. japonica populations is very high: 66.2% of loci were polymorphic on average per population, with a mean number of 2.16 alleles per locus; the mean observed and panmictic heterozygosities were 0.230 and 0.265, respectively. Genotypic proportions at most loci in most populations fit Hardy-Weinberg expectations. However, small heterozygote deficiencies were commonly observed (mean population fixation index = 0.129). It is suggested that the most likely cause of the observed deficiencies is population subdivision into genetically divergent subpopulations. The overall level of population differentiation is greater than is typically observed in out-breeders: The mean genetic distance and identity (Nei's D and I) between pairs of populations were 0.073 and 0.930, respectively, and Wright's Fst was 0.144. Differences among populations appeared to be manifested as variation in gene frequencies at many loci rather than variation in allelic composition per se. However, the patterns of variation were not random. Reciprocal clinal variation of gene frequencies was observed for allele pairs at six loci. In addition, principal components analysis revealed that populations tended to genetically cluster into four regions representing the geographic areas Kyushu, Shikoku, western Honshu, and eastern Honshu. There was a significant relationship between genetic and geographic distance (r = 0.61; P < 0.01). Analysis of variance on allozyme frequencies showed that there was approximately four times as much differentiation among populations within regions, as among regions. It is likely that the observed patterns of population relationships result from the balance between genetic drift in small subpopulations and gene flow between them.     

Genetic variability, population size and reproduction potential in Ligularia sibirica (L.) populations in Estonia

Ligularia sibirica (L.) Cass. (Asteraceae) is a EU Habitats Directive Annex II plant species that has suffered a lot from human-caused major changes in quality and availability of habitats in Estonia. The aim of this study was to find out if the observed decline in population size is reflected in the amount of genetic variation and fertility in remnant populations of this species. AFLP technique was used for that purpose. Genetic diversity within populations was assessed as the percentage of polymorphic loci in a given population and average gene diversity over loci. The degree of genetic differentiation among populations and genetic differentiation between pairs of populations was estimated. The amount of viable seeds per flower stem was compared among populations and between years (2007 and 2008). Average genetic diversity over loci and proportion of polymorphic loci in L. sibirica populations were significantly correlated with population size, suggesting the action of genetic drift and/or inbreeding. No correlation was found between genetic and geographic distances. Natural barriers like forests may have been efficiently preventing seed migration even between geographically closer populations. Results of this study suggest that genetic erosion could be partially responsible for the lower fitness in smaller populations of this species.     

Vulnerability and determinants of reproductive success in the narrow endemic Antirrhinum microphyllum (Scrophulariaceae)

The breeding system and flowering phenology of the narrow endemic Antirrhinum microphyllum (Scrophulariaceae) were studied in order to assess the main factors affecting female reproductive success and to identify existing or potential threats to the viability of its populations. Hand-pollination experiments showed that A. microphyllum is an allogamous self-incompatible species. In both populations studied, the flowering season was 4 mo long and mean flowering duration per plant was about 1 mo. Peak flower production took place between mid-April and mid-May and overlapped with the period of activity of the main pollinator, Rhodanthidium sticticum (Megachilidae). Estimated mean number of seeds produced per plant was 9391, showing that population viability is not presently limited by seed output. The study of the direct and indirect effects of plant size, phenological traits (first flowering date, flowering duration, and flowering synchrony), and distance to neighbor plants on reproductive success was performed using structural equation modeling (SEM). In both populations, number of flowers and plant size were the main factors that determined the total number of fruits produced by a plant. First flowering date and flowering synchrony also affected fruit production. Multisample comparison of path coefficients for the two populations rejected the possibility that reproductive patterns could be described by one single model.     

Consequences of prairie fragmentation on the progeny sex ratio of a gynodioecious species, Lobelia spicata (Campanulaceae)

Habitat fragmentation of prairie ecosystems has resulted in increased isolation and decreased size of plant populations. In large populations, frequency-dependent selection is expected to maintain genetic diversity of sex determining factors associated with gynodioecy, that is, nuclear restorer genes that reverse cytoplasmic male sterility (nucleocytoplasmic gynodioecy). However, genetic drift will have a greater influence on small isolated populations that result from habitat fragmentation. The genetic model for nucleocytoplasmic gynodioecy implies that the proportion of female progeny produced by hermaphroditic and female plants will show more extreme differences in populations with reduced allelic diversity, and that restoration of male function will increase with inbreeding. We investigated potential impacts of effects resulting from reduced population sizes by comparison of progeny sex ratios produced by female and hermaphroditic plants in small and large populations of the gynodioecious prairie species, Lobelia spicata. A four-way contingency analysis of the impact of population size, population sex ratio, and maternal gender on progeny sex ratios showed that progeny sex ratios of hermaphroditic plants were strongly influenced by population size, whereas progeny sex ratios of female plants were strongly influenced by population sex ratio. Further, analysis of variation in progeny-type distribution indicated decreased restoration and increased loss of male function in smaller and isolated populations. These results are consistent with reduced allelic diversity or low allelic frequency at restorer loci in small and isolated populations. The consequent decrease in male function has the potential to impede seed production in these fragmented prairies.     

Genetic diversity of natural and cultivated populations ofOenanthe javanica in Korea

Enzyme electrophoresis was used to estimate genetic diversity and population structure in natural and cultivatedOenanthe javanica (Blume) DC. In the six natural populations, 8 of the 22 loci showed polymorphisms. Cultivated populations had fewer alleles per locus (1.84 vs. 1.91), fewer effective alleles per locus (1.47 vs. 1.52), a lower percentage of polymorphic loci (42.3 vs. 50.0), and lower diversity (0.210 vs. 0.228) than did natural populations. These parameters of genetic diversity indicate that the cultivated populations are genetically depauperate relative to their presumptive progenitor, and that the domestication process has partly eroded the level of genetic variation of this species. Nevertheless, the diversity of this species has higher-than-average values compared with other species having similar life-history traits. We propose that the mix-mating system; perennial, high gene flow; and large population sizes are possible factors contributing to this high diversity, which seemed to increase with distance from the coastlines.     

Genetic diversity and hierarchical population structure of a rare autotetraploid plant,Aster kantoensis (Asteraceae)

We sampled 17 populations of a rare autotetraploid Aster kantoensis (Asteraceae) from three river systems located in central Japan, and studied them for allelic variation at 22 enzyme loci. There was no significant correlation between the actual population size and three genetic diversity parameters, suggesting that the effective population size was very small even for the large populations, i.e., even large populations may still have a high probability of being of recent origin and remain influenced by the founder effect. Compared to other autotetraploid species, the total genetic variation of A. kantoensis is small. The number of alleles and gene diversity of a population were not significantly different among the river systems, although the percentage of polymorphic loci was different. Genetic differentiation among river systems was larger than between populations within the river systems, thereby indicating that gene flow between river systems is small, especially between the Kinu River system and Tama or Sagami River systems.     

Morphological variation and female reproductive success in two sympatric Trillium species: evidence for phenotypic selection in Trillium erectum and Trillium grandiflorum (Liliaceae)

I investigated the mating systems and phenotypic variation of two sympatric spring ephemerals, Trillium erectum and T. grandiflorum (Liliaceae), and phenotypic selection acting through female reproductive success for 11 morphological characters in five sympatric populations of the two species. I examined the degree of self-compatibility, pollinator-visitation rates, and pollen limitation of fruit and seed production in both species. Both Trillium species were self-compatible, but outcrossed flowers produced more successful fruits and seeds than self-pollinated flowers. Pollinator-visitation rates to the two species were low compared to other insect-pollinated spring ephemerals. In addition, both T. erectum and T. grandiflorum experienced pollen limitation in fruit and/or seed production; however, levels of fecundity in both species may be influenced by resource availability as well. I found significant phenotypic variation in 11 morphological characters within and among the five study populations. The sizes of all morphological characters were positively correlated. In general, larger T. erectum and T. grandiflorum produced more seeds. Phenotypic selection analysis revealed that direct and indirect selection acted on the size of morphological characters for both species. But there was no detectable selection acting on plant shape. This study reveals that variation in plant size exists within and among populations of both species, and this variation is associated with variance in female reproductive success. Spatial and temporal variation in pollinator and/or resource abundance may play a role in the phenotypic variation exhibited by both Trillium species.     

No genetic diversity at molecular markers and strong phenotypic plasticity in populations of Ranunculus nodiflorus, an endangered plant species in France

BACKGROUND AND AIMS: Although conservation biology has long focused on population dynamics and genetics, phenotypic plasticity is likely to play a significant role in population viability. Here, an investigation is made into the relative contribution of genetic diversity and phenotypic plasticity to the phenotypic variation in natural populations of Ranunculus nodiflorus, a rare annual plant inhabiting temporary puddles in the Fontainebleau forest (Paris region, France) and exhibiting metapopulation dynamics. METHODS: The genetic diversity and phenotypic plasticity of quantitative traits (morphological and fitness components) were measured in five populations, using a combination of field measurements, common garden experiments and genotyping at microsatellite loci. KEY RESULTS: It is shown that populations exhibit almost undetectable genetic diversity at molecular markers, and that the variation in quantitative traits observed among populations is due to a high level of phenotypic plasticity. Despite the lack of genetic diversity, the natural population of R. nodiflorus exhibits large population sizes and does not appear threatened by extinction; this may be attributable to large phenotypic plasticity, enabling the production of numerous seeds under a wide range of environmental conditions. CONCLUSIONS: Efficient conservation of the populations can only be based on habitat management, to favour the maintenance of microenvironmental variation and the resulting strong phenotypic plasticity. In contrast, classical actions aiming to improve genetic diversity are useless in the present case.     

How do reproductive skew and founder group size affect genetic diversity in reintroduced populations?

Reduced genetic diversity can result in short-term decreases in fitness and reduced adaptive potential, which may lead to an increased extinction risk. Therefore, maintaining genetic variation is important for the short- and long-term success of reintroduced populations. Here, we evaluate how founder group size and variance in male reproductive success influence the long-term maintenance of genetic diversity after reintroduction. We used microsatellite data to quantify the loss of heterozygosity and allelic diversity in the founder groups from three reintroductions of tuatara ( Sphenodon ), the sole living representatives of the reptilian order Rhynchocephalia. We then estimated the maintenance of genetic diversity over 400 years (∼10 generations) using population viability analyses. Reproduction of tuatara is highly skewed, with as few as 30% of males mating across years. Predicted losses of heterozygosity over 10 generations were low (1–14%), and populations founded with more animals retained a greater proportion of the heterozygosity and allelic diversity of their source populations and founder groups. Greater male reproductive skew led to greater predicted losses of genetic diversity over 10 generations, but only accelerated the loss of genetic diversity at small population size (<250 animals). A reduction in reproductive skew at low density may facilitate the maintenance of genetic diversity in small reintroduced populations. If reproductive skew is high and density-independent, larger founder groups could be released to achieve genetic goals for management.     

Microsatellite and major histocompatibility complex variation in an endangered rattlesnake,the Eastern Massasauga (Sistrurus catenatus)

Genetic diversity is fundamental to maintaining the long‐term viability of populations, yet reduced genetic variation is often associated with small, isolated populations. To examine the relationship between demography and genetic variation, variation at hypervariable loci (e.g., microsatellite DNA loci) is often measured. However, these loci are selectively neutral (or near neutral) and may not accurately reflect genomewide variation. Variation at functional trait loci, such as the major histocompatibility complex (MHC), can provide a better assessment of adaptive genetic variation in fragmented populations. We compared patterns of microsatellite and MHC variation across three Eastern Massasauga (Sistrurus catenatus) populations representing a gradient of demographic histories to assess the relative roles of natural selection and genetic drift. Using 454 deep amplicon sequencing, we identified 24 putatively functional MHC IIB exon 2 alleles belonging to a minimum of six loci. Analysis of synonymous and nonsynonymous substitution rates provided evidence of historical positive selection at the nucleotide level, and Tajima's D provided support for balancing selection in each population. As predicted, estimates of microsatellite allelic richness, observed, heterozygosity, and expected heterozygosity varied among populations in a pattern qualitatively consistent with demographic history and abundance. While MHC allelic richness at the population and individual levels revealed similar trends, MHC nucleotide diversity was unexpectedly high in the smallest population. Overall, these results suggest that genetic variation in the Eastern Massasauga populations in Illinois has been shaped by multiple evolutionary mechanisms. Thus, conservation efforts should consider both neutral and functional genetic variation when managing captive and wild Eastern Massasauga populations.