Genetics and the conservation of natural populations: allozymes to genomes.

I consider how the study of genetic variation has influenced efforts to conserve natural populations over the last 50 years. Studies with allozymes in the 1970s provided the first estimates of the amount of genetic variation within and between natural populations at multiple loci. These early studies played an important role in developing plans to conserve species. The description of genetic variation in mitochondrial DNA in the early 1980s laid the foundation for the field of phylogeography, which provided a deeper look in time of the relationships and connectivity among populations. The development of microsatellites in the 1990s provided much more powerful means to describe genetic variation at nuclear loci, including the ability to detect past bottlenecks and estimate current effective population size with a single temporal sample. In the 2000s, single nucleotide polymorphisms presented a cornucopia of loci that has greatly improved power to estimate genetic and population demographic parameters important for conservation. Today, population genomics presents the ability to detect regions of the genome that are affected by natural selection (e.g. local adaptation or inbreeding depression). In addition, the ability to genotype historical samples has provided power to understand how climate change and other anthropogenic phenomena have affected populations. Modern molecular techniques provide unprecedented power to understand genetic variation in natural populations. Nevertheless, application of this information requires sound understanding of population genetics theory. I believe that current training in conservation genetics focuses too much on the latest techniques and too little on understanding the conceptual basis which is needed to interpret these data and ask good questions.

Cryptic speciation in allotetraploids: Lessons from the Botrychium matricariifolium complex.

PREMISE OF THE STUDY: Cryptic species are a challenge for botanists and taxonomists. To improve species delineation in the genus Botrychium (Ophioglossaceae), which includes multiple instances of allotetraploid speciation, we examined a cryptic species complex using genetics and morphology. METHODS: We sampled species in the B. matricariifolium complex, concentrating on the Upper Peninsula of Michigan and including multiple proposed morphospecies. We analyzed over 1500 samples using 10 enzyme systems, measured 42 quantitative and qualitative morphological characters for over 650 individuals, and analyzed 145 samples using AFLPs. We tested for diagnostic enzymes in the morphospecies and calculated the correlation between morphological and genetic distances to determine whether putatively distinct morphotypes warrant taxonomic recognition. KEY RESULTS: Allozyme allelic variation corresponded loosely to some morphotypes of B. matricariifolium, but with lower genetic distinction among them than found between B. matricariifolium and B. michiganense. Botrychium michiganense contains unique alleles, indicating a different hybrid origin from that of B. matricariifolium and supporting its status as a genetically distinct species. CONCLUSIONS: We showed that B. acuminatum morphology and genetics are accommodated taxonomically within B. matricariifolium; B. matricariifolium and B. michiganense likely represent hybridization events between related species; and morphotypes within B. matricariifolium likely represent repeated hybridization events between the same two parental species. These hybridizations have resulted in the array of morphotypes observed by field botanists. By helping to identify diagnostic morphological characters, genetic analyses also help us understand and resolve morphological variation observed in the field.

Evolutionary history of Ichthyosaura alpestris (Caudata, Salamandridae) inferred from the combined analysis of nuclear and mitochondrial markers.

Widespread species with morphologically and ecologically differentiated populations are key to understand speciation because they allow investigating the different stages of the continuous process of population divergence. The alpine newt, Ichthyosaura alpestris, with a range that covers a large part of Central Europe as well as isolated regions in all three European Mediterranean peninsulas, and with strong ecological and life-history differences among populations, is an excellent system for such studies. We sampled individuals across most of the range of the species, and analyzed mitochondrial (1442 bp) and nuclear (two nuclear genes -1554 bp- and 35 allozyme loci) markers to produce a time-calibrated phylogeny and reconstruct the historical biogeography of the species. Phylogenetic analyses of mtDNA data produced a fully resolved topology, with an endemic, Balkan clade (Vlasina) which is sister to a clade comprising an eastern and a western group. Within the former, one clade (subspecies I. a. veluchiensis) is sister to a clade containing subspecies I. a. montenegrina and I. a. serdara as well as samples from southern Romania, Bosnia-Herzegovina, Serbia and Bulgaria (subspecies I. a. reiseri and part of I. a. alpestris). Within the western group, populations from the Italian peninsula (subspecies I. a. apuana and I. a. inexpectata) are sister to a clade containing samples from the Iberian Peninsula (subspecies I. a. cyreni) and the remainder of the samples from subspecies I. a. alpestris (populations from Hungary, Austria, Poland, France, Germany and the larger part of Romania). Results of (∗)BEAST analyses on a combined mtDNA and nDNA dataset consistently recovered with high statistical support four lineages with unresolved inter-relationships: (1) subspecies I. a. veluchiensis; (2) subspecies I. a. apuana+I. a. inexpectata; (3) subspecies I. a. cyreni+part of subspecies I. a. alpestris (the westernmost populations, plus most Romanian populations); and (4) the remaining populations, including subspecies I. a. serdara, I. a. reiseri and I. a. montenegrina and part of subspecies I. a. alpestris, plus samples from Vlasina. Our time estimates are consistent with ages based on the fossil record and suggest a widespread distribution for the I. alpestris ancestor, with the split of the major eastern and western lineages during the Miocene, in the Tortonian. Our study provides a solid, comprehensive background on the evolutionary history of the species based on the most complete combined (mtDNA+nDNA+allozymes) dataset to date. The combination of the historical perspective provided by coalescent-based analyses of mitochondrial and nuclear DNA variation with individual-based multilocus assignment methods based on multiple nuclear markers (allozymes) also allowed identification of instances of discordance across markers that highlight the complexity and dynamism of past and ongoing evolutionary processes in the species.

Autotetraploids of Vicia cracca show a higher allelic richness in natural populations and a higher seed set after artificial selfing than diploids.

BACKGROUND AND AIMS: Despite the great importance of autopolyploidy in the evolution of angiosperms, relatively little attention has been devoted to autopolyploids in natural polyploid systems. Several hypotheses have been proposed to explain why autopolyploids are so common and successful, for example increased genetic diversity and heterozygosity and the transition towards selfing. However, case studies on patterns of genetic diversity and on mating systems in autopolyploids are scarce. In this study allozymes were employed to investigate the origin, population genetic diversity and mating system in the contact zone between diploid and assumed autotetraploid cytotypes of Vicia cracca in Central Europe. METHODS: Four enzyme systems resolved in six putative loci were investigated in ten diploid, ten tetraploid and five mixed-ploidy populations. Genetic diversity and heterozygosity, partitioning of genetic diversity among populations and cytotypes, spatial genetic structure and fixed heterozygosity were analysed. These studies were supplemented by a pollination experiment and meiotic chromosome observation. KEY RESULTS AND CONCLUSIONS: Weak evidence of fixed heterozygosity, a low proportion of unique alleles and genetic variation between cytotypes similar to the variation among populations within cytotypes supported the autopolyploid origin of tetraploids, although no multivalent formation was observed. Tetraploids possessed more alleles than diploids and showed higher observed zygotic heterozygosity than diploids, but the observed gametic heterozygosity was similar to the value observed in diploids and smaller than expected under panmixis. Values of the inbreeding coefficient and differentiation among populations (ρST) suggested that the breeding system in both cytotypes of V. cracca is mixed mating with prevailing outcrossing. The reduction in seed production of tetraploids after selfing was less than that in diploids. An absence of correlation between genetic and geographic distances and high differentiation among neighbouring tetraploid populations supports the secondary contact hypothesis with tetraploids of several independent origins in Central Europe. Nevertheless, the possibility of a recent in situ origin of tetraploids through a triploid bridge in some regions is also discussed.

Comparative genetic structure between Sedum ussuriense and S. kamtschaticum (Crassulaceae), two stonecrops co-occurring on rocky cliffs.

• Premise of the study: Geographic isolation due to discontinuities of suitable habitat may have significant effects on the genetic structure of plant populations. Even within a few kilometers, physical barriers to gene flow may lead to considerable genetic differentiation among populations.• Methods: Sedum ussuriense is a boreal species that in Korea occurs only in four valleys separated by mountain ranges in Juwangsan National Park and its vicinity (a range of ∼15 km). Its congener S. kamtschaticum, by contrast, co-occurs in the four valleys but also on the intervening mountains. Using 12 allozyme loci, we comparatively assessed genetic variability and structure in 12 population pairs of the two stonecrops.• Key results: While we found high and comparable levels of within-population genetic variation for the two species, among-population divergence was significantly higher in S. ussuriense (FST = 0.261 vs. FST = 0.165). Sedum ussuriense also showed a much higher percentage of among-valley variation (19%) than S. kamtschaticum (4%).• Conclusions: High levels of genetic diversity in the two Sedum species are consistent with the previous hypothesis that mountains of the Korean Peninsula served as glacial refugia for many boreal species. Given that the two congeners have similar life-history traits, the lower among-population differentiation in S. kamtschaticum is attributable to its higher abundance and more continuous distribution in the study area. This study confirms the central role of geographic isolation in the genetic structure of plant species even at very small scales.

Population genetics of Braun's Rockcress (Boechera perstellata, Brassicaceae), an endangered plant with a disjunct distribution.

Boechera perstellata is an endangered plant found only in middle Tennessee and north central Kentucky. After sampling 4 Tennessee and 3 Kentucky populations, genetic variability and population structure were examined for this species using isozymes, chloroplast DNA, and microsatellites (averaging 35, 29, and 27 individuals per population per class of marker, respectively). The only genetic variability detected for 23 isozymes was a fixed difference between Tennessee and Kentucky populations at 1 locus. Fixed differences between populations of the 2 states were also observed for 3 chloroplast markers. Polymorphism at 19 nuclear microsatellites was 74% at the species level and averaged 21% at the population level. However, observed heterozygosity was extremely low in all populations, ranging from 0.000 to 0.005. High F IS values (0.93) suggest that Boechera perstellata is a primarily selfing species. Tennessee populations have more genetic diversity than Kentucky populations of B. perstellata. Microsatellite markers revealed substantial genetic divergence between the states and genetic differences among populations within each state. Analysis of molecular variance indicates that most variability in this species occurs between the 2 states (49%) and among populations within each state (42%), with relatively little variation found within populations (9%). These data indicate that there is very little gene flow among populations of B. perstellata and that it is important to protect as many populations as possible in order to conserve the genetic diversity of this rare species.

Significant spatial aggregation and fine-scale genetic structure in the homosporous fern Cyrtomium falcatum (Dryopteridaceae).

Spores of homosporous ferns are small, wind-borne and thus have the potential for long-distance dispersal. This common perception has led to a prediction of near-random spatial genetic structure within fern populations. Spore dispersal and spore bank studies, however, indicate that most spores fall close to the maternal plant (< 5 m), supporting a prediction of significant fine-scale genetic structure (FSGS) within populations. To determine which of these two hypotheses is more likely to occur in nature, we measured inbreeding and quantified the spatial distribution of individuals and allozyme-based genotypes using spatial autocorrelation methods within four populations of the fern Cyrtomium falcatum in southern South Korea. Inbreeding levels were low, and all populations exhibited significant aggregation of individuals and strong FSGS. The present results support the second hypothesis, and the substantial FSGS in C. falcatum could reflect the unique features of most homosporous ferns (outcrossing mating systems that lead a majority of spores to occur at short distances and a very limited dispersal distance of male gametes). Although fern spores are physically analogous to orchid seeds, the intensity of FSGS exhibited in C. falcatum is four times stronger than that in 16 terrestrial orchid species.

How complex is the Bufo bufo species group?

Species delineation remains one of the most challenging tasks in the study of biodiversity, mostly owing to the application of different species concepts, which results in contrasting taxonomic arrangements. This has important practical consequences, since species are basic units in fields like ecology and conservation biology. We here review molecular genetic evidence relevant to the systematics of toads in the Bufo bufo species group (Anura, Bufonidae). Two studies recently published in this journal (Recuero et al., MPE 62: 71-86 and García-Porta et al., MPE 63: 113-130) addressed this issue but reached opposing conclusions on the taxonomy of the group (four versus two species). In particular, allozyme data in the latter paper were interpreted as evidence for hybridization across species (between B. bufo-B. spinosus and B. bufo-B. verrucosissimus). We tested claims for hybridization through re-analysis of allozyme data for individuals instead of populations, to be able to distinguish between sympatry with and without admixture, and found no evidence of hybridization across taxa. We propose alternative explanations for the observed patterns that García-Porta et al. (2012) failed to consider. In the absence of unequivocal evidence for hybridization and introgression, we reject the proposal to downgrade Bufo spinosus and Bufo verrucosissimus to the subspecies level.

A review of the allozyme data set for the Canarian endemic flora: causes of the high genetic diversity levels and implications for conservation.

Background and Aims Allozyme and reproductive data sets for the Canarian flora are updated in order to assess how the present levels and structuring of genetic variation have been influenced by the abiotic island traits and by phylogenetically determined biotic traits of the corresponding taxa; and in order to suggest conservation guidelines. Methods Kruskal-Wallis tests are conducted to assess the relationships of 27 variables with genetic diversity (estimated by A, P, Ho and He) and structuring (GST) of 123 taxa representing 309 populations and 16 families. Multiple linear regression analyses (MLRAs) are carried out to determine the relative influence of the less correlated significant abiotic and biotic factors on the genetic diversity levels. Key Results and Conclusions The interactions between biotic features of the colonizing taxa and the abiotic island features drive plant diversification in the Canarian flora. However, the lower weight of closeness to the mainland than of (respectively) high basic chromosome number, partial or total self-incompatibility and polyploidy in the MLRAs indicates substantial phylogenetic constraint; the importance of a high chromosome number is feasibly due to the generation of a larger number of linkage groups, which increase gametic and genotypic diversity. Genetic structure is also more influenced by biotic factors (long-range seed dispersal, basic chromosome number and partial or total self-incompatibility) than by distance to the mainland. Conservation-wise, genetic structure estimates (FST/GST) only reflect endangerment under intensive population sampling designs, and neutral genetic variation levels do not directly relate to threat status or to small population sizes. Habitat protection is emphasized, but the results suggest the need for urgent implementation of elementary reproductive studies in all cases, and for ex situ conservation measures for the most endangered taxa, even without prior studies. In non-endangered endemics, multidisciplinary research is needed before suggesting case-specific conservation strategies. The molecular information relevant for conservation should be conserved in a standardized format to facilitate further insight.

Population history of the two carnivorous plants Drosera peltata var. nipponica and Drosera rotundifolia (Droseraceae) in Korea.

PREMISE OF THE STUDY: Drosera peltata var. nipponica, an element of the East Asia warm-temperate vegetation, and D. rotundifolia, a widely distributed boreal species, reach one of their northernmost and southernmost limits, respectively, on the Korean Peninsula. Because the Last Glacial Maximum (LGM)-Holocene dynamics of warm-temperate and boreal paleovegetation differed considerably on the Peninsula, the population history of these two sundews is expected to be different, leaving differential imprints in their genetic structure. METHODS: We investigated population genetic structure of D. peltata var. nipponica and D. rotundifolia in South Korea (10 populations of each for 20 allozyme loci) to infer their population history in this region. In addition, we compared the genetic variation harbored in the two sundews to those reported for other carnivorous and wetland plants. KEY RESULTS: Drosera peltata var. nipponica showed no genetic diversity, whereas D. rotundifolia exhibited extremely low within-population variation (He = 0.005) and considerable among-population divergence (FST = 0.817). CONCLUSIONS: Our results suggest that extant populations of D. peltata var. nipponica likely originated from a single ancestral population from southern Japan or southern China through postglacial dispersal. On the contrary, D. rotundifolia probably survived the LGM in situ, with extant populations derived from either one or several small source populations. We argue that separate conservation strategies should be employed, given that the two taxa have different ecological and demographic traits and harbor different levels of genetic diversity.

Microsatellites as Molecular Markers with Applications in Exploitation and Conservation of Aquatic Animal Populations.

A large number of species and taxa have been studied for genetic polymorphism. Microsatellites have been known as hypervariable neutral molecular markers with the highest resolution power in comparison with any other markers. However, the discovery of a new type of molecular marker-single nucleotide polymorphism (SNP) has put the existing applications of microsatellites to the test. To ensure good resolution power in studies of populations and individuals, a number of microsatellite loci from 14 to 20 was often used, which corresponds to about 200 independent alleles. Recently, these numbers have tended to be increased by the application of genomic sequencing of expressed sequence tags (ESTs) and the choice of the most informative loci for genotyping depends on the aims of research. Examples of successful applications of microsatellite molecular markers in aquaculture, fisheries, and conservation genetics in comparison to SNPs are summarized in this review. Microsatellites can be considered superior markers in such topics as kinship and parentage analysis in cultured and natural populations, the assessment of gynogenesis, androgenesis and ploidization. Microsatellites can be coupled with SNPs for mapping QTL. Microsatellites will continue to be used in research of genetic diversity in cultured stocks, and also in natural populations as an economically advantageous genotyping technique.

Sulfation of 12-hydroxy-nevirapine by human SULTs and the effects of genetic polymorphisms of SULT1A1 and SULT2A1.

Nevirapine (NVP) is an effective drug for the treatment of HIV infections, but its use is limited by a high incidence of severe skin rash and liver injury. 12-Hydroxynevirapine (12-OH-NVP) is the major metabolite of nevirapine. There is strong evidence that the sulfate of 12-OH-NVP is responsible for the skin rash. While several cytosolic sulfotransferases (SULTs) have been shown to be capable of sulfating 12-OH-NVP, the exact mechanism of sulfation in vivo is unclear. The current study aimed to clarify human SULT(s) and human organs that are capable of sulfating 12-OH-NVP and investigate the metabolic sulfation of 12-OH-NVP using cultured HepG2 human hepatoma cells. Enzymatic assays revealed that of the thirteen human SULTs, SULT1A1 and SULT2A1 displayed strong 12-OH-NVP-sulfating activity. 1-Phenyl-1-hexanol (PHHX), which applied topically prevents the skin rash in rats, inhibited 12-OH-NVP sulfation by SULT1A1 and SULT2A1, implying the involvement of these two enzymes in the sulfation of 12-OH-NVP in vivo. Among five human organ cytosols analyzed, liver cytosol displayed the strongest 12-OH-NVP-sulfating activity, while a low but significant activity was detected with skin cytosol. Cultured HepG2 cells were shown to be capable of sulfating 12-OH-NVP. The effects of genetic polymorphisms of SULT1A1 and SULT2A1 genes on the sulfation of 12-OH-NVP by SULT1A1 and SULT2A1 allozymes were investigated. Two SULT1A1 allozymes, Arg37Asp and Met223Val, showed no detectable 12-OH-NVP-sulfating activity, while a SULT2A1 allozyme, Met57Thr, displayed significantly higher 12-OH-NVP-sulfating activity compared with the wild-type enzyme. Collectively, these results contribute to a better understanding of the involvement of sulfation in NVP-induced skin rash and provide clues to the possible role of SULT genetic polymorphisms in the risk of this adverse reaction.

From allozymes to NGS: population genetics of forest trees in Slovakia in the past 40 years.

This review summarizes the development of population genetics and population genomics studies of forest trees in Slovakia during the past 40 years. Various protein and DNA markers have been applied during this period to address several topics in evolutionary genetics and biogeography of trees: allozymes, uniparentally inherited chloroplast and mitochondrial markers, simple sequence repeats and single nucleotide polymorphisms. The main object of studies of phylogeny and postglacial migration were Fagus sylvatica s.l. and eastern-Mediterranean firs (Abies Mill. section Abies), where the divergence of genetic lineages (species and subspecific taxa) in time, as well as colonization of the current ranges during the Holocene were reconstructed. The studies on intraspecific gene flow and homoploid hybridization focused on hybrid swarms Pinus sylvestris/P. mugo and firs. Unusual maternal inheritance of chloroplast DNA was revealed in P. mugo × P. sylvestris crosses. Contrasting geographical structures of hybrid zones were revealed in wind-dispersed vs. animal-dispersed trees. Within the studies of adaptation, signals of selection were identified both in field observations and common-garden experiments on Picea abies, F. sylvatica and A. alba. Perspectives of ongoing research employing next-generation sequencing were shortly outlined.

Biochemical comparison of two Hypostomus populations (Siluriformes, Loricariidae) from the Atlântico Stream of the upper Paraná River basin, Brazil.

Two syntopic morphotypes of the genus Hypostomus - H. nigromaculatus and H. cf. nigromaculatus (Atlântico Stream, Paraná State) - were compared through the allozyme electrophoresis technique. Twelve enzymatic systems (AAT, ADH, EST, GCDH, G3PDH, GPI, IDH, LDH, MDH, ME, PGM and SOD) were analyzed, attributing the score of 20 loci, with a total of 30 alleles. Six loci were diagnostic (Aat-2, Gcdh-1, Gpi-A, Idh-1, Ldh-A and Mdh-A), indicating the presence of interjacent reproductive isolation. The occurrence of few polymorphic loci acknowledge two morphotypes, with heterozygosity values He = 0.0291 for H. nigromaculatus and He = 0.0346 for H. cf. nigromaculatus. F(IS) statistics demonstrated fixation of the alleles in the two morphotypes. Genetic identity (I) and distance (D) of Nei (1978) values were I = 0.6515 and D = 0.4285. The data indicate that these two morphotypes from the Atlântico Stream belong to different species.

Allozyme comparison of two populations of Rineloricaria (Siluriformes, Loricariidae) from the Ivaí River, upper Paraná River basin, Brazil.

Two allopatric morphotypes of the genus Rinelocaria were compared through the allozyme electrophoresis technique: one morphotype, R. pentamaculata, from the Keller River in the middle stretch of the Ivaí River basin and the other, R. aff. pentamaculata, from the São João River in the upper portion of the Ivaí River basin. The morphotype from the São João River was collected upstream from the São João waterfall, which is about 80 m deep. Twelve enzymatic systems (AAT, ADH, EST, GCDH, G3PDH, GPI, IDH, LDH, MDH, ME, PGM and SOD) were analyzed, which allowed to score 22 loci. Only lociAat-2, Est-3 and Mdh-C showed polymorphism. The two samples differed in allele frequencies at the three polymorphic loci. The average expected heterozygosity for all loci was 0.0806 ± 0.0447 in the Keller River sample. For the São João River morphotype, this value was 0.0489 ± 0.0350. Nei' s genetic identity and distance between the two populations were respectively 0.9789 and 0.0213. Wright's F(IS) , F(IT) and F(ST) over all loci were estimated as 0.3121, 0.4021 and 0.1309, respectively. We consider that the two morphotypes represent species in statunascendi.

Integrative taxonomic reassessment of Odontophrynus populations in Argentina and phylogenetic relationships within Odontophrynidae (Anura).

Amphibians are the most vulnerable vertebrates to biodiversity loss mediated by habitat destruction, climate change and diseases. Informed conservation management requires improving the taxonomy of anurans to assess reliably the species' geographic range. The genus Odontophrynus that is geographically refined to Argentina, Bolivia, Brazil, Uruguay and Paraguay includes currently 12 nominal species with many populations of uncertain taxonomic assignment and subsequently unclear geographic ranges. In this study, we applied integrative taxonomic methods combining molecular (mitochondrial 16S gene), allozyme, morphological and bioacoustic data to delimit species of the genus Odontophrynus sampled from throughout Argentina where most species occur. The combined evidence demonstrates one case of cryptic diversity and another of overestimation of species richness. The populations referred to as O. americanus comprise at least three species. In contrast, O. achalensis and O. barrioi represent junior synonyms of the phenotypically plastic species O. occidentalis. We conclude that each of the four species occurring in Argentina inhabits medium to large areas. The Red List classification is currently "Least Concern". We also propose a phylogenetic hypothesis for the genus and associated genera Macrogenioglottus and Proceratophrys (Odontophrynidae).

Chromosomal Localization and Contribution of Three Homoeologous Genes to Biosynthesis of Cytosolic Aspartate Aminotransferase in Common Wheat.

Chromosomal localization of the three homoeologous genes encoding cytosolic aspartate aminotransferase in common wheat (Triticum aestivum cv. Chinese Spring, 2n = 6x = 42, AABBDD) was specified to: 3AL (0.42÷0.61), 3BL (0.38÷0.41) and 3DL (0.23÷0.81) by a comparative zymographic analysis of the enzymatic activities in deletion lines. It was also attempted to precisely explain the nature of the relationship between a number of genes encoding α and ß subunits and a distribution of staining intensity of cytosolic aspartate aminotransferase allozyme activity bands using aneuploid lines of common wheat with modified third pair of homoeologous chromosomes from genomes A, B and D, on which the genes encoding subunit α (genome A) and ß (genome B and D) are localized. The highest consistency between the experimental results and the theoretical distributions was achieved by substituting values of α = 0.57 and ß = 0.43 in a theoretical model. These results demonstrate that the individual participation of the diploid genome A in the biosynthesis of the cytosolic aspartate aminotransferase allozymes subunits is greater than the individual participation of the diploid genomes B and D.

Genetic variation in natural Melandrium album populations exposed to chronic ionizing radiation.

The effect of radiation pollution on genetic variation in natural populations of Melandrium album was investigated at the head part of the East-Ural Radioactive Trace (EURT) and background areas. The highest genetic differentiation estimated using F ST was revealed between compared pairs of the background and impact samples in populations of M. album. The highest rate of polymorphism was observed at the closest to nuclear accident, Impact-1 site. The unique alleles (Mdh-3104, Pgi-2106, Lap 105, Mdh-296, and Dia 94) were discovered at the EURT. Individuals from chronically low-level irradiated sites were genetically closer than to plants from background sites using Nadhdh locus. The increase of the frequency of unique homozygous and heterozygous genotypes was identified in populations of M. album growing under chronic radiation exposure conditions. The largest contribution to the group of unique heterozygous genotypes at the EURT was made by three loci - Lap, Pgi-2, and Nadhdh; the main role in interpopulation differentiation of samples was made by the alleles Sod-2115, Skdh 100, and Nadhdh 100. Our results provide evidence for the correlation between the increase of genetic variation other than the «genetic erosion¼ and chronic radiation exposure factor in natural plant populations.

Temporal genetic variation of Fasciola hepatica from sheep in Galicia (NW Spain).

We found low genetic differentiation between two temporal samples of Fasciola hepatica (2006 and 2008) collected from nine sheep of the same flock that shared the same pasture for at least 2 years. However, each sample, represented by four and five infrapopulations respectively, showed strong heterozygote deficits regarding Hardy-Weinberg expectations and a high degree of genetic structure at infrapopulation level. This is an unexpected result since genetic drift should increase temporal variation among years. Our findings are most likely explained by the fact that the parasite can survive many years in the definitive host. Temporal gene flow favored by high longevity probably increases levels of genetic variability of the population but could also contribute to the observed heterozygote deficits within temporal samples and infrapopulations if it favors the Wahlund effect. Despite the homogenizing effect of gene flow, the high genetic divergence observed between infrapopulations is most likely a consequence of strong genetic drift associated to the complexity of the life cycle.

Temporal stability of parasite distribution and genetic variability values of Contracaecum osculatum sp. D and C. osculatum sp. E (Nematoda: Anisakidae) from fish of the Ross Sea (Antarctica).

The Ross Sea, Eastern Antarctica, is considered a "pristine ecosystem" and a biodiversity "hotspot" scarcely impacted by humans. The sibling species Contracaecum osculatum sp. D and C. osculatum sp. E are anisakid parasites embedded in the natural Antarctic marine ecosystem. Aims of this study were to: identify the larvae of C. osculatum (s.l.) recovered in fish hosts during the XXVII Italian Expedition to Antarctica (2011-2012); perform a comparative analysis of the contemporary parasitic load and genetic variability estimates of C. osculatum sp. D and C. osculatum sp. E with respect to samples collected during the expedition of 1993-1994; to provide ecological data on these parasites. 200 fish specimens (Chionodraco hamatus, Trematomus bernacchii, Trematomus hansoni, Trematomus newnesi) were analysed for Contracaecum sp. larvae, identified at species level by allozyme diagnostic markers and sequences analysis of the mtDNA cox2 gene. Statistically significant differences were found between the occurrence of C. osculatum sp. D and C. osculatum sp. E in different fish species. C. osculatum sp. E was more prevalent in T. bernacchii; while, a higher percentage of C. osculatum sp. D occurred in Ch. hamatus and T. hansoni. The two species also showed differences in the host infection site: C. osculatum sp. D showed higher percentage of infection in the fish liver. High genetic variability values at both nuclear and mitochondrial level were found in the two species in both sampling periods. The parasitic infection levels by C. osculatum sp. D and sp. E and their estimates of genetic variability showed no statistically significant variation over a temporal scale (2012 versus 1994). This suggests that the low habitat disturbance of the Antarctic region permits the maintenance of stable ecosystem trophic webs, which contributes to the maintenance of a large populations of anisakid nematodes with high genetic variability.