Evaluation of the state of the gene pool of natural populations dwelling in the fragmented landscape of Moscow and Moscow region (with special reference to brown frogs)

A system of urban ecological genetic monitoring of vertebrates has been developed for the first time in Russia and in the world. As a model, two species of brown frogs, Rana arvalis Nilss. and R. temporaria L. (16 populations, seven isozyme loci) were used. An evaluation of the gene pool state in urban frog populations has shown that the diversity in the Moscow frog populations was lower than that in natural populations (up to 80 and 50% in respectively R. arvalis and R. temporaria). Mean heterozygosities per locus were higher in large natural populations than in small urban isolates: in R. arvalis, these values were 0.16 and 0.06; in R. temporaria, 0.34 and 0.18, respectively. The number of polymorphic loci was also higher in natural populations than in the urban ones: 4 versus 2 in R. arvalis and 5 versus 4 in R. temporaria. Using superoxiddismutase as an example, fixation of different alleles of the same locus in different small isolates was shown. The gene pool condition of all but one urban populations of brown frog was evaluated as unsatisfactory, and that of the R. arvalis populations, as critical. These changes of the gene pool are explained mainly by gene drift accompanied by inbreeding, which was caused by human-induced fragmentation of the range and a decrease in population size of the species. The results of this study was employed in the development of the Moscow governmental program on restoration of the gene pools of vanishing animal species on specially protected natural urban territories. The series of works on long-term monitoring and assessment of the state of natural populations of model species in anthropogenic landscapes of Moscow and Moscow region has laid a foundation for a new branch of science, gene urbanology.     

Genetic structure of the critically endangered plant Tricyrtis ishiiana (Convallariaceae) in relict populations of Japan

Tricyrtis ishiiana is a relic endemic plant taxon of the Convallariaceae that inhabits two nearby gorges in Kanagawa Prefecture, Japan. The distribution range and number of populations are thought to have been reduced to the present refugial populations during the Quaternary climatic oscillations. Because of its showy flowers, this plant has faced illegal removal from its natural habitats for horticultural use and has been designated a critically endangered species (class IA). In this study, we analyzed the genetic structure of the relict populations of T. ishiiana in order to contribute to the conservation strategies of the prefectural government. Our analyses of nine nuclear microsatellite loci detected high genetic diversity (H _E = 0.704 and H _O = 0.541) for the two populations. The two populations were slightly differentiated (R _ST = 0.032), accompanied by faint substructure across the populations (K = 3). In addition, each population exhibited spatial genetic structuring. The relatively low inbreeding coefficient for both populations together (F _IS = 0.233) and each population separately (F _IS = 0.217?C0.246) may be attributable to crossing among descendants within a population along with occasional gene flow between the populations. These results suggested that the extant populations have not experienced a severe bottleneck. The two extant populations were genetically differentiated at a very low level, accompanied by occasional pollen flow via pollinators and/or seed dispersal by gravity in the mountainous environment. Occasional gene exchange between the populations has allowed T. ishiiana to harbor high genetic diversity despite being a relic plant confined to two small refugial populations.     

Genetic Diversity and Population Structure of the Rare and Endangered Plant Species Pulsatilla patens (L.) Mill in East Central Europe

Pulsatilla patens s.s. is a one of the most endangered plant species in Europe. The present range of this species in Europe is highly fragmented and the size of the populations has been dramatically reduced in the past 50 years. The rapid disappearance of P. patens localities in Europe has prompted the European Commission to initiate active protection of this critically endangered species. The aim of this study was to estimate the degree and distribution of genetic diversity within European populations of this endangered species. We screened 29 populations of P. patens using a set of six microsatellite primers. The results of our study indicate that the analyzed populations are characterized by low levels of genetic diversity (H_o = 0.005) and very high levels of inbreeding (F_IS = 0.90). These results suggest that genetic erosion could be partially responsible for the lower fitness in smaller populations of this species. Private allelic richness was very low, being as low as 0.00 for most populations. Average genetic diversity over loci and mean number of alleles in P. patens populations were significantly correlated with population size, suggesting severe genetic drift. The results of AMOVA point to higher levels of variation within populations than between populations.The results of Structure and PCoA analyses suggest that the genetic structure of the studied P. patens populations fall into three clusters corresponding to geographical regions. The most isolated populations (mostly from Romania) formed a separate group with a homogeneous gene pool located at the southern, steppic part of the distribution range. Baltic, mostly Polish, populations fall into two genetic groups which were not fully compatible with their geographic distribution.Our results indicate the serious genetic depauperation of P. patens in the western part of its range, even hinting at an ongoing extinction vortex. Therefore, special conservation attention is required to maintain the populations of this highly endangered species of European Community interest.     

Genetic and Phylogeographic Structures of the Symbiotic Fungus Tuber magnatum

The quality and market price of truffles vary with the species and, traditionally, the place of origin. The premium species Tuber magnatum produces white truffles and has a patchy distribution restricted to Italy and some Balkan areas. We used polymorphic microsatellites to evaluate 316 specimens grouped into 26 populations sampled across the species' geographic range to determine if natural populations of T. magnatum are genetically differentiated. We found that the southernmost and the northwesternmost populations were significantly differentiated from the rest of the populations. The simple sequence repeat data also could be used to make inferences about the postglacial T. magnatum expansion pattern. This study is the first to identify a genetic and phylogeographic structure in T. magnatum. The presence of a genetic structure can be of practical interest in tracing truffle populations according to their geographic origin for marketing strategies. Evidence for extensive outcrossing in field populations of T. magnatum also is provided for the first time.     

Clonal structure and reduced diversity of the invasive alien plant Erigeron annuus in Lithuania

The alien species Erigeron annuus (L.) Pers. is in an intensive spreading phase in Lithuania. Random amplified polymorphic DNA (RAPDs) and inter-simple sequence repeats (ISSRs) assays were used to study the genetic structure of old and new invasive populations and to determine the most spread genotypes of this species in Lithuania. Pairwise genetic distances between populations established using RAPD and ISSR markers significantly correlated (r=0.91, P<0.05). Our study indicates that there are two genetically different types of E. annuus populations. The first type is represented by a widely spread main clone and related monomorphic populations. The second type is represented by polymorphic populations, some of them present at sites where E. annuus has not been previously observed. Main clone predominates in nine populations and is from the region where this species was first described in natural ecosystems of Lithuania. UPGMA cluster analysis revealed genetic relationships between the main clone and accessions from old cemeteries where E. annuus has been grown as an ornamental plant. We found high genetic differentiation among populations (G _ST=0.58 for RAPDs, G _ST=0.64 for ISSRs). Taken together, our results will contribute to the monitoring of E. annuus spread in Lithuania.     

Molecular phylogenetic study of the Iranians based on polymorphic markers

The application of polymorphic markers in construction of phylogenetic trees has been documented. Five polymorphic markers located in the PAH gene region including PAH-BglII, PAH-PvuII(A), PAH-EcoRI, PAH-MspI and PAH-STR were selected for analysis of phylogenetic relationships of the Iranians with 15 other populations of the world. The lowest genetic distance was observed between the Iranians and populations residing in Adygei (an ethnic group of the Russian Caucasus), Russia and Druze (a Middle Eastern group). However, East Asian populations including Han, Japanese and Cambodians, Khmer or the Oceanians (Melanesian, Nasioi) showed high genetic distance with the Iranians. The data suggested that the Iranians might have relatively close evolutionary history with the populations residing in Russia rather than East Asian populations. This study provided the first new molecular insight into the evolutionary history of the Iranian population.     

Range expansion and lineage admixture of the Japanese evergreen tree Machilus thunbergii in central Japan

We investigated the range expansion histories of Machilus thunbergii populations in the Kinki region of central Japan on the basis of nuclear microsatellite data. In the Kinki region, M. thunbergii is typically found in the coastal area, with some fragmented populations inland, around Lake Biwa. Phylogenetic and Bayesian clustering analysis (STRUCTURE analysis) revealed that the inland populations have different genetic components between the west and east sides of Lake Biwa. The population located on the north side of the lake has an admixture of the two genetically differentiated lineages, contributing to an increase in the genetic diversity of the population. Populations around Lake Biwa had lost rare alleles and the F value obtained from STRUCTURE analysis was lower in the coastal populations than in the lake populations. These results suggest that populations around Lake Biwa experienced a bottleneck due to a founder effect during the initial migration to the lake and that glacial refugia of M. thunbergii in the Kinki region existed along the coast.     

Near Fixation of 374l Allele Frequencies of the Skin Pigmentation Gene SLC45A2 in Africa

The L374F polymorphism of the SLC45A2 gene, encoding the membrane-associated transporter protein that plays an important role in melanin synthesis, has been suggested to be associated with skin color in human populations. In this study, the detailed distribution of the 374f and 374l alleles has been investigated in 2,581 unrelated subjects from 36 North, East, West, and Central African populations. We found once more the highly significant (p < 0.001) correlation coefficient (r = 0.957) cline of 374f frequencies with degrees of latitude in European and North African populations. Almost all the African populations located below 16° of latitude are fixed for the 374l allele. Peul, Toucouleur, and Soninké populations have 374l allele frequencies of 0.06, 0.03, and 0.03, respectively.     

Molecular evidence for the origin and evolutionary history of the rare American desert monotypic family Setchellanthaceae

Setchellanthus caeruleus, which has disjunct populations in the north of the Chihuahuan Desert and in the Tehuacán-Cuicatlán valley, was selected to understand the evolutionary history of plants in this desert and its southerly relicts. This species constitutes the monotypic family Setchellanthaceae, which forms part of a group of plants that produce mustard-oil glucosides or glucosinolates. Molecular phylogenetic analyses based on DNA plastid sequences of plants of S. caeruleus from both areas, including representative taxa of the order Brassicales, were carried out to estimate the time of origin of the family (based on matK?+?rcbL) and divergence of populations (based on psbI-K, trnh-psbA, trnL-trnF). In addition, comparative ecological niche modelling was performed to detect if climate variables vary significantly in northern and southern populations. Analyses revealed that Setchellanthaceae is an ancient lineage that originated between 78 and 112 Mya during the mid-late Cretaceous—much earlier than the formation of the Chihuahuan Desert. The molecular data matrix displayed a few indel events as the only differences of plastid DNA sequences between northern and southern populations. It is suggested that due to climate changes in this desert in the Pliocene, populations of Setchellanthus remained in the Sierra de Jimulco and in Cuicatlán, in climatically stable locations. Ecological niche models of northern populations predict niches of southern populations and identity niche tests indicate that there are no differences in their ecological niches.     

Variation in metallothionein gene expression is associated with adaptation to copper in the earthworm Dendrobaena octaedra

Evolution of resistance to heavy metals has been reported for several populations of soil living organisms occurring at metal contaminated sites. Such genetically based and heritable resistance contribute to the persistence of populations in contaminated areas. Here we report on molecular responses to experimental copper in populations of the earthworm, Dendrobaena octaedra, originating from copper contaminated soil near Gusum (Sweden) where heavy metal pollution has been present for several decades. We studied gene expression of six genes potentially involved in resistance to copper toxicity using F2-generations of D. octaedra populations, originating from reference sites and contaminated (High, Medium and Low) sites around Gusum. The main result was different expression patterns of genes encoding for two different isoforms (mt1 and mt2) of metallothionein proteins during experimental exposure to copper contaminated soil. Expression of mt1 showed a fast and significant upregulation in the High population and a slower, albeit significant, upregulation in Medium and Low populations. However, in the three reference populations no upregulation were seen. In comparison, a fast upregulation was also seen for the High population in the isoform mt2, whereas, gene expression of all other populations, including reference populations, showed slower upregulation in response to experimental copper. The results indicate that copper resistance in D. octaedra from contaminated areas is related to an increased expression of metallothioneins.     

Genetic structure in two meadow varieties ofEuphrasia stricta on the Baltic island of Gotland (Sweden) and implications for conservation

The annual hemiparasiteEuphrasia stricta occurs on Gotland in two early-flowering meadow varieties.E. stricta var.suecica is on the Swedish red-list as endangered, occurring in Sweden only on the Baltic island of Gotland. It probably has near relatives east of the Baltic proper. The other variety,E. stricta var.tenuis, has a wider distribution occurring almost all over Sweden, but is declining in abundance. Both varieties have close morphological similarities and habitat preferences, raising questions about which level conservation efforts should be focused on, varieties or populations. In this study we describe the genetic structure between and within these two varieties using amplified fragment length polymorphism, AFLP.F _ST between varieties is 0.14, and between populations within the varietiesE. stricta var.suecica and var.tenuis F _ST is 0.60 and 0.26 respectively. The partitioning of gene diversity to different levels shows that 14% of the genetic diversity occurs between varieties, 42% between populations within varieties, and 44% within populations. Significant genetic differentiation was detected between varieties, populations and subpopulations within populations using a constrained principal coordinate analysis. We suggest that all of the existing populations of these two varieties on Gotland should be preserved, since they are few (6E. stricta var.suecica and 5E. stricta var.tenuis) and much of the genetic diversity is partitioned between populations.     

Genetic variation and Wolbachia infection status of the lycaenid butterfly Zizina emelina (Lepidoptera,Lycaenidae) from Korean populations

The lycaenid butterfly Zizina emelina emelina (de l’Orza) (previously Zizina otis emelina; Lepidoptera, Lycaenidae) has populations distributed across Japan and Korea. Zizina emelina is listed on Japan’s Red List as an endangered species, owing to the loss of its principal plant food and habitat. Researchers previously investigated the genetic diversity and structure in Japanese populations by using mitochondrial and nuclear DNA analyses alongside their Wolbachia infection status to consider the conservation strategy. In this study, we investigated the genetic variation and Wolbachia infection status of Korean populations and compared our findings with these traits in the Japanese populations. Four novel mitochondrial haplotypes were found in the Korean populations, whereas nuclear DNA analysis revealed one new haplotype and another that matched the one reported previously in Japan. The two known strains of Wolbachia in the Japanese populations were also found in the Korean ones. Mitochondrial DNA analysis demonstrated that no gene flow had occurred between the Japanese and Korean populations. The fact that they share the same nuclear haplotype and Wolbachia strains, however, indicates a low level of population differentiation between the two.     

Systematics of the genus Floridichthys

Samples representing the three nominal subspecies of Floridichthys carpio were examined electrophoretically. Although the populations in Florida could not be distinguished completely from the populations in Yucatan by morphology, 5 of the 30 electrophoretic characters demonstrated fixed differences between Florida and Yucatan populations. Based on the observed genetic differentiation between Florida and Yucatan populations and the absence of genetic differentiation within those populations, we conclude that the Yucatan population has diverged to the species level. We, therefore, propose to elevate the nominal Yucatan subspecies Floridichthys carpio polyommus to a species status.     

Analysis of genetic diversity in the endangered Hucho taimen from China

Hucho taimen are listed as endangered in China. The population size has declined recently, prompting an increase in the level of listing from grade three in 2002 to grade five in 2006. We analyzed the genetic diversity of wild populations using 17 microsatellite markers to establish a scientific basis for conservation of this species. We collected tissue samples from four populations in the Heilongjiang River basin: Huma River (HM), Hutou (HT), Haiqing (HQ), and Zhuaji (ZJ). A total of 21 loci were amplified, 18 of which were polymorphic. The number of alleles per locus ranged from 2 to 9 (mean: 4.1905). There were 13 highly polymorphic loci and 5 moderately polymorphic loci. Analysis of five genetic diversity parameters (Na, Ne, Ho, He, and PIC) suggested moderate levels of diversity within the populations. The populations were ranked HT > HQ > ZJ > HM, but the differences in diversity were not statistically significant (P > 0.05). A comparison of variation among all four populations suggested Hardy–Weinberg disequilibrium at 20% of the loci. Genetic differentiation (Fst) was 0.0644 and the gene flow among populations was estimated at 3.36 individuals per generation. The majority of diversity (93.88%) occurred among individuals within a population. In contrast, relatively little (6.12%) of the genetic diversity was distributed between the populations. An analysis of genetic differentiation and genetic distance between pairs of populations revealed that both parameters were higher in comparisons of the HM population to the HT, HQ, and ZJ populations than among the three latter populations. This suggests that the HM population has a distinct genetic structure. We hypothesize that habitat degradation and excessive fishing, not low genetic diversity, has caused the decline in H. taimen populations. However, this species should be protected from further declines in genetic diversity.     

Inferring the invasion history of coral berry Ardisia crenata from China to the USA using molecular markers

Genetic comparisons between native and invasive populations of a species can provide insights into its invasion history information, which is useful for guiding management and control strategies. The coral berry Ardisia crenata was introduced to Florida last century as a cultivated ornament plant, and has since spread widely throughout the southern regions of the USA. Previously, the genetic variation among 20 natural populations of A. crenata across its distribution center in southern China was quantified using seven microsatellite markers. Here we expand on that work by additionally sampling individuals from four other native populations in Taiwan and Japan, and from five invasive populations in the USA. We also examined the results from one chloroplast intergenic spacer region (trnF-trnL) in all 29 populations. Our aim is to identify the invasion source and subsequent history of the species?? spread throughout the southern USA. We observed lower genetic diversity in the invasive populations based on both microsatellite and chloroplast markers. Our data show that the invasive populations can be clustered with native populations in southeastern China, inferring this region as the geographic origin of A. crenata cultivars invading the USA. We further classified invasive individuals into invasive I and invasive II clusters. Nantou in Taiwan and Xihu in mainland China are the most closely related populations to those, which identify the former as potential sources for host-specific control agents. Our results, combined with the known introduction records, suggest that A. crenata was first multiply introduced into Florida and then secondarily colonized Louisiana and Texas from Florida.     

Naturally Occurring Incompatibilities between Different Culex pipiens pallens Populations as the Basis of Potential Mosquito Control Measures

Background

Vector-borne diseases remain a threat to public health, especially in tropical countries. The incompatible insect technique has been explored as a potential control strategy for several important insect vectors. However, this strategy has not been tested in Culex pipiens pallens, the most prevalent mosquito species in China. Previous works used introgression to generate new strains that matched the genetic backgrounds of target populations while harboring a new Wolbachia endosymbiont, resulting in mating competitiveness and cytoplasmic incompatibility. The generation of these incompatible insects is often time-consuming, and the long-term stability of the newly created insect-Wolbachia symbiosis is uncertain. Considering the wide distribution of Cx. pipiens pallens and hence possible isolation of different populations, we sought to test for incompatibilities between natural populations and the possibility of exploiting these incompatibilities as a control strategy.

Methodology/Principal Findings

Three field populations were collected from three geographic locations in eastern China. Reciprocal cross results showed that bi-directional patterns of incompatibility existed between some populations. Mating competition experiments indicated that incompatible males could compete with cognate males in mating with females, leading to reduced overall fecundity. F1 offspring from incompatible crosses maintained their maternal crossing types. All three populations tested positive for Wolbachia. Removal of Wolbachia by tetracycline rendered matings between these populations fully compatible.

Conclusions/Significance

Our findings indicate that naturally occurring patterns of cytoplasmic incompatibility between Cx. pipiens pallens populations can be the basis of a control strategy for this important vector species. The observed incompatibilities are caused by Wolbachia. More tests including field trials are warranted to evaluate the feasibility of this strategy as a supplement to other control measures.     

Range and variation in the genusCeratozamia (Zamiaceae)

The genusCeratozamia has an extensive distribution that starts in northeastern Mexico and continues southward and southeastward into Guatemala, Belize, and Honduras. Over this extensive distributional rangeCeratozamia is found in numerous disjunct populations. These populations are never very widespread and are often restricted to a single mountain or canyon. Because of the restricted range of these populations, and the lack of genetic exchange between them, many of them have evolved into easily identifiable groups. At the present time there are 16 validly published species ofCeratozamia, with several more possible new species under investigation.     

Divergence and gene flow between the East Sea and the Southeast Atlantic populations of North Pacific light fish Maurolicus japonicus Ishikawa

In the present study, we examined the divergence time and the magnitude of gene flow between two distantly separated populations of North Pacific light fish Maurolicus japonicus, one in the southern part of the East Sea (off Korea) and the other in the Southeast Atlantic Ocean (off Namibia). The mitochondrial 16SrDNA sequences (524 base pairs) obtained from the two populations were analyzed using the isolation with migration (IM) coalescent method as well as the conventional F _ST statistic and a phylogeographic method. A significant nonzero F _ST value (0.176, P<0.05) indicated genetic differentiation between the two populations. The low level of nucleotide diversity compared to the moderately high level of haplotype diversity implied that the populations have experienced a bottleneck followed by rapid growth in both populations. IM analysis suggested that these two populations most likely split approximately 500?C800 K years ago during the Pleistocene climatic oscillations and that gene flow has occurred unidirectionally from the Southeast Atlantic population to the East Sea population. Nested clade phylogeographic analysis supports restricted gene flow between the two populations.     

Monophyly of Opisthorchis viverrini populations in the lower Mekong Basin,using mitochondrial DNA nad1 gene as the marker

The liver fluke, Opisthorchis viverrini, causes serious public-health problems in the Lower Mekong Basin. This study aimed to clarify whether O. viverrini populations may be genetically divided into sub-specific taxa. We collected 6 populations of O. viverrini from different places in Cambodia, Lao PDR, and Thailand, along both sides of the Mekong River, and analyzed the population structure of these using the mitochondrial nad1 gene as a marker. The results of the DNA polymorphism measurements, by theta-w (θw) and -π (θπ) values, neutrality tests, and mismatch distribution, suggested that the population of O. viverrini has expanded under the influence of purifying selection and selective sweep. The analysis of molecular variance (AMOVA) test revealed no significant genetic differences among the O. viverrini populations on opposite sides of the Mekong River. O. viverrini haplotypes occurred in multiple populations, and no distinct geographical clade. The star-like haplotype network confirmed a demographic expansion of the O. viverrini population. Overall, the genetic data from these populations suggested that the postulated existence of an O. viverrini species complex should be rejected. The bio-geographical diversity of O. viverrini populations should be explored further, using other appropriate markers and a wider range of samples from geographically different areas.     

Demography of the upward-shifting temperate woody species of the Rhododendron pseudochrysanthum complex and ecologically relevant adaptive divergence in its trailing edge populations

Trailing edge populations of upward migrating species induced by postglacial climatic warming can be evolutionarily significant in the face of global warming. We tested for population size changes between lower- and higher-elevation populations located in the same mountainous regions of the upward-shifting temperate woody species of the Rhododendron pseudochrysanthum complex in Taiwan. We also tested whether natural selection evoked adaptive divergence in trailing edge populations of this species complex. We genotyped 26 expressed sequence tag-simple sequence repeat (EST-SSR) loci of 185 individuals from nine populations of the R. pseudochrysanthum complex including Rhododendron rubropunctatum, Rhododendron hyperythrum, Rhododendron morii, and R. pseudochrysanthum. R. rubropunctatum populations in the R. pseudochrysanthum complex possessed the lowest estimates of genetic diversity and effective population size. Higher-elevation R. pseudochrysanthum populations had lower effective population sizes than lower-elevation R. morii populations in Hohuanshan and Tahsueshan, as revealed by estimates using both MIGRATE-N and approximate Bayesian computation (ABC). R. rubropunctatum populations diverged significantly from populations of other members of the R. pseudochrysanthum complex. An outlier potentially under positive selection specific to R. rubropunctatum populations was identified and strongly associated with ecologically relevant environmental variables. Postglacial climatic warming has a significant influence on population isolation in the R. pseudochrysanthum complex. The results indicate adaptive evolution in the trailing edge populations, i.e., R. rubropunctatum populations of the upward shifting R. pseudochrysanthum complex.