The systematic position of Phaeocryptopus gaeumannii

Phaeocryptopus gaeumannii, causal agent of the Douglas-fir foliar disease Swiss needle cast, is the only known pathogenic species of the genus. Current classifications place Phaeocryptopus in the Venturiaceae (Pleosporales), typified by the apple-scab pathogen Venturia inaequalis. All core members of this family have hyphomycetous anamorphs. We sought to confirm these relationships by means of phylogenetic analyses of the small (SSU) and large (LSU) subunits and internal transcribed spacer (ITS) region of nuclear ribosomal gene sequences (nrDNA). Analyses indicated that both the genus Phaeocryptopus and the family Venturiaceae, as currently defined, are unnatural groups. Phaeocryptopus nudus, type of the genus, is aligned in the Dothioraceae (Dothideales) and P. gaeumannii in the Mycosphaerellaceae (Capnodiales) near species of Mycosphaerella and Rasutoria. Core representatives of Venturiaceae formed an unambiguous clade but ordinal placement was unresolved. The family apparently is not included in the Pleosporales, Dothideales, Myriangiales or Capnodiales. Coelomycetous Rhizosphaera form-species are accepted generally as anamorphic states of Phaeocryptopus, however the relationship never has been established conclusively. Species of Rhizosphaera are closely related to P. nudus but not to P. gaeumannii, supporting an anamorph-teleomorph connection between Rhizosphaera and Phaeocryptopus and providing further evidence that P. gaeumanii is not congeneric with P. nudus.     

Population structure and dynamics of Magnaporthe grisea in the Indian Himalayas.

The population genetics of Magnaporthe grisea, the rice blast pathogen, were analyzed in a center of rice diversity (the Uttar Pradesh hills of the Indian Himalayas) using multilocus and single-, or low-copy, DNA markers. Based on DNA fingerprinting with the multilocus probe MGR586 and single-locus probes, 157 haplotypes clustered into 56 lineages (at >/=70% MGR586 band similarity, each with unique single-locus profiles) and high diversity indices were detected among 458 isolates collected from 29 sites during 1992-1995. Most valleys sampled had distinct populations (73% of the lineages were site specific) with some containing one or a few lineages, confirming the importance of clonal propagation, and others were very diverse. Widely distributed lineages suggested that migration occurs across the region and into the Indo-Gangetic plains. Repeated sampling at one site, Matli, (170 isolates, 1992-1995) yielded 19 lineages and diversity significantly greater than that reported from similar samples from Colombia and the Philippines. Analysis of allelic associations using pairwise comparisons and multilocus variance analysis failed to reject the hypothesis of gametic phase equilibrium. The Matli population shifted from highly diverse in 1992 to almost complete dominance by one lineage in 1995. Such population dynamics are consistent with recombination followed by differential survival of clonal descendants of recombinant progeny. At another site, Ranichauri, population (n = 84) composition changed from 2 to 11 lineages over 2 yr and yielded additional evidence for equilibrium. Sexually fertile and hermaphrodite isolates of both mating types were recovered from rice in both Matli and Ranichauri. We demonstrate that Himalayan M. grisea populations are diverse and dynamic and conclude that the structure of some populations may be affected to some extent by sexual recombination.     

Population structure of and mycotoxin production by Fusarium graminearum from maize in South Korea

Fusarium graminearum (Gibberella zeae) is an important pathogen of wheat, maize, barley, and rice in South Korea, and harvested grain often is contaminated with trichothecenes such as deoxynivalenol and nivalenol. In this study, we examined 568 isolates of F. graminearum collected from maize at eight locations in South Korea. We used amplified fragment length polymorphisms (AFLPs) to identify four lineages (2, 3, 6, and 7); lineage 7 was the most common (75%), followed by lineage 6 (12%), lineage 3 (12%), and lineage 2 (1%). The genetic identity among populations was high (>0.98), and the effective migration rate between locations was higher than that between lineages. Female fertility varied by lineage: all lineage 7 isolates were fertile, while 70%, 26%, and 14% of the isolates in lineages 6, 3, and 2, respectively, were fertile. All lineage 3 and lineage 7 isolates produced deoxynivalenol, whereas most lineage 2 and 6 isolates produced nivalenol. Genotypic diversity in lineage 3 and lineage 6 populations is similar to that found in previously described Korean rice populations, but genotypic diversity in lineage 7 is much lower, even though similar levels of gene flow occur between lineage 7 populations. We conclude that lineage 7 was relatively recently introduced into South Korea, perhaps accompanying imported maize seeds.     

Neither connectivity nor genetic diversity matter in the conservation of a rare fern and a moss on insular erratic boulders

Erratic boulders provide habitat for rock-dwelling species and contribute to the biodiversity of landscapes. In the calcareous Swiss lowlands, siliceous erratic boulders are exclusive habitat islands for the regionally critically endangered fern Asplenium septentrionale, about 20 bryophyte species and numerous lichens. Focusing on island biogeographical processes, we analysed the conservation genomics of A. septentrionale and the moss Hedwigia ciliata on insular erratic boulders in the Swiss lowlands and the adjacent “mainland” in siliceous mountains. We genotyped both species using double digest restriction associated DNA sequencing (ddRAD). For the tetraploid A. septentrionale, abundant identical multilocus genotypes within populations suggested prevalent intragametophytic selfing, and six out of eight boulder populations consisting of a single multilocus genotype each indicated single spore founder events. The genetic structure of A. septentrionale mainland populations coincided with Pleistocene glacial refugia. Four genetic lineages of H. ciliata were identified, and populations consisting of a single multilocus genotype were less common than in A. septentrionale. For both taxa, multilocus genotype diversity on boulders was lower than in mainland populations. The absence of common genetic groups among boulder populations, and the absence of isolation by distance patterns, suggested colonisation of boulders through independent long-distance dispersal events. Successful boulder colonisation of A. septentrionale seems to be rare, while colonisation by H. ciliata appears to be more frequent. We conclude that pivotal principles of conservation biology, such as connectivity and genetic diversity, are of less importance for the studied cryptogams on insular erratic boulders because of long-distance dispersal, intragametophytic selfing and polyploidy.

     

Characterisation of large and small subunit rRNA and mini-exon genes further supports the distinction of six Trypanosoma cruzi lineages

It has been proposed that isolates of Trypanosoma cruzi, the agent of American trypanosomiasis, can be ordered into two primary phylogenetic lineages, first based on multilocus enzyme electrophoresis and random amplified polymorphic DNA, and subsequently based on the 24Salpha rRNA and mini-exon genes. Recent multilocus enzyme electrophoresis and random amplified polymorphic DNA data have additionally shown that the major multilocus enzyme electrophoresis/random amplified polymorphic DNA lineage II is further subdivided into five smaller lineages, designated IIa-IIe. In this study, the precise correspondence between the multilocus enzyme electrophoresis/random amplified polymorphic DNA and rRNA/mini-exon lineages was investigated. Using the 24Salpha rRNA and mini-exon markers in combination, five sets of strains were distinguished, corresponding to the multilocus enzyme electrophoresis/random amplified polymorphic DNA lineages I, IIa, IIc, IId and to lineages IIb/IIe together, respectively. The previous categorisation into only two primary lineages based on 24Salpha rRNA and mini-exon characterisation is explained, in part, by the lack of representativeness of the breadth of T. cruzi diversity in earlier study samples. Additionally, a PCR assay based on a length-variable region of the 18S rRNA gene distinguished lineage IIe from lineage IIb. Thus, the six multilocus enzyme electrophoresis/random amplified polymorphic DNA lineages could be readily identified by combining data from the 24Salpha rRNA, mini-exon and 18S rRNA characterisation assays, further supporting the relevance of these genetic units for T. cruzi strain classification and subspecific nomenclature. The recently proposed groups T. cruzi I and T. cruzi II correspond to multilocus enzyme electrophoresis/random amplified polymorphic DNA lineages I and IIb, respectively. Our findings show that T. cruzi lineage characterisation based on a single marker (either mini-exon or 24Salpha rRNA) has insufficient resolution, and leads to important reinterpretations of recent epidemiological and evolutionary studies based on the oversimplified rRNA/mini-exon dichotomic classification of T. cruzi isolates.     

Range-wide chloroplast and mitochondrial DNA imprints reveal multiple lineages and complex biogeographic history for Douglas-fir

The contemporary genetic structure of species offers key imprints of how organisms responded to past geological and climatic events, which have played a crucial role in shaping the current geographical distribution of north-temperate organisms. In this study, range-wide patterns of genetic variation were examined in Douglas-fir (Pseudotsuga menziesii), a dominant forest tree species distributed from Mexico to British Columbia in western North America. Two organelle DNA markers with contrasting modes of inheritance were genotyped for 613 individuals from 44 populations. Two mitotypes and 42 chlorotypes were recovered in this survey. Both genomes showed significant population subdivision, indicative of limited gene flow through seeds and pollen. Three distinct cpDNA lineages corresponding to the Pacific Coast, the Rocky Mountains, and Mexico were observed. The split time of the two lineages from the Rockies lineage was dated back to 8.5 million years (Ma). The most recent common ancestors of Mexican and coastal populations were estimated at 3.2 and 4.8 Ma, respectively. The northern populations of once glaciated regions were characterized by a high level of genetic diversity, indicating a large zone of contact between ancestral lineages. A possible northern refugium was also inferred. The Mexican lineage, which appeared established by southward migration from the Rockies lineage, was characterized by the lowest genetic diversity but highest population differentiation. These results suggest that the effects of Quaternary climatic oscillations on the population dynamics and genetic diversity of Douglas-fir varied substantially across the latitudinal section. The results emphasize the pressing need for the conservation of Mexican Douglas-fir.     

Genetic Variation within Clonal Lineages of Phytophthora infestans Revealed through Genotyping-By-Sequencing,and Implications for Late Blight Epidemiology

Genotyping-by-sequencing (GBS) was performed on 257 Phytophthora infestans isolates belonging to four clonal lineages to study within-lineage diversity. The four lineages used in the study were US-8 (n = 28), US-11 (n = 27), US-23 (n = 166), and US-24 (n = 36), with isolates originating from 23 of the United States and Ontario, Canada. The majority of isolates were collected between 2010 and 2014 (94%), with the remaining isolates collected from 1994 to 2009, and 2015. Between 3,774 and 5,070 single-nucleotide polymorphisms (SNPs) were identified within each lineage and were used to investigate relationships among individuals. K-means hierarchical clustering revealed three clusters within lineage US-23, with US-23 isolates clustering more by collection year than by geographic origin. K-means hierarchical clustering did not reveal significant clustering within the smaller US-8, US-11, and US-24 data sets. Neighbor-joining (NJ) trees were also constructed for each lineage. All four NJ trees revealed evidence for pathogen dispersal and overwintering within regions, as well as long-distance pathogen transport across regions. In the US-23 NJ tree, grouping by year was more prominent than grouping by region, which indicates the importance of long-distance pathogen transport as a source of initial late blight inoculum. Our results support previous studies that found significant genetic diversity within clonal lineages of P. infestans and show that GBS offers sufficiently high resolution to detect sub-structuring within clonal populations.     

NONRANDOM GENOTYPIC ASSOCIATIONS IN A LEGUME—BRADYRHIZOBIUM MUTUALISM

Genetically divergent lineages often coexist within populations of the annual legume Amphicarpaea bracteata. At one site dominated by two such lineages (termed biotypes “C” and “S”), isolates of root-nodule bacteria (Bradyrhizobium sp.) were sampled from both hosts and analyzed by enzyme electrophoresis. Symbiont populations on the two plant biotypes were highly distinct. Out of 15 bacterial multilocus genotypes detected (among 51 isolates analyzed), only one was shared in common by the two plant biotypes. Cluster analysis revealed three bacterial lineages (designated I, II, and III), with lineage I found exclusively on biotype C plants, and the two other lineages almost completely restricted to biotype S hosts. Laboratory inoculation tests indicated that lineage I bacteria were strictly specialized on biotype C hosts, forming few or no nodules on plants of the other host biotype. Bacterial lineages II and III were capable of forming nodules on both kinds of plants, but nodule numbers were often significantly higher on biotype S hosts. The nonrandom association between plant and bacterial lineages at this site implies that genetic diversity of hosts is an important factor in the maintenance of polymorphism within the symbiont population.     

Evidence for the dispersal of a unique lineage from Asia to America and Africa in the sugarcane fungal pathogen Ustilago scitaminea

The basidiomycete Ustilago scitaminea Sydow, which causes sugarcane smut disease, has been spreading throughout Africa and America since the 1940s. The genetic diversity and structure of different populations of this fungus worldwide was investigated using microsatellites. A total of 142 single-teliospore were isolated from 77 distinct whips (sori) collected in 15 countries worldwide. Mycelium culture derived from on generation of selfing of these single teliospores were analysed for their polymorphisms at 17 microsatellite loci. All these strains but one were homozygous at all loci, indicating that selfing is likely the predominant reproductive mode of U. scitaminea. The genetic diversity of either American or African U. scitaminea populations was found to be extremely low and all strains belong to a single lineage. This lineage was also found in some populations of Asia, where most U. scitaminea genetic diversity was detected, suggesting that this fungal species originated from this region. The strong founder effect observed in U. scitaminea African and American populations suggests that the fungus migrated from Asia to other continents on rare occasions through movement of infected plant material.     

Population structure and diversity in sexual and asexual populations of the pathogenic fungus Melampsora lini

Many pathogens undergo both sexual and asexual reproduction to varying degrees, yet the ecological, genetic and evolutionary consequences of different reproductive strategies remain poorly understood. Here we investigate the population genetic structure of wild populations of the plant pathogen Melampsora lini on its host Linum marginale , using amplified fragment length polymorphism (AFLP) markers, two genes underlying pathogen virulence, and phenotypic variation in virulence. In Australia, M. lini occurs as two genetically and geographically divergent lineages (AA and AB), one of which is completely asexual (AB), and the other able to reproduce both clonally and sexually (AA). To quantify the genetic and evolutionary consequences of these different life histories, we sampled five populations in each of two biogeographical regions. Analysis of AFLP data obtained for 275 isolates revealed largely disjunct geographical distributions for the two different lineages, low genetic diversity within lineages, and strong genetic structure among populations within each region. We also detected significant divergence among populations for both Avr genes and virulence phenotypes, although generally these values were lower than those obtained with AFLP markers. Furthermore, isolates belonging to lineage AA collectively harboured significantly higher genotypic and phenotypic diversity than lineage AB isolates. Together these results illustrate the important roles of reproductive modes and geographical structure in the generation and maintenance of virulence diversity in populations of M. lini .     

Reticulated and epidemic population genetic structure of Rhizobium etli biovar phaseoli in a traditionally managed locality in Mexico

We conducted a multilocus enzyme electrophoresis (MLEE) study to assess the genetic structure of the nitrogen-fixing bacteria Rhizobium etli bv. phaseoli . We analysed the genetic variation at 10 enzyme-encoding chromosomal loci of 482 isolates from root nodules of Phaseolus vulgaris and P. coccineus bean plants. The isolates were obtained from six traditionally managed agricultural plots in two localities in the State of Puebla, in Central Mexico. The total mean genetic diversity ( H _E) for the six plots was 0.531. Among the 482 isolates collected, 126 distinctive multilocus genotypes (electrophoretic types [Ets]) were obtained, and approximately half of the isolates are represented by five widespread ETs. A significant degree of genetic differentiation among the six plots ( G _ST = 0.072) and between the two localities ( G _ST = 0.022) was detected. The main part of the observed variability (70%) was found among the isolates within the plants. The cluster analysis revealed two deeply diverging lineages, separated at a genetic distance of 0.7. When a multilocus linkage disequilibrium analysis was performed at different hierarchical levels, we found significant linkage disequilibrium, but when the analysis was performed for the genotypes within the two diverging lineages, we found evidence of recombination. We propose for R. etli bv. phaseoli a reticulated and epidemic genetic structure, in which few genotypes increase in frequency to produce numerically dominant clones, and genetic exchange occurs mainly among genotypes within each lineage.     

Polymorphic microsatellite markers for the Douglas‐fir pathogen Phaeocryptopus gaeumannii,causal agent of Swiss Needle Cast disease

Ten polymorphic microsatellite markers were isolated from Phaeocryptopus gaeumannii, causal agent of the Douglas‐fir foliage disease Swiss Needle Cast. The primer sets were tested on 60 isolates that had, with more conservative markers, previously segregated into three reproductively isolated lineages comprising nine genotypes. The microsatellite data yielded 46 multilocus genotypes that segregated into three groups identical to the three lineages previously recovered. Microsatellite genotypic diversity ranged from 0.85 to 0.99 within lineages and was lowest where both pathogen and host are exotic.     

The demography and population genomics of evolutionary transitions to self-fertilization in plants

The evolution of self-fertilization from outcrossing has occurred on numerous occasions in flowering plants. This shift in mating system profoundly influences the morphology, ecology, genetics and evolution of selfing lineages. As a result, there has been sustained interest in understanding the mechanisms driving the evolution of selfing and its environmental context. Recently, patterns of molecular variation have been used to make inferences about the selective mechanisms associated with mating system transitions. However, these inferences can be complicated by the action of linked selection following the transition. Here, using multilocus simulations and comparative molecular data from related selfers and outcrossers, we demonstrate that there is little evidence for strong bottlenecks associated with initial transitions to selfing, and our simulation results cast doubt on whether it is possible to infer the role of bottlenecks associated with reproductive assurance in the evolution of selfing. They indicate that the effects of background selection on the loss of diversity and efficacy of selection occur rapidly following the shift to high selfing. Future comparative studies that integrate explicit ecological and genomic details are necessary for quantifying the independent and joint effects of selection and demography on transitions to selfing and the loss of genetic diversity.     

Molecular diversity of native bradyrhizobia isolated from lima bean (Phaseolus lunatus L.) in Peru

The diversity of a collection of 21 bradyrhizobial isolates from Lima bean (Phaseolus lunatus L.) was assayed by molecular methods. Moderately high to high genetic diversity was revealed by multilocus enzyme electrophoresis (MLEE) analysis of seven enzyme loci and genomic fingerprints with ERIC and BOX primers. Two groups with differences in growth rate were found among the isolates and their differentiation as two divergent bradyrhizobial lineages was supported by PCR-RFLP of the rpoB gene and sequence analysis of the 16S rDNA and dnaK genes. Isolates with slow growth (SG) were identified as Bradyrhizobium yuanmingense, while extra-slow growing isolates (ESG) constitute a new lineage different from all described Bradyrhizobium species. Three distinct symbiotic genotypes were detected among Lima bean bradyrhizobia by PCR-RFLP and sequence analysis of the nifH and nodB genes. One genotype was found in the ESG lineage and two in B. yuanmingense. Another symbiotic genotype was detected in B. yuamingense isolated from Lespedeza plants. The identified bradyrhizobial lineages constitute sympatric species effectively nodulating Lima bean on the coast of Peru.     

Putative origin of clonal lineages of Amylostereum areolatum, the fungal symbiont associated with Sirex noctilio, retrieved from Pinus sylvestris, in eastern Canada

The Eurasian Sirex noctilio-Amylostereum areolatum complex was discovered and has become established close to the North American Great Lakes in the 2000s. This invasive forest insect pest represents a very high risk to native and exotic pines in North America. We investigated the geographical origin of clonal lineages of the fungal symbiont A. areolatum in the recently pest-colonized eastern Canadian region by analyzing mitochondrial and nuclear sequence variations and comparing the genetic diversity of a worldwide collection of fungal symbionts among six countries where the Sirex complex is native and four countries from which the insect-fungal complex has been introduced. In total, 102 isolates were analyzed. While 12 multilocus genotypes (MLGs) are observed in the areas where S. noctilio is native, only two MLGs are retrieved from areas where S. noctilio is not native, indicating the wide spread of clonal lineages in the introduced fungal symbiont of S. noctilio. MLG2 comprises 26% of the Canadian isolates and is also observed in Chile and South Africa, where the insect-fungal complex has also been introduced. MLG3 comprises 74% of the Canadian isolates and is also observed in the USA, but nowhere else in our worldwide collection. Thus, at least one of the Canadian clonal lineages shares a common origin with A.?areolatum isolates from the Southern Hemisphere. The source of the second clonal lineage is still unknown, but phylogenetic analyses show that MLG3 is isolated. More extended sampling is necessary to determine the origin of this fungal clonal lineage and investigate its probable symbiotic association with native North American Sirex.     

Genotypic Analysis of Russian Isolates of Phytophthora infestans from the Moscow Region, Siberia and Far East

Phytophthora infestans samples were collected during 1997 and 1998 at multiple sites in Russia from Sakhalin Island in the Far East across Siberia (nine sites, 160 isolates) to the Moscow region (four sites, 325 isolates). In addition, 12 isolates that were obtained previously were included. All isolates were analysed for mating type, and sensitivity to metalaxyl. Isolates from within any of the nine sites outside of the Moscow region were monomorphic for mating type and nearly monomorphic for metalaxyl resistance. In contrast, both A1 and A2 isolates were detected in the Moscow region, and these isolates were also polymorphic for metalaxyl resistance. In two sites in Siberia only A2 mating type strains were detected, in the other six sites in Siberia and in Sakahlin Island, only A1 mating types were detected. A subset of isolates ( n =191) was also analysed for pathotype (virulence to 10 R-genes, each in a distinct differential genotype). All isolates were highly complex (mean number of virulences approximately 8.4 of a maximum number of 10). All isolates ( n =43) from Sakahlin Island were virulent to all 10 of the R-genes tested. A further subset of isolates ( n =70, including 12 isolates collected before 1997) was analysed for genotype at the Glucose- 6-phosphate isomerase and Peptidase loci, mtDNA haplotypes, and RFLP pattern using the RG57 probe. The US-1 clonal lineage (previously dominant) was not detected in the 1997–98 sample. The populations of P. infestans near Moscow in 1997 and 1998 was highly diverse with 15 unique genotypes (including both mating types) among a sample of 18 isolates. In contrast, the populations of P. infestans in Siberia had limited diversity, with only three multilocus genotypes detected and most populations were dominated by the SIB-1 clonal lineage. This lineage accounted for 31 of the 39 strains collected in Siberia that were assayed for multilocus genotype.     

Inferring a population structure for Staphylococcus epidermidis from multilocus sequence typing data

Despite its importance as a human pathogen, information on population structure and global epidemiology of Staphylococcus epidermidis is scarce and the relative importance of the mechanisms contributing to clonal diversification is unknown. In this study, we addressed these issues by analyzing a representative collection of S. epidermidis isolates from diverse geographic and clinical origins using multilocus sequence typing (MLST). Additionally, we characterized the mobile element (SCCmec) carrying the genetic determinant of methicillin resistance. The 217 S. epidermidis isolates from our collection were split by MLST into 74 types, suggesting a high level of genetic diversity. Analysis of MLST data using the eBURST algorithm revealed the existence of nine epidemic clonal lineages that were disseminated worldwide. One single clonal lineage (clonal complex 2) comprised 74% of the isolates, whereas the remaining isolates were clustered into 8 minor clonal lineages and 13 singletons. According to our evolutionary model, SCCmec was acquired at least 56 times by S. epidermidis. Although geographic dissemination of S. epidermidis strains and the value of the index of association between the alleles, 0.2898 (P < 0.05), support the clonality of S. epidermidis species, examination of the sequence changes at MLST loci during clonal diversification showed that recombination gives rise to new alleles approximately twice as frequently as point mutations. We suggest that S. epidermidis has a population with an epidemic structure, in which nine clones have emerged upon a recombining background and evolved quickly through frequent transfer of genetic mobile elements, including SCCmec.     

Polyphasic analysis of Thermus isolates from geothermal areas in Iceland

Genetic relationships and diversity of 101 Thermus isolates from different geothermal regions in Iceland were investigated by using multilocus enzyme electrophoresis (MLEE) and small subunit ribosomal rRNA (SSU rRNA) sequence analysis. Ten polymorphic enzymes were used and seven distinct and genetically highly divergent lineages of Thermus were observed. Six of seven lineages could be assigned to species whose names have been validated. The most diverse lineage was Thermus scotoductus. In contrast to the other lineages, this lineage was divided into very distinct genetic sublineages that may represent subspecies with different habitat preferences. The least diverse lineage was Thermus brockianus. Phenotypic and physiological analysis was carried out on a subset of the isolates. No relationship was found between growth on specific single carbon source to the grouping obtained by the isoenzyme analysis. The response to various salts was distinguishing in a few cases. No relationship was found between temperature at the isolation site and the different lineages, but pH indicated a relation to specific lineages.     

Mitochondrial DNA phylogeography of the three-spined stickleback (Gasterosteus aculeatus) in Europe-evidence for multiple glacial refugia

An analysis of mitochondrial DNA sequence variation in 172 three-spined sticklebacks (Gasterosteus aculeatus) sampled across the European distribution range revealed three major evolutionary lineages occupying relatively large and separate geographic areas. The trans-Atlantic lineage comprised of populations spanning from the East Coast of USA to the continental Europe and was basal group to the other European lineages in the phylogeny. The European lineage included populations located in the Western and Eastern Europe, British Isles, Scandinavia as well as some parts of the Mediterranean region. The third lineage was specific to the Black Sea drainages. The within lineage structure was characterized by significant excess of low frequency haplotypes and star-like mtDNA genealogies, which suggest a recent population expansions to the formerly glaciated marine and freshwater environments. A coalescent-based method dated the splits between the major lineages to have occurred during the Saalian and Weichselian glaciations in the late Pleistocene, depending on the molecular clock calibration. The coalescent simulations further indicate high degree of genetic diversity within the lineages and a substantial increase in the genetic diversity in the European lineage relative to the ancestral level. In addition to the three major lineages, the freshwater populations in R. Neretva and L. Skadar in the Adriatic Sea coast region harboured unique and highly divergent haplotypes suggesting long independent histories of these populations. Evidence from mtDNA analyses suggests that these populations deserve a status of an evolutionary significant unit.     

Genetic evidence for a high diversity and wide distribution of endemic strains of the pathogenic chytrid fungus Batrachochytrium dendrobatidis in wild Asian amphibians

Population declines and extinctions of amphibians have been attributed to the chytrid fungus Batrachochytrium dendrobatidis (Bd), especially one globally emerging recombinant lineage (‘Bd‐GPL’). We used PCR assays that target the ribosomal internal transcribed spacer region (ITS) of Bd to determine the prevalence and genetic diversity of Bd in South Korea, where Bd is widely distributed but is not known to cause morbidity or mortality in wild populations. We isolated Korean Bd strains from native amphibians with low infection loads and compared them to known worldwide Bd strains using 19 polymorphic SNP and microsatellite loci. Bd prevalence ranged between 12.5 and 48.0%, in 11 of 17 native Korean species, and 24.7% in the introduced bullfrog Lithobates catesbeianus. Based on ITS sequence variation, 47 of the 50 identified Korean haplotypes formed a group closely associated with a native Brazilian Bd lineage, separated from the Bd‐GPL lineage. However, multilocus genotyping of three Korean Bd isolates revealed strong divergence from both Bd‐GPL and the native Brazilian Bd lineages. Thus, the ITS region resolves genotypes that diverge from Bd‐GPL but otherwise generates ambiguous phylogenies. Our results point to the presence of highly diversified endemic strains of Bd across Asian amphibian species. The rarity of Bd‐GPL‐associated haplotypes suggests that either this lineage was introduced into Korea only recently or Bd‐GPL has been outcompeted by native Bd strains. Our results highlight the need to consider possible complex interactions among native Bd lineages, Bd‐GPL and their associated amphibian hosts when assessing the spread and impact of Bd‐GPL on worldwide amphibian populations.