Genetic homogenization of the nuclear ITS loci across two morphologically distinct gentians in their overlapping distributions in the Qinghai-Tibet Plateau.
ABSTRACT: Interspecific hybridization and introgression can lead to partial genetic homogenization at certain neutral loci between morphologically distinct species and may obscure the species delimitation using nuclear genes. In this study, we investigated this phenomenon through population genetic survey of two alpine plants (Gentiana siphonantha and G. straminea) in the Qinghai-Tibet Plateau, where the distributions of two species are partly overlapped. We identified two clusters of chloroplast DNA haplotypes which correspond to the two species, and three clusters of ITS ribotypes. In addition to clusters specific to each species, the third ITS cluster, which was most likely derived from hybridization between the other two clusters and subsequent recombination and concerted evolution, was widely shared by two species in their adjacent areas. In contrast to the morphological distinctiveness of the two species, interspecific gene flow possibly led to genetic homogenization at their ITS loci. The new ITS lineage recovered for species in adjacent areas is distinctly different from original lineages found in allopatric areas. These findings may have general implications for our understanding of cryptic changes at some genetic loci caused by interspecific gene flow in the history, and they indicate that species delimitation should be based on a combination of both nuclear and chloroplast DNA sequence variations.
Project description:The genus Dioscorea is widely distributed in tropical and subtropical regions, and is economically important in terms of food supply and pharmaceutical applications. However, DNA barcodes are relatively unsuccessful in discriminating between Dioscorea species, with the highest discrimination rate (23.26%) derived from matK sequences. In this study, we compared genic and intergenic regions of three Dioscorea chloroplast genomes and found that the density of SNPs and indels in intergenic sites was about twice and seven times higher than that of SNPs and indels in the genic regions, respectively. A total of 52 primer pairs covering highly variable regions were designed and seven pairs of primers had 80%-100% PCR success rate. PCR amplicons of 73 Dioscorea individuals and assembled sequences of 47 Dioscorea SRAs were used for estimating intraspecific and interspecific divergence for the seven loci: The rpoB-trnC locus had the highest interspecific divergence. Automatic barcoding gap discovery (ABGD), Poisson tree processes (PTP), and generalized mixed Yule coalescence (GMYC) analysis were applied for species delimitation based on the seven loci and successfully identified the majority of species, except for species in the Enantiophyllum section. Phylogenetic analysis of 51 Dioscorea individuals (28 species) showed that most individuals belonging to the same species tended to cluster in the same group. Our results suggest that the variable loci derived from comparative analysis of plastid genome sequences could be good DNA barcode candidates for taxonomic analysis and species delimitation.
Project description:Genetic variations from few chloroplast DNA fragments show lower discriminatory power in the delimitation of closely related species and less resolution ability in discerning interspecific relationships than from nrITS. Here we use Orychophragmus (Brassicaceae) as a model system to test the hypothesis that the whole chloroplast genomes (plastomes), with accumulation of more variations despite the slow evolution, can overcome these weaknesses. We used Illumina sequencing technology via a reference-guided assembly to construct complete plastomes of 17 individuals from six putatively assumed species in the genus. All plastomes are highly conserved in genome structure, gene order, and orientation, and they are around 153 kb in length and contain 113 unique genes. However, nucleotide variations are quite substantial to support the delimitation of all sampled species and to resolve interspecific relationships with high statistical supports. As expected, the estimated divergences between major clades and species are lower than those estimated from nrITS probably due to the slow substitution rate of the plastomes. However, the plastome and nrITS phylogenies were contradictory in the placements of most species, thus suggesting that these species may have experienced complex non-bifurcating evolutions with incomplete lineage sorting and/or hybrid introgressions. Overall, our case study highlights the importance of using plastomes to examine species boundaries and establish an independent phylogeny to infer the speciation history of plants.
Project description:Delimitation of species is an important and controversial area within evolutionary biology. Many species boundaries have been defined using morphological data. New genetic approaches now offer more objective evaluation and assessment of the reliability of morphological variation as an indicator that speciation has occurred. We examined geographic variation in morphology of the continuously distributed skink Chalcides mionecton from Morocco and used Bayesian analyses of nuclear and mitochondrial DNA (mtDNA) loci to examine: (i) their concordance with morphological patterns, (ii) support for species delimitation, (iii) timing of speciation, and (iv) levels of gene flow between species. Four digit individuals were found at sites between Cap Rhir (in the south) and the northern extreme of the range, whereas five-digit individuals were found in two disjunct areas: (i) south of Cap Rhir and (ii) the north of the range where they were often syntopic with four-digit individuals. The pattern of variation in generalized body dimensions was largely concordant with that in digit number, suggesting two general morphotypes. Bayesian analyses of population structure showed that individuals from sites south of Cap Rhir formed one genetic cluster, but that northern four- and five-digit individuals clustered together. Statistical support for delimitation of these genetic clusters into two species was provided by a recent Bayesian method. Phylogenetic-coalescent dating with external time calibrations indicates that speciation was relatively recent, with a 95% posterior interval of 0.46-2.66 mya. This postdates equivalent phylogenetic dating estimates of sequence divergence by approximately 1 Ma. Statistical analyses of a small number of independent loci provide important insights into the history of the speciation process in C. mionecton and support delimitation of populations into two species with distributions that are spatially discordant with patterns of morphological variation.
Project description:Accurately delimiting species is fundamentally important for understanding species diversity and distributions and devising effective strategies to conserve biodiversity. However, species delimitation is problematic in many taxa, including 'non-adaptive radiations' containing morphologically cryptic lineages. Fortunately, coalescent-based species delimitation methods hold promise for objectively estimating species limits in such radiations, using multilocus genetic data. Using coalescent-based approaches, we delimit species and infer evolutionary relationships in a morphologically conserved group of Central American freshwater fishes, the Poecilia sphenops species complex. Phylogenetic analyses of multiple genetic markers (sequences of two mitochondrial DNA genes and five nuclear loci) from 10/15 species and genetic lineages recognized in the group support the P. sphenops species complex as monophyletic with respect to outgroups, with eight mitochondrial 'major-lineages' diverged by ?2% pairwise genetic distances. From general mixed Yule-coalescent models, we discovered (conservatively) 10 species within our concatenated mitochondrial DNA dataset, 9 of which were strongly supported by subsequent multilocus Bayesian species delimitation and species tree analyses. Results suggested species-level diversity is underestimated or overestimated by at least ~15% in different lineages in the complex. Nonparametric statistics and coalescent simulations indicate genealogical discordance among our gene tree results has mainly derived from interspecific hybridization in the nuclear genome. However, mitochondrial DNA show little evidence for introgression, and our species delimitation results appear robust to effects of this process. Overall, our findings support the utility of combining multiple lines of genetic evidence and broad phylogeographical sampling to discover and validate species using coalescent-based methods. Our study also highlights the importance of testing for hybridization versus incomplete lineage sorting, which aids inference of not only species limits but also evolutionary processes influencing genetic diversity.
Project description:Pityopsis ruthii (Ruth's golden aster) is a federally endangered herbaceous perennial endemic to the Hiwassee and Ocoee Rivers in southeastern Tennessee, United States. Comprehensive genetic studies providing novel information to conservationists for preservation of the species are lacking. Genetic variation and gene flow were evaluated for 814 individuals from 33 discrete locations using polymorphic microsatellites: seven chloroplast and twelve nuclear. A total of 198 alleles were detected with the nuclear loci and 79 alleles with the chloroplast loci. Gene flow was estimated, with the Hiwassee River (Nm = 2.16; FST = 0.15) showing higher levels of gene flow and lower levels of population differentiation than the Ocoee River (Nm = 1.28; FST = 0.19). Population structure was examined using Bayesian cluster analyses. Nuclear and chloroplast analyses were incongruent. From the chloroplast microsatellites, three clusters were identified; all were present in sampling sites at both rivers, indicating a lack of allele fixation along rivers. Nuclear markers revealed two clusters and separated by river. When the Hiwassee River locations were analyzed, four clusters were identified for both the chloroplast and nuclear microsatellites, though the individuals clustered differently. Analysis of the Ocoee River revealed two clusters for the chloroplast microsatellites and three for the nuclear microsatellites. We recommend P. ruthii be managed as four populations for the Hiwassee River and three populations for the Ocoee River. Our results provide critical genetic information for P. ruthii that can be used for species management decisions to drive future population augmentation/reintroduction and ex situ conservation efforts.
Project description:Accurate species delimitation and description are necessary to guide effective conservation of imperiled species, and this synergy is maximized when multiple data sources are used to delimit species. We illustrate this point by examining Drymarchon couperi (Eastern Indigo Snake), a large, federally-protected species in North America that was recently divided into two species based on gene sequence data from three loci and heuristic morphological assessment. Here, we re-evaluate the two-species hypothesis for D. couperi by evaluating both population genetic and gene sequence data. Our analyses of 14 microsatellite markers revealed 6-8 genetic population clusters with significant admixture, particularly across the contact zone between the two hypothesized species. Phylogenetic analyses of gene sequence data with maximum-likelihood methods suggested discordance between mitochondrial and nuclear markers and provided phylogenetic support for one species rather than two. For these reasons, we place Drymarchon kolpobasileus into synonymy with D. couperi. We suggest inconsistent patterns between mitochondrial and nuclear DNA are driven by high dispersal of males relative to females. We advocate for species delimitation exercises that evaluate admixture and gene flow in addition to phylogenetic analyses, particularly when the latter reveal monophyletic lineages. This is particularly important for taxa, such as squamates, that exhibit strong sex-biased dispersal. Problems associated with over-delimitation of species richness can become particularly acute for threatened and endangered species, because of high costs to conservation when taxonomy demands protection of more individual species than are supported by accumulating data.
Project description:BACKGROUND:The genus Ostrya (Betulaceae) contains eight species and four of them are distributed in China. However, studies based on limited informative sites of several chloroplast markers failed to resolve interspecific delimitation and relationships among the four Chinese species. In this study, we aimed to use the whole chloroplast genomes to address these two issues. RESULTS:We assembled and annotated 33 complete chloroplast genomes (plastomes) of the four Chinese species, representing 17 populations across most of their geographical distributions. Each species contained samples of several individuals that cover most of geographic distributions of the species. All plastomes are highly conserved in genome structure and gene order, with a total length of 158-159?kb and 122 genes. Phylogenetic analyses of whole plastomes, non-coding regions and protein-coding genes produced almost the same topological relationships. In contrast to the well-delimitated species boundary inferred from the nuclear ITS sequence variations, three of the four species are non-monophyletic in the plastome trees, which is consistent with previous studies based on a few chloroplast markers. CONCLUSIONS:The high incongruence between the ITS and plastome trees may suggest the widespread occurrences of hybrid introgression and incomplete lineage sorting during the divergence of these species. In addition, the plastomes with more informative sites compared with a few chloroplast markers still failed to resolve the phylogenetic relationships of the four species, and further studies involving population genomic data may be needed to better understand their evolutionary histories.
Project description:Although Aspergillus fumigatus is the major agent of invasive aspergillosis, an increasing number of infections are caused by its cryptic species, especially A. lentulus and the A. viridinutans species complex (AVSC). Their identification is clinically relevant because of antifungal drug resistance and refractory infections. Species boundaries in the AVSC are unresolved since most species have uniform morphology and produce interspecific hybrids in vitro. Clinical and environmental strains from six continents (n = 110) were characterized by DNA sequencing of four to six loci. Biological compatibilities were tested within and between major phylogenetic clades, and ascospore morphology was characterised. Species delimitation methods based on the multispecies coalescent model (MSC) supported recognition of ten species including one new species. Four species are confirmed opportunistic pathogens; A. udagawae followed by A. felis and A. pseudoviridinutans are known from opportunistic human infections, while A. felis followed by A. udagawae and A. wyomingensis are agents of feline sino-orbital aspergillosis. Recently described human-pathogenic species A. parafelis and A. pseudofelis are synonymized with A. felis and an epitype is designated for A. udagawae. Intraspecific mating assay showed that only a few of the heterothallic species can readily generate sexual morphs in vitro. Interspecific mating assays revealed that five different species combinations were biologically compatible. Hybrid ascospores had atypical surface ornamentation and significantly different dimensions compared to parental species. This suggests that species limits in the AVSC are maintained by both pre- and post-zygotic barriers and these species display a great potential for rapid adaptation and modulation of virulence. This study highlights that a sufficient number of strains representing genetic diversity within a species is essential for meaningful species boundaries delimitation in cryptic species complexes. MSC-based delimitation methods are robust and suitable tools for evaluation of boundaries between these species.
Project description:<h4>Background</h4>Populus is an ecologically and economically important genus of trees, but distinguishing between wild species is relatively difficult due to extensive interspecific hybridization and introgression, and the high level of intraspecific morphological variation. The DNA barcoding approach is a potential solution to this problem.<h4>Methodology/principal findings</h4>Here, we tested the discrimination power of five chloroplast barcodes and one nuclear barcode (ITS) among 95 trees that represent 21 Populus species from western China. Among all single barcode candidates, the discrimination power is highest for the nuclear ITS, progressively lower for chloroplast barcodes matK (M), trnG-psbK (G) and psbK-psbI (P), and trnH-psbA (H) and rbcL (R); the discrimination efficiency of the nuclear ITS (I) is also higher than any two-, three-, or even the five-locus combination of chloroplast barcodes. Among the five combinations of a single chloroplast barcode plus the nuclear ITS, H+I and P+I differentiated the highest and lowest portion of species, respectively. The highest discrimination rate for the barcodes or barcode combinations examined here is 55.0% (H+I), and usually discrimination failures occurred among species from sympatric or parapatric areas.<h4>Conclusions/significance</h4>In this case study, we showed that when discriminating Populus species from western China, the nuclear ITS region represents a more promising barcode than any maternally inherited chloroplast region or combination of chloroplast regions. Meanwhile, combining the ITS region with chloroplast regions may improve the barcoding success rate and assist in detecting recent interspecific hybridizations. Failure to discriminate among several groups of Populus species from sympatric or parapatric areas may have been the result of incomplete lineage sorting, frequent interspecific hybridizations and introgressions. We agree with a previous proposal for constructing a tiered barcoding system in plants, especially for taxonomic groups that have complex evolutionary histories (e.g. Populus).
Project description:The use of subtle features as species diagnostic traits in taxa with high morphological similarity sometimes fails in discriminating intraspecific variation from interspecific differences, leading to an incorrect species delimitation. A clear assessment of species boundaries is particularly relevant in disease vector organisms in order to understand epidemiological and evolutionary processes that affect transmission capacity. Here, we assess the validity of the recently described Rhodnius taquarussuensis (da Rosa et al., 2017) using interspecific crosses and molecular markers. We did not detect differences in hatching rates in interspecific crosses between R. taquarussuensis and R. neglectus (Lent, 1954). Furthermore, genetic divergence and species delimitation analyses show that R. taquarussuensis is not an independent lineage in the R. prolixus group. These results suggest that R. taquarussuensis is a phenotypic form of R. neglectus instead of a distinct species. We would like to stress that different sources of evidence are needed to correctly delimit species. We consider this is an important step in understanding vectorial Chagas disease spread and transmission.