Median-joining Haplotype Network Research Articles

Population genetics of Babesia vogeli based on the mitochondrial cytochrome b gene

The current study aimed at population genetic characterization of B. vogeli based on the cytochrome b (cyt b) gene sequences (≥ 685 bp) available in the GenBank. Phylogenetic trees placed all the sequences of B. vogeli in a single large monophyletic clade; however, it was further divided into two subclades (Bv1 and Bv2). Out of seven nucleotide variations observed between Bv1 and Bv2 subclades, four were synonymous (G92A, C170T, T488C and A659G), and three were non-synonymous (G324A, C438A and G465A) resulting in amino acid substitutions at three places (V108I, L146I and V155I). Within different B. vogeli populations, the nucleotide and haplotype diversities were low. The median-joining haplotype network revealed only two haplotypes (Hap_1 and Hap_2). A geographical sub-structuring was noticed in the B. vogeli populations, with moderate genetic differentiation (FST = 0.05000; P < 0.05) and a very high gene flow (Nm = 4.75) between Indian and Chinese populations. Neutrality tests and mismatch distributions for the Indian population and the overall dataset of B. vogeli indicated a constant population size. This study provides the first insight into the genetic characterization, population genetics and haplotype network of B. vogeli based on the cyt b gene.

First Insight into the Phylogenetic Diversity of Bovicola caprae Infesting Goats of Different Agro-climatic Locations in India.

Bovicola caprae is an important obligate ectoparasite of goats worldwide including India. The present study aimed at the molecular confirmation, phylogenetics and population structure analyses of B. caprae infesting goats of three different agro-climatic locations in India, by targeting the mitochondrial cytochrome C oxidase subunit 1 (cox1) genetic marker. The phylogenetic tree exhibited the presence of two different lineages of B. caprae. The sequences generated herein clustered in lineage 2 along with theGenBank™ archived sequences from China and Iran. The sequences generated herein also showed the circulation of sub-lineages of B. caprae in India based on the analysis of pairwise genetic distances between sequences and median-joining haplotype network. The population structure analyses revealed low nucleotide (0.00353 ± 0.00291 and 0.02694 ± 0.00363) and high haplotype (0.667 ± 0.314 and 0.618 ± 0.104) diversities for the present study isolates as well as for the complete dataset, respectively, which evinced a recent demographic expansion. High genetic differentiation (FST value = 0.97826) and low gene flow (Nm = 0.00556) were also recorded in the different lineages/populations. In conclusion, the present study addressed the research gap and provided the first insight into the phylogenetics of the goat louse B. caprae and highlighted the circulation of sub-lineages of the ectoparasite in India.

Phylogenetic Diversity, Host Specificity, and Distribution of the Wood-Decaying Fungus Phellinotus teixeirae in Western Colombia’s Seasonally Dry Tropical Forest

Phellinotus (Polyporales) is a common genus of wood-decay fungi in tropical and subtropical areas, endemic to the Seasonally Dry Tropical Forest (SDTF) biome. However, Phellinotus diversity remains unexplored, despite being a major threat to living trees. Therefore, this study is aimed at confirming and characterizing through morphological and molecular data the first isolates of Phellinotus teixeirae in Pithecellobium dulce (Fabaceae) trees (locally referred to as ‘Chiminango’) from the endangered Colombian SDTF biome. Fifteen fungal specimens were recovered from living P. dulce trees, in the urban area and at the Universidad del Valle campus, and classified as P. teixeirae based on taxonomical descriptors. Phylogenetic relationships were inferred from a four-loci dataset (ribosomal and gene-coding regions), including 82 taxa covering 3991 nucleotide positions. The analysis recovered seven highly supported (&gt;90% bootstrapping) monophyletic taxa of the ‘Phellinotus Clade’, and confirmed the new distribution range of P. teixeirae (100% bootstrap support), which extends approx. 1000 km north in the Neotropics. Hierarchical stratified Analysis of MOlecular VAriance (AMOVA) provided a clear genetic distinction between species (70% of variation, p-value = 0.001) and low differentiation among country of origin within species (11%, p-value = 0.044). Discriminant Analysis for Principal Components (DAPC) indicated complex clustering including closely related species, probably a signal of recent radiation and weak species boundaries. Median-joining haplotype network analysis identified unique haplotypes, which may correlate with new host colonization and population expansion (Tajima’s D ≤ −0.5). In conclusion, this study provides the first assessment of the genetic diversity of P. teixeirae in a novel geography (SDTP) and host tree (P. dulce). However, increasing the number of isolates remains critical to understand further the genus’ distribution patterns and drivers of genetic diversity.

Phylogenetic, phylogeographic and divergence time analysis of Anopheles subpictus species complex using ITS2 and COI sequences

Objective: To address the phylogenetic and phylogeographic relationship between different lineages of Anopheles (An.) subpictus species complex in most parts of the Asian continent by maximum utilization of Internal Transcriber Spacer 2 (ITS2) and cytochrome C oxidase I (COI) sequences deposited at the GenBank. Methods: Seventy-five ITS2, 210 COI and 26 concatenated sequences available in the NCBI database were used. Phylogenetic analysis was performed using Bayesian likelihood trees, whereas median-joining haplotype networks and time-scale divergence trees were generated for phylogeographic analysis. Genetic diversity indices and genetic differentiation were also calculated. Results: Two genetically divergent molecular forms of An. subpictus species complex corresponding to sibling species A and B are established. Species A evolved around 37-82 million years ago in Sri Lanka, India, and the Netherlands, and species B evolved around 22-79 million years ago in Sri Lanka, India, and Myanmar. Vietnam, Thailand, and Cambodia have two molecular forms: one is phylogenetically similar to species B. Other forms differ from species A and B and evolved recently in the above mentioned countries, Indonesia and the Philippines. Genetic subdivision among Sri Lanka, India, and the Netherlands is almost absent. A substantial genetic differentiation was obtained for some populations due to isolation by large geographical distances. Genetic diversity indices reveal the presence of a long-established stable mosquito population, at mutation-drift equilibrium, regardless of population fluctuations. Conclusions: An. subpictus species complex consists of more than two genetically divergent molecular forms. Species A is highly divergent from the rest. Sri Lanka and India contain only species A and B.

Understanding the role of pigs in the transmission of zoonotic Echinococcus ortleppi in Haryana, India.

The etiological agents of zoonotic cystic echinococcosis comprise the Echinococcus granulosus sensu lato (s.l.) species complex. The present study was aimed at investigating the zoonotic genotypes of Echinococcus granulosus s.l. circulating in the pig population of Haryana, India. Out of 253 slaughtered pigs screened, 5 showed the presence of hydatid cysts. The amplification of the partial mitochondrial NADH dehydrogenase subunit 1 (nad1) gene for the molecular confirmation and phylogenetics of the retrieved metacestodes (n = 2) revealed the presence of E. ortleppi. The sequences generated herein exhibited 99.80% homology to the GenBank archived E. ortleppi sequences. Cladistics targeting genetic diversity and haplotype network analysis involved 37 E. granulosus s.l. GenBank archived sequences from India corresponding to different hosts (large and small ruminants and humans) along with the sequences (n = 2) generated in the present study. Overall, 14 haplotypes with high haplotype (0.780 ± 0.059) and low nucleotide (0.033 ± 0.010) diversities were recorded for the overall data set, which evinced a population expansion. The median-joining haplotype network revealed a stellate shape of E. granulosus sensu stricto (s.s.) sequences, which was indicative of rapid population expansion. High genetic differentiation (FST = 0.840 - 0.983) and low gene flow (Nm = 0.003 - 0.047) were recorded between the pig intermediate hosts infected with E. ortleppi and other hosts infected with E. granulosus s.s. The findings are of paramount significance for the formulation of effective control strategies considering the public health and economic impact of cystic echinococcosis.

Molecular and morphometric data provide evidence of intraspecific variation in shape and pigmentation pattern in Otocinclus cocama (Siluriformes: Loricariidae) across major river drainages.

Otocinclus cocama, a uniquely colored species of the loricariid catfish genus Otocinclus described solely from the type locality in the lower Ucayali River in northern Peru, is reported occurring in the Tigre River, a tributary to the Marañón River that drains a different section of the Andean Mountain range in the western Amazon. Both populations differ in the number of dark bars spanning the flanks of the body, and we investigated whether these morphotypes constitute distinct species. The body shapes of populations from the Tigre and Ucayali rivers were compared using geometric morphometrics. Although principal component analysis detected a broad overlap between populations, multivariate analysis of variance and linear driscriminat analysis revealed a subtle differentiation between the populations of the two hydrographic basins. Average body shape of the Ucayali River population tend to be slightly higher than that of the Tigre River, with the caudal peduncle stretched vertically in the Ucayali population. Multivariate regression of shape and centroid size revealed an allometric effect of 10.7% (p < 0.001), suggesting that the variation between Tigre and Ucayali populations was purely shape variation. Molecular data of coI, cytb, nd2, and 16S mitochondrial genes indicated a nucleotide diversity range from 0.001 to 0.003, and haplotypic diversity range from 0.600 ± 0.11 to 0.79 ± 0.07. The median-joining haplotype network for the concatenated matrix exhibited two divergent haplogroups related to the geographic area and separated by <10 mutational steps. The molecular species delimitation methods based on distance (automatic barcode gap discovery and assemble species by automatic partitioning) recovered two molecular lineages evolving independently, being one of the lineages formed by individuals from both populations. Tree-based methods (generalized mixed Yule coalescent and Bayesian implementation of the Poisson tree process) recovered similar topologies and supported single lineage recognition. Methods of molecular delimitation of species disclosed the high similarity between the two populations of Otocinclus cocama, further supported by the presence of old haplotypes common to both groups which could indicate that the populations still maintain gene flow. Although the morphological data reveal a subtle variation between both river basins, the molecular data suggest a weak population structuration based on hydrographic areas, but not different species lineages, therefore Otocinclus cocama is composed of a single lineage with two distinct morphotypes.

Population genetic characterization of Theileria annulata based on the cytochrome b gene, with genetic insights into buparvaquone susceptibility in Haryana (India)

The present investigation was aimed at population genetic characterization of Theileria annulata on the basis of the cytochrome b (cyt b) gene along with the evaluation of status of buparvaquone resistance in Haryana (India). The sequences originating from China, Egypt, India, Iran, Iraq, Tunisia, Turkey and Sudan were included in the analysis. The maximum likelihood tree based on the Tamura-Nei (TN93+G) model placed all the sequences of T. annulata into a single clade. The median-joining haplotype network exemplified geographical clustering between T. annulata haplotypes originating from each country. Only five haplotypes (7.81 %) were shared between any two countries, while the remaining 59 haplotypes (92.19 %) were singleton and unique to one country. The values of pairwise genetic distance (FST) between all the populations indicated huge genetic differentiation (> 0.25) between different T. annulata populations, barring the FST value between Iraq and Turkey (0.14454) which suggested a moderate differentiation. Contrary to the FST index, the values of gene flow (Nm) between T. annulata populations were very low. The neutrality indices and mismatch distributions indicated a population expansion in the Indian T. annulata population. Furthermore, the secondary structure and homology modeling of the partial cyt b protein is also reported. The molecular analysis of newly generated sequences for buparvaquone resistance revealed that all the isolates were susceptible to buparvaquone treatment. However, two novel mutations at positions V203I and V219I in between the Q01 and Q02 drug-binding regions of the cyt b gene were observed for the first time.

Co-occurrence of dual lineages within Simulium (Gomphostilbia) atratum De Meijere in the Indonesian Archipelago along Wallace's Line

Mitochondrial cytochrome c oxidase subunit I (COI) sequences were utilized to infer the population genetic structure of Simulium (Gomphostilbia) atratum De Meijere, an endemic simulid species to Indonesia. Both median-joining haplotype network and maximum-likelihood tree revealed two genetic lineages (A and B) within the species, with an overlap distribution in Lombok, which is situated along Wallace's line. Genetic differentiation and gene flow with varying frequencies (FST = 0.02–0.967; Nm = 0.01–10.58) were observed between populations of S. (G.) atratum, of which population pairs of different lineages showed high genetic differentiation. Notably, the high genetic distance of up to 5.92 % observed within S. (G.) atratum in Lombok was attributed to the existence of two genetically distinct lineages. The co-occurrence of distinct lineages in Lombok indicated that Wallace's line did not act as faunistic border for S. (G.) atratum in the present study. Moreover, both lineages also exhibited unimodal distributions and negative values of neutrality tests, suggesting a pattern of population expansion. The expansion and divergence time estimation suggested that the two lineages of S. (G.) atratum diverged and expanded during the Pleistocene era in Indonesia.

Cryptic Diversity in Sympatric Migonemyia migonei (Diptera: Psychodidae), Eventual Meaning for Leishmaniasis Transmission.

Migonemyia migonei (Franҫa, 1920) (Diptera: Psychodidae) belongs to the subfamily Phlebotominae, of epidemiological importance due to its role as a vector in leishmaniasis transmission cycles and its broad geographic distribution in South America. Few morphometric and genetic studies have demonstrated the existence of variability among geographically distant populations in Brazil. The aim of the study was to estimate the genetic distance within the morphospecies Mg. migonei through the analysis of cytochrome C oxidase subunit I (COI) sequences of specimens captured in Argentina and those available in online databases. The COI sequences from specimens collected in different localities of Argentina and sequences available in online databases were utilized. Genetic distances were analyzed and a median-joining haplotype network was constructed. Finally, phylogenetic reconstruction was performed according to Bayesian inference. The analyses led to the identification of at least two haplogroups: haplogroup I with sequences of specimens from Colombia, Brazil and Argentina, and haplogroup II with sequences of specimens from Argentina. Interestingly, specimens from Argentina whose haplotypes corresponded to both haplogroups, were collected in sympatry. The results suggest that Mg. migonei could be a species complex with at least two distinct members. This hypothesis could explain the known characteristics of adaptability and vector permissiveness of the species, as the putative cryptic species of the complex could differ in traits of epidemiological importance.

Phylogeography of sharks and rays: a global review based on life history traits and biogeographic partitions.

Considerable research exists on the life history traits, evolutionary history, and environmental factors that shape the population genetic structure of marine organisms, including sharks and rays. Conservation concerns are particularly strong for this group as they are highly susceptible to anthropogenic stressors due to a combination of life history traits including late maturity and low fecundity. Here, we provide a review and synthesis of the global phylogeography of sharks and rays. We examined existing data for 40 species of sharks belonging to 17 genera and 19 species of rays belonging to 11 genera. Median joining haplotype networks were constructed for each species for the mtDNA cytochrome C oxidase subunit I (COI), and an Analysis of Molecular Variance (AMOVA) was conducted to understand patterns of genetic diversity and structure across the three major ocean basins-the Indian, Atlantic and Pacific Oceans. Haplotype networks showed very shallow coalescence in most species, a finding previously reported for marine teleosts. Star topologies were predominant among sharks while complex mutational topologies predominated among rays, a finding we attribute to extremely limited dispersal in the early life history of rays. Population structuring varied amongst species groups, apparently due to differences in life history traits including reproductive philopatry, site fidelity, pelagic habitat, migratory habits, and dispersal ability. In comparison to reef-associated and demersal species, pelagic and semi pelagic species showed lower levels of structure between and within ocean basins. As expected, there is variation between taxa and groups, but there are also some broad patterns that can guide management and conservation strategies.

Genetic diversity and haplotype analysis of Leishmania tropica identified in sand fly vectors of the genera Phlebotomus and Sergentomyia using next-generation sequencing technology.

Next-generation sequencing (NGS) was used to investigate the genetic diversity of Leishmania tropica in the sand fly vector, targeting the internal transcribed spacer 1 (ITS1) of the genus Leishmania. Bioinformatics analyses were conducted using Galaxy, MEGA version X, DnaSP ver. 6.12.03, and PopART 1.7 software for NGS analysis, phylogenetic tree, genetic diversity, and haplotype networking, respectively. A total of 307 engorged sand flies were trapped, with an overall Leishmania infection rate of 9.4 (29/307) and 6.8% by NGS and ITS1-PCR, respectively. Two Leishmania-infectedsand fly genera were identified: Phlebotomus (10.2%, 26/254) and Sergentomyia (5.7% (3/53). The phylogenetic tree showed two clusters, cluster I included the four study sequences along with 25 GenBank-retrieved DNA sequences. Cluster II consisted of three sequences from Iran and Pakistan. The genetic diversity analysis for the 29 L. tropica sequences showed high haplotype (gene) diversity index (Hd) (0.62 ± 0.07) but low nucleotide diversity index (π) (0.04 ± 0.01). Tajima's D, a neutrality test, is more negative in cluster I (D = - 2.0) than in total population (D = - 1.83), but both are equally significant (P < 0.001), indicating that observed variation in cluster I and whole population is less frequent than expected. The median-joining haplotype network produced a total of 11 active haplotypes. In conclusion, L. tropica from sand flies in Palestine is monophyletic that assembled in one main phylogroup and one haplotype.

Integrative taxonomy of two morphologically similar species of the subgenus Cryobius Chaudoir, 1838 (Coleoptera: Carabidae: Pterostichus Bonelli, 1810) from northern Eurasia and North America

In this study, we synonymized Pterostichus (Cryobius) kolymensis Erjiomin, 1998 with P. (C.) mandibularoides Ball, 1966 by an integrative approach. Geometric morphometric analysis revealed no obvious differences in the shape of the pronotum and median lobe of the aedeagus between P. (C.) mandibularoides and P. (C.) kolymensis. According to DNA analysis, all examined specimens belong to P. (C.) mandibularoides with 11 unique haplotypes of COI from northern Canada, USA (Alaska) and Russia (Yakutia, Chukotka and Wrangel Island) and 1 haplotype of 28S rDNA from northern Eurasia (Russia). The median-joining haplotype network and the uncorrected COI p-distance values between different geographic populations of P. (C.) mandibularoides suggest the trans-Beringian origin of the species and its subsequent dispersal across North America and northern Eurasia is the most likely scenario of the biogeographic history of the species.

Genetic Variation and Phylogeography of Lumbriculus variegatus (Annelida: Clitellata: Lumbriculidae) Based on Mitochondrial Genes

Lumbriculus variegatus is a typical cold-water worm and is mainly distributed in the Tibetan Plateau and Northeast in China. The current study aimed to explore the genetic diversity and phylogeography of L. variegatus sampled from different geographical regions based on concatenated (COI + 16S rRNA, 879 bp) genes. Among 63 L. variegatus specimens, 29 haplotypes were identified with high haplotype diversity (h = 0.923) and nucleotide diversity (π = 0.062). The Bayesian phylogenetic analysis and Median-joining haplotype network revealed two lineages, or species, of L. variegatus. Taxa belonging to lineage I was mainly distributed in the Tibetan Plateau of China, North America, and Sweden, while lineage II composed taxa from Northeast China, southern China, and Sweden. The analysis of molecular variance indicated that the genetic difference was mainly due to differences between lineages. Neutrality tests showed that the overall L. variegatus have a stable population since the time of origin. Divergence time analysis suggested that L. variegatus originated from the Triassic period of Mesozoic in 235 MYA (95%HPD: 199–252 MYA), and the divergence between different lineages of L. variegatus began from the next 170 million years.

Genetic Polymorphism and Phylogenetics of Aedes aegypti from Sudan Based on ND4 Mitochondrial Gene Variations.

This study investigated the genetic differences between Aedes aegypti subspecies (Aedes aegypti aegypti (Aaa) and Aedes aegypti formosus (Aaf)) from Sudan using the NADH dehydrogenase subunit 4 (ND4) mitochondrial gene marker. Nineteen distinct haplotypes of the ND4 were identified in female Aedes aegypti mosquitoes from the study sites. The phylogenetic relationship of the 19 ND4 haplotypes was demonstrated in a median-joining haplotype network tree with Aaa and Aaf populations found to share three haplotypes. The genetic variance (Pairwise FST values) was estimated and found to range from 0.000 to 0.811. Isolation by distance test revealed that geographical distance was correlated to genetic variation (coefficient value (r) = 0.43). The Polar maximum likelihood tree showed the phylogenetic relationship of 91 female Aaa and Aaf from the study sites, with most of the Aaf haplotypes clustered in one group while most of the Aaa haplotypes gathered in another group, but there was an admixture of the subspecies in both clusters, especially the Aaa cluster. The Spatial Analysis of Molecular Variance (SAMOVA) test revealed that the eight populations clustered into two phylogeographic groups/clusters of the two subspecies populations. The 2 Aedes aegypti subspecies seemed not to be totally separated geographically with gene flow among the populations.

Survivors and colonizers: Contrasting biogeographic histories reconciled in the Antarctic freshwater copepod Boeckella poppei

Two main hypotheses have been proposed to explain the contemporary distribution of Antarctic terrestrial biota. We assess whether the current distribution of maritime Antarctic populations of the freshwater copepod Boeckella poppei is the result of (1) a post-Last Glacial Maximum (LGM) colonization, or whether (2) the species survived in regional glacial refugia throughout the LGM and earlier glaciations. Using 438 specimens from 34 different sampling sites across Southern South America, South Georgia, South Orkney Islands, South Shetland Islands, and the Antarctic Peninsula, we analyzed mitochondrial and nuclear sequences to uncover patterns of genetic diversity and population structure. We also performed median-joining haplotype network, phylogenetic reconstruction, and divergence time analyses. Finally, we evaluated past demographic changes and historical scenarios using the Approximate Bayesian Computation (ABC) method. Our data support the existence of two clades with different and contrasting biogeographic histories. The first clade has been present in maritime Antarctica since at least the mid-Pleistocene, with the South Orkney Islands the most likely refugial area. The second clade has a broader distribution including southern South America, South Georgia, South Shetland Islands, and the Antarctic Peninsula. The ABC method identified long-distance dispersal (LDD) colonization event(s) from southern South America to South Georgia and the maritime Antarctic after the LGM deglaciation, supporting more recent colonization of Antarctic locations. The current Antarctic and sub-Antarctic distribution of B. poppei is likely derived from two independent biogeographic events. The combination of both (1) post-LGM colonization from southern South America and (2) longer-term persistence in in situ regional refugia throughout glacial periods challenges current understanding of the biogeographic history of Antarctic freshwater biota. Re-colonization of ice-impacted Antarctic areas would have occurred following a LDD and Establishment model, pointing to the existence of possible post-dispersal barriers, despite widely assumed high passive dispersal capacity in freshwater invertebrates.

Assessment of genetic markers for multilocus sequence typing (MLST) of Fasciola isolates from Iran.

Several markers have been described to characterise the population structure and genetic diversity of Fasciola species (Fasciola hepatica (F. hepatica) and Fasciola gigantica (F. gigantica). However, sequence analysis of a single genomic locus cannot provide sufficient resolution for the genetic diversity of the Fasciola parasite whose genomes are ∼1.3 GB in size. To gain a better understanding of the gene diversity of Fasciola isolates from western Iran and to identify the most informative markers as candidates for epidemiological studies, five housekeeping genes were evaluated using a multilocus sequence typing (MLST) approach. MLST analysis was developed based on five genes (ND1, Pepck, Pold, Cyt b and HSP70) after genomic DNA extraction, amplification and sequencing. Nucleotide diversity and phylogeny analysis were conducted on both concatenated MLST loci and each individual locus. A median joining haplotype network was created to examine the haplotypes relationship among Fasciola isolates. Thirty-three Fasciola isolates (19 F. hepatica and 14 F. gigantica) were included in the study. A total of 2971bp was analysed for each isolate and 31 sequence types (STs) were identified among the 33 isolates (19 for F. hepatica and 14 for F. gigantica isolates). The STs produced 44 and 42 polymorphic sites and 17 and 14 haplotypes for F. hepatica and F. gigantica, respectively. Haplotype diversity was 0.982 ± 0.026 and 1.000 ± 0.027 and nucleotide diversity was 0.00200 and 0.00353 ± 0.00088 for F. hepatica and F. gigantica, respectively. There was a high degree of genetic diversity with a Simpson's index of diversity of 0.98 and 1 for F. hepatica and F. gigantica, respectively. While HSP70 and Pold haplotypes from Fasciola species were separated by one to three mutational steps, the haplotype networks of ND1 and Cyt b were more complex and numerous mutational steps were found, likely due to recombination. Although HSP70 and Pold genes from F. gigantica were invariant over the entire region of sequence coverage, MLST was useful for investigating the phylogenetic relationship of Fasciola species. The present study also provided insight into markers more suitable for phylogenetic studies and the genetic structure of Fasciola parasites.

Population Structure of Wild Schizothorax kozlovi in the Upper Yangtze River Based on mtDNA and Stable Isotopes, and Their Relationship with Ambient Temperature

Schizothorax kozlovi, as an endemic and vulnerable fish of the upper Yangtze River in China, faces many threats. In order to expose the population structure of wild S. kozlovi, the carbon and oxygen isotopic ratios in the otoliths, and the gene sequences of two common mitochondrial markers (Cytb and COI) were investigated in four sampling locations, and then their relationship with ambient temperature was further investigated. In general, it exhibits limited geographic population structuring of S. kozlovi in the upper Yangtze River by both mtDNA and stable isotopes. The values of otolith stable isotope ratios varied from −15.30‰ to −12.37‰ for δ18O and from −10.10‰ to −6.13‰ for δ13C. Significant relationships were revealed between stable isotope ratios and specific mean monthly water temperature variables (from November to March), indicating low temperature effect on otolith stable isotope ratios. Haplotype diversity and nucleotide diversity were 0.928 and 0.00778, both exhibiting high levels. A median-joining haplotype network indicated a mixture of geographical distribution but exhibited two distinct haplotype lineages (Clade I and Clade II). AMOVA detected that the higher percentage of genetic variance was within sampling locations (96.94%) and between two haplotype lineages (72.82%). Most FST values between sampling locations showed small levels of genetic differentiation except the differentiation between population SJ (Sanjiangkou) and JP (Jinping). Therefore, two haplotype lineages and population JP of S. kozlovi in the upper Yangtze River are suggested as three management units for conservation due to their moderate-to-great genetic differentiation and isolated habitat.

Molecular analysis of knockdown resistance (kdr) mutations in the voltage-gated sodium channel gene of Aedes aegypti populations from Saudi Arabia

BackgroundThe Aedes aegypti mosquito is the primary vector for dengue, chikungunya, yellow fever and Zika viruses worldwide. The first record of Ae. aegypti in southwestern Saudi Arabia was in 1956. However, the first outbreak and cases of dengue fever were reported in 1994, and cases have increased in recent years. Vector control for Ae. aegypti mainly uses pyrethroid insecticides in outdoor and indoor space spraying. The constant use of pyrethroids has exerted intense selection pressure for developing target-site mutations in the voltage-gated sodium channel (vgsc) gene in Ae. Aegypti against pyrethroids—mutations that have led to knockdown resistance (kdr).MethodsAedes aegypti field populations from five regions (Jazan, Sahil, Makkah, Jeddah and Madinah) of southwestern Saudi Arabia were genotyped for known kdr mutations in domains IIS6 and IIIS6 of the vgsc gene using polymerase chain reaction (PCR) amplification and sequencing. We estimated the frequency of kdr mutations and genotypes from Saudi Arabia as well as from other countries, Thailand, Myanmar (Southeast Asia) and Uganda (East Africa). We constructed haplotype networks to infer the evolutionary relationships of these gene regions.ResultsThe three known kdr mutations, S989P, V1016G (IIS6) and F1534C (IIIS6), were detected in all five regions of Saudi Arabia. Interestingly, the triple homozygous wild genotype was reported for the first time in two individuals from the highlands of the Jazan region and one from the Al-Quoz, Sahil region. Overall, nine genotypes comprising four haplotypes were observed in southwestern Saudi Arabia. The median-joining haplotype networks of eight populations from Saudi Arabia, Southeast Asia and East Africa for both the IIS6 and IIIS6 domains revealed that haplotype diversity was highest in Uganda and in the Jazan and Sahil regions of Saudi Arabia, whereas haplotype diversity was low in the Jeddah, Makkah and Madinah regions. Median-joining haplotype networks of both domains indicated selection acting on the kdr-mutation containing haplotypes in Saudi Arabia.ConclusionsThe presence of wild type haplotypes without any of the three kdr mutations, i.e. that are fully susceptible, in Saudi Arabia indicates that further consideration should be given to insecticide resistance management strategies that could restore pyrethroid sensitivity to the populations of Ae. aegypti in Saudi Arabia as part of an integrative vector control strategy.Graphical

Genetic and population diversity of Toxocara cati (Schrank, 1788) Brumpt, 1927, on the basis of the internal transcribed spacer (ITS) region.

The present investigation was aimed to study the sequence, phylogenetic and haplotype analyses of Toxocara cati based on the ITS region, along with the genetic diversity, demographic history and population-genetic structure. The maximum likelihood tree based on Kimura 2-parameter model was constructed using the complete ITS region of all the nucleotide sequences (n = 57) of Toxocara spp. and other related ascarid worms available in the GenBank™. It placed all the sequences of T. cati into four major clades designated as T. cati genotypes 1-4 (TcG1-G4). A total of 66 signature nucleotides were identified in the ITS region between genotypes. The median-joining haplotype network displayed a total of 24 haplotypes, with China exhibiting the highest number of haplotypes (h = 20) followed by India (h = 4), and Japan and Russia (h = 1). It indicated a clear distinction between all the four genotypes. The pairwise FST values between all the genotypes indicated huge genetic differentiation (> 0.25) between different T. cati genotypes. Moreover, the gene flow (Nm) between T. cati genotypes was very low. Results of AMOVA revealed higher genetic variation between genotypes (92.82%) as compared to the variation within genotypes (7.18%). The neutrality indices and mismatch distributions for the G1-G4 genotypes, Indian isolates and the overall dataset of T. cati indicated either a constant population size or a slight population increase. The geographical distribution of all the genotypes of T. cati is also reported. This is the first report of genotyping of T. cati on the basis of the ITS region.

Incorporation of Turkish Hyalopterus spp. into recent species reassessment based on their molecular and morphometric features.

The Hyalopterus group is a Mediterranean originated genus and Turkey is one of the important distribution pathways of the species belonging to this genus. Up to date, Hyalopterus amygdali (from host plant Prunus dulcis), Hyalopterus pruni (from Prunus domestica) and Hyalopterus arundiniformis (from Prunus armeniaca and Prunus persica) are species distinctively defined in Turkey. Samples of Hyalopterus were collected on P. dulcis, P. domestica, P. armeniaca, and P. persica from Afyonkarahisar, Ktahya, Nide and Uak provinces from May 2012 to October 2013. To detect morphological variations related with the preferred host plant and locality, 755 individuals morphological features were measured. Maximum likelihood and Bayesian analyses of COI (658 bp.) sequences of Hyalopterus spp were applied to evaluate phylogenetic relationships of the genus members. A median joining haplotype network was evaluated to determine the relationships among haplotypes. The molecular and morphometric analyses indicated that the host plant utilization affects the speciation of Hyalopterus spp. There are no comprehensive molecular studies about Hyalopterus in Turkey although previous studies conducted on this genus emphasized the importance of studying the Turkish populations to understand host plant relations, distribution route and speciation of this economically important aphid group. Therefore this study aimed to give information about phylogenetic and morphological variability of Hyalopterus spp. and criticize how these findings are incorporated into the recent species reassesment.