High Haplotype Diversity Research Articles

Phylogeography and demographic history of the cyprinid fish Barbodes semifasciolatus: implications for the history of landform changes in south mainland China, Hainan and Taiwan

Hainan Island and Taiwan Island are adjacent to the southern margin of mainland China and Vietnam. During glacial periods, global sea levels dropped, allowing that the land bridges connected the continental island and mainland, connecting rivers and providing dispersal opportunities that shaped the origin and diversification of freshwater fishes. Barbodes semifasciolatus is distributed in various water systems of Vietnam, Hainan, Taiwan, and southern mainland China and is restricted to the southern region of the Min River. Our study aimed to evaluate the genetic diversity and phylogeography of B. semifasciolatus using the mtDNA cyt b gene (1,141 bp). A total of 107 haplotypes were identified from 395 specimens in 23 populations, and high haplotype diversity (1.000) and low nucleotide diversity (0.0134) were detected. Mitochondrial phylogenetic analysis and haplotype network analyses revealed three major lineages according to geographical distribution. Lineage A was mainly distributed in Hainan Island, Vietnam and the southern region of the Pearl River in mainland China. Lineage B was distributed only in southeastern Hainan Island. Lineage C was distributed in the coastal rivers of mainland China and Taiwan. We suggest that the river in the Guangdong region is a colonization route in South Taiwan and that the populations distributed in the Pearl River region moved southward to Hainan Island and Vietnam based on the network and Bayesian binary MCMC (BBM) analysis. Our demographic history results indicated that the populations of B. semifasciolatus experienced a bottleneck event following a recent population expansion (DECINC model) supported by ABC analysis. We suggest that sea-level changes exerted pronounced effects on the demography of B. semifasciolatus on the continental island and in the mainland during the late Pleistocene glacial cycles.

Intrabreed Diversity and Relationships between Races of Honey Bee <i>Apis mellifera carpathica</i> and <i>Apis mellifera caucasica</i>

Based on the analysis of 94 sequences of the mtDNA CO1 gene, the intrabreed diversity and evolutionary relationships of bees from Russia, Ukraine, Armenia, Kyrgyzstan, Tajikistan, and Poland were studied. We revealed two main haplotypes corresponding to the races A. m. carpathica and A. m. caucasica, common in the studied regions. However, in Armenia bees of the Caucasian race predominate, and in Poland – Carpathian. A. m. caucasica has higher haplotype diversity: one of its haplotypes is formed by samples of bees from the Crimean mountain breed, possibly an independent race A. m. taurica. There are 2 clusters on the phylogenetic tree: one cluster includes A. m. mellifera and A. m. iberica (evolutionary line M), the other (line C) ‒ A. m. ligustica, A. m. carpathica, A. m. caucasica. Haplotypes of A. m. caucasica have more substitutions per site, indicating their earlier origin compared to A. m. ligustica and A. m. carpathica. According to the time of divergence of A. mellifera and A. cerana at 6 Ma, the divergence of all races of line C is estimated from 1.3 to 0.6 Ma; races A. m. caucasica 0.35–0.25 million, A. m. carpathica – 0.2–0.04 million. When using universal primers with optimization of amplification conditions for the mtDNA CO1 gene region, a nonsynonymous G/A SNP was detected in position 4 (680 bp) which can be used to identify A. m. carpathica from A. m. ligustica.

Genetic Variations of Malaysian And Golden Thai Strains of Climbing Perch and Their Hybrids Based On The Partial Mitochondrial Cytochrome Oxidase Subunit 1 Gene

Rapid genetic improvements from selective breeding are anticipated in many aquaculture species and mitogenome is used to complement the morphological taxonomy of hybrids to evaluate its genetic structures. A study of the genetic variations within the two strains (Malaysian strain and Golden Thai strain) of Anabas testudineus (Bloch, 1792) and their hybrids, phylogenetic trees based on cytochrome oxidase subunit 1 (CO1) of the partial mitochondrial DNA gene were constructed using Maximum Likelihood and Neighbor Joining approaches. The findings support the monophyletic status of the genus with only one haplogroup from which other haplotypes were descended and of a single common ancestor. The individual fishes’ phylogenetic relationships revealed two major clades and Saurida undosquamis as an outgroup. All the groups had high haplotype diversity, except for the hybrid Malaysian strain × golden Thai strain (0.1540±0.126). This suggests that different fish species in each of these studies had different nucleotide compositions in their mitochondrial genomes. The highest number of haplotypes and the presence of distinct haplotypes in the Golden Thai strain × Malaysian strain hybrid point to the absence of recent or regular gene flow as well as high genetic diversity within the hybrids. This study demonstrated that the mtDNA diversity of Anabas testudineus from Malaysia and Thailand had been preserved. Studies on the population genetic diversity and molecular evolution of anabas fish species can benefit from the data provided by this work.

Genetic diversity of mtDNA in the grey wolf population of Belarus threatened by wolf-dog admixture

Hybridisation with domestic species is one of the prominent threats to conser-vation of numerous valuable species in Belarus, in particular the grey wolf (Canis lupus Linnaeus, 1758), the population of which has previously shown unusually divergent genotypes, indicating a possible admixture with free-ranging dogs (Canis lupus familiaris Linnaeus, 1758). Such admixture could threaten the future of the species already weakened by anthropogenic pressure. Reports of hybrids in Belarus based on morphological characteristics have been steadily increasing in frequency since 2010, which can be tied with a growing food availability for feral dogs and low population density of grey wolf facili-tating hybridisation. A limited number of genetic studies with sampling that partially covered Belarus did not detect any traces of hybridisation. Here we report our estimate of genetic diversity in the Belarusian population of grey wolf according to control region (D-loop) mtDNA sequence analysis. We ana-lysed tissue samples from 35 specimens harvested during legal hunts across 25 administrative districts from all 6 regions of Belarus between 2009 and 2022, with 9 of the specimens reported to have morphological characteristics of wolf–dog hybrids. We detected 6 haplotypes among those 35 specimens, 4 of which were found among the likely wolf–dog hybrids. Clade analysis of the obtained sequences with 100 confirmed sequences of wolves, dogs, and their hybrids from the NCBI Genbank database has shown the presence of possible wolf–dog hybrids in our sample. The Belarusian wolf population has shown a low nucleotide and a relatively high haplotype diversity. The discovered genet-ic diversity data of the Belarusian wolf population is mostly in line with studies of similar wolf populations across Europe. The persistence of large wolf meta-populations across Eastern Europe and Russia, combined with the high mobility of the animal, seems to be the probable cause of genetic diversity of wolf populations in Belarus, but some degree of wolf–dog hybridisation could also explain the observed genetic heterogeneity, which invites further research based on nuclear markers.

SARS-CoV-2 variants in Vietnam: A comprehensive analysis of nucleotide changes in the spike gene

SARS-CoV-2, causing the COVID-19 pandemic, has claimed millions of lives worldwide. SARS-CoV-2 has a high mutation rate in its genome, resulting in thousands of variants. The success of SARS-CoV-2 is attributed to a mutation in the S gene, which encodes the spike protein that interacts directly with hACE2. Mutations in this gene are known to increase the transmission rate and the ability to escape the immune system of the virus. This study focused on analyzing the nucleotide changes in the S gene of the SARS-CoV-2 variants that appeared in Vietnam. The results showed that Vietnam recorded many VOC variants, including Alpha, Beta, Delta, and Omicron, with Delta and Omicron being the most prevalent. The S1 region of the S gene had the highest mutation rate, with missense and C to T mutations being the most common. The NTD region contained all deletion and insertion mutations, with the nucleotide at 22198 to 22206 being the hotspot for insertion. The RBD region showed a positive selection during evolution, indicating that it had undergone harsh missense mutation. Overall, this study demonstrates that the S gene in Vietnam has high haplotype and nucleotide diversity. The Omicron variant in Vietnam had the highest nucleotide, haplotype diversity indexes, and average number of mutations in the S gene. These findings provide insights into the genetic diversity of SARS-CoV-2 variants in Vietnam and the impact of the S gene mutations on the evolution of the virus.

Genetic Diversity and Phylogeography of Taenioides cirratus in Five Geographical Populations Based on Mitochondrial COI and Cytb Gene Sequences

The genetic diversity of Taenioides cirratus was investigated on the basis of mitochondrial cytochrome c oxidase subunit I (COI) and cytochrome b (Cytb) gene sequences. A total of 159 specimens collected from the Chaohu Lake (CL), Nansihu Lake (NL), Taihu Lake (TL), Pearl River (PR), and Nandu River (NR) were sequenced. The total length of the sequence was 2485 bp with 412 polymorphic sites. A total of 73 haplotypes were identified, with Hap1 being the most widely distributed. The PR and NR populations showed high genetic diversity, while the CL population showed low genetic diversity. TL and NL showed high haplotype diversity but low nucleotide diversity. The analysis of molecular variance demonstrated that the sequence variations were mainly occurred among populations. T. cirratus populations are declining, and rare alleles are present at low frequencies, as analysed using a neutral test and a mismatched distribution analysis. There was a relatively high level of genetic differentiation among the populations of the Yangtze River basin (including NL), PR, and NR (Fst > 0.15). The two similar phylogenetic trees constructed by the maximum likelihood (ML) and Bayesian inference (BI) methods presented three major lineages, of which lineage II contains haplotypes from PR and NR, lineage III contains haplotypes from CL, NL, TL, and PR, whereas lineage I contains only a portion of haplotypes from NR. Based on the neutral test, mismatch analysis, and Bayesian Skyline Plot (BSP), geological and climatic events were inferred to have played an important role in the historical dynamics of T. cirratus population. Hap1, Hap25, and Hap58 were inferred as possible ancestral haplotypes by network analysis. Our study offers an essential foundation for resource preservation and additional taxonomic clarification of T. cirratus.

Revealing novel cytb and nad5 genes-based population diversity and benzimidazole resistance in Echinococcus granulosus of bovine origin.

Cystic echinococcosis (CE) is a neglected zoonotic disease caused by Echinococcus granulosus (sensu stricto). The parasite affects a wide range of livestock and wild animals. In this study, the population diversity of the Echinococcus species was investigated based on mitochondrial cytochrome b (cytb) and NADH dehydrogenase subunit 5 (nad5) genes. In addition to this, β-tubulin gene isoforms of Echinococcus granulosus were amplified to determine the resistance against benzimidazoles. For this purpose, 40 cyst samples from cattle (n = 20) and buffaloes (n = 20) were collected from the main abattoir of Sialkot. DNA extraction was performed using Qiagen Blood and Tissue Kits. Amplification was performed through PCR. Each amplicon was confirmed by GelRed™ stained agarose gel (2%). Samples were sequenced in a DNA analyzer and viewed for any misread nucleotide by using MEGA (v.11). Corrections in nucleotide sequence and multiple sequence alignment were made through the same software. NCBI-BLAST was used for sample specific sequences to identify them as belonging to a particular species. Diversity indices were estimated using DnaSP (v.6) while phylogenetic analysis was inferred using the Bayesian method using MrBayes (v.1.1). β-tubulin gene isoforms sequence analysis was performed to find out the candidate gene causing benzimidazole resistance. All 40 isolates were found positive for E. granulosus. BLAST-based searches of sequences of each isolate for each gene (nad5 and cytb) confirmed their maximum similarity with the G1 genotype. Overall, high haplotype diversity (Hd nad5 = 1.00; Hd cytb = 0.833) and low nucleotide diversity (π nad5 = 0.00560; π = cytb = 0.00763) was identified based on diversity indices. For both the genes, non-significant values of Tajima's D (nad5 = -0.81734; cytb = -0.80861) and Fu's Fs (nad5 = -1.012; cytb = 0.731) indicate recent population expansion. Bayesian phylogeny-based results of nad5 and cytb sequences confirmed their genotypic status as distinct from other Echinococcus species. This study shed light on the status of benzimidazole resistance in Echinococcus granulosus for the very first time from Pakistan. The findings of this study will significantly add in the information available on genetic diversity of Echinoccous granulosus based on cytb and nad5 genes sequences.

Genetic variation of giant freshwater prawns Macrobrachium rosenbergii wild population of South Sulawesi, Indonesia

Abstract. Wahidah, Omar SBA, Trijuno DD, Nugroho E, Amrullah, Khatimah K. 2023. Genetic variation of giant freshwater prawns Macrobrachium rosenbergii wild population of South Sulawesi, Indonesia. Biodiversitas 24: 3081-3090. Quality giant freshwater prawn broodstock can be provided using broodstock sources from wild populations with good morphological and genetic performance. This study aimed to analyze the genotypes of the wild population of giant freshwater prawns in South Sulawesi, hence it can be used as basic information for quality broodstock. Evaluation of giant prawn haplotypes, Macrobrachium rosenbergii, from 3 rivers in South Sulawesi: 26 samples from the Waelawi River (11 males, 15 females); 25 samples from the Kalibone River (10 males, 15 females); and 26 samples from the Kariango River (12 males, 14 females), so there are 6 population groups. Analysis used mt-DNA and obtained an amplification of 700 bp. The amplified restriction uses four enzymes (Hae III, Mbo I, Rsa I, and Taq I). The identified male and female Kariango populations each had 3 haplotypes: female Kalibone had 2 haplotypes; male and female Waelawi had 1 haplotype; and male Kalibone had 1 haplotype. Female Kariango had the highest haplotype diversity (0.3651), followed by male Kariango (0.1449) and female Kalibone (0.1011). Kariango females have the farthest genetic distance from Kalibone males, Waelawi males, and Waelawi females. The male Waelawi, female Waelawi, and male Kalibone are in one cluster and form a new cluster with the male Kariango, then with the female Kalibone, and the last cluster is formed with the female Kariango. The Kariango female population can be used as potential broodstock, which can be mated with Kalibone males, Waelawi males, or other populations.

Habitat quality affects the social organization in mouse lemurs (Microcebus griseorufus)

Species can either maintain a certain social organization in different habitats or show different social organizations in similar habitats. The reasons underlying this variability are not always clear but might have consequences for population dynamics, especially under changing environmental conditions. Among mammals, the primate genus Microcebus lives in small groups of closely related females, derived from female philopatry and dispersed males, as illustrated by the well-studied Microcebus murinus. Here, we studied the genetic structure of a population of the congeneric Microcebus griseorufus, inhabiting three adjacent habitats with different resource availabilities. In order to learn more about the plasticity of the species’ social organization under these different conditions, we analyzed the spatial arrangement of mitochondrial haplotypes of 122 individuals. The study revealed high haplotype diversity and a pronounced difference in spatial distribution between the sexes. Females exhibited spatial aggregation of haplotypes, suggesting a system of female philopatry and matrilines, similar to M. murinus. Male haplotypes were dispersed, and males were more likely to carry rare haplotypes, indicating higher dispersal activity. These findings hint towards the unity of the social organization across the genus Microcebus, suggesting a phylogenetic origin of the social organization. Yet, with decreasing resources, the clustering of female haplotypes declined and approached a random distribution in the marginal habitat, with cluster sizes correlating with resource availability as predicted by the socioecological model. Our study supports the notion that social organization is shaped by both phylogenetic origin and ecological conditions, at least in these small primates.Significance statementImpacts of habitat degradation are mostly described in terms of changes in population densities in relation to the reduction of resources. This neglects the possible effects of altered social organizations due to declining resources or population densities. Using a genetic sampling of three subpopulations of mouse lemurs in Madagascar along a gradient of food availability up to the limit of the species’ ecological tolerance, we show that their social organization consisting of spatial clusters of closely related females and overdispersed males converges towards random spatial distributions of both sexes with declining food availability.

Species diversity of phlebotomine sand flies and sympatric occurrence of Leishmania (Mundinia) martiniquensis, Leishmania (Leishmania) donovani complex, and Trypanosoma spp. in the visceral leishmaniasis focus of southern Thailand

Autochthonous leishmaniasis in Thailand has recently been a public health concern due to an increasing number of new clinical cases. Most indigenous cases were diagnosed with Leishmania (Mundinia) martiniquensis, and Leishmania (Mundinia) orientalis. However, some doubts regarding vector misidentification have arisen and need to be elucidated. Accordingly, we aimed to assess the species composition of sand flies and determine the molecular prevalence of trypanosomatids in the transmission area of leishmaniasis in southern Thailand. In the present study, a total of 569 sand flies were caught from the vicinity of a visceral leishmaniasis patient's house in Na Thawi District, Songkhla Province. Of these, 229 parous and gravid females consisted of Sergentomyia khawi, Se. barraudi, Phlebotomus stantoni, Grassomyia indica, and Se. hivernus, accounting for 31.4%, 30.6%, 29.7%, 7.9%, and 0.4%, respectively. However, Se. gemmea, which has previously been proposed as the most abundant species and putative vector of visceral leishmaniasis, was not found in the present study. Based on ITS1-PCR and sequence analysis, two specimens of Gr. indica and Ph. stantoni showed positive amplification of L. martiniquensis and L. donovani complex, respectively, the first one being presumed indigenous and the second one being not. Anuran Trypanosoma was also molecularly detected using SSU rRNA-PCR and ubiquitously found in 16 specimens of four dominant sand fly species except for Se. hivernus. The obtained sequences could be phylogenetically categorized into the two major amphibian clades (An04/Frog1 and An01+An02/Frog2). The existence of the monophyletic subgroup and distinct lineage suggests them as novel Trypanosoma species. The TCS network analysis of these anuran Trypanosoma sequences also revealed high haplotype diversity (Hd = 0.925 ± 0.050), but low nucleotide diversity (π = 0.019 ± 0.009). Furthermore, the living anuran trypanosomes were microscopically demonstrated in a single specimen of Gr. indica, supporting the vectorial capacity. Importantly, our data confirmed the scarcity of Se. gemmea and also uncovered, for the first time, the co-circulation of L. martiniquensis, L. donovani complex, and suspected novel anuran Trypanosoma spp. in phlebotomine sand files, implicating their potential role as vectors of trypanosomatid parasites. Therefore, the novel data from this study would greatly facilitate the comprehension of the complexity of trypanosomatid transmission and the establishment of prevention and control measures for this neglected disease more effectively.

Genetic Evidence for Colletotrichum gloeosporioides Transmission Between the Invasive Plant Ageratina adenophora and Co-occurring Neighbor Plants.

To understand the disease-mediated invasion of exotic plants and the potential risk of disease transmission in local ecosystems, it is necessary to characterize population genetic structure and spatio-temporal dynamics of fungal community associated with both invasive and co-occurring plants. In this study, multiple genes were used to characterize the genetic diversity of 165 strains of Colletotrichum gloeosporioides species complex (CGSC) isolated from healthy leaves and symptomatic leaves of invasive plant Ageratina adenophora, as well as symptomatic leaves of its neighbor plants from eleven geographic sites in China. The data showed that these CGSC strains had a high genetic diversity in each geographic site (all Hd > 0.67 and Pi > 0.01). Haplotype diversity and nucleotide diversity varied greatly in individual gene locus: gs had the highest haplotype diversity (Hd = 0.8972), gapdh had the highest nucleotide diversity (Pi = 0.0705), and ITS had the lowest nucleotide diversity (Pi = 0.0074). Haplotypes were not clustered by geographic site, invasive age, or isolation source. AMOVA revealed that the genetic variation was mainly from within-populations, regardless of geographic or isolation origin. Both AMOVA and neutrality tests indicated these CGSC strains occurred gene exchange among geographic populations but did not experience population expansion along with A. adenophora invasion progress. Our data indicated that A. adenophora primarily accumulated these CGSC fungi in the introduced range, suggesting a high frequency of CGSC transmission between A. adenophora and co-occurring neighbor plants. This study is valuable for understanding the disease-mediated plant invasion and the potential risk of disease transmission driven by exotic plants in local ecosystems.

Studying Genetic Diversity and Relationships between Mountainous Meihua Chickens Using Mitochondrial DNA Control Region.

The Mountainous Meihua chicken is a unique regional germplasm resource from Tongjiang County, Bazhong City, China, but its genetic structure and evolutionary relationships with other native chicken breeds in the Sichuan region remain unclear. Here, we analyzed a total of 469 sequences, including 199 Mountainous Meihua chicken sequences generated in this study, together with 30 sequences representing 13 clades and 240 sequences from seven different Sichuan local chicken breeds downloaded from NCBI. These sequences were further used to analyze genetic diversity, patterns of population differentiation, and phylogenetic relationships between groups. We show that Mountainous Meihua chicken mtDNA sequences have high haplotypic and nucleotide diversity (0.876 and 0.012, respectively) and with a T bias that is suggestive of good breeding potential. Phylogenetic analysis showed that Mountainous Meihua chickens belong to clades A, B, E, and G and have a low affinity to other chicken breeds, with a moderate degree of differentiation. A non-significant Tajima's D indicates that no demographic expansions occurred in the past. Finally, the four maternal lineages identified in Mountainous Meihua chicken showed unique genetic characteristics.

Rapid Detection and Identification of Fasciola spp. and Dicrocoelium spp. Isolated from the Ruminant Livestock of Northwest Iran Using High-Resolution Melting Analysis (HRM).

The liver flukes of the Fasciola species and Dicrocoelium spp. are recognised as parasites of domestic and wild herbivores. Both species of F. hepatica and F. gigantica as well as D. dendriticum are distributed in Iran. The present study aimed to identify Fasciola spp. and Dicrocoelium spp. using mitochondrial Cox1 (cytochrome c oxidase I) gene by HRM method. Totally, thirty infected liver specimens were collected from the sheep (n:23) and cattle (n:7) at the abattoirs of Qazvin Province, northwest Iran in 2022. DNA extraction and PCR amplification of Cox1 gene were conducted by HRM technique. DnaSP v.5.0 was used for compression of diversity indices of ribosomal 28S rDNA and mitochondrial Cox1 markers of Dicrocoelium spp. The taxonomic status of Dicrocoelium spp. was performed by sequencing and phylogenetic analysis. Overall, 26 and 4 isolates were identified as F. hepatica and F. gigantica, respectively. D. dendriticum was the sole infecting species of Dicrocoelium revealed by HRM analysis. Genomic analysis showed a moderate (28S rDNA genes: 0.600±0.215) to high (Cox1: 0.733±0.155) haplotype diversity for D. dendriticum. The parasite-dependent mitochondrial gene (Cox1) could identify a higher genetic diversity of D. dendriticum compared to nuclear 28S rDNA gene. HRM technique in the present study found to be a reliable technique for identification and genetic diversity of liver flukes but more comprehensive and in-depth studies in different parts of the country are needed.

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.

Shedding light on variation in reproductive success through studies of population genetic structure in a Southeast Pacific Coast mussel.

Phylogeography often focuses on the spatial dimension of genetic diversity, rarely including the temporal dynamics occurring interannually among local populations, which can provide insight into past variations in reproductive success. Currently, there is an intense aquaculture industry of Mytilus spp. on the Southeast Pacific Coast which depends entirely on the spat released by natural populations forming a relevant and sensitive social-ecological system. Temporal and spatial spat variability from natural mussel beds could be related to interannual reproductive dynamics with variable reproductive success and recruitment, which leave genetic signatures. Temporal and spatial genetic structure was evaluated in six natural beds in the Southeast Pacific (from 39°25'S to 43°07'S) on the most abundant and widespread Mytilus lineage detected, Mytilus cf. chilensis, in 4 consecutive years. Analyses included data from >180 individuals per year, with a total of 751 (mitochondrial COI) and 747 (nuclear H1) individuals, respectively. Overall, both markers showed high haplotype diversity and low spatial and temporal genetic differentiation. Likely, the high dispersal capacity of Mytilus cf. chilensis maintains population homogeneity and prevents diversity erosion. The slight differences in genetic variance of COI were better explained by differences among sites (space), and conversely, the H1 genetic variance was better explained by interannual (temporal) comparisons, which could explain temporal variability in spat availability. This study highlights the important insights achieved with the evaluation of both temporal and spatial population genetic structures in marine species with high reproductive output, which can condition the success and sustainability of the relevant social-ecological system.

Divergent evolution drives high diversity of toll-like receptors (TLRs) in passerine birds: Buntings and finches

Toll-like receptors (TLRs) form a key component of animal innate immunity, being responsible for recognition of conserved microbial structures. As such, TLRs may be subject to diversifying and balancing selection, which maintains allelic variation both within and between populations. However, most research on TLRs in non-model avian species is focused on bottlenecked populations with depleted genetic variation. Here, we assessed variation at the extracellular domains of three TLR genes (TLR1LA, TLR3, TLR4) across eleven species from two passerine families of buntings (Emberizidae) and finches (Fringillidae), all having large breeding population sizes (millions of individuals). We found extraordinary TLR polymorphism in our study taxa, with >100 alleles detected at TLR1LA and TLR4 across species and high haplotype diversity (>0.75) in several species. Despite recent species divergence, no nucleotide allelic variants were shared between species, suggesting rapid TLR evolution. Higher variation at TLR1LA and TLR4 than TLR3 was associated with a stronger signal of diversifying selection, as measured with nucleotide substitutions rates and the number of positively selected sites (PSS). Structural protein modelling of TLRs showed that some PSS detected within TLR1LA and TLR4 were previously recognized as functionally important sites or were located in their proximity, possibly affecting ligand recognition. Furthermore, we identified PSS responsible for major surface electrostatic charge clustering, which may indicate their adaptive importance. Our study provides compelling evidence for the divergent evolution of TLR genes in buntings and finches and indicates that high TLR variation may be adaptively maintained via diversifying selection acting on functional ligand binding sites.

Incidence, distribution, and genetic diversity of ‘Candidatus Phytoplasma luffae’-related strain (16SrVIII) associated with the cassava witches' broom (CWB) disease in the Philippines

Cassava (Manihot esculenta Crantz) is widely grown in the Philippines mainly for human consumption, animal feed, and industrial purposes. However, the cassava industry is recently challenged by the cassava witches' broom (CWB) disease which is known to be caused by phytoplasmas. The extent of the disease and the causal phytoplasma strain in the country are not known until this study. Surveys in major cassava areas in the Philippines conducted from 2017 to 2019 revealed that CWB is present in 13 out of 19 provinces assessed in this study, with symptoms ranging from leaf proliferation, little leaf, shortened internodes, stunted growth, discoloration of the vascular system, and small tubers. Molecular detection (through nested PCR) and sequence analysis of phytoplasma 16S ribosomal RNA gene from CWB-infected plants showed high genetic similarity of >98.65% to Candidatus Phytoplasma luffae 16SrVIII. In addition, the 18 phytoplasma isolates from five provinces successfully grouped to Ca. Phytoplasma luffae 16SrVIII clade in the constructed phylogenetic tree. Analysis on the genetic diversity and structure of Ca. Phytoplasma luffae 16SrVIII sequences from cassava and other host plant species revealed low genetic diversity indices and a rudimentary genetic structure with high haplotype diversity. Meanwhile, neutrality tests showed evidence of recent population expansion in Ca. Phytoplasma luffae 16SrVIII. This is the first study to identify ‘Ca. Phytoplasma luffae’-related strain (16SrVIII) as the causal pathogen of CWB in the Philippines. The information generated in this study can help in better understanding the etiology of the disease and in formulating proper and effective strategies to detect and manage the disease in the Philippines.

Phylogenetic and Haplotype Analyses Offer Insights Towards Conservation of Pteroptyx tener Olivier, 1907 (Coleoptera: Lampyridae) Populations from Malaysia

The synchronous-flashing firefly, Pteroptyx tener Olivier, 1907 is a congregating firefly with high economic value for the ecotourism industry. However, the populations of P. tener are increasingly being threatened due to habitat destruction. The assessment of genetic variation within and among populations of the species from Malaysia might play a major role in conservation and management programs. Objectives of this study were to investigate the relationships among P. tener from five populations in Malaysia (Peninsular Malaysia and Borneo (Sabah) using cytochrome oxidase subunit I (COI), and to obtain its haplotype information. A total of 50 sequences of P. tener were used in the Neighbour-Joining (NJ), Maximum Parsimony (MP), and Bayesian Inference (BI) phylogenetic analyses and successfully formed several distinct clades of West and East Malaysia (East Coast and West Coast of Peninsular Malaysia) with moderately to highly supported bootstrap values and posterior probabilities. Several haplotype analysis parameters are presented; low nucleotide diversity (PiT, 0.01893), haplotype number (Hn, 19), and high haplotype diversity (Hd, 0.89469). Our findings assumed that the geographical isolation due to the inability for flying long distances without help by the wind and flooding in tidal rivers. The data presented here are important genetic information on the P. tener populations and are important for the conservation programme in Malaysia by conducting the breeding program among populations that have similar genetic information.

Population structure and genetic diversity of tobacco mild green mosaic virus variants in Western Anatolia of Turkey

Tobamoviruses have been pioneers in understanding the population genetic structure, host-vector interactions, and evolutionary processes of viruses. To better understand the population genetic structure and molecular evolutionary relationships of tobacco mild green mosaic virus (TMGMV), a significant species of tobamoviruses, extensive analyses were performed using bioinformatics tools in this study. 300 samples were collected from plants exhibiting viruses and virus-like symptoms from Turkey's largest tobacco cultivation areas during the plant vegetation period between 2019 and 2020. Samples were tested using conventional molecular techniques for tobamoviruses, including ToBRFV, TMV, TMGMV, ToMV, and ToMMV species. Single and double tobamovirus infections were determined in 258 of 300 samples. Single infections were 225 (75%), 10 (3.33%), and 1 (0.33%) for TMGMV, TMV, and ToMV, respectively. Double infections were 21 (7.0%) and 1 (0.33%) for TMGMV + TMV and TMV + ToMV, respectively. TMGMV-infected samples from each region were chosen, and their complete genomes were revealed. Detailed analyses were carried out from Turkey, with global variants available in GenBank. Molecular evolutionary analyses revealed three main lineages (Clades I, II, and III) at the p126, p183, MP, and complete genome levels and two main lineages at the CP gene (Clades I and II). Variants from Turkey were distributed in two different branches in other phylogenetic trees, except for the CP gene. High haplotype diversity and low nucleotide diversity were determined in each gene region, indicating consistent genetic stability. In addition, purifying selection pressures were determined in specific gene regions. With AMOVA (analysis of molecular variance), it was determined that the source of genetic variation came from within the main lineages, including various hosts and geographies. The differences in lineages were confirmed by independent test statistics. While neutrality tests revealed population expansions in the CP and MP genes, other p183 and p126 genes revealed bottlenecks or balancing selection. The fact that TMGMV was more common than TMV in Turkey strongly supported the phenomenon called “mutational melting” or “Müller latch,” which presumably causes TMV to disappear from its niche.

Determination of genetic diversity between natural and cultured populations of Common Dentex (Dentex dentex) fish in the East Aegean Sea

Common dentex (Dentex dentex) is a commercial species of fish that is a highly valuable food source living naturally near Mediterranean and Atlantic Coasts. Therefore, monitoring and maintaining common dentex habitats are of high importance. A total of 53 specimens were collected from 6 natural habitats (from the Eastern Aegean and from the Antalya coast, which is the closest coast to the Western Mediterranean), and one aquaculture facility in the Eastern Aegean Sea. The mitochondrial Cytochrome Oxidase I (COI) gene was used to detect a total of 26 haplotypes. Along with the COI gene sequence, real-time PCR and highresolution melting analyses were performed as rapid and inexpensive alternatives. Sequence analysis showed that the highest haplotype diversity was obtained from the aquaculture facility in Karaburun and Antalya Locations, Turkey. Analysis of Molecular Variance (AMOVA) based on the haplotype frequencies resulted in 92.54% genetic variation within localities and 7.46% genetic variation between/among localities. The mean fixation index (Fst) was calculated as 0.0746 (p &lt; 0.001). Genetic distances were primarily in collaboration with geographical distances and were efficiently confirmed by high resolution melting (HRM) analysis. The results will be valuable in monitoring and maintaining natural habitats as well as aquaculture facilities where common dentex are grown.