Molecular Haplotyping Research Articles

Molecular characterization, haplotype analysis and development of markers specific to dzs18 gene regulating methionine accumulation in kernels of subtropical maize.

The online version contains supplementary material available at 10.1007/s13205-024-04088-2.

Molecular characterization and haplotypes of hydatid cyst isolates collected from humans and ruminants in Setif Province (northeast of Algeria) based on mitochondrial cytochrome C oxidase subunit 1 (mt-CO1) gene sequences.

Cystic echinococcosis (CE) is a disease that can be transmitted from animals to humans, caused by the metacestode of Echinococcus granulosus. The disease has significant health and economic impacts worldwide, particularly in endemic areas. The study aimed to evaluate the prevalence of hydatid cysts in ruminants (cattle and sheep) (n = 2060) from the Setif Province of Algeria using microscopy. The results showed that hydatid cysts were detected in 9.6% (198/2060) of ruminants, with a higher prevalence in cattle (16.8%; 56/333) compared to sheep (8.2%; 142/1727). Molecular techniques were used to analyze a subset of animals consisting of 30 sheep and 4 cattle. Specifically, a fragment of the mitochondrial cytochrome c oxidase subunit 1 (mt-CO1) gene was sequenced and compared to sequences from seven humans from the same region. The results indicated that all isolates were identified as E. granulosus sensu stricto. Haplotype analysis identified 19 E. granulosus s.s. haplotypes arranged like a star, with the dominant haplotype (Hap04) at the center. Hap04 has been assigned a total of 17 positives, including positives from sheep, cattle, and two humans. This study is noteworthy for being the first to use a molecular approach to human and ruminant echinococcosis in Setif, a significant breeding region in Algeria.

Delineation of genes for a major QTL governing heat stress tolerance in chickpea.

Chickpea (Cicer arietinum) is a cool season grain legume experiencing severe yield loss during heat stress due to the intensifying climate changes and its associated gradual increase of mean temperature. Hence, understanding the genetic architecture regulating heat stress tolerance has emerged as an important trait to be addressed for enhancing yield and productivity of chickpea under heat stress. The present study is intended to identify the major genomic region(s) governing heat stress tolerance in chickpea. For this, an integrated genomics-assisted breeding strategy involving NGS-based high-resolution QTL-seq assay, QTL region-specific association analysis and molecular haplotyping was deployed in a population of 206 mapping individuals and a diversity panel of 217 germplasm accessions of chickpea. This combinatorial strategy delineated a major 156.8 kb QTL genomic region, which was subsequently narrowed-down to a functional candidate gene CaHSFA5 and its natural alleles associated strongly with heat stress tolerance in chickpea. Superior natural alleles and haplotypes delineated from the CaHSFA5 gene have functional significance in regulating heat stress tolerance in chickpea. Histochemical staining, interaction studies along with differential expression profiling of CaHSFA5 and ROS scavenging genes suggest a cross talk between CaHSFA5 with ROS homeostasis pertaining to heat stress tolerance in chickpea. Heterologous gene expression followed by heat stress screening further validated the functional significance of CaHSFA5 for heat stress tolerance. The salient outcomes obtained here can have potential to accelerate multiple translational genomic analysis including marker-assisted breeding and gene editing in order to develop high-yielding heat stress tolerant chickpea varieties.

DNA barcoding, an effective approach for the identification of thrips (Thysanoptera: Thripidae) in citrus orchards

AbstractThrips are serious crop pests with a worldwide distribution. The first step in the design of control methods is accurate species identification. However, the morphological identification of thrips can be complex, especially as there is morphological variation amongst individuals of the same species. Analyzing a fragment of cytochrome c oxidase subunit I (COI) has been used successfully in the identification of insects and other arthropods. Citrus is among the most important crops in the world, and Mexico is one of the largest producers. Despite this, formal reports on the species diversity of thrips found in Mexican citrus orchards are scarce. We explored the species diversity of thrips in selected citrus orchards (of Mexican orange, Key lime and Persian lime), based on morphological taxonomy and on partial sequences of COI. We analyzed 111 thrips specimens, distributed in two genera: Frankliniella, including Frankliniella insularis, Frankliniella invasor and Frankliniella occidentalis; and Scirtothrips, including Scirtothrips citri and Scirtothrips dorsalis. Frankliniella insularis and F. invasor were the most abundant species in Mexican orange (Citrus × sinensis) and Persian lime (Citrus × latofolia) orchards. Scirtothrips dorsalis was the most abundant species in Key lime orchards (Citrus × aurantiifolia). Analysis of molecular variance (AMOVA) and haplotype analyses performed on F. insularis sequences (the most abundant species) showed no significant effect of geographical origin or plant host on its genetic population structure. DNA barcoding is a valuable method to complement the morphological identification of thrips in citrus orchards, allowing more accurate identification to species level.

Sample design in biodiversity studies matters: a fine-scale study of Lawrence's velvet worm, Peripatopsis lawrencei (Onychophora: Peripatopsidae), reveals hidden diversity.

A fine-scale phylogenetic and phylogeographic analysis of Peripatopsis lawrencei s.l. was conducted with both mitochondrial and nuclear DNA sequence data, using both external morphology and scanning electron microscopy of taxonomically important characters. A total of 119 sequences were used for the mitochondrial cytochrome c oxidase subunit I (COI ) whereas a single representative specimen from each locality was sequenced for the nuclear 18S rRNA locus. Phylogenetic analyses were conducted on the total COI data set and the combined COI + 18S rRNA data set using a Bayesian analysis and maximum likelihood analyses. For the combined DNA sequence data set, a divergence time estimation was further undertaken in BEAST and specimens placed in a phylogenetic framework including all the described Peripatopsis species from South Africa. In addition, a phylogeographic study was conducted exclusively on P. lawrencei s.s. (clade A) using an analysis of molecular variance and haplotype network. Phylogenetic results indicated that, at the Oubos sample locality, two highly distinct genetic lineages were present (clades A and B), whereas a divergence time estimation suggests a Miocene cladogenesis of the novel Oubos lineage. Marked phylogeographic structure was observed for P. lawrencei s.s. (restricted to clade A) across the distribution range with limited maternal dispersal. Morphologically, the two sympatric lineages at Oubos A and B differed in leg pair number, ventral colour and dorsal scale rank counts, as evident from scanning electron microscopy. Our results support the recognition of a distinct species that occurs in sympatry with P. lawrencei s.s. The new species, P. aereus sp. nov. (clade B) is described and the implication for fine-scale taxonomic studies on saproxylic taxa is discussed. ZooBank: urn:lsid:zoobank.org:pub:AB6E0BDA-7B5F-4FD3-A863-BA7C814E278C.

Contribution to the knowledge of the distribution of <i>Parmelia</i> species (Parmeliaceae, Ascomycota) – New records confirmed by molecular data

The first records of<i> Parmelia ernstiae</i> from Madeira, as well as the new localities of <i>P. encryptata</i> from Poland and <i>P. sulcata</i> from Chile, are presented. All records are confirmed by molecular data, BLAST search, and haplotype network analysis of the nucITS rDNA sequences. The discussion on the morphology and secondary chemistry of all three species is provided with notes on their distribution and similar taxa. <i>Parmelia encryptata</i> is a rarely reported cryptic species that is morphologically indistinguishable from <i>P. sulcata</i>, and its identification requires molecular data. New localities of <i>P. encryptata</i> from northern Poland suggest that the taxon may be widespread in the country and not confined to large forests but also present in open areas on free-standing trees. Moreover, a new nucITS rDNA haplotype of the species is reported. <i>Parmelia ernstiae</i> is a near-cryptic species, morphologically and chemically very similar to <i>P. saxatilis</i> and <i>P. serrana</i>. Two specimens of the species from Madeira have sublinear lobes with marginal and laminal isidia, unlike most specimens of P. ernstiae, having short, broad lobes with laminal isidia, and thus are morphologically closer to <i>P. saxatilis</i> in lobe shape and to <i>P. serrana</i> in placement of isidia. However, the BLAST search and haplotype analysis of nucITS rDNA sequences placed these specimens in <i>P. ernstiae</i>. <i>Parmelia sulcata</i>, a common species in the Northern Hemisphere, is rare elsewhere, and only two records confirmed by molecular data were known from South America. The nucITS rDNA haplotype of two new and two previous records of the species represent the most common haplotype of the species known from the Northern Hemisphere.

Seasonal malaria chemoprevention in a context of high presumed sulfadoxine-pyrimethamine resistance: malaria morbidity and molecular drug resistance profiles in South Sudan

BackgroundSeasonal malaria chemoprevention (SMC) using sulfadoxine-pyrimethamine plus amodiaquine (SP-AQ), is a community-based malaria preventive strategy commonly used in the Sahel region of sub-Saharan Africa. However, to date it has not been implemented in East Africa due to high SP resistance levels. This paper is a report on the implementation of SMC outside of the Sahel in an environment with a high level of presumed SP-resistance: five cycles of SMC using SPAQ were administered to children 3–59 months during a period of high malaria transmission (July–December 2019) in 21 villages in South Sudan.MethodsA population-based SMC coverage survey was combined with a longitudinal time series analysis of health facility and community health data measured after each SMC cycle. SMC campaign effectiveness was assessed by Poisson model. SPAQ molecular resistance markers were additionally analysed from dried blood spots from malaria confirmed patients.ResultsIncidence of uncomplicated malaria was reduced from 6.6 per 100 to an average of 3.2 per 100 after SMC administration (mean reduction: 53%) and incidence of severe malaria showed a reduction from 21 per 10,000 before SMC campaign to a mean of 3.3 per 10,000 after each cycle (mean reduction: 84%) in the target group when compared to before the SMC campaign. The most prevalent molecular haplotype associated with SP resistance was the IRNGE haplotype (quintuple mutant, with 51I/59R/108N mutation in pfdhfr + 437G/540E in pfdhps). In contrast, there was a low frequency of AQ resistance markers and haplotypes resistant to both drugs combined (< 2%).ConclusionsThe SMC campaign was effective and could be used as an additional preventive tool in seasonal malaria settings outside of the Sahel, especially in areas where access to health care is unstable. Malaria case load reduction was observed despite the high level of resistance to SP.

Invasive flower thrips, Thrips parvispinus (Karny) occurrence, host expansion and genetic diversification in a tropical poly-crop ecosystem.

Invasive black flower thrips Thrips parvispinus (Karny) has recently emerged as a significant threat to Indian chilli production. Identifying T. parvispinus became difficult due to the complex presence of thrips species in Indian chilli and allied ecosystems. Pest management success depends on assessing invasive pests genetic populations and their distribution in newly habituated areas. The current study investigated the genetic diversity and phylogeographic structure of T. parvispinus across major chilli-growing zones representing different agro-climatic conditions in Tamil Nadu. The species-specific chaetotaxy characteristics of T. parvispinus and molecular analysis of the mtCOI gene were used to confirm that the species T. parvispinus has expanded rapidly in three regions (North Western, Western and Cauvery delta), sparsely in one (Southern) and absent from two (hilly and high rainfall). Fifteen allied crops in chilli/capsicum growing tracts served as host plants for T. parvispinus. A shrub species, namely Littleleaf boxwood, Buxus microphylla Siebold & Zucc., is described as a host plant for the first time. On capsicum and chilli, T. parvispinus and Scirtothrips dorsalis coexisted. Thrips palmi, T. tabaci, Frankliniella schultzei, and Microcephalothrips abdominalis co-occurring alongside T. parvispinus on allied crops. Molecular characterization and haplotype identification help define the genetic composition of T. parvispinus and serve as a foundation for efficient monitoring and creation of Integrated Pest Management (IPM) strategies. As a result, the genetic data presented in this work strongly argues that T. parvispinus as a population is resolving itself towards a fixed state through natural selection that spans its native range globally along with low genetic diversity [Hd: 0.771].

Functional allele of a MATE gene selected during domestication modulates seed color in chickpea.

Seed color is one of the key target traits of domestication and artificial selection in chickpeas due to its implications on consumer preference and market value. The complex seed color trait has been well dissected in several crop species; however, the genetic mechanism underlying seed color variation in chickpea remains poorly understood. Here, we employed an integrated genomics strategy involving QTL mapping, high-density mapping, map-based cloning, association analysis, and molecular haplotyping in an inter-specific RIL mapping population, association panel, wild accessions, and introgression lines (ILs) of Cicer gene pool. This delineated a MATE gene, CaMATE23, encoding a Transparent Testa (TT) and its natural allele (8-bp insertion) and haplotype underlying a major QTL governing seed color on chickpea chromosome 4. Signatures of selective sweep and a strong purifying selection reflected that CaMATE23, especially its 8-bp insertion natural allelic variant, underwent selection during chickpea domestication. Functional investigations revealed that the 8-bp insertion containing the third cis-regulatory RY-motif element in the CaMATE23 promoter is critical for enhanced binding of CaFUSCA3 transcription factor, a key regulator of seed development and flavonoid biosynthesis, thereby affecting CaMATE23 expression and proanthocyanidin (PA) accumulation in the seed coat to impart varied seed color in chickpea. Consequently, overexpression of CaMATE23 in Arabidopsis tt12 mutant partially restored the seed color phenotype to brown pigmentation, ascertaining its functional role in PA accumulation in the seed coat. These findings shed new light on the seed color regulation and evolutionary history, and highlight the transcriptional regulation of CaMATE23 by CaFUSCA3 in modulating seed color in chickpea. The functionally relevant InDel variation, natural allele, and haplotype from CaMATE23 are vital for translational genomic research, including marker-assisted breeding, for developing chickpea cultivars with desirable seed color that appeal to consumers and meet global market demand.

Mitochondrial DNA reveals two recent diverged lineages in Amphioctopusaegina (Gray, 1849) (Cephalopoda, Octopodidae) across the Leizhou Peninsula: a marine ecoregion barrier.

Amphioctopusaegina is an economically important species that has been intensively exploited in the marine areas along the Chinese coast. However, the genetic variation and population genetic structure, which would provide valuable information for their fisheries management, have rarely been investigated. In this study, the genetic variation within and among four A.aegina populations throughout its full distribution range were estimated based on mitochondrial cytochrome b DNA sequences. Our results indicated low (Qinzhou) to high (Dongshan) genetic diversities among the four populations. Analysis of molecular variance (AMOVA), ΦST statistics, phylogenetic tree and haplotype networks revealed two significant (p < 0.01) divergent lineages with a ΦST value of 0.7116 between them, one from a population in Qinzhou and the other from the remaining three populations of Dongshan, Huizhou and Zhanjiang. However, the low genetic distance (0.0032) and only two fixed substitutions between them suggest their recent divergence is possibly due to the last glacial period barriers to gene flow produced by the Leizhou Peninsula. The observed lineage divergence suggests that populations of A.aegina in China are genetically subdivided and may represent evolutionary lineages that should be managed individually.

Natural alleles of Mediator subunit genes modulate plant height in chickpea.

SUMMARYPlant height (PH) is an important plant architectural trait targeted during Green Revolution to enhance crop yields. Identification of genes and natural alleles governing plant height without compromising agronomic performance can fill the lacuna of knowledge connecting ideal plant architecture with maximum achievable yield in chickpea. Through coherent strategy involving genome‐wide association study, QTL/fine mapping, map‐based cloning, molecular haplotyping, and downstream functional genomics, the current study identified two Mediator subunit genes namely, CaMED23 and CaMED5b and their derived natural alleles/haplotypes underlying the major QTLs and trans‐acting eQTLs regulating plant height in chickpea. Differential accumulation of haplotype‐specific transcripts of these two Mediator genes in corresponding haplotype‐introgressed near‐isogenic lines (NILs) correlates negatively with the plant height trait. Quantitative as well as qualitative estimation based on histology, scanning electron microscopy, and histochemical assay unraveled the reduced lengths and cell sizes of internodes along with compromised lignin levels in dwarf/semi‐dwarf chickpea NILs introgressed with superior CaMED23 and CaMED5b gene haplotypes. This observation, supported by global transcriptome profiling‐based diminished expression of various phenylpropanoid pathway genes upstream of lignin biosynthesis in dwarf/semi‐dwarf NILs, essentially links plant height with lignin accumulation. The identified molecular signatures in the Mediator subunit genes can be efficiently utilized to develop desirable dwarf/semi‐dwarf‐type chickpea cultivars without affecting their yield per plant via modulating lignin/phenylpropanoid biosynthesis.

Contrasting results of multiple species delimitation approaches cause uncertainty in synecological studies: A case study on Sri Lankan chafers

Abstract Biodiversity patterns are the sum of multiple overlapping species distributions. Their analysis therefore requires proper species inference. DNA‐based species delimitation has become increasingly popular for such assessments, but their robustness is often problematic due to incongruence between multiple delimitation approaches. Here, we explored how contrasting results of different species’ delimitations translate into conclusions of synecological studies, exemplified by assemblages of phytophagous scarab beetles in Sri Lanka from different elevations and forest types. Particularly, we compared estimates based on complete assemblages and on cumulated species inventories inferred from individually analysed subclades. The patterns of assemblage similarity were analysed across different spatial scales with reference to morphospecies, molecular operational taxonomic units (MOTUs) and haplotypes. Method‐related ambiguity of species (MOTUs) estimates, which also included subclade inferences, severely affected the certainty of apparent biodiversity patterns at most spatial scales. Even more contrasting patterns resulted from individual clade‐wise analyses of faunal similarity or even from cumulated species inventories from individual clade‐wise species delimitation analysis. In this case study of tropical chafer beetles, haplotypes provided only very little explanatory information, since genetically highly diverse populations widely lacked shared haplotypes. Therefore, searching proper species boundaries should be the ultimate goal of biodiversity assessments to lend an enduring meaning to biodiversity research and its sustainable application. Our study underlines the need for awareness when synecological observations from different studies are integrated, which use different species delimitation methods for their biodiversity assessment, and its potential impact on conservation management.

Global phylogeographical distribution of Gloeoporus dichrous.

Phylogeographic analyses are efficient in ecological and evolutionary studies to discover the origin of a lineage, its dispersal routes, and the divergence of ancestral traits. Studies on widespread wood-decay fungi have revealed the phylogenetic division of several polypores based on geographical distribution. In this study, specimens of Gloeoporus dichrous, a cosmopolitan polypore species, were collected globally and analyzed for their geographic distribution. Multi-marker Bayesian molecular clock and haplotype analyses revealed a clear division of G. dichrous populations by continent. The species diverged from its neighboring clades 10.3 (16.0-5.6) million years ago, with Asian and North American populations at the center of divergence. Possible dispersal mechanisms and pathways are predicted and discussed based on the evaluated transfer routes. The biogeography of G. dichrous analyzed in this study represents a fraction of the polypore evolution and may advance the understanding of the overall evolution of wood-decay fungi.

Molecular Characterization and Haplotype Analysis of Low Phytic Acid-1 (lpa1) Gene Governing Accumulation of Kernel Phytic Acid in Subtropically-Adapted Maize

Maize is an important food, feed, fodder and industrial crop in addition to being a valuable source of micronutrients. Phytic acid (PA), an anti-nutritional factor in maize, makes crucial minerals inaccessible to monogastric animals. The low phytic acid-1 (lpa1) gene located on chromosome-1S is 7292 bp long with 11 exons, and the recessive lpa1-1 allele reduces the accumulation of PA thereby enhances the bioavailability of essential minerals in maize kernels. Here, we characterized the full-length Lpa1 gene sequence in three mutants (lpa1-1) and seven wild-type (Lpa1) maize inbreds. Sequence analysis revealed 607 polymorphic sites across Lpa1 sequences, indicating wide variability for Lpa1 among the inbreds. Further, SNP from “C” to “T” differentiated wild-type and mutant-type alleles at 1432 amino acid position. Gene-based diversity among 48 diverse maize inbreds using 15 InDel markers revealed the formation of 42 distinct haplotypes; six of which (Hap6, Hap16, Hap17, Hap19 Hap27 and Hap31) were shared by more than one genotype. The number of exons in Lpa1 ranged from 11–19 among maize genotypes and 6–14 among 26 orthologues. Major functional motifs of Lpa1 detected were ATP-binding Cassette (ABC) transporter trans-membrane region and ABC transporter. Phylogenetic tree using nucleotide and protein sequences revealed a closer relationship of maize Lpa1 sequences with Sorghum bicolor, Panicum hallii, Setaria italica and S. viridis. This study offered newer insights into the understanding of the genetic diversity of the Lpa1 gene in maize and related crop-species, and information generated here would further help in exploiting the lpa1 mutant for the enhancement of nutritional value in maize kernels.

Lasiodiplodia: Generic revision by providing molecular markers, geographical distribution and haplotype diversity

Lasiodiplodia: Generic revision by providing molecular markers, geographical distribution and haplotype diversity

A superior gene allele involved in abscisic acid signaling enhances drought tolerance and yield in chickpea.

Identifying potential molecular tags for drought tolerance is essential for achieving higher crop productivity under drought stress. We employed an integrated genomics-assisted breeding and functional genomics strategy involving association mapping, fine mapping, map-based cloning, molecular haplotyping and transcript profiling in the introgression lines (ILs)- and near isogenic lines (NILs)-based association panel and mapping population of chickpea (Cicer arietinum). This combinatorial approach delineated a bHLH (basic helix-loop-helix) transcription factor, CabHLH10 (Cicer arietinum bHLH10) underlying a major QTL, along with its derived natural alleles/haplotypes governing yield traits under drought stress in chickpea. CabHLH10 binds to a cis-regulatory G-box promoter element to modulate the expression of RD22 (responsive to desiccation 22), a drought/abscisic acid (ABA)-responsive gene (via a trans-expression QTL), and two strong yield-enhancement photosynthetic efficiency (PE) genes. This, in turn, upregulates other downstream drought-responsive and ABA signaling genes, as well as yield-enhancing PE genes, thus increasing plant adaptation to drought with reduced yield penalty. We showed that a superior allele of CabHLH10 introgressed into the NILs improved root and shoot biomass and PE, thereby enhancing yield and productivity during drought without compromising agronomic performance. Furthermore, overexpression of CabHLH10 in chickpea and Arabidopsis (Arabidopsis thaliana) conferred enhanced drought tolerance by improving root and shoot agro-morphological traits. These findings facilitate translational genomics for crop improvement and the development of genetically tailored, climate-resilient, high-yielding chickpea cultivars.

Low Genetic Diversity Study on Leopard Coral Grouper Plectropomus leopardus (Perciformes: Serranidae) from Sulawesi, Indonesia

Highlight ResearchThe leopard coral grouper Plectropomus leopardus was identified and analysed based on molecular approach.Genetic diversity within two regions in Gorontalo, Sulawesi successfully performed using connectivity analysis.Three haplotypes of Plectropomus leopardus from two region in Gorontalo as one of economical important marine fish species. AbstractBar-cheek coral trout (P. leopardus) is the flagship of the grouper in the live fish market in Asia. Unfortunately, the potential of the grouper is still partly produced from natural catches. Even though hybridisation activities have also started to be carried out, there still have not been many studies on the genetic diversity of these fish. The application of molecular identification has been widely applied in marine aquatic animal species, which are very likely to occur due to errors in terms of shape and colour in the morphological character. DNA information has been beneficial in efforts to the breeding program and develop grouper aquaculture activities. DNA barcoding was used for the molecular identification and haplotype analysis of P. leopardus from two locations in Gorontalo, Sulawesi, Indonesia. A total of 14 fish samples were collected from two traditional fish markets around Kwandang and Sumalata Gulf in the northern part of Gorontalo Province, Sulawesi. This study identified and found three haplotypes from both regions. Molecular identification using Cytochrome c Oxidase subunit I (COI) gene region on mitochondrial DNA. Besides Mega7 for phylogenetic reconstruction, the data analysis using DnaSP6, Arlequin Ver.3.5.2.2, and Network 5.0.1.1. The first Haplotype is a mixed population between the Kwandang Gulf and the Sumalata Gulf, then the Kwandang Gulf haplotype and the Sumalata Gulf haplotype. The genetic distance between Kwandang Gulf haplotype and Sumalata Gulf haplotype is 0.003984, classified as a shallow genetic distance and needs more samples from another region to figure out leopard coral grouper around Indonesia.

Molecular diversity, haplotype distribution and genetic variation flow of Bipolaris sorokiniana fungus causing spot blotch disease in different wheat-growing zones.

Bipolaris sorokiniana (BS) is an economically important fungal pathogen causing spot blotch of wheat (Trtiticum aestivum) and found in all wheat-growing zones of India. Very scanty and fragmentary information is available on its genetic diversity. The current research is the first detailed report on the geographic distribution and evolution of BS population in five geographically distinct wheat-growing zones (North Western Plain Zone (NWPZ), North Eastern Plain zone (NEPZ), North Hill Zone (NHZ), Southern Hill Zone (SHZ) and Peninsular Zone (PZ)) of India, studied by performing nucleotide sequence comparison of internal transcribed spacer region of 528 isolates. A moderate to low levels of haplotypic diversity was noticed in different wheat-growing zones. Phylogenetic analysis suggests that B. sorokiniana exist in two distinct lineages as all isolates under study were grouped in two different clades and found analogous to the findings of haplotypic and TCS network analysis. The genetic parameters revealed the existence of 40 haplotypes with three major haplotypes (H-1, H-2 and H-3) which showed star-like structure network surrounded by several single haplotypes, revealing high frequency of the mutations (Eta = 2 - 158) in total analyzed population. H-1 was observed as a predominant haplotype and prevalent in all the five zones. Moderate level of genetic differentiation was found between NHZ and other zones like NWPZ (Fst = 0.332) and SHZ (Fst = 0.382) and PZ (Fst = 0.299), whereas it was low between NEPZ and PZ (Fst = 0.034). Higher transfer rate of genetic variation was noticed between NEPZ and PZ (Nm = 7.06), while it was found minimum between NHZ and SHZ (Nm = 0.40). Moreover, negative score of neutrality statistics (Tajima's D and Fu's FS test) for NWPZ population suggested recent population expansion. However, positive score for both the neutrality tests observed in NEPZ indicated the dominance of balancing selection in structuring their population. Recombination events were observed in the NWPZ and NHZ population, while it was absent in SHZ, NEPZ and PZ population. Thus, the lack of any specific genetic population structure in all the zones indicates for the expansion history only from one common source population, i.e. NWPZ, a mega zone of wheat production in India. Overall, it seems that the predominance of individual haplotypes with a moderate level of genetic variation and human-mediated movement of contaminated seed and dispersal of inoculum, mutations and recombination as prime evolutionary processes play essential role in defining the genetic structure of BS population.

Molecular Identification and Novel Mitochondrial COI Gene Haplotypes of Nesting Kemp’s Ridley Turtles (Lepidochelys kempii) in Rancho Nuevo Sanctuary, Mexico

One hundred new COI sequences of nesting female Kemp’s ridley turtles (Lepidochelys kempii) were obtained in the Rancho Nuevo Sanctuary (RNS). The COI sequences were analyzed and contrasted with others retrieved from BOLD and GenBank with the aim of investigating genetic variability, genetic divergence, and haplotypes of the nesting female population at RNS. Four new COI haplotypes for Kemp’s ridley were described; two are redundant with (LK-RN01) 97 and (LK-COI-01) 17 specimens belonging to the RNS and other localities, respectively. Nucleotide (0.00052) and haplotype (0.303) diversity showed low and conserved COI values The fixation index (FST) between these main redundant haplotypes showed a high degree of differentiation with ~1. Genetic divergence clearly demonstrated two different Kemp’s ridley nesting populations, one from RNS and a second outside Mexico. Phylogenetic COI analysis was useful to differentiate these redundant (LK-COI-01 and RNS LK-RN01) haplotypes and, therefore, these different Kemp’s ridley populations. In addition, phylogenetic COI analysis clearly separates Kemp’s ridley turtles from other sea turtle species, supporting its use as a barcode marker.

Characterization of a novel activating protein-1 (AP-1) gene and the association of its single nucleotide polymorphisms with vibrio resistance in Tegillarca granosa

The blood clam Tegillarca granosa is a commercial marine bivalve of economic value, accounting for approximately 50% of clam production in China. In recent years, the yield of blood clams has been threatened by bacterial infections caused by marine Vibrio species that thrive under a rising sea temperature. The transcription factor activating protein-1 (AP-1) is emerging as an important player in the innate immunity of marine bivalves against viral or bacterial infections. In this study, the full-length cDNA of a novel T. granosa AP-1 (TgAP-1) was cloned for the first time. The 1591-bp cDNA encoded a protein of 292 amino acid residues with a calculated molecular weight of 32.8 kDa. The TgAP-1 protein contained an N-terminal Jun domain and a C-terminal basic region leucine zipper domain typically found in Jun proteins (a subfamily of AP-1 proteins). TgAP-1 was ubiquitously expressed in T. granosa, with the highest expression detected in the gill and foot, followed by the mantle, hemolymph, and hepatopancreas. Exposure to Vibrio harveyi induced TgAP-1 expression in gill tissues and the expression levels of TgAP-1 of resistant blood clams were always lower than that of control population whether Vibro infection or not. A total of 18 single nucleotide polymorphisms (SNPs) of TgAP-1 were detected in T. granosa. SNP-typing and haplotyping of resistant and susceptible populations revealed that six SNPs (AG type of TgSNP-1, GA type of TgSNP-2, TG type of TgSNP-4, CT type of TgSNP-7, AG type of TgSNP-11, and GA type of TgSNP-12) and four haplotypes (fHap2, fHap3, fHap6, and fHap7) were significantly associated with V. harveyi resistance. Risk assessment showed that fHap2 (CG) and fHap7 (GA) were associated with an increased resistance, while fHap3 (CT) and fHap6 (AG) were associated with an increased susceptibility. The results from this study supported a potential role of TgAp-1 in the anti-Vibro immunity of T. granosa. The discovery of the genetic molecular markers and haplotypes related to Vibrio resistance can provide guidance for selective breeding of T. granosa in the future.