Intraspecific Genetic Distances Research Articles

Combination of c oxidase subunit I based deoxyribonucleic acid barcoding and HPLC techniques for the identification and quality evaluation of Pheretima aspergillum.

This study aimed to identify Pheretima aspergillum (Guang-Pheretima) and its adulterants using the cytochrome c oxidase subunit I based deoxyribonucleic acid barcoding technology, and further to evaluate their quality using an optimized high-performance liquid chromatography method. For deoxyribonucleic acid barcoding identification, the Kimura-2-Parameter model was used to analyze genetic distance, and phylogenetic neighbor-joining tree was constructed for species identification of 20 labeled Guang-Pheretima samples. A high-performance liquid chromatography method was developed for the simultaneous determination of seven nucleoside components for quality evaluation. Compared with the GenBank database, 10 samples were identified as real Guang-Pheretima (P. aspergillum), and the others as the adulterants-Metaphire magna. The maximum intraspecific genetic distances of c oxidase subunit I sequence for P. aspergillum were smaller than the minimum interspecific genetic distances between P. aspergillum and M. magna. Ten P. aspergillum and 10 M. magna samples were clearly clustered in the neighbor-joining tree. The contents of seven nucleosides components in P. aspergillum were significantly higher than that in its adulterant-M. magna. The incidence of adulterants for Guang-Pheretima was high (up to 50%) with an alarming quality. This study provided a powerful idea for the quality evaluation of other highly valuable plant- or animal-derived products for safety concerns to avoid misidentification.

Assessment of Genetic Variation in Apis nigrocincta (Hymenoptera: Apidae) in Sulawesi Revealed by Partial Mitochondrial Cytochrome Oxidase I Gene Sequences.

Asian cavity-nesting honey bee Apis nigrocincta, a native bee species from Sulawesi and the Philippines, plays a vital role in pollinating flowering plants in local ecosystem and agriculture. In this study, we assessed the intraspecific genetic variation of A. nigrocincta using the sequence of cytochrome c oxidase subunit I (COI). Molecular phylogenetic analysis showed that there were three main clades in A. nigrocincta specimens from Sulawesi based on their respective locations (North, Central, and South Sulawesi). Genetic distance analysis using the Kimura 2-parameter (K2P) model showed that the intraspecific genetic distance in Sulawesi specimens ranged from 0.000 to 0.055. There are 26 nucleotide polymorphic sites within Sulawesi A. nigrocincta. The variation was dominated by transition T ↔ C. The molecular identification result was supported by morphological identification. The results of the two methods agree that the specimen under study was A. nigrocincta. The result of genetic distance calculation indicated that although the tested specimens were derived from remote locations, the genetic variation was still within the range of intraspecific variation.

Identification of Black Bullhead (Ameiurus melas) and Brown Bullhead (A. nebulosus) from the Western Great Lakes: Recommendations for Small Individuals

Distinguishing Black Bullhead (Ameiurus melas) from Brown Bullhead (A. nebulosus) can be difficult, in part, due to conflicting evidence regarding reliable meristic and morphological characters with which to base identifications. The possibility of hybrids with intermediate traits further complicates diagnosis. Identification guides have typically focused on adults, leaving guidance for juveniles largely undescribed. We compared morphological identifications of Black Bullhead and Brown Bullhead with a cytochrome c oxidase I (COI) barcode and microsatellite assignments. Specifically, we examined: (1) gill-raker count, (2) anal-fin ray count, (3) prominence of pectoral-spine serrae, (4) DNA barcoding using the COI gene, and (5) cluster analysis of nine microsatellite loci. With the exception of anal-fin ray counts, we found a high degree of agreement among all methods. Microsatellite analysis identified two clearly distinct genetic groups, and these groups corresponded to individuals identified as Black Bullhead and Brown Bullhead based on morphology. Although Black Bullhead and Brown Bullhead did not share COI haplotypes (except for one individual), we found greater intraspecific genetic distances among individuals identified as Brown Bullhead than between Brown Bullhead and Black Bullhead, demonstrating that caution is warranted against using COI haplotypes as a barcode to solely distinguish the two species. Pectoral-spine serrae prominence was a useful character to distinguish the two species and worked especially well on individuals <125 mm total length. Gill-raker counts of euthanized fish in the laboratory were appropriate for the identification of all sizes of Black Bullhead and Brown Bullhead (using the lowest number of gill rakers on the first gill arch if asymmetry was observed). Finally, microsatellites did not show evidence of recent hybridization at our sampling sites. We recommend using pectoral-spine serrae prominence to distinguish between Black Bullhead and Brown Bullhead in the field and using gill-raker counts for laboratory based identification.

Disentangling the taxonomy of the subfamily Rasborinae (Cypriniformes, Danionidae) in Sundaland using DNA barcodes

Sundaland constitutes one of the largest and most threatened biodiversity hotspots; however, our understanding of its biodiversity is afflicted by knowledge gaps in taxonomy and distribution patterns. The subfamily Rasborinae is the most diversified group of freshwater fishes in Sundaland. Uncertainties in their taxonomy and systematics have constrained its use as a model in evolutionary studies. Here, we established a DNA barcode reference library of the Rasborinae in Sundaland to examine species boundaries and range distributions through DNA-based species delimitation methods. A checklist of the Rasborinae of Sundaland was compiled based on online catalogs and used to estimate the taxonomic coverage of the present study. We generated a total of 991 DNA barcodes from 189 sampling sites in Sundaland. Together with 106 previously published sequences, we subsequently assembled a reference library of 1097 sequences that covers 65 taxa, including 61 of the 79 known Rasborinae species of Sundaland. Our library indicates that Rasborinae species are defined by distinct molecular lineages that are captured by species delimitation methods. A large overlap between intraspecific and interspecific genetic distance is observed that can be explained by the large amounts of cryptic diversity as evidenced by the 166 Operational Taxonomic Units detected. Implications for the evolutionary dynamics of species diversification are discussed.

Molecular identification of Cerithiidae (Mollusca: Gastropod) in Hainan island, China

A number of same species of Cerithiidae are morphologically unlike, whereas most of species in the same genus are morphologically similar and just exhibit subtle differences. It is difficult to identify them by morphological methods alone. DNA barcoding is a modern molecular technique that can be used to identify species accurately, and is particularly helpful when distinguishing morphologically similar species. In order to identify species of Cerithiidae using DNA barcoding technology based on mitochondrial cytochrome oxidase subunit I (COI) and 16S ribosomal RNA (16S rRNA) genes, this study calculated intraspecific and interspecific genetic distance and constructed the phylogenetic trees. A total of 80 COI and 16S rRNA barcode sequences were obtained from 10 species and 3 genera. Some unknown specimens were further identified and a cryptic species may exist in Cerithium traillii, showing that DNA barcoding technology has the potential to discover new species and cryptic species. The phylogenetic trees revealed that all of the cerithiids could converge upon a monophyly with high support values and two genera (Cerithium and Clypeomorus) maybe support the reclassification. It is necessary for traditional morphological methods to combine with the DNA barcoding for classification and identification of Cerithiidae.

African freshwater eel species (Anguilla spp.) identification through DNA barcoding

Freshwater eels (Anguilla spp.) have a long and complex catadromous life cycle. This unique feature, coupled with difficulty in separating species based on morphology, makes them complex targets for conservation. In this study we evaluated the utility of DNA barcoding using cytochrome oxidase I (COI) to delimit the four species of African eels found in the western Indian Ocean region. We collected 76 individual fin clips from the four eel species (A. mossambica, n=51; A. marmorata, n=17; A. bengalensis, n=6; A. bicolor, n=2) in the rivers of KwaZulu–Natal, South Africa, in 2016–18. Phylogenetic analysis of the COI sequences recovered all four species as monophyletic. Barcoding gap analyses were performed and there was no found overlap in inter- and intraspecific genetic distances. Consequently, the use of COI barcoding as an identification tool was found to be reliable for identifying African eels to the species level, which suggests that this marker should be included in future environmental DNA or metabarcoding studies.

Morphological characteristics identification and molecular DNA barcoding analysis of Hippocampus spinosissimus

The combination of morphological characteristics and DNA barcodes was used to a systematic study of Hippocampus spinosissimus,laying the foundation for rapid and accurate identification for the medical seahorse species. According to the reported literature and observation on seahorse samples,the typical characteristics of the H. spinosissimus include highly developed spiny,much short nose,single or double cheeks and strongly developed spines bordering pouch. Genomic DNAs of H. spinosissimus and other related seahorse species were extracted using the TIANamp Marine Animals DNA Kit. The COⅠ and ATP6 genes were amplified and sequenced in both directions. After the verification by Blast,the GC content,intraspecific and interspecific genetic distance,and the Neighbor joining( NJ) phylogenetic trees were analyzed by MEGA 7. The lengths of the COⅠ and ATP6 genes were 649 bp and 602-603 bp,respectively,with the average GC content of 39. 96% and 35. 37%. The maximum intraspecific genetic distances in H. spinosissimus based on COⅠ and ATP were both far less than the minimum interspecific genetic distance between H. spinosissimus and other seahorses,suggesting a significant barcoding gap. NJ analysis results of COⅠ and ATP6 exhibited that all H. spinosissimus species clustered together,indicating that the two DNA barcode could identify H. spinosissimus from other seahorses accurately and quickly. In addition,H. spinosissimus shared a close genetic relationship between H. kelloggi according to the NJ tree. Furthermore,there exits three stable subgroup structure of H. spinosissimus,indicating that COⅠ and ATP6 barcodes could be applied the indicator for the geographical ecology research of H. spinosissimus. The results obtained the typical morphological and molecular identification characteristics of H. spinosissimus,which played central roles for the development of species identification. This study provides an important basis data for expanding the medical seahorse resources and ensuring the safety of clinical medicine.

Population genetic structure and intraspecific genetic distance of Periplaneta americana (Blattodea: Blattidae) based on mitochondrial and nuclear DNA markers.

The American cockroach (Periplaneta americana) is a globally invasive pest that can cause significant economic loss and threaten human health. Although it is abundant and lives in close proximity to humans, few studies have investigated the genetic diversity of P. americana. Our study analyzed 1,053 P. americana and other Periplaneta species' samples from different locations in China and the United States. A traditional tree‐based method using 17 unique mitochondrial COI haplotypes of P. americana and 20 haplotypes of the other Periplaneta species accurately identified P. americana with a barcoding threshold of 5.1%. To identify the population genetic structure of P. americana, we investigated wingless gene and pooled them with obtained mtDNA data for a combined analysis. Although the genetic diversity of the USA group was relatively higher than the China group, the number of haplotypes and alleles of both groups was small. The analysis of molecular variance (AMOVA), intraspecific phylogeny, and haplotype networks indicated that P. americana had very little global genetic differentiation. The weak geographic genetic structure might reflect the human‐mediated dispersal of P. americana. Despite no apparent phylogeographic assignment of mtDNA and nuclear lineages was observed in both BI trees, the integrated COI sequence data identified four distinct P. americana haplotype groups, showing four ancient maternal lineages of P. americana in China and the United States.

DNA barcoding of aphid-associated ants (Hymenoptera, Formicidae) in a subtropical area of southern China.

As one of the most abundant and complex groups of terrestrial insects, ants have associations with many other organismal groups, such as hemipteran insects producing honeydew. With the aim of expanding the knowledge base of ant species associated with aphids, this study analyzed mitochondrial COI barcodes of 301 ant samples for 37 aphid-associated ant species in a subtropical area of southern China. Sequence analyses revealed that the intraspecific and interspecific distances ranged from zero to 7.7%% and 0.2 to 31.7%, respectively. Three barcoding approaches – Automatic Barcode Gap Discovery, Bayesian Poisson Tree Processes and Generalized Mixed Yule-coalescent – were used to help delimit ant species based on COI sequences, and their results corresponded well with most of the morphospecies. All three approaches indicate cryptic diversity may exist within Tetramorium bicarinatum and Technomyrmex albipes, with intraspecific genetic distances of 7.7% and 6.24%, respectively. Our analyses also reported five species for the first time from Fujian Province of China, and the COI sequences of nine species are newly added into the GenBank. This study provides information about species diversity of aphid-associated ants in subtropical China and compiles a DNA barcode reference library for future ant barcoding work.

DNA barcoding of coastal ray-finned fishes in Vietnam.

DNA barcoding based on a fragment of the cytochrome c oxidase subunit I (COI) gene is widely applied in species identification and biodiversity studies. The aim of this study was to establish a comprehensive barcoding database of coastal ray-finned fishes in Vietnam. A total of 3,638 specimens were collected from fish landing sites in northern, central and southern Vietnam. Seven hundred and sixty-five COI sequences of ray-finned fishes were generated, belonging to 458 species, 273 genera, 113 families and 43 orders. A total of 59 species were newly recorded in Vietnam and sequences of six species were new to the Genbank and BOLD online databases. Only 32 species cannot be annotated to species level because difficulty in morphological identifications and their Kimura-2-Parameter (K2P) genetic distances to most similar sequences were more than 2%. Moreover, intra-specific genetic distances in some species are also higher than 2%, implying the existence of putative cryptic species. The mean K2P genetic distances within species, genera, families, orders and classes were 0.34%, 12.14%, 17.39%, 21.42%, and 24.80, respectively. Species compositions are quite different with only 16 common species among northern, central and southern Vietnam. This may attribute to multiple habitats and environmental factors across the 3,260 km Vietnamese coastline. Our results confirmed that DNA barcoding is an efficient and reliable tool for coastal fish identification in Vietnam, and also established a reliable DNA barcode reference library for these fishes. DNA barcodes will contribute to future efforts to achieve better monitoring, conservation, and management of fisheries in Vietnam.

Taxonomic review of Dendrophryniscus brevipollicatus Jiménez de la Espada, 1870, with revalidation of D. imitator (Miranda-Ribeiro, 1920) br />and D. lauroi Miranda-Ribeiro, 1926, and description of four new related species(Anura, Bufonidae).

Dendrophryniscus brevipollicatus Jiménez de la Espada is a Neotropical bufonid endemic to a small range of the Brazilian Coastal Atlantic Forest, with reduced body size for the family and bromeligenous habit. We reviewed the taxonomic status of populations of D. brevipollicatus from states of Rio de Janeiro and São Paulo, including some continental islands, based on external morphology. We tested our morphological species by a DNA-barcoding approach with sequences of 16S RNA ribosomal gene fragment. DNA-barcoding analysis included other recognized Dendrophryniscus species and was concordant with morphological species diagnosed in our review. Intraspecific genetic distances ranged from 0 to 2.72 % (± 0.91 %). Interspecific distances ranged from 3.35 % (± 0.90 %) to 20.15 (± 2.23 %). Optimal threshold values ranged from 2.8 % to 3.0 % and barcode gap analysis showed that for all individuals the furthest intraspecific distances was always lower than the closest non-conspecific individual. Seven distinct species were recognized. A neotype for D. brevipollicatus was designated and described; the type locality was determined for the Açude da Solidão (22º57'S, 43º17'W, Datum WGS 84; 410 m altitude), Parque Nacional da Tijuca, Municipality of Rio de Janeiro, State of Rio de Janeiro, Brazil. Two species were revalidated: D. imitator (Miranda-Ribeiro), with designation of a lectotype and descriptions of the lectotype and of a topotype from the restricted type locality, Alto da Serra (23o46'S, 46o19'W, Datum WGS 84, 800 m altitude), Municipality of Paranapiacaba, State of São Paulo, Brazil; and D. lauroi (Miranda-Ribeiro), with descriptions of the lectotype and of a topotype from the type locality, Municipality of Angra dos Reis (22o54'S, 44o20'W, Datum WGS 84; 25 m altitude), State of Rio de Janeiro, Brazil. Four new species were described: D. davori sp. nov., from Baixo Caledônia (22º21'S, 42º35'W, Datum WGS 84; 1600 m altitude), Municipality of Nova Friburgo, State of Rio de Janeiro, Brazil; D. haddadi sp. nov., from Parque Estadual da Serra do Mar, Núcleo Santa Virgínia (23o21'S, 45o08'W, Datum WGS 84; 970 m altitude), Municipality of São Luís do Paraitinga, State of São Paulo, Brazil; D. izecksohni sp. nov., from Campo de Fruticultura da Bocaina (currently Núcleo Senador Vergueiro), Municipality of São José do Barreiro (22º38'S, 44º34'W, Datum WGS 84, 540 m altitude), State of São Paulo, Brazil; and D. jureia sp. nov., from Estação Ecológica da Juréia-Itatins-Núcleo Rio Verde (24º22'S, 47º04'W, Datum WGS 84; 32 m altitude), Municipality of Iguape, State of São Paulo, Brazil. Geographical distributions of all species are provided.

Indication of ongoing amphipod speciation in Lake Baikal by genetic structures within endemic species

BackgroundThe ancient Lake Baikal is characterized by an outstanding diversity of endemic faunas with more than 350 amphipod species and subspecies. We determined the genetic diversity within the endemic littoral amphipod species Eulimnogammarus verrucosus, E. cyaneus and E. vittatus and investigated whether within those species genetically separate populations occur across Lake Baikal. Gammarus lacustris from water bodies in the Baikal area was examined for comparison.ResultsGenetic diversities within a species were determined based on fragments of cytochrome c oxidase I (COI) and for E. verrucosus additionally of 18S rDNA. Highly location-specific haplogroups of E. verrucosus and E. vittatus were found at the southern and western shores of Baikal that are separated by the Angara River outflow; E. verrucosus from the eastern shore formed a further, clearly distinct haplotype cluster possibly confined by the Selenga River and Angarskiy Sor deltas. The genetic diversities within these haplogroups were lower than between the different haplogroups. Intraspecific genetic diversities within E. verrucosus and E. vittatus with 13 and 10%, respectively, were similar to interspecies differences indicating the occurrence of cryptic, morphologically highly similar species; for E. verrucosus this was confirmed with 18S rDNA. The haplotypes of E. cyaneus and G. lacustris specimens were with intraspecific genetic distances of 3 and 2%, respectively, more homogeneous indicating no or only recent disruption of gene flow of E. cyaneus across Baikal and recent colonization of water bodies around Baikal by G. lacustris.ConclusionsOur finding of separation of subgroups of Baikal endemic amphipods to different degrees points to a species-specific ability of dispersal across areas with adverse conditions and to potential geographical dispersal barriers in Lake Baikal.

Fish diversity of the largest deltaic formation in the Americas - a description of the fish fauna of the Parna\xedba Delta using DNA Barcoding

Deltas are dynamic and productive systems of enormous ecological significance, encompassing unique and biologically diverse wetland habitats. Here, we present the first data on the molecular diversity of the fish fauna of the Parnaíba Delta, the largest deltaic formation of the Americas. Partial sequences (626 bp) of the mitochondrial COI gene (Cytochrome c oxidase subunit I) were used to barcode 402 individuals, representing 128 species, belonging to 98 genera, 57 families, 17 orders and two classes. The most abundant orders were the Perciformes, Siluriformes, Gobiiformes, and Pleuronectiformes. The Neighbor-Joining (NJ), Bayesian Inference (BI), and BIN analyses produced 103 molecular clusters, while the Automatic Barcode Gap Discovery (ABGD) and Maximum Likelihood (ML) approaches revealed 102 clusters. The mean conspecific, congeneric and confamilial genetic distances were 0.33%, 14.37%, and 18.60%, respectively. Intraspecific divergence ranged from 0.0% to 1.4%, and all species presented barcode gaps, with the exception of two clusters of Cathorops spixii (OTU 96 and OTU 103), which were separated by a low interspecific distance (1.2%), which overlaps the maximum intraspecific genetic distance (1.4%). The barcode data provide new insights into the fish diversity of the Parnaíba Delta, which will be important for the development of further research on this fauna.

A new species of black fly (Diptera: Simuliidae) in the Simulium (Simulium) multistriatum species-group from Thailand.

A new black fly species of the Simulium multistriatum species-group of the subgenus Simulium Latreille is described from the mountainous area in northeastern Thailand, based on morphology and mitochondrial DNA sequences. The new species is morphologically similar to S. laui Takaoka and Sofian-Azirun and S. lacduongense Takaoka and Ya'cob originally described from Vietnam, S. fenestratum Edwards originally described from Indonesia and S. chaliowae Takaoka and Boonkemtong originally described from Thailand, but can be distinguished in the adult stage by the number of upper eye facets and globular shape of the spermatheca and in the pupal stage by the cocoon and shape of thoracic tubercles. Genetic distance and phylogenetic analyses of mitochondrial cytochrome c oxidase I sequences differentiated the new species from other members of S. multistriatum species-group. All specimens of the new species formed a monophyletic clade with strong support in all phylogenetic analyses. The minimum interspecific genetic distance of 4.9% is considerably greater than the new species maximum intraspecific genetic distance (2.7%).

Shedding Light on a Secretive Tertiary urodelean Relict: Hynobiid salamanders (Paradactylodon persicus s.l.) from Iran, Illuminated by Phylogeographic, Developmental and Transcriptomic Data.

The Hyrcanian Forests present a unique Tertiary relict ecosystem, covering the northern Elburz and Talysh Ranges (Iran, Azerbaijan), a poorly investigated, unique biodiversity hotspot with many cryptic species. Since the 1970s, two nominal species of Urodela, Hynobiidae, Batrachuperus (later: Paradactylodon) have been described: Paradactylodon persicus from northwestern and P. gorganensis from northeastern Iran. Although P. gorganensis has been involved in studies on phylogeny and development, there is little data on the phylogeography, systematics, and development of the genus throughout the Hyrcanian Forests; genome-wide resources have been entirely missing. Given the huge genome size of hynobiids, making whole genome sequencing hardly affordable, we aimed to publish the first transcriptomic resources for Paradactylodon from an embryo and a larva (9.17 Gb RNA sequences; assembled to 78,918 unigenes). We also listed 32 genes involved in vertebrate sexual development and sex determination. Photographic documentation of the development from egg sacs across several embryonal and larval stages until metamorphosis enabled, for the first time, comparison of the ontogeny with that of other hynobiids and new histological and transcriptomic insights into early gonads and timing of their differentiation. Transcriptomes from central Elburz, next-generation sequencing (NGS) libraries of archival DNA of topotypic P. persicus, and GenBank-sequences of eastern P. gorganensis allowed phylogenetic analysis with three mitochondrial genomes, supplemented by PCR-amplified mtDNA-fragments from 17 museum specimens, documenting <2% uncorrected intraspecific genetic distance. Our data suggest that these rare salamanders belong to a single species P. persicus s.l. Humankind has a great responsibility to protect this species and the unique biodiversity of the Hyrcanian Forest ecosystems.

The First Attempt at Studying the Species Diversity of Fish in Lake Khanka Using DNA Barcoding Techniques

Lake Khanka is the largest lake in the northeastern part of Asia, being a valuable part of the resources of freshwater fisheries. Lake Khanka has a low value of endemism owing to the recent origin and connection to large river systems of the Amur and Ussuri. The list of fish species present in the lake has been constantly updated owing to invasions that occur both unintentionally and because of the introduction of hydrobionts to maximize the potential of natural productivity. In this regard, the Lake Khanka is a suitable testing ground for the development and implementation of complex methods for monitoring species diversity. The cases of invasion have become more frequent in the lake recently. Moreover, the introduction of some fish species for stabilization of relationships among communities and exhaustive utilization of the natural productivity is now being carried out, which makes challenging the development and implementation of powerful techniques for monitoring of the local fauna species diversity. For the first time, we present a complex approach to the solution of that problem. We collected 64 fish specimens from 16 species representing four families of ray-finned fishes and analyzed their taxonomy and species diversity using both classic methods and DNA barcoding techniques. Specimens have been genotyped based on the mitochondrial Co-1 gene marker. The mean values of K2P-corrected intraspecific genetic distances were 0.15 ± 0.01%; the distances between different species within the same genera were 4.08 ± 0.01%. Phylogenetic analysis revealed monophyletic origin of all species clusters with high support (98–100% of bootstrap values). Thus, reciprocal complement of morphological and genetic techniques for the purpose of species delimitation and high effectiveness of DNA barcoding in application for documentation of species diversity in Lake Khanka has been demonstrated. New molecular genetic data generally are in agreement with taxonomic relationships previously shown for these fish taxa, but an additional investigation is required to verify the composition of genus Acheilognathus in Lake Khanka.

The molecular identification and phylogenetic reconstruction of Palaemonid and Penaeid shrimp from the southern part of Bangladesh

The study aimed to reveal the molecular identification and relationships between Palaemonid prawns and Penaeid shrimp in Bangladesh. Combined, they form an important economic part of tropical and subtropical fisheries. These species have a wide distribution range around the mangrove ecosystems, rivers, estuaries, flood plains, inundated natural depression water bodies, rice fields and other inland water bodies. Some of these species have a lack of information available about their biological and molecular characteristics. This study revealed the identification of prawn and shrimp species based on the molecular approach employed using mitochondrial COI gene markers, PCR, sequencing and matching the sequence to that stored in the NCBI database; four Palaemonid species (Macrobrachiumrosenbergii, M. lamarrei, M. kistnense,Palaemonstyliferus) and one Penaeid shrimp (Penaeus monodon) were confirmed with 99-100% confirmation of identity with multiple sequencing alignments. The inter-specific genetic divergence (K2P distance) among the family Palaemonid (Macrobrachium and Palaemon) is 0.171-0.257 %, and intraspecific genetic distance was revealed to be 0.243-0.302%. This result will be useful for obtaining the basic genetic information of these species and it will be helpful for future studies looking into the genetic biodiversity of the population structure, and finally, for the conservation and management of these resources in the southern part of Bangladesh.

DNA barcoding of cyprinids (Labeo rohita, Catla catla and Cirrhinus mrigala), mitochondrial CO1-based study

For DNA barcoding, the Cytochrome C Oxidase 1 gene (CO1) is a genetic marker used as uniform, testimonial authentication and reliable evidence for a universal species-level bio-cataloguing system compared with the morphological identification. The barcoding of Catla catla, Cirrhinus mrigala and Labeo rohita of family Cyprinidae was accomplished in the present study. Amplified CO1 gene through PCR was sequenced and figured out using bioinformatic tools. Conspecific and congeneric, K2P nucleotide deviation and nucleotide composition, the number of haplotypes and haplotype diversity were calculated. All three species of studied fish (C. catla, L. rohita and C. mrigala) were G-deficient (17.2, 17.8 and 18.5%) than C (28.3, 27.0, 27.3), A (25.2, 27.3, 26.8) and T (28.0, 28.5, 28.1), respectively. The mean intraspecific and intergeneric K2P genetic distance was .031% and 1.23%, respectively, with a high rate of transitional substitutions. Study revealed that C. catla followed the neutral evolution (R = .5), while C. mrigala (R = 2.01) and L. rohita (R = 2.84) deviated from it and offered very low genetic diversity (<.55).

First molecular identification of Euphlyctis ehrenbergii (Anura: Amphibia) inhabiting southwestern Saudi Arabia

The dicroglossid Euphlyctis ehrenbergii inhabiting southwestern Saudi Arabia has been identified morphologically and molecularly in this study. The morphometric measurements and their indices indicate that females are slightly bigger than males in some characters. Approximately 550 nucleotides of 16S rDNA gene were compared to the Yemeni haplotype available in the GenBank database. The phylogenetic inference revealed a strong sister relationship between the two haplotypes [bootstrap = 100% for maximum parsimony (MP), neighbor joining (NJ) and maximum likelihood (ML)]. The intra-specific genetic distance (D = 0.9%) was lower than the observed variation between southern and northern haplotypes of E. cyanophlyctis (D = 4.6%) currently ascribed to distinct lineages. Nonetheless, results suggest great intra-specific variation within E. ehrenbergii, deserving further investigation. The genetic distance between Yemeni and Saudi haplotypes of E. ehrenbergii (D = 0.9%) resembles intra-specific distances between haplotypes of E. cyanophlyctis ascribed to the same cryptic lineages (e.g. as observed within the lineages including haplotypes from southern India and Sri Lanka). More samples and molecular data from both haplotypes are needed to better clarify their phylogenetic relationships. This study adds a clue for forensic herpetology by highlighting the efficiency of 16S rDNA gene in animal barcoding.

The diversity of hydras (Cnidaria: Hydridae) in the Baikal region

We have studied the fauna of Hydra the waters of the Baikal region using morphological and molecular genetic methods. By the external morphological features of the structure of the polyp and the microscopic study of nematocysts, we have identified four species belonging to three different genetic groups: “oligactis group” (Hydra oligactis, H. oxycnida), “braueri group” (H. circumcincta) and “vulgaris group” (H. vulgaris). Molecular phylogenetic analysis has confirmed the species status of the hydras studied. The intraspecific genetic distances between the Baikal and European hydras are 1.5–4.2% and 0.4–2.7% of the substitutions in COI and ITS1–5.8S–ITS2 markers, respectively, while, interspecific distances for different species significantly exceed intraspecific and amount to 9.8−16.1% and 6.4−32.1% of substitutions for the markers COI and ITS1–5.8S–ITS2, respectively. The temperature regime of the water and the availability of food resources, which play a key role in the reproduction of hydras, determine the habitats of the identified species. The research performed has replenished the regional fauna with species whose findings were previously considered presumptive or doubtful. The first record of H. vulgaris in the artificial reservoir near the Angara River (Lake Kuzmikhinskoye) will help to clarify the distribution of this species.