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This work used small (12S rRNA) mitochondrial rRNA gene to assess the evolutionary connections of some species of the cardinal fishes belonging to order Kurtiformes. Small mitochondrial rRNA sequences varied in length from 920 to 944 base pairs. Of the 947 characters in the aligned 12S rRNA data set, 834 were consistent sites, 19 were parsimony informative, and 111 were variable. Small mitochondrial rRNA sequences' nucleotides were uploaded to GenBank with accession numbers (PP830791- PP830794). The P-distances among the understudied cardinalfish species, expanded from 0.0060 to 0.0130. The highest value (0.0130) was found between Cheilodipterus novemstriatus and Ostorhinchus fasciatus. While, the lowest P-distance (0.0060) was found between Cheilodipterus quinquelineatus and Cheilodipterus macrodon. Our results demonstrated the utility of 12S rRNA in the phylogenetic analysis of the cardinalfish species. [ABSTRACT FROM AUTHOR]

In recent decades, medicinal leeches of the genus Hirudo have experienced a sharp decline throughout their distribution range due to their overexploitation for medical use and habitat alteration. Such a phenomenon is not fully understood because of the complex taxonomy of the genus, which remained unsettled until the beginning of the XXI century, when the implementation of DNA taxonomy allowed for a better understanding of the diversity of the genus and the distribution ranges of Hirudo species. Only the Mediterranean medicinal leech Hirudo verbana Carena, 1820 is currently reported to occur in Italy and its major islands, although records of other Hirudo species have been reported in the literature. In this study, we report the occurrence of the Dragon leech Hirudo troctina Johnson, 1816 in Sardinia and provide its molecular characterization. In addition, based on a review of the existing literature and museum collections, we provide an ample synopsis of the available evidence regarding the occurrence of the species on the island. Hirudo troctina proved to be widespread and abundant in Sardinia during the XIX century but became rarer and rarer in the XX and XXI centuries possibly due to its overexploitation for medicinal use and export in mainland Europe, the abandonment of traditional grazing practices, and the decline of wetland habitats and amphibian populations. Currently, the only recently validated occurrence sites of the species are the water bodies of the “Giara di Gesturi”, a basaltic plateau in central-southern Sardinia. No corroborated evidence of the possible occurrence of other Hirudo species on the island was found. A better understanding of Hirudo troctina distribution in Sardinia will allow the amelioration of its protection and management practices, also under the provisions of the European “Habitats Directive”, where the H. medicinalis species complex is listed in Annex V.

The escalating environmental changes are imposing multiple pressures on biodiversity conservation, thereby, more attention has been paid to fish diversity and community assembly patterns. Fish diversity in Liaohe River has deteriorated significantly due to overfishing, environmental pollution and other reasons, in addition, no fish survey related research has been published in Liaohe National Park in recent years. In order to study the diversity of fish species in the area and to investigate the presence of any invasive species. This study used eDNA metabarcoding to explore the distribution and diversity of fish communities in the Liaohe National Park. The study showed that a total of 54 species of fish from six orders, 14 families, and 39 genera were detected, meantime, the invasive species Lepomis cyanellus was found at two sampling sites. The α-diversity results showed that estuarine sites had the highest species diversity. The spatial structure of fish communities in the upstream and downstream reaches were similar as indicated by non-metric multidimensional scaling. In our study, the fish community is influenced by both deterministic and stochastic processes, with the stochastic processes being the main factor. The potential existence of L. cyanellus in Liaohe National Park might be resulted from aquaculture, ornamental pet trade, other economic fish farming, artificial release. The relatively low number of fish species in this study may be attributed to the limited number of sampling events and sampling sites. In addition to these factors, over exploitation of resources, water pollution, and the invasion of alien species have directly impacted the biodiversity of aquatic life. The results of this study lay a foundation for the future biodiversity investigation in National Parks of China, meantime, the use of eDNA monitoring is crucial to prevent, to control and limit the spreading of invasive species in advance. [ABSTRACT FROM AUTHOR]

Abstract Background Rhipicephalus (Boophilus) microplus (Canestrini, 1888), the Asian blue tick, is a highly invasive and adaptable ectoparasite. This tick species has successfully established itself in most regions of the world, with movement of cattle being a major driver for its spread. In the recent past, R. microplus ticks have been reported in three districts of Uganda. Information on its spread and distribution are vital in deepening our understanding of the ecological scenarios that lead to tick persistence and in the formulation of control strategies. This is especially important in the cattle-dense districts. Methods We randomly collected tick specimens from 1,461cattle spread across seven cattle dense districts located in the Central, Karamoja and West Nile regions of Uganda from January to September 2020. The ticks were identified using standard morpho-taxonomic keys and the R. microplus tick species identities were confirmed by sequencing of the ITS2 region, 12S rRNA and 16S rRNA genes and phylogenetic analyses. Results Adult ticks (n = 13,019) were collected from 1,461 cattle. Seventeen tick species were identified based on morpho-taxonomic keys and the majority (47.4%; n=6184) of these were R. appendiculatus. In total, 257 R. microplus ticks were found infesting cattle in 18 study sites in the districts of Amudat, Kaabong, Napak (Karamoja region) and Arua (West Nile region). The identity of R. microplus was confirmed using molecular technics. No R. microplus tick was recorded in the districts of Lyantonde and Nakaseke (Central region). Arua district accounted for 82.1% (n=211) of the R. microplus ticks recorded followed by Napak district at 16.3% (n=42), while Amudat and Kaabong districts accounted for 1.5% (n=4). Rhipicephalus microplus and R. decoloratus co-existed in 6 of the 13 study sites in Arua district, while in another 6 study sites, no R. decoloratus was recorded. In the Karamoja region districts R. decoloratus co-existed with R.microplus. Of the total 618 ticks belonging to four species of the subgenus Boophilus recorded in this study, R. decoloratus accounted for 50.04% (n=334), followed by R. microplus at 41.58% (n=257), R. geigyi at 2.75% (n=17) and R. annulatus at 1.61% (n=10). In the districts of Amudat, Kaabong and Napak, R. decoloratus was more dominant (76.1%; n=179) of the three Rhipicephalus (Boophilus) tick species recorded, followed by R. microplus (19.5%; n=46) and R. geigyi (4.2%; n=10). Contrariwise, R. microplus was more dominant (84%; n=211) in Arua district followed by R. decoloratus (10.7%; n=27), R. annulatus (3.9%; n=10) and R. geigyi (1.1%; n=3). Phylogenetic analyses of the ITS2 region, 12S rRNA and 16S rRNA genes revealed subgrouping of the obtained sequences with the previously published R. microplus sequences from other parts of the world. Conclusion Rhipicephalus microplus ticks were found infesting cattle in four districts of Uganda. The inability to find R. decoloratus, an indigenous tick, from six sites in the district of Arua is suggestive of its replacement by R. microplus. Rhipicephalus microplus negatively affects livestock production, and therefore, there is a need to determine its distribution and to deepen the understanding of the ecological factors that lead to its spread and persistence in an area.

Background: Rhipicephalus (Boophilus) microplus (Canestrini, 1888), the Asian blue tick, is a highly invasive and adaptable ectoparasite. This tick species has successfully established itself in most regions of the world, with movement of cattle being a major driver for its spread. In the recent past, R. microplus ticks have been reported in three districts of Uganda. Information on its spread and distribution are vital in deepening our understanding of the ecological scenarios that lead to tick persistence and in the formulation of control strategies. This is especially important in the cattle-dense districts. Methods: We randomly collected tick specimens from 1,461cattle spread across seven cattle dense districts located in the Central, Karamoja and West Nile regions of Uganda from January to September 2020. The ticks were identified using standard morpho-taxonomic keys and the R. microplus tick species identities were confirmed by sequencing of the ITS2 region, 12S rRNA and 16S rRNA genes and phylogenetic analyses. Results: Adult ticks (n = 13,019) were collected from 1,461 cattle. Seventeen tick species were identified based on morpho-taxonomic keys and the majority (47.4%; n=6184) of these were R. appendiculatus. In total, 257 R. microplus ticks were found infesting cattle in 18 study sites in the districts of Amudat, Kaabong, Napak (Karamoja region) and Arua (West Nile region). The identity of R. microplus was confirmed using molecular technics. No R. microplus tick was recorded in the districts of Lyantonde and Nakaseke (Central region). Arua district accounted for 82.1% (n=211) of the R. microplus ticks recorded followed by Napak district at 16.3% (n=42), while Amudat and Kaabong districts accounted for 1.5% (n=4). Rhipicephalus microplus and R. decoloratus co-existed in 6 of the 13 study sites in Arua district, while in another 6 study sites, no R. decoloratus was recorded. In the Karamoja region districts R. decoloratus co-existed with R.microplus. Of the total 618 ticks belonging to four species of the subgenus Boophilus recorded in this study, R. decoloratus accounted for 50.04% (n=334), followed by R. microplus at 41.58% (n=257), R. geigyi at 2.75% (n=17) and R. annulatus at 1.61% (n=10). In the districts of Amudat, Kaabong and Napak, R. decoloratus was more dominant (76.1%; n=179) of the three Rhipicephalus (Boophilus) tick species recorded, followed by R. microplus (19.5%; n=46) and R. geigyi (4.2%; n=10). Contrariwise, R. microplus was more dominant (84%; n=211) in Arua district followed by R. decoloratus (10.7%; n=27), R. annulatus (3.9%; n=10) and R. geigyi (1.1%; n=3). Phylogenetic analyses of the ITS2 region, 12S rRNA and 16S rRNA genes revealed subgrouping of the obtained sequences with the previously published R. microplus sequences from other parts of the world. Conclusion: Rhipicephalus microplus ticks were found infesting cattle in four districts of Uganda. The inability to find R. decoloratus, an indigenous tick, from six sites in the district of Arua is suggestive of its replacement by R. microplus. Rhipicephalus microplus negatively affects livestock production, and therefore, there is a need to determine its distribution and to deepen the understanding of the ecological factors that lead to its spread and persistence in an area. [ABSTRACT FROM AUTHOR]

As the largest lake in South China, Wanlv Lake is also an important drinking water source for Guangdong and Hong Kong and is responsible for water supplying of more than 40 million people in the Guangdong-Hong Kong-Macao Greater Bay Area. The study of fish diversity in Wanlv Lake can help us to effectively understand the changes and states of its aquatic ecosystem and provide scientific basis for natural ecological protection, biodiversity, sustainable use and scientific management of fishery resources. However, research on the lake's fish is scarce. The high throughput environmental DNA metabarcoding technology (eDNA technology) of the mitochondrial 12S rRNA gene sequence was herein used to conduct a preliminary exploration of Wanlv Lake fish diversity. A total of 10 sampling sites were set up to monitor fish diversity and analyze the composition and richness of the species. The results revealed a total of 83 genera in 42 families and 17 orders of fish in the lake, with the highest proportion found in Cypriniformes (38 species), accounting for 45.24% of the total. Five alien species and eight rare and endangered species were also detected. The proportion of invasive species was 5.95%, and the combined proportion of rare and endangered species was 9.52%. The results indicated that Wanlv Lake serves as an important drinking water source, and overall, the condition of fish is satisfactory. However, proactive measures should be implemented to control the overpopulation of invasive species, as their unchecked proliferation may lead to a decline in species abundance, particularly among endangered species. This was the first overall assessment report on fish of the Wanlv Lake via 12S rRNA; the results herein lay a foundation for water quality assessment of Wanlv Lake as a drinking water source. [ABSTRACT FROM AUTHOR]

Turkey is among the countries where sand flies (Diptera: Psychodidae) are responsible for the endemic presence of leishmaniasis. Understanding the feeding behaviour of haematophagous arthropods is crucial for revealing the transmission cycles of vector-borne diseases. However, research on the host preferences of sand fly species in Turkey has been limited. This study aims to determine the blood meal sources of sand fly species collected from a wide range of geographical areas and to identify their feeding patterns. For the blood meal analysis, Sanger sequencing targeting the vertebrate mitochondrial cytochrome b or mammalian 12 S rRNA genes was performed on 265 female specimens collected between 2006 and 2019. The feeding patterns of sand flies were determined using Null model analysis. Host assignments of 218 individuals revealed nine different vertebrate hosts for 14 sand fly taxa. Bos taurus was the predominant host (81.19%), followed by Gallus gallus (8.88%), Capra hircus (5.14%), Mus musculus (1.81%), Ovis aries (0.93%), Homo sapiens (0.47%), Canis lupus familiaris (0.47%), Felis catus (0.47%) and Sitta spp. (0.47%). We discovered an aggregated feeding pattern among sand fly taxa instead of random or segregated feeding. Most sand fly taxa predominantly fed on bovines as their hosts, highlighting the need for further research on the role of domestic animals in the transmission of sand fly-borne diseases. Paraphlebotomus specimens displayed a relatively broad host range, particularly favouring Avian hosts, which requires further investigation into their life history traits. [ABSTRACT FROM AUTHOR]

Ticks are vectors for transmitting tick-borne pathogens (TBPs) in animals and humans. Therefore, tick identification is necessary to understand the distribution of tick species and the pathogens they carry. Unfortunately, data on dog ticks and the TBPs they harbor in Malawi are incomplete. This study aimed to identify dog ticks and the TBPs they transmit in Malawi. One hundred thirty-two ticks were collected from 87 apparently healthy but infested domestic dogs in four districts of Malawi, which were pooled into 128 tick samples. The ticks were morphologically identified under a stereomicroscope using identification keys, and species identification was authenticated by polymerase chain reaction (PCR) through the amplification and sequencing of 12S rRNA and cytochrome c oxidase subunit I (CO1) genes. The tick species identified were Rhipicephalus sanguineus sensu lato (58.3%), Haemaphysalis elliptica (32.6%), and Hyalomma truncatum (9.1%). Screening for TBPs using species-specific PCR assays revealed that 48.4% of the ticks were infected with at least one TBP. The TBP detection rates were 13.3% for Anaplasma platys, 10.2% for Babesia rossi, 8.6% for B. vogeli, 6.3% for Ehrlichia canis, 3.9% for A. phagocytophilum, 3.1% for B. gibsoni, 2.3% for B. canis and 0.8% for Hepatozoon canis. Co-infections of up to three pathogens were observed in 48.4% of the positive samples. This is the first study to identify dog ticks and the TBPs they harbor in Malawi. These findings provide the basis for understanding dog tick distribution and pathogens they carry in Malawi. This study necessitates the examination of ticks from more study locations to have a better picture of tick challenge, and the development of ticks and tick-borne disease control methods in Malawi. [ABSTRACT FROM AUTHOR]

The study of ichthyoplankton is paramount to understanding fish assemblages' reproductive dynamics. DNA metabarcoding has been applied as a rapid, cost‐effective, and accurate taxonomy tool, allowing the identification of multiple individuals simultaneously. However, there remain significant challenges when using DNA metabarcoding, such as molecular marker choice according to the taxonomic resolution and length of the fragment to be sequenced, primer bias, incomplete reference databases, and qualitative inference incongruences. Here, 30 ichthyoplankton pools collected from a Neotropical river were identified at a molecular level using DNA metabarcoding to compare the resolution, sensibility, specificity, and relative read abundance (RRA) recovery of three molecular markers: the standard COI fragment (650 pb, with each end analyzed individually) and two short 12S rRNA genes markers (≅200 bp – NeoFish and MiFish markers). The combined use of the three markers increased the genera detection rates by 25%–87.5%, allowing an increased taxonomic coverage and robust taxonomic identification of complex Neotropical ichthyoplankton communities. RRA is marker‐dependent, indicating caution is still needed while inferring species abundance based on DNA metabarcoding data when using PCR‐dependent protocols. [ABSTRACT FROM AUTHOR]

AbstractIn Lar National Park (Caspian Sea basin, Iran), the Caspian trout (Salmo caspius) population faces different threats, including introduced fish species. Due to the harsh environmental conditions and limited accessibility, monitoring of fish species via conventional approaches proves difficult. Hence, environmental DNA metabarcoding may prove an appropriate tool for monitoring fishes within the park. Environmental DNA samples from eight stream sites in the National Park were sequenced via metabarcoding of the 12S rRNA gene, and the species identified via eDNA metabarcoding were compared to the results of electrofishing performed at the same localities on the same day. No significant difference in the number of Caspian Sea trout DNA sequence reads was detected among the collection sites (p > 0.05). The highest number of reads was detected in Dalichay Stream, but the highest population density determined via electrofishing was in Siahpalas Stream. The discrepancy between the eDNA read count and trout population density, as well as the limited sampling scheme within this study, limit our ability to provide a robust conclusion about the application of environmental DNA metabarcoding for assessment of fish density in Lar National Park. Environmental DNA metabarcoding detected more species than electrofishing, but no significant differences in the composition of the local fish community were observed. Introduced fish species were all observed or detected in Siahpalas Stream, which is characterized by high water temperature, muddy substrate, and lower flow rate. A significant effect of flow rate and total dissolved solids on the presence of introduced fish species (p = 0.02) and of flow rate alone on relative abundance of introduced fish species (p = 0.03) was detected. To standardize the application of eDNA metabarcoding as a biodiversity assessment tool in Lar National Park, future studies should characterize the parameters that affect eDNA persistence and detectability in the system.

Abstract To successfully implement environmental DNA‐based (eDNA) diversity monitoring, the completeness and accuracy of reference databases used for taxonomic assignment of eDNA sequences are among the challenges to be tackled. Here, we have developed a workflow that evaluates the current status of GenBank for marine fishes. For a given combination of species and barcodes, a gap analysis is performed and potentially erroneous sequences are identified. Our gap analysis based on the four most used genes (cytochrome c oxidase subunit 1, 12S rRNA, 16S rRNA, and cytochrome b) for fish eDNA metabarcoding found that COI, the universal choice for metazoans, is the gene covering the highest number of Northeast Atlantic marine fishes (70%), while 12S rRNA, the preferred region for fish‐targeting studies, only covers about 50% of the species. The presence of too close and too distant barcode sequences as expected by their taxonomic classification confirms the existence of erroneous sequences in GenBank that our workflow can detect and eliminate. Comparing taxonomic assignments of real marine eDNA samples with raw and clean reference databases for the most used 12S rRNA barcodes (teleo and MiFish), we confirmed that both barcodes perform differently and demonstrated that the application of the database cleaning workflow can result in drastic changes in community composition. Besides providing a tool for reference database curation, this study confirms the need to increase 12S rRNA reference sequences for European marine fishes and evidences the dangers of taxonomic assignments by directly querying GenBank. We have developed a workflow that evaluates the current status of GenBank for marine fishes. For a given combination of species and barcodes, a gap analysis is performed and potentially erroneous sequences are identified.

Accurate species identification is a key component of biodiversity research. DNA barcoding is an effective molecular method used for fish species identification. We aimed to study the DNA barcoding of fish in Zhoushan coastal waters, explore the differences and applicability of two gene fragments (12S rRNA and COI) of DNA barcoding in fish species identification, and established a comprehensive fish barcoding reference database. Two hundred and eighty-seven captured fish samples from Zhoushan coastal waters were identified using morphological characteristics and DNA barcoding. A total of 264 12S rRNA sequences (belonging to eight orders, 31 families, 55 genera, and 66 species) and 188 COI sequences (belonging to seven orders, 30 families, 48 genera, and 58 species) were obtained. The lengths of the 12S rRNA sequences ranged from 165 to 178 bp, and the guanine-cytosine (GC) content was 45.37%. The average 12S rRNA interspecific and intraspecific genetic distances (K2P) were 0.10% and 26.66%, respectively. The length of the COI sequence ranged 574–655 bp, and the content of GC was 45.97%. The average 12S rRNA interspecific and intraspecific genetic distances (K2P) were 0.16% and 27.45%, respectively. The minimum interspecific genetic distances of 12S rRNA and COI (1.23% and 1.86%) were both greater than their maximum intraspecific genetic distances (2.42% and 8.66%). Three molecular analyses (NJ tree, ABGD, and GMYC) were performed to accurately identify and delineate species. Clustering errors occurred when the 12S rRNA sequences were delimited using the NJ tree method, and the delimitation results of ABGD and GMYC are consistent with the final species identification results. Our results demonstrate that DNA barcoding based on 12S rRNA and COI can be used as an effective tool for fish species identification, and 12S rRNA has good application prospects in the environmental DNA (eDNA) metabarcoding of marine fish. [ABSTRACT FROM AUTHOR]

Abstract The study of ichthyoplankton is paramount to understanding fish assemblages' reproductive dynamics. DNA metabarcoding has been applied as a rapid, cost‐effective, and accurate taxonomy tool, allowing the identification of multiple individuals simultaneously. However, there remain significant challenges when using DNA metabarcoding, such as molecular marker choice according to the taxonomic resolution and length of the fragment to be sequenced, primer bias, incomplete reference databases, and qualitative inference incongruences. Here, 30 ichthyoplankton pools collected from a Neotropical river were identified at a molecular level using DNA metabarcoding to compare the resolution, sensibility, specificity, and relative read abundance (RRA) recovery of three molecular markers: the standard COI fragment (650 pb, with each end analyzed individually) and two short 12S rRNA genes markers (≅200 bp – NeoFish and MiFish markers). The combined use of the three markers increased the genera detection rates by 25%–87.5%, allowing an increased taxonomic coverage and robust taxonomic identification of complex Neotropical ichthyoplankton communities. RRA is marker‐dependent, indicating caution is still needed while inferring species abundance based on DNA metabarcoding data when using PCR‐dependent protocols.

As the largest lake in South China, Wanlv Lake is also an important drinking water source for Guangdong and Hong Kong and is responsible for water supplying of more than 40 million people in the Guangdong-Hong Kong-Macao Greater Bay Area. The study of fish diversity in Wanlv Lake can help us to effectively understand the changes and states of its aquatic ecosystem and provide scientific basis for natural ecological protection, biodiversity, sustainable use and scientific management of fishery resources. However, research on the lake’s fish is scarce. The high throughput environmental DNA metabarcoding technology (eDNA technology) of the mitochondrial 12S rRNA gene sequence was herein used to conduct a preliminary exploration of Wanlv Lake fish diversity. A total of 10 sampling sites were set up to monitor fish diversity and analyze the composition and richness of the species. The results revealed a total of 83 genera in 42 families and 17 orders of fish in the lake, with the highest proportion found in Cypriniformes (38 species), accounting for 45.24% of the total. Five alien species and eight rare and endangered species were also detected. The proportion of invasive species was 5.95%, and the combined proportion of rare and endangered species was 9.52%. The results indicated that Wanlv Lake serves as an important drinking water source, and overall, the condition of fish is satisfactory. However, proactive measures should be implemented to control the overpopulation of invasive species, as their unchecked proliferation may lead to a decline in species abundance, particularly among endangered species. This was the first overall assessment report on fish of the Wanlv Lake via 12S rRNA; the results herein lay a foundation for water quality assessment of Wanlv Lake as a drinking water source.

DNA barcoding is a powerful tool that provides rapid, accurate, and automatable species identification using standardized genetic region(s), such as for revealing the existence of cryptic species and/or rare species in biodiversity monitoring. DNA barcoding techniques require the development of sets of universal PCR primers for DNA barcoding. We tried to develop universal primer sets, and succeeded in designing not only universal primer sets for DNA barcoding regions of almost all insects, which were designed to include a hypervariable site between highly conserved sites, but also primer sets for longer fragment sequences for registration in a database. We confirmed successful amplification for 14 orders, 43 families, and 68 species with DNA barcoding in the mtDNA 16S rRNA region, and for 13 orders, 42 families, and 66 species with DNA barcoding in the mtDNA 12S rRNA region, including Apterygota and Pterygota (Paleoptera, Polyneoptera, Paraneoptera, and Oligoneoptera). A key feature is that the DNA fragments of the DNA barcoding regions amplified by these primer sets are both short at about 200-bp, and longer fragment sequences will increase the level of data registration in the DNA database. In addition, we evaluated the sensitivity of these newly developed primers using Epeorus aesculus (Heptageniidae), which inhabits a relatively wide range of river systems. The results of this study revealed the existence of a cryptic species or an undescribed species. Such resulting database enhancements will provide opportunities for increasingly accurate assessment of biodiversity and genetic diversity. [ABSTRACT FROM AUTHOR]

An adult Indian buffalo (Bubalus bubalis) presented with corneal opacity, irritation, and excessive lacrimation from the left eye in the Referral Veterinary Polyclinic-Teaching Veterinary Clinical Complex (RVC-TVCC), Indian Veterinary Research Institute, Izatnagar. Clinical examination revealed a whitish thread-like worm in the left eye's anterior chamber. The worm was surgically removed from the eye with supportive nerve blocks. Light microscopy was used for parasite morphological identification, which provided insight into the worm as female Setaria sp. Genomic DNA was isolated, and polymerase chain reaction amplification of 12S rRNA was conducted for molecular confirmation of the parasite. The amplicon was sequenced and analysed by bioinformatics software. Sequence data showed an amplicon size of 243 bp. Phylogenetic analysis with reference data from the NCBI Genbank database revealed the worm was S. digitata , with a similarity of 99.17%. The common predilection site of S. digitata is in the peritoneal cavity of natural hosts like cattle and buffalo and is mostly non-pathogenic. The aberrant migration of the parasite larva to the brain and eye commonly occurs in goats, sheep, and horses, causing clinical conditions like cerebrospinal nematodiasis (lumbar paralysis) and ocular setariasis, respectively. Nevertheless, until now, there have been no reports of ocular setariasis in buffalo. This report is the first unusual occurrence of ocular setariasis in buffalo and its molecular confirmation and phylogenetic analysis using 12S rRNA. [ABSTRACT FROM AUTHOR]

Argas persicus (the fowl tick) is a species of soft tick commonly associated with poultry farms. It has a wide geographic distribution and colonizes different climate regions. Morphological identification of A. persicus has been reported worldwide, but genetic data regarding its molecular characterization is limited. The present study provides data for morphological identification and genetic characterization of A. persicus collected from domestic birds in traditional farms from east Algeria (Setif region). Additionally, A. persicus samples originating from Gansu province in China were included for comparative molecular study. In total, 1518 ticks collected from 30 infested farms were examined and morphologically identified as A. persicus. Furthermore, the 14 tick samples obtained from China were morphologically identified as A. persicus. Molecular analysis of 30 ticks from Algeria (one tick from each infested farm) and the 14 Chinese samples based on PCR, sequencing, and phylogenetic analysis of three mitochondrial genetic markers (16S rRNA, 12S rRNA, and cox1) confirmed morphological results where all samples belonged to the A. persicus group. However, phylogenetic analysis showed that all Algerian samples and two Chinese samples belong to A. persicus sensu stricto (s.s.), while the remaining Chinese samples represented A. persicus sensu lato (s.l.) (divergent lineage). The present study confirms the occurrence of A. persicus s.s. both in Algeria and China, as well as provides novel molecular data for a distinct Chinese lineage of A. persicus. [ABSTRACT FROM AUTHOR]

Environmental DNA (eDNA) metabarcoding emerges as a powerful method, allowing a more exhaustive investigation of fish fauna than any other methods. Yet, the general use of eDNA as a replacement of traditional methods such as visual surveys or physical sampling remains debatable. Therefore, a prior understanding of eDNA’s spatial resolution is necessary. This study aimed to compare the assessments of fish diversity at three spatial scales by eDNA, underwater visual census (UVC), and diver-operated video (DOV) surveys across 21 reef sites in northern Taiwan. The specific objectives were to explore the regional species pool (γ-diversity), reveal spatial patterns of fish assemblages (β-diversity), and disentangle the relationships between fish assemblages and benthic composition (α-diversity). At the γ-diversity level, a total of 438 marine fish species were detected across methods. eDNA exhibits an extraordinary power to explore the regional species pool given sufficient replication, a power which is unachievable by DOV and UVC. At the β-diversity level, all the methods successfully revealed the same spatial patterns of beta diversity and the distance decay of similarity in fish assemblages. At the α-diversity level, none of the methods is capable of investigating the entire resident fish fauna, but visual surveys are more suitable for scrutinizing interactions between fish and benthos. Instead of undiscriminatingly recommending a combination of eDNA with traditional survey methods, we suggest implementing specific surveys in accordance with the ecological questions of interest.

This study was conducted to assess the role of the mitochondrial 12S rRNA variations in the phylogenetic discrimination between two Iraqi breeds of sheep that differ in geographical distribution. A total of 122 animals (68 Awassi and 54 Hamdani) were included in the study. Direct sequencing of amplicons followed by the construction of a median-joining network and several trees were performed to identify the possible phylogenetic differences between both involved breeds. Genetic diversity, relative frequencies, and analysis of molecular variance (AMOVA) were performed to assess the genetic correlation between both populations. The median-joining network and minimized tree values showed that all observed haplotypes were separated into two groups according to their breed. Comprehensive phylogenetic data revealed only one Asian ancestor for all observed haplotypes. As indicated by AMOVA, the observed diversity was mostly due to between-population variation (1.24836%), while within-population variation (0.91221%) accounted for much less. The currently investigated rRNA amplicons exhibited different mitochondrial manifestations between Awassi and Hamdani breeds. Due to the ability of these 12S rRNA amplicons to mimic the geographical diversity for the currently investigated breeds, it is highly recommended to be used as potent mitochondrial genetic markers among broader ovine sequences. [ABSTRACT FROM AUTHOR]

The availability of a reliable molecular assay in species recognition in forensic cases is of paramount importance when visual inspection or morphological methods are not exhaustive, especially from challenging samples. Here, two different caseworks involving bone samples founded during medico-legal outdoor investigations are presented. In order to exclude the human nature of the specimens and to determine the exact species they belong to, we proceeded with the molecular approach trying to generate sequences from the classical mtDNA markers cyt b and COI. However, they both gave critical results. For this reason, a short amplicon of ~ 150 bp of the 12S rRNA gene was used as an alternative.This short fragment was sufficient to identify the biological origin of the bone specimens with a high degree of certainty leading to the exclusion of their human nature. This work highlights the utility of the 12S rRNA and underlines the importance of deepen the choice of alternative shorter markers with respect to the classical ones, in order to achieve species identification even from challenging and degraded material in forensic criminal and wildlife caseworks. [ABSTRACT FROM AUTHOR]

Limpets of the genera Erginus Jeffreys, 1877 (previously in Lottiidae) and Rhodopetala Dall, 1921 (previously in Acmaeidae), unlike other patellogastropods, brood larvae and juveniles in the modified pallial cavity. A phylogenetic analysis using three mitochondrial gene markers (cytochrome c oxidase subunit I, 16S rRNA, and 12S rRNA) has shown that Rhodopetala rosea (Dall 1872) and four Erginus species form a highly supported clade that cannot be assigned to Acmaeidae or Lottiidae. The transition from external fertilization and pelagic development to internal fertilization with brooded larvae and juveniles in the special pallial cavity occurred in the evolution of patellogastropods only once. The genera Erginus and Rhodopetala belong to Rhodopetalidae Lindberg, 1981 stat. nov. (= Erginidae Chernyshev 2018 syn. nov.) elevated from subfamily Rhodopetalinae to family status. Erginus puniceus Lindberg, 1988a is restored from synonym of Erginus sybariticus (Dall 1871) for both typical purple forms and the ' sybariticus ' forms with small shell. [ABSTRACT FROM AUTHOR]

A single specimen of Spotted frillgoby, Bathygobius cyclopterus (Valenciennes, 1837), was collected from a tidal pool on Jeju Island, Korea, in October 2017. The species is characterized by five upper free rays of the pectoral fin, a lower jaw with a mental frenum with a slightly curved posterior margin, and a small flap on the anterior nostril. Mitochondrial DNA 12S rRNA revealed that it corresponds to B. cyclopterus than other congeneric species by 100% identity. This species is the third Bathygobius species of the fish fauna of Korea, and the new Korean name proposed for the species is “Eol-lug-mu-nui-mang-dug”.

Abstract Large carnivores play a pivotal regulating role in maintaining healthy and balanced ecosystems; however, most of them are rare and elusive, and knowledge about their resource consumption is scarce. Traditional methods based on morphological identification of undigested remains are labor intensive and often not sufficiently accurate, leading to errors and biased ecological inferences. Here, we developed a multi‐marker DNA metabarcoding approach to analyze the dietary diversity of giant otters (Pteronura brasiliensis) from fecal DNA while controlling predator species identity. We combined two mitochondrial markers, 12S rRNA and cytochrome c oxidase 1 (COI) gene, that target the full range of potential vertebrate and invertebrate prey. We compiled a local reference database of DNA barcodes for most potentially ingested fish, which were used to evaluate the specificity of the metabarcoding primers in silico. Most prey are identified at the species level (>90%) and the dietary profiles provided independently by the two markers are highly similar, whether in terms of list of prey or frequency of occurrences, hence validating the approach. We detected a higher number of rare fish prey with the 12S primers that amplified solely Teleost species while the degenerate COI primers revealed non‐fish prey (e.g., amphibians, snakes, birds, and earthworms) and confirmed predator species identity. This study demonstrated that scat DNA metabarcoding is particularly useful to provide in‐depth information on elusive carnivorous dietary profile. Our methodology opens up new opportunities to understand how top carnivores diet cope with the effects of anthropogenic alteration of ecosystems and identify conflicts with humans and livestock.

The taxonomy of 185 species of Afrotropical freshwater crabs is revised to conform to the updated phylogenetic relationships within this large assemblage, based on the largest taxonomic sampling to date (that includes almost all genera and 57% of the species). Four DNA loci were sequenced including three mitochondrial loci (COI , 12S rRNA, 16S rRNA) and one nuclear locus (histone 3), and evolutionary relationships were estimated using maximum likelihood and Bayesian approaches. The divergence times of the major clades were estimated with a Yule tree prior and an uncorrelated lognormal relaxed molecular clock. The taxonomic acts proposed here include the recognition of two Afrotropical families of freshwater crabs (Potamonautidae and Deckeniidae) each with two subfamilies. In the revised Potamonautidae, three new tribes and three new genera (Maritimonautes gen. nov. Nesonautes gen. nov. Occidensonautes gen. nov.), are established, three previously published genera are revised and four subgenera are raised to genus (Arcopotamonautes , Lirrangopotamonautes , Longipotamonautes , Rotundopotamonautes). An updated species list and identification keys for the new taxa, revised distribution maps, digital images and illustrations of diagnostic characters are provided. [ABSTRACT FROM AUTHOR]

Gobioidei is one of the largest vertebrate taxa with over 2000 species observed around the globe. The largest group in Gobioidei is gobies that had been classified as one family, Gobiidae, based on morphological features. Molecular phylogenetic studies revealed that gobies consisted of two monophyletic families, Gobiidae and Oxudercidae, in which 19 lineages have been proposed, despite some claims arisen about the relationship among these lineages or species. We analyzed 58 Gobioidei species, including 45 East Asian oxudercids, based on 12S rRNA sequences to reconstruct the spatiotemporal diversification history of gobies. Our analysis yielded the results compatible with the previous reports in a large framework. The common ancestor of Gobiidae and Oxudercidae were estimated to appear at 38.66 Mya. Genus-level splits occurred in Gobiidae and Oxudercidae predominantly at Miocene and late Miocene to early Pleistocene, respectively. Gobies have likely originated in many parts of the northern and western Pacific Ocean, of which a large proportion of Oxudercidae have adapted to various environments in the North Pacific.

Species detection and identification is a crucial steps in biodiversity assessment. Traditional methods are often invasive and resource intensive. The number of studies demonstrating successful of eDNA metabarcoding approach in species identification has increased rapidly in recent years. Some of large terrestrial mammals have reportedly utilize natural salt licks as a source of minerals in the diet and its genetic material left in the environment can be used to identify species from this site. An eDNA metabarcoding protocol had been carried out to identify Sulawesi mammals from Adudu natural salt-licks, Nantu Wildlife Reserve, Gorontalo. Environmental DNA were extracted from water samples, Amplicon libraries were prepared by PCR amplification and Illumina MiSeq high throughput sequencing. Reads processing and taxonomic assignment carried out in two bioinformatics packages, PipeCraft-1.0 and OBITools-2.11. Two endangered Sulawesi mammals species had been identified, i.e. lowland anoa (Bubalus depressicornis) and babirusa (Babyrousa babyrussa). The accuracy of mammal species identification using eDNA metabarcoding is affected by rigorous experimental procedures, DNA marker reliability, and availability of reference sequence database. [ABSTRACT FROM AUTHOR]

In India and some of the African countries, one of the unconventional meats receiving the latest attention in meat adulteration is camel meat. So, the objective of this study was to develop a species-specific PCR based on mitochondrial cytochrome b (CYTB) gene and a PCR-RFLP assay of mitochondrial 12S rRNA to identify camel meat in suspected samples. Known sample of camel meat, samples suspected to be from illegally slaughtered camel and known samples of cattle, buffalo, sheep, goat, pork and chicken were used in the study. DNA were extracted from all samples following spin column method and PCR amplification were carried out using both CYTB and 12S rRNA gene primers. The CYTB gene amplification produced amplicon with a size of 435 bp without any non-specific spurious amplification towards other species studied. Further, the 12S rRNA PCR products were analysed both by sequencing and by RFLP using enzyme AluI. On BLAST analysis the 448 bp sequence obtained from suspected samples showed > 99% sequence homology to previously reported Camelus dromedaries (accession no: AM 9369251.1). On AluI digestion of the 448 bp product from both known and suspected camel samples, a specific RFLP pattern with three distinct products of 90, 148 and 210 bp size were evident, which were significantly different from the pattern of cattle, sheep, goat, chicken and buffalo. Further, after in-house validation, this cost effective and rapid method of camel meat identification is placed into practice for regular screening of vetero-legal samples in the lab. [ABSTRACT FROM AUTHOR]

Abstract Background The family Labridae made up of 519 species in the world. The functional evolution of the feeding-related jaws leaded to differentiation of species, and the pharyngeal jaw apparatus evolved independently, but evolutionary mechanism still remain unaddressed in wrasses. Mitogenomes data can be used to infer genetic diversification and investigate evolutionary history of wrasses, whereas only eight complete mitogenomes in this family have been sequenced to date. Here, we sequenced the complete mitogenomes of Iniistius trivittatus to investigate genetic differentiation among wrasse species. Results We sequenced the complete mitogenomes of I. trivittatus using a novel PCR strategy. The I. trivittatus mitogenomes is 16,820 bp in length and includes 13 protein -coding genes, 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes, and a control region. Compared to eight known mitochondrial genome, 2 additional noncoding regions (lengths of 121 and 107 bp), or so-called inserts, are found in the intergenic regions 12S rRNA - tRNAVal - 16S rRNA. The presumed origin of the two rare inserts is from tRNA- related retrotransposons. Compared with cytochrome b gene, the two insert sequences are highly conserved at the intraspecies level, but they showed significant variation and low similarity (

Abstract Background Adult worms of Setaria equina mainly found in the peritoneal cavity of equine. They were nonpathogenic but might induce varied degrees of peritonitis and might migrate to the eye, brain, lung, and scrotum causing lacrimation, blindness, paraplegia, locomotor, and neurological disturbances. Identification by light microscopy is insufficient to differentiate Setaria species, and so scanning electron microscopy (SEM) is required to observe their ultrastructures. The study was performed on 80 donkeys from May 2018 to January 2019 for the detection of microfilaria in blood and the adult worms in the peritoneal cavity. The blood samples were either stained with Giemsa stain or examined by modified Knott’s technique for the detection of microfilariae. Adult worms were morphologically characterized based on light microscope and scanning electron microscopy (SEM). PCR was performed targeting the 12S rRNA gene followed by sequencing and phylogenetic analysis. Results The current study recorded 21.6% and 16.2% prevalence rates for adult worms and microfilariae, respectively. By using SEM, this study was able to clarify the detailed structure of amphids, predeirids, vulva, arrangement, and number of male caudal papillae. PCR amplified products for 12S rRNA gene (408 bp) for adult worm and microfilaria. Sequence and phylogenetic analysis revealed that S. equina isolated in the current study from donkeys in Egypt (accession no., MH345965) shared 100% identity with isolates from horse and man in Italy and Iran, respectively and clustered in the same clade with S. digitata, S. tundra and S. labiatopapillosa. Conclusions Identification with light microscopy lacked the ability to characterize different Setaria species, and so using scanning electron microscopy is considered a good choice to distinguish the ultrastructures. In addition, performing the phylogenetic analysis was necessary to detect relationships between different filarial worms, which could not detect by the morphological characterization of adult worms.

Objective A molecular study of the buffalo genetic structure can be effective for better understanding the origin of this animal. Among the molecular markers, mitochondrial genomic sequencing is one of the best and most commonly used methods for genetic classification of populations and species close together, studying the possibility of deriving different species from a common ancestor, studying the phylogenic relationship of each species with other species and races, and obtaining solutions for conservation genetic resources. The purpose of this research was to determine the sequence of 12s rRNA and 16s rRNA regions of Khouzestan buffalo. Materials and methods For this study, 30 blood samples collected from both sexes of unrelated Khuzestan buffalo. After DNA extraction, the target regions amplified by corresponding specific primers by PCR technique and sequenced after purification. Results The results of matching the sequences of 12srRNA and 16srRNA regions of Khuzestan buffalo showed that there was no mutation in this population, which indicates low diversity in Khuzestan buffalo population. According to this study, the 12srRNA and 16srRNA regions are encoding regions and the mutation and diversity are low. The results of the phylogenetic test using UPGMA for both sites showed that the buffaloes of Iran with Indian and Italian buffaloes are in a closely spaced cluster. Conclusions Finally, the sequences generated from these regions were the first recorded in the gene bank with MG650115 access code and the name of the Khuzestan buffalo brought to the World Bank Gene, and this race introduced to international associations.

Copyright of Journal of Hydrobiology is the property of Editorial Department of Journal of Hydrobiology and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The phylogeny of European crayfish fauna, especially with respect to Eastern European species, is still far from being completely resolved. To fill this gap, we analyzed most of the European crayfish species focusing on the phylogenetic position of the endemic crayfish Astacus colchicus, inhabiting Georgia. Three mitochondrial and one nuclear marker were used to study evolutionary relationships among European crayfish species, resulting in the unique phylogenetic position of A. colchicus indicating independent species status to A. astacus. Phylogenetic analyses revealed a deep molecular divergence of A. colchicus in comparison to A. astacus (6.5–10.9% in mtDNA and 1.1% in nDNA) as well as to Pontastacus leptodactylus and P. pachypus (5.5–10.0% in mtDNA and 1.4–2.4% in nDNA). Absent ventral process on second male pleopod and abdominal somites II and III with pleura rounded lacking prominent spines clearly indicate taxonomic assignment to the genus Astacus; however, the species is distributed almost in the middle of Ponto‐Caspian area typical by occurrence of the genus Pontastacus. Several morphological indices linked to head length, carapace, and total body length and width were found to demonstrate apparent differences between A. colchicus and A. astacus. Although this study provides a novel insight into European crayfish phylogeography, we also point out the gaps in comprehensive study of the P. leptodactylus species complex, which could reveal details about the potential species status of particular species and subspecies within this genus. [ABSTRACT FROM AUTHOR]

The 12S rRNA gene is one of unique regions in mitochondrial genome usually used for phylogenetic studies and species identification. The objective of present study was to develop species specific primers from mitochondrial 12S rRNA gene for identification of dog and rat in beef by using multiplex PCR assay. Three primer pairs of mitochondrial 12S rRNA gene specific for bovine, dog and rat were designed and selected to evaluate their specificity and fidelity. Moreover, a total of twelve DNA samples extracted from meat tissue were also prepared to test those primers using simplex and multiplex PCR. The PCR products were then visualized using 2% of agarose gel under the UV light and three of them were sequenced. In addition, sequence data were analyzed using Clustal Omega software and BLAST. The result showed that simplex PCR assay successfully amplified DNA targets which are respectively indicated by 155 bp (bovine), 244 bp (dog), and 491 bp (rat) of DNA bands. Furthermore, DNA sample sequences were identically similar to reference sequence used in this study. Multiplex and simplex PCR analyses also indicated that these primer pairs specifically amplified DNA target for each species in the samples containing various species. The results suggested that designed primers in this study could be used to identify dog and rat in raw beef containing these species meat. Further experiment should be conducted using meat-processed products and commercial meat products as samples.

Molecular tools have been used in wildlife forensics for species identification of indiscernible samples and to provide evidence against poaching of wildlife and illegal trade of their products. This study was undertaken in identifying a confiscated material that has been seized under the Wildlife (Protection) Act, 1972 of India suspecting that accused has poached a deer species of wild origin for local consumption. The sample was processed for DNA sequencing of the partial fragments of three mitochondrial genes, that is, Cytochrome b, 12S rRNA, and 16S rRNA genes. Surprisingly, generated sequences from the confiscated sample showed 99% homology with Bos indicus, and phylogenetic analysis also clustered the confiscated material with Bos indicus with high bootstrap support. This study upholds the evidence for accused as the suspicion of confiscated material being an illegal product was negated, and the raw meat of cattle was merely for consumption through reliable assessment and validation from a public database. This case study exhibits the value of DNA barcoding and molecular diagnostic techniques to decipher the morphologically altered materials in wildlife forensic cases.

In this study we investigated morphological and molecular variation within the predatory mite Typhlodromus (Anthoseius) recki , and their relationships to ecological features. In total, 42 morphological characters were measured on 87 specimens from seven populations in the south of France and Sicily living on plants of four families. DNA sequences (two mitochondrial markers) and the amino acid sequences of the CytB protein were assessed. A relationship between morphological variation and plant families was observed. The 12S rRNA gene showed differentiation that appeared to be related to feeding habit, in agreement with the findings for two other Phytoseiidae species. CytB mitochondrial DNA showed variation related to geographical location. Four amino acid mutations separated the Sicilian and the French populations. CytB amino acid sequences were analysed for three other Phytoseiidae species, and again diagnostic mutations associated with geographical location were observed, as already shown for Phytoseiulus macropilis. The population differentiation observed for each marker (morphological, DNA fragments) appeared to be related to ecological/biological features, revealing new perspectives for forecasting functional characteristics based on morphotypes and genotypes. However, additional studies are needed to confirm these observations and to explain such functional relationships. [ABSTRACT FROM AUTHOR]

Gobioid larvae collected from the coast of Okinawa Island, southern Japan, were identified on the basis of a combination of morphological characters and a sequence of a hypervariable region of mitochondrial 12S ribosomal RNA (12S rRNA) gene (138–145 base pairs). A short-term formalin fixation technique enabled identification using both morphological and genetic methods. Thirteen of the 21 types of gobioid larvae assessed were identified to the species level. Additionally, we described the morphology of the postflexion larvae of wormfish Paragunnellichthys sp. and shrimp-associated goby Ctenogobiops feroculus, identified for the first time in their respective genera. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Hydrobiology is the property of Editorial Department of Journal of Hydrobiology and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract Background The zooplanktonic cladocerans Daphnia, present in a wide range of water bodies, are an important component of freshwater ecosystems. In contrast to their high dispersal capacity through diapausing eggs carried by waterfowl, Daphnia often exhibit strong population genetic differentiation. Here, to test for common patterns in the population genetic structure of a widespread Holarctic species, D. galeata, we genotyped two sets of populations collected from geographically distant areas: across 13 lakes in Eastern China and 14 lakes in Central Europe. The majority of these populations were genotyped at two types of markers: a mitochondrial gene (for 12S rRNA) and 15 nuclear microsatellite loci. Results Mitochondrial DNA demonstrated relatively shallow divergence within D. galeata, with distinct haplotype compositions in the two study regions but one widely distributed haplotype shared between several of the Chinese as well as European populations. At microsatellite markers, clear separation was observed at both large (between China and Europe) and small (within Europe) geographical scales, as demonstrated by Factorial Correspondence Analyses, Bayesian assignment and a clustering method based on genetic distances. Genetic diversity was comparable between the sets of Chinese and European D. galeata populations for both types of markers. Interestingly, we observed a significant association between genetic distance and geographical distance for D. galeata populations in China but not in Europe. Conclusions Our results indicate relatively recent spread of D. galeata across wide expanses of the Palaearctic, with one mtDNA lineage of D. galeata successfully establishing over large distances. Despite a clear differentiation of Chinese and European D. galeata at a nuclear level, the pattern of genetic variation is nevertheless similar between both regions. Overall, our findings provide insights into the genetic population structure of a cladoceran species with extremely wide geographical range.

The Sumatran Striped Rabbit (Nesolagus netscheri ) is a naturally rare, which is a protected animal based on Government Regulation Number 7 of 1999 and renewal of Permen LHK No. P.106 which are categorized as Data Deficient by the IUCN (International Union of Conservation of Nature) since 2019. Samples were obtained from Pagar Alam, South Sumatra. Most commonly found at elevations above 600 m in montane and sub-montane primary habitat. Molecular genetic characterization of sumatran striped rabbit is to know genetic information and genetic identification of N. netscheri. Primers 12S rRNA were used in this study to characterize Sumatran striped rabbit. A 1 band were detected ranged from 900 to 1000 bp. Molecular markers represent reliable tools which may have a great impact in rabbit breeding and genetic improvement of rabbits. Molecular markers on Nesolagus netscheri is expected to tools in the identification of Sumatran striped rabbits from South Sumatra.

Abstract Background The ixodid tick Rhipicephalus appendiculatus is the main vector of Theileria parva, wich causes the highly fatal cattle disease East Coast fever (ECF) in sub-Saharan Africa. Rhipicephalus appendiculatus populations differ in their ecology, diapause behaviour and vector competence. Thus, their expansion in new areas may change the genetic structure and consequently affect the vector-pathogen system and disease outcomes. In this study we investigated the genetic distribution of R. appendiculatus across agro-ecological zones (AEZs) in the African Great Lakes region to better understand the epidemiology of ECF and elucidate R. appendiculatus evolutionary history and biogeographical colonization in Africa. Methods Sequencing was performed on two mitochondrial genes (cox1 and 12S rRNA) of 218 ticks collected from cattle across six AEZs along an altitudinal gradient in the Democratic Republic of Congo, Rwanda, Burundi and Tanzania. Phylogenetic relationships between tick populations were determined and evolutionary population dynamics models were assessed by mismach distribution. Results Population genetic analysis yielded 22 cox1 and 9 12S haplotypes in a total of 209 and 126 nucleotide sequences, respectively. Phylogenetic algorithms grouped these haplotypes for both genes into two major clades (lineages A and B). We observed significant genetic variation segregating the two lineages and low structure among populations with high degree of migration. The observed high gene flow indicates population admixture between AEZs. However, reduced number of migrants was observed between lowlands and highlands. Mismatch analysis detected a signature of rapid demographic and range expansion of lineage A. The star-like pattern of isolated and published haplotypes indicates that the two lineages evolve independently and have been subjected to expansion across Africa. Conclusions Two sympatric R. appendiculatus lineages occur in the Great Lakes region. Lineage A, the most diverse and ubiquitous, has experienced rapid population growth and range expansion in all AEZs probably through cattle movement, whereas lineage B, the less abundant, has probably established a founder population from recent colonization events and its occurrence decreases with altitude. These two lineages are sympatric in central and eastern Africa and allopatric in southern Africa. The observed colonization pattern may strongly affect the transmission system and may explain ECF endemic instability in the tick distribution fringes.

Poaching of wildlife for local consumption and commercial purposes is a major threat to the loss of the world's biodiversity which has even resulted in the extirpation of the numerous species at local or regional scale. Law enforcement agencies have been working in the implementation of the Wildlife Protection Act to combat illegal trade of wildlife for parts and products all across the world. Nevertheless, several times confiscated materials have been altered and fabricated so much that their morphological identification often got difficult. In the last few decades, DNA technology has revolutionized species identification in forensic investigations and has shown its power in the firm identification of those fabricated materials. We received a confiscated material, appeared to be chopped raw meat with no intact morphological identity. The accused was caught by the officials of the State Forest Department after he put pictures on social media depicting his involvement in cooking and eating of animal meat, suspected to be of wildlife origin. On homology search, we found a 99% identity of the confiscated material with Asian Palm Civet (Paradoxurus hermaphroditus) and neighbour joining trees based on genetic distances clustered the query sequence with the Asian Palm Civet with 100% bootstrap support. The present study exhibits hope for ascertaining species identification from the processed meat in reliable assessment for wildlife forensics. This case study extends, the role of the authenticated references as available on the public database, highlighting the application of DNA forensics in identifying species even from the phenotypically altered materials.

A list of terrestrial enchytraeids of the Russian Far East is compiled based on literature and extensive field data collected by the authors in 2019. A database has been created consisting of geographic coordinates, habitat type, species, and data source. For some species collected by the authors, barcoding using COI, 16s, and 12s rRNA genes has been performed. In total, there are at least 62 species of enchytraeids belonging to 12 genera. Seven species (Achaeta macroampullacea, Cognettia sphagnetorum, Enchytraeus dichaetus, Fridericia cusanica, Globulidrilus riparius, Marionina southerni, Mesenchytraeus gigachaetus) are reported in the Russian Far East for the first time. Cognettia sphagnetorum and F. cusanica are most probably introduced. Taxonomic and biogeographical remarks on some of the species found and differences from the original descriptions are provided. Some of the specimens may be undescribed species, but this requires a more in-depth examination. The Russian Far East, especially its southeastern part, is of great interest as a possible location for new species of enchytraeids. [ABSTRACT FROM AUTHOR]

The clam Meretrix lamarckii is an ecologically and economically important species in the coastal regions of China. In order to characterize the genetic diversity and population structure of M. lamarckii, a 750 bp region of mitochondrial COIII gene and an 822 bp region of 12S rRNA gene were sequenced and analyzed for 118 and 105 individuals from five populations (ZS, WZ, ZP, ST and ZJ) in the Southeast coastal areas of China, respectively. Results revealed that 33 haplotypes were defined in COIII gene, and 30 haplotypes were defined in 12S rRNA gene. The pairwise Fst values between the ZS population and other four populations were range from 0.122~0.154 (COIII, P< 0.05) and 0.052~0.228 (12S rRNA, P<0.05), respectively. The revealed that ZS population was significant divergence from other four populations, and no significant genetic divergence among WZ, ZP, ST and ZJ populations. Moreover, results from the median-joining network, plot of STRUCTURE and the UPGMA trees were similar to pairwise Fst values. Results from AMOVA indicated that the genetic variation of M. lamarckii populations was mainly from the variation within populations (COIII: 93.27%, 12S rRNA: 92.32%). The results of neutrality tests combined with the mismatch distribution indicated recent population expansion of M. lamarckii on large spatial scales in the period of late Pleistocene (0.1-0.12 Ma). [ABSTRACT FROM AUTHOR]

The true phylogenetic relation among Epinephelus species is under debate. The present investigation was designed for evaluation of DNA polymorphism in some Red sea Epinephelus species using 12s rRNA and Inter Simple Sequence repeats. DNA polymorphism values were detected and evaluated within all estimated fishes. Both applied techniques revealed that E. malabaricus is closely related to E. summana. The distance value between E. chlorostigma and E. areolatus is lower than the distance value between E. chlorostigma and E. radiatus. The Serranidae evolutionary variations were evaluated comparatively with other ray-finned fishes belonging to four fish genera (Carangidae, Labridae, Mugilidae and Cichlidae) based on 12s rRNA sequence variations (obtained from NCBI). The developed DNA markers were reliably branched in the evaluated fishes. More molecular investigations using more spatial and temporal fish samples should be carried out in the future for exploring the true Epinephelus species biodiversity in the Red sea. [ABSTRACT FROM AUTHOR]

Codfish is an essential part of the marine fishery resources, and contains several commercially important species, including Gadus species (G. morhua, G. macrocephalus, and G. chalcogrammus), and other five Gadiformes species, including Pollachius virens, Melanogrammus aeglefinus, Merluccius merluccius, Merluccius australis and Albatrossia pectoralis. According to our previous studies, Gadus species represents the most frequently used Gadiformes species for codfish products, with other five Gadiformes species also occasionally found. However, due to the legislative shortcomings, there is still not a harmonization around the definition of codfish in China, leading to great chaos. To help prevent this episode, the present study aimed to develop a one-tube fast method for codfish species identification, with particular attention to differentiate Gadus species, and the other five Gadiformes species, using the high resolution melting (HRM) analysis of a 12S rRNA mini barcode. The specificity of the newly designed primers was successfully confirmed for the eight common Gadiformes species. The novel HRM assay allowed the full discrimination between Gadus species and the other five Gadiformes species. 19 commercial codfish products were included in the Gadus cluster, cross-confirmed by the real-time PCR and DNA barcoding. The identification of less valuable U. japonicas is essentially a consumer fraud, while great health concern could also be highlighted by the identified Lagocephalus species. The established HRM of a 12S rRNA mini barcode method in the present study allows a fast and accurate identification of codfish species in processed fish products. [ABSTRACT FROM AUTHOR]

Summary: Using species outside the order Gadiformes to prepare codfish products has been widely revealed as a new type of fraud in China, highlighting the necessity to establish proper methods for codfish species identification. This work aimed to develop a new SYBR Green real‐time PCR assay for the detection of codfish ingredient (mainly the eight Gadiformes species, Gadus morhua, Gadus macrocephalus, Theragra chalcogramma, Pollachius virens, Melanogrammus aeglefinus, Merluccius merluccius, Merluccius australis and Albatrossia pectoralis) in processed fish products based on mitochondrial 12S rRNA gene. Results highlighted the positive Cq (cycle quantification value, average value 19.18 ± 1.49) only from the eight Gadiformes species, with no fluorescent signal (Cq > 32) for the other sixty‐seven non‐Gadiformes species. The absolute limit of detection was 0.048 ng genomic DNA. Methodology validation succeeded in twenty‐six commercial products. These results demonstrated that the newly developed method is suitable for the identification of codfish ingredient in processed fish products. [ABSTRACT FROM AUTHOR]

Background: The blood samples of spots typing regards most important step in forensic analysis. Determining the blood affiliation as human on non-human is not enough as a forensic tool leading to the truth . The mitochondrial 12S rRNA gene are suitable for taxonomy and species identification, especially for the discrimination of even closely related species. The current study aimed to showed the validity of designed 12S rRNA species specific primer pairs for human and nonhuman animal blood typing as a forensic tool. Methodology: Seventy two blood samples were collected from Homo sapiens, Ovis aries, Capra hircus and Bos taurus (18 blood samples from each). Five milliliter were withdrawn and placed in EDTAtube and stored in refrigerator for further processing. Four sets of species specific primer pairs targeting mitochondrial 12S rRNA were designed and checked. PCR and sequencing were performed and sequences were analyzed and register in GenBank. Results: the results revealed that, the amplicon of 12S rRNA gene of Homo sapiens were 800bp, Ovis aries were 560bp, Capra hircus were 460bp, and Bos taurus were 600bp The first confirmatory test for validity and specificity of designed primer pairs is no amplification for intraspecies primer pairs (i.e. using homo12S rRNA primer pairs to amplify Ovis aries, Capra hircus, and Bos taurus 12S rRNA will not give product and vice versa). The second confirmatory test is the results of sequencing. The identity percentage and alignment of sequences results of amplified 12S rRNA gene of homo sapiens, Ovis aries, Capra hircus and Bos taurus revealed that, the similarity percentage ranged from 98.03% to 100%.Conclusion: The current study conclude that, validity and accuracy of designed 12S rRNA species specific primer pairs for human and nonhuman animal blood typing as a forensic tool and there is no intraspecies cross amplification. [ABSTRACT FROM AUTHOR]

Although a significant unexploited potential of the geometric morphometrics in taxonomy and systematics has been documented, only a few studies so far have focused on the utility of this method in the detection of population genetic structure in turtles. For the first time, we employed geometric morphometrics to explicitly analyse whether populations possessing divergent haplotypes evolved different patterns of plastral shape in any Testudo species using the Greek populations of Hermann's tortoise as a case study. To molecularly characterize the Greek populations of Testudo hermanni boettgeri used for plastron shape analyses, 12s rRNA gene of tortoises sampled from seven localities in Greece was sequenced. Three divergent haplotypes were observed, regionally corresponding to the north‐western (HI), south‐western (HII) and Peloponnese (HIII) areas of Greece. In both females and males, haplotypes HI and HII showed the lack of plastral shape divergence, but were both statistically significantly different relative to HIII found in Peloponnese. The Peloponnesian populations (HIII) had longer bridges and the wider and shorter anterior part of the plastron, relative to populations from the mainland Greece (HI and HII). Hence, both morphological and molecular markers suggested Peloponnese as a distinct lineage in relation to mainland populations. Finally, we consider geometric morphometrics as an efficient method to detect and characterize subtle interpopulation differences. Findings in this study have conservation applications as well, implying the presence of cryptic genetic and taxonomic diversity, and, thus, filling the crucial gap in the assessment of the conservation status of T. hermanni. [ABSTRACT FROM AUTHOR]