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The genus Rhonciscus Jordan & Evermann, 1896 (Haemulidae) was recently revalidated, and it brought together four species allocated along with Pomadasys Lacepède, 1802, two from the Western Atlantic and two from the Eastern Pacific. However, the status of some of the species in the western South Atlantic was still doubtful. This study aims to review Rhonciscus species from the Western Atlantic based on morphological and genetic data. Our results bring the re-creation and redefinition of the limits of Rhonciscus crocro (Cuvier, 1830) and R. approximans Bean & Dresel, 1884 with the recently described R. pauco Tavera, Schärer-Umpierre & Acero P., 2022 as a junior synonym, with a description of a new species of Rhonciscus. The new species, distinguished by a very long snout and short upper jaw, is from the Brazilian coast. A further evaluation of the status of Pristipoma cultriferum Poey, 1860 and Pristipoma ramosum Poey, 1860, described from Cuba, is also provided. [ABSTRACT FROM AUTHOR]

Baikal amphipods are one of the most unique freshwater species flocks of over 350 morphological species and subspecies playing multiple ecological roles in this ancient lake. However, molecular phylogeny studies accumulate evidence that this number is underestimated. Previously, we showed that an abundant littoral species Eulimnogammarus verrucosus comprised at least three genetic lineages: southern (S), western (W), and eastern (E), inhabiting corresponding parts of Baikal shore. The Angara River is the only outflow of the lake and is inhabited by amphipods originating from Baikal, which were previously analysed only morphologically. The river has parts with strong current or closed bays as opposed to sea‐like conditions of Lake Baikal, which raises the question of which species were able to adapt here. Here we studied the phylogeny of E. verrucosus from the Angara River, which could be reached by two lineages, S and W. We analysed 17 sites from 0.6 to 650 km from the outflow. Cytochrome c oxidase subunit I (COI) and 18S rRNA analysis showed that the sites closest to Baikal were inhabited by the S and W lineages sympatrically. The other sites were inhabited by the S lineage at both banks and also by the novel A (Angara) lineage. The A lineage was the first to split up from the last common ancestor of all E. verrucosus. Our data confirm the importance of considering cryptic diversity in ecological monitoring, and in order to simplify the identification of the genetic lineage, we developed a web‐app speCOIdent. [ABSTRACT FROM AUTHOR]

The Ili River Valley, located in the northwest of China, serves as a vital repository for fish genetic resources. Its extensive water network and diverse climate have given rise to a unique fish composition and endemic species. In this study, we collected the cytochrome c oxidase subunit I (COI) sequences from 660 fish specimens in the Ili River Valley. The effectiveness of DNA barcoding in identifying fish species in the area was assessed by examining genetic distances, constructing phylogenetic trees, and performing ABGD (Automatic Barcode Gap Discovery) analyses, among other methods. In total, 20 species were identified, including one unidentified species (Silurus sp.). Except for Silurus asotus and Hypophthalmichthys molitrix (only one sample), the maximum intraspecific genetic distance among the remaining species was smaller than the minimum interspecific distance, which proves that the species exhibit obvious barcode gaps. In the Neighbor‐Joining trees, 20 species formed separate monophyletic branches. According to ABGD analysis, 660 sequences were categorized into 19 Operational Taxonomic Units, with Silurus sp. and S. asotus grouped into a single OTU. The Silurus in this study exhibits shared haplotypes and significant genetic divergence, suggesting the potential presence of cryptic species. Furthermore, the nucleotide diversity across all species fell below the threshold level, indicating that the local fish population is gradually declining. In conclusion, this study has demonstrated the effectiveness of DNA barcoding in identifying fish species in the Ili River Valley, providing valuable data to support the conservation of local fish resources. [ABSTRACT FROM AUTHOR]

Sciadonus alphacrucis Melo, Gomes, Møller & Nielsen, 2022 is a rare deep‐sea species, previously known from only two specimens collected off São Paulo State, southeastern Brazil, in the western South Atlantic. Herein, we report a new specimen of S. alphacrucis collected on the continental slope off Santa Catarina State, southern Brazil, thereby extending its known distribution by 420 km. Additionally, we provide the new meristic and morphometric data, the molecular identification using sequences of the cytochrome c oxidase subunit I (COI), an updated distribution map, and a discussion of troglomorphic traits. [ABSTRACT FROM AUTHOR]

Identification of fish larvae based on morphology is typically limited to higher taxonomic ranks (e.g., family or order), as larvae possess few morphological diagnostic characters for precise discrimination to species. When many samples are presented at any one time, the use of morphology to identify such specimens can be laborious and time‐consuming. Using a reverse workflow for specimen sorting and identification leveraging high‐throughput DNA sequencing, thousands of fish larvae can be DNA barcoded and sorted into molecular operational taxonomic units (mOTUs) in a single sequencing run with the nanopore sequencing technology (e.g., MinION). This process reduces the time and financial costs of morphology‐based sorting and instead deploys experienced taxonomists for species taxonomic work where they are needed most. In this study, a total of 3022 fish larval specimens from plankton tows across four sites in Singapore were collected and sorted based on this workflow. Eye tissue from individual samples was used for DNA extraction and sequencing of cytochrome c oxidase subunit I. We generated a total of 2746 barcodes after quality filtering (90.9% barcoding success), identified 2067 DNA barcodes (75.3% identification success), and delimited 256 mOTUs (146 genera, 52 families). Our analyses identified specific challenges to species assignment, such as the potential misidentification of publicly available sequences used as reference barcodes. We highlighted how the conservative application and comparison of a local sequence database can help resolve identification conflicts. Overall, this proposed approach enables and expedites taxonomic identification of fish larvae, contributing to the enhancement of reference barcode databases and potentially better understanding of fish connectivity. [ABSTRACT FROM AUTHOR]

Backgroud: The Northeast India, being part of two global biodiversity hotspot namely the Indo-Burma and Eastern Himalayan Hotspots supports a wide variety of rich aquatic biodiversity including fishes. The family Danionidae is a widely diverse group inhabiting the upper colder stretches of river although few are abundant in the lower stretches. The persisting similarity in the morphological appearance and body colouration within the members of this family seeks an integrated method to identify the species correctly. Methods and results: In the present study, the mt-DNA barcode was generated for correct identification and confirmation of the species. A total of nine mitochondrial cytochrome c oxidase subunit I gene sequences were generated for each species under the study. The pairwise distance values ranged from 0.09 to 9.11% within species and 9.06–32.71% between species. A neighbour-joining tree was constructed based on the Kimura 2 parameter model. Two major groups were observed where Danioninae formed a sister group to the Chedrinae and Rasborinae. Conclusion: The present study is a preliminary work to document and identify the species under the family Danionidae from Brahmaputra basin, Assam, using molecular tools and establish the phylogenetic relationship. [ABSTRACT FROM AUTHOR]

Abstract The Ili River Valley, located in the northwest of China, serves as a vital repository for fish genetic resources. Its extensive water network and diverse climate have given rise to a unique fish composition and endemic species. In this study, we collected the cytochrome c oxidase subunit I (COI) sequences from 660 fish specimens in the Ili River Valley. The effectiveness of DNA barcoding in identifying fish species in the area was assessed by examining genetic distances, constructing phylogenetic trees, and performing ABGD (Automatic Barcode Gap Discovery) analyses, among other methods. In total, 20 species were identified, including one unidentified species (Silurus sp.). Except for Silurus asotus and Hypophthalmichthys molitrix (only one sample), the maximum intraspecific genetic distance among the remaining species was smaller than the minimum interspecific distance, which proves that the species exhibit obvious barcode gaps. In the Neighbor‐Joining trees, 20 species formed separate monophyletic branches. According to ABGD analysis, 660 sequences were categorized into 19 Operational Taxonomic Units, with Silurus sp. and S. asotus grouped into a single OTU. The Silurus in this study exhibits shared haplotypes and significant genetic divergence, suggesting the potential presence of cryptic species. Furthermore, the nucleotide diversity across all species fell below the threshold level, indicating that the local fish population is gradually declining. In conclusion, this study has demonstrated the effectiveness of DNA barcoding in identifying fish species in the Ili River Valley, providing valuable data to support the conservation of local fish resources.

Faunal studies of oceanic islands provide unique opportunities to study ecology and evolution. Four Armadillidae taxa were recorded from the Izu Islands, a group of oceanic islands off the coast of Japan. The distributions of these taxa and a phylogenetic tree are based on molecular data for 249 specimens from 88 sites. The Spherillo russoi (Arcangeli, 1927) species complex is distributed widely throughout these islands, and dispersal may have occurred via oceanic currents. On the nearby Izuoshima and Toshima Islands, Spherillo dorsalis (Iwamoto, 1943) may have recently migrated from the Izu Peninsula either via currents or been transported by human activity. In the middle islands, Shikinejima, Kozushima, Miyakejima, and Mikurajima, Venezillo hasegawai (Nunomura, 1991) was found, which may be derived from Spherillo obscurus (Budde-Lund, 1885) distributed in the Kanto region on the mainland. On the most southern and distant Hachijojima Island, Spherillo punctatus Nunomura, 2007 was found, and it does not differ genetically from specimens from Izenajima and Tokunoshima Islands in the Nansei Island Group, despite an approximate 1,000-km distance between them; this species may have been transferred by human activity. Our study showed that the Armadillidae assemblages in the Izu Islands were formed by several migration routes.

Studies were conducted in four consecutive seasons from kharif, 2022 to rabi, 2023, on onion crop to understand the thrips, Thrips tabaci Linderman incidence. The identity of the species was confirmed through morphological characteristics as well as sequence amplified product of Cytochrome c oxidase subunit I (COT) gene (GenBank No. PP838743). The phylogenetic analysis revealed that the T. tabaci population is closely related to the sequence of MZ882441 and MT991561 from China and New Delhi, India, respectively. Incidence in the Kharif season has experienced minimal thrips infestation compared to the Rabi season, where severe infestation was noted. The peak thrips population occurred in the 6th Standard Meteorological Week (SMW) of 2022 and 2023, highlighting the role of weather in thrips population dynamics. Correlation analysis indicated that maximum temperature exhibited a significant positive correlation, while relative humidity showed a significant negative correlation. Additionally, rainfall was found to have a cleansing effect on thrips populations, resulting in lower incidences during the Kharifseason. However, during the Rabi season, the coincidence of higher temperatures with low relative humidity contributed to the proliferation of thrips, making them a significant threat to bulb onion production. This study provides a comprehensive information on the occurrence, species confirmation, nature of damage, seasonal incidence and associated weather parameters with T. tabaci in onion crops. [ABSTRACT FROM AUTHOR]

Subterranean termites (Family Rhinotermitidae) and drywood termites (Family Kalotermitidae) can be pest species as they have the potential to inflict damage to wooden constructions and can therefore cause a significant economic impact. Some species of these families are highly destructive and are invasive in many parts of the world. New introductions and/or the spread of termites are often caused by import of infested wood, plants or soils. The present study reports the first records of three exotic termite species in Belgium, viz. Cryptotermes brevis and Reticulitermes banyulensis in Brussels (Brussels Capital Region), and Reticulitermes flavipes in Brugelette (Hainaut province; Walloon region). The morphological identification of the specimens (N = 8) was validated by DNA-barcoding. Cryptotermes brevis and Reticulitermes banyulensis infestations are probably of little concern as local climatological conditions make it unlikely for either species to survive in the temperate climate in Belgium. In contrast, it is likely that Reticulitermes flavipes may become established and will spread to other sites in Belgium. [ABSTRACT FROM AUTHOR]

Carassius gibelio (Prussian carp) is the latest invader of Canada's fresh waters, with reports of its presence in Alberta and Saskatchewan. Morphological similarities with introduced C. auratus (goldfish) and the putatively introduced (but likely misidentified) C. carassius (crucian carp) make it difficult to distinguish these species without careful examination. DNA barcoding of the mitochondrial cytochrome c oxidase I (COI) gene is a potential tool to identify Carassius individuals to species, but incorrect annotations of publicly-available sequences can confound attempts at species identification. Here we use morphology and DNA barcoding to identify putative C. gibelio specimens collected from two Alberta locations that constituted new records for the province. Morphologically, specimens were consistent with C. gibelio but were in the range for both C. gibelio and C. auratus. Genetically, our samples could not be identified to species level, being a 100% match with multiple Carassius species. Haplotype networks, however, supported the identification of Alberta carp as C. gibelio, in agreement with statistical analysis. Furthermore, the Alberta haplotype was shared with a fish reported from the seafood trade, suggesting a possible source of entry into Alberta. Therefore, although the BOLD algorithm would suggest the COI gene is not a strong candidate for species-level identification of Carassius species, a haplotype network approach and statistical examination of variability among haplotypes can be used to make reasonable inferences as to species identity. Early detection and management of Carassius species are of critical importance due to their projected ecological impact in Canada; DNA barcoding is one important tool for species identification, especially when specimens are within the phenotypic range expected for multiple species. [ABSTRACT FROM AUTHOR]

Oxynoemacheilus chaboras, new species, from the stream Beyazsu in the Euphrates drainage, belongs to the O. persa species group, being closely related to O. shehabi from the Orontes, O. sarus from the Seyhan and Ceyhan, O. euphraticus from the Euphrates and Tigris, O. karunensis from the Karkheh, and O. persa from Central Iran. The new species is distinguished from others in the O. persa group by having 8–9 pores in the supraorbital canal, two distinct black blotches at the caudal-fin base, a rudimentary and shallow pelvic axillary lobe, 6–10 irregularly shaped bars on the flank, and a deep head, body, and caudal peduncle. Oxynoemacheilus chaboras sp. nov. is most closely related to O. euphraticus, from which it is differentiated by a mean uncorrected p-distance of 3.24% (min. 3.09%) in its COI barcode gene. [ABSTRACT FROM AUTHOR]

Simple Summary: The catfish of the genus Pimelodus are amply distributed in the Neotropical region, although the species-level taxonomy and phylogenetic relationships of these fish are still poorly resolved. In the present study, we used a molecular approach to delimit the Pimelodus species from the different river basins of the Neotropical region. For this, we analyzed sequences of the mitochondrial Cytochrome c oxidase subunit I (COI) gene from 13 nominal species, which generated 24 consensus Molecular Operational Taxonomic Units (MOTUs). Only six of the nominal species were recovered as well-defined molecular entities, while seven presented cryptic diversity or taxonomic uncertainties. The DNA barcode analysis presented here represents an important step toward the definition of the species of this economically important group of fish, which will be fundamental to the conservation of its diversity. Pimelodus is the most speciose genus of the family Pimelodidae, and is amply distributed in the Neotropical region. The species-level taxonomy and phylogenetic relationships within this genus are still poorly resolved, however. These taxonomic problems and the general lack of data have generated major uncertainties with regard to the identification of specimens from different localities. In the present study, we applied a single-locus species delimitation approach to identify the MOTUs found within the genus Pimelodus and provide sound evidence for the evaluation of the species richness of this genus in the different river basins of the Neotropical region. The study was based on the analysis of sequences of the mitochondrial COI gene of 13 nominal species, which resulted in the identification of 24 consensus MOTUs. Only six nominal species were recovered as well-defined molecular entities by both the traditional barcoding analysis and the molecular delimitation methods, while the other seven presented cryptic diversity or persistent taxonomic uncertainties. The lineages identified from the Parnaíba ecoregions, Amazonas Estuary and Coastal Drainages may represent a much greater diversity of Pimelodus species than that recognized currently, although a more detailed study of this diversity will be necessary to provide a more definitive classification of the genus. [ABSTRACT FROM AUTHOR]

Purpose: Myiases are infestations of human and animal tissues by fly larvae. These conditions are widespread in tropical countries and travelers in those areas are at risk of becoming infested. Although Cordylobia anthropophaga (Blanchard & Berenger-Feraud, 1872) is one of the most common myiasis-causing species, few high-quality images and molecular sequences are available for this fly. We present a case of C. anthropophaga infestation in an Italian patient returning from Senegal, with the aim of increasing both visual and molecular data for this species. Methods: After removal, the larva was determined following standardized morphological keys and photographed under a digital microscope. Molecular characterization of the Cytochrome c oxidase subunit I (COI) was performed using universal primers. Results: The general appearance, the structural organization of the cephalic region, of the cephaloskeleton, and of the posterior tracheal spiracles suggested that the causative agent of the myiasis was a third instar larva of C. anthropophaga. The morphological data are further supported by the molecular data: the COI sequence showed high levels of identity with the already published verified COI sequences of C. anthropophaga. Conclusion: We provide high-quality morphological and molecular data useful for the identification of larvae of C. anthropophaga. We highlight that myiasis might be common in Senegal and better data about its prevalence in travelers and in the endemic countries are needed to understand the burden of this condition. [ABSTRACT FROM AUTHOR]

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

Simple Summary: Pinna nobilis, a species of marine shellfish living in the Mediterranean Sea, is at a high risk of extinction due to a not-entirely-known disease that started affecting its populations in 2016. In this paper, we reported the main traits of its evolutionary history to understand how this species evolved over time and space from the moment its ancestor entered the Mediterranean. To achieve this goal, we analysed a total of 469 sequences from all over the Mediterranean Sea. Our research showed that P. nobilis evolved from its ancestor about 2.5 million years ago, following a rapid and catastrophic entry of waters from the Atlantic Ocean that pushed the P. nobilis ancestor into the Mediterranean around 5.3 million years ago. Our results also suggest that the central part of the western Mediterranean was the first marine area where this species settled and, later on, it spread to the Adriatic and the eastern part of the basin. This information is of twofold importance, as it helps us to understand how this species adapted to the Mediterranean over time and may be the basis of present and future restocking plans which want to take into consideration the reconstruction of pre-existing genetic variability. Pinna nobilis, commonly known as the noble pen shell, is a marine bivalve endemic to the Mediterranean Sea. Unfortunately, due to a multifactorial disease that began affecting its populations in 2016, the species is currently facing the threat of extinction. To gain insights into the evolutionary history of P. nobilis before the mass mortality event (MME), and to obtain a comprehensive understanding of how evolutionary processes led to the adaptation of the species into the Mediterranean Sea, phylogenetic and phylogeographic analyses were carried out. The dataset analysed includes 469 sequences of COI gene fragment both from GenBank and the present study (100). The analysis performed evidenced that P. nobilis diverged about 2.5 mya, after the entrance of its ancestor into the Mediterranean Sea following the Zanclean flood (5.33 mya). Moreover, our results suggest that the starting point of colonisation was the central part of the western Mediterranean basin, with the eastern basin being populated subsequently. From a conservational viewpoint, these results provide important hints for present and future restocking plans, helping to reconstruct the pre-existing genetic variability in sites where the species became extinct. [ABSTRACT FROM AUTHOR]

Three species of Hemiscorpius were identified in Hormozgan province, for which the available antivenoms lack the efficacy required for treating patients. Consequently, an exact identification of the existing species was deemed necessary as the first step in managing treatment procedures. Considering the morphological similarities among the species, the aim of this research was the molecular study of the samples to accurately identify the species. Hemiscorpius specimens were collected from various locations in Hormozgan province between 2021 to 2023. The Cytochrome c oxidase subunit I gene was amplified and sequenced. Four sequences were obtained from Hemiscorpius specimens collected from Hormozgan province, and three sequences were sourced from the NCBI for analysis. Bayesian inference and Maximum likelihood phylogenetic trees showed similar results, positioning the base of Hemiscorpius enischnochela tree as an older species and Hemiscorpius lepturus adjacent to Hemiscorpius acanthocercus, identified as the newest species at the tree's tip. The results confirmed the validity of three species, namely H. acanthocercus, H. enischnochela, and H. lepturus. Hemiscorpius acanthocercus and H. lepturus are known for having dangerous venom for humans with reported deaths due to their stings. Considering the importance of the members of this genus from the medical point of view, a comprehensive examination of all species is imperative. [ABSTRACT FROM AUTHOR]

Simple Summary: The routine monitoring of zooplankton is difficult due to their small size and morphological ambiguity. Also, the eggs and larva of meroplankton resemble one another, and therefore it is challenging to identify their species. Alternatively, DNA-based tools can provide precise species identifications, regardless of the size or the developmental stage of a specimen. We developed a protocol for testing the potential of DNA metabarcoding for assessing the seasonal and spatial dynamics of zooplankton in a temperate estuary. Both the seasonal and spatial gradients influenced recovered richness, composition, and taxonomic distinctness, confirming the great aptitude of DNA metabarcoding for providing higher density monitoring and shedding new light on the composition and dynamics of complex zooplankton. Zooplankton are key components of estuarine trophic networks. However, routine monitoring is hindered by the difficulty of morphology-based identification. DNA-based methods allow us to circumvent some of these hurdles, providing precise species identifications regardless of the taxonomic expertise of the investigator or the developmental stage of the specimens. However, the process is dependent on the completeness of the reference libraries. In this study, we sought to evaluate the potential of DNA metabarcoding to assess the seasonal (summer, autumn, and early spring) and spatial dynamics of zooplankton (four locations spanning ca. 6 km) in the Lima estuary (NW Portugal). Two genetic markers were used: the cytochrome c oxidase subunit I and the V4 hypervariable region of the ribosomal 18S rRNA genes. Overall, 327 species were recovered, and both markers displayed minute overlap (7% were detected with both markers). Species richness, composition, and taxonomic distinctness were majorly influenced by the season, with a declining tendency from summer (highest number of exclusive species, n = 74) to spring. Second to season, the taxa composition was influenced by spatial variation where the most downstream site displayed the highest number of exclusive species, n = 53. A total of 16 non-indigenous species were detected using metabarcoding, but only one (Austrominus modestus) has been documented out in the estuary. In conclusion, both the seasonal and spatial gradients influenced the recovered richness, composition, and taxonomic distinctness, confirming the great aptitude of DNA metabarcoding for providing higher density monitoring and shedding new light on the composition and dynamics of complex zooplankton communities. [ABSTRACT FROM AUTHOR]

Charopidae is a highly speciose family of land snails mostly distributed in the main landmasses and archipelagos of the Southern Hemisphere, with a few genera in the Northern Hemisphere. DNA sequencing is making substantial contributions to understanding the phylogeography of Charopidae and the systematic relationships of its subfamilies and related families in the Punctoidea. Here, new data from 28S rRNA are combined with available GenBank sequences to investigate whether Australian Charopidae comprise a single clade, exclusive of species from other regions. Phylogenetic analyses show that this is not the case, with most species from Australia and other regions included in a bootstrap-supported clade that excludes some Australian species. Analyses of concatenated 28S rRNA data and available cytochrome c oxidase subunit I (COI) also contradicted monophyly of Australian species, as did analyses of inferred COI protein sequences. In DNA analyses, the genus Hirasea and Australian taxa with an incomplete secondary ureter were basal within Charopidae. We also examined whether known doubts about the taxonomic status of supra-generic clades in Punctoidea could be resolved by considering autapomorphic COI amino acid changes. Such variation could not generally be applied as an objective criterion for taxonomic categorization but did identify some clades for taxonomic investigation. [ABSTRACT FROM AUTHOR]

DNA barcoding represents a handy tool for species identification. In addition, it serves as a complementary approach that improves the characterisation of evolutionary lineages and facilitates the detection of potentially undescribed and cryptic species. Based on the case study in the Western Carpathians, which belong to the Carpathian biodiversity hotspot, we have compiled the first DNA barcode reference library for molecular identification of invertebrates associated with epikarst, a unique, yet understudied, shallow subterranean aquatic habitat that extends at the interface between the soil and carbonate rocks. We analysed invertebrates collected in 2019-2020 from epikarst water that continuously seeps into four caves of the Demänovský Cave System in northern Slovakia. The standard barcode marker of the mitochondrial COI gene was amplified in more than 920 individuals of aquatic, semi-aquatic, and terrestrial invertebrates. The final data set consisted of 784 barcode sequences representing 36 morphospecies, the majority (98.3%) belonging to Arthropoda. Automated cluster delineation using the Barcode of Life Data System (BOLD) revealed 60 Barcode Index Numbers (BINs), of which 43 BINs were new to BOLD, representing mostly typical subterranean species. Almost 20% of the morphospecies displayed high intraspecific variation (>2.2%), suggesting the need for further investigation to assess potential taxonomic problems or cryptic diversity. Our results also indicated the existence of several yet undescribed invertebrate species and possible heteroplasmy or COI numts in the collembolan Megalothorax sp. (incertus species group). The resulting DNA barcode library represents a significant advance not only in the characterisation of epikarst biodiversity but also in the understanding of subterranean biodiversity in general, paving the way for future complex evolutionary and biogeographical studies. [ABSTRACT FROM AUTHOR]

Lake Skadar with its surrounding springs, wetlands and larger affluents is among the most diverse freshwater ecosystems in the Mediterranean region and a key biodiversity/endemism hotspot in Europe. It is also highly endangered due to climate change and rapid tourism development in the area. Being abundant, diverse and mostly predatory, true aquatic bugs play an important role in the functioning of freshwater ecosystems and are used as indicators of aquatic habitat quality. Nevertheless, this taxonomic group has been scarcely studied in the area. Our survey provides the first comprehensive DNA barcode library for 24 out of 25 species of aquatic Heteroptera collected in the Skadar Lake basin and adjacent regions. By this, we extend the list of species known from the area by 60%. In the case of three species, Notonecta maculata, Hydrometra stagnorum and Nepa cinerea, we detected multiple highly divergent, and also new BINs indicating possible taxonomic inconsistencies, the potential for (pseudo)cryptic diversity and intricate phylogeographic patterns. We show that presumably well-known hotspots, such as Lake Skadar region, are heavily understudied regarding even the prominent insect taxa and, thus, particularly vulnerable to undocumented biodiversity loss. Finally, we underline the value of simple DNA-barcoding-based surveys for providing reference barcode libraries for effective biomonitoring and signalling taxonomic and biogeographic issues. [ABSTRACT FROM AUTHOR]

Pelvic spine polymorphism occurs in several species in the stickleback family (Gasterosteidae). Given the similar phenotypic polymorphisms in multiple stickleback species, we sought to determine the extent of parallelism in the ecological correlates of pelvic spine reduction. Based on a metabarcoding analysis of brook stickleback gut contents in two polymorphic populations, we found that significant diet differences were associated with pelvic spine reduction, but we found no clear or consistent trend supporting a tendency for benthic feeding in pelvic‐reduced brook sticklebacks. These results contrast with those found in threespine sticklebacks where pelvic spine reduction is often associated with a benthic diet. Hence, we found non‐parallel consequences of spine polymorphism across species. Furthermore, a difference in gill raker morphology has been frequently observed between ecomorphs with different diets in many fish species. However, we found no evidence of any difference in gill raker morphology associated with pelvic spine polymorphism in brook sticklebacks. [ABSTRACT FROM AUTHOR]

The zoonotic nematode Thelazia callipaeda infects the eyes of domestic and wild animals and uses canids as primary hosts. It was originally described in Asia, but in the last 20 years it has been reported in many European countries, where it is mainly transmitted by the drosophilid fruit fly Phortica variegata. We report the autochthonous occurrence of T. callipaeda and its vector P. variegata in Austria. Nematodes were collected from clinical cases and fruit flies were caught using traps, netting, and from the conjunctival sac of one dog. Fruit flies and nematodes were morphologically identified and a section of the mitochondrial cytochrome c oxidase subunit I gene (COI) was analysed. A DNA haplotype network was calculated to visualize the relation of the obtained COI sequences to published sequences. Additionally, Phortica spp. were screened for the presence of DNA of T. callipaeda by polymerase chain reaction. Thelazia callipaeda and P. variegata were identified in Burgenland, Lower Austria, and Styria. Thelazia callipaeda was also documented in Vienna and P. variegata in Upper Austria and South Tyrol, Italy. All T. callipaeda corresponded to haplotype 1. Twenty-two different haplotypes of P. variegata were identified in the fruit flies. One sequence was distinctly different from those of Phortica variegata and was more closely related to those of Phortica chi and Phortica okadai. Thelazia callipaeda could not be detected in any of the Phortica specimens. [ABSTRACT FROM AUTHOR]

Indonesian leaffish Pristolepis grooti, an endemic species, are distributed in the region of Sumatra, Riau, Bangka Belitung and Kalimantan. However, there has been a decline in the population recently. This research purposed to investigate the mitochondrial DNA cytochrome c oxidase subunit I (COI) gene, the genetic distance, the genetic tree of the leaffish and characterize the chemical physics of water of its habitat in the Kelekar River Muara Enim Regency and the Ogan River, Ogan Ilir Regency. The method used in species authentication was DNA isolation, amplification using PCR (Polymerase Chain Reaction) and sequencing of COI gene. The size of the COI mtDNA gene fragment was 704 bp (PM 1, PM 4, PP 2 and PP 4) and 723 bp (PM 2, PM 3, PP 1 and PP 3). A cryptic diversity of the species P. grooti is found based on the genetic distance value of 4.5-6%, both in the Kelakar and Ogan Rivers. The phylogenetic tree of the leaffish of this study formed 2 separate sub-clusters with a bootstrap value of 50%. The properties of water qualities in the two rivers included temperatures 28.3-31.8°C, pH 5.6-8.3, dissolved oxygen 4.82-10.89 mg L-1, alkalinity 10-28 mg L-1 CaCO3, water transparency 16-45 cm, ammonia 0.47-0.70 mg L-1, water current 0.17-0.30 m s-1 and TDS 7-44 mg L-1. [ABSTRACT FROM AUTHOR]

The nematode Angiostrongylus cantonensis has been reported worldwide. However, some basic questions remain unanswered about A. cantonensis in Ecuador: (1) Was the invasion of A. cantonensis in Ecuador unique, or did it occur in different waves? (2) Was this invasion as recent as historical records suggest? (3) Did this invasion come from other regions of South America or elsewhere? To address these issues, we assessed the genetic diversity of MT-CO1 gene sequences from isolates obtained in 11 of Ecuador's 24 provinces. Our Bayesian inference phylogenetic tree recovered A. cantonensis as a well-supported monophyletic group. All 11 sequences from Ecuador were identical and identified as AC17a. The haplotype AC17a, found in Ecuador and the USA, formed a cluster with AC17b (USA), AC13 (Thailand), and AC12a-b (Cambodia). Notably, all the samples obtained in Ecuadorian provinces' different geographic and climatic regions had no genetic difference. Despite the lack of genetic information on A. cantonensis in Latin America, except in Brazil, our finding differs from previous studies by its absence of gene diversity in Ecuador. We concluded that the invasion of A. cantonensis in Ecuador may have occurred: (1) as a one-time event, (2) recently, and (3) from Asia via the USA. Further research should include samples from countries neighboring Ecuador to delve deeper into this. [ABSTRACT FROM AUTHOR]

Females of the genus Mansonia feed on the blood of humans, livestock, and other vertebrates to develop their eggs. The females' biting behavior may cause severe disturbance to blood hosts, with a negative impact on public health and economics. Certain species have been identified as potential or effective disease vectors. The accurate species identification of field-collected specimens is of paramount importance for the success of monitoring and control strategies. Mansonia (Mansonia) morphological species boundaries are blurred by patterns of intraspecific heteromorphism and interspecific isomorphism. DNA barcodes can help to solve taxonomic controversies, especially if combined with other molecular tools. We used cytochrome c oxidase subunit I (COI) gene 5′ end (DNA barcode) sequences to identify 327 field-collected specimens of Mansonia (Mansonia) spp. The sampling encompassed males and females collected from three Brazilian regions and previously assigned to species based on their morphological characteristics. Eleven GenBank and BOLD sequences were added to the DNA barcode analyses. Initial morphospecies assignments were mostly corroborated by the results of five clustering methods based on Kimura two-parameter distance and maximum likelihood phylogeny. Five to eight molecular operational taxonomic units may represent taxonomically unknown species. The first DNA barcode records for Mansonia fonsecai, Mansonia iguassuensis, and Mansonia pseudotitillans are presented. [ABSTRACT FROM AUTHOR]

Mischief Reef is located in the eastern Nansha Islands of the South China Sea. With increasingly intense anthropogenic disturbance, Pinctada fucata, a previously unrecorded species in the reef, has occurred in the region. In this study, we identified and described the occurrence of P. fucata in Mischief Reef based on morphology and molecular markers. Furthermore, we performed a population genetics analysis of seven P. fucata populations of the South China Sea. All P. fucata populations showed significant high-level genetic diversity, but the differentiation among P. fucata populations was small. There was an FST value close to zero (−0.0083) between the Lingshui and Mischief Reef populations. Our results hint that Lingshui may be one of the potential sources of P. fucata to Mischief. In addition, we discussed the possible cause of the mass occurrence of P. fucata. The present study serves as a warning that anthropogenic disturbances have disrupted the local ecosystem in Mischief Reef. [ABSTRACT FROM AUTHOR]

Basic knowledge on the biology and epidemiology of equine strongylid species still needs to be improved to contribute to the design of better parasite control strategies. Nemabiome metabarcoding is a convenient tool to quantify and identify species in bulk samples that could overcome the hurdle that cyathostomin morphological identification represents. To date, this approach has relied on the internal transcribed spacer 2 (ITS-2) of the ribosomal RNA gene, with a limited investigation of its predictive performance for cyathostomin communities. Using DNA pools of single cyathostomin worms, this study aimed to provide the first elements to compare performances of the ITS-2 and a cytochrome c oxidase subunit I (COI) barcode newly developed in this study. Barcode predictive abilities were compared across various mock community compositions of two, five and 11 individuals from distinct species. The amplification bias of each barcode was estimated. Results were also compared between various types of biological samples, i.e., eggs, infective larvae or adults. Bioinformatic parameters were chosen to yield the closest representation of the cyathostomin community for each barcode, underscoring the need for communities of known composition for metabarcoding purposes. Overall, the proposed COI barcode was suboptimal relative to the ITS-2 rDNA region, because of PCR amplification biases, reduced sensitivity and higher divergence from the expected community composition. Metabarcoding yielded consistent community composition across the three sample types. However, imperfect correlations were found between relative abundances from infective larvae and other life-stages for Cylicostephanus species using the ITS-2 barcode. While the results remain limited by the considered biological material, they suggest that additional improvements are needed for both the ITS-2 and COI barcodes. [ABSTRACT FROM AUTHOR]

In this study, we document the first recorded range extension of the genus Synaphobranchus from the East Mariana Basin in the Western Pacific Ocean. We sequenced the complete mitochondrial (mt) genome of two deep-sea eels (Synaphobranchus brevidorsalis and S. affinis) collected in the East Mariana Basin in the Western Pacific Ocean. The complete mt genomes of S. brevidorsalis and S. affinis were 16,686 bp and 16,677 bp in length, respectively, and consisted of 13 protein-coding genes, 22 tRNA genes, and two rRNA genes. Molecular phylogenetic analysis of the two deep-sea eel species was performed, based on the mt cytochrome c oxidase subunit I (COI) gene using the maximum likelihood method. The molecular phylogenetic tree demonstrated that S. brevidorsalis and S. affinis were congeneric species of S. brevidorsalis and S. affinis reported in previous studies with bootstrap values of 100% and 100%, respectively. This is the first report on the complete mt genomes of S. brevidorsalis and S. affinis collected in the East Mariana Basin in the Western Pacific Ocean. Overall, our study highlights the potential of molecular approaches in identifying species diversity and distribution in the Western Pacific Ocean. [ABSTRACT FROM AUTHOR]

Abstract Pelvic spine polymorphism occurs in several species in the stickleback family (Gasterosteidae). Given the similar phenotypic polymorphisms in multiple stickleback species, we sought to determine the extent of parallelism in the ecological correlates of pelvic spine reduction. Based on a metabarcoding analysis of brook stickleback gut contents in two polymorphic populations, we found that significant diet differences were associated with pelvic spine reduction, but we found no clear or consistent trend supporting a tendency for benthic feeding in pelvic‐reduced brook sticklebacks. These results contrast with those found in threespine sticklebacks where pelvic spine reduction is often associated with a benthic diet. Hence, we found non‐parallel consequences of spine polymorphism across species. Furthermore, a difference in gill raker morphology has been frequently observed between ecomorphs with different diets in many fish species. However, we found no evidence of any difference in gill raker morphology associated with pelvic spine polymorphism in brook sticklebacks.

Pimelodus is the most speciose genus of the family Pimelodidae, and is amply distributed in the Neotropical region. The species-level taxonomy and phylogenetic relationships within this genus are still poorly resolved, however. These taxonomic problems and the general lack of data have generated major uncertainties with regard to the identification of specimens from different localities. In the present study, we applied a single-locus species delimitation approach to identify the MOTUs found within the genus Pimelodus and provide sound evidence for the evaluation of the species richness of this genus in the different river basins of the Neotropical region. The study was based on the analysis of sequences of the mitochondrial COI gene of 13 nominal species, which resulted in the identification of 24 consensus MOTUs. Only six nominal species were recovered as well-defined molecular entities by both the traditional barcoding analysis and the molecular delimitation methods, while the other seven presented cryptic diversity or persistent taxonomic uncertainties. The lineages identified from the Parnaíba ecoregions, Amazonas Estuary and Coastal Drainages may represent a much greater diversity of Pimelodus species than that recognized currently, although a more detailed study of this diversity will be necessary to provide a more definitive classification of the genus.

Severe aphid outbreaks were recorded on sorghum (Sorghum bicolor (L.) Moench) and Johnson grass (Sorghum halepense (L.) Pers.) during the 2020/2021 and 2021/2022 cropping seasons in Argentina. The general aspect of the aphids resembled that of Melanaphis sacchari (Zehntner, 1897), a sugarcane pest occasionally attacking sorghum, reported in the country almost forty years ago. However, its rapid spread and extraordinary abundance on Sorghum spp. raised doubts about the species identity. To address this issue, aphids were sampled from heavily infested sorghum crops and volunteer Johnson grass plants throughout northern and central Argentina and subjected to direct partial sequencing of cytochrome c oxidase subunit I (COI) and elongation factor 1 alpha (EF1-α) genes. Sequence analyses provided evidence that the species causing the outbreaks on Sorghum spp. was, in fact, the closely related Melanaphis sorghi (Theobald, 1904), which had not been previously recorded in Argentina. The lack of genetic variability among the newly obtained sequences suggests the presence of a predominant clonal lineage. A broader population genetics study including further molecular markers, and the evaluation of pest management practices deserve future research. [ABSTRACT FROM AUTHOR]

A subterranean millipede species, Antrokoreana gujoensis Masuda, 2010, is redescribed based on specimens newly collected around its type locality, in the limestone area of Mino Terrane on Honshu Island, Japan. Males of A. gujoensis were originally described as having no penes, but nonetheless, the examined specimens clearly demonstrate that this species possesses the penes behind leg-pair 2. An emended diagnosis of this species and a key to all eight currently recognized species of Antrokoreana is provided. [ABSTRACT FROM AUTHOR]

Rapid and accurate insect identification is crucial for effective plant quarantine procedures. This study introduced a reverse transcription recombinase-assisted amplification-lateral flow strip (RT RAA-LFS), specifically designed for on-site detection of the globally quarantined pest, Tuta absoluta , within a 25-minute timeframe. A pivotal strategy involved integrating three locked nucleic acid (LNA) modifications into a species-specific probe, enabling RT RAA to selectively and efficiently amplify the unique haplotype of T. absoluta 's mitochondrial cytochrome c oxidase subunit I (COI) gene. After determining the amplification product dilution factor and the volumes of primers, magnesium acetate, and nucleic acid lysate in the amplification system, the finalized RT RAA-LFS system is capable of direct visual detection, complemented by handheld fluorescence detection for quantum dots (QDs) fluorescence on the lateral flow strip. This technique successfully identifies various developmental stages of T. absoluta in crude nucleic acid lysates without the need for genomic DNA extraction. Moreover, it detected the target gene in the genomic solution, reaching concentrations as low as 1.6 pg or 51 copies per 50-µL reaction system, aligning with qPCR results. Field validation with 50 insect samples supports the correlation between RT RAA-LFS quantum dots' fluorescence intensity and LFS color intensity, affirming the system's reliability in estimating the target gene copy number (R =0.584, p < 0.01). This validation emphasizes the practical efficacy of the RT RAA-LFS system for T. absoluta detection, highlighting its potential as a rapid on-site tool in agricultural pest management. • Streamlined RT RAA-LFS detection across Tuta absoluta developmental stages. • on-site visual detection in just 25 minutes without nucleic acid extraction. • Optimizing probe with 3 LNAs for efficient species-specific haplotype amplification. • Detection sensitivity at 1.6 pg or 51 copies, consistent with qPCR assay. [ABSTRACT FROM AUTHOR]

Zooplankton are key components of estuarine trophic networks. However, routine monitoring is hindered by the difficulty of morphology-based identification. DNA-based methods allow us to circumvent some of these hurdles, providing precise species identifications regardless of the taxonomic expertise of the investigator or the developmental stage of the specimens. However, the process is dependent on the completeness of the reference libraries. In this study, we sought to evaluate the potential of DNA metabarcoding to assess the seasonal (summer, autumn, and early spring) and spatial dynamics of zooplankton (four locations spanning ca. 6 km) in the Lima estuary (NW Portugal). Two genetic markers were used: the cytochrome c oxidase subunit I and the V4 hypervariable region of the ribosomal 18S rRNA genes. Overall, 327 species were recovered, and both markers displayed minute overlap (7% were detected with both markers). Species richness, composition, and taxonomic distinctness were majorly influenced by the season, with a declining tendency from summer (highest number of exclusive species, n = 74) to spring. Second to season, the taxa composition was influenced by spatial variation where the most downstream site displayed the highest number of exclusive species, n = 53. A total of 16 non-indigenous species were detected using metabarcoding, but only one (Austrominus modestus) has been documented out in the estuary. In conclusion, both the seasonal and spatial gradients influenced the recovered richness, composition, and taxonomic distinctness, confirming the great aptitude of DNA metabarcoding for providing higher density monitoring and shedding new light on the composition and dynamics of complex zooplankton communities.

Pinna nobilis, commonly known as the noble pen shell, is a marine bivalve endemic to the Mediterranean Sea. Unfortunately, due to a multifactorial disease that began affecting its populations in 2016, the species is currently facing the threat of extinction. To gain insights into the evolutionary history of P. nobilis before the mass mortality event (MME), and to obtain a comprehensive understanding of how evolutionary processes led to the adaptation of the species into the Mediterranean Sea, phylogenetic and phylogeographic analyses were carried out. The dataset analysed includes 469 sequences of COI gene fragment both from GenBank and the present study (100). The analysis performed evidenced that P. nobilis diverged about 2.5 mya, after the entrance of its ancestor into the Mediterranean Sea following the Zanclean flood (5.33 mya). Moreover, our results suggest that the starting point of colonisation was the central part of the western Mediterranean basin, with the eastern basin being populated subsequently. From a conservational viewpoint, these results provide important hints for present and future restocking plans, helping to reconstruct the pre-existing genetic variability in sites where the species became extinct.

Charopidae is a highly speciose family of land snails mostly distributed in the main landmasses and archipelagos of the Southern Hemisphere, with a few genera in the Northern Hemisphere. DNA sequencing is making substantial contributions to understanding the phylogeography of Charopidae and the systematic relationships of its subfamilies and related families in the Punctoidea. Here, new data from 28S rRNA are combined with available GenBank sequences to investigate whether Australian Charopidae comprise a single clade, exclusive of species from other regions. Phylogenetic analyses show that this is not the case, with most species from Australia and other regions included in a bootstrap-supported clade that excludes some Australian species. Analyses of concatenated 28S rRNA data and available cytochrome c oxidase subunit I (COI) also contradicted monophyly of Australian species, as did analyses of inferred COI protein sequences. In DNA analyses, the genus Hirasea and Australian taxa with an incomplete secondary ureter were basal within Charopidae. We also examined whether known doubts about the taxonomic status of supra-generic clades in Punctoidea could be resolved by considering autapomorphic COI amino acid changes. Such variation could not generally be applied as an objective criterion for taxonomic categorization but did identify some clades for taxonomic investigation.

Introduction: Calliphoridae plays a key role in forensic entomology research, which is one of the first insects to decompose animal carcasses.The mitochondrial cytochrome c oxidase subunit I and the ribosomal internal transcribed spacer 2 (ITS2) are among the most widely used molecular markers for insect taxonomic characterization. Aim: The aim of the study was to test the suitability of two genetic markers based on conducting the molecular identification of six necrophagous Calliphorid flies. Materials and Methods: Fourteen Calliphoridae flies were collected and classified with traditional morphological characteristics. The DNA of flies was extracted and the fragments of COI and ITS2 were amplified and sequenced. All the sequences were aligned and analyzed by MEGA 7 software for NCBI BLAST, nucleotide composition, intra- and inter-specific divergence calculation, and phylogenetic tree inference successively. Results: The results indicated that COI and ITS2 genes were robust in the identification of Calliphoridae at the species level and ITS2 gene sequence possessed a strong resolution power as it showed higher variation values between Lucilia sericata and Lucilia cuprina, Calliphora vomitoria and Triceratopyga calliphoroides, C.vomitoria and Aldrichina grahami, but inferior to COI for T. calliphoroides and A. grahami. Conclusions: Our results showed that combination of COI + ITS2 genes yields more accurate identification and diagnoses and better agreement with morphological data than the mitochondrial barcodes alone. As a supplementary method for morphological identification, we advocated for the combination of nuclear and mitochondrial gene approaches to address the taxonomy and phylogeny of forensic relevant flies, especially of closely related species and populations.

The two congeneric hyperiids Themisto libellula and T. abyssorum provide an important trophic link between lower and higher trophic levels in the rapidly changing Arctic marine ecosystem. These amphipods are characterized by distinct hydrographic affinities and are hence anticipated to be impacted differently by environmental changes, with major consequences for the Arctic food web. In this study, we applied DNA metabarcoding to the stomach contents of these Themisto species, to comprehensively reveal their prey spectra at an unprecedented-high-taxonomic-resolution and assess the regional variation in their diet across the Fram Strait. Both species feed on a wide variety of prey but their diet strongly differed in the investigated summer season, showing overlap for only a few prey taxa, such as calanoid copepods. The spatially structured prey field of T. libellula clearly differentiated it from T. abyssorum, of which the diet was mainly dominated by chaetognaths. Our approach also allowed the detection of previously overlooked prey in the diet of T. libellula, such as fish species and gelatinous zooplankton. We discuss the reasons for the differences in prey spectra and which consequences these may have in the light of ongoing environmental changes. [ABSTRACT FROM AUTHOR]

To date, only two Cherax species have been reported to occur in Italy, i.e., C. destructor Clark, 1936 and C. quadricarinatus (von Martens, 1868), both in the wild and in aquaculture farms. Therefore, we aimed to update their current status and distribution in Italian mainland and Sicily. In addition, we investigated the origin of their known populations, and their possible routes of invasion. In order to genetically characterize the Cherax populations occurring in Italian inland waters and aquaculture facilities, the barcode region of the mtDNA gene cytochrome oxidase subunit I was sequenced in the available specimens originating from an aquaculture facility and a museum collection. The sampling campaigns conducted in 2021 did not lead to the collection of any Cherax individuals in the sites where the species was formerly reported to occur. The recent failures to detect Cherax spp. from Italian inland waters might be due to the inability of the species to cope with the Italian climate, which is likely for C. quadricarinatus but less so for C. destructor, to the possible impact of the alien red swamp crayfish Procambarus clarkii (as well as its indirect impact i.e., the spread of the crayfish plague for which yabby is susceptible), which occurs in both the sites where C. destructor was reported in Italy, or to the burrowing habits of the species, which might lead to overlook their presence when present at low densities. In the light of the well-known impact of invasive crayfish and considering the scant knowledge about the current distribution and status of Cherax species in Italy, a regular monitoring of their possible presence is recommended. [ABSTRACT FROM AUTHOR]

Dryadaulaepischista (Meyrick, 1936), initially described from a single male specimen in Japan, is herein redescribed based on newly collected specimens from the type locality. Furthermore, we describe Dryadaulaorientalis Park & Yagi, sp. nov. , a new species from Japan that closely resembles D.epischista . The adults and genitalia of the two species are illustrated. The genitalia of D.epischista from a specimen collected at the type locality are shown for the first time. DNA barcodes of the two Dryadaula species and the genetic distances of barcode regions among them and other congeners are provided., Competing Interests: The authors have declared that no competing interests exist., (Jinhyeong Park, Sadahisa Yagi, Shigeki Kobayashi, Toshiya Hirowatari.)

Abstract DNA metabarcoding is an increasingly popular technique to investigate biodiversity; however, many methodological unknowns remain, especially concerning the biases resulting from marker choice. Regions of the cytochrome c oxidase subunit I (COI) and 18S rDNA (18S) genes are commonly employed “universal” markers for eukaryotes, but the extent of taxonomic biases introduced by these markers and how such biases may impact metabarcoding performance is not well quantified. Here, focusing on macroeukaryotes, we use standardized sampling from autonomous reef monitoring structures (ARMS) deployed in the world's most biodiverse marine ecosystem, the Coral Triangle, to compare the performance of COI and 18S markers. We then compared metabarcoding data to image‐based annotations of ARMS plates. Although both markers provided similar estimates of taxonomic richness and total sequence reads, marker choice skewed estimates of eukaryotic diversity. The COI marker recovered relative abundances of the dominant sessile phyla consistent with image annotations. Both COI and the image annotations provided higher relative abundance estimates of Bryozoa and Porifera and lower estimates of Chordata as compared to 18S, but 18S recovered 25% more phyla than COI. Thus, while COI more reliably reflects the occurrence of dominant sessile phyla, 18S provides a more holistic representation of overall taxonomic diversity. Ideal marker choice is, therefore, contingent on study system and research question, especially in relation to desired taxonomic resolution, and a multimarker approach provides the greatest application across a broad range of research objectives. As metabarcoding becomes an essential tool to monitor biodiversity in our changing world, it is critical to evaluate biases associated with marker choice.

ObjectiveTo identify the common sarcosaprophagous flies in the Yangtze River Delta based on mitochondrial cytochrome c oxidase subunit Ⅰ（COⅠ） gene sequence and verify the reliability of this method.MethodsSeven common genetically stable sarcosaprophagous flies in three families and six genera were collected from large domestic pig carcasses placed in the field and cultured in the laboratory for many generations. The whole genome DNA was extracted and the COⅠ gene fragment was amplified. The forward and reverse sequencing was followed by splicing. The base composition of the amplified fragment and the rate of interspecific evolutionary divergence were analyzed by software such as Mega 7.0.26. The phylogenetic tree of COⅠ gene sequence of common necrophagous flies in the Yangtze River Delta was established by neighbor joining （NJ） method and unweighted pair-group method with arithmetic means （UPGMA） method.ResultsThe average base composition of different flies was A（30.14%）, T（38.23%）, C（15.98%）, G（15.65%）. The rate of interspecific evolutionary divergence ranged from 2.2% to 15.3%, the lowest rate was between Chrysomya megacephala and Chrysomya pinguis, the highest rate was between Muscina stabulans and Boettcherisca peregrina.ConclusionCOⅠ gene can be used to identify the common necrophagous flies in the Yangtze River Delta.

Hybridization is an important evolutionary process that can fuel diversification via formation of hybrid species or can lead to fusion of previously separated lineages by forming highly diverse species complexes. We provide here the first molecular evidence of hybridization in wild populations of ciliates, a highly diverse group of free-living and symbiotic eukaryotic microbes. The impact of hybridization was studied on the model of Plagiotoma, an obligate endosymbiont of the digestive tube of earthworms, using split decomposition analyses and species networks, 2D modeling of the nuclear rRNA molecules and compensatory base change analyses as well as multidimensional morphometrics. Gene flow slowed down and eventually hampered the diversification of Lumbricus-dwelling plagiotomids, which collapsed into a single highly variable biological entity, the P. lumbrici complex. Disruption of the species boundaries was suggested also by the continuum of morphological variability in the phenotypic space. On the other hand, hybridization conspicuously increased diversity in the nuclear rDNA cistron and somewhat weakened the host structural specificity of the P. lumbrici complex, whose members colonize a variety of phylogenetically closely related anecic and epigeic earthworms. By contrast, another recorded species, P. aporrectodeae sp. n., showed no signs of introgression, no variability in the rDNA cistron, and very high host specificity. These contrasting eco-evolutionary patterns indicate that hybridization might decrease the alpha-diversity by dissolving species boundaries, weaken the structural host specificity by broadening ecological amplitudes, and increase the nuclear rDNA variability by overcoming concerted evolution within the P. lumbrici species complex.

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