Marine Invertebrates Research Articles

Review of two marine vertebrate assemblages from the Arauco Basin (central Chile) reveals diversity changes throughout the Maastrichtian

Upper Cretaceous vertebrate records from Chile are mostly known by historical mentions with unknown repositories and uncertain stratigraphic provenance. This contribution reviews and complements two marine vertebrate assemblages from the Upper Cretaceous of central Chile, which were part of the ancient Arauco Basin. The oldest assemblage (lower Maastrichtian) comprises abundant condrichthyans referred to Carcharias gracilis, Odontaspis cf. winkleri, Scapanorhynchus sp., Centrophoroides appendiculatus, Squatina sp., Cretorectolobus sp., Orectolobidae indet., Paraorthacodus sp., Ischyrhiza chilensis and Biropristis landbecki, which adds to the previously reported occurrences of Echinorhinus sp. and Myledaphus araucanus. In addition, chimeroids referred to as Edaphodon kawai and remains of a leatherback turtle referable to Mesodermochelys sp. are here described, the latter being its first occurrence outside Japan. The younger assemblage (upper Maastrichtian) includes similar chondrichthyans and a higher diversity of marine reptiles, including plesiosaurians (Aristonectes sp., Aristonectinae indet., and Elasmosauridae indet.), sea turtles (Pancheloniidae indet.) and diverse mosasaurs (Halisaurus sp., Tylosaurinae indet., and the first local occurrence of Plioplatecarpinae indet.). Throughout the Maastrichtian, the local marine vertebrates likely suffered a declination in abundance but a rise in diversity, with evidence of a marked alteration in middle levels of the trophic web during the upper Maastrichtian. This fauna shows a main influence from the northern hemisphere (especially from the Western Interior Sea), acquiring a more marked Weddellian influence during the end of the Maastrichtian. The studied material allows a better understanding of the Upper Cretaceous vertebrate marine fauna in lower latitudes of the southeastern Pacific.

Calcium-rich seawater affects the mechanical properties of echinoderm skeleton

Shifts in the magnesium to calcium ratio of seawater in the geological history are thought to have profoundly affected biomineralization of marine invertebrates, including some echinoderms, which changed their skeletal mineralogy from high-magnesium to low-magnesium calcite and vice versa. Here we report on experiments that aimed to investigate the effect of ambient seawater magnesium to calcium ratio on magnesium to calcium ratio and nanomechanical properties in the spines of two echinoid species (Arbacia lixula and Paracentrotus lividus). We found that echinoids cultured in seawater with a low magnesium to calcium ratio produced a skeleton with lower both magnesium content and nanohardness than those of the control specimens incubated under normal (high) magnesium to calcium ratio conditions. These results may suggest that at certain times in the geological past (during the so-called calcite seas) sea urchins with decreased skeletal magnesium contents were more susceptible to damage due to physical disturbances, predation and post-mortem taphonomic processes. Increased skeletal hardness of echinoids from the so-called aragonite seas is expected to enhance their taphonomic potential, thus, to some extent, mitigates the preservation bias related to increased solubility of high-magnesium calcite.

Slow recovery in trophic structure of restored wetlands in Northeast China

Restoration measures have been widely implemented in wetland ecosystems globally to bend the curve of biodiversity loss and restore associated ecological functions. However, assessments of the effectiveness of wetland restoration have predominantly focused on the recovery of taxonomic composition, while few studies have assessed the effectiveness of these efforts from a food web perspective. Here, we incorporated stable isotope approach to investigate trophic structure in natural and restored wetlands in Northeast China. The investigated consumers, including zooplankton, macroinvertebrates, and fish, exhibited lower δ15N and higher δ13C values in restored wetlands than in natural wetlands. Natural wetlands exhibited higher trophic positions and a wider range of trophic levels compared to restored wetlands. Primary consumers in natural wetlands relied more on particulate organic matter (POM, 42.9 % ± 24.1 %), while those in restored wetlands were more dependent on substrate organic matter (SOM, 42.3 % ± 23.9 %). Compared to natural wetlands, isotopic richness was significantly lower in restored wetlands, with smaller isotopic variation (SEAs) in basal resources, aquatic invertebrates, and fish. Our findings reveal that the recovery of trophic structures in restored wetlands lags behind that of taxonomic composition. Future restoration efforts should prioritize enhancing habitat heterogeneity and resource availability to support a diverse range of trophic levels. Monitoring trophic dynamics is essential for assessing the progress of wetland restoration and should be integrated into monitoring schemes.

Tubeworms as bioindicators of paleoenvironmental conditions

Tubeworms comprise an informal designation for a biological group of marine invertebrates, associated with producing rigid tubes around their body with varied compositions and functions. The main representatives of this group currently comprise organisms from different phyla and have a wide stratigraphic distribution, taxonomic and biostratigraphic discussions have always been related to the group, not presenting themselves in a distinct way for the Devonian of the Paraná Basin. In the present work, we recognize the taxa Annulitubus mutveii, Cornulites auxilium, and a polychaete agglutinating of the order Terebellida occurring in all lithofacies of the lower shoreface to the offshore from Late Praguian to Givetian. In addition, we discuss the potential of Annulitubus mutveii as a paleoenvironmental bioindicator associated with ecologically stressed environments caused by post-biotic crises caused by extinction events.

Foraging Habitat Availability and the Non-Fish Diet Composition of the Grey Heron (Ardea cinerea) at Two Spatial Scales.

Habitat structure on foraging ground is one of the crucial factors determining diet diversity in colonially breeding avian predators. Quantifying the habitat and diet composition at different spatial scales (regional and local inter-colonial) can help provide understanding of the drivers of diet composition. In this study, we examined the composition of the non-fish diet of an opportunistic predator, the Grey Heron (Ardea cinerea), based on pellets. We compared pellets from 21 colonies in two different regions of Poland that differing in composition of foraging habitats. Multivariate statistical techniques were used to analyze the relationship between diet and habitat compositions in a 20 km radius around these colonies. Significant inter-regional and inter-colony differences in Grey Heron diet and habitat composition were detected in foraging areas. However, some prey were present in the diet only from one of the regions. Around the heronries with a predominance of farmland habitats, the European water vole (Arvicola amphibius) and terrestrial invertebrates were present in relatively low frequencies, and the striped field mouse (Apodemus agrarius) was present in high frequencies. Voles (Microtus sp.) were more frequent in colonies with prevalence of non-irrigated arable land around the colony, in contrast to the bank vole (Myodes glareolus), which was less common in heronries with a higher contribution of this habitat type. Remains of aquatic invertebrates were less abundant in colonies surrounded by extensive forests. The results of our research indicate the opportunistic character of the non-fish part of the diet of the Grey Heron adapted to the local foraging habitat and prey availability.

Temperature influences immune cell development and body length in purple sea urchin larvae

Anthropogenic climate change has increased the frequency and intensity of marine heatwaves that may broadly impact the health of marine invertebrates. Rising ocean temperatures lead to increases in disease prevalence in marine organisms; it is therefore critical to understand how marine heatwaves impact immune system development. The purple sea urchin (Strongylocentrotus purpuratus) is an ecologically important, broadcast-spawning, omnivore that primarily inhabits kelp forests in the northeastern Pacific Ocean. The S. purpuratus life cycle includes a relatively long-lived (∼2 months) planktotrophic larval stage. Larvae have a well-characterized cellular immune system that is mediated, in part, by a subset of mesenchymal cells known as pigment cells. To assess the role of environmental temperature on the development of larval immune cells, embryos were generated from adult sea urchins conditioned at 14 °C. Embryos were then cultured in either ambient (14 °C) or elevated (18 °C) seawater. Results indicate that larvae raised in an elevated temperature were slightly larger and had more pigment cells than those raised at ambient temperature. Further, the larval phenotypes varied significantly among genetic crosses, which highlights the importance of genotype in structuring how the immune system develops in the context of the environment. Overall, these results indicate that larvae are phenotypically plastic in modulating their immune cells and body length in response to adverse developmental conditions.

Optical Temperature Sensing and Bioimaging of Aquatic Invertebrates With Nd3+‐ Sensitized Core@Shell Nanoparticles

Optical Temperature Sensing and Bioimaging of Aquatic Invertebrates With Nd³⁺‐ Sensitized Core@Shell Nanoparticles

Characterization of water microbiota and their relationship with resident oysters during an oyster mortality event.

Microorganisms are vital for the health of marine invertebrates, and their assembly is driven by both deterministic and stochastic factors that regulate residents (innate to the host) and transients (from ambient water). However, the role of water microbiota and the significance of deterministic and stochastic processes in aquatic hosts facing mortality threats are largely unknown. This study examines the shifts in water microbiota during an oyster mortality event using amplicon sequencing and compared with those of resident oysters to disentangle the balance of the deterministic and stochastic factors involved. Water temperature and dissolved oxygen significantly shape the microbial community with a distinct monthly pattern, and Cyanobacteria blooms might exacerbate oyster mortality. The comparative analysis of microbial communities in oysters and water revealed that ≤ 21% of the genera were shared between oysters and water, implying that water microbiota cannot easily transfer into oysters. Furthermore, these shared genera had different functions, with oysters more involved in promoting host digestion and nutrient acquisition and water bacteria enriched more in functions promoting their own growth and survival. These findings illustrate that oysters may possess specific selection or barrier mechanisms that permit a small percentage of transients, controlled by stochastic factors and having a minimal effect on oyster mortality, to enter, whereas the majority of oyster microbiota are residents governed by deterministic factors. Consequently, oysters exhibit some plasticity in their symbiotic microbiota, enabling them to maintain microbial homeostasis and adapt to complex microbial surroundings. This may be a shared mechanism among marine invertebrates for survival in complex marine environments.IMPORTANCEPacific oysters are widely cultured and play vital ecological roles. However, the summer mortality hinders sustainable oyster farming. Untangling causative mechanisms of oyster mortality is a complex task due to the intricate "interactome" involving environmental factors, hosts, and pathogens. Interactions between hosts and microorganisms offer an ideal avenue for investigating the truth. We systematically investigated the microbial community in water and resident oysters during a summer mortality event and proposed that the assembly of oyster microbiota is primarily governed by deterministic processes independent of mortality. Pathogens mainly originate from resident members of the oyster microbiota, with a limited influence from the microbial community in the water. Additionally, environmental degraders, such as Cyanobacteria blooms, cannot be overlooked as a contributing factor of oyster mortality. This study evaluated the weight of deterministic and stochastic factors in microbial assembly during an oyster mortality event and greatly broadened our understanding of the "interactome" through the interaction between oysters and water in microbiota.

Myocyte enhancer factor 2 upregulates expression of myostatin promoter in Yesso scallop, Patinopecten yessoensis

Myostatin (MSTN) plays an important role in muscle development in animals, especially for mammals and fishes. However, little information has been reported on the regulation of MSTN in marine invertebrates, such as bivalves. In the present study, we cloned the MSTN promoter sequence of Yesso scallop Patinopecten yessoensis, identifying 4 transcription start sites, eleven TATA boxes and one E-box. Additionally, transcription factor binding sites, including myocyte enhancer factor 2 (MEF2) and POU homeodomain protein, were identified. The interaction between the MSTN promoter and MEF2 was analyzed to reveal the transcriptional activity of different fragment sizes of promoters through the dual-luciferase reporter assays. The highest transcriptional activity was found in recombinant plasmids with the most MEF2 binding sites, indicating that this transcription factor upregulates MSTN in Yesso scallop. This study provides new insight into the regulation of muscle growth and development in this species.

Ecology of Intertidal Rocky Shores Related to Examples of Coastal Geology across Phanerozoic Time

The boundary between land and sea is among the most extensive and extreme ecological barriers on planet Earth. Intertidal organisms living at this junction are subject to potentially lethal conditions related to desiccation, temperature fluctuations, and wave shock. Although difficult to quantify at a global level, rocky coasts account for between a third and three-quarters of all shorelines today. Resistant to coastal erosion, rocky coasts also exhibit adaptations by marine invertebrates and marine algae within intertidal zones that are geographically widespread. Biotic composition is variable depending on the tidal range at any given place and on climatic differences between tropical and more temperate latitudes. Contemporary granite shores having low recession rates with well-established biotic zones are reviewed in three regions: Mount Desert Island in Maine, USA (44° N latitude), Mahé and Praslin islands in Seychelles (4°30′ S Latitude), and Lizard Island off the coast of Queensland, Australia (14°40′ S Latitude). The zonal composition and overall biotic diversity at these locations are compared, as well as the fossilization potential of key biota. Over the Phanerozoic time period during the last 538 million years, the ratio between land and ocean varied as a function of plate tectonics depending on the maximum dispersal or coalescence of continents and on relative changes in global sea level. Fossil biotas from a dozen paleoislands with dominant rocky shores that range from 485 million to 125,000 years in age are reviewed to show diversification through time, taking into account episodic mass extinctions. Relationships between storm tracks and volcanic eruptions that factor into physical disruptions are also considered as means of rapid burial and preservation. Themes related to rocky-shore ecology and paleoecology benefit from the cross-fertilization of ideas by marine biologists and marine paleontologists. It is hoped this review may attract a cohort of new students to these allied fields of study, especially in Asia, Africa, and South America.

Population genetics of two sibling species of fiddler crabs (Decapoda, Ocypodidae): Brazilian homogeneity and West Atlantic heterogeneity

Abstract Fiddler crabs are a widespread group of marine invertebrates. Their wide distribution is due to the great capacity of their larvae to disperse among estuarine and shelf waters. Therefore, it is expected that all populations along their distribution show molecular similarities due to extensive gene flow. Thus, we assessed the intra- and interspecific genetic divergence of two sibling species of fiddler crabs, Minuca panema and Minuca mordax, based on COI mtDNA sequences. The molecular data resulted in a highly mixed haplotype network for M. panema, while M. mordax sequences were clustered in two groups, one with Brazilian and other with Costa Rican haplotypes. The highest variance in M. panema was found within populations, while in M. mordax it was between groups and populations. The analyses of nucleotide divergence showed small genetic variance within populations of both species and between Brazilian populations, and higher variance between Brazilian and Costa Rican populations of M. mordax. The highest genetic divergence was found between the two species. Thereby, our results show an intense gene flow along the Brazilian coast, while there is a clear divergence among Brazil and Costa Rica, which may be a result of the great distance between these populations.

Effects of Land Use on the Community Structure of Aquatic Invertebrate in Subtropical Streams

Streams constitute the water supply of the watersheds and provide the transfer of energy along the course of water, ensuring water biodiversity. Due to the different uses of the earth, the benthic community is being suppressed. The objective of this study is to investigate the effects of changes in land use on the abundance, richness, and diversity of benthic macroinvertebrates in subtropical streams, considering the physical and chemical variables of streams seasonally. In total, 144 samples were collected between February 2022 and November 2023 at 12 sites distributed in a watershed in southern Brazil, four sites for each land use. Herein, 83.520 individuals were identified, comprising 67 taxa. There was a decrease in richness along the forested–rural–urban gradient and a greater abundance in the urban environment, without the significant influence of seasonality. We conclude that the diversity of benthic macroinvertebrate in the streams is influenced not only by environmental variables but also by the spatial distribution between streams. The proximity of the streams generates a similarity in dominance. This study contributed to understanding the relationships of the environment with the benthic community and considering the spatial distribution among the streams sampled in future investigations.

Hemal sinus basal laminae contact sites: a possible route between gonadal lumen and myoepithelial cells in the gonad of the sea star Patiria pectinifera.

Sea stars are a group of marine invertebrates suitable for studying the hormonal regulation of reproduction and spawning. In spite of substantial progress in understanding how various substances such as 1-methyladenine act in their gonads, there are still many gaps concerning the fine details of their action. One such gap is how the gonadal wall contraction is induced. Recent literature data suggest that, upon 1-methyladenine stimulation, some cells within the gonadal lumen produce non-neuronal acetylcholine that, upon contact with the gonadal wall, induces contraction of myoepithelial cells. Our ultrastructural study of the gonads in the sea star Patiria pectinifera has shown, for the first time, that there are sites where the basal laminae bordering the hemal sinus directly contact one another and appear at this contact site as a single entity. These contact sites are often associated with hemidesmosome-like junctions that anchor male accessory cells or female follicle cells on one side of the site and myoepithelial cells on the opposite. We suggest that contraction-inducing substance is secreted from an accessory or follicle cell, passes through a basal lamina contact site, and on the opposite side of the contact site acts on a myoepithelial cell to induce its contraction.

A quantitative assessment of continuous versus structured methods for the detection of marine mammals and seabirds via opportunistic shipboard surveys

Marine monitoring efforts are increasingly supported by opportunistic shipboard surveys. However, opportunistic survey methods often require adaptation to suit the vessel and the operations being conducted onboard. Whilst best-practice techniques for surveying marine wildlife on vessels of opportunity are yet to be established, testing and development of alternative methods can provide means for capturing ecological information in otherwise under-surveyed areas. Explicitly, survey methods can be improved while baseline ecological data for new regions are gathered simultaneously. Herein, we tested different survey approaches on a vessel of opportunity in a remote offshore area where little is known about the community composition of top-order marine vertebrate predators: western and south-western Tasmania, Australia. We found that continuous surveys provide greater species counts than structured “snapshot” surveys over the course of a voyage, but that structured surveys can be more practical when managing factors such as observer fatigue. Moreover, we provide a baseline dataset on the marine vertebrate community encountered in western and south-western Tasmania. This information will be critically important for industry and conservation management objectives, and is key to our understanding of the offshore ecosystem around Tasmania.

Dual impacts of hydrology and damming on eutrophication: Comparison of two Ramsar wetlands in the middle Yangtze floodplain

Dam construction for social-economic benefits has raised substantial biological and ecological concerns. However, contrasting findings have been reported regarding the role of hydrological modification in floodplain lake ecosystems. Here, we evaluated the influence of hydrology and dam construction on the eutrophication of floodplain lakes over the last 200 years by studying Zn/Al (an indicator of industrial activities), nutrient influxes (total phosphorus, TP), cyanobacteria production (canthaxanthin) and aquatic invertebrate dynamics (chironomids) in 210Pb dated sediment cores from two Ramsar Wetlands of International Importance in the middle Yangtze floodplain, one dammed (Wanghu) and the other freely connected (Poyang) with the Yangtze River. The results show that Wanghu Lake transitioned to a macrophyte-dominated clear water state (as indicated by the increases in pigment derived ultraviolet radiation index and dominance of macrophyte-related chironomid taxa) after local dam construction when anthropogenic nutrient loadings were relatively low. With increases in nutrient loadings, phytoplankton increased in both lakes, but water clarity declined and macrophyte-related chironomids decreased only in the lake which was locally dammed. Our study reveals that local damming facilitates the response of floodplain lake ecosystems to eutrophication and decouples it from the effects of hydrological variability. This study highlights the potential influence of hydrology and damming on the eutrophication of floodplain lakes by influencing water clarity and macrophyte coverage, implying that evaluating the role of local dam construction on ecosystem states should be based on knowledge of nutrient conditions in floodplain lakes.

A coordinated attack by a bacterial secretion system and a small molecule drives prey specificity

Vibrio species are recognized for their role in food- and water-borne diseases in humans, fish, and aquatic invertebrates. We screened bacterial strains isolated from raw food shrimp for those that are bactericidal to Vibrio strains. Here we identify and characterize Aeromonas dhakensis strain A603 which shows robust bactericidal activity specifically towards Vibrio and related taxa but less potency toward other Gram-negative species. Using the A603 genome and genetic analysis, we show that two antibacterial mechanisms account for its vibriocidal activity -- a highly potent Type Six Secretion System (T6SS) and biosynthesis of a vibriocidal phenazine-like small molecule, named here as Ad-Phen. Further analysis indicates coregulation between Ad-Phen and a pore-forming T6SS effector TseC, which potentiates V. cholerae to killing by Ad-Phen.

Aquatic invertebrate diversity in isolated and permanent pools along an urban ephemeral stream in Central Texas

Aquatic invertebrate diversity in isolated and permanent pools along an urban ephemeral stream in Central Texas

Holocene ecosystem and temperature development inferred from invertebrate remains in Zminje Jezero (Dinaric Alps, Montenegro)

AbstractLake Zminje Jezero (1535 m a.s.l.) in Montenegro was studied for chironomid and other aquatic invertebrate remains in a sediment sequence dating back to 12,000 calibrated 14C years before present (cal yr BP), providing, to our knowledge, the first lake-sediment record studied for chironomids and other associated chitinous aquatic invertebrate remains in the Dinaric Alps. Changes in chironomid and other invertebrate remains along the record make it possible to constrain changes of relevant environmental variables for aquatic invertebrates in the lake including temperature, oxygen availability, trophic status and water depth. The results suggest moderate changes in trophic conditions with chironomid assemblages indicating meso- to eutrophic conditions during the analysed interval. Invertebrate assemblages were typical for lakes with relatively high oxygen availability with a minor trend during the middle to late Holocene to conditions observed in lakes that are more hypoxic. A change in temperature is suggested in the earlier part of the record when the share of chironomid taxa adapted to warmer conditions increased, at the Younger Dryas to Holocene transition, whereas later no particularly pronounced shifts in temperature-sensitive taxa were observed. July air temperatures were estimated based on chironomid assemblages using a transfer function based on calibration data consisting of 117 lakes sampled in the Swiss Alps and northern Switzerland. The results suggest that temperatures rapidly increased by 5 °C at the onset of the Holocene (11,500 cal yr BP) leading to a relatively warm early to mid-Holocene and thereafter slightly decreased during the late Holocene. Reconstructed temperatures are discussed together with a previously published pollen record for our study site and are broadly consistent with other chironomid-based temperature reconstructions from Central, Eastern and Southern Europe. From ca. 3000 cal yr BP sedimentation rates increased and from ca. 500 cal yr BP onwards pollen data suggest that the vegetation and lake catchment were affected by human activities, possibly influencing chironomid and other invertebrate assemblages in the lake and thereby also reconstructed temperatures. Overall, our results show that combining analyses of chironomid and other invertebrate assemblages can provide valuable insights into long-term environmental changes and can provide temperature reconstructions for small mountain lakes in the Balkans, such as Zminje Jezero.

Exploring Biodiversity and Food Webs in Sulfur Cave in the Vromoner Canyon on the Greek–Albanian Border

Sulfidic caves support diverse and abundant subterranean communities, including numerous endemic species and complex food webs, though the full extent of species diversity and resource utilization in these ecosystems remains largely unexplored. This paper presents the results of biological surveys conducted from 2023 to 2024 in Sulfur Cave, located in the Vromoner Canyon on the Greek–Albanian border, focusing on microbial, vertebrate, and invertebrate communities and investigating the structure of the subterranean food web. The microbial communities from the different biofilms are dominated by chemosynthetic sulfur-oxidizing microorganisms, specifically filamentous bacteria such as Thiotrix and Beggiatoa. Two species of fish, an eel (Anguilla sp.) and a Cyprinid (Alburnoides sp.), and six bat species from three families (Rhinolophidae, Miniopteridae, and Vespertilionidae) were documented. The invertebrate fauna includes five aquatic species, 25 terrestrial species, and four amphibiotic species. Among these, eight species are endemic, and seven species exhibit troglomorphic traits. Stable isotope analysis showed light carbon and nitrogen values for the terrestrial and aquatic invertebrates, suggesting that subterranean communities rely on food produced in situ by chemoautotrophic microorganisms. Our results identified cave areas of significant biological relevance and provided reference data to inform conservation actions aimed at preserving the biodiversity of this sulfidic cave.

Cost-efficient PCR based DNA barcoding of marine invertebrate specimens with NovaSeq amplicon sequencing.

The marine environment harbors high biodiversity; however, it is poorly understood. Nucleotide sequence data of all marine organisms should be accumulated before natural and/or anthropogenic environmental changes jeopardize the marine environment. In this study, we report a cost-effective and easy DNA barcoding method. This method can be readily adopted without using library preparation kits. It includes multiplex PCR of short targets, indexing PCR, and outsourcing to a sequencing service using the NovaSeq system. We targeted four mitochondrial genes [cytochrome c oxidase subunit I (COI), COIII, 16S rRNA (16S), and 12S rRNA (12S)] and three nuclear genes [18S rRNA (18S), 28S rRNA (28S), internal transcribed spacer 2 (ITS2)] in 95 marine invertebrate specimens, which were primarily annelids. The primers, including adapters and indices for NovaSeq sequencing, were newly designed. Two PCR runs were conducted. The 1st PCR amplified specific loci with universal primers and the 2nd added sequencing adapters and indices to the 1st PCR products. The gene sequences obtained from the FASTQ files were subjected to BLAST search and phylogenetic analyses. One run using 95 specimens yielded sequences averaging 2816bp per specimen for a total length of six loci. Nuclear genes were more successfully assembled compared with mitochondrial genes. A weak but significantly negative correlation was observed between the average length of each locus and success rate of the assembly. Some of the sequences were almost identical to the sequences obtained from specimens collected far from Japan, indicating the presence of potentially invasive species identified for the first time. We obtained gene sequences efficiently using next-generation sequencing rather than Sanger sequencing. Although this method requires further optimization to increase the success rate for some loci, it is used as a first step to select specimens for further analyses by determining the specific loci of the targets.