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Human interaction with marine creatures holds both positive and negative dimensions. Coastal communities benefit from marine environments, relying on them for sustenance and livelihoods. Fishing activities support economies, and marine biodiversity contributes to overall ecosystem health. However, challenges like overfishing, habitat destruction, and pollution pose threats to both marine life and human communities. Recently, there has been widespread concern regarding the potential increase in jellyfish populations across global marine ecosystems, attributed mainly to environmental factors such as climate drivers and anthropogenic forces, or their complex interactions. Encounters with hazardous marine species, such as box jellyfish, exemplify the dangers associated with coastal activities. Unintended interactions may lead to stings, injuries, and even fatalities, necessitating proactive measures and advanced technologies. This study addresses the inadequacies of existing measures in preventing box jellyfish incidents by introducing environmental DNA (eDNA) assays for detecting the deadly Chiropsoides buitendijki and focuses on developing qPCR and dPCR-based eDNA assays. Emphasising prevention over treatment, the study establishes a proactive system to assess C. buitendijki distribution across 63 tourist beaches in the Gulf of Thailand. Comparative analysis highlights the superior performance of dPCR over qPCR and traditional surveys. The dPCR experiment yielded positive results for all eDNA samples collected at sites where C. buitendijki had previously been identified. Remarkably, the eDNA testing also detected positive results in 16 additional sample locations where no physical specimens were collected, despite reported jellyfish stings at some of these sites. These findings underscore the precision and efficacy of the proposed eDNA detection technology in the early detection and assessment of box jellyfish distribution. This advancement therefore not only aids ecological research but also serves as a valuable tool for safeguarding public health, providing an early warning system for potential jellyfish encounters. Balancing positive human-marine interactions with effective risk mitigation strategies is crucial for sustainable coexistence, the preservation of marine ecosystems, and human well-being., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Morphologically, species of fanged frogs (Limnonectes) are exceedingly similar, making it difficult to distinguish them within the complex. In Thailand, it has been difficult to distinguish between the sympatric species L. bannaensis and L. taylori, particularly among tadpoles, adolescents, and adult females. A precise identification contributes to a greater understanding of biodiversity, particularly for assessing distributions and population dynamics. Therefore, a novel approach is required. The objective of this study was to develop a high resolution melting analysis (HRM) for the rapid and accurate identification of six species of Limnonectes of the L. kuhlii complex found in Thailand, particularly the two sympatric fanged frogs. Here, HRM assays using 16S rRNA mitochondrial primers were designed and developed. There was as much as a 25.3% variation in the nucleotide sequence of the fragment amplified by HRM16S primers among the six species of Limnonectes. Prior to conducting an in vitro HRM, the DNA sequences were used in a simulation HRM, uMELT Quartz, to predict the melting curve for each species of Limnonectes. There were discrepancies between the predicted melting curves of each species generated by the programme. Consequently, in vitro HRM tests were conducted. The obtained melting curve and T m values were consistent with those predicted, albeit with a slightly different T m value and a more distinct melting curve. All evaluated species of Limnonectes could be easily distinguished from one another by comparing the melting curve shapes. The HRM assay was then used to confirm the species of 18 Limnonectes samples in comparison to the reference samples (confidence interval > 90%). In addition, the results of HRM were consistent with those of experts who used morphological analysis to identify species. The HRM was found to be useful, and therefore the method would also contribute to future ecological and systematic studies on the target species., (© 2023. The Author(s).)

Capture-based aquaculture is now gaining much attention in Southeast Asia. This system was used to produce several fish species with social and economic implications, including the giant snakehead (Channa micropeltes). As wild harvesting of organisms for seed stock is one of main practices in capture-based aquaculture, abundance and distribution of the wild stock are essential for both environmental impact evaluation and stock management. Mark and recapture, visual observation and physical capture of target species are costly, ineffective, and labour intensive for fish surveys in several cases. Detection of target organisms using eDNA (environmental DNA) could be a good alternative as it has proved to be a non-invasive, rapid, and sensitive method for aquatic species monitoring and surveying. Here, we developed a TaqMan assay that targets the 16S region of giant snakehead DNA to amplify eDNA captured in water samples. 300 µl of water samples were collected from 15 sites located in the Chao Phraya River Basin (Ping, Wang, Yom, Nan, and Chao Phraya River) and filtered with 0.7 µm glass fibre membrane filter. Giant snakehead eDNA was detected in most tributaries (60%) with concentrations ranging from 74.0 copies/ml in Wang River sites to 7.4 copies/ml in Nan River sites. As intensification of capture-based aquaculture could lead to depleting of wild fish stocks, urgent management is needed. However, the existing conventional approaches for assessment of fish overexploitation, survey and monitoring have several limitations., Competing Interests: The authors have declared that no competing interests exist.

Large tropical and subtropical rivers are among the most biodiverse ecosystems worldwide, but also suffer from high anthropogenic pressures. These rivers are hitherto subject to little or no routine biomonitoring, which would be essential for identification of conservation areas of high importance. Here, we use a single environmental DNA multi-site sampling campaign across the 200,000 km 2 Chao Phraya river basin, Thailand, to provide key information on fish diversity. We found a total of 108 fish taxa and identified key biodiversity patterns within the river network. By using hierarchical clustering, we grouped the fish communities of all sites across the catchment into distinct clusters. The clusters not only accurately matched the topology of the river network, but also revealed distinct groups of sites enabling informed conservation measures. Our study reveals novel opportunities of large-scale monitoring via eDNA to identify relevant areas within whole river catchments for conservation and habitat protection.

A lack of reliable tools for determining the presence and distribution of fish species can impede understanding of predator-prey interactions and fishery management. Conventional fish survey methods are invasive, and can be size or species selective. Combining netting and electrofishing is a current method used to monitor fish species in Phayao Lake (Kwan Phayao), Thailand. However, the methods are inefficient and time-consuming. Recently, locals who rely on inland fisheries in Kwan Phayao expressed their deep concerns about the giant snakehead, Channa micropeltes (Cuvier, 1831) destroying other fish there. The giant snakehead prey on many commercially important fish species, as the prey species is reduced, negative effects on both biodiversity and the fishery sector could follow. Here, an eDNA-based survey was developed to detect the presence of the giant snakehead. Water samples were collected from six sites within Kwan Phayao and 17 sites in Ing River where water flowed into and out of Kwan Payao. The eDNA of the giant snakehead was detected in water samples from all collection sites using the developed qPCR assay with various concentrations. The eDNA was shown here to be a sensitive and reliable tool for fish surveillance so there will be a better chance for developing an effective management strategy.

Background: A variety of plants in Acanthaceae have long been used in traditional Thai ailment and commercialised with significant economic value. Nowadays medicinal plants are sold in processed forms and thus morphological authentication is almost impossible. Full identification requires comparison of the specimen with some authoritative sources, such as a full and accurate description and verification of the species deposited in herbarium. Intake of wrong herbals can cause adverse effects. Identification of both raw materials and end products is therefore needed., Methods: Here, the potential of a DNA-based identification method, called Bar-HRM (DNA barcoding coupled with High Resolution Melting analysis), in raw material species identification is investigated. DNA barcode sequences from five regions (matK, rbcL, trnH-psbA spacer region, trnL and ITS2) of Acanthaceae species were retrieved for in silico analysis. Then the specific primer pairs were used in HRM assay to generate unique melting profiles for each plants species., Results: The method allows identification of samples lacking necessary morphological parts. In silico analyses of all five selected regions suggested that ITS2 is the most suitable marker for Bar-HRM in this study. The HRM analysis on dried samples of 16 Acanthaceae medicinal species was then performed using primer pair derived from ITS2 region. 100% discrimination of the tested samples at both genus and species level was observed. However, two samples documented as Clinacanthus nutans and Clinacanthus siamensis were recognised as the same species from the HRM analysis. Further investigation reveals that C. siamensis is now accepted as C. nutans., Conclusions: The results here proved that Bar-HRM is a promising technique in species identification of the studied medicinal plants in Acanthaceae. In addition, molecular biological data is currently used in plant taxonomy and increasingly popular in recent years. Here, DNA barcode sequence data should be incorporated with morphological characters in the species identification.

Medicinal plants are used as a popular alternative to synthetic drugs, both in developed and developing countries. The economic importance of the herbal and natural supplement industry is increasing every year. As the herbal industry grows, consumer safety is one issue that cannot be overlooked. Herbal products in Thai local markets are commonly sold without packaging or labels. Plant powders are stored in large bags or boxes, and therefore buying local herbal products poses a high risk of acquiring counterfeited, substituted and/or adulterated products. Due to these issues, a reliable method to authenticate products is needed. Here DNA barcoding was used in combination with High Resolution Melting analysis (Bar-HRM) to authenticate three medicinal Acanthaceae species (Acanthus ebracteatus, Andrographis paniculata and Rhinacanthus nasutus) commonly used in Thailand. The rbcL barcode was selected for use in primers design for HRM analysis to produce standard melting profiles of the selected species. Melting data from the HRM assay using the designed rbcL primers showed that the three chosen species could be distinguished from each other. HRM curves of all fifteen test samples indicated that three of tested products did not contain the indicated species. Two closely related species (A. paniculata and R. nasutus), which have a high level of morphological similarity, were interchanged with one another in three tested products. Incorrect information on packaging and labels of the tested herbal products was the cause of the results shown here. Morphological similarity among the species of interest also hindered the collection process. The Bar-HRM method developed here proved useful in aiding in the identification and authentication of herbal species in processed samples. In the future, species authentication through Bar-HRM could be used to promote consumer trust, as well as raising the quality of herbal products.