Multilocus DNA Barcoding Resolves Area-Based Genetic Variation in Freshwater Shrimps (Macrobrachium sp. and Caridina sp.) from Saline and Non-Saline Habitats in Northeastern Thailand

Freshwater prawns are important components of freshwater ecosystems, yet their diversity and genetic structure in northeastern Thailand remain insufficiently documented, particularly across habitats differing in soil salinity. This study assessed freshwater prawn diversity and phylogenetic relationships in saline and non-saline habitats using an integrative taxonomic approach combining morphological examination with mitochondrial and nuclear DNA barcoding. A total of 225 mature specimens were collected from 15 sampling sites in Maha Sarakham province (saline habitats) and Mukdahan province (non-saline habitats). Representative specimens were examined morphologically and characterized using four barcode loci: cytochrome c oxidase subunit I (COI), 16S ribosomal RNA (16S rRNA), 18S ribosomal RNA (18S rRNA), and the internal transcribed spacer (ITS). Morphological assessment identified three taxa, Macrobrachium lanchesteri, Macrobrachium niphanae and Caridina sp. Molecular analyses recovered three principal lineages broadly corresponding to these taxonomic groups, although the resolution varied among loci. COI provided consistent species-level discrimination, while 16S rRNA revealed additional genetic variation within Macrobrachium. The conserved 18S rRNA marker supported separation of the major taxonomic lineages, whereas ITS contributed complementary sequence variation for lineage assessment. The concatenated multilocus dataset provided the most stable phylogenetic reconstruction, with strong support for the principal lineages. Samples from saline and non-saline habitats were not consistently separated, suggesting that salinity was not the primary factor underlying the observed genetic structure. Overall, integrating morphological and multilocus molecular evidence provided a robust framework for assessing freshwater prawn diversity and phylogenetic relationships and establishes a baseline for future taxonomic, phylogeographic, and conservation studies in the Mekong region.