Multi-omics analysis of the gill–gut axis in Scylla paramamosain under acute low-salinity stress: Implications for ion and osmotic regulation

The mud crab Scylla paramamosain is a euryhaline species with significant potential for aquaculture in low-salinity environments. However, abrupt exposure of this species to very low saline conditions may exceed its osmoregulatory capacity. Here, we evaluated the physiological, gill transcriptomic, and gut microbiome responses of S. paramamosain after 48 h of acute exposure to salinities of 1–5‰, with a control salinity of 15‰. The survival rate exhibited a sharp decline at salinities ≤5‰ (After 48 h of acute exposure, the survival rates were 33.3% at 1‰, 56.7% at 2‰, 70% at 3‰, 76.7% at 4‰, 86.7% at 5‰, and 100.0% at 15‰), accompanied by increased gill Na⁺/K⁺-ATPase activity, reduced hemolymph osmolality, and elevated hemolymph ammonia content. These results indicate that ion-regulatory responses were activated but were insufficient to sustain osmotic homeostasis under severe acute low-salinity stress. Gill transcriptome analysis between the 1‰ and 15‰ groups identified 1853 differentially expressed genes. Upregulated genes were enriched in ion transport, ABC transporters, calcium signaling, and mitochondrial energy-related pathways, whereas downregulated genes were mainly associated with chitin metabolism, steroid hormone biosynthesis, and molting-related transcriptional processes. Gut 16S rRNA sequencing showed that acute exposure to 1‰ salinity reduced microbial alpha diversity and altered community composition, including reduced abundance of several marine-associated genera such as Photobacterium and enrichment of Lactobacillus. BugBase and PICRUSt2 based analyses indicated differences in predicted microbial phenotypes and inferred functional potential between the 1‰ and 15‰ groups; these predictions should not be interpreted as direct evidence of microbial functional activity. Overall, S. paramamosain showed coordinated physiological, gill transcriptional, and gut microbial responses to acute low salinity. Nevertheless, severe hypoosmotic exposure impaired osmotic balance and reduced survival. These findings provide baseline information for evaluating low-salinity tolerance and suggest that gradual acclimation should be considered when introducing this species into low-salinity culture systems.