Comparative Physiological and Metabolomic Responses of Procambarus clarkii and Macrobrachium nipponense to Combined Salinity-pH and NaHCO3 Alkalinity Stress.

Procambarus clarkii and Macrobrachium nipponense are two freshwater shrimp species used in rice-farming systems. To guide their cultivation in saline-alkaline waters, two 96 h experiments were conducted on juvenilels of each species using 540 juveniles per species (initial body length: 1.51 ± 0.16 cm for P. clarkii and 0.88 ± 0.03 cm for M. nipponense): combined salinity-pH stress (0/15.0/25.0‱ at pH 7.4/9.0/10.0) and NaHCO3 alkalinity stress (0/10/20 mmol/L). Under salinity-pH stress, survival of P. clarkii was 100%, 72.23%, and 53.33%, respectively; for M. nipponense, it was 93.33%, 61.13%, and 23.33%. Under alkalinity stress, survival of P. clarkii was 92.22%, 82.22%, and 61.11%; for M. nipponense, it was 91.11%, 65.55%, and 46.67%. In both species, superoxide dismutase, acid phosphatase, and alkaline phosphatase decreased while malondialdehyde increased with stress intensity. Metabolomics revealed species-specific reprogramming: P. clarkii upregulated protective lipids (e.g., glycerophosphoethanolamines, fatty acid esters), whereas M. nipponense showed depletion of critical osmolytes (glycerophosphocholines) and accumulation of apoptosis-related ceramides. KEGG analysis confirmed that energy metabolism, amino acid homeostasis, and ABC transporters were primary pathways affected. In conclusion, extreme stress significantly reduced survival, suppressed immunity, and triggered metabolic reprogramming in both species, with P. clarkii showing substantially higher tolerance. For cultivation, P. clarkii is preferred in moderately high saline-alkaline waters, while M. nipponense requires low-alkalinity freshwater with strict monitoring. These findings support species-specific management of saline-alkaline rice-shrimp systems.