Refeeding effects on physiological functions and gut microbiota composition in starved red swamp crayfish (Procambarus clarkii)

Abstract

This study investigated the effects of starvation and refeeding on physiological responses and gut microbiota composition in Procambarus clarkii. Four treatment groups were established: a regular feeding group (RF), a 6-day starvation followed by 6-day refeeding group (SF6), a 12-day starvation followed by 12-day refeeding group (SF12), and a continuous starvation group (NS). A total of 360 healthy juvenile crayfish with intact appendages and uniform body size (mean weight: 9.25 ± 0.05 g) were randomly allocated to the four groups, with three replicates per group and 30 individuals per replicate. The feeding trial lasted 24 days. Starvation followed by refeeding significantly improved growth performance, restored physiological functions, and enhanced digestive capacity. Starvation increased body moisture and ash content, while decreasing crude protein and lipid levels. Energy utilization during starvation followed a sequential depletion of glycogen, lipids, and proteins. Elevated SOD activity post-starvation indicated enhanced antioxidative defense against reactive oxygen species and lipid peroxidation. Moreover, starvation markedly upregulated the expression of TGF-β and Mstn, while refeeding—particularly in the SF6 group—led to higher expression levels of ACTL, TNT, Myhb, and MLC. Histological analysis revealed that starvation induced degeneration of intestinal villi, thinning of intestinal walls, and atrophy of R-cells in the hepatopancreas. 16S rRNA gene sequencing showed that starvation significantly reduced gut microbial diversity and richness, increased the relative abundance of Bacteroidota, Shewanella, and Citrobacter, and decreased the Firmicutes/Bacteroidota (F/B) ratio. In contrast, refeeding reversed these changes, enriching the microbiota with Firmicutes, Erysipelotrichaceae, Dysgonomonas, and Rhodobacter. In conclusion, starvation imposed significant physiological and microbial alterations in P. clarkii, including impaired growth traits and increased abundance of potentially pathogenic bacteria. However, refeeding effectively mitigated these effects, demonstrating the resilience of P. clarkii to nutritional stress.