Cold stress reshapes gut microbiota and antibiotic resistance in ornamental fish (Cyprinus carpio)

Antimicrobial overuse in ornamental fish farming drives antibiotic resistance genes (ARGs) proliferation, posing public health risks. However, the effects of cold stress on gut microbiota and ARGs in ornamental fish remain poorly understood. Here, we used 16S rRNA gene sequencing and high-throughput quantitative PCR techniques to clarify the difference of gut microbes and ARGs in Cyprinus carpio exposed to temperatures of 4 °C and 25 °C. Tetracycline and sulfonamide resistance genes dominated carp intestinal ARGs at both 4 °C and 25 °C. Five high-risk ARGs (aadA-01, aadA-02, floR, dfrA1, tetM-02) were identified in carp intestine, though cold stress did not significantly alter their relative abundance. Notably, cold stress reduced the relative abundance of mobile genetic elements (intI-1(clinic)), suggesting that low temperature may reduce the potential of horizontal transfer of ARGs. Two gut microbial phyla (Actinobacteria and Planctomycetes) remarkably increased as temperature decreased. Temperature significantly reduced microbial diversity (P < 0.001) and restructured community composition (R² = 0.674, P = 0.001). The PICRUSt analysis showed that low temperature enriched pathways involved in Synthesis and degradation of ketone bodies, geraniol degradation and fatty acid degradation. In addition, Gut microbial network analysis showed that low-temperature stress enhanced community stability characterized by increased modularity and decreased complexity. Moreover, correlation analysis revealed eight opportunistic pathogens (e.g. Comamonas, Streptococcus) within the carp intestine as putative reservoirs of high-risk ARGs. This study offers critical insights into cold stress effects on ornamental fish gut microbiota and ARGs, informing public health strategies.