Synergistic toxic effects of copper sulfate and trichlorfon on the health and intestinal microbiota of gibel carp Carassius auratus gibelio

In the high -density pond aquaculture of gibel carp ( Carassius auratus gibelio ) in China, copper sulfate (CuSO 4 ) and trichlorfon (Tri) are commonly used, either in combination or alternately, to prevent and control various diseases and harmful algae. Despite their widespread use, there is a lack of studies on the toxic effects of CuSO 4 and Tri on gibel carp, particularly regarding their combined application. This study assessed the effects of exposure to CuSO 4 (0.5 mg/L) and Tri (0.5 mg/L), both individually and in combination, on gibel carp over 21 days. Histopathological assessments revealed significant structural damage to the liver and intestines in gibel carp exposed to either CuSO 4 or Tri alone, with co -exposure notably intensifying these effects. Analysis of antioxidant enzyme activities revealed a substantial reduction in the liver and intestines, concurrent with a significant rise in the oxidative stress marker malondialdehyde in both single and combined exposure groups. Moreover, quantitative PCR analyses indicated a downregulation of immune -related genes post -exposure, with the combined treatment causing more severe suppression. This suppressed immune response correlated with a reduced ability to resist infection by the pathogenic bacterium Aeromonas hydrophila , particularly in the combined exposure group. This indicates that long-term exposure to CuSO 4 and Tri, particularly in combination, induces significant oxidative stress damage in the liver and intestinal tissues, suppresses immune function, and reduces disease resistance. Additionally, 16S rRNA sequencing of the intestinal microbiota revealed that CuSO 4 and Tri exposure altered the diversity and composition of intestinal microbial communities, potentially bringing both benefits and drawbacks. These findings underscore the need for careful management of CuSO 4 and Tri in aquaculture to mitigate their adverse impacts, especially when used in combination for an extended period.