The direct and gut microbiota-mediated effects of dietary bile acids on the improvement of gut barriers in largemouth bass(Micropterus salmoides)

Fish gut barrier damage under intensive culture model is a significant concern for aquaculture industry.This study aimed to investigate the effects of bile acids(BAs)on gut barriers in Micropterus salmoides.A germ-free(GF)zebrafish model was employed to elucidate the effects of the direct stimulation of BAs and the indirect regulations mediated by the gut microbiota on gut barrier functions.Four diets were formulated with BAs supplemented at 0,150,300 and 450 mg/kg,and these 4 diets were defined as control,BA150,BA300 and BA450,respectively.After 5 weeks of feeding experiment,the survival rate of fish fed with BA300 diet was increased(P<0.05).Histological analysis revealed an improvement of gut structural integrity in the BA150 and BA300 groups.Compared with the control group,the expression of genes related to chemical barrier(mucin,lysozyme and complement 1)and physical barrier(occludin and claudin-4)was increased in the BA150 and BA300 groups(P<0.05),and the expression of genes related to immunological barrier(interleukin[IL]-6,tumor growth factorβ,IL-10,macrophage galactosetype lectin and immunoglobulin M[Ig M])was significantly increased in the BA300 group(P<0.05),but the expression of genes related to chemical barrier(hepcidin)and immunological barrier(IL-1β,tumor necrosis factor-a,IL-6 and arginase)was significantly decreased in the BA450 group(P<0.05).Gut microbiota composition analysis revealed that the abundance of Firmicutes was augmented prominently in the BA150 and BA300 groups(P<0.05),while that of Actinobacteriota and Proteobacteria showed a downward trend in the BA150 and BA300 groups(P>0.05).The results of the gut microbiota transferring experiment demonstrated an upregulation of gut barrier-related genes,including immunoglobulin Z/T(Ig Z/T),IL-6,IL-1βand IL-10,by the gut microbiota transferred from the BA300 group compared with the control(P<0.05).Feeding the BA300 diet directly to GF zebrafish resulted in enhanced expression of Ig M,Ig Z/T,lysozyme,occludin-2,IL-6 and IL-10(P<0.05).In conclusion,BAs can improve the gut barriers of fish through both direct and indirect effects mediated by the gut microbiota.