Bacillus methylotrophicus influences on the virulence of Aeromonas hydrophila and probiotic applications in Ctenopharyngodon idella

Abstract

Probiotics are alternatives to antibiotics that resist infection and benefit the host in several aspects. Here, we isolated and identified a novel strain of Bacillus methylotrophicus (XA-8) with a broad spectrum of activity against Aeromonas hydrophila AhX040 and other aquatic bacterial pathogens. The strain was non-pathogenic as it revealed no cytotoxic effects in hepatic L8824 cells or pathophysiological signs in the tissue of Ctenopharyngodon idella (grass carp). Using a co-culture approach, XA-8 (1×109 CFU mL−1) when co-cultivated with AhX040 (1×108 CFU mL−1) revealed a reduction in the cell density, cytotoxicity, and infection rate of AhX040. More interestingly, genes such as aer, act, hylA, lafA, hcp, and luxS, associated with the virulence of AhX040, were significantly down-regulated during co-cultivation (P<0.05). Furthermore, XA-8 as a feed supplement to grass carp up-regulated the immune-related genes in the liver, spleen, kidney, and intestine. The villi length and expression level of growth-related genes in the intestine were significantly higher (P<0.05) in the XA-8 supplemented grass carp. In addition to resisting infection (52.63 %), probiotic XA-8 modulated the gut microbiota of grass carp by increasing Fusobacteriota and Bacteroidota at the phylum level, while decreasing Aeromonas at the genus level. The whole genome sequence analysis showed the strain possesses multiple gene clusters for antimicrobial substances and metabolism. Our results demonstrate that B.methylotrophicus XA-8 is a novel probiotic with potential applications for aquaculture.