Effects of Combined Exposure to Ammonia and Microcystin-LR on Intestinal Immunity and Microbial Community in Zebrafish(Danio rerio)
Ammonia and microcystin-LR(MCLR)are commonly found pollutants in aquatic environments around the world.To explore the potential synergistic effects of ammonia and MCLR on the zebrafish intestine,adult female zebrafish were exposed to total ammonia nitrogen(30 mg·L-1)、MCLR(10 μg·L-1)and a mixture of both(30 mg·L-1+10 μg·L-1)for a period of 30 d.Histopathological analysis showed that ammonia exposure led to a decrease in intestinal villis area,while MCLR exposure caused villus rupture and increased the ratio of vacuoliza-tion in intestines.The combination of ammonia and MCLR resulted in more serious damage in the intestine.Ammonia alone and in combination with MCLR significantly reduced the contents and related gene expression levels of intestinal lysozyme and β-defensin compared with the control group,indicating suppressed intestinal immunity in zebrafish.Moreover,the single and combined exposure to ammonia and MCLR increased the mRNA expression levels and contents of intestinal inflammatory factors IL-1β and TNF-α by activating NOD1/2 and TLR4a/4b signal pathways,leading to intestinal inflammation.Results of intestinal microbiota showed that both ammonia and MCLR caused changes in diversity.Ammonia nitrogen increased the richness of Firmicutes,while MCLR increased the richness of Actinobacteria and decreased the richness of Proteobacteria.The combined exposure to ammonia and MCLR significantly increased the abundance of Pseudomonas and Mycobacterium.Furthermore,their combined exposure reduced the number of OTUs of short-chain fatty acids producers and the contents of short-chain fatty acids in the intestine.In conclusion,the combination of ammonia and MCLR has a synergistic effect on the intestinal immunity and structure of intestinal flora in zebrafish,and their combined effects on the health of aquatic animals and the water ecosystem's integrity should not be underestimated.
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