| The mammalian and fish intestinal mucosal surface is continuously exposed to a complex and dynamic community of microorganisms,therefore,one of the major functions of the intestinal mucosal surface is to act as a protective barrier against the enormous amount of bacteria that resides in the lumen.However,researchers mainly focused on the mucosal intestinal epithelium and tight junctions,ignored the mucus layer as an important defense system,where was the first place for the host-microbial interaction.This inevitable colonization by the microbiota has shaped an intimate relationship where host and microbes have co-evolved for various effective adaptation mechanism,and showed different characteristics between different gut regions.Studies on the characteristics and relationships between host,mucin and adhesive gut microbiota may provide more basis for a better understanding of the interaction between microorganisms and hosts,and are also important for the gut health of fish and discovery of new probiotics.In this paper,proteomics,transcriptomics and metagenomic were presented to investigated the differences in gut function,mucus proteins and genes associated with gut health,and adhesive gut microbiota between different zebrafish gut sections as its naturally folded into three segments(foregut,midgut and distal gut),and revealed the importance of intestine adhesive microbial homeostasis to intestine and host health.Finally,an attempt was made to illustrate the complex relationship between gut health and adhesive gut bacteria in breeding culture,taking tilapia as an example.The main conclusions are as follows:1.We stained the gut mucus of different gut segments of zebrafish as its natural folds,and found that the mucus layer covered in foregut,midgut,and distal gut were all coated with only one layer of mucus,the mean mucus thickness of foregut,midgut,and distal gut were 10.92 ?m,12.56 ?m,and 9.67 ?m,respectively.Screening of significant differential proteins by proteomics,there were 353 in the comparison between midgut and foregut,329 between distal gut and foregut,but only 201 between distal gut and midgut,indicating the foregut harbored a highest differentially expressed proteins.Pathway enrichment analysis of these proteins found that they were mainly involved in metabolic pathways(carbon fixation,nitrogen metabolism),biosynthesis of amino acids,carbon metabolism and so on,indicating that these proteins were mainly associated with nutritional metabolism.Finally,we screened the proteins with different barrier function and the distribution of mucins in the different gut section,such as mechanical barrier,chemical barrier,immune barrier and microbial barrier,respectively.we revealed that mucus from different gut sections took different barrier strategies.2.Differentially expressed genes(DEGs)between different zebrafish gut sections were investigated by transcriptomics,it's obviously that the gene expression patterns in distal gut was more different than the foregut and midgut,both the significant up-regulated and down-regulated genes were more than the other two parts.A total of 8 significant different pathways in comparison between midgut and foregut,23 between distal gut and foregut,24 between distal gut and midgut were enriched by pathway analysis of the DEGs,among the latter two comparisons,18 pathways were the same,which also illustrate the functional differences of the distal.Similarly,we screened DEGs related to gut health,and found that the immune system process and epithelial development related genes are mostly distributed in the distal gut,followed by foregut,and then midgut,indicating regional differences in gut immunoregulatory function and cell barrier function.However,gata5,which was a critical gene in regulating mucus secretion and intestinal epithelial differentiation,was most abundant in the midgut,showed a great correlation with the thickness of intestinal mucus,suggesting that this gene is positively correlated with the mucous barrier in the gut.3.Comparison of the adhesive bacteria composition by meta 16 S seq between different gut segments of normal breeding zebrafish showed a closer microbial community structure between foregut and distal gut.Proteobacteria,Fusobacteria,Firmicutes were 3 predominant phylums with significantly different proportions in each segments.At the genus level,Cetobacterium and Plesiomonas were the two predominant groups without considering the “unknown”,and the relative abundance of Cetobacterium reflected a significant high level in midgut(31.35±1.21%),but with no statistically significant difference between foregut(8.67±0.50%)and distal gut(13.14±3.43%).Streptococcus and Lactococcus presented significant high abundance in foregut and decreased gradually along the intestinal tract,Pseudomonas and Shewanella with no statistically difference between foregut and midgut but significantly reduced in the distal gut.In contact with the thickest mucus in the midgut,we believed that the Cetobacterium had a better colonization ability and played an important role in the microbial barrier function in fish.It also showed that the adhesive microbiota can not be ignored and mixed with the content flora in evaluation of the gut and the host's health,but should be presented separately as a health assessment marker.4.In this chapter,the adhesive microbiota composition and health related indicators in gut were used to evaluate the effects of dietary microbial phytase on tilapia.The diversity of adhesive gut bacterial communities were strikingly increased that the relative abundance of Proteobacteria,Firmicutes and Thermus were all significantly enhanced by microbial phytase with 1,000 U/kg.At the genus level,the abundances of Clostridium sp.and Thermus sp.also showed significant increase by microbial phytase.At the same time,the microbial phytase had obviously improved the phosphorus bioavailability and microvilli density but reduced microvilli length.Stimulated intestinal inflammation and stress status were observed in the diet supplemented with microbial phytase,as indicated by the up-regulated expressions of the cytokine genes(Tnf-? and Tgf-?)and Hsp70.Therefore,this study suggested that the effect mediated by change in adhesive microbiota was putatively beneficial,and the observed adverse impacts were probably attributed to the direct effect of microbial phytase on the host,which deserves further investigations,but in the assessment of fish health,gut adhesive microbiota which can provide a new perspective need to be taken more into account. |
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