The Relationship Between Intestinal Microbioata And The Clinical Symptoms Of Irritable Bowel Syndrome With Diarrhea And Its Mechanism

Aim:The pathogenesis of irritable bowel syndrome with diarrhea(IBS-D)remains unclear,in recent years the role of intestinal microbiota on the pathogenesis of IBS has attracted much attention.The intestinal microbiota could divided into mucosal-associated microbiota(MAM)and luminal microbiota(LM)because of different position,it was thought that MAM affect the intestinal barrier and immune function through intestinal epithelial cells and immune cells,and LM mainly take charge of the digestion and absorption of carbohydrates.Due to the role of intestinal microbiota in disease depends on their functions,what a kind of role that these two intestinal microbiota composition and function played in the clinical symptoms of IBS-D is unclear,this study investigates the composition and the function of MAM and LM,and their relationship with the clinical symptoms of IBS-D,we explore the possible mechanism between intestinal micreobiota and IBS-D and provide new thoughts for clinic.Methods:(1)Clinical datas were collected from 69 IBS-D patients fulfilling RomanⅢand 20 healthy volunteers.Without intestinal preparation,6 mucosal tissues were taken as MAM specimens by rectoscope,and feces from the same day were taken as LM specimens.The composition of MAM and LM and their correlation with clinical symptoms of IBS-D were analyzed by 16s RNA pyrosequencing.(2)Based on the first part,PICRUSt was used to predict and classify the gene function of the microbiota,to analyze the correlation between the functions of MAM and LM and the clinical symptoms of IBS-D,as well as the relationship between the predominant genera and functional genes in MAM related to clinical symptoms.(3)77 IBS-D patients fulfilling Rome-III criteria were recruited,SIBO was identified according to standard glucose hydrogen breath test.For 16S-r RNA gene sequencing,samples of duodenal mucosa,duodenal fluid,rectal mucosa and fresh feces were collected and performed.The differences in microbial composition and function,in SIBO+and SIBO-IBS-D subjects were evaluated.Results:(1)Our results showed significant differences between MAM and LM both in IBS-D patients and HCs;however,there were greater alterations of MAM than in LM in IBS-D patients compared to HCs.While the composition of MAM were related to IBS-D clinical symptoms,the overall composition of LM did not change significantly and the structure of LM were not related to clinical symptoms.We also found that changes in intestinal MAM in IBS-D patients were closely related to abdominal pain and bloating.Four dominant genera in MAM were associated with abdominal pain and bloating.the proportion of Lachnospira and Collinsella was decreased in IBS-D patients and was negatively correlated with bloating,while the proportion of Lactococcus and Sphingobium was increased in IBS-D patients and was positively correlated with bloating and abdominal pain,respectively.These four bacteria significantly changed in MAM but not in LM.(2)The results of this part were similar to the results of the microbial composition.Our results showed significant differences between predicted function genes of MAM and LM both in IBS-D patients and HCs;however,there were greater alterations of MAM than in LM.While the predicted function genes of MAM were related to IBS-D clinical symptoms,the predicted function of LM genes were not related to clinical symptoms.We also found that five predicted function genes were closely related to clinical symptoms.Those functional genes significantly changed in MAM but not in LM.(3)The microbial diversity and composition obviously differed between SIBO~+and SIBO~-IBS-D in duodenal and rectal mucosa,but not in duodenal fluid and fresh feces.For rectal-mucosal microbiota,it displayed markedly reduced aerobe and Gram-negative bacteria,and increased facultative anaerobe and Gram-positive bacteria,moreover,altered functions of microbial metabolism in SIBO~+IBS-D.Significantly higher rectal mucosa-related MDI was observed in SIBO~+IBS-D,and a cutoff value at-0.37 had a sensitivity of 56.55%and specificity of 90.91%to identify the SIBO in IBS-D subjects.Conclusions:The MAM in IBS-D patients have a more significant dysbiosis than that in LM,and the composition and function of MAM are closely related to the clinical symptoms of IBS-D,which can also be used as a potential indicator for the prediction of SIBO.It’s suggested that we should pay more attention to the role of MAM in the development of IBS-D.