In Vitro Study Of The Effect Of Food Ingredients On Gut Microbiome Of Health Person And IBS-D Patients

Background:Prebiotic is a substrate that is selectively utilized by host microorganisms conferring a health benefit.Recent study showed that the prebiotics have different effects on the microbiota of the different enterotypes.Isomaltooligosaccharides(IMO)as an oligosaccharide with prebiotic activity is often used as a dietary supplement to regulate intestinal flora.However,in other hand,oligosaccahrides may impose the detrimental effects on Irritable bowel syndrome(IBS-D)patients.IBS is a commonly diagnosed functional gastrointestinal disorders(FGIDs)characterized by chronic abdominal pain associated with changes in stool form and/or relieved by defecation.Recent study showed that most of IBS patients' symptoms were associated with food and the dietary therapies,and restricting intake of FODMAPs(fermentable short-chain carbohydrates,termed fermentable oligosaccharides,disaccharides,monosaccharides and polyols)may reduce GI symptoms especially for patients who are diarrhea-predominant(IBS-D).Therefore,food ingredients containing prebiotics may have different effects on the gut microorganisms of healthy person and IBS-D patients.Aim:This study is to explore the fermentation performance of intestine microorganisms from normal healthy person and IBS-D patients by using the vitro batch fermentation model in the medium containing oligosaccahrides.The first part of thesis is to study microbiota from healthy person fermenting IMO substrates with five different length of polymerization(DP).The second part of thesis is to study the differences in metabolic status of low-FODMAPs dietary therapy for Chinese IBS-D patients.Methods and results:Eleven fecal slurries collected from three enterotypes,including the Bacteroides-,Prevotella-and Mixed-type,inoculated into fermenting medium containing different length of IMO.The different enterotypes of microbiota exhibited different degradation rates for long chain IMOs(DP7to16).In contrast,the degradation rates for short chain IMOs(DP 2 to 6)were not affected by enterotypes.Both 16S rDNA sequencing and quantitative PCR demonstrated that,after fermentation,the Bifidobacterium growth with IMOs was primarily enhanced in the Bacteroides-and Mixed-type(non-Prevotella-type),and to a lesser degree in the Prevotella-type.Interestingly,the Prevotlla-type microbiome had higher levels of propionic acid and butyric acid production than non-Prevotella-type microbiome after IMOs fermentation.Moreover,principal coordinate analysis(PCoA)of both denaturing gradient gel electrophoresis(DGGE)profiling and 16S rRNA sequencing data demonstrated that the microbiome community compositions were separately clustered based on IMO chain length,suggesting significant impact of DP of IMO on the bacterial community structure.In the second part of study,the fecal SCFA was also measured by Gas chromotograpgy(GC).Total SCFA(P?0.001),acetic acid(P?0.001),propionic acid(P<0.001)increased in IBS group compared to healthy controls.However,in term of molar ratio of SCFA,the percentage of acetic acid in the feces of IBS-D patients showed statistical significantly increase than healthy controls.The percentage of butyric acid,isobutyric acid,valeric acid,isovaleric acid and sum of isobutyric acid and isovaleric acid was found statistical significant higher in healthy controls than IBS group(P?0.05).When calculating the P-B index using the IBS-D P-B equation,"propionic acid-butyric acid",IBS-D P-B index was significantly higher than healthy controls(P?0.05).According to the existing literature,Index A of SCFA is anr index,calculated as(acetic acid-propionic acid-butyric acid)/total SCFAs and Index B is an sum of isobutyric acid and isovaleric acid..After 3-week dietary therapy,LDF-E group showed decreased index A in their feces,whiles LFD-N group showed increased index A.Furthermore,LFD-E showed significantly higher index B than LFD-N group(P<0.05).We then conducted fermentation with fresh stools from IBS-D patients.Fermentation media contained defatted soybean(SOY),skimmed milk powder(MILK),mucin(MUCH),low complex saccharides(MSSL)and high complex saccharides(MSSH)as carbon source,and YCFA using as control.Fermentation was conducted before and after dietary therapy.A trend of decline of index A from LFD-E group was observed in YCFA,SOY,MILK,MSSL and MSSH containing media.The index B from LFD-E group increased after dietary therapy,especially in SOY and MSSH containing medium(P<0.05).Gas production was also detected after 24 hours fermentation.In general,the gas production increased after the dietary therapy.but no statistically significance.The current results clearly demonstrated that the chain length of IMO could modulate the structure and composition of the human colonic microbiome.Different responses to short and long chain IMOs were observed from three human enterotypes,indicating that IMOs may be used as therapeutic substrates for directly altering human colonic bacteria.In addition,patients showed improved syptoms with Low FODMAPS foods had significant changes in the metabolic of SCFAs.It suggests that SCFAs metabolism may play an important role in the pathogenesis and treatment of IBS.The specific mechanism needs further study.