Diarrhoea-predominant irritable bowel syndrome distinguishable by 16S rRNA gene phylotype quantification

AIM:To study whether selected bacterial 16S ribosomal RNA(rRNA)gene phylotypes are capable of disting- uishing irritable bowel syndrome(IBS). METHODS:The faecal microbiota of twenty volunteers with IBS,subdivided into eight diarrhoea-predominant (IBS-D),eight constipation-predominant(IBS-C)and four mixed symptom-subtype(IBS-M)IBS patients,and fifteen control subjects,were analysed at three time-points with a set of fourteen quantitative real-timepolymerase chain reaction assays.All assays targeted 16S rRNA gene phylotypes putatively associated with IBS,based on 16S rRNA gene library sequence analysis. The target phylotypes were affiliated with Actinobac-teria,Bacteroidetes and Firmicutes.Eight of the target phylotypes had less than 95%similarity to cultured bacterial species according to their 16S rRNA gene sequence.The data analyses were made with repeated-measures ANCOVA-type modelling of the data and principle component analysis(PCA)with linear mixed-effects models applied to the principal component scores. RESULTS:Bacterial phylotypes Clostridium cocleatum 88%,Clostridium thermosuccinogenes 85%,Coprobacillus catenaformis 91%,Ruminococcus bromii-like, Ruminococcus torques 91%,and R.torques 93%were detected from all samples analysed.A multivariate analysis of the relative quantities of all 14 bacterial 16S rRNA gene phylotypes suggested that the intestinal microbiota of the IBS-D patients differed from other sample groups.The PCA on the first principal component(PC1),explaining 30.36%of the observed variation in the IBS-D patient group,was significantly altered from all other sample groups(IBS-D vs control, P=0.01;IBS-D vs IBS-M,P=0.00;IBS-D vs IBS-C, P=0.05).Significant differences were also observed in the levels of distinct phylotypes using relative values in proportion to the total amount of bacteria.A phy- lotype with 85%similarity to C.thermosuccinogenes was quantified in significantly different quantities among the IBS-D and control subjects(-4.08±0.90 vs -3.33±1.16,P=0.04)and IBS-D and IBS-M subjects (-4.08±0.90 vs-3.08±1.38,P=0.05).Furthermore,a phylotype with 94%similarity to R.torques was more prevalent in IBS-D patients'intestinal micro- biota than in that of control subjects(-2.43±1.49 vs -4.02±1.63,P=0.01).A phylotype with 93%simi- larity to R.torques was associated with control sam- ples when compared with IBS-M(-2.41±0.53 vs -2.92±0.56,P=0.00).Additionally,a R.bromii-like phylotype was associated with IBS-C patients in com- parison to control subjects(-1.61±1.83 vs-3.69± 2.42,P=0.01).All of the above mentioned phylotype specific alterations were independent of the effect of time. CONCLUSION:Significant phylotype level alterationsin the intestinal microbiotas of IBS patients were observed,further emphasizing the possible contribution of the gastrointestinal microbiota in IBS.

Correlation analysis of breast fibroadenoma and the intestinal flora based on 16S rRNA sequencing

Objective To analyze the characteristics of the intestinal microflora in patients with breast fibroadenoma using 16S ribosomal RNA(rRNA)high-throughput sequencing.Methods Fecal samples from 20 patients with breast fibroadenoma and 36 healthy subjects were randomly collected and analyzed using high-throughput sequencing technology for 16S rRNA V4 region sequencing,and the alpha diversity(Chao index,Shannon index)was calculated using Mothur(v.1.39.5)software.Beta diversity was analyzed using QIIME(v1.80).SPSS software(version 23.0)and the t-test of two independent samples were used to analyze differences in the abundance of bacteria between the two groups.Results Compared with that in the healthy control group,theαdiversity of the intestinal microflora in breast fibroadenoma patients increased,but the difference was not statistically significant(P>0.05).At the phylum level,significant differences were observed between the two groups.The abundance of Firmicutes was higher in the breast fibroadenoma group(P<0.05),whereas the abundance of Synergistetes was higher in the healthy control group(P<0.005).A total of five bacterial genera showed significant differences between the two groups:the breast fibroadenoma group showed higher levels of Bautia(P<0.005),Coprococcus(P<0.005),Roseburia(P<0.05),and Ruminococcus(P<0.005),whereas Sutterella was more abundant in the healthy control group than in the breast fibroadenoma group(P<0.05).Conclusion The diversity and abundance of the intestinal flora in patients with breast fibroadenoma are significantly different from those in healthy subjects,suggesting that an imbalance in the intestinal flora is correlated with the occurrence of breast fibroadenoma.

Novel mitochondrial 16S rRNA mutation, 3200T→C, associated with adult-onset type 2 diabetes

Objective To investigate the role of a potential diabetes related mitochondrial region, which includes two previously reported mutations, 3243AG and 3316GA, in Chinese patients with adult onset type 2 diabetes Methods A total of 277 patients and 241 normal subjects were recruited for the study Mitochondrial nt 3116-3353, which spans the 16S rRNA, tRNA leu(UUR) and the NADH dehydrogenase 1 gene, were detected using polymerase chain reaction (PCR), direct DNA sequencing, PCR restriction fragment length polymorphism and allele specific PCR Variants were analyzed by two tailed Fisher exact test The function of the variants in 16S rRNA were predicted for minimal free energy secondary structures by RNA folding software mfold version 3 Results Four homoplasmic nucleotide substitutions were observed, 3200TC, 3206CT, 3290TC and 3316GA Only the 3200TC mutation is present in the diabetic population and absent in the control population No statistically significant associations were found between the other three variants and type 2 diabetes The 3200TC and 3206CT nucleotide substitutions located in 16S rRNA are novel variants The 3200TC caused a great alteration in the minimal free energy secondary structure model while the 3206CT altered normal 16S rRNA structure little Conclusions The results suggest that the 3200TC mutation is linked to the development of type 2 diabetes, but that the other observed mutations are neutral In contrast to the Japanese studies, the 3316GA does not appear to be related to type 2 diabetes

Changes in the gut microbiota of osteoporosis patients based on 16S rRNA gene sequencing:a systematic review and meta-analysis

Background:Osteoporosis(OP)has become a major public health issue,threatening the bone health of middle-aged and elderly people from all around the world.Changes in the gut microbiota(GM)are correlated with the maintenance of bone mass and bone quality.However,research results in this field remain highly controversial,and no systematic review or meta-analysis of the relationship between GM and OP has been conducted.This paper addresses this shortcoming,focusing on the difference in the GM abundance between OP patients and healthy controls based on previous 16S ribosomal RNA(rRNA)gene sequencing results,in order to provide new clinical reference information for future customized prevention and treatment options of OP.Methods:According to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses(PRISMA),we comprehensively searched the databases of Pub Med,Web of Science,Embase,Cochrane Library,and China National Knowledge Infrastructure(CNKI).In addition,we applied the R programming language version 4.0.3 and Stata 15.1 software for data analysis.We also implemented the Newcastle-Ottawa Scale(NOS),funnel plot analysis,sensitivity analysis,Egger’s test,and Begg’s test to assess the risk of bias.Results:This research ultimately considered 12 studies,which included the fecal GM data of 2033 people(604 with OP and 1429 healthy controls).In the included research papers,it was observed that the relative abundance of Lactobacillus and Ruminococcus increased in the OP group,while the relative abundance for Bacteroides of Bacteroidetes increased(except for Ireland).Meanwhile,Firmicutes,Blautia,Alistipes,Megamonas,and Anaerostipes showed reduced relative abundance in Chinese studies.In the linear discriminant analysis Effect Size(LEfSe)analysis,certain bacteria showed statistically significant results consistently across different studies.Conclusions:This observational meta-analysis revealed that changes in the GM were correlated with OP,and variations in some advantageous GM might involve regional differences.

Microbial community changes in the digestive tract of the clam Meretrix petechialis in response to Vibrio parahaemolyticus challenge

Disease in clams frequently occurred over the last decade and has become a serious threat to the clam aquaculture industry and natural stocks.Mass clam mortality events were reported to be associated with the presence of opportunistic pathogen vibrio.However,the complexity of infection that occurs in the natural environment remains poorly understood.In this study,we smulated a natural disease outbreak by vibrio immersion infection to study the diversity and dynamics of microbiota in the digestive tract of clam Meretrix petechialis during the infection process.Dramatic changes in operational taxonomic unit richness and phylum composition of the bacterial communities were observed during pathogen invasion.In addition,we investigated the potential relationship between microbiota dynamics and host status during disease progression.Results reveal that,at the end stage of vibrio infection,interindividual variation in the digestive tract microbiota increased,as did the diff erence in individual health status.The moribund clams displayed signs of microbial community shifts to low diversity,and the microbial community was characterized by mass proliferation of a few operational taxonomic units.

Application of PCR-DGGE in Research of Bacterial Diversity in Drinking Water

Objective To analyze the structure of bacteria in drinking water by molecular biological techniques, Methods DNA of bacteria in drinking water was directly extracted without culture. 16S ribosomal DNA fragments, including V-6, -7, and -8 regions, were amplified with universal primers （EUBf933CJC and EUBr1387） and analyzed by DGGE. Results DGGE indicated that amplification products could be separated, The results showed that DGGE could be used in the separation of different microbial 16SrRNA genes extracted from drinkng water. Though there were special bacteria in different water samples, the predominant bacteria were essentially the same. Three sequences of the reclaimed specific bands were obtained, and phylogenetic tree of these bands was made. Conclusion Bacterial diversity in drinking water is identified by molecular biological techniques.

Rapid detection of sepsis complicating acute necrotizing pancreatitis using polymerase chain reaction

INTRODUCTIONAcute narcotizing pancreatitis usually takes a severe clinical course and is associated with multiple organ dysfunction .With the further understanding of pathophysiological events of acute pancreatisis and the therapeutic measuses taken by the clinicians ,the patients can pass through the critical carry stages ,and then the septic complication caused by rtanslocated bacteria, mostly gram-negative microbes from the intestines ensues[1].

Microbial community structure of different tongue fur types in patients with chronic gastritis

OBJECTIVE:To explore the correlation between tongue and oral microbiota,we studied the microbial community structure of different tongue coating types in patients with chronic gastritis.METHODS:16S rDNA gene sequencing and bioinformatics analysis were used to study the dynamic changes and correlation of microbial flora in patients with chronic gastritis,healthy people,and patients with different tongue fur.In addition,it was also discussed between the severity of gastritis and the microflora of tongue fur.RESULTS:The microbial diversity of tongue fur in patients with chronic gastritis was significantly different from healthy controls.There were significant changes in bacterial communities’diversity and relative abundance between extra tongue fur in patients but not in healthy people.Oral bacteria with relative abundance>1%and P<0.05 among different tongue fur flora were dominant bacteria,including 12 phyla such as Bacteroidetes,Proteobacteria and Firmicutes,and 256 genera such as Neisseria,Prevotella_7 and Haemophilus.CONCLUSIONS:The changes in oral flora in patients with chronic gastritis were related to tongue fur.Therefore,the significant microbiota might enlighten further study on the correlation between tongue inspection and oral microbiota in patients with chronic gastritis.

Dietary supplementation with Clostridium butyricum improves growth performance of broilers by regulating intestinal microbiota and mucosal epithelial cells

Clostridium butyricum has been widely considered an antibiotic substitute in recent years.It can promote growth performance,improve the immune response and enhance the intestinal barrier function of the host.In the present study,1-d-old Arbor Acres(AA)broilers were fed C.butyricum(1×109 cfu/kg)for 28 d.The transcriptomic characteristics of epithelial cells of the cecal mucosa were determined by RNA-sequence,and the cecal microbiota composition was explored by 16 S ribosomal RNA gene sequencing.The changes in the intestinal mucosa of broilers were then analyzed by tissue staining.Gene Ontology(GO)annotations identified substance transport and processes and pathways that might participate in intestinal development and cell viability.Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis revealed that the differentially expressed genes are involved in numerous pathways related to amino acid and vitamin metabolism and antioxidant and defensive functions,among others.The relative expression of some genes associated with intestinal barrier function(claudins 2,15,19,and 23,tight junction proteins 1,2,and 3 and mucin 1)was significantly increased in the treatment group(P<0.05 or P<0.01).Moreover,the proportion of Firmicutes was higher in the C.butyricum-treated group,whereas the proportion of Proteobacteria was higher in the control group.At the genus level,the relative abundances of Butyricicoccus and Lactobacillus,among other bacteria,were increased after C.butyricum supplementation.The tissue staining analysis showed that the cecal mucosa of broilers was significantly ameliorated after the addition of C.butyricum(P<0.05 or P<0.01).These results showed that dietary supplementation with C.butyricum can enhance the antioxidant capacity,mucosal barrier function,and stabilize the cecal microbiota,resulting in improving the growth performance.

Antibiotic resistance mechanisms of Myroides sp.

Bacteria of the genus Myroides （Myroides spp.） are rare opportunistic pathogens. Myroides sp. infections have been reported mainly in China. Myroides sp. is highly resistant to most available antibiotics, but the resistance mechanisms are not fully elucidated. Current strain identification methods based on biochemical traits are unable to identify strains accurately at the species level. While 16S ribosomal RNA （rRNA） gene sequencing can accurately achieve this, it fails to give information on the status and mechanisms of antibiotic resistance, because the 16S rRNA sequence contains no information on resistance genes, resistance islands or enzymes. We hypothesized that ob- taining the whole genome sequence of Myroides sp., using next generation sequencing methods, would help to clarify the mechanisms of pathogenesis and antibiotic resistance, and guide antibiotic selection to treat Myroides sp. infec- tions. As Myroides sp. can survive in hospitals and the environment, there is a risk of nosocomial infections and pandemics. For better management of Myroides sp. infections, it is imperative to apply next generation sequencing technologies to clarify the antibiotic resistance mechanisms in these bacteria.