Modulation of the gut microbiota composition by rifaximin in non-constipated irritable bowel syndrome patients: a molecular approach

Abstract

Rifaximin, with its low systemic absorption, may represent a treatment of choice for irritable bowel syndrome (IBS), mainly due to its ability to act on IBS pathogenesis, through the influence on gut microbiota. The aim of the present study was to assess, by biomolecular tools, the rifaximin active modulation exerted on gut microbiota of non-constipated IBS patients. Fifteen non-constipated IBS subjects were treated with 550 mg rifaximin three times a day for 14 days. Stool samples were collected before starting the treatment, at the end of it, and after a 6-week washout period. Real-time polymerase chain reaction, denaturing gradient gel electrophoresis, and next-generation sequencing were applied to all the samples to verify and quantify possible microbial fluctuations. Rifaximin treatment did not affect the overall composition of the microbiota of the treated subjects, inducing fluctuations in few bacterial groups, balanced by the replacement of homologs or complementary bacterial groups. Rifaximin appeared to influence mainly potentially detrimental bacteria, such as Clostridium, but increasing the presence of some species, such as Faecalibacterium prausnitzii. A decrease in the Firmicutes/Bacteroidetes ratio after 14 days of treatment and bacterial profiles with higher biodiversity were observed during the follow-up compared to baseline. Rifaximin treatment, although effective on IBS symptom relief and normalization of lactulose breath test, did not induce dramatic shifts in the microbiota composition of the subjects, stimulating microbial reorganization in some populations toward a more diverse composition. It was not possible to speculate on differences of fecal microbiota modification between responders vs nonresponders and to correlate the quali-/quantitative modification of upper gastrointestinal microbiota and clinical response.

Keywords: IBS; microbiota; next-generation sequencing; rifaximin.

Figure 1

Hierarchical clustering of classified sequences, according to the order–family taxonomy levels for taxa participating with ≥10% in at least one sample. Notes: Different colors indicate the bacterial taxa identified for each sample. No specific clustering was observed. Abbreviations: HS, healthy subjects; T0, baseline; T14, at the end of 14 days of treatment; T56, at the end of the 6-week follow-up period.

Figure 2

Pie charts of the community compositions of samples derived from IBS patients and healthy subjects. Notes: The mean values of the major order/family level taxa (A) and genera (B) are presented for each study group. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Abbreviations: IBS, irritable bowel syndrome; HS, healthy subjects; IS, incertae sedis; Unc, unclassified.

Figure 3

Boxplots of diversity index (Simpson’s D) grouped according to samplings and non-constipated IBS/HS subjects. Notes: The corresponding mean values and also the mean values of the IBS-associated sampling groups are also provided on the top right-hand side. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Abbreviations: IBS, irritable bowel syndrome; HS, healthy subjects.

Figure 4

Distance-based redundancy analysis (RDA) showing the combined effect of subject and sampling time on the composition of the bacterial communities. Notes: The total variance explained by the model along with its partition into the model components are reported in the plot above. Model explained variance: 68.6% (P=0.005) of total: subject variance 94.3% (P=0) of explained, time variance 5.7% (P=0.028) of explained. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Abbreviation: S, subject.

Figure 5

Pairwise comparisons according to treatment and time. Notes: (A) HS, T0, T14, T56 for dominant OTUs; (B) Taxonomic lineages of the OTUs displayed in the bar plot (A), accounting for a participation of at least 1% in the analyzed samples. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Letters ‘a’ and ‘b’ are used to indicate the groups of significance according to the performed pairwise comparisons test. ‘ab’ show that no statistically significant differences were shown between the particular sample group and the sample groups classified in ‘a’ or ‘b’. The data in bold show the OTUs showing differential abundance between treatments. Abbreviations: OTUs, operational taxonomic units; HS, healthy subjects.

Figure 5

Pairwise comparisons according to treatment and time. Notes: (A) HS, T0, T14, T56 for dominant OTUs; (B) Taxonomic lineages of the OTUs displayed in the bar plot (A), accounting for a participation of at least 1% in the analyzed samples. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Letters ‘a’ and ‘b’ are used to indicate the groups of significance according to the performed pairwise comparisons test. ‘ab’ show that no statistically significant differences were shown between the particular sample group and the sample groups classified in ‘a’ or ‘b’. The data in bold show the OTUs showing differential abundance between treatments. Abbreviations: OTUs, operational taxonomic units; HS, healthy subjects.

Figure 6

Spearman’s rank correlation test between bacterial taxonomical annotations followed by hierarchical clustering of the obtained correlation values. Abbreviations: IS, incertae sedis; Unc, unclassified.

Figure 7

Firmicutes/Bacteroidetes ratio box-plot (left) and density plot (right). Notes: The box-plot provides the median values obtained from the samples according to the collection moment. T0: baseline; T14: at the end of 14 days of treatment; T56: at the end of the 6-week follow-up period. Abbreviation: HS, healthy subjects.