The Effect Of Conventional Anti-tuberculosis Drug Therapy On Sputum And Gut Microbiota

Objectives(1)To explore the changes of sputum microbiota in newly diagnosed pulmonary tuberculosis patients during the conventional anti-tuberculosis chemotherapy;(2)To investigate the effects of Mycobacterium tuberculosis infection and conventional anti-tuberculosis chemotherapy on gut microbiome and metabolome.Methods(1)103 pulmonary tuberculosis(TB)patients were recruited in this study,including 54 newly diagnosed TB patients(T0 group)and 32 TB patients who received first-line drug treatment for 2 months(T2 group),and 17 TB patients who received treatment for 6 months and were cured(T6 group).The 16 S r DNA were sequenced by high throughput sequencing,and the sequencing data were analyzed by bioinformatics.(2)We collected 168 stool specimens from 49 healthy volunteers without Mycobacterium tuberculosis infection(HV group),30 healthy volunteers with latently infected by Mycobacterium tuberculosis(LTBI group),41 patients with active TB(ATB group),28 patients with 2-month treatment(A2 group)and 20 patients with 6-month treatment(A6 group).After high-throughput sequencing of the intestinal flora of the above five groups,the differences of the intestinal flora among the HV,LTBI and ATB groups were first analyzed,and then the differences of the gut microbiome among the ATB,A2 and A6 groups were also analyzed.An untargeted Liquid Chromatograph Mass Spectrometer-based metabolomics was used to investigate the alteration of fecal metabolome in HV,ATB,A2 and A6 groups.Finally,Spearman correlation analysis was performed on the differential metabolites and the differential flora.Results(1)Compared with T0 group,α diversity indexes decreased in T2 and T6 group(except Pielou＿e index),and the decrease of α diversity index was more obvious with the prolongation of treatment time.The results of β diversity analysis showed that compared with the T0 group,there were significant differences in the bacterial community structure between T2 and T6.LEf Se analysis results showed that a total of 61 different bacterial communities were identified in the three groups at different taxonomic levels.PICRUSt analysis showed that fatty acid metabolism,butanoate metabolism,bacterial motility proteins and other functional pathways changed in the sputum microbiota of patients after anti-TB chemotherapy compared with T0.(2)There was no significant difference in α diversity indexes between the LTBI and HV groups.The α diversity indexes of ATB group decreased significantly compared with HV and LTBI groups.The results of β diversity analysis showed that the structural composition of intestinal flora in ATB group was significantly different from the other two groups.The LEf Se analysis results showed that a total of 48 different bacterial communities were identified in the HV group and the ATB group.Compared with ATB and A2,the α diversity of A6 group was higher than them,and the difference was statistically significant.The results of β diversity analysis showed that only ATB group and A6 group had differences in intestinal microflora structure.LEf Se analysis results showed that a total of 18 different bacterial communities were obtained at different taxonomic levels.Fecal stool metabolomics analysis showed that compared with the HV group,there were 63 metabolites in the ATB group,142 metabolites in A2 and 53 metabolites in the A6.Spearman correlation analysis showed that changes in fecal metabolites caused by Mycobacterium tuberculosis infection or anti-tuberculosis drug treatment were related to the changes in gut microbiota.Conclusions(1)Anti-TB treatment largely disrupts the composition of the sputum flora,which may lead to changes in the metabolism of intestinal flora and ultimately affect the body’s immune response to Mycobacterium tuberculosis.(2)The diversity of gut microbiome in active TB patients were significantly decreased.With the prolongation of anti-tuberculosis treatment,the diversity of gut microbiome recovered to a certain extent.Some significantly altered microflora in the gut microbiota of TB patients may serve as diagnostic markers for TB.Mycobacterium tuberculosis infection and anti-tuberculosis drug therapy can cause changes in intestinal microbiota and metabolome of TB patients,revealing the potential correlation between intestinal flora and metabolites and the occurrence and development of TB,which can provide potential biomarkers for the diagnosis of tuberculosis or the evaluation of treatment effect.