Abstract:

Objective: To explore the structural differences in the respiratory tract flora community in pulmonary tuberculosis patients with rifampicin (RFP) resistance or sensitivity using 16S high-throughput sequencing technology. Methods: Sputum samples (80 patients) from Beijing Center for Disease Control and Prevention (70 patients) and Hunan Chest Hospital (10 patients) were included in this prospective study. Using Xpert assay, high-throughput targeted rpoB sequencing and RFP susceptibility assay on these sputum samples, the 80 patients were divided into RFP-resistant group (RFP-R, with RFP resistance based on the RFP-resistance determining region (RRDR) mutation; 41 patients) and RFP-sensitive group (RFP-S, no RRDR mutation; 39 patients). High-throughput sequencing of the 16S V4 region, bioinformatics analysis, and statistical analysis of the sequencing results were performed. Results: In total, 41950224 original 16S rRNA reads were obtained from the RFP-R and RFP-S groups, with 344 and 342 average operational classification units (OTUs) for the RFP-R group and RFP-S group, respectively. Firmicutes (47.72%), Bacteroidota (22.36%), Proteobacteria (14.67%), Actinobacteriota (7.57%), and Fusobacteriota (6.46%) were the main bacterial phyla detected, accounting for 98.78% of the microbial community. Compared with the RFP-S group, the relative abundances of the Fusobacteriota (7.531% vs. 5.167%; t=2.218, P=0.030) and Spirochaeta (0.596% vs. 0.246%; t=2.128, P=0.038) in the RFP-R group were significantly higher, the same as for Oribacterium (0.690% vs. 0.326%; t=3.112, P=0.003), Filifactor (0.135% vs. 0.030%; t=2.171, P=0.035), Allisonella (0.046% vs. 0.006%; t=3.237, P=0.002), Pseudomonas (0.205% vs. 0.054%; t=2.040, P=0.046), and Treponema (0.601% vs. 0.242%; t=2.155, P=0.036), whereas Capnocytophaga (0.351% vs. 0.772%; t=―2.044, P=0.046) and Pauljensenia (2.314% vs. 3.706%; t=―2.660, P=0.010) in the RFP-R group showed decreased relative abundances compared with those in the RFP-S group. Conclusion: RFP resistant and sensitive Mycobacterium tuberculosis infection did not affect the overall structure of the respiratory tract flora in pulmonary tuberculosis patients. However, significant differences were observed in the relative abundance of two Phyla (Fusobacteriota, Spirochaeta) and seven Genera (Oribacterium, Filifactor, Allisonella, Pseudomonas, Treponemas, Capnocytophaga, Pauljensenia) between the RFP-resistant and sensitive groups. Specifically, Capnocytophaga and Pauljensenia exhibited higher abundance in the RFP-sensitive group while the remaining taxa showed higher abundance in the RFP-resistant group.

Key words: Tuberculosis, pulmonary, Sputum, Rifampin, Drug resistance, Biodiversity, Bacteria