Upper respiratory tract commensal Streptococcus oralis inhibits mycobacterial growth by producing bioactive molecules and inducing macrophage autophagy.

BACKGROUND

Tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb) and remains a major global health challenge. Although host immunity is critical for controlling infection, the role of upper respiratory tract commensal microbiota in anti-mycobacterial defense remains unclear. This study investigated whether commensal bacteria from close contacts of patients with TB, who remained persistently tuberculin skin test- (TST) and QuantiFERON-TB (QFT) Gold-negative after long-term exposure, produce factors capable of inhibiting mycobacterial growth.

METHODS

Throat swabs were collected from close contacts of patients with TB and subjected to ex vivo enrichment culture. Cell-free microbial culture supernatants were tested for activity against Bacillus Calmette-Guérin (BCG), H37Rv, and Mycobacterium smegmatis (M. smegmatis) in cell-free culture and THP-1-derived macrophage infection models. Microbiota composition was analyzed by 16S ribosomal ribonucleic acid (rRNA) gene amplicon sequencing, and functional strains were isolated and identified by whole-genome sequencing. The physicochemical properties of active components were examined using ethanol precipitation and proteinase K digestion. Autophagy-related proteins were assessed by Western blotting, and in vivo activity was evaluated in mouse models of M. smegmatis and BCG infection.

RESULTS

Supernatants from selected throat swab cultures significantly inhibited mycobacterial growth and reduced intracellular bacterial burdens in macrophages. Microbiota analysis and functional screening identified Streptococcus oralis (S. oralis) as a candidate anti-mycobacterial commensal. The culture supernatant of an isolated S. oralis strain inhibited BCG, H37Rv, and M. smegmatis, and decreased intracellular mycobacterial survival. Its activity was enriched in ethanol-precipitated fractions and was markedly reduced after proteinase K digestion, suggesting proteinaceous or peptide-like active components. Treatment with S. oralis supernatant reduced mTOR and p62 expression and increased LC3B-II abundance, consistent with activation of autophagy-related pathways. Oral administration of the supernatant reduced bacterial loads in mouse infection models.

CONCLUSIONS

Upper respiratory tract-derived S. oralis may contribute to anti-mycobacterial defense by producing protease-sensitive bioactive molecules and modulating macrophage autophagy-related pathways. These findings support further investigation of respiratory commensal-derived products as potential host-directed anti-TB strategies.