Molecular Analysis of Tuberculosis Transmission in Household Contacts Using a Culturomics Approach

Abstract

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), continues to be a major global public health concern. While molecular diagnostics have advanced, culture is still the gold standard for TB diagnosis. However, certain sub-populations of Mtb in clinical samples, termed differentially culturable tubercule bacteria (DCTB), cannot grow under standard culture conditions, resulting in them being missed during diagnosis, particularly in paucibacillary disease. This also impacts transmission studies, which are based on the premise that all bacteria in a clinical sample have equivalent abilities to grow therefore, complicating transmission network mapping. This study employed a culturomics approach, integrating various culture techniques, quantitative polymerase chain reaction (qPCR), and whole genome sequencing (WGS) to investigate TB transmission dynamics within households of TB patients. In 247 household contacts (HHCs) of 100 index participants, 112 (45%) tested positive for Mtb in at least one assay. The standard Mycobacterial Growth Indicator Tube (MGIT) assay identified the majority of Mtb-positive cases, however, the most probable number (MPN) assay used to detect DCTB, and colony-forming unit (CFU) assays detected 55 additional cases missed by MGIT culture across the 16-month timeline. Combining MGIT culture and MPN assays increased the yield of TB detection in HHCs from 14.2% to 22.7%. MPN assays also identified an additional 15 (26%) transmission links within households, which were missed by routine culture. An in-house molecular diagnostic method, qPCR identified 59 Mtb-positive HHCs that were culture negative. These findings highlight the limitations of the current culture techniques in identifying all populations of Mtb. Integrating routine culture with MPN/CFU assays provides a more comprehensive understanding of Mtb-positive HHCs and the communities they reside in thereby, contributing to improved TB control strategies.