Molecular epidemiology of resistant M. tuberculosis

The emergence and spread of resistant tuberculosis (TB) pose a threat to public health, so it is necessary to diagnose these forms in a clinically short time frame and closely monitor their transmission. The aim of this study was the molecular characterization of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains of Mycobacterium (M.) tuberculosis using whole genome sequencing (WGS). In overall, 65 isolates phenotypically resistant to at least rifampicin and isoniazid collected in the Czech Republic in 2005-2020 were enrolled for further analysis. The sensitivity and specificity of WGS in the diagnosis of MDR-TB were 100 % and 100 %, respectively. Additional mutations encoding resistance to other first-line and second-line antituberculosis drugs (including delamanid) were observed in 96.2% of isolates (n = 63/65). Although most mutations are high-confidence and known from other studies, we have described novel indels in the pncA gene potentially encoding pyrazinamide resistance. Phylogenetic analysis of WGS data revealed the the majority of MDR M. tuberculosis isolates were the Beijing lineage 2.2.1 (n = 46/65; 70.8%), while the remaining strains belonged to Euro-American lineage. Cluster analysis with a predefined cut-off distance of less than 12 single nucleotide polymorphisms between isolates showed 19 isolates in 6 clusters (clustering rate 29.2%), located mainly in the region of the capital city of Prague. This study highlights the utility of WGS as a high-resolution approach in the diagnosis, characterization of resistance patterns, and molecular-epidemiological analysis of resistant TB in the country.