Diverse Lineages and Complex Genomic Variation Associated with Pyrazinamide Resistance in Mycobacterium tuberculosis in Oman Revealed by Whole-Genome Sequencing

Tuberculosis (TB) in Oman remains above the pre-elimination threshold of the End TB Strategy, largely due to cases among the expatriate population. Pyrazinamide (PZA), a key first-line anti-TB drug that shortens treatment duration, is increasingly threatened by rising PZA resistance (PZA-R). This study analyzed 241 Mycobacterium tuberculosis (MTB) isolates collected across Oman between 2018 and 2023, including PZA-resistant (PZA-R) (n = 105) and PZA-susceptible (PZA-S) (n = 74) isolates. Drug susceptibility testing was performed using BD MGIT SIRE/PZA kits. Mutations in candidate PZA resistance genes (pncA, panD, and rpsA) and spoligotype profiles were assessed, while a subset of 96 isolates underwent whole-genome sequencing (WGS). MTB isolates showed high genetic diversity, comprising ten clades across four major lineages, with PZA-R predominating in lineage 1. Mutations in pncA, panD, and rpsA showed low sensitivity (≈10%) but high specificity (97.5%) for detecting PZA-R. Consistent with these findings, WGS showed poor concordance with phenotypic PZA resistance (κ = 0.15), but accurately predicted resistance to rifampicin (κ = 0.85) and isoniazid (κ = 0.79). Genome-wide association analysis identified 16 genes associated with PZA-R; however, the detected variants were not consistently linked to the resistant phenotype, suggesting lineage-specific polymorphisms or compensatory adaptations. Phylogenetic analysis revealed substantial diversity, with PZA-R isolates distributed across multiple lineages differing by more than 50 SNPs. These findings highlight substantial genotype–phenotype discordance in PZA resistance and support further studies integrating quantitative phenotyping and genomic analysis. Further genomic studies can therefore improve resistance prediction and monitoring of the spread of drug resistance.