P018

1:San Diego State University

Five New and Repurposed Drugs (NRDs) were approved in the last decade for treatment of multi-drug resistant tuberculosis: bedaquiline, clofazimine, linezolid, delamanid, and pretomanid. Unfortunately, resistance to these drugs emerged faster than anticipated, partly from preexisting resistance in naïve strains. Previous investigations into the rapid emergence mostly used short variants. For the first time, we utilize de novo-assembled genomes, and systematically include Structural Variations (SV) and heterogeneity to comprehensively study this rapid emergence. First, a systematic literature review revealed 313 NRD resistance variants in 13 genes. Next, 409 globally diverse clinical isolates collected prior to the drugs’ programmatic use (308 were MDR, 106 had de novo assembled genomes) were utilized to study the 13 genes comprehensively for conventional, structural, and heterogeneous variants. We identified 5 previously reported and 67 novel putative NRD resistance variants. These variants were 2 promoter mutations (in 8/409 isolates), 13 frameshifts (21/409), 6 SVs (9/409), 35 heterogeneous frameshifts (32/409) and 11 heterogeneous SVs (12/106). Delamanid and pretomanid resistance mutations were most prevalent (48/409), while linezolid resistance mutations were least prevalent (8/409). Preexisting NRD resistance was highly prevalent in these strains (85/409, 21%), primarily through loss-of-function mutations in genes responsible for prodrug activation and efflux pump regulation. Loss-of-function mutations were similarly frequent throughout the genome, suggesting a wide bet-hedging strategy. Hence, any future drugs resistible through loss-of-function in non-essential genes may similarly suffer from preexisting resistance, which will then spread rapidly upon programmatic use. This effect might be mitigated at early-stage drug design through comprehensive preexisting-resistance assessment.