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Evolution of high-level ethambutol-resistant tuberculosis through interacting mutations in decaprenylphosphoryl-β-D-arabinose biosynthetic and utilization pathway genes

2013; Nature Portfolio; Volume: 45; Issue: 10 Linguagem: Inglês

10.1038/ng.2743

1546-1718

Hassan Safi, Subramanya Lingaraju, Anita G. Amin, Soyeon Kim, Marcus B. Jones, Michael H. Holmes, Michael McNeil, Scott N. Peterson, Delphi Chatterjee, Robert Fleischmann, David Alland,

Biochemical and Molecular Research

David Alland and colleagues report sequencing of Mycobacterium tuberculosis strains under ethambutol drug selection. They follow the multistep progression in acquisition of ethambutol resistance in clinical isolates, finding that an interaction of mutations in several genes in DPA biosynthetic and utilization pathways influence the level of resistance as measured by the minimal inhibitory concentration. To study the evolution of drug resistance, we genetically and biochemically characterized Mycobacterium tuberculosis strains selected in vitro for ethambutol resistance. Mutations in decaprenylphosphoryl-β-D-arabinose (DPA) biosynthetic and utilization pathway genes Rv3806c, Rv3792, embB and embC accumulated to produce a wide range of ethambutol minimal inhibitory concentrations (MICs) that depended on mutation type and number. Rv3806c mutations increased DPA synthesis, causing MICs to double from 2 to 4 μg/ml in a wild-type background and to increase from 16 to 32 μg/ml in an embB codon 306 mutant background. Synonymous mutations in Rv3792 increased the expression of downstream embC, an ethambutol target, resulting in MICs of 8 μg/ml. Multistep selection was required for high-level resistance. Mutations in embC or very high embC expression were observed at the highest resistance level. In clinical isolates, Rv3806c mutations were associated with high-level resistance and had multiplicative effects with embB mutations on MICs. Ethambutol resistance is acquired through the acquisition of mutations that interact in complex ways to produce a range of MICs, from those falling below breakpoint values to ones representing high-level resistance.