Molecular Analysis of theembCABLocus andembRGene Involved in Ethambutol Resistance in Clinical Isolates of Mycobacterium tuberculosis in France

Abstract

ABSTRACTModification of codon 306 inembBis regarded as the main mechanism leading to ethambutol (ETB) resistance in clinical isolates ofMycobacterium tuberculosis. However, numerous mutations elsewhere in theembCABlocus and inembR, a putative transcriptional activator of this locus, have been reported to be involved in ETB resistance. Here, we investigated the diversity of nucleotide variations observed inembCABandembRinM. tuberculosiscomplex isolates from France. These regions were sequenced in 71 ETB-resistant (ETB-R) and 60 ETB-susceptible (ETB-S) clinical isolates of known phylogenetic lineages. The 131 isolates had 12 mutations corresponding to phylogenetic markers. Among the 60 ETB-S isolates, only 3 (5%) had nonsynonymous mutations that were not phylogenetic markers. Among the 71 ETB-R isolates, 98% had mutations inembCABthat likely contribute to ETB resistance: 70% had mutations located inembBcodon 306, 406, or 497; 13% had mutations located outside these three positions between codons 296 and 426; and 15% had mutations corresponding to mutations in theembC-embAintergenic region. We found a strong association between resistance to ETB and the presence of mutations inembBand theembC-embAintergenic region (P< 0.001). In contrast, the mutations detected inembCandembAwere not involved in ETB resistance, and no mutation was detected inembR. These results strongly suggest that the sensitivity of diagnostic assays for detecting ETB resistance based on testing ofembBcodon 306 can be increased by testing of theembBregion between codons 296 and 497 and by including theembC-embAintergenic region between positions −8 and −21.