Perceived ethionamide resistance in isoniazid susceptible isolates of mycobacterium tuberculosis.

Abstract

In Mycobacterium tuberculosis, resistance to ethionamide (ETH) is usually combined with isoniazid (INH) resistance due to a number of mutations in genes that are involved in the biosynthesis of mycolic acids. ETH resistance in INH susceptible isolates is rare. Ten such isolates were identified from patients participating in other studies. Genotyping by means of IS6110 was performed to compare the relatedness of these isolates to each other. In attempts to identify the molecular basis for the resistance to ETH, the ethA, mshA and mshC genes were amplified and the amplicons sequenced using an ABI 3730 DNA Analyser. INH and ETH minimum inhibitory concentrations (MICs) were determined alone and in combination by means of checkerboard titrations in Middlebrook 7H9 broth, using the 3-(4,5-dimethylthiozol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and microplate alamarblue assays (MABA) for detection of growth. Seven isolates were not related to each other and their INH susceptibility was confirmed. No mutations were observed in all the sequenced genes. One out of seven isolates was found to be co-resistant to INH nad ETH. The MIC for the remaining isolates was 1μg/ml for ETH. MABA revealed a paradoxical susceptibility of the isolates to ETH, where mycobacterial growth was observed in ETH concentrations higher than the MIC for the six isolates. For combination of the two drugs, MABA revealed an antagonism between INH and ETH, where the isolates grew in high ETH concentrations regardless of the concentration of INH. The paradoxical effect of ETH and antagonism between ETH and INH in our isolates does not result from mutations in ethA, mshA or mshC.