Molecular Analysis of Isoniazid-Resistant Mycobacterium tuberculosis Isolates from England and Wales Reveals the Phylogenetic Significance of the ahpC −46A Polymorphism

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Abstract
The present study investigated the prevalence and diagnostic potential of the most commonly reported mutations associated with isoniazid resistance, katG 315Thr, katG 315Asn, inhA −15T, inhA −8A, and the oxyR - ahpC intergenic region, in a population sample of 202 isoniazid-resistant Mycobacterium tuberculosis isolates and 176 randomly selected fully sensitive isolates from England and Wales identified by using a directed oligonucleotide array and limited DNA sequencing. The strains were recovered from patients originating from 29 countries; 41 isolates were multidrug resistant. Mutations affecting katG 315, the inhA promoter, and the oxyR - ahpC intergenic region were found in 62.7, 21.9, and 30% of 169 genotypically distinct isoniazid-resistant isolates, respectively, whereas they were found in 0, 0, and 8% of susceptible strains, respectively. The frequency of mutation at each locus was unrelated to the resistance profile or previous antituberculous drug therapy. The commonest mutation in the oxyR - ahpC intergenic region, ahpC −46A, was present in 23.7% of isoniazid-resistant isolates and 7.5% of susceptible isolates. This proved to be a phylogenetic marker for a subgroup of M. tuberculosis strains originating on the Indian subcontinent, which shared IS 6110 -based restriction fragment length polymorphism and spoligotype features with the Delhi strain and Central Asian strain CAS1; and this marker is strongly associated with isoniazid resistance and the katG 315Thr mutation. In total, 82.8% of unrelated isoniazid-resistant isolates could be identified by analysis of just two loci: katG 315 and the inhA promoter. Analysis of the oxyR - ahpC intergenic region, although phylogenetically interesting, does not contribute significantly to further identification of isoniazid-resistant isolates.