Mutations in gidB Confer Low-Level Streptomycin Resistance in Mycobacterium tuberculosis

Abstract

The global threat posed by drug-resistant strains of Mycobacterium tuberculosis demands a greater understanding of the genetic basis and molecular mechanisms that govern how such strains develop resistance against various antituberculous drugs. In this report, we examine a new genetic basis for resistance to one of the oldest and most widely used second-line drugs employed in tuberculosis therapy, streptomycin (SM). This marker for SM resistance was first discovered on the basis of genomic data obtained from drug-resistant M. tuberculosis strains collected in Japan, wherein an association was observed between SM resistance and a mutation in gidB, a putative 16S rRNA methyltransferase. By evaluating an isogenic ⌬gidB mutant strain constructed from strain H37Rv, we demonstrate the causal role of gidB in conferring a low-level SM-resistant phenotype in M. tuberculosis with a 16-fold increase in the MIC over the parent strain. Among clinical isolates, the modest increase in SM resistance conferred by a gidB mutation leads to an MIC distribution of gidB mutation-containing strains that spans the recommended SM breakpoint concentration currently used in drug susceptibility testing protocols. As such, some gidB mutation-containing isolates are found to be SM sensitive, while others are SM resistant. On the basis of a pharmacodynamic analysis and Monte Carlo simulation, those isolates that are found to be SM sensitive should still respond favorably to SM treatment, while nearly half of those found to be SM resistant will likely respond poorly. This report provides the first microbiological evidence for the contribution of gidB in streptomycin resistance and examines the clinical implications of mutations in the gidB gene.The World Health Organization (WHO) 2010 report estimates that 440,000 cases of multidrug-resistant (MDR) tuberculosis (TB) emerged in 2008, with 5.4% of these cases being extensively drug resistant (XDR) TB (36). The strains responsible for producing these forms of TB incur far greater mortality and morbidity to TB patients than drug-sensitive strains (12, 13). If left undetected, these drug-resistant strains can further propagate throughout a community with the potential to cause an epidemic of untreatable, drug-resistant TB. Yet if the patients receive a proper diagnosis and are properly cared for, the success of treatment of MDR-TB can be as high as 60% (36). Therefore, there is an urgent need for faster, more accurate diagnostic alternatives to the current, culture-based drug susceptibility test (DST) methods, which require at least 6 to 9 weeks to determine the resistance pattern of the organism. Fortunately, molecular-based diagnostic methods have already begun to emerge as a viable alternative (2, 23, 38). The most successful of these are based on detecting genotypic polymorphisms associated with drug resistance. In order for these improved methods to supplant culture-based methods, the molecular mechanisms and genetic bases that underlie drug resistance must be understood.In many parts of the w...

Analysis of Mutations in Streptomycin-Resistant Strains Reveals a Simple and Reliable Genetic Marker for Identification of the Mycobacterium tuberculosis Beijing Genotype

“…Among the non-Beijing strains, the gidB C413T mutation has been found in both STR-resistant and -susceptible M. tuberculosis isolates (20,21,23). One STR-resistant isolate carried the mutation G248C, indicating that it might lead to resistance to this drug; however, other mutations in this position (G248T) have been found in STR-susceptible strains (21).…”

“…Many missense mutations have been described in STR-susceptible strains, such as the mutation T47G (the most frequent mutation found in our study), which has been reported as being specific to the Latin American-Mediterranean (LAM) family strains (22); in fact, the 6 strains carrying this mutation in our study belong to the LAM family of strains (data not shown). The four Beijing strains carried two mutations: A276C (E92D), which has been described as an SNP specific to this family (22), and A615G; both mutations can be found in STR-susceptible strains (20). In the Beijing strains, these gidB mutations occur simultaneously with the mutation A128G in the rpsL gene, as mentioned above.…”

“…The role of the GidB protein in conferring high-level STR resistance in M. tuberculosis has been fully characterized (20), although the contribution of certain point mutations in STR resistance is still controversial; it has been speculated that some mutations in the gidB gene might promote the acquisition of mu- (20,21,23). c "InsC 580-581" indicates insertion of a C nucleotide between nucleotides 580 and 581.…”

Regulation of expression and catalytic activity of Escherichia coli RsmG methyltransferase

“…In spite of participation of the 530 loop in the decoding process being clear, the precise role of the m 7 G527 modification has remained elusive. Loss of RsmG activity confers low-level streptomycin resistance and inexplicably prompts the appearance of high-level streptomycin resistance mutations in the S12-encoding gene, rpsL (Nishimura et al 2007a,b;Okamoto et al 2007;Gregory et al 2009;Wong et al 2011). Moreover, the effects of rsmG mutations on bacterial fitness and translational accuracy have been reported as being variable, depending on the bacterial species (Nishimura et al 2007b;Okamoto et al 2007;Gregory et al 2009;Ochi et al 2009).…”

Old and New TB Drugs: Mechanisms of Action and Resistance

“…On the other hand, mechanisms of low-level resistance to streptomycin (MIC = 4-32 mg/L) remain largely undefined but may be attributable to changes in cell envelope permeability or diminished drug uptake (Honore and Cole 1994;Cooksey, Morlock et al 1996). More recently, it has been shown that mutations in gidB, which encodes a conserved S-adenosylmethionine-dependent 16S rRNA methyltransferase, can confer low-level resistance to streptomycin (Wong, Lee et al 2011). …”