Mycobacterium tuberculosis Strains with Highly Discordant Rifampin Susceptibility Test Results

Abstract

The objectives of this study were to investigate the origin of highly discordant rifampin (rifampicin) (RMP) drug susceptibility test results obtained for Mycobacterium tuberculosis strains during proficiency testing. Nine Supra-National Tuberculosis Reference Laboratories tested the RMP susceptibilities of 19 selected M. tuberculosis strains, using standard culture-based methods. The strains were classified as definitely resistant (R) (n ‫؍‬ 6) or susceptible (S) (n ‫؍‬ 2) or probably resistant (PR) (n ‫؍‬ 8) or susceptible (PS) (n ‫؍‬ 3) based on rpoB mutations and treatment outcome. All methods yielded a susceptible result for the two S and three PS strains lacking an rpoB mutation and a resistant result for one R strain with a Ser531Leu mutation and one PR strain with a double mutation. Although the remaining 12 R and PR strains had rpoB mutations (four Asp516Tyr, three Leu511Pro, two Leu533Pro, one each His526Leu/Ser, and one Ile572Phe), they were all susceptible by the radiometric Bactec 460TB or Bactec 960 MGIT methods. In contrast, only one was susceptible by the proportion method on Löwenstein-Jensen medium and two on Middlebrook 7H10 agar. Low-level but probably clinically relevant RMP resistance linked to specific rpoB mutations is easily missed by standard growth-based methods, particularly the automated broth-based systems. Further studies are required to confirm these findings, to determine the frequency of these low-level-resistant isolates, and to identify technical improvements that may identify such strains.The prevalence of multidrug-resistant (MDR) tuberculosis (TB) is rising globally, posing a serious threat to TB control. (25) MDR TB does not respond to treatment with first-line drugs, (3), and its management using second-line drugs has not yet been properly organized by most control programs (25). Although MDR TB is defined as resistance to at least isoniazid and rifampin (rifampicin) (RMP), the key determinant for treatment failure is RMP resistance. Detection of RMP resistance has thus been proposed as a proxy for MDR TB diagnosis, as well as for epidemiological monitoring (14,20,24). RMP drug susceptibility testing (DST), by conventional methods based on growth as well as by newer genetic techniques, is generally considered the most reliable (1, 8). Highly consistent results were obtained during the early proficiency testing (PT) rounds among the Supra-National TB Reference Laboratories (SRLS) of the World Health Organization (WHO)/International Union against Tuberculosis and Lung Disease network. Consequently, Laszlo et al. proposed a 99% efficiency target for RMP DST by the SRLs (8). However, 15 of 240 quality control strains (6.2%) distributed from 1999 to 2007 yielded less than 80% agreement for RMP resistance among the SRLs, insufficient for a judicial result. The panels were designed to contain approximately 50% resistance to all first-line TB drugs in various combinations. This precondition resulted in overrepresentation of rare profiles. The SRLs employed one of the four recogni...

Assessment of the GenoType MTBDRplus assay for rifampin and isoniazid resistance detection on sputum samples in Cote d'Ivoire

“…These cases of false-susceptibility results were already evocated [23,24] in particular with dysgonic M. tuberculosis strains. Probably, the proportion method in solid medium used as a gold standard would have given better results.…”

Rapid molecular TB diagnosis: evidence, policy making and global implementation of Xpert MTB/RIF

“…DOI: 10.1183/09031936.00157212methods, particularly the automated broth-based systems [97]. Sequencing, albeit limited, has largely resolved discordant results in favour of Xpert MTB/RIF, although a few truly false-positive results have been reported [17,18,84].…”

Rifampin Heteroresistance in Mycobacterium tuberculosis Cultures as Detected by Phenotypic and Genotypic Drug Susceptibility Test Methods

“…For example, 314 selected strains were retrospectively sequenced for resistance mutations in various genes, including rpoB, and compared with previous DST results (17). In the other study (18), discordant rifampin DST results in a proficiency panel were analyzed. In future heteroresistance prevalence studies, the analyses need to be done on the primary samples or primary culture.…”