Discordance across Phenotypic and Molecular Methods for Drug Susceptibility Testing of Drug-Resistant Mycobacterium tuberculosis Isolates in a Low TB Incidence Country.

Suhail Ahmad,Hanaa S Eldeen,Noura Al-Mutairi,Shirin Mohammadi,Eiman Mokaddas,Riccardo Manganelli

doi:10.1371/journal.pone.0153563

Suhail Ahmad, Hanaa S Eldeen + Show 4 more

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https://doi.org/10.1371/journal.pone.0153563

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Journal: PloS one	Publication Date: Apr 20, 2016
Citations: 51	License type: CC BY 4.0

Affiliation: Kuwait University, Kuwait Cancer Control Center

Abstract

With increasing incidence of multidrug-resistant tuberculosis (MDR-TB), accurate drug susceptibility testing (DST) of Mycobacterium tuberculosis to first-line drugs has become crucial for proper patient management. We evaluated concordance of DST results for 70 M. tuberculosis isolates across two phenotypic and two molecular methods: BACTEC 460TB, MGIT 960 system, GenoType MTBDRplus and DNA sequencing of gene segments most commonly implicated in conferring resistance to anti-TB drugs. Most (84%) M. tuberculosis isolates were multidrug-resistant. Twenty-four isolates yielded discrepant DST results. For rifampicin, isoniazid and streptomycin, 96%, 97% and 93% of isolates, respectively, were susceptible or resistant by all four methods, whereas for ethambutol, this agreement was observed for only 76% of isolates (P <0.05 for rifampicin or isoniazid or streptomycin versus ethambutol). Occurrence of rare mutations in three isolates that confer low-level resistance caused lower agreement for rifampicin among the four methods (kappa coefficient (κ) range, 0.84 to 0.95). For isoniazid, there was perfect agreement among phenotypic methods and molecular methods (κ, 1.00) but lower agreement between phenotypic and molecular methods. Three isolates were detected as polydrug-resistant by MGIT 960 system but as multidrug-resistant by DNA sequence-based method. The agreement was higher for streptomycin among the two phenotypic methods (κ, 0.97) while targeted sequencing yielded lower agreement (κ range, 0.86 to 0.89). The discrepancy for ethambutol resulted largely due to lower concordance of MGIT 960 results (κ range, 0.53 to 0.64). The MGIT 960 system is an accurate method for DST of M. tuberculosis against isoniazid and streptomycin while the results of rifampicin susceptibility should be complemented with DNA sequencing-based method when the suspicion for resistance is high. The possibility of false susceptibility to ethambutol with MGIT 960 system suggests that molecular or other phenotypic methods may be more useful when accurate ethambutol susceptibility results are warranted.

Highlights

Despite declining trends in the incidence of tuberculosis (TB) in recent years, the morbidity and mortality associated with TB is still enormous and drug-resistant TB is a growing problem worldwide
All isolates were identified as M. tuberculosis complex by the AccuProbe DNA probe assay as well as by the multiplex PCR assay based on specific amplification of two DNA fragments of ~473 bp and ~235 bp, as described previously [46, 48]
Based on drug susceptibility testing (DST) by the 460TB, 63, 59, 34 and 43 M. tuberculosis isolates were resistant to isoniazid, rifampicin, streptomycin and ethambutol, respectively

Summary

Introduction

Despite declining trends in the incidence of tuberculosis (TB) in recent years, the morbidity and mortality associated with TB is still enormous and drug-resistant TB is a growing problem worldwide. 3.3% of new cases and 20% of previously treated cases are identified as multidrug-resistant TB (MDR-TB, infection with Mycobacterium tuberculosis strain resistant at least to rifampicin and isoniazid, the two most effective first-line anti-TB drugs) [1,2,3]. MDR-TB is a risk factor for extensively drug-resistant TB (XDR-TB, defined as infection with MDR-TB strains resistant to a fluoroquinolone and injectable agent such as kanamycin, amikacin or capreomycin), a virtually untreatable disease in most of the developing countries [5,6,7]. Compared to the global average, much higher rates of MDR-TB (15.7% among new and 45.3% among previously treated TB cases) and XDR-TB (11.4% of all MDR-TB cases) have been reported from several countries in the European Region presenting new challenges for tuberculosis control [8,9,10]. Rapid and accurate laboratory diagnosis of MDR-TB is crucial for effective treatment which will limit transmission of MDR-TB and development of XDR-TB [3, 6, 11]

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