Drug susceptibility testing of Mycobacterium tuberculosis by two phenotypic methods

ABSTRACTForMycobacterium tuberculosis, phenotypic methods for drug susceptibility testing of second-line drugs are poorly standardized and technically challenging. The Sensititre MYCOTB MIC plate (MYCOTB) is a microtiter plate containing lyophilized antibiotics and configured for determination of MICs to first- and second-line antituberculosis drugs. To evaluate the performance of MYCOTB forM. tuberculosisdrug susceptibility testing using the Middlebrook 7H10 agar proportion method (APM) as the comparator, we conducted a two-site study using archivedM. tuberculosisisolates from Uganda and the Republic of Korea. Thawed isolates were subcultured, and dilutions were inoculated into MYCOTB wells and onto 7H10 agar. MYCOTB results were read at days 7, 10, 14, and 21; APM results were read at 21 days. A total of 222 isolates provided results on both platforms. By APM, 106/222 (47.7%) of isolates were resistant to at least isoniazid and rifampin. Agreement between MYCOTB and APM with respect to susceptibility or resistance was ≥92% for 7 of 12 drugs when a strict definition was used and ≥96% for 10 of 12 drugs when agreement was defined by allowing a ± one-well range of dilutions around the APM critical concentration. For ethambutol, agreement was 80% to 81%. For moxifloxacin, agreement was 83% to 85%; incorporating existing DNA sequencing information for discrepant analysis raised agreement to 91% to 96%. For MYCOTB, the median time to plate interpretation was 10 days and interreader agreement was ≥95% for all drugs. MYCOTB provided reliable results forM. tuberculosissusceptibility testing of first- and second-line drugs except ethambutol, and results were available sooner than those determined by APM.

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Comparison of Proportion and Resistance Ratio Methods for Drug Susceptibility Testing of Mycobacterium tuberculosis Isolated from Patients Attending National Tuberculosis Centre, Nepal

SAARC Journal of Tuberculosis Lung Diseases and HIV/AIDS ◽

10.3126/saarctb.v5i1.3078 ◽

2010 ◽

Vol 5 (1) ◽

pp. 13-20

Author(s):

S Acharya ◽

P Ghimire ◽

DK Khadka ◽

S Nepali

Keyword(s):

Mycobacterium Tuberculosis ◽

Fluorescence Microscopy ◽

Susceptibility Testing ◽

Drug Susceptibility ◽

Drug Susceptibility Testing ◽

Resistance Ratio ◽

National Tuberculosis ◽

Smear Negative ◽

Culture Positive ◽

Good Agreement

Background: Tuberculosis (TB) is among the most serious infectious cause of global morbidity and mortality. Emergence of Multi-drug resistant tuberculosis (MDR-TB) is posing an increased threat to TB control programs. Drug susceptibility testing (DST) of Mycobacterium tuberculosis (M. tuberculosis) isolates is important for tackling such problems. Setting: National Tuberculosis Centre (NTC), Thimi, Bhaktapur, Nepal. Objectives: Comparative evaluation of two in vitro DST methods in determining susceptibility of M. tuberculosis isolates from patients attending NTC, to front-line anti-TB drugs: (Isoniazid-INH, Rifampicin-RFP, Streptomycin-SM, and Ethambutol-EMB). Methodology: This study was conducted from Sep 2006-Jun 2007. A total of 862 sputum samples (diagnosis or follow up cases) collected from patients (type of patients or their categories was not differentiated in this study) attending NTC bacteriology lab for sputum direct smear microscopy were analyzed using fluorescence microscopy. All smear positive samples, smear negative samples requested for culture were cultured. All culture positive samples confirmed as M. tuberculosis by biochemical tests were processed for DST by both proportion (PR) and resistance ratio (RR) methods. Results: Out of 862 sputum samples analyzed, 226 (26.2%) samples were positive for Acid Fast Bacilli (AFB) by fluorescence microscopy. Among 323 samples 226 smear positive samples and 97 smear negative samples requested for culture), 221 (68.4%) were culture positive, 92 (28.5%) were culture negative and 10 (3.1%) were contaminated. Out of 221 isolates of M. tuberculosis, 57.5% were resistant to one or more drugs by the PR method and 56.6% by the RR method. Similarly, MDR isolates were 29.9% and 29% by PR and RR methods respectively. On correlation analysis using Mc Nemar Chi-square test, no significant difference between the two tests were observed (p>0.05). The results showed high agreement between both methods and agreement rates to INH, RFP, SM and EMB were 93.2%, 93.7%, 93.2% and 94.1% respectively. Similarly, the agreement rates between both methods using kappa analysis showed kappa (k) value of 0.86, 0.85, 0.86 and 0.84 for INH, RFP, SM and EMB respectively, which is believed to be good agreement between both methods (k=0.80 to 1.00: Very good agreement). Conclusion: In conclusion, this study showed that both the Proportion and Resistance ratio methods are equally good for determining drug susceptibility of M. tuberculosis. Keywords: Mycobacterium tuberculosis; Drug Susceptibility Testing; Proportion Method; Resistance Ratio Method. DOI: 10.3126/saarctb.v5i1.3078 SAARC J. Tuber. Lung Dis. HIV/AIDS 2008 Vol.5(1) 13-20

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Comparison of the Performances of Two In-House Rapid Methods for Antitubercular Drug Susceptibility Testing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00960-08 ◽

2008 ◽

Vol 53 (2) ◽

pp. 808-810 ◽

Cited By ~ 6

Author(s):

Agustina I. de la Iglesia ◽

Emma J. Stella ◽

Héctor R. Morbidoni

Keyword(s):

Drug Resistance ◽

Mycobacterium Tuberculosis ◽

Sensitivity And Specificity ◽

Susceptibility Testing ◽

Drug Susceptibility ◽

Drug Susceptibility Testing ◽

Antitubercular Drug ◽

Rapid Methods ◽

Resistance Assessment

ABSTRACT Resistance to rifampin (rifampicin), isoniazid, and streptomycin of 69 Mycobacterium tuberculosis isolates was analyzed by an in-house method based on mycobacteriophage D29 and a colorimetric micromethod. Both methods showed sensitivity and specificity values ranging from 93% to 100%. These simple methods offer an option for drug resistance assessment of M. tuberculosis.

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