Evaluation of the Sensititre MycoTB Plate for Susceptibility Testing of the Mycobacterium tuberculosis Complex against First- and Second-Line Agents

Abstract

bThe Sensititre MycoTB plate (TREK Diagnostic Systems, Cleveland, OH) uses a microtiter plate MIC format for susceptibility testing of Mycobacterium tuberculosis complex isolates against first-and second-line antituberculosis agents. Categorical agreement versus the agar proportion method for 122 M. tuberculosis complex isolates was 94% to 100%. T he gold standard method for Mycobacterium tuberculosis complex susceptibility testing is the 1% indirect agar proportion method (APM), which is laborious and requires 2 to 3 weeks for results (4, 7). This two-site study evaluated a new system, the Sensititre MycoTB plate (MycoTB), for susceptibility testing of M. tuberculosis complex. The plate uses a 96-well microtiter broth format and contains 12 lyophilized first-and second-line antimycobacterial drugs. In contrast to other M. tuberculosis complex susceptibility methods, which test one or two critical concentrations of a drug, the MycoTB plate examines a range of drug concentrations and produces an MIC result. Recently, Abuali et al. evaluated the MycoTB plate using 37 M. tuberculosis complex isolates (1). Our study confirms and extends these results by testing a larger number of isolates, including singly and multiply resistant isolates, and provides precision data for the MycoTB plate. We also compared a manual, mirror plate-reading method with a commercially available, manual plate reader with data management software (Vizion System, TREK Diagnostics).(This study was presented in part at the 50th Annual Interscience Conference on Antimicrobial Agents and Chemotherapy Boston, MA, 12 to 15 September 2010.)One hundred twenty-two M. tuberculosis complex isolates (59 from site 1 and 63 from site 2) were tested using the APM and the MycoTB plate. An additional 15 M. tuberculosis complex "challenge" strains with known susceptibility patterns were provided by the Centers for Disease Control and Prevention (CDC) and tested using the MycoTB plate. Inoculum preparation and plating were performed using a biological safety cabinet (BSC) inside a biosafety level 3 (BSL3) laboratory as previously described (5). Several colonies were selected from Middlebrook 7H10 medium using a sterile loop and inoculated into a test tube containing saline-Tween and glass beads (TREK Diagnostics). After being vortexed for 30 to 60 s, the inoculum was allowed to settle for 15 min and adjusted to a 0.5 McFarland standard equivalent using a nephelometer. The numbers of CFU were determined for each isolate tested to verify that the inoculum was within a specific targeted amount (ϳ10 5 CFU/ml). One hundred microliters of the inoculum was transferred to 11 ml of Middlebrook 7H9 broth containing oleic acid-albumin-dextrose-catalase (TREK Diagnostics) and vortexed for 20 s. One hundred microliters was transferred to the MycoTB plate wells containing the lyophilized antibiotics. MycoTB plates were covered with plastic seals provided by the manufacturer, and the entire outer surface of the plate was disinfected with a tuberculocidal agent. Plates were in...

Discordance across Several Methods for Drug Susceptibility Testing of Drug-Resistant Mycobacterium tuberculosis Isolates in a Single Laboratory

“…S1 in the supplemental material). For the MIC results, we used the breakpoints described by Hall, which utilize the breakpoint nearest to the 7H10 agar proportion critical concentration (8). We noted few differences whether we used the MIC breakpoints described by Hall or the empirical MIC breakpoint that optimized the Youden indices for INH, RIF, EMB, and STR (see Fig.…”

Plasma Drug Activity in Patients on Treatment for Multidrug-Resistant Tuberculosis

“…Susceptibility testing was repeated with a 1.0 McFarland suspension of newly cultured preparation according to the 1% proportion method using established critical concentrations for all drugs except pyrazinamide for which susceptibility testing was performed in pyrazinamide-specific Bactec MGIT media. MIC testing was performed for all drugs except pyrazinamide on MYCOTB Sensititre plates (TREK Diagnostics, Cleveland, OH) according to the manufacturer's protocol (11).…”

Sensititre MYCOTB MIC Plate for Testing Mycobacterium tuberculosis Susceptibility to First- and Second-Line Drugs

“…Notably, in our study, using the liquid MYCOTB method, the direction of discordance (driven mostly by isolates categorized as susceptible by MYCOTB and resistant by APM) was the same as that observed in the CDC program. In light of the findings of others and the fact that our (14,15). A limitation of our study was that we assessed MYCOTB reproducibility only for the 69/222 (31.1%) isolates for which one or more drugs had an absence of conditional (or categorical) agreement between MYCOTB and APM.…”

“…1). Previous studies comparing use of the MYCOTB plate to solid-medium drug susceptibility testing (DST) methods have shown good agreement between methods (14,15). However, there is little published information about test performance with a large and diverse isolate population that includes highly resistant M. tuberculosis isolates.…”