Nitrate Reductase Assay for Rapid Determination of Methicillin-Resistant Staphylococcus aureus Clinical Isolates.

OBJECTIVE: To evaluate the performance of nitrate reductase assay (NRA), a rapid, colorimetric method for the determination of methicillin resistance in Staphylococcus aureus isolates obtained from the culture collection of the Akdeniz University Hospital Central Laboratory, Antalya, Türkiye. MATERIALS AND METHODS: Identification for all 290 S aureus isolates at the species level was performed via matrix-assisted laser desorption/ionization-time of flight. Isolates were tested with NRA for methicillin resistance. The cefoxitin broth microdilution (BMD) method recommended by the Clinical and Laboratory Standards Institute was used as the reference method in the study. S aureus ATCC 29213 and S aureus ATCC 43300 strains were used for quality control. RESULTS: According to Food and Drug Administration criteria, the category agreement between NRA and BMD was found to be 100%. The essential agreement between both methods was determined to be 96.20%. There is no minor, major, or extremely major discrepancy between both methods. CONCLUSION: The results show that NRA is a rapid, practical, and reliable colorimetric method for detecting MRSA.

The Effect of Inoculum Size on Antimicrobial Susceptibility Testing of Mycobacterium tuberculosis.

Phenotypic drug susceptibility testing (DST) requires a standardized amount of inoculum to produce reproducible susceptibility results. The most critical step in the application of DST in Mycobacterium tuberculosis isolates is the preparation of the bacterial inoculum. In this study, the effect of bacterial inoculum prepared in various McFarland turbidities on primary antituberculosis drug susceptibility of M. tuberculosis strains was investigated. Five standard ATCC strains (ATCC 27294 [H37Rv], ATCC 35822 [izoniazid-resistant], ATCC 35838 [rifampicin-resistant], ATCC 35820 [streptomycin-resistant], ATCC 35837 [ethambutol-resistant]) were tested. Inoculums of McFarland standard of 0.5, 1, 2, 3, and 1:100 dilutions of 1 McFarland standard of each strain were used. The effect of inoculum size on DST results was determined by the proportion method in Lowenstein-Jensen (LJ) medium and nitrate reductase assay (NRA) in the LJ medium. In both test methods, the increase in inoculum size did not affect the DST results of the strains. On the contrary, DST results were obtained more rapidly as a result of the use of dense inoculum. DST results obtained in all McFarland turbidities were found to be 100% compatible with the recommended amount of inoculum, 1:100 dilution of 1 McFarland standard (inoculum size of gold standard method). In conclusion, the use of a high amount of inoculum did not change the drug susceptibility profile of tuberculosis bacilli. Minimizing manipulations during the inoculum preparation phase of susceptibility testing, this outcome will decrease the need for equipment and make the test application easier, particularly in developing countries. IMPORTANCE During DST application, it can be challenging to evenly homogenize TB cell clumps with lipid-rich cell walls. These experiments must be carried out under Biosafety Level-3 (BSL-3) laboratory conditions with personal protective equipment and taking safety precautions because the procedures applied at this stage cause the formation of bacillus-laden aerosols and carry a serious risk of transmission. Considering this situation, this stage is important given that it is not possible to establish a BSL-3 laboratory in poor and developing countries. Reducing the manipulations to be applied during the preparation of bacterial turbidity will minimize the risk of aerosol formation. Perhaps there will be no need to do these steps for susceptibility tests in these countries or even in developed countries.