title: [Nitrate reductase assay for the rapid detection of Staphylococcus aureus methicillin resistance: a breakpoint susceptibility testing method].
abstract: Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of hospital- and community-acquired infections. Therefore rapid and accurate detection of MRSA is essential for infection control and prevention of nosocomial spread. In this study, the efficacy of a nitrate reductase assay (NRA) as a breakpoint susceptibility testing method was evaluated for the rapid detection of methicillin resistance in S.aureus A total of 135 S.aureus clinical isolates from our collection were tested for methicillin susceptibility by NRA breakpoint susceptibility method and by broth microdilution method. For NRA breakpoint susceptibility testing three tubes including growth control tube (without drug), test tube (with 4 mg/L cefoxitin) and lyophilized test tube (with 4 mg/L cefoxitin) were used. 50 µl of 0.5 McFarland bacterial suspension of each isolate was inoculated into the tubes. All tubes were incubated at 35ºC. After five-hour incubation, 500 µl of freshly prepared reagent [2 units of 0.2% sulfanilamide, 2 units of 0.1% N-(1-naphthyl) ethylenediamine dihydrochloride and 1 unit of concentrated hydrochloric acid] was added into each tube and a color change was watched for. The color changed to violet/purple, if there was bacterial growth. If the color changed to violet/purple in all three tubes, the isolate was identified as methicillin-resistant. If the color changed in growth control tube but not in the test and lyophilized tube, the isolate was identified as methicillin-susceptible. Among 135 isolates tested, 97 had mecA gene and were methicillin-resistant by both microdilution and NRA breakpoint susceptibility method. The remaining 38 clinical isolates did not have this gene and were susceptible to methicillin by both methods used. All results were concordant to the PCR which was considered as the gold standard method. Specificity, sensitivity, positive and negative predictive values were 100%. NRA breakpoint susceptibility test in tubes is an inexpensive and reproducible method. This method can easily be used in many laboratories and does not require skilled personnel. In addition, test tubes are prepared by lyophilisation to provide long shelf-life which gives an important advantage.
authors: Ondokuz Mayis University Faculty of Medicine, Medical Microbiology Department, Samsun, Turkey. cobanay2003@gmail.com.
authors: Ahmet Yılmaz
authors: AY
authors: Coban
authors: Uğur
authors: U
authors: Demirpek
authors: Alper
authors: A
authors: Ciftçi
authors: Bülent
authors: B
authors: Bozdoğan
mesh_terms: D003124:Colorimetry; D017714:Community-Acquired Infections / Q000175:diagnosis / Q000382:microbiology / Q000517:prevention & control; D003428:Cross Infection / Q000175:diagnosis / Q000382:microbiology / Q000517:prevention & control; D005612:Freeze Drying; D006801:Humans; D016106:Methicillin Resistance; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects / Q000302:isolation & purification*; D008826:Microbial Sensitivity Tests / Q000379:methods* / Q000592:standards; D050901:Nitrate Reductase; D013203:Staphylococcal Infections / Q000175:diagnosis / Q000382:microbiology* / Q000517:prevention & control
chemical_list: D050901:Nitrate Reductase