CHARACTERIZATION OF METHICILLIN - RESISTANT STAPHYLOCOCCUS AUREUS FROM ORTHOPAEDIC PATIENTS IN UNIVERSITY TEACHING HOSPITAL IN NIGERIA
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ABSTRACT
Methicillin resistant Staphylococcus aureus (MRSA) is now a threat to both the hospitalized patients and community globally. This work was aimed at detecting molecularly, methicillin resistant Staphylococcus aureus from the orthopaedic patients in Ahmadu Bello University Teaching Hospital, Zaria, Nigeria. Conventional biochemical methods were used to identify the isolates while API STAPH identification test kit further characterized the isolates to species level. The susceptibility test was carried out using disc agar diffusion method while beta – lactamase production was tested for using nitrocefin. Methicillin resistance was detected phenotypically using cefoxitin 30 µg disc and oxacillin agar screen test. Multiplex polymerase chain reaction (PCR) was used to detect mecA gene, the gene coding methicillin resistance and blaZ gene, the gene coding for beta- lactamase production with 16SrRNA gene being the internal control. Sequencing was carried out for the amplified isolates. A total number of 126 samples were collected from wound, skin and bed of orthopaedic patients for 5 months. With the conventional biochemical method of identification, 100(79.4%) isolates were identified as S. aureus while with the use of API STAPH identification test kit 39(39%) of the 100 were characterized as S. aureus. The susceptibility test of the S. aureus isolates showed that gentamicin had the greatest activity: 100%, 100% and 93.8% in wound, bed and skin respectively, followed by ciprofloxacin (100%, 94.1% and 93.8%) and pefloxacin (100%, 88.2% and 75%) respectively. However, the greatest level of resistance was observed with ampicillin: 100%, 100% and 87.5% in wound, bed and skin respectively followed by ceftriaxone: 100%, 76.5% and 75%; and amoxicillin - clavulanate: 66.7%, 58.8%, 56.2%. Phenotypic detection of MRSA with the use of cefoxitin disc diffusion gave a MRSA prevalence of 83.3%, 64.5% and 56.3% from wound, bed and skin viii respectively. These MRSA isolates were generally resistant to the beta lactam antibiotics used, while 11/25 (44%) were multi-drug resistant. However, vancomycin, gentamicin and ciprofloxacin were most active against the MRSA isolates, 15% of these phenotypic MRSA isolates were hyper-producers of beta-lactamase. The gold standard for detecting MRSA using polymerase chain reaction is detection of mecA gene and only 2 (5.1%) S. aureus isolates were positive. Fifteen (78.9%) of the phenotypic MRSA tested carried plasmid with molecular weight ranging from 9.23 to 13.27 kilobase pairs. The presence of plasmid and hyper- production of betalactamase can be suggested to be responsible for the phenotypic detection of MRSA observed in this study; 33.3% of the S. aureus isolates amplified blaZ gene. The nucleotide sequence of 16SrRNA gene of isolate S41 in comparison with those from GenBank database showed that the S. aureus isolate has 99% identity with Staphylococcus aureus strain KIBGE-MB01 with sequence ID (accession) number HM061132.1.
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