Investigation on virulence factor profile and bacteriophage detection of methicillin-resistant Staphylococcus aureus from clinical isolates in Baghdad

Any part of the body can be infected with Staphylococcus aureus, including the skin, soft tissues, lungs, urinary tract and bones. A staph infection can cause opportunistic infections, sepsis and septic shock. Methicillin-resistant S. aureus is an important pathogen associated with human cases. Methicillin-resistant S. aureus (MRSA) is known to live in the noses, skin and gastrointestinal tract of humans. Colonization increases the risk of MRSA infections. In this study. Different clinical samples were collected from patients attending private laboratories and the National Center for Teaching Laboratory (NCTL), Baghdad Teaching Hospital and Gazzi Al-Harrii Hospital. A total of 200 different clinical samples (wound swabs, urine, burn, ear, sputum) were collected from both sexes. Identification of S. aureus from clinical samples was determined by culture and biochemical tests, followed by confirmation with the VITEK-2 compact system. The results showed that S. auerus was most recovered from wound and urine samples, 96/200 (48.0%) and 32/200 (16.0%), respectively. 50/200 (25%) of S. auerus were confirmed to be MRSA by testing the isolates against CLSI recommended oxacillin (1µg) and cefoxitin (30µg) using the disk diffusion method. These 50 MRSA isolates showed high susceptibility to Gentamicin (88%) and Rifampin (72.5%) and resistance to Penicillin (100%), Azithromycin (80.4%) and Erythromycin (80.4%). Regarding phenotyping resistance to macrolide antibiotic groups, only 10/50 (20%) isolates were S-phenotype, 13/50 (26%) isolates were R-phenotype and 27/50 (54%) isolates were D-phenotype. Biofilm detection of MRSA was performed using Congo red agar (CRA), tube method and microtiter plate. The results of the CRA method showed that 35/50 (70%) MRSA were biofilm producers compared to 18/50 (36%) using the tube method. Using the microtiter plate method, only 8/50 (18%) isolates were classified as strong biofilm producers, 14/50 (28%) as moderate biofilm producers, 12/50 (24%) as weak biofilm producers and 16/50 (32%) as non-biofilm producers. The results of this study also reported a non-significant difference between the antibiotic susceptibility status of Pencillin and Cephalosporins antibiotics against MRSA isolates (N=50) and biofilm formation score. The antimicrobial effect of Cyprus rotundus, Oreganum vulgare and Syzygium aromaticum oil was evaluated using disk diffusion method and microtiter plate iii method for (N=21) MRSA isolates. In the disk diffusion methods at 3 concentrations of 100%, 50% and 25%, the results showed an increase in zone inhibition diameters (ZIDs) with increasing dilution of cyperus and clove oil and highly significant differences between the three concentrations (P-value≤0.0001). In the microtiter plate methods, the results between 50% and 0.3% concentrations were highly significant (p-value≤0.0001) at all concentrations. Regarding the effect of the plant extract on MRSA isolates (n=8), the results of the disk diffusion and microtiter plate methods were highly significant (p value≤0.0001) at 100% and 50% concentrations in the disk diffusion method, and highly significant (p-value≤0.0001) at all concentrations (50%-0.3%) in the microtiter plate methods. All MRSA isolates were screened for harboring mecA, blaDHA-1 genes. The results showed that 50/50 (100%) of all MRSA isolates carried the mecA gene, while only 17/50 isolates carried the blaDHA-1 gene. Considering phenotyping resistance to macrolide antibiotic groups, 10/50 (20%), 13/50 (26%), 27/50 (54%) MRSA isolates showed S, R, D phenotyping, respectively. Only 6 isolates from each group (S, R, D) were phenotyped for genotyping detection. ermA, B, C genes showed high prevalence in three MRSA isolate groups. Regarding biofilm formation, only 8 out of 50 MRSA isolates were assessed as strong biofilm producers, harboring five genes (fnbA, fnbB) and icaA, C and D. All MRSA isolates (N=50) were subjected to the presence of phages using the plaque test method. The results showed that none of the MRSA isolates could carry phages.