Evaluation of the antibacterial sensitivity properties of mannitol salt agar

Background: Staphylococcus spp. are major pathogens causing diverse clinical infections. Evaluating their phenotypic characteristics and antimicrobial response across different culture media is essential for accurate diagnosis. Objective: To assess vancomycin susceptibility testing performance of Mannitol Salt Agar (MSA) compared with Mueller–Hinton Agar (MHA) against Staphylococcus spp. Methods: Thirty clinical specimens (urine, wound, ear swabs) were collected. Isolates were identified using standard microbiological methods including MSA for colony color, blood agar for hemolysis, and VITEK-2 confirmation. Vancomycin susceptibility was evaluated on both media. Statistical analysis used t-test, chi-square, and Pearson correlation (p < 0.05). Results: Staphylococcus spp. were predominantly isolated from urine (60%; p = 0.007). On MSA, 40% showed yellow colonies and 60% pinkish-white (p = 0.27). Hemolysis patterns were 46.7% beta and 53.3% alpha/gamma (p > 0.05). Vancomycin produced significantly larger inhibition zones on MSA (25.3 ± 6.1 mm) than MHA (17.5 ± 5 mm; p < 0.0001). Strong correlation existed between colony color and hemolysis pattern (r = 0.873, p < 0.001). Conclusions: Vancomycin showed significantly larger inhibition zones on MSA compared to MHA, indicating that high salt content and mannitol alter antibiotic diffusion and/or bacterial phenotypic expression. Larger zones on MSA represent greater apparent susceptibility rather than enhanced resistance detection. Therefore, MSA cannot replace MHA for standardized antimicrobial susceptibility testing, as altered medium composition leads to misleading susceptibility profiles.