Analysis of the hemolysin and virulence-related genes of incomplete hemolysis Staphylococcus aureus

Objective To analyze hemolysin and virulence-related genes in incomplete hemolytic Staphylococcus aureus. Methods Fifty strains of incomplete hemolytic Staphylococcus aureus were isolated from patients admitted in the Second Affiliated Hospital of Soochow University during 2013 and 2014, and the isolates with complete hemolytic phenotype were also collected at the same period as the control strains. All the strains were inoculated and subcultured on four kinds of sheep blood agar plates supplied by different manufacturers to compare their hemolytic phenotype. The relative mRNA expressions of hemolysin genes (hla, hlb, hlc, hld) in standard strain, complete and incomplete hemolytic phenotype strains were detected by real-time quantitative polymerase chain reaction (RT-qPCR), and valued by 2-△△Ct method. t test was used to compare mRNA expressions of hemolysin genes. Western blot was performed to analyze the expression of α-hemolysin. Antibiotic susceptibility test of incomplete hemolytic strains was performed using broth microdilution method. Resistant gene mecA and virulence genes pvl, tst were detected by PCR. Results The steady and hereditary incomplete hemolysis was observed in 50 strains of incomplete hemolytic Staphylococcus aureus on the sheep blood agar plates from different suppliers. Taking mRNA expression of hla, hlb, hlc, hld in standard strain as 1, the relative mRNA expressions of hemolysin genes in incomplete hemolytic strains were 0.02, 7.51, 0.06 and 0.12 respectively, there were statistical differences between standard strain and incomplete hemolytic strains (t=8.46, -56.40, 8.12 and 7.61, all P<0.05). And the expression of α-hemolysin was decreased in incomplete hemolytic strains. All the strains were identified as methicillin resistant Staphylococcus aureus (MRSA). Three strains exhibited different minimum inhibitory concentrations of teicoplanin and linezolid after subcultured, but the differences had no impact on the final results of antibiotic susceptibility test. mecA, pvl and tst genes were positive in incomplete hemolytic strains. Conclusion Staphylococcus aureus with incomplete hemolytic phenotype is methicillin resistant with higher expression of β-hemolysin and lower expressions of α-hemolysin, γ-hemolysin and δ-hemolysin. It carries plv and tst virulence genes and is of high virulence. Key words: Staphylococcus aureus; Incomplete hemolysis; Hemolysin; Drug resistance; Virulence