β-Hemolysin, not mutation, inhibits the hemolysis of α-hemolysin in laboratory and clinical strains.

UNLABELLED

produces various hemolysins regulated by the Agr-QS system, except β-hemolysin encoded by the gene . A classical laboratory strain RN4220 displays only the β-hemolysin phenotype. It was suspected that the 8A mutation at the end of its gene delayed the expressions of and , then failed to express α- and δ-hemolysins. However, gene expression was detected at the later culture time without α-hemolysin phenotype, the reason for such a phenotype has not been clearly understood. We created knockout and complementary mutants via homologous recombination in RN4220 and NRS049, two strains that normally produce β-hemolysin and carry mutation. We found interestingly that the presence or absence of α-hemolysin phenotype in such strains depended on the expression of β-hemolysin instead of mutations, which only inhibited δ-hemolysin expression. The hemolysis phenotype was verified by the Christie-Atkinson-Munch-Peterson (CAMP) test. Quantitative reverse transcription PCR was carried out to evaluate the relative gene expressions of , , and . The construction of mutants did not affect the mutation status. We demonstrate that the absence of α-hemolysin in RN4220 and NRS049 strains is attributed to their production of β-hemolysin instead of mutation. Our findings broaden the understanding of the molecular mechanisms that control hemolysin expression in that is crucial for the development of new therapeutic strategies to combat infections.

IMPORTANCE

α-Hemolysin is a critical virulence factor in and its expression is largely controlled by the Agr-QS system. Nonetheless, the hemolysis phenotype and the regulation of the Agr-QS system in still hold many mysteries. Our study finds that it is the expression of β- hemolysin rather than the mutation that inhibits the function of the α-hemolysin in an important strain RN4220 and a clinical strain presents a similar phenotype, which clarifies the misunderstood hemolytic phenotype and mechanism of . Our findings highlight the interactions among different toxins and their biological roles, combined with QS system regulation, which is ultimately the true underlying cause of its virulence. This emphasizes the importance of considering the collaborative action of various factors in the infection process caused by this significant human pathogen.