Nonhemolysis of epidemic El Tor biotype strains of Vibrio cholerae is related to multiple functional deficiencies of hemolysin A

Background Hemolysis of bacteria is an important phenotype used for typing and characterizing strains with specific biomarkers and even a virulence factor in bacterial pathogenesis. In Vibrio cholerae , hemolysin HlyA is responsible for hemolysis of sheep red blood cells, and this hemolytic phenotype is used as a biotyping indicator and considered one of the virulence factors. At the beginning of the seventh cholera pandemic, the El Tor biotype strains of serogroup O1 were distinguished by hemolysis from the sixth pandemic O1 classical biotype strains, whereas during the following epidemics, nonhemolytic El Tor strains appeared, suggesting phenotypic and genetic variations in these strains. This study aimed to investigate the possible mechanisms involved in nonhemolysis of El Tor strains. Results Five sequence types of hlyA genes were found in the studied O1 El Tor strains isolated during the seventh pandemic. A 4-base deletion in hlyA caused the HlyA protein mutation and non-hemolytic phenotype. Some strains carry wildtype hlyA genes but are still non-hemolytic, and greatly reduced hlyA transcription and blocked secretion of hemolysin were observed in hemolysis tests of the subcellular components and transcription/expression analysis of hlyA . Conclusions Mechanisms responsible for nonhemolysis of the epidemic O1 El Tor strains are complex and not only confined to gene mutation but also deficiencies of transcription and extracellular transport of HlyA. Mutations in gene regulation and protein secretion systems of HlyA in the nonhemolytic V. cholerae strains should be areas of concern in future studies.