Identification and molecular analysis of the gene encoding Rickettsia typhi hemolysin

Rickettsia typhi, the causative agent of murine typhus, grows directly with in the host cell cytoplasm, accumulating a large number of progeny, and eve ntually lyses the cells, Typhus group rickettsiae (R. typhi and R. prowazek ii) adhere to and lyse human and sheep erythrocytes, However, the molecular mechanism underlying erythrocyte lysis by R. typhi has not been defined, H ere we describe the cloning and nucleotide sequence analysis of the gene (t lyC) encoding a hemolysin from R, typhi. DNA sequence analysis of R. typhi tlyC revealed an open reading frame of 912 bp, which encodes a protein of 3 04 amino acids with a predicted molecular mass of 38 kDa, To associate the R typhi tlyC gene product with hemolytic activity, we performed complementa tion studies with hemolysin-negative Proteus mirabilis WPM111 (a HpmA(-) mu tant of BA6163) transformed with R. typhi tlyC or R. typhi GFPuv-tlyC const ructs. We demonstrated that the cloned tlyC gene conferred a hemolytic phen otype on an otherwise nonhemolytic mutant of P. mirabilis. The availability of the cloned R. typhi tlyC will permit further characterization and defin ition of its role in rickettsial virulence.