Genetic analysis of isolates of Rickettsia rickettsii that differ in virulence

We examined the degree of intraspecies genetic polymorphisms present among 13 R. rickettsii strains isolated from the blood of patients and ixodid ticks from North and South America. Preliminary confirmation of these isolates as R. rickettsii was done by evaluating RsaI and PstI restriction fragment length polymorphic (RFLP) sites in a 631 bp fragment of the rOmpA gene amplified with primers encompassing the Rr190.70-701 nt region. All isolates had the same profile, which was identical to that previously described for strain Bitterroot. The AluI RFLP analysis of the rOmpA fragment differentiated only Hlp#2 isolate from the other strains. Two types of RFLP patterns were obtained for a 381 bp gltA fragment. Strains Bitterroot, Sheila Smith, Lost Horse Canyon, and Morgan contained an additional AluI fragment of 40 bp, compared to the four-band profile of 122, 87, 81, and 43 bp determined for other isolates. These differences correlated with the absence of a CGC insert in the former isolates as determined by comparative DNA sequence analysis. To measure the levels of deletion/mutation events among isolates of R. rickettsii, DNA sequence analysis was performed on a portion of a 2,672 bp region spanning the 3'-end of polA to the 5'-end of dnaE. Except for Hlp#2 strain these isolates were highly conserved in the regions sequenced.