I~ UTTERWORTH I N E M A N N Vaccine, VoL 13, No. 1, pp. 29-35, 1995 Copyright © 1994 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0264-410X/95 $10.00 + 0.00 Protection of guinea-pigs from experimental Rocky Mountain spotted fever by immunization with baculovirus-expressed Rickettsia rickettsii rOmpA protein John W. Sumner*, Kim G. Sims, Dana C. Jones and Burt E. Anderson Baculovirus recombinants that express the Rickettsia rickettsii rOmpA protein were constructed. Monoclonal antibodies (mAbs) against the rOmpA protein reacted with recombinant-infected Spodoptera frugiperda (Sf9) cells in indirect immunofluorescence assays. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of infected Sfl) cell lysates with a mAb against rOmpA showed that the recombinant- expressed rOmpA protein migrated slightly below rOmpA extracted from R. rickettsii. Guinea-pigs immunized with lysates of recombinant-infected Sfl) cells developed antibodies reactive with R. rickettsii and were protected against challenge, indicating that the baculovirus-expressed rOmpA protein could be useful in subunit vaccines and for studies of the immune response to R. rickettsii infection. Keywords: rOmpA protein; Rickettsia rickettsii; baculovirus expression system Rickettsia rickettsii is the aetiological agent that causes Rocky Mountain spotted fever (RMSF). From a public health viewpoint, RMSF is the most important rickettsial disease in the United States. In 1990, state health departments reported 603 cases of RMSF, a 7.6% increase over the previous year. The case-fatality ratio for persons with laboratory-confirmed RMSF was 5.2%. The disease is transmitted by the bite of an infected tick and is most prevalent in the South Atlantic and West South Central regionsL Prophylactic vaccination would be beneficial for persons who work in or visit tick-infected areas, and for laboratory workers occupationally exposed to R. rickettsii 2. Vaccines have been prepared from phenolized suspensions of infected ticks 3'4, and from formalin-inactivated R. rickettsii grown in either yolk sa--cs of hens' eggs 5 or chick embryo fibroblastr'7. These vaccines were thought to be effective based on data from animal protection studies and field experience, but were subsequently shown to be only partially effective for human protection and were withdrawn from the market 8-~°. A commercial RMSF vaccine has not been available since 1978. More recently, efforts have focused Viral and Rickettsial Zoonoses Branch, Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, 16130 Clifton Road, Mailstop G13, Atlanta, GA 30333, USA. *To whom correspondence should be addressed. (Received 17 December 1993; revised 25 April 1994; accepted 25 April 1994) on vaccination with less virulent live rickettsia or with recombinant-expressed rickettsial proteins. Walker et al.z vaccinated guinea-pigs with R. conorii, a closely related but less virulent rickettsia, and Gage and Jerrells 11 used the more distantly related R. rhipicephali. In each case, the guinea-pigs were protected against challenge with R. rickettsiL The guinea-pig is considered the best animal model for protection experiments because they develop a clinical illness similar to RMSF among humans. However, success in the guinea-pig model is not equivalent to success among humans. The phenol- or formalin-killed vaccines previously mentioned protected or partially protected guinea-pigs, but failed to adequately protect humans. Two R. rickettsii surface proteins are protective antigens 12'13, indicating that they may be useful for the production of subunit vaccines. These proteins, the 190kDa antigen and the 135kDa antigen, are now referred to as the rOmpA and rOmpB proteins 14'~5, respectively, in order to avoid confusion caused by different estimates of apparent molecular mass determined by relative migration in gel electrophoresis. The rOmpA antigen gene was partially cloned by McDonald et al. 16. A truncated version of the protein was expressed in Escherichia coli and was shown to partially protect guinea-pigs from challenge with R. rickettsii ~7. By determining the nucleotide sequence of the same clone, Anderson et al.~ 8 concluded that the 5' end of the coding sequence was missing, that portion was cloned and a Vaccine 1995 Volume 13 Number 1 29