Detection of Rickettsia prowazekii in body lice and their feces by using monoclonal antibodies

Abstract

In order to identify Rickettsia prowazekii in lice, we developed a panel of 29 representative monoclonal antibodies selected from 187 positive hybridomas made by fusing splenocytes of immunized mice with SP2/0-Ag14 myeloma cells. Immunoblotting revealed that 15 monoclonal antibodies reacted with the lipopolysaccharide-like (LPS-L) antigen and 14 reacted with the epitopes of a 120-kDa protein. Only typhus group rickettsiae reacted with the monoclonal antibodies against LPS-L. R. felis, a recently identified rickettsial species, did not react with these monoclonal antibodies, confirming that it is not antigenically related to the typhus group. Monoclonal antibodies against the 120-kDa protein were highly specific for R. prowazekii. We successfully applied a selected monoclonal antibody against the 120-kDa protein to detect by immunofluorescence assay R. prowazekii in smears from 56 wild and laboratory lice, as well as in 10 samples of louse feces infected or not infected with the organism. We have developed a simple, practical, and specific diagnostic assay for clinical specimens and large-scale epidemiological surveys with a sensitivity of 91%. These monoclonal antibodies could be added to the rickettsial diagnostic panel and be used to differentiate R. prowazekii from other rickettsial species.

FIG. 1.

Western blotting profiles of MAbs against R. prowazekii. Lane 1, molecular size markers (numbers on the left are in kilodaltons); lane 2, Coomassie brilliant blue-stained, native R. prowazekii antigen (SDS-PAGE); lanes 3, 5, and 7, MAb P11A12; lanes 4, 6, and 8, MAb P9G1; lane 9, polyclonal antisera against native antigen (lanes 3, 4, and 9), heated antigen (lanes 5 and 6), and proteinase K-digested antigen (lanes 7 and 8).

FIG. 2.

Fluorescent antibody-stained isolate of R. prowazekii. (a) Feces from an uninfected louse; (b) feces of an experimentally infected louse; (c) cell smear prepared from intestines of an experimentally infected louse.