IMMUNOCYTOCHEMICAL METHODS TO STUDY THE DISTRIBUTION OF ORIENTIA-TSUTSUGAMUSHI IN LEPTOTROMBIDIUM (ACARI, TROMBICULIDAE) CHIGGERS

Immunocytochemical methods were developed and tested for their ability to detect the distribution of Orientia tsutsugamushi in paraffin sect ions of adult chiggers (Leptotrombidium imphalum Vercammen-Grandjean & Langston). Rickettsial antigen was detected by application of a simpl e direct or amplified immunocytochemistry procedure and an indirect im munofluorescent procedure. In the direct procedure alkaline phosphatas e conjugation to the mouse polyclonal antibody to the Karp strain was followed by the HistoMark Red test system to detect rickettsial antige n. The amplification procedure used a similar method hut used an unlab eled primary antibody followed by secondary biotinylated antimouse IgG , streptavidin-alkaline phosphatase, and the HistoMark Red test system . The immunofluorescent procedure included a biotinylated secondary an tibody followed by addition of a streptavidin-FITC conjugate. Specific tissue tropisms in infected chiggers were observed in the salivary gl ands, nervous tissue, and ovaries of adult female mites in all procedu res; however, nonspecific fluorescence of the chigger limited definiti ve identification of tissue tropisms with the indirect immunofluoresce nt procedure.