Changes in immunoferritin labeling of Rickettsia tsutsugamushi after serial cultivation in 60Co-irradiated BHK cells.

Abstract

The immunolabeling characteristics of Rickettsia tsutsugamushi (Gilliam strain) were examined by using a purified immunoglobulin G fraction of antibody to R. tsutsugamushi raised in rabbits. Formalin-fixed rickettsiae were reacted with this antibody and then with ferritin-conjugated goat anti-rabbit Fc antibody. R. tsutsugamushi cultivated in yolk sacs was used to raise antibody for this study. When rickettsiae in BHK-21 cells infected from yolk sac seed material were immunoferritin labeled, the binding of ferritin was found to be dense and uniform on the outer surface of the rickettsiae in disrupted host cells. Immunolabeling of purified suspensions of extracellular rickettsiae resulted in the uniform ferritin labeling of the microorganism. Aggregation of these rickettsiae by antibody appeared to depend upon the purity of the pellets. Immunoferritin labeling examined at high magnification revealed ferritin very close to the outer dense leaflet of the outer membrane. On some rickettsiae or on focal sites of others, the labelin; was several ferritin particles thick, suggesting the presence of a thick coating. The immunoferritin labeling of R. tsutsugamushi during successive serial passages in BHK-21 cells revealed decreased labeling with each passage, and by the 10th passage there was no detectable labeling. However, these rickettsiae inoculated back into yolk sacs regained their immunoferritin labeling. R. tsutsugamushi passed back into yolk sacs after four serial propagations in BHK-21 cells regained their labeling on the first passage in yolk sacs. However, rickettsiae from the 20th serial passage in BHK-21 cells required five passages in yolk sacs to reestablish their previous labeling affinity. Rickettsiae which did not label after 20 passages in BHK cells regained some of their labeling characteristics when sonicated. Antibody against rickettsiae cultivated in BHK-21 cells continued labeling rickettsiae even after 9 serial passages in BHK-21 cells.