The influence of temperature and pH on the growth of Rickettsia conorii in irradiated mammalian cells.

The temperature range for optimum growth of Rickettsia conorii in suspension culture of gamma-irradiated L cells was 32-38 degrees C, resulting in rickettsial doubling times between 4.1 and 6.0 hr. An asynchronous release of R. conorii from host cells was suggested by the constant increase in percent cells infected over a 36 hr period. Rickettsial growth was optimal at neutral to slightly alkaline extracellular pH levels. A moderately acidic pH, however, resulted in an increase in doubling time from 4.1 to 7.8 hr.

Plaque assay and cloning of scrub typhus rickettsiae in irradiated L-929 cells.

It was demonstrated that gamma-irradiated L-929 cells support plaque formation by three strains of Rickettsia tsutsugamushi and representative species of the spotted fever and typhus group rickettsiae. Sensitivity of the plaque assay for detection of viable scrub typhus rickettsiae was similar to that achieved with intraperitoneal inoculation of random-bred mice. The concentration of irradiated cells and the temperature and length of incubation were all found to affect plaque size. A technique combining terminal dilution and plaque purification was used to obtain clones of three strains of scrub typhus rickettsiae.

Laboratory animal models for human scrub typhus.

A wide variety of animals have been utilized in an attempt to provide the information necessary to bring scrub typhus to the point where it is no longer a threat to man. The laboratory mouse is usually the animal of choice for the study of this disease. The discovery that certain strains of inbred mice are genetically resistant to Rickettsia tsutsugamushi, the agent of scrub typhus, has opened new avenues in the study of the immune response to the disease. The cynomolgus monkey, Macaca fascicularis, appears to be the best animal model for the study of scrub typhus as it occurs in humans and should be useful in the development of an efficacious vaccine.

A scanning and transmission electron microscopic examination of Rickettsia tsutsugamushi-infected human endothelial, MRC-5, and L-929 cells.

Monolayers of primary human endothelial cells were infected with the Karp strain of Rickettsia tsutsugamushi and examined by scanning and transmission electron microscopy. The results were compared with those obtained with similarly infected L-929 and MRC-5 cells and with uninfected cells of all three types. The rickettsiae grew to slightly higher titers in the human endothelial cells. Transmission electron microscopy revealed significant changes in the host cell organelles; a reduction in ribosome-coated endoplasmic reticulum and in Golgi activity, swelling of mitochondria, and an increase in vacuolation within the cytoplasm. Since human endothelial cells are known to retain their in vivo structural and functional qualities when cultured in vitro, it is likely that these effects are similar to those which occur during the infectious process in human scrub typhus.

Pathology of toxic death in mice following intravenous injection of Rickettsia tsutsugamushi strain gilliam: examination by light and scanning electron microscopy.

The pathologic manifestations of the toxic death elicited by the iv injection of mice with Rickettsia tsutsugamushi strain Gilliam were examined by light and scanning electron microscopic techniques to ascertain the cause of death. Light microscopic examinations of hematoxylin and eosin-stained sections of organs suggested the presence of fluid in the lungs; this was confirmed by an increase in the weight of the lungs of moribund mice. Scanning electron microscopy of blood cells demonstrated a marked crenation of erythrocytes from these mice. Death appears to be the result of shock brought on by vascular collapse secondary to plasma leakage.