Penetration of Rickettsia tsutsugamushi into Cultured Mouse Fibroblasts (L Cells): An Electron Microscopic Observation

Abstract

The mechanism of penetration of purified Rickettsia tsutsugamushi (Gilliam strain) into cultured mouse fibroblasts (L cells) was examined by electron microscopy. After 10-40 min of infection, rickettsiae in the process of being phagocytized were often seen on the cell surface. These were restricted to the rickettsiae which seemed to be intact in morphology, while heavily plasmolyzed ones were never phagocytized. Additionally, rickettsiae were taken up individually into a phagosome, and phagocytosis of several rickettsiae together was rarely observed, except in the case of heat-inactivated microorganisms. In the cells, phagosomes whose membranes enclosed rickettsiae either tightly or loosely were seen. Rickettsiae in the loose phagosomes often showed signs of plasmolysis and were rarely released into the cell cytoplasm. Partial disintegration of phagosomal membranes and the escape of rickettsiae from the phagosomes were seen only in tight phagosomes. Large phagosomes containing a clump of several rickettsiae were observed occasionally, in which case the microorganisms were deformed and seemed to be denatured. From the above observations and the frequency of appearance of these different penetration stages in the specimens 10, 20, and 40 min after infection, it was concluded that the rickettsiae enter initially into a tight phagosome by phagocytosis and are then released into the cell cytoplasm by disruption of the phagosomal membrane. No other mechanisms of penetration were found. On the other hand, rickettsiae inactivated by trypsin did not attach to host cells. Inactivation by heat or UV irradiation resulted in reduction of phagocytosis, and rickettsiae treated with rifamycin could penetrate into the host cell cytoplasm to the same extent as in the case of infection with intact rickettsiae.Although rickettsiae are recognized as obligatory intracellular microparasites which propagate in the host cell cytoplasm, studies on the mechanism of penetration of this microorganism across the host cell membrane are scarce. Cohn et al (I) first showed that the entrance of Rickettsia tsutsugamushi into cultured mouse MB III lymphoblasts requires active participation of both the rickettsiae and the host, and a similar result was reported by Walker and Winkler (10) in the R. protoazekiimouse fibroblast (L cell) system. Wisseman et al (13) indicated rapid, bidirectional traffic of R. rickettsii directly across the host cell membrane. However Wisse-251

Intrasperm vertical symbiont transmission

“…For example, the biparental symbiont transmission may entail occasional mixing of different symbiont lineages, which would potentially lead to the evolution of some virulent phenotypes of the symbiont (29). Considering that some Rickettsia and allied pathogenic bacteria exhibit nuclear infections in somatic cells of their hosts (18)(19)(20)(21), it seems plausible, although speculative, that the Rickettsia symbiont of N. cincticeps has coopted the pathogenic nuclear infection mechanism for establishing the sperm head infection and enabling the efficient paternal transmission. The molecular and cellular mechanisms underlying the sperm head infection without disturbing sperm functioning are of not only basic but also applied interest, which would potentially provide insights into the development of sperm-mediated genetic transformation and/or material delivery technologies that have long been anticipated but not yet realized (30,31).…”

“…Intranuclear bacterial symbionts, such as Holospora and Caedibacter, have been relatively well-documented from unicellular ciliates (6,15), but reported only rarely from multicellular metazoans (16,17). In insects and other arthropods, intracellular Rickettsia and Orientia pathogens/symbionts are sometimes observed to localize not only to the cytoplasm, but also to the nucleus of the host cells (18)(19)(20)(21). In bathymodiolin mussels inhabiting hydrothermal vents and cold seeps, intranuclear bacterial parasites "Candidatus Endonucleobacter bathymodioli" have been described (16).…”

Isolation of Rickettsia tsutsugamushi Antigenically Different from Kato, Karp, and Gilliam Strains from Patients

“…Samples were fixed, dehydrated, and embedded by the method previously reported (16), and thin sections prepared by an LKB-IV ultra tomewere observed in a JEM-100CX electron microscope.…”

Rickettsia rickettsiiInfection of Cultured Human Endothelial Cells Induces Heme Oxygenase 1 Expression

“…Invasion of endothelial cells by rickettsiae depends on the active participation of both organisms and endothelial cells (63). Heat-or UVinactivated rickettsiae can bind to host cell surfaces but cannot penetrate the cells (60). To evaluate whether active rickettsia infection is a requisite for HO-1 mRNA induction, we utilized …”