Rebecca Wattam 07-08-2004 0.83 The methods of cell invasion and bacterial multiplication within vertebrate cells are described for the family Rickettsiaceae, which includes the three tribes Rickettsieae, Ehrlichieae, and Wolibachieae. Specifically, this interaction deals with Rickettsia prowazekii. TEXT TEXT TEXT TEXT wattam@vbi.vt.edu TEXT

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Adhesin TEXT Extracellular Ligand binding or carrier >Evidence that rickettsial outer membrane protein A (OmpA) is an adhesin of R. rickettsii and that OmpA and OmpB are adhesins of R. japonica has been documented. Rickettsiae attach to a protein-dependent receptor on the host cell membrane and induce focal host cell cytoskeletal rearrangements at the site of attachment, resulting in their entry into the host cell even in nonprofessional phagocytes by a mechanism requiring rickettsial metabolic activity (Walker, et al., 2003). Protein-dependent Receptor TEXT Cell membrane Ligand binding or carrier Rickettsia attach to a protein dependent receptor on the host cell membrane and induce focal host cell cytoskeletal rearrangements at the site of attachment, resulting in their entry into the host cell even in nonprofessional phagocytes by a mechanism requiring rickettsial metabolic activity (Walker et al., 2003). Important unidentified elements of the rickettsia-host cell interaction include the host cell membrane receptor for the rickettsial adhesin(s) (Walker et al., 2003). Rickettsia in Phagosome TEXT Phagosome Other The entry of rickettsiae into eukaryotic cells is mediated by an induced phagocytosis, but rickettsiae have never been observed in a closed phagocytic vacuole. In this study, Rickettsia conorii entry into Vero cells was observed by transmission electron microscopy during a period of 3 to 20 min after bacterium-cell contact. The entry occurred within 3 min after bacterium-cell contact, and R. conorii was observed in the process of engulfment, within a phagocytic vacuole, or free in the cytosol. Escape from the phagosome is a very rapid step since phagosome lysis was only occasionally observed. By 12 min, 90% of bacteria were internalized and half were free in the cytosol. This report confirms that rickettsiae penetrate nonphagocytic cells by induced phagocytosis and is the first demonstration of rickettsiae within a complete phagocytic vacuole (Teysseire et al., 1995). Internalization of obligate intracellular bacteria belonging to the genus Rickettsia by eukaryotic cells requires participation of both the parasitized host and the microorganism. The term "induced phagocytosis" has been used specifically to describe the entry of Rickettsia prowazekii, although a similar mechanism is likely for R. rickettsii. A role for a phospholipase in the internalization process has been proposed for both of these organisms, with the strongest supporting evidence provided for R. prowazekii. Despite general acceptance of the notion that phospholipase activity is involved in the internalization process of these bacteria, the origin of the enzyme is not known (Silverman et al., 1992). Rickettsia in Cytoplasm TEXT Cytoplasm Other Rickettsia rapidly lyse the phagosomal membrane and escape into the cytosol prior to phagolysomal fusion avoiding exposure to the lysosomal enzymes. In the cytosol they acquire their nutrients (e.g., glutamate), a part of their energy requirements (ADP/ATP transporter), and many components required for growth (e.g., amino acids) (Walker et al., 2003). Whereas R. rickettsii rarely grows to number greater than 100 organisms per cell before lysis occurs, R. prowazekii will achieve levels of greater than 800 organisms per cell (Hackstadt, 1996). Extracellular Rickettsia TEXT Extracellular Other During the exponential growth of R. prowazekii, very few bacteria exit from the infected cell to set up secondary infections in neighboring cells. Instead, R. prowazekii grows and fills the cytoplasm until, presumably, the host can no longer support the growth of the parasite. The host cell then bursts and a bolus of hundreds of R. prowazekii organisms are released to initiate infections in many new host cells. The mechanism by which R. prowazekii exits from the host cell as a burst, late in infection, has not been elucidated (Winkler and Daugherty, 1989). The typhus-group rickettsia do not produce such dramatic effects on the host cell and characteristically accumulate to very large numbers within the cytoplasm until lysis occurs. The ultrastructural damage seen in R. rickettsii-infected cells is not observed in R. prowazekii-infected cells (Hackstadt, 1996). TEXT Rickettsia attach to a protein dependent receptor on the host cell membrane and induce focal host cell cytoskeletal rearrangements at the site of attachment, resulting in their entry into the host cell even in nonprofessional phagocytes by a mechanism requiring rickettsial metabolic activity (Walker et al.,2003). Rickettsial invasion of susceptible host cells is an active process that requires active participation of both the host cell and the rickettsiae. Starved rickettsiae or rickettsiae inhibited by heat, formalin, KCN, 2,4-dinitrophenol, N-ethylmaleimide,or UV-irradiation do not penetrate cells efficiently or even adhere as well. Similarly, inhibition of the host cell by NaF or N-ethylmaleimide treatment or inhibition of actin polymerization by cytochalasin B or D, greatly reduced rickettsial entry, although the rickettsiae were able to adhere to the treated cells. The requirement for active involvement of both the parasite and the host has lead to this process being termed induced phagocytosis, or parasite-directed endocytosis (Hackstadt, 1996). TEXT The genus Rickettsia differs from most obligate and facultative intracellular parasites in that rickettsia grow in the cytoplasm of their host unbounded by phagosomal or phagolysosomal membranes. Walker and Winkler demonstrated that successful entry of R. prowazekii into L cells requires that rickettsiae be metabolically active and the host cells be capable of phagocytosis, even though they are not professionals phagocytes. Similar results were observed much earlier by Cohn et al. working with R. tustusgamushi. These observations suggest that rickettsiae induce their own phagocytosis by host cells, but they leave unexplained the mechanisms by which rickettsia escape from the phagosomes to the cytoplasm (Winkler and Miller, 1982). In this study we found that the pospholipase A activity can also be observed in the interaction of rickettsiae and competent host cells. We hypothesize that this activity may be the basis of one or more of the following: The induction of phagocytosis of rickettsiae, the lysis and escape from phagosomes by rickettsiae, and the rupture of the host cell membrane at the termination of infection of one cell, or allowing the egress of rickettsiae to infect other cells (Winkler and Miller, 1982). TEXT In this study we found that the pospholipase A activity can also be observed in the interaction of rickettsiae and competent host cells. We hypothesize that this activity may be the basis of one or more of the following. The induction of phagocytosis of rickettsiae, the lysis and escape from phagosomes by rickettsiae, and the rupture of the host cell membrane at the termination of infection of one cell, allowing the egress of rickettsiae to infect other cells (Winkler and Miller, 1982). It seems reasonable to postulate that either the activity responsible for the burst is induced only late in infection or the burst is due to the accumulation of damage to the host cell during the entire course of the infection, an accumulation that eventually overcomes the ability of the host cell to repair such damage (Winkler and Daugherty, 1989). TEXT TEXT TEXT TEXT Walker DH, Valbuena GA, Olano JP. Ann N Y Acad Sci. 2003 990 1 11 Teysseire N, Boudier JA, Raoult D. Infect Immun. 1995 63 1 366 374 Silverman DJ, Santucci LA, Meyers N, Sekeyova Z. Infect Immun 1992 60 7 2733 2740 Hackstadt T. Infect Agents Dis 1996 5 3 127 143 Winkler HH, Miller ET. Infect Immun. 1982 38 1 109 113 Winkler HH, Daugherty RM. Infect Immun. 1989 57 1 36 40 Gaywee J, Radulovic S, Higgins JA, Azad AF. Infect Immun. 2002 70 11 6346 6354 TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT TEXT

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