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PubMed1992

Demonstration and partial characterization of antigens of Rickettsia rhipicephali that induce cross-reactive cellular and humoral immune responses to Rickettsia rickettsii.

The relatively unrelated spotted fever group rickettsia Rickettsia rhipicephali conferred on guinea pigs protective immunity against challenge with virulent R. rickettsii. Immunity was conferred at all doses of R. rhipicephali used in the study. Because of the serologic unrelatedness of these two rickettsiae, determined by the use of microimmunofluorescence and other serological assays, further studies were performed to define the nature of the immune response elicited by R. rhipicephali and the characteristics of the rickettsial antigens that evoke cross-reactive antibody responses. Animals immune to R. rhipicephali tested at the time of challenge showed a complete cross-reactive lymphocyte proliferative response to rickettsial antigens prepared from each species. In fact, spleen cells from R. rhipicephali-immune animals responded better to R. rickettsii antigens than to homologous immunizing antigens. Serum samples were obtained from R. rhipicephali-infected animals at various times after infection and tested by the use of Western immunoblot assay for antibodies that were cross-reactive with antigens of R. rickettsii. By 10 days after infection with R. rhipicephali, antibodies to antigens of both species were noted, and by 37 days after infection, sera from immune animals showed strong reactivity to antigens of R. rhipicephali with apparent molecular masses of 107 and 151 kDa. The cross-reactive antibody response to antigens of R. rickettsii was relatively strong and involved predominantly the rOmpB protein and the rickettsial lipopolysaccharide. These findings establish the presence of T-cell-dependent epitopes associated with antigens of R. rhipicephali, which confer protective immunity against challenge with R. rickettsii. Results of Western immunoblot assays support the contention that the R. rickettsii rOmpB surface antigen contains important protective epitopes.

Animals↗

PubMed1997

Characterization of and application of monoclonal antibodies against Rickettsia africae, a newly recognized species of spotted fever group rickettsia.

Rickettsia africae is a newly described species which causes African tick bite fever. Mediterranean spotted fever caused by R. conorii is endemic in the same regions of Africa as tick bite fever, and differentiation of the two syndromes by characterization of their etiological agents is important for epidemiological studies. R. africae and R. conorii are, however, difficult to distinguish, and therefore, our aim was to produce monoclonal antibodies to address this problem. Monoclonal antibodies were produced against R. africae by fusing splenocytes from BALB/C mice immunized with purified rickettsial organisms and SP2/0-Ag14 myeloma cells. A total of 355 hybridomas producing monoclonal antibodies to R. africae were identified by initial screening with six different antigens by microimmunofluorescence assay. A panel of 23 representative monoclonal antibodies were selected and subcloned. This panel was screened with a further 17 different spotted fever group (SFG) rickettsial reference antigens. Of these 23 monoclonal antibodies, 1 cross-reacted with only R. parkeri, whereas the others cross-reacted with more than two different antigens. Immunoblotting indicated that all the monoclonal antibodies were directed against the epitopes on two major high-molecular-mass heat-labile proteins, of which the molecular masses were 128 and 135 kDa, respectively. This monoclonal antibody panel was used successfully to identify R. africae in the blood culture of an infected patient, in infected cells within shell vials, and in infected ticks collected from Africa. Furthermore, the cross-reactivity of each SFG rickettsia with each of these 23 monoclonal antibodies was scored and was used to build a dendrogram of taxonomic relatedness between R. africae and the other SFG rickettsiae on the basis of Jaccard coefficients and unweighted pair group method with arithmetic mean analysis. The relatedness was generally consistent with that obtained by other methods of comparison.

Africa↗

PubMed2003

Detection of antibodies against spotted fever group Rickettsia (SFGR), typhus group Rickettsia (TGR), and Coxiella burnetii in human febrile patients in the Philippines.

A total of 157 sera from febrile patients in the Philippine General Hospital in Manila, Luzon, and the Northern Samar Provincial Hospital, the Philippines, were used. Serum antibodies against spotted fever group Rickettsia (SFGR) and typhus group Rickettsia (TGR) were detected by indirect immunofluorescence test. Antibody positive rates were 1.3% for SFGR (Rickettsia japonica) and 2.5% for TGR (R. typhus), respectively. Rickettsial antibodies in humans in the Philippines were found for the first time. These results underscore the need for further epidemiological study of clinical rickettsioses in the Philippines.

Antibodies, Bacterial↗

PubMed2003

Antibody prevalence of Orientia tsutsugamushi, Rickettsia typhi and TT118 spotted fever group rickettsiae among Malaysian blood donors and febrile patients in the urban areas.

The seroprevalence of Orientia tsutsugamushi (OT), Rickettsia typhi (RT) and TT118 spotted fever group rickettsiae (SFGR) among blood donors and febrile Malaysian patients in the urban areas was determined. Of the 240 blood donors, 5.4%, 9.2% and 1.7% had either present or previous exposure to OT, RT and SFG rickettsiae, respectively. Patients admitted to an urban hospital had high seroprevalences of OT (43.5%) and RT (22.9%), as compared to SFGR (11.6%). Antibody levels suggestive of recent infections of scrub typhus, murine typhus and tick typhus were detected in 16.8%, 12.7% and 8.2% of patients respectively. No significant difference was noted in the distribution of rickettsial antibodies among urban patients from 2 geographical locations. However, the serologic patterns of rickettsial infection in the urban areas were different form those of rural areas.

Adolescent↗

PubMed1988

Clinical, hematologic, and humoral immune response in female dogs inoculated with Rickettsia rickettsii and Rickettsia montana.

Female Beagles were inoculated intradermally with a sublethal dose of Rickettsia rickettsii and R montana. Three dogs (group 1) were inoculated with 2 X 10(2) plaque-forming units (PFU) of R rickettsia and were treated with tetracycline beginning on postinoculation day (PID) 12; 3 dogs (group 2) were inoculated with 2 X 10(2) PFU of R rickettsii but were not treated; 3 dogs (group 3) were inoculated with 2 X 10(2) PFU of R montana. Group-3 dogs failed to seroconvert and were inoculated a second time on PID 68. Groups 1 and 2 dogs inoculated with R rickettsii became depressed and developed occasional inappetence, fever, hematochezia, and ocular lesions. These dogs had a decrease in PCV and RBC count, an initial decrease in WBC count followed by leukocytosis, and a decrease in platelet count. Group-3 dogs inoculated with R montana remained healthy. After R rickettsii inoculation, the serologic response to spotted fever group (SFG) rickettsial antigens (R rickettsii, R rhipicephali, R montana, and R bellii) was similar. The antibody response to R rickettsii was first detected on PID 9, with peak titers reached by PID 20. Serum titers to R rickettsii remained stable or decreased one dilution through PID 120. Of 4 SFG rickettsial antigens, the highest serologic response was to R rickettsii. A cross-reacting antibody response with R rhipicephali and R montana was nearly identical and was only slightly less than the response to R rickettsii. Cross-reacting antibodies to R belli were of lower mean titer and of shorter duration than were cross-reacting antibodies to other SFG rickettsiae.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1980

Sexual transmission of spotted fever group rickettsiae by infected male ticks: detection of rickettsiae in immature spermatozoa of Ixodes ricinus.

Electron microscopic evidence is provided showing that the newly described "Swiss agent," a spotted fever group rickettsia, is incorporated into the reproductive cells of its male tick vector, Ixodes ricinus. Rickettsiae were found in spermatogonia, spermatocytes, and maturing spermatids. The potential significance of these observations is briefly discussed in relation to published data on sexual transmission of rickettsiae by ticks.

Animals↗

PubMed1990

Selection of alpha/beta interferon- and gamma interferon-resistant rickettsiae by passage of Rickettsia prowazekii in L929 cells.

The ability of endogenously produced alpha/beta interferon (IFN-alpha/beta) to inhibit rickettsial growth in infected L929 cell cultures was evaluated by comparing the growth of Rickettsia prowazekii Madrid E in untreated cultures and cultures treated with anti-mouse IFN (alpha + beta) serum. The endogenously produced IFN was neutralized, and rickettsial growth was enhanced in the antiserum-treated cultures. This inhibitory effect of endogenously produced IFN-alpha/beta was used to select rickettsiae resistant to IFN-alpha/beta. Rickettsiae were screened for resistance to IFN-alpha/beta after being cultured in untreated L929 cells for several weeks to several months. Two isolates derived from R. prowazekii Madrid E and two isolates derived from plaque-purified R. prowazekii Madrid E were plaque-purified twice, grown in embryonated hen eggs, and evaluated for resistance to IFN-alpha/beta and IFN-gamma. Compared with the parental rickettsial strain, all four isolates were significantly resistant to IFN-alpha/beta and IFN-gamma. In addition, they were as resistant or more resistant to IFN-gamma when compared with two previously described IFN-gamma resistant isolates that were selected in IFN-gamma-treated L929 cells. One of the two isolates from IFN-gamma-treated L929 cells was also resistant to IFN-alpha/beta; the other isolate was similar to the parental Madrid E strain in sensitivity to IFN-alpha/beta.

Animals↗

PubMed1990

Secretion of tissue-type plasminogen activator and plasminogen activator inhibitor by Rickettsia conorii- and Rickettsia rickettsii-infected cultured endothelial cells.

Hemostasis abnormalities have been described in patients with Mediterranean spotted fever and Rocky Mountain spotted fever. Evidence of the activation of the fibrinolytic system has been obtained in both diseases. After experimental Rocky Mountain spotted fever, an elevated level of fibrinogen was found in parallel with the activation of the fibrinolytic system and transient elevation of the tissue-type plasminogen activator. Later protein is mainly synthesized by endothelial cells. The ability to culture human endothelial cells in vitro provides a unique system to study the secretion of tissue-type plasminogen activator and of plasminogen activator inhibitor after rickettsial infection. Human vascular endothelial cells derived from the umbilical vein, when infected with Rickettsia conorii or Rickettsia rickettsii, secreted as much tissue-type plasminogen activator as control cells. The activity of plasminogen activator inhibitor however, was higher in the supernatants of infected cells than in those of control cells. This rickettsia-induced imbalance of the tissue-type plasminogen activator-inhibitor pair was a very early event after in vitro infection. The involvement of this system during Mediterranean spotted fever and Rocky Mountain spotted fever remains to be demonstrated.

Antibodies, Monoclonal↗

PubMed1991

Comparative susceptibility to mouse interferons of Rickettsia tsutsugamushi strains with different virulence in mice and of Rickettsia rickettsii.

Three strains of Rickettsia tsutsugamushi (Karp, Gilliam, and TA716, representing three virulence types in mice) were examined for their sensitivity to the inhibitory effects of recombinant gamma interferon (IFN-gamma) and purified IFN-alpha/beta in two cultured mouse fibroblast cell lines. The susceptibilities of another species, Rickettsia rickettsii, and of encephalomyocarditis virus (EMCV) were also tested for comparative purposes. IFN-gamma inhibited rickettsial replication in only one of the six combinations of R. tsutsugamushi strains and mouse cells (strain Gilliam and the BALB/c mouse-derived cell line). In contrast, R. rickettsii and EMCV replication were markedly inhibited in both cell types, but to a greater extent in the BALB/c line than in the C3H cells. IFN-alpha/beta (300 to 450 U/ml) was uniformly ineffective in three of the combinations of R. tsutsugamushi strains and mouse cells (Gilliam in C3H cells and Karp in both C3H and BALB/c cells); in the remaining sets, IFN-alpha/beta-mediated inhibition of rickettsial replication was variable and in no case was it very pronounced. The tests with R. rickettsii in both cell types also indicated slight, variable sensitivity to IFN-alpha/beta. EMCV, on the other hand, was very susceptible to IFN-alpha/beta, confirming the potency of the preparation used; as with IFN-gamma, virus replication was inhibited to a greater degree in the BALB/c cell line than in the C3H cultures. These results are discussed in terms of their relationship to the virulence properties of the R. tsutsugamushi strains in BALB/c and C3H mice and to the known IFN-sensitivities of the more widely studied Rickettsia prowazekii.

3T3 Cells↗

PubMed1991

Tick-bite fever in Zimbabwe. Survey of antibodies to Rickettsia conorii in man and dogs, and of rickettsia-like organisms in dog ticks.

As part of a study of tick-bite fever in Zimbabwe, the prevalence of antibodies to Rickettsia conorii in dogs and humans was examined. Blood samples were obtained from 184 dogs in different parts of the country and tested by indirect immunofluorescence. In all, 150 (82%) were positive at a titre of 1/40 or higher. Dogs from the south and east of the country showed very high seroprevalence compared with dogs from the main urban centre, Harare. Human serum was obtained from blood donors, from people attending clinics with non-febrile illness and from agricultural workers. Antibodies to R. conorii were found in a high proportion of samples from the south and east, with 100% of blood donor samples in one area being positive. As with the dogs, a much lower proportion of samples from Harare were positive. The exact role of dogs and their ticks (Rhipicephalus sanguineus, R. simus and Haemaphysalis leachi) in the epidemiology of human tick-bite fever remains unclear, since dog ticks were seldom found to harbour rickettsia-like organisms and man is not one of their preferred hosts. While dogs may be of little importance in disease transmission, their value as sentinels of spotted-fever group rickettsias in the environment was confirmed.

Animals↗

PubMed1984

[Electron microscopic characteristics of the action of penicillin and vancomycin on Rickettsia conorii and Rickettsia akari in vitro].

The possibility of L-transformation for two species of Rickettsia of the tick group (R. conorii and R. akari) was studied with light and electron microscopy. The experiments were performed on the cultures of L-cells (murine fibroblasts). Penicillin and vancomycin were used as the transforming agents. Electron microscopy revealed clear changes in the morphological structure of both species when the doses of penicillin and vancomycin were 50-100 and 250-1500 micrograms/ml, respectively. The antibiotics had a morphologically analogous effect on both species of Rickettsia. The resulting morphological structures were large oval spheroplast-like cells with significant expansion of the periplasmic space up to 1 X 1.75 micron under the effect of penicillin and 0.5 X 0.7 micron under the effect of vancomycin. R. akari was somewhat more sensitive to the antibiotics. Electron microscopy of the morphological changes in the Rickettsia induced by penicillin and vancomycin showed that they were similar to the changes observed in gram-negative bacteria on their L-transformation and to the penicillin forms of chlamydia (galprovia) and R. prowazekii. They may be considered as a phase of unbalanced growth and represent the initial stage of L-transformation.

Cell Wall↗

PubMed1991

In vitro susceptibilities of Coxiella burnetii, Rickettsia rickettsii, and Rickettsia conorii to the fluoroquinolone sparfloxacin.

In vitro susceptibilities of Rickettsia rickettsii, Rickettsia conorii, and Coxiella burnetii to the new fluoroquinolone sparfloxacin (AT-4140; RP 64206) were determined. Plaque and dye uptake assays were used to measure the MICs against R. rickettsii and R. conorii. The susceptibilities of C. burnetii Nine Mile and Q 212 were determined in two acute-infection models and in two chronic-infection models. The MICs were 0.125 to 0.25 microgram/ml for R. rickettsii and 0.25 to 0.5 microgram/ml for R. conorii. Sparfloxacin (1 microgram/ml) cured cells recently infected with C. burnetii Nine Mile and Q 212 within 4 to 9 days and cured multiplying, persistently infected cells within 10 days. As previously described with other fluoroquinolones (D. Raoult, M. Drancourt, and G. Vestris, Antimicrob. Agents Chemother. 34:1512-1514, 1990), sparfloxacin failed to cure cells persistently infected with C. burnetii and blocked from dividing with cycloheximide. As determined by the dye uptake assay, no cellular toxicity was noted with sparfloxacin at up to 128 micrograms/ml. These results are consistent with those previously obtained with fluoroquinolones (D. Raoult, M. Yeaman, and O. Baca, Rev. Infect. Dis. 11[Suppl. 5]:S986, 1989), although sparfloxacin may be slightly more active.

4-Quinolones↗

PubMed2004

Complete genome sequence of Rickettsia typhi and comparison with sequences of other rickettsiae.

Rickettsia typhi, the causative agent of murine typhus, is an obligate intracellular bacterium with a life cycle involving both vertebrate and invertebrate hosts. Here we present the complete genome sequence of R. typhi (1,111,496 bp) and compare it to the two published rickettsial genome sequences: R. prowazekii and R. conorii. We identified 877 genes in R. typhi encoding 3 rRNAs, 33 tRNAs, 3 noncoding RNAs, and 838 proteins, 3 of which are frameshifts. In addition, we discovered more than 40 pseudogenes, including the entire cytochrome c oxidase system. The three rickettsial genomes share 775 genes: 23 are found only in R. prowazekii and R. typhi, 15 are found only in R. conorii and R. typhi, and 24 are unique to R. typhi. Although most of the genes are colinear, there is a 35-kb inversion in gene order, which is close to the replication terminus, in R. typhi, compared to R. prowazekii and R. conorii. In addition, we found a 124-kb R. typhi-specific inversion, starting 19 kb from the origin of replication, compared to R. prowazekii and R. conorii. Inversions in this region are also seen in the unpublished genome sequences of R. sibirica and R. rickettsii, indicating that this region is a hot spot for rearrangements. Genome comparisons also revealed a 12-kb insertion in the R. prowazekii genome, relative to R. typhi and R. conorii, which appears to have occurred after the typhus (R. prowazekii and R. typhi) and spotted fever (R. conorii) groups diverged. The three-way comparison allowed further in silico analysis of the SpoT split genes, leading us to propose that the stringent response system is still functional in these rickettsiae.

Chromosome Inversion↗

PubMed1987

Reactivity of monoclonal antibodies to Rickettsia rickettsii with spotted fever and typhus group rickettsiae.

Analysis of 15 spotted fever group (SFG) and 2 typhus group strains of rickettsiae with a panel of monoclonal antibodies revealed a number of shared and unique epitopes of the 120- and 155-kilodalton surface proteins. All of the SFG strains but neither of the typhus group strains reacted with antibody to the lipopolysaccharidelike antigen of Rickettsia rickettsii; possibly the lipopolysaccharidelike antigen is the common antigen which defines the SFG. North Carolina and Montana strains of R. rickettsii known to differ slightly in virulence for guinea pigs differed in at least one epitope of the 120-kilodalton protein.

Animals↗

PubMed1994

The role of tumor necrosis factor in host defense against scrub typhus rickettsiae. I. Inhibition of growth of Rickettsia tsutsugamushi, Karp strain, in cultured murine embryonic cells and macrophages by recombinant tumor necrosis factor-alpha.

Recombinant murine tumor necrosis factor-alpha (TNF-alpha) inhibited intracellular growth of Rickettsia tsutsugamushi, Karp strain, in the mouse embryo cell line C3H/10T1/2 clone 8 at doses of 100 to 10 U/ml. The growth inhibitory effect of TNF-alpha was also evident when peritoneal exudate macrophages or bone marrow-derived macrophages were used as the host cell for rickettsial growth. Interferon-gamma (IFN-gamma), at doses up to 1,000 U/ml, did not affect the growth of this strain of rickettsiae in the mouse embryo cell line but, as expected, profoundly inhibited rickettsial growth in peritoneal exudate macrophages and bone marrow-derived macrophages. The effect of TNF-alpha on rickettsial growth in the mouse embryo cell line was not reproducibly enhanced by IFN-gamma. Treatment of the cell line with TNF-alpha delayed rickettsial cytopathic effects, but the rickettsiae ultimately grew to high numbers in the cells and caused cell death. These findings show that, at least in our system, R. tsutsugamushi is resistant to IFN-gamma-mediated antirickettsial effects in cells other than macrophages. The results of this study support the suggestion that TNF-alpha may inhibit rickettsial growth in cells other than macrophages.

Animals↗

PubMed1990

Prevalence of antibodies to Rickettsia conorii Coxiella burnetii and Rickettsia typhi in Salamanca Province (Spain). Serosurvey in the human population.

The Salamanca Province of Spain is an endemic zone for Mediterranean spotted fever. In this area, only one case of Q fever has been reported and typhus group rickettsial diseases have never been diagnosed. To obtain a panoramic view of the presence of antibodies to the most ubiquitous rickettsial agents, 400 sera specimens from a statistically representative sample of the human population of Salamanca City and its surrounding province were subjected to indirect microimmunofluorescent test for antibodies against Rickettsia conorii, Coxiella burnetti, phase II, and Rickettsia typhi antigens. Titers greater than or equal to 1:40 to R. conorii were found in 73.5% of the sera. Positivity was more common in subjects who reported closer contact with the natural environment and/or with domestic animals. Seropositivity to C. burnetti, phase II, was detected in 50.2% of sera. These positive cases were related to rural environmental factors and to previous contact with animals. The frequency of antibodies increased with age showing a progressive exposure to the rickettsial antigen. The prevalence of antibodies to R. conorii and C. burnetii in the human population of Salamanca Province is higher than that reported from any other geographic zone. The study of antibodies to R. typhi showed that 12.5% of the sera had titers greater than or equal to 1:40. According to our results, seropositivity to this rickettsial antigen cannot be related to any particular group of population nor interpreted totally as cross reactivity with R. conorii. Our data show a wide distribution of R. conorii and C. burnetii antigens in Salamanca Province, and also indicate the presence of R. typhi antigens in this area.

Adolescent↗

PubMed1959

Study on growth of Rickettsia. V. Penetration of Rickettsia tsutsugamushi into mammalian cells in vitro.

A system has been described in which the penetration of Rickettsia tsutsugamushi (Karp strain) into tissue culture cells can be quantitated, and the factors affecting this process studied. The results indicated that rickettsial penetration in vitro depended largely on the viability of the organisms. Certain components of the fluid environment such as the divalent cations and protein were found to be of importance. The temperature dependence of the penetration process was found to vary with the nature of the suspending medium. A number of compounds related to L-glutamic acid enhanced penetration, whereas metabolic inhibitors depressed this process. Aureomycin at concentrations between 50 and 250 microg./ml. inhibited the penetration of rickettsiae while chloramphenicol at similar concentrations was ineffective. The results are discussed in terms of the biological and biochemical properties of this group of agents.

Animals↗

PubMed2004

T cells mediate cross-protective immunity between spotted fever group rickettsiae and typhus group rickettsiae.

Rickettsioses are severe infections caused by obligately intracellular bacteria that preferentially infect the endothelium lining the vasculature. The causative agents, rickettsiae, have been divided according to biological, genetic, and antigenic parameters into 2 main groups: spotted fever and typhus. They have not been thought to stimulate cross-reactive protective immune responses; however, in this study, we show that, in relevant animal models that mimic human rickettsial infections, there is reciprocal immunological cross-protection between spotted fever group and typhus group rickettsiae. Furthermore, we present evidence that T cells are responsible for this cross-immunity and that cross-stimulation of T cells also occurs in humans.

Adoptive Transfer↗