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Genomic identification of Rickettsia slovaca among spotted fever group rickettsia isolates from Dermacentor marginatus in Armenia.

Restriction fragment length polymorphism (RFLP) analysis of polymerase chain reaction (PCR) amplified genes was used for genomic identification of Armenian isolates of the Spotted fever group (SFG) rickettsiae with unclear taxonomic position. Analysis was performed by using one genus-specific primer pair derived from R. prowazekii citrate synthase gene and two species-specific primer pairs derived from R. rickettsii genes for 190 K and 120 K antigens following AluI, PstI and RsaI digestion of amplicons. All tested rickettsial SFG Armenian isolates from Dermacentor marginatus were identified as R. slovaca. The geographic distribution and genetic homogeneity of R. slovaca strains are discussed.

Animals↗

Studies on rickettsia-like organism disease of the tropical marine pearl oyster I: the fine structure and morphogenesis of pinctada maxima pathogen rickettsia-like organism

The present paper reports for the first time the discovery of a rickettsia-like organism (RLO) in the cultured tropical marine pearl oyster Pinctada maxima with mass mortality in the Hainan Province of China. This organism parasitizes the cytoplasm of host cells and forms intracytoplasmic eosinophilic inclusions. These organisms are extremely pleiomorphic in shape and average 967 x 551 nm in size, as measured in cross sections of transmission electron micrographs. The organisms exhibit clearly recognizable ultrastructural characteristics of prokaryotic bacteria-like cells, including two trilaminar membranes, an increasing electron-dense periplasmic ribosome zone, and a thread-like DNA nucleoidal structure. In addition to the above prokaryotic characteristics, the following unique biological characteristics were confirmed by TEM: (i) These organisms are usually located in host cells in two ways, namely, free in the cell cytoplasm and involved within membrane-limited phagolysosomes; (ii) The organisms exist in two morphological cell types, namely a small cell variant (SCV) and a large cell variant (LCV). The most important morphological difference between two cell types is that the SCV is obviously ribosome-rich in the periphery of the body, which makes SCV more electron-dense in the cytoplasm and narrower in the central nucleoid area than the LCV; (iii) Two propagative modes of the organisms, transverse binary fission and budding, are observed in cytoplasm and phagolysosomes of host cells under TEM, in which the budding is more often seen in phagolysosomes. These characteristics indicate that the organism is a separate species in the family Rickettsiaceae and should be classified into the genus Rickettsia. On the basis of the existence of the two propagative modes and two cell types, and intracellular location, we propose a developmental cycle for this organism which includes a vegetative differentiation stage to develop LCV by transverse binary fisson and a budding differentiation stage to develop resistant SCV. Copyright 1999 Academic Press.

Journal Article↗

Sensitive microplate enzyme-linked immunosorbent assay for detection of antibodies against the scrub typhus rickettsia, Rickettsia tsutsugamushi.

A microtiter enzyme-linked immunosorbent assay (ELISA) has been developed for the titration of antibodies against scrub typhus in human and animal sera. Scrub typhus rickettsiae were grown in monolayers of irradiated mouse LM3 cells and separated from host cell materials by differential centrifugation, filtration through a glass filter (AP-20, Millipore Corp.), and isopycnic banding in Renografin density gradients. The scrub typhus ELISA antigens were obtained from the purified viable rickettsiae by French pressure cell disruption and addition of 0.2% Formalin to the soluble extract. Antisera prepared in rabbits against the prototype Karp, the Kato, and the Gilliam strains of scrub typhus were used to standardize the ELISA and to compare its sensitivity and specificity to that of the indirect fluorescent antibody test (IFA). ELISA titers were measured as the greatest serum dilution showing an optical density 0.25 above controls or by the optical density achieved at a fixed serum dilution. The IFA and ELISA end point titers were quite similar, and all three measures of titer had comparable specificity for the strains of scrub typhus. No cross-reactions between the typhus and scrub typhus wera were observed by ELISA. Both the immunoglobulin M (IgM) and IgG antibody titers of 12 sequential sera from four patients with scrub typhus were obtained by IFA and ELISA. The IFA and ELISA end point titers for IgM and IgG had correlation coefficients of 0.91 and 0.97, respectively, whereas the ELISA optical density values at a serum dilution of 1:100 had slightly lower correlations with IFA titers (0.80 and 0.94). Early rising IgM titers followed by rising IgG titers were demonstrated by ELISA in three patients with primary scrub typhus infections, whereas the IgG response predominated in a patient with a reinfection. It is concluded that the ELISA for scrub typhus is a very satisfactory alternative to the IFA test.

Animals↗

The effects of x-rays and beta rays (tritium) on the growth of Rickettsia mooseri and Rickettsia akari in embrvonate eggs.

The growth of Rickettsia mooseri was accelerated and quantitatively increased in embryonate eggs containing tritium oxide at levels of 180, 90, and 45 mc./egg during the growth period. The eggs of a group containing 22.5 mc./egg showed only a slight increase in the rate of growth of organisms; the infections in the eggs of a group given 11.2 mc./egg did not differ significantly from those of the control group. On the other hand, growth of R. akari was inhibited in embryonate eggs containing tritium oxide at levels of 180, 90, and 45 mc./egg, and partially inhibited in groups containing 22.5 and 11.2 mc./egg. The patterns of growth of R. mooseri and of R. akari exposed to tritium oxide for 6 hours prior to inoculation into embryonate eggs did not differ significantly from that of the control group. Single and divided doses of x-rays to the host resulted in partial inhibition of the growth of R. akari.

Beta Particles↗

Rickettsia honei: a spotted fever group Rickettsia on three continents.

Rickettsia honei (also known as strain TT-118) has been detected on three continents. Originally isolated in Thailand in 1962 (and confirmed in 2001), it has also been detected on Flinders Island (Australia) in 1993 and in Texas (USA) in 1998. On each continent it has been associated with a different species of tick. The original isolate (Thai Tick Typhus strain TT-118) was from a pool of larval Ixodes and Rhipicephalus ticks. Later it was detected in I. granulatus from Rattus rattus. Its pathogenicity for humans has not yet been confirmed in Thailand, but it is possibly responsible for the Spotted Fever Group human rickettsiosis in Thailand. The strain from Texas (USA) was isolated from Amblyomma cajennense ticks taken from cattle. Its pathogenicity for humans has not yet been confirmed in Texas. However, this tick is known to bite humans. The strain from Flinders Island (Australia) described as R. honei, has been isolated from patients with "Flinders Island Spotted Fever" and from Aponomma hydrosauri ticks taken from blue-tongue lizards (Tiliqua nigrolutea), tiger snakes (Notechis ater humphreysi), and copperhead snakes (Austrelaps superbus) on Flinders Island. The ecology of R. honei in this location is unusual in that reptiles, rather than mammals, are the vertebrate hosts.

Animals↗

Ultrastucture of the surface of Rickettsia prowazeki and Rickettsia akari.

Negative-contrast electron microscopy revealed that the outer layer of the envelope of rickettsiae is composed of a matrix of tetragonally arranged subunits. The layer projects approximately 7 nm from the cell wall. It is suggested that this outer layer is analogous to the structure considered capsule-like in morphology.

Antigens, Bacterial↗

Evaluation of IgG antibody response against Rickettsia conorii and Rickettsia slovaca in patients with DEBONEL/TIBOLA.

The aim of the study was to determine the IgG antibody response to spotted fever group Rickettsia (SFGR) R. conorii and R. slovaca, and its specificity and sensitivity in patients with DEBONEL/TIBOLA. A prospective study of 31 patients with DEBONEL was carried out from January 2001 to May 2004. The SFGR serology testing (IgG IFA) for the diagnosis of DEBONEL/TIBOLA showed 61% sensitivity and 100% specificity. The R. slovaca antigen allowed the diagnoses in 18 of the 31 patients (58%), and 17 patients (55%) were diagnosed with this disease using R. conorii antigen. Therefore, using R. slovaca as antigen did not improve the sensitivity of the assay.

Antibodies, Bacterial↗

A comparative view of Rickettsia tsutsugamushi and the other groups of rickettsiae.

Recent researches on the rickettsial group microorganisms are summarized in their comparative aspects of morphology, cultivation and multiplication, susceptibility to chemotherapeutics, chemical structure of envelopes, nucleic acid, protein constitution, and gene structures. From this overview, Rickettsia tsutsugamushi seems to have different properties from the others and should be reclassified into a new genus, and a new species name as Orientia tsutsugamushi is proposed.

Bacterial Proteins↗

In vitro studies on Rickettsia-host cell interactions: lag phase in intracellular growth cycle as a function of stage of growth of infecting Rickettsia prowazeki, with preliminary observations on inhibition of rickettsial uptake by host cell fragments.

Two Rickettsia prowazeki seeds, an "early" seed in the logarithmic or exponential growth phase and a "late" seed in the stationary or possibly early decline phase, were prepared in chicken embryo (CE) cell cultures and compared with respect to morphology and infection cycle in CE cells in culture. Differences in size and ultrastructure of the organisms in the two seeds were similar to those seen in other gram-negative bacteria at comparable stages to growth. Vacuolar structures, rare in log-phase organisms, were common in stationary-phase organisms. Minute spherical forms reminiscent of minicells were seen in the stationary-phase preparations. In quantitative uptake experiments, organisms, typical in size and morphology of each preparation, had comparable capacity per plaque-forming unit to penetrate into CE cells in suspension when the seeds had been depleted of host cell membrane fragments and other debris. This suggests that host cell fragments, presumably of membrane origin, competitively inhibit rickettsial uptake by intact CE cells. Organisms of the log-phase organisms displayed a lag phase of about 7.5 h, during which they enlarged and increased in intensity of staining, before entering the log phase of growth.

Cells, Cultured↗

Absorption of Rickettsia rickettsii antibodies by Rickettsia rickettsii antigens in four diagnostic tests.

Immunoabsorption profiles were determined for Rickettsia rickettsii antigens used in four confirmatory tests for detection of antibodies to Rocky Mountain spotted fever. A human serum reactive in the four tests was absorbed with each test antigen and then reexamined by all four tests. The results indicated that the whole organism and complement-fixation antigen absorbed the whole array of antibodies to R. rickettsii, whereas erythrocyte-sensitizing substance coated on latex or erythrocytes did not. With erythrocyte-sensitizing substance coated on latex there was a decrease in titer when tested by the latex, indirect hemagglutination, and complement fixation tests but none when tested by the microimmunofluorescence test. With erythrocyte-sensitizing substance coated on sheep erythrocytes there was a decrease in titer when tested by indirect hemagglutination or complement fixation but none when tested by latex or microimmunofluorescence. These findings confirm that erythrocyte-sensitizing substance contains more than a single antigenic substance and plays a major role in the complement fixation test and a minor role in the microimmunofluorescence test.

Antibodies, Bacterial↗

Flying squirrel-associated Rickettsia prowazekii (epidemic typhus rickettsiae) characterized by a specific DNA fragment produced by restriction endonuclease digestion.

The DNA from flying squirrel-associated Rickettsia prowazekii was characterized by using a specific DNA fragment produced by digestion with the enzyme BamHI. The DNA fragment was cloned into a plasmid vector and used to readily distinguish between available human- and flying squirrel-associated R. prowazekii DNAs derived from crude cytoplasmic extracts.

Animals↗

Antibodies to Rickettsia rickettsii, Rickettsia typhi, Coxiella burnetii, Bartonella henselae, Bartonella quintana, and Ehrlichia chaffeensis among healthy population in Minas Gerais, Brazil.

Rickettsial diseases except those belonging to spotted fever group rickettsioses are poorly studied in South America particularly in Brazil where few epidemiological reports have been published. We describe a serosurvey for Rickettsia rickettsii, R. typhi, Coxiella burnetii, Bartonella henselae, B. quintana, and Ehrlichia chaffeensis in 437 healthy people from a Brazilian rural community. The serum samples were tested by indirected micro-immunoflourescence technique and a cutoff titer of 1:64 was used. The seroprevalence rates for R. rickettsii, R. typhi, C. burnetii, B. henselae, B. quintana, and E. chaffeensis were respectively 1.6% (7 samples); 1.1% (5 samples); 3.9% (17 samples); 13.7% (60 samples); 12.8% (56 samples), and 10.5% (46 samples). Frequent multiple/cross-reactivity was observed in this study. Age over 40 years old, urban profession, and rural residence were significantly associated with some but not all infections rate. Low seropositivity rates for R. rickettsii, R. typhi, and C. burnetii contrasted with higher rates of seropositivity for B. quintana, B. henselae, and E. chaffeensis. These results show that all tested rickettsial species or antigenically closely related possible exist in this particular region.

Adolescent↗