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PubMed2000

Reducing the risk of transfusion-transmitted rickettsial disease by WBC filtration, using Orientia tsutsugamushiin a model system.

BACKGROUND: Careful donor screening and infectious disease marker testing have significantly reduced the incidence of transfusion-transmitted diseases and improved the safety of the blood supply. However, transfusion-transmitted diseases resulting from the use of asymptomatic yet infectious donors continue to put patients at risk. This study was undertaken to determine if third-generation WBC filters could remove Orientia tsutsugamushi-infected cells from contaminated blood. STUDY DESIGN AND METHODS: Packed RBCs were inoculated with human MNCs infected with O. tsutsugamushi at levels estimated to occur in asymptomatic infectious donors. WBC reduction was accomplished with a third-generation WBC filter. Prefiltration and postfiltration specimens were collected, serially diluted, and injected into mice to determine the infectivity of the samples. RESULTS: Mice receiving WBC-reduced packed RBCs showed no signs of illness or markers of infectivity, which suggested that a reduction of as much as 10(5) infectious rickettsiae could be achieved by filtration. CONCLUSION: The high-efficiency, third-generation, WBC-reduction filters that were tested may provide protection against the transfusion transmission of scrub typhus rickettsiae by removing from contaminated blood cells that contain intracellular bacteria.

Animals↗
PubMed1994

Detection of Rickettsia tsutsugamushi in Experimentally infected mice by PCR.

We developed a rapid procedure for the detection of Rickettsia tsutsugamushi DNA by the PCR technique. The primer pair used for the PCR was designed from the DNA sequence of the gene encoding a 120-kDa antigen, which was proven to be group specific by immunoblot analysis with mouse hyperimmune sera against various rickettsial strains. This PCR method was able to detect up to 10 ag of plasmid DNA (pKT12). Specific PCR products were obtained with DNAs from R. tsutsugamushi Kato, Karp, Gilliam, TA716, TA1817, and Boryong, but not with DNAs from other rickettsiae, such as R. prowazekii, R. typhi, R. akari, and strain TT118. In a study with experimentally infected mice, the PCR method could detect rickettsial DNA from 2 days after inoculation (DAI), whereas serum antibody against R. tsutsugamushi could be detected from 6 to 8 DAI by an immunofluorescence test. Although clinical manifestations subsided after 14 DAI, rickettsial DNA in blood samples could be detected by PCR for up to 64 DAI. These results suggest that this PCR method can be applied to the early diagnosis of scrub typhus and can also be used to detect the residual rickettsiae after clinical symptoms subside.

Animals↗
PubMed2006

An epitope shared by cellular cytokeratin and Orientia tsutsugamushi.

Many microbial pathogens have epitopes shared with host cell components and these epitopes may induce transient or longer-term tissue-damaging autoantibody responses. We observed that several mouse monoclonal antibodies (MAbs) raised against Orientia tsutsugamushi were also reactive with host cells. One such antibody, MAb Rb105, cross-reacted with the cytoskeleton, as shown by immunofluorescent staining. Biochemical studies identified the cross-reacting component as a cytokeratin protein. These results identify an epitope shared by O. tsutsugamushi and the cytokeratins of host cells. In addition, antibodies cross-reactive with the cytoskeleton were detected in the sera of scrub typhus patients, suggesting that an epitope similar to that detected by MAb Rb105 may be recognized by human antibodies.

Antibodies, Bacterial↗
PubMed1998

Molecular and serologic survey of Orientia tsutsugamushi infection among field rodents in southern Cholla Province, Korea.

Field rodents were collected from six areas in southern Cholla Province, Korea from October to December 1993. Twenty-eight (24%) of the 119 Apodemus agrarius were seropositive (> 1:10) for Orientia tsutsugamushi by the passive hemagglutination assay (PHA). Of the seropositive cases, 11 specimens had antibody titers greater than 1:80. No seropositive specimens were found among the eight Crocidura lasiura collected. On the other hand, the polymerase chain reaction (PCR) amplified about 520 basepairs of a gene encoding the 56-kD protein from the genomic DNA of 12 strains of O. tsutsugamushi tested. This target DNA sequence was amplified from the 11 (8.7%) blood specimens of A. agrarius, and one of the eight C. lasiura also showed evidence of O. tsutsugamushi infection by PCR. Only one of the PCR-positive samples was also PHA-positive. These results suggest that the PCR combined with a serologic assay more accurately detects the degree of infection of rodents with rickettsiae-causing scrub typhus in epidemiologic surveys.

Animals↗
PubMed1992

Gelatin particle agglutination test for early serodiagnosis of Japanese spotted fever.

A gelatin particle agglutination (PA) test for Japanese spotted fever has been developed. Gelatin particles were sensitized with a sonicated causative rickettsia and used as antigens. The antibodies by PA test were detected as early as days 4-7 after the onset, whereas those by indirect immunoperoxidase (IP) test were detected after days 8-11. In addition, PA titers were higher than IP titers before days 20-23. The agglutinins detected by PA test were proven to be IgM because they were all sensitive to dithiothreitol. PA test was, however, less specific than IP test, giving a little nonspecific reaction to the sera from patients with scrub typhus and from individuals unrelated to those two rickettsioses. Nevertheless, PA test, which is simple, rapid, and easy to interpret the results, is useful for the early serodiagnosis of Japanese spotted fever.

Agglutination Tests↗
PubMed1984

Rickettsial diseases of the Far East: new perspectives.

The rickettsial disease of man found only in Asia is mite-borne (scrub) typhus, caused by Rickettsia tsutsugamushi. Unique to southern Japan is a little-known human mononucleosis-like disease caused by Rickettsia sennetsu. In 1981 and 1982, there was a remarkable resurgence in the number of reported cases of mite-borne typhus in Japan after some years of virtual absence. Recent studies of R sennetsu have resulted in its reclassification to the genus Ehrlichia, members of which until now have been exclusively pathogens of animals. The historical background of ecologic investigations, in Malaysia and elsewhere, of these two developments suggest directions for future research.

Animals↗
PubMed1977

Indirect immunofluorescence antibodies in natural and acquired Rickettsia tsutsugamushi infections of Philippine rodents.

Antibodies against Rickettsia tsutsugamushi detected by the indirect fluorescent-antibody test (IFAT) were present in most rats trapped from a human focus of scrub typhus in the Philippines. Rickettsiae were isolated only from rats with positive IFATs. Naturally acquired antibodies persisted for at least 11 months, and antibodies resulting from experimental infections of rats persisted for at least 7 months. A common Philippine rodent, Rattus mindanensis, tolerated experimental infections with both local and standard Karp strains of R. tsutsugamushi, and such infections always produced a positive IFAT.

Animals↗
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↗
PubMed1997

Antibodies to Orientia tsutsugamushi in soldiers in northeastern Thailand.

The prevalence and incidence of antibodies to Orientia tsutsugamushi, the etiologic agent of scrub typhus, in Thai soldiers living and working near the Thai-Cambodian border in Si Sa Ket Province was investigated. The point prevalence of antibodies varied from 0 to 4.1%. The incidence of antibodies, calculated from individuals who seroconverted following a negative result in a previous bleeding 3 to 5 months earlier, was 4.21% (9/214) in January 1992, 0 in April 1992 and 3.76% (8/213) in September 1992. An annual infection rate of 2.66% was estimated.

Antibodies, Bacterial↗
PubMed1993

High-level expression of a 56-kilodalton protein gene (bor56) of Rickettsia tsutsugamushi Boryong and its application to enzyme-linked immunosorbent assays.

The 56-kDa protein of Rickettsia tsutsugamushi, which is located on the rickettsial surface, has been shown to be an immunodominant antigen. The gene that encodes the 56-kDa protein of R. tsutsugamushi Boryong (bor56) was cloned. Sequencing revealed an open reading frame of 1,602 bp encoding 534 amino acids with a molecular weight of 56,803. The 56-kDa protein of R. tsutsugamushi Boryong (Bor56) was expressed as a fusion protein with the maltose-binding protein of Escherichia coli by deleting 252 bp from the 5' end of the open reading frame and subcloning it into the StuI site of pIH821. The recombinant fusion protein was purified by amylose column chromatography for application in an enzyme-linked immunosorbent assay to evaluate the ability of the method to detect the antibody to R. tsutsugamushi in human patient sera. By using sera from 100 patients with scrub typhus and 70 patients with other febrile diseases, a high diagnostic sensitivity (95%) and a high diagnostic specificity (100%) were demonstrated, suggesting the suitability of the recombinant antigen for use as an immunodiagnostic tool.

ATP-Binding Cassette Transporters↗
PubMed1996

Comparison of serological reactions of Rickettsiae-infected patients and rabbit anti-Proteus OX antibodies with Proteus OX2, OX19 and OXK lipopolysaccharides.

The reactivity of anti-Rickettsiae human antibodies with Proteus OX cells is used as a presumptive rickettsial diseases diagnostic test Weil-Felix reaction. In presented studies we compare the reactivity of human anti-Rickettsiae and rabbit anti-Proteus antibodies with series of Proteus OX2, OX19 and OXK lipopolysaccharides (LPS). Polyclonal rabbit anti-OX2, anti-OX19 and anti-OXK sera reacted only with the homologous LPS--OX2, OX19 and OXK, respectively. The antibodies of Japanese spotted fever patients were less specific and reacted with OX2 as well as OX19 LPS. The antibodies of scrub typhus patients recognized Proteus OXK LPS, only. The serological reactions of O-antigen of P. mirabilis S1959 indicated that this, previously serologically not classified, strain may belong to the OXK group. Bacteria, used in the studies, came from the American, Japanese and European strain collections. The series of OX2, OX19 and OXK LPS, isolated from these Proteus strains, presented pattern of electrophoretic mobility and serological reactivities specific for each of OX-group types.

Animals↗
PubMed1996

Antibodies to Orientia tsutsugamushi in Thai soldiers.

Thai soldiers who were conscripted, Royal Thai Army forces, professional Border Patrol Police, or local militia (Thai Rangers) located in any of seven provinces of Thailand were bled in April and again, four months later, in July 1989. In 1991, soldiers from five different locations in southern Thailand were bled once, in July. Serum samples were tested by indirect fluorescent antibody assay for antibody to Orientia (formerly Rickettsia) tsutsugamushi, etiologic agent of scrub typhus, with any titer > or = 1:50 considered positive. Prior to field exercises, prevalence of antibody varied significantly between different types of units, ranging between 18.6% for Thai Rangers and 6.8% for the Royal Thai Army. The April prevalence, July prevalence, and incidence varied significantly by province in 1989, with highest incidence being 14.5% in Kanchanaburi and the lowest 0% in Utraladit. The prevalence in southern Thailand in 1991 varied between 1.6% and 6.8%. The data demonstrate that O. tsutsugamushi is widely distributed in Thailand and that military activity consisting of field exercises that simulate combat conditions significantly expose soldiers to infection.

Antibodies, Bacterial↗
PubMed1986

Production and characterization of monoclonal strain-specific antibodies against prototype strains of Rickettsia tsutsugamushi.

We have developed 18 hybridoma cell lines which secrete murine monoclonal strain-specific antibodies to prototype strains of Rickettsia tsutsugamushi: nine anti-Gilliam, four anti-Karp and five anti-Kato antibodies. All the monoclonal antibodies reacted only with their homologous strains in direct and indirect immunofluorescence (IF), or indirect immunoperoxidase (IP) test. By IF and IP tests with the monoclonal antibodies, 22 strains of R. tsutsugamushi, which were newly isolated from mites, field rodents and patients with Tsutsugamushi disease (scrub typhus) in Japan, were all clearly identified as either Gilliam or Karp type. Analysis by polyacrylamide gel electrophoresis and immunoblotting techniques revealed that the monoclonal antibodies recognized primarily the polypeptides of an apparent molecular weight of 54 to 56 kilodaltons of the homologous rickettsial surface. The monoclonal antibodies produced in the present study should enhance the serotyping and further analytical investigation of the rickettsial antigens since they recognize the strain- or type-specific polypeptides and do not show any cross-reaction among strains.

Animals↗
PubMed1996

ELISA-based colorimetric detection of Rickettsia tsutsugamushi DNA from patient sera by nested polymerase chain reaction.

A rapid diagnostic system for scrub typhus was established using colorimetric detection of nested polymerase chain reaction (PCR). This system relied on binding the amplified DNA via a sequence in one of oligodeoxyribonucleotide to the DNA-binding protein GCN4 coated on the well of a micotiter dish. The primer pairs used for the nested PCR were designed on the basis of the homologous nucleotide sequence of the gene that encodes the 56 kDa antigen of serovariants. With this colorimetric PCR, diagnosis can be performed easily from serum samples of patients before the antibody titer increases or in the early stage of the disease. Furthermore, these positive results are able to be confirmed by pathogenic isolation.

Base Sequence↗
PubMed1991

Use of monoclonal antibodies against Rickettsia tsutsugamushi Kawasaki for serodiagnosis by enzyme-linked immunosorbent assay.

Monoclonal antibodies (MAbs) against Rickettsia tsutsugamushi Kawasaki were prepared. The crossreactivity tests of the MAbs performed by using antigenically distinct strains of R. tsutsugamushi in immunofluorescence and immunoblotting analyses indicated that the Kawasaki strain contains a strain-specific epitope and also contains a common epitope on the 56-kDa polypeptide cross-reactive with the Gilliam strain, group- and subgroup-specific epitopes on the 46-kDa polypeptide, and a subgroup-specific epitope on the 25-kDa polypeptide. By using the strain-specific MAb for serodiagnosis of tsutsugamushi disease (or scrub typhus fever), we have established a method which was designated the inhibition enzyme-linked immunosorbent assay. The principle of the method is to measure the percentage of inhibition of antigen absorption on a MAb-coated plate by antibody-positive sample sera which were mixed with the antigen suspension. The advantages of this test for practical use are that (i) crude antigen can be used, i.e., purification of the antigen is not required; (ii) the test is more sensitive than immunofluorescence; (iii) the final judgment of plus or minus is clear-cut; and (iv) rickettsial antigenic types in the patients can be distinguished by this test.

Antibodies, Bacterial↗
PubMed1997

Isolation of a new Orientia tsutsugamushi serotype.

Orientia tsutsugamushi, the etiological agent of scrub typhus, is an antigenically diverse organism and many serologically distinct strains have been identified. The 56 kDa protein of O. tsutsugamushi, a major protein in the outer membrane, has been thought to be responsible for this antigenic variability. A strain of O. tsutsugamushi isolated in Korea cross-reacted with both Gilliam strain-specific and Karp strain-specific monoclonal antibodies. When its 56 kDa protein gene was cloned and analyzed, its sequence showed variation especially between 1,200 and 1,250 bp, showing that this isolate is a new O. tsutsugamushi strain.

Amino Acid Sequence↗
PubMed1980

New paper enzyme-linked immunosorbent technique compared with microimmunofluorescence for detection of human serum antibodies to Rickettsia tsutsugamushi.

A new paper enzyme-linked immunosorbent assay was developed for the screening and titration of human serum antibodies against the scrub typhus rickettsia, Rickettsia tsutsugamushi. The objective [corrected] was to provide a relatively simple method for antibody screening which required neither sophisticated laboratory equipment nor a high degree of technological skill. The technique develops an enzyme product from filter paper saturated with a 5-aminosalicylic acid substrate and enzymatically reacted with a commerically available anti-human immunoglobulin G peroxidase conjugate. The product of the enzymatic reaction can be interpreted visually. Comparison of 351 human sera tested by the immunofluorescent and paper enzyme-linked immunosorbent assays against a three-antigen pool of the Karp, Kato, and Gilliam strains of R. tsutsugamushi demonstrated an agreement of 96%. The sensitivity of the paper enzyme-linked immunosorbent assay as compared to immunofluorescence was 98.2%, and the specificity was 94.4%.

Antibodies, Bacterial↗
PubMed1990

[Imported tsutsugamushi fever].

Scrub typhus rarely figures among the imported tropical diseases in western Europe. Rickettsia tsutsugamushi is transmitted by larval mites. A typical eschar develops at the site of the mite bite during the incubation period, after which systemic symptoms (remittent or continuous high fever, severe headache, tender lymphatic glands and symptoms of bronchitis) develop with sudden onset. Antibodies appear during the second week and can be shown by a positive and increasing titer against Prot. OXK (Weil-Felix) or by specific rickettsial agglutination. The Weil-Felix reaction is neither very sensitive nor very specific. Therefore, treatment should be started as soon as suspicion arises whether a positive serology is available or not. Tetracycline drugs are effective treatment and fever subsides in less than 24 hours in most patients. - We report the rare observation of a tourist who imported tsutsugamushi fever from India.

Animals↗