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PubMed1994

Prevalence of antibodies to rickettsiae in the human population of suburban Bangkok.

Following a report of three cases of scrub typhus in suburban Bangkok, we performed a serosurvey in the patients' communities. Both IgG and IgM antibodies were measured in an indirect immunoperoxidase assay, using separate spots of antigen from Rickettsia tsutsugamushi (scrub typhus), R. typhi (murine typhus), and TT-118 spotted fever group rickettsiae. Of 215 people donating blood, antibody levels indicative of most recent exposure to R. tsutsugamushi were most prevalent (21%), followed by R. typhi (8%), and TT-118 (4%). Seroprevalence suggesting most recent exposure to R. tsutsugamushi varied by location (range 13-31%), gender (26% of females and 13% of males), age (61-80-year-old individuals were the highest, with a prevalence of 38%), and contact with orchards and orchid farms (29% of those with extensive contact, 38% of those with occasional contact, and 10% of those with no contact). These patterns indicated that exposure to R. tsutsugamushi was related to occupation and behavior, as has been observed in areas of rural transmission. Expansion of metropolitan Bangkok has created a situation in which people employed in agriculture live with people employed in the city. As a result, a serosurvey in suburban Bangkok reveals evidence of murine typhus, which is usually transmitted in urban areas, as well as scrub and tick typhus, which are usually transmitted in rural areas.

Adolescent↗
PubMed1991

Specific amplification of Rickettsia tsutsugamushi DNA from clinical specimens by polymerase chain reaction.

Polymerase chain reaction (PCR) was used to detect Rickettsia tsutsugamushi-specific DNA in clinical specimens. The primer pair used for PCR was designed from the nucleotide sequence of the gene encoding the 56-kDa antigen of the Gilliam strain. Theses primers led to a 78-bp fragment by amplifying the genomic DNAs from five serovariants, i.e., the Gilliam, Karp, Kato, Kawasaki, and Kuroki strains of R. tsutsugamushi, and also the DNA from blood clots of patients with scrub typhus, even at the early stage of onset of the disease. This indicates that this method is suitable for the diagnosis of scrub typhus.

Base Sequence↗
PubMed2002

Comparative evaluation of selected diagnostic assays for the detection of IgG and IgM antibody to Orientia tsutsugamushi in Thailand.

We compared the performance of 2 commercially available dipstick assays, 2 enzyme-linked immunosorbent assays (ELISAs), and an indirect immunofluorescent antibody (IFA) assay for the diagnosis of scrub typhus, using the indirect immunoperoxidase (IIP) test as the reference standard. The dipstick assays were the Integrated Diagnostics (Baltimore, MD) Dip-S-Ticks Scrub Recombinant (r56) dipstick test (INDX assay) and the PanBio (Brisbane, Australia) Scrub Typhus IgM and IgG Rapid Immunochromatographic test (PanBio assay). One of the ELISAs used pooled cell lysates of Karp, Kato, and Gilliam strain Orientia tsutsugamushi as antigen (pooled-antigen ELISA), and the other used a recombinant r56 protein as the antigen (recombinant ELISA). With a panel of 123 positive and 227 negative sera, sensitivity and specificity of the assays were as follows: INDX assay, IgG, 60% and 95%, IgM, 60% and 97%; PanBio assay, IgG, 94% and 96%, IgM, 83% and 93%; IFA (1:400 cutoff), IgG, 91% and 96%, IgM, 85% and 98%; pooled-antigen ELISA, IgG (1:1600 cutoff), 97% and 89%, IgM (1:400 cutoff), 94% and 91%; recombinant ELISA, IgG (1:1600 cutoff), 97% and 92%, IgM (1:400 cutoff), 93% and 94%. Because of its excellent performance and use of a standardized, commercially available antigen, the recombinant ELISA is suitable for use in a diagnostic laboratory, where it may be able to replace the IFA and IIP assays. In contrast, the PanBio dipstick assay was easy to perform and did not require sophisticated equipment, making it suitable for use in rural areas where more sophisticated diagnostic tests such as the ELISA and IFA may not be available.

Antibodies, Bacterial↗
PubMed1995

[Isolation of Rickettsia tsutsugamushi from the blood samples of patients in Taiwan].

Scrub typhus, an acute febrile eruptive disease caused by Rickettsia tsutsugamushi, is one of the most commonly reported communicable diseases in Taiwan. However, information about the epidemic trends of scrub typhus in Taiwan is still very limited. Therefore, this study was performed to isolate the causative agent and determine the prevalence of three different serotypes. From June 1992 to December 1994, lymphocyte cell cultures were grown from different regions of Taiwan. Cell cultures were assayed for R. tsutsugamuhi via immuno-fluorescence, using antisera against Gillian, Karp and Kato strains. Thirty-three of the specimens were positive for R. tsutsugamushi, including one from Keelung, 23 from Taitung, 2 from Kinmen and 7 from Lienjan. Of these, 15 were classified as Gillian-related types, 10 were Karp-related types and 8 were not positively classified. These results indicate Gilliam-and Karp-related serotype are more prevalent than the Kato-related serotype in Taiwan.

Base Sequence↗
PubMed1993

Epitope mapping of the Sta58 major outer membrane protein of Rickettsia tsutsugamushi.

Binding studies of 160 overlapping, synthetic octapeptides from the hydrophilic regions of the Sta58 major outer membrane protein of Rickettsia tsutsugamushi with sera from patients with scrub typhus revealed 15 immunodominant peptides which are recognized by all the sera tested. Further analysis of the specificity of peptide binding with five of these peptides indicated that the peptides showed significantly stronger binding to scrub typhus patients' sera than they did to sera from patients with other febrile illnesses common in the region, i.e., malaria, dengue fever, typhoid fever, and leptospirosis. The main antibody class binding to these peptides appears to be immunoglobulin M, and there appears to be little correlation between reactivity with peptides and antibody titers measured by the indirect immunoperoxidase test.

Amino Acid Sequence↗
PubMed2006

Causes of acute, undifferentiated, febrile illness in rural Thailand: results of a prospective observational study.

The adult patients who, between July 2001 and June 2002, presented at any of five hospitals in Thailand with acute febrile illness in the absence of an obvious focus of infection were prospectively investigated. Blood samples were taken from all of the patients and checked for aerobic bacteria and leptospires by culture. In addition, at least two samples of serum were collected at different times (on admission and 2-4 weeks post-discharge) from each patient and tested, in serological tests, for evidence of leptospirosis, rickettsioses, dengue and influenza. The 845 patients investigated, of whom 661 were male, had a median age of 38 years and a median duration of fever, on presentation, of 3.5 days. Most (76.5%) were agricultural workers and most (68.3%) had the cause of their fever identified, as leptospirosis (36.9%), scrub typhus (19.9%), dengue infection or influenza (10.7%), murine typhus (2.8%), Rickettsia helvetica infection (1.3%), Q fever (1%), or other bacterial infection (1.2%). The serological results indicated that 103 (12.2%) and nine (1%) of the patients may have had double and triple infections, respectively. Leptospirosis and rickettsioses, especially scrub typhus, were thus found to be major causes of acute, undifferentiated fever in Thai agricultural workers.

Acute Disease↗
PubMed1997

Production of monoclonal antibodies against Rickettsia massiliae and their use in antigenic and epidemiological studies.

Rickettsiae are gram-negative, obligate intracellular bacteria which have historically been divided into three groups: the typhus group, the scrub typhus group, and the spotted fever group (SFG). Recently, several new SFG rickettsiae have been characterized, and most of these species are associated with ticks and have, as yet, no known pathogenicity toward humans. Rickettsia massiliae, which is widely distributed in Europe and Africa, is one such rickettsia. In order to investigate the antigenic relationships between R. massiliae and other rickettsial species and to develop a more convenient methodology for identifying R. massiliae, we produced monoclonal antibodies against the type strain (Mtu1T) of R. massiliae by fusing immunized splenocytes with SP2/0-Ag14 myeloma cells. A panel of 16 representatives were selected from the 163 positive hybridomas identified on initial screening, and their secreted monoclonal antibodies were further characterized. The reactivities of these 16 monoclonal antibodies with a large panel of rickettsial species were assessed by the microimmunofluorescence assay. All species of the SFG rickettsiae reacted with the monoclonal antibodies directed against epitopes on lipopolysaccharide, which is the common antigen among the SFG rickettsiae. Some closely related species of the SFG, such as Bar29, "R. aeschlimanni," and R. rhipicephali, showed strong cross-reactivities with the monoclonal antibodies directed against epitopes on the two major high-molecular-mass heat-labile proteins (106 and 120 kDa). In addition, species-specific monoclonal antibodies demonstrated that R. massiliae is antigenically different from other rickettsial species. Moreover, these species-specific monoclonal antibodies were successfully used for identifying R. massiliae in the ticks collected from southern France, and are therefore potentially useful tools in the identification and investigation of R. massiliae in ticks in large-scale field work.

Antibodies, Bacterial↗
PubMed2006

A review of endoparasitic acarines of Malaysia with special reference to novel endoparasitism of mites in amphibious sea snakes and supplementary notes on ecology of chiggers.

Some 2,000 species of mites of the family Trombiculidae are known in the world. The 6-legged larvae are mostly ectoparasites of reptiles, birds, mammals and invertebrates. Their 8-legged active nymphs and adults are free-living predators. In the Asia-Pacific region, a few species in various genera are vectors of scrub typhus and scrub-itch. In this a paper, a very bizarre trombiculid species, Vatacarus ipoides Southcott 1957, endoparasitic in the trachea of the amphibious sea snake, Laticauda colubrina (Schenider) is re-described based mostly on new-born larvae reared in the laboratory. Life history study of the mite produced very novel and interesting results. A brief account of the life-cycle was presented at the first laboratory demonstration of the Malaysian Society of Parasitology and Tropical Medicine Meeting by Nadchatram and Audy (1965). The life history is illustrated and described here in greater detail. The active nymphal, and the akinetic teleiophane stages are bypassed, which is unusual in the life-cycle of the family Trombiculidae. Also, the larva is the only stage in the life-cycle that feeds. The sexes are predetermined in the larval neosomatic stage and give rise to small males and bigger females. Having obtained adults of the species, by rearing, it is deemed unnecessary for the original proposal by Southcott to erect a new family, Vatacaridae, because the adults share all the attributes of the family Trombiculidae. The male and female obtained through laboratory rearing are illustrated for the first time. Relationship of V. ipoides with Laticauda snakes show close host-specificity, in a group of acarines that are generally habitat-specific. Possible explanations for their association are discussed. The unusual morphology and the formation of new structures during an instar is of ontogenetic and evolutionary importance. The hypertrophic larvae are superficially vermiform, rather than typically acarine in shape. This, and other biological features, necessitated the proposal of new morphological terms, and they are discussed here.

Animals↗
PubMed1984

Febrile illness in Malaysia--an analysis of 1,629 hospitalized patients.

We studied 1,629 febrile patients from a rural area of Malaysia, and made a laboratory diagnosis in 1,025 (62.9%) cases. Scrub typhus was the most frequent diagnosis (19.3% of all illnesses) followed by typhoid and paratyphoid (7.4%); flavivirus infection (7.0%); leptospirosis (6.8%); and malaria (6.2%). The hospital mortality was very low (0.5% of all febrile patients). The high prevalence of scrub typhus in oil palm laborers (46.8% of all febrile illnesses in that group) was confirmed. In rural Malaysia, therapy with chloramphenicol or a tetracycline would be appropriate for undiagnosed patients in whom malaria has been excluded. Failure to respond to tetracycline within 48 hours would usually suggest a diagnosis of typhoid, and indicate the need for a change in therapy.

Adolescent↗
PubMed1993

[Detection antigen of Rickettsia tsutsugamushi by using monoclonal antibody].

Dot-ELISA was used with HRP-labelled monoclonal antibodies Kp3 (IgGi) and Kp4-6 (IgM) against antigen of the Karp strain of Rickettsia tsutsugamushi to detect the antigen of R. tsutsugamushi. The positive rate of sera of 56 acute scrub typhus patients and 37 wild rats caught from the countryside of the endemic area as well as 29 batches of chiggers collected from the wild rats were 92.9%, 100% and 89.7%, respectively. No cross reaction was found with antigens of other rickettsia groups such as R. mooseri, R. prowazeki, R.burneti and R. rickettsia. In patients suffering from acute scrub typhus the antigen of R. tsutsugamushi could be detected earlier in the course of illness than the antibody. This method is very sensitive that an amount of antigen of 6.7ng/microliters can be detected. It is easy to perform and the results can be read by naked eye.

Animals↗
PubMed2003

Evaluation of an immunoglobulin M capture enzyme-linked immunosorbent assay for diagnosis of Orientia tsutsugamushi infection.

To differentiate scrub typhus from other acute febrile diseases, a rapid and reliable serological diagnosis is important. We developed an immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (ELISA) for diagnosis of recent Orientia tsutsugamushi infections in humans. The 56-kDa major outer membrane protein of O. tsutsugamushi is well known as the most immunodominant antigen in scrub typhus. The test is based on the use of the biotinylated recombinant 56-kDa protein of O. tsutsugamushi Boryong, Bor56, which was expressed as a fusion protein with a maltose-binding protein in Escherichia coli. In the test, the serum IgM antibodies were captured by anti-human IgM antibodies coated onto a microtiter plate. The captured IgM antibodies were revealed through sequential addition of biotinylated Bor56 antigen and peroxidase-conjugated streptavidin to the plate. The IgM capture ELISA was compared with the immunofluorescence antibody assay (IFA) by testing 176 serum samples from patients with diagnosed cases of rickettsial disease and patients with other acute febrile diseases. Of the 81 IgG IFA-positive samples, 78 tested positive (sensitivity, 96.3%) and all 31 IgM IFA-positive samples tested positive (sensitivity, 100%) by the IgM capture ELISA. The specificity of the IgM capture ELISA was 99%, and 1 of the 95 IFA-negative samples was positive in the assay. These results strongly suggest that IgM capture ELISA using the recombinant Bor56 antigen is a reliable and detailed method for the detection of early O. tsutsugamushi infection.

Antibodies, Bacterial↗
PubMed1978

Enzyme immunoassay of antibody to Rochalimaea quintana: diagnosis of trench fever and serologic cross-reactions among other rickettsiae.

Enzyme immunoassay (EIA) tests were used to diagnose trench fever and to determine cross-reactions of Rochalimaea quintana with other rickettsiae. The results were compared with those obtained by counterimmunoelectrophoresis (CIE). All sera from cases of primary or relapsed forms of trench fever were positive both in EIA, with serum antibody titers of 1:20-1:640, and in CIE, giving one to three precipitin lines. Sera from patients with other rickettsial infections were also tested for reactivity with R. quintana antigen: typhus group (Rickettsia prowazekii, Ricketsia mooseri), 15 sera; spotted fever group (Ricketsia ricketsii, Rickettsia akari), eight sera; and scrub typhus (Rickettsia tsutsugamushi), six sera. Strong reactions occurred with four sera from patients with scrub typhus, giving one or two lines in CIE and EIA titers of 1:40-1:160; these results were extended to guinea pig antisera to R. tsutsugamushi. About 50% of typhus group sera reacted with a single line in CIE and had antibody titers of 1:20-1:80 by EIA. The results show that EIA is accurate for the diagnosis of trench fever and, with the results obtained by CIE, suggest that R. quintana is antigenically related to R. tsutsugamushi and possibly to rickettsiae in the typhus group as well.

Antibodies, Bacterial↗
PubMed2005

Fatal septicemic melioidosis in a young military person possibly co-infected with Leptospira interrogans and Orientia tsutsugamushi.

Concurrent melioidosis, leptospirosis, and scrub typhus after rural activities is rarely reported. A 19-year-old previously healthy man had fever onset after 2 weeks of military training. Pneumonia became evident on the fifth day of fever under intravenous penicillin and oral minocycline therapy. Acute respiratory failure developed the next day with shock and acute renal and liver function deterioration, which resulted in death. Blood cultures on the third and fifth days grew Burkholderia pseudomallei. Serology revealed leptospirosis and scrub typhus. The emergence of melioidosis in Taiwan and this death without antibiotic treatment for melioidosis alert us that B. pseudomallei should be included as a possible pathogen of pneumonia and sepsis, especially after rural activities.

Adult↗
PubMedDate not supplied

Imported rickettsioses in German travelers.

Twenty-two cases of rickettsiosis imported to Germany (13 men, nine women, average age 42 years) in a 5-year period were analyzed retrospectively regarding the travel histories, symptoms and clinical findings, laboratory features and course of the disease. The two primary rickettsial diseases were boutonneuse fever (18 patients) and scrub typhus (three patients). One patient had murine typhus. The main symptom was fever in 91% followed by headache (64%), myalgia (40%), arthralgia (50%) and diarrhea (36%). The most frequent clinical finding was lymphadenopathy in 65%. Eschar was detectable in 55% of patients with Rickettsia conori infection and in one patient with Rickettsia tsutsugamushi infection. All patients with R. tsutsugamushi infection as well as 33% of the patients with R. conori infection had a macular exanthema. One patient with scrub typhus had pleural and pericardial effusions. Seventy-three percent had an increased ESR. Three patients had leucocytosis, three increased transaminases and two normochromic anemia. The incubation period for R. conori infection was 5 to 28 days (average 14 days), for R. tsutsugamushi infection 7 to 21 days (average 16 days). Twenty-one patients were treated with tetracycline or doxycycline, one with erythromycin. All patients were cured. One patient had a relapse. Due to the fact that the symptoms are often not characteristic and that the routine laboratory findings are of only marginal help, the diagnosis of rickettsial diseases is often not easy. A detailed travel history sometimes gives an important hint for diagnosis.

Adult↗
PubMed1993

Serotype-specific amplification of Rickettsia tsutsugamushi DNA by nested polymerase chain reaction.

Polymerase chain reaction (PCR) with nested primer pairs was used to diagnose scrub typhus and identify the Rickettsia tsutsugamushi serotype. The primer pairs used for PCR were designed on the basis of the nucleotide sequence of the gene that encodes the 56-kDa antigen. Serotype-specific primers were used in the second PCR amplification. Five serovariants, the Gilliam, Karp, Kato, Kawasaki, and Kuroki strains of R. tsutsugamushi, were identified by nested PCR. In addition, the serotype identified by PCR with DNA from blood clots was the same as that of the strain isolated from five patients with scrub typhus. These findings indicate that this method is useful for diagnosis and identification of the rickettsial serotype in infected patients.

Base Sequence↗
PubMed2005

Cloning and sequence analysis of the 22-kDa antigen genes of Orientia tsutsugamushi strains Kato, TA763, AFSC 7, 18-032460, TH1814, and MAK 119.

The 22-kDa protein antigen is one of several antigens recognized by sera from scrub typhus patients infected with Orientia tsutsugamushi. The 22-kDa protein genes from six O. tsutsugamushi strains (Kato, TA763, AFSC 7, 18-032460, TH1814, MAK119) were cloned and their sequences were determined and compared to each other and to the Karp strain sequence listed in GenBank. The sequence alignment revealed that the promoter regions of these seven strains were highly conserved. However, the ORFs exhibited some variation. The phylogenetic analysis of the DNA sequences indicated that among the seven strains assessed, Kato and TA763 were the most closely related, while Karp and TH1814 were the most distantly related. The information gained from this analysis will facilitate our selection of O. tsutsugamushi strains from which antigens should be derived to be included in a multivalent vaccine candidate for scrub typhus.

Amino Acid Sequence↗
PubMed2002

Antigenic types of Orientia tsutsugamushi in Malaysia.

The seroprevalence of various Orientia tsutsugamushi (OT) strains among Malaysian patients with suspected scrub typhus infections was determined using an indirect immunoperoxidase (IIP) assay. IgG against a single OT strain were detected in six sera (3 Karp, 1 Gilliam and 2 TC586), whereas IgM antibodies against a single OT strain (Gilliam) were noted in 3 sera (Gilliam). IgG reactive to all OT strains were present in 33 (47.1%) of the 70 sera and IgM reactive to all OT strains were present in 22 (78.6%) of the 28 sera. The fact that most sera were reactive to multiple OT strains suggests that group-specific antigens are involved in scrub typhus infections, whereas very few were due to strain-specific epitopes present on these strains. Peak IgG and IgM titers were noted more frequently against Gilliam, Karp, and TA763 strains: this suggests that these strains may be the commonest infecting strains among Malaysian patients. Two predominant OT polypeptides consistently reacted with patients' sera were the 70 kDa and 56 kDa proteins.

Antigens, Bacterial↗
PubMed2025

Comparative virulence analysis of seven diverse strains of Orientia tsutsugamushi reveals a multifaceted and complex interplay of virulence factors responsible for disease.

Orientia tsutsugamushi is an obligate intracellular bacterium found in Leptotrombidium mites that causes the human disease scrub typhus. A distinguishing feature of O. tsutsugamushi is its extensive strain diversity, yet differences in virulence between strains are not well defined nor well understood. We sought to determine the bacterial drivers of pathogenicity by comparing seven strains using murine infections combined with epidemiological human data to rank each strain in terms of relative virulence. Murine cytokine expression data revealed that the two most virulent strains, Ikeda and Kato, induced higher levels of IL-6, IL-10, IFN-γ and MCP-1 than other strains, consistent with increased levels of these cytokines in patients with severe scrub typhus. We sought to identify the mechanistic basis of the observed differential virulence between strains by comparing their genomes, in vitro growth properties and cytokine/chemokine induction in host cells. We found that there was no single gene or gene group that correlated with virulence, and no clear pattern of in vitro growth rate that predicted disease. However, microscopy-based analysis of the intracellular infection cycle revealed that the only fully avirulent strain in our study, TA686, differed from all the virulent strains in its subcellular localisation and expression of its surface protein ScaC. This leads us to a model whereby drivers of pathogenicity in Orientia tsutsugamushi are distributed throughout the genome, likely in the large and varying arsenal of effector proteins encoded by different strains, and that these interact in complex ways to induce differing immune responses and thus differing disease outcomes in mammalian hosts.

Orientia tsutsugamushi↗