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Serologic diagnosis of epidemic typhus fever.

Microagglutination (MA), microcomplement-fixation (CF), indirect microimmunofluorescence (micro-IF) and Weil-Felix (WF) tests were compared for sensitivity and specificity in detection of antibodies against Rickettsia prowazekii and for serologic diagnosis of epidemic typhus fever. Paired sera from hospitalized Ethiopian patients suffering from febrile illness were examined. Purified particulate rickettsial antigens used in MA, CF and micro-IF tests were made from infected yolk sacs by differential centrifugation from molar NaCl and from NaCl-sucrose, D20=1.1491, pH 5.5. In sensitivity the tests ranked micro-IF greater than MA greater than CF greater than WF. Use of specific anti-IgG and anti-IgM sera in parallel micro-IF tests made it possible to differentiate cases of recrudescent epidemic typhus (Brill-Zinsser disease) from primary epidemic typhus cases. Antibodies reacting in high titer with R. canada were produced by 86% of patients with rising antibody titers against R. prowazekii.

Agglutination Tests↗

Rickettsia tsutsugamushi infection and scrub typhus incidence among Chinese military personnel in the Pescadores Islands.

Personnel assigned to infantry units of the army of the Republic of China (Taiwan) stationed in the Pescadores Islands experienced the highest incidence rate of scrub typhus (4%) and the highest rate of Rickettsia tsutsugamushi infection (12%) of the units studied during 1976. The ratio of scrub typhus to infection with R. tsutsugamushi was 30%. Non-immune personnel, when infected, were more likely to develop scrub typhus than were those who had previously been infected. There was a correlation between clinical illness and high antibody titers developed in response to the infection.

Humans↗

Changing risk of scrub typhus in relation to socioeconomic development in the Pescadores islands of Taiwan.

Long term changes in risk of Rickettsia tsutsugamushi infection among civilian residents of the Pescadores Islands of Taiwan were associated with changing social conditions. Age specific incidence rates of scrub typhus in the Pescadores before 1940 were highest among children under 5 years of age. Rickettsiae can be demonstrated in vectors, wild animals continue to be infected and scrub typhus occurs in military personnel. A serologic survey for antibody to R. tsutsugamushi was conducted during 1975 and 1977 and failed to show evidence of previous infection among children. Two events appear to be associated with the decreased incidence of rickettsial infection in the young: increased urbanization and increased enrollment in schools. Both changes accompanied socioeconomic development which took place in the islands during the past 50 years. Prevalence of antibody to R. tsutsugamushi continues to be equal in the sexes and is undoubtedly due to similar occupational exposure of both sexes in fields and farms where vectors are numerous. The absence of apparent morbidity due to scrub typhus among the civilian populace was attributed to the mild nature of the disease caused by Pescadores strains, misdiagnosis and a lack of obligatory reporting.

Adolescent↗

Diagnosis of typhus infection with Rickettsia tsutsugamushi by polymerase chain reaction.

Two sets of oligonucleotide primers were used to amplify the genomic DNA of Rickettsia tsutsugamushi, the causative agent of scrub typhus (tsutsugamushi disease), by the polymerase chain reaction. Each set of primers amplified 538-bp and 109-bp products, representing part of a gene encoding a possible major 58-kDa immunogenic protein, from whole genomic DNA extracted from R. tsutsugamushi strains Karp, Kato, Gilliam, Kuroki and Kawasaki. No amplification was observed from R. sibirica, R. rickettsii, mouse and human genomic DNA. DNA amplification was observed from crude lysates of peripheral whole blood, tissue homogenates and paraffin-embedded skin biopsy sections obtained from patients with scrub typhus disease. Southern blot analysis demonstrated the specificity of the amplified DNA fragments following hybridisation with a DNA probe generated from R. tsutsugamushi strain Karp. By means of this procedure, a rapid and sensitive diagnosis of scrub typhus disease can be made during the acute stage of this infection.

Acute Disease↗

Seroepidemiology of spotted fever group and typhus group rickettsioses in humans, South Korea.

The prevalence of spotted fever group (SFG) and typhus group (TG) rickettsioses was investigated in 3,362 sera by immunofluorescence assay. The serum samples were obtained from patients with acute febrile episodes in South Korea from December 1992 to November 1993. The number of polyvalent positive sera against SFG rickettsial agents at the level of 1: 40 dilution was 269 (8%) in Rickettsia sibirica, 482 (14.34%) in R. conorii, and 546 (16.24%) in R. akari. Many of the positive sera contained immunoglobulin (Ig) M antibodies rather than IgG antibodies. These results strongly suggest that SFG rickettsioses are prevalent in Korea. For TG rickettsial agents, the number of positive sera was 1,096 (32.60%) in R. typhi and 951 (28.29%) in R. prowazekii. Only a few epidemic typhus positive sera contained IgM antibodies. The result suggests that recent and/or primary infections of epidemic typhus were very rare in Korea during the said period. Among seven patients who had high titers (1:5,120) of IgG antibody to R. prowazekii, six were over 50 years old. The result suggests that Brill-Zinsser disease was prevalent in Korea.

Adolescent↗

Pregnancy with scrub typhus and vertical transmission: a case report.

Scrub typhus is a rickettsian disease which is seldom found in pregnancy. A 31-year-old, 34 weeks pregnant woman presented with fever, chill and cough for 6 weeks. Fetal jeopardy was found then a cesarean section was performed to deliver a 2,200 g male with hepatosplenomegaly. The mother's diagnosis was confirmed by positive Weil-Felix (OXK titer 1:320) and scrub typhus (titer 1:1600) tests. Vertical transmission was also demonstrated by a positive scrub typhus IgM in her child.

Adult↗

[Serodiagnosis of scrub typhus by dot-blot method].

Microimmunofluorescence (IF) and immunoperoxidase tests are generally used for the serodiagnosis of scrub typhus. While these tests give satisfactory results in the hands of experienced personnel, they can be troublesome for inexperienced technicians. To develop a simpler diagnostic method, dot-blot assay was examined in this study. Six antigenically distinctive strains of Gilliam, Karp, Kato, Shimokoshi, Kawasaki and Kuroki of Rickettsia tsutsugamushi and a strain of Rickettsia sibirica were dotted on a nitrocellulose sheet by a dot-blot instrument. The sheets were treated with patient sera, followed by peroxidase-conjugated anti-human immunoglobulin-antibody and then with the substrate of the enzyme, and the color development on the dots was compared by naked eye observation. By this procedure, anti-rickettsial antibody-positive patient sera showed clear color development on at least one, usually several dots, while the very faint color was observed by the treatment with antibody-negative sera. On the other hand, it was ascertained that the antigens on nitrocellulose sheets were stable for at least 4 months at room temperature. Therefore the diagnostic kits were prepared, and the practical application of this procedure for diagnosis of scrub typhus were tested in Shizuoka and Miyazaki Prefectural Pabulic Health Laboratories. The results indicated a very good comparability between the dot-blot assay and IF-tests, and this dot-blot method was ascertained as a simple and useful method for the scrub typhus serodiagnosis.

Adolescent↗

Epidemic typhus infection in cynomolgus monkeys (Macaca fascicularis).

A nonhuman primate model of clinical Rickettsia prowazekii infections was developed in cynomolgus monkeys (Macaca fascicularis). Monkeys infected intravenously with 10(7) plaque-forming units developed clinical signs of illness and pathological changes characteristic of epidemic typhus infection in humans. Increases in total leukocyte counts, serum alkaline phosphatase, blood urea nitrogen, and serum glutamic pyruvate transaminase values were observed. Microscopic examination revealed typical typhus nodules in the brains of two monkeys that died. These data indicated that the cynomolgus monkey is a suitable model for study of the pathogenesis of epidemic typhus infection and may prove valuable in the evaluation of candidate R. prowazekii vaccines.

Animals↗

Typhus and typhuslike rickettsiae associated with opossums and their fleas in Los Angeles County, California.

The recent discovery of cat fleas (Ctenocephalides felis) infected with a typhuslike rickettsia (designated the ELB agent) raises the question of whether similar rickettsial infections exist in wild cat flea populations. We verified the presence of the ELB agent and Rickettsia typhi in urban and suburban areas of Los Angeles, Calif. Opossums trapped in close proximity to the residences of human murine typhus cases in Los Angeles county and other areas within the city of Los Angeles were tested for the presence of typhus group rickettsiae by the polymerase chain reaction (PCR). The presence of rickettsiae in the spleen tissues of three opossums (n = 9) and in 66 opossum fleas (n = 205) was determined by PCR and was verified by dot blot and Southern transfer hybridization. Further analysis of the amplified PCR products generated by a series of primer pairs derived from either the 17-kDa antigen gene or the citrate synthase gene revealed that both R. typhi and the ELB agent were present in the tested samples. Dual infection was not noted in the samples; however, the fleas were infected with either R. typhi or the ELB agent. The presence of the ELB agent in the cat flea population may have implications for public health. Whether this agent is responsible for the mild cases of human murine typhus in urban and suburban areas of Los Angeles or in other endemic foci remains to be determined.

Animals↗

Rapid diagnosis of scrub typhus by a passive hemagglutination assay using recombinant 56-kilodalton polypeptides.

The genes encoding the 56-kDa polypeptides were amplified by polymerase chain reaction from the genomic DNAs of three serotypes of Rickettsia tsutsugamushi, Gilliam, Karp, and Boryong. The amplified products were cloned into expression vector pIH821, and the recombinant antigens were expressed in Escherichia coli as fusion proteins with maltose-binding protein. The recombinant 56-kDa polypeptides were purified by affinity chromatography for the sensitization of sheep erythrocytes. The recombinant 56-kDa polypeptides were evaluated with 89 serum specimens from health blood donors, 94 serum specimens from scrub typhus patients, and 31 serum specimens from patients with other febrile diseases by a passive hemagglutination assay (PHA). Among the scrub typhus patients diagnosed by indirect immunofluorescent-antibody testing, the antibodies to R. tsutsugamushi were detected in 93 patients (99%). One serum specimen from a healthy person showed a false-positive reaction by this method. The recombinant PHA showed no cross-reactions with sera obtained from other febrile patients with diseases such as murine typhus, hemorrhagic fever with renal syndrome, and leptospirosis. In conclusion, this recombinant PHA could be substituted for the conventional indirect immunofluorescent-antibody test and the immunoperoxidase test.

Antibodies, Bacterial↗

Identification of a novel rickettsial infection in a patient diagnosed with murine typhus.

Identification of ELB agent-infected fleas and rodents within several foci of murine typhus in the United States has prompted a retrospective investigation for this agent among human murine typhus patients. This agent is a recently described rickettsia which is indistinguishable from Rickettsia typhi with currently available serologic reagents. Molecular analysis of the 17-kDa antigen gene and the citrate synthase gene has discriminated this bacterium from other typhus group and spotted fever group rickettsiae. Current sequencing of its 16S ribosomal DNA gene indicates a homology of 98.5% with R. typhi and 99.5% with R. rickettsii. Through a combination of restriction fragment length polymorphism and Southern hybridization analysis of rickettsia-specific PCR products, one of five tested patient blood samples was shown to be infected with ELB while R. typhi infections were confirmed in the remaining samples. This is the first reported observation of a human infection by the ELB agent and underscores the utility of PCR-facilitated diagnosis and discrimination of these closely related rickettsial infections.

Adult↗

Usefulness of eschar PCR for diagnosis of scrub typhus.

We report here on the case of a child who was infected with scrub typhus, and we made the diagnosis according to the serology and by performing PCRs on the child's eschar. The patient was treated with azithromycin, and he did not experience any complications. Performing nested PCR on the eschar might be both a rapid diagnostic test for scrub typhus in the early acute stage and a differential test as to whether or not a scab is a scrub typhus eschar.

Antigens, Bacterial↗

Typhus-induced facial necrosis.

A patient with open wounds on his hands developed a severe case of typhus following exposure to a wild rabbit. Typhus resulted in extensive necrosis of the mid-face, including the nasal alae, pinna, and lips. The patient also sustained extensive extremity necrosis. A brief review of the epidemiology, pathophysiology, and treatment of this cause of facial necrosis is presented, as well as a discussion of dopamine gangrene. While this uncommon entity probably played a large part in the necrosis of this patient's extremities, these peripheral lesions are also compatible with typhus.

Adult↗

A stranger to our camps: Typhus in American history.

Medical observers during the American Civil War were happily surprised to find that typhus fever rarely made an appearance, and was not a major killer in the prisoner-of-war camps where the crowded, filthy, and malnourished populations appeared to offer an ideal breeding ground for the disease. Through a review of apparent typhus outbreaks in America north of the Mexican border, this article argues that typhus fever rarely if ever extended to the established populations of the United States, even when imported on immigrant ships into densely populated and unsanitary slums. It suggests that something in the American environment was inhospitable to the extensive spread of the disease, most likely an unrecognized difference in the North American louse population compared to that of Europe.

American Civil War↗

Temporal changes in prevalence of scrub typhus rickettsia (Orientia tsutsugamushi) infecting the eggs of Leptotrombidium imphalum (Acari: Trombiculidae).

Eggs from seven colony lines of the chigger mite Leptotrombidium imphalum (Vercammen-Grandjean & Langston) were examined for infection with Orientia tsutsugamushi (Hyashi), the etiologic agent of scrub typhus. The polymerase chain reaction (PCR) using primers OtP 56.809 and OtM 56.1221, which amplify a 291 bp region of the P56 gene of O. tsutsugamushi, was used to detect scrub typhus within single eggs. All seven chigger mite lines produced infected eggs with varying rates of infection (Li1 = 8.1%, n = 124; Li2 = 45.6%, n = 90; Li3 = 30.1%, n = 144; Li4 = 31.7%, n = 145; Li5 = 21.3%, n = 136; Li6 = 41.6% n = 77; Li7 = 22.5%, n = 110). The 3 wk with the highest infection rates for each line using Fourier analysis were as follows: Li1 = 2, 7, 14; Li2 = 4, 6, 12; Li3 = 3, 6, 12; Li4 = 4, 6, 12; Li5 = 5, 7, 14; Li7 = 4, 6, 12. Li6 only had nine measurements over time; therefore, Li6 was excluded from individual analysis. Infection rates of scrub typhus in eggs occurred in a 2-wk 2-d cycle, using Fourier analysis of combined data. Not only did infection rates vary among the progeny of females, but temporal variation also occurred.

Animals↗

Geographic association of Rickettsia felis-infected opossums with human murine typhus, Texas.

Application of molecular diagnostic technology in the past 10 years has resulted in the discovery of several new species of pathogenic rickettsiae, including Rickettsia felis. As more sequence information for rickettsial genes has become available, the data have been used to reclassify rickettsial species and to develop new diagnostic tools for analysis of mixed rickettsial pathogens. R. felis has been associated with opossums and their fleas in Texas and California. Because R. felis can cause human illness, we investigated the distribution dynamics in the murine typhus-endemic areas of these two states. The geographic distribution of R. felis-infected opossum populations in two well-established endemic foci overlaps with that of the reported human cases of murine typhus. Descriptive epidemiologic analysis of 1998 human cases in Corpus Christi, Texas, identified disease patterns consistent with studies done in the 1980s. A close geographic association of seropositive opossums (22% R. felis; 8% R. typhi) with human murine typhus cases was also observed.

Adolescent↗

Acute respiratory distress syndrome associated with scrub typhus: diffuse alveolar damage without pulmonary vasculitis.

Pathologic findings of scrub typhus have been characterized by vasculitis of the microvasculature of the involved organ resulting from a direct invasion by Orientia tsutsugamushi. We experienced a case of acute respiratory distress syndrome (ARDS) associated with scrub typhus. The case was proven by eschar and high titer of serum IgM antibody (positive at 1:1280). Open lung biopsy showed diffuse alveolar damage (DAD) in the organizing stage without evidence of vasculitis. Immunofluorescent antibody staining and polymerase chain reaction for O. tsutsugamushi failed to demonstrate the organism in the lung tissue. The patient expired due to progressive respiratory failure despite doxycycline therapy. Immunologic mechanism, without direct invasion of the organism, may participate in the pathogenesis of ARDS associated with scrub typhus.

Acute Disease↗

Diagnostic criteria for scrub typhus: probability values for immunofluorescent antibody and Proteus OXK agglutinin titers.

The sensitivities and specificities of the indirect microimmunofluorescent antibody (IFA) and Weil-Felix (OXK) tests for scrub typhus were established for a range of titers using groups of diseased and control (other febrile illnesses) patients diagnosed by other methods. At a cut-off point of greater than or equal to 1:400, the IFA test was 0.96 specific, and at greater than or equal to 1:320, the OXK was 0.97 specific. Using either these highly specific levels of antibody or other rigorous diagnostic criteria (isolation or 4-fold rising titers), the prevalence of scrub typhus infection was determined to be 0.22 in an unselected population of febrile patients in a rural Malaysian hospital. Probability values (Pr) for the correct diagnosis of scrub typhus were then calculated from the specificity, sensitivity and prevalence determination for a range of titers. The Pr for an OXK titer of greater than or equal to 1:320 was 0.79, and the Pr for an IFA titer of greater than or equal to 1:400 was 0.78. When both these titers were present in a single specimen, the Pr increased to 0.96.

Agglutination Tests↗