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Clinical observations of scrub typhus on Penghu (the Pescadores Islands).

Between May and September 1973, 68 cases of scrub typhus in Chinese military personnel on the Pescadores Islands were studied. The common symptoms and signs were fever, chills, headache, eschar, myalgia, and lymph node enlargement. Most eschars were located in the axilla, waist, groin and genitals, and neck. These lesions were painless and not noticed by the patients themselves. Regional lymph node enlargement at the site of eschar drainage was common. Relative bradycardia with fever was observed in 40%, a skin rash in 35% of the patients. Leucopenia was noted more frequently in the febrile than in the convalescent stage, but more than half of the patients had a normal count. Lymphocytosis was prominent, especially during the convalescent period. An acceleration of ESR was noted. Instead of depression of the erythroid series in the marrow which was reported previously, 47% of examined patients were found to have erythroid hyperplasia. Two patients showed marked hypocellularity of the marrow in the acute febrile stage; later on became normocellular. Albuminuria was present in 15 and BUN increased in 12 patients. Elevation of serum bilirubin and SGOT was also noted. Biologic false positive VDRL tests were observed in nine patients. In 30 tests elevation of Proteus OX-K titres between 1:160 and 1:640 was noted. A geometric mean OX-K titre rise in the patients is presented; the mean titre reached a peak in the third week of illness, and then fell off. Most of the patients were treated with tetracycline 500 mg every six hours for about nine days. The fever usually subsided within 36 hours. Complications or mortality were not encountered.

Adult

Host defenses in experimental scrub typhus: effect of chloramphenicol.

The effect of chloramphenicol treatment on the development of immunity to scrub typhus in mice was studied. Chemotherapy was administered either shortly before infection and for 14 days thereafter (group I), or from 7 to 21 days postinfection (group II). Although the full course of either regimen resulted in complete protection of the mice against subsequent challenge with the homologous strain of Rickettsia tsutsugamushi, initiation of chemotherapy at 7 days postinfection resulted in more rapid development of immunity against both the original infection and subsequent challenge. In both treatment groups, a 1- to 2- day hiatus was observed between immunity to challenge in the treated animal and the ability to transfer this immunity to syngeneic recipients with lymphocyte-enriched spleen cells. Similarly, complement-fixing antibodies were not detectable until shortly after the animals were able to resist challenge. These data supported the conclusion that the rickettsiostatic effect of chloramphenicol allows the infected animal time to mount an effective immune response and, further, that initiation of chemotherapy early in the infection may delay development of this response.

Animals

Gamma-irradiated scrub typhus immunogens: development and duration of immunity.

The development and duration of immunity to lethal scrub typhus infection was studied in BALB/c mice vaccinated with gamma-irradiated Rickettsia tsutsugamushi, strain Karp. One intraperitoneal injection containing approximately 10(8) 50% mouse lethal doses (MLD(50)) of irradiated organisms elicited an immune response protective against challenge with 10(5) MLD(50) of viable Karp. The same mass of immunogen given in three injections at 5-day intervals increased homologous (Karp strain) protection 25-fold and heterologous (Kato strain) protection 60-fold. Further temporal expansion of the immunization regimen did not increase protection. Subcutaneous vaccination provided significant, but lower, levels of protection than were achieved by intraperitoneal immunization, but the levels of cell-transferable immunity elicited by the two routes were approximately the same. Immunologically specific protection after intraperitoneal vaccination developed rapidly enough to provide resistance against simultaneous challenge with 200 MLD(50) of Karp. Homologous immunity was protective against a 10(6)-MLD(50) challenge 7 days after completion of the three-injection regimen, remained at that level for 3 months, dropped to 10(4) MLD(50) by 9 months, and was effective against a 50-MLD(50) Karp challenge at 12 months. Protection against heterologous challenge was first observed on day 17 and peaked on day 38, when the mice resisted a 10(5)-MLD(50) Kato challenge. Thereafter, heterologous protection waned rapidly and was not significant at 6 months.

Animals

Host defenses in experimental scrub typhus: role of cellular immunity in heterologous protection.

The relative contributions of cellular and humoral immunity in scrub typhus infections were studied in inbred mice employing paired strains of Rickettsia tsutsugamushi differing in virulence. An infectious dose (100 MID50) of the less virulent Gilliam strain resulted in heterologous immune protection against an otherwise lethal challenge (1,000 MLD50) of the virulent Karp strain. Partial heterologous protection against lethal Karp challenge was observed in animals preimmunized with the Gilliam strain as early as 3 days prior to challenge, whereas complete protection against illness and death existed in animals immunized at least 7 days prior to challenge. In the heterologous protection provided by prior Gilliam infection, the role of humoral immunity was not of primary importance for the following reasons: (i) significant levels of complement-fixing antibody against R. tsutsugamushi were not detectable until long after animals were solidly immune; (ii) antibody eventually appearing after Gilliam immunization exhibited a consistently low complement-fixing titer against the immunizing homologous (Gilliam) strain and contained no detectable activity against the heterologous challenge (Karp) strain; and (iii) passive transfer of large quantities of serum from Gilliam immune mice, themselves immune to Karp challenge, failed to protect recipients against a similar challenge. However, protection was afforded by the passive transfer of serum containing antibody against Karp, suggesting a major role for antibody in protection against homologous infection. This heterologous challenge system was particularly useful because it minimized the role of humoral immunity, at least early in the course of infection, and allowed a definitive examination of the cellular response. Cell-mediated immunity played a major role in the heterologous protection observed after Gilliam immunization. This was evidenced by the significant protection against Karp challenge afforded by the passive transfer of spleen cells from animals immunized with Gilliam 7 to 63 days previously. Of the immune spleen cells, only those which were nonadherent, presumably lymphocytes, were capable of transferring passive heterologous protection. This protective effect of nonadherent cells could be ablated by depleting the cell population of thymus-derived or T cells with anti-theta serum and complement prior to transfer but not by use of anti-immunoglobulin serum and complement, which selectively removes bone marrow-derived or B cells. These results suggested that the cell in immune spleens capable of conferring heterologous protection was a T lymphocyte.

Animals

Presence of antibodies to scrub typhus and murine typhus in dogs from Selangor, Peninsular, Malaysia.

Dog sera, collected from different communities throughout Selangor, Peninsular Malaysia, were investigated for the presence of antibodies to R. tsutsugamushi and R. typhi. Scrub typhus antibodies were present in animals from the rural areas only, whereas murine typhus antibodies were observed in equal numbers of dogs from both rural and metropolitan areas. Greater percentage of dogs from suburban areas had demonstrable antibody titers to murine typhus than from the urban area.

Animals

The prevalence of scrub typhus antibodies in residents of West Malaysia.

Based on the prevalence of antibody, an estimated 3% of the population of rural Malaysia is infected with Rickettsia tsutsugamushi each year, resulting in positive antibody rates in focal areas of 6 to 69%. Most of these infections do not appear to produce clinical scrub typhus. A wide range of seropositivity rates was found in areas otherwise resembling each other in predominant occupation, terrain, and nearby habitat. The prevalence rates however were significantly higher in people who worked in forested areas and significantly lower in people with urban occupations.

Adult

Serologic classification of scrub typhus isolates from Pakistan.

Detailed serologic characterization of 79 isolates of Rickettsia tsutsugamushi from Pakistan was performed by using complement fixation and direct immunofluorescence techniques for comparison with three prototype strains. Seventy-one were of the Karp serotype and one was of a Gilliam serotype. Seven isolates were multiply-reactive: one with Karp and Gilliam, 3 with Karp and Kato, and 3 with Karp, Gilliam and Kato, No strain failed to react with at least 1 of the 3 prototype antisera. No correlation of antigenic types with local geographic factors, animal hosts, vecotr mites, or specific strain virulence was observed. These findings suggest the possibility that antigenic heterogeneity may be more restricted in areas peripheral to the geographic epicenter of scrub typhus endemicity, that the serotypes are relatively stable genetically and that new geographic variants are probably infrequent. The importance of these findings to the practical problems of immunoprophylaxis and serodiagnosis are discussed.

Animals

Host defenses in experimental scrub typhus: role of normal and activated macrophages.

Resident peritoneal macrophage from BALB/c mice were infected in vitro with Rickettsia tsutsugamushi strain Gilliam, and rickettsial growth was estimated by microscopic examination of Giemsa-stained cells. Both number of infected macrophage per culture and number of intracellular rickettsiae per cell increased with time during culture. Treatment of rickettsiae with immune serum before infection macrophage cultures reduced the number of infected macrophage by 50%. Macrophage treated in vitro with lymphokines were able to suppress rickettsial growth in the absence of detectable antibody and exhibited a 75% reduction in infection compared with normal macrophage. We also obtained activated macrophage from immune mice and found that they were refractory to in vitro rickettsial infection. Macrophage populations activated in vitro or in vivo contained a small percentage of cells which supported unrestrained growth of rickettsiae. These data suggest that an early immunological event in experimental scrub typhus infection may be the development of activated macrophage capable of suppressing rickettsial proliferation before the appearance of circulating antibody.

Animals

Host defenses in experimental scrub typhus: histopathological correlates.

Intraperitoneal (i.p.) infection of BALB/c mice with 1,000 50% mouse lethal doses of the Karp strain of Rickettsia tsutsugamushi was inevitably lethal, and associated pathological alterations were confined to the peritoneal cavity. These included: (i) continuous proliferation of rickettsial organisms in peritoneal macrophages until death; (ii) hepatic granulomas appearing 6 days after infection and increasing in size and number until death; (iii) splenomegaly, resulting principally from proliferation of lymphoid tissue, and (iv) terminal peritonitis. Under two circumstances, i.p. infections with R. tsutsugamushi were not lethal: (i) infection with 100 50% mouse infectious doses of the Gilliam strain, which, in fact, resulted in immune protection against otherwise lethal Karp challenge; and (ii) Karp infection of animals immunized with the Gilliam strain. In both cases, the associated pathological abnormalities were, as with primary Karp infection, restricted to the peritoneal cavity. Also similar was the striking splenomegaly due to lymphoid proliferation, which was particularly prominent in immunized animals. In contrast to primary and lethal Karp infection, however, these infections were characterized by: (i) minimal and transient proliferation of rickettsial organisms in peritoneal macrophages; (ii) disappearance of hepatic granulomas; and (iii) absence of peritonitis. It was concluded that the survival of an animal bearing an i.p. infection of scrub typhus depended on its ability to concentrate a sufficiently vigorous immune response in the peritoneal cavity, resulting in the evolution of rickettsiacidal macrophages capable of suppressing the infection.

Animals

Protection against scrub typhus infection engendered by the passive transfer of immune sera.

The passive transfer of convalescent sera did not protect the majority of mice against challenge with the homologous strain and was completely ineffective against challenge with strains unrelated by fluorescent antibody techniques. When the immune sera was incubated with the rickettsia in vitro and then inoculated into the mice a dramatic increase occurred in the number of surviving mice. The importance of these data in relation to published results with other species of rickettsia is discussed.

Animals

Distribution of rats infected with Rickettsia tsutsugamushi (scrub typhus) in an edge habitat.

No focalization of rats (Rattus tiomanicus and R. argentiventer) infected with Rickettsia tsutsugamushi could be discerned over a 500 m trapping transect at the border between a forest and lalang grass (Imperata cylindrica). R. tiomanicus appeared to occupy 250 m of the transect on the average and had periods during which infections were observed which averaged 97 days. Calulations indicated that more than 50% of individuals become infected over their life-time. The high rate of infection in this and other areas described in earlier publications and the habits of the rats suggest that infected mites are densely and widely dispersed in the areas studied in Malaysia.

Animals

Experimental infection of mouse peritoneal mesothelium with scrub typhus rickettsiae: an ultrastructural study.

The infection cycle of Rickettsia tsutsugamushi in mouse peritoneal mesothelial cells, observed late in the course of an established infection, intimately involved the host cell plasma membrane. Organisms multiplied in the cytoplasm, moved to the cell periphery, and acquired a host-membrane coat as they budded from the cell surface. Rickettsiae enveloped by this membrane entered other mesothelial cells, apparently by a phagocytic mechanism. Organisms escaped from the phagocytic vacuole as the vacuole membrane and host membrane coat disintegrated. Free rickettsiae replicated by binary fission in the cell cytoplasm. Rickettsial infection of mesothelial cells induced conspicuous cellular hypertrophy with increased numbers of unaltered cytoplasmic organelles.

Animals

Sero-epidemiological survey of scrub typhus.

A total of 1017 human sera from ecologically different areas in the states of Rajasthan and Jammu & Kashmir were subjected to the CF test using the Gilliam strain antigen of R. tsutsugamushi. Results revealed that 2.4% and 3.5% of the populations examined in the two states respectively, were found to possess antibodies to the test antigen. The titres of positive sera varied from 1:8 to 1:32. Findings of this survey could be correlated with the local topography, presence or absence of vector and the exposure of susceptible population to the rickettsial agent. However, the presence of hidden foci in the neighbouring areas not covered by the current survey cannot be ruled out.

Antibodies, Bacterial