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Soluble antigen obtained by the treatment of Coxiella burneti of the I phase with trichloracetic acid could be used in the capacity of a precipitinogen in the band-precipitation reaction in the diagnosis of Q-rickettsiosis and in mass epidemiological studies.
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BACKGROUND: Long-term persistence of C. burnetii in infected animals was established in the 1950s and 60s, but the implications for human Q fever are not fully explored. AIM: To compare the prevalence of markers of infection in a cohort of Q fever patients in Australia (up to 5 years after infection) with those in the 1989 Birmingham cohort (12 years after infection). DESIGN: Case follow-up study. METHODS: C. burnetii was tested for by: (i) antibodies to Phase 1 and 2 antigens in the three immunoglobulin classes; (ii) detection of DNA in bone marrow and peripheral blood mononuclear cells by PCR assays directed against several different targets in the genome; and (iii) attempts to isolate coxiellas in cell culture or mice from PCR-positive samples. Amplicon specificity was verified by fluorometric probing and by sequencing. Cross-contamination was excluded by extensive use of non-template controls, and in particular by the use of certain IS1111a target sequences. RESULTS: Irrespective of clinical state, both groups remained seropositive, principally exhibiting medium levels of IgG antibody against C. burnetii Phase 2 antigen. C. burnetii genomic DNA was detected by PCR in 65% of bone marrow aspirates from Australian patients and approximately 88% of Birmingham patients. No coxiella were isolated from PCR positive samples. DISCUSSION: We propose a provisional model for persistence. In Q fever without sequelae, the process is largely confined to the bone marrow. In Q fever fatigue syndrome (QFS), it is modulated by the patient's immunogenetic background to give higher levels of coxiella genomes in bone marrow and increased shedding into the peripheral blood. In Q fever endocarditis, late pregnancy, or during iatrogenic or other immunosuppression, the multiplication cycle is prolonged, and a potential source of live organisms.
The lysosomal response of a murine macrophage-like tumor cell line (J774) during persistent infection with Coxiella burnettii was examined. By using acid phosphatase as a lysosomal marker, it was shown that phagosome-lysosome fusion occurred in J774 cells persistently infected with C. burnetii. This observation was verified using thorium dioxide, an electron-dense compound that is sequestered in secondary lysosomes. The phagolysosomes contained viable replicating rickettsiae. Spectrofluorometric analysis indicated that the phagolysosomal pH of persistently infected cells was acidic. In attempts to correlate rickettsial survival with lysosome function, the activities of several lysosomal enzymes were assayed in both infected and uninfected cells. Activities of acid phosphatase and beta-acetylglucosaminidase were not significantly altered during infection. However, infected cells appeared to display slightly higher intracellular lysozyme, beta-glucuronidase, and beta-galactosidase activities.
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Between 1987-1989 almost 7000 adult Ixodes ricinus, Dermacentor reticulatus, Dermacentor marginatus, Haemaphysalis concinna, Haemaphysalis punctata and Haemaphysalis inermis ticks collected in all 38 districts of Slovakia were screened for the presence of Coxiella burnettii. The proportion of ticks containing C. burnetii as indicated by the haemocyte test was less than 3%. Attempts to recover C. burnetii by inoculation of yolk sacs of embryonated hen eggs from pools of 1-6 specimens of haemocyte test positive ticks resulted in the isolation of 10 rickettsial strains. Six strains were recovered from I. ricinus, the remaining ones from single pools of D. reticulatus, D. marginatus. H. concinna and H. inermis ticks. In addition to the previous recovery of C. burnetii from H. punctata ticks, the agent was thus isolated from all important ticks living in Slovakia. The agent was found in tick habitats regardless of the latitude and altitude in the entire country. These results are not consistent with the negligible number of Q fever cases occurring in past years in Slovakia.
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The development of humoral and cell-mediated immune responses was studied in guinea pigs infected with Coxiella burnetti administered in small-particle aerosols. Direct macrophage migration inhibition was observed in cultured peritoneal exudate cells as early as 3 days after exposure. Maximum inhibition of macrophages cultured with phase I or II antigen occurred 14 to 21 days postexposure and persisted through 35 days. This inhibitory action was no longer detectable at 42 days. Serum antibody to the phase II antigen of C. burnetii was detected at 14 days, and serum antibody to phase I antigen was detected at 21 days, 18 days after the cell-mediated immune response.
L929 mouse fibroblast cells and J774 macrophage-like cells are both susceptible to persistent infection with the Q fever agent Coxiella burnetti. Previously this laboratory has shown that persistently infected cell populations multiply with unaltered generation times or cell cycle progression. It has also been reported by others and us that highly infected cells typically exhibit one large parasite-containing vacuole. We now report that lightly and heavily infected cells are capable of division and in the process segregate the parasite-containing vacuole into one of the emerging daughter cells; the companion daughter cell emerges parasite-free. This asymmetric division of infected cells, revealed via photomicrography of stained cells, accounts for the appearance of uninfected cells within persistently infected host cell populations that were previously 100% infected. Some of the persistently infected L929 populations were maintained in culture for over two years without the addition of normal cells.
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