Biomedical subjects

F Jongejan

Publications and source records attributed to F Jongejan.

At least 55 records · Page 3Linked to original sources

PubMed1993

Correlation between antibodies to Cowdria ruminantium (rickettsiales) in cattle and the distribution of Amblyomma vector ticks in Zimbabwe.

Cowdriosis, caused by Cowdria ruminantium, is transmitted by Amblyomma ticks, which are widely distributed in Zimbabwe. To assess the distribution of this disease in Zimbabwe, cattle either exposed to Amblyomma ticks or maintained in areas free from these ticks were tested for antibodies to Cowdria. A total of 324 sera were tested using competitive ELISA and the indirect fluorescent antibody test (IFAT). At diptanks in Amblyomma-infested areas 52% (n = 95) and 26% (n = 47) of sera were positive by cELISA and IFAT, respectively. At diptanks in Amblyomma-free areas 11% (n = 125) and 10% (n = 134) of sera were positive by cELISA and IFAT, respectively. The results were significantly different between Amblyomma-infested and tick-free areas (chi 2 = 24.73, P < or = 0.005 for IFAT and chi 2 = 57.53, P < or = 0.005 for cELISA). High background readings in field sera, possibly due to cross-reactive antibodies to Ehrlichia spp., complicated the determination of a realistic cut-off point, especially in cELISA. On the basis of the distribution of Amblyomma ticks, currently a large part of Zimbabwe can be considered endemic for the disease.

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PubMed1993

Detection of the protozoan parasite Theileria annulata in Hyalomma ticks by the polymerase chain reaction.

Adult Hyalomma ticks were examined for the presence of Theileria annulata infection using the Polymerase Chain Reaction (PCR). A 372 bp DNA fragment derived from the small ribosomal RNA gene of T. annulata was amplified from 45 out of 50 (90%) H. dromedarii ticks and from 36 out of 50 (72%) H. marginatum marginatum ticks. No product was amplified from non-infected control ticks. Restriction enzyme digestion with Sac II confirmed that the product was derived from the targeted T. annulata gene. As a further confirmation it was shown that both species of Hyalomma ticks were able to transmit T. annulata to experimental calves. PCR detection of Theileria parasites in ticks was compared with conventional staining of dissected salivary glands using methyl green pyronin and its comparative advantages are discussed.

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PubMed1993

Antibodies to Cowdria ruminantium in Mozambican goats and cattle detected by immunofluorescence using endothelial cell culture antigen.

Endothelial cell cultures, established from bovine umbilical cord arteries and subsequently infected with Cowdria ruminantium, were used as antigen in the indirect fluorescent antibody test. Bovine sera (374) and caprine sera (388) collected in 6 provinces of Mozambique were tested. Overall, 30.4% of goat sera had antibodies to Cowdria, and 43% of sera collected from cattle. North of the River Save, where the tick Amblyomma variegatum is highly prevalent, overall percentages of positive sera were low, 10% in goats and 20% in cattle. However, south of the river where the tick Amblyomma hebraeum is abundant percentages were much higher, 63.5% in goats and 59.4% in cattle. These results are discussed in relation to field observations that clinical disease is rare or absent in the north with enzootic instability in goats and Friesian calves in the south.

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PubMed1993

Detection of antibodies to Cowdria ruminantium in the serum of domestic ruminants by indirect ELISA.

A solid phase enzyme immunoassay for the detection of antibodies to Cowdria ruminantium in the serum of domestic ruminants was developed by using microorganisms cultivated on bovine umbilical endothelial cells as antigen. When the culture showed 90% lysis, the supernatant was centrifuged, sonicated and coated on polystyrene microtiter plates. Antibodies were detected as early as 9 days after experimental immunization of goats. The sensitivity of the ELISA calculated with 73 ruminant sera ranged between 97.3% and 98.6%. The overall specificity of the test was 97% (N = 2925). However, the specificity was far lower for sheep (94.4%, N = 881) than for goats (98.6%, N = 651) and cattle (97.8%, N = 1393). Crossreactivity which could explain some of the false positive reactions, was found between Cowdria antigen and sera raised against Ehrlichia bovis (1 bovine positive out of 2 inoculated) or E. ovina (2 sheep out of 2 inoculated became positive) but not with E. phagocytophila. The intra-assay and inter-assay variability were 7.5% and 7.8% respectively, indicating a good reproducibility of the ELISA.

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PubMed1993

The sero-diagnosis of heartwater: a comparison of five tests.

Five serological tests, the indirect and competitive ELISA, the indirect fluorescent antibody (IFA) test with 2 different antigens and the Western blot technique were compared and applied to sera that were known to be either negative or positive against Cowdria ruminantium or that were collected from animals in heartwater-free regions. No false positive reactions were recorded with any of the tests against the known negative sera. Except for minor variations in the sensitivity of the 5 tests, there was good correlation between them. Their specificity, however, remains in dispute since in all 5 tests extensive cross-reactions were recorded with antibodies in response to an as yet unidentified agent, probably Ehrlichia.

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PubMed1993

The immunodominant 32-kilodalton protein of Cowdria ruminantium is conserved within the genus Ehrlichia.

Serological tests for cowdriosis are hampered by cross-reacting antibodies from animals suspected to be infected with Ehrlichia species. We have monitored infections with Ehrlichia bovis, E. ovina, E. canis and E. phagocytophila in experimental animals by competitive ELISA, Western blotting and immunofluorescence using Cowdria-infected endothelial cell culture antigens. Cross-reactions due to Ehrlichia antibodies could be attributed to the recognition of epitopes on the immunodominant Cr32 Cowdria protein. This was especially true for E. canis and E. ovina, much less for E. bovis, but not at all for E. phagocytophila. In addition, strong cross-reactivity between Cowdria and antibodies to E. Chaffeenis were demonstrated. These findings are in agreement with the phylogenetic relationships, recently reported by van Vliet et al. in 1992, between Cowdria and other members of the tribe Ehrlichieae, which showed Cowdria to be closely related to E. canis and also to E. chaffeensis. Although the tests used in this study remain valuable tools under laboratory conditions, their specificity requires improvement. It is suggested to study recombinant Cowdria antigens for the development of second generation serological tests for cowdriosis.

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PubMed1993

Cloning and partial characterization of the Cr32 gene of Cowdria ruminantium.

Cowdria organisms were purified by density gradient centrifugation. The DNA was used to construct expression libraries. The immunodominant Cr32 protein was purified and its N-terminal amino acid sequence was determined. The expression libraries were screened with Cr32-specific monoclonal antibodies, but did not yield Cr32-positive clones. Therefore a part of the Cr32-gene was amplified using primers derived from the N-terminal and an internal amino acid sequence. This DNA was used as a probe to detect the genomic DNA fragment encoding the Cr32 protein. This fragment was cloned, using genomic DNA of the Senegal strain of Cowdria ruminantium. A part of the gene comprising two third of its total length has been expressed in vector pGEX2T. This expression product is recognized by Cr32-specific monoclonal antibodies.

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PubMed1993

Use of serological response to evaluate heartwater immunization of cattle.

Frozen blood vaccine containing the Ball 3 strain of Cowdria ruminantium is prepared by an FAO/DANIDA Project in Malawi for the immunization of improved dairy cattle against heartwater. The immunogenicity of vaccine batches for cattle has been quantified and different regimens for immunization have been evaluated using indirect immunofluorescence to assess antibody responses. Infected endothelial cells grown in culture are used as antigen. The proportion of animals responding serologically has varied between different batches of vaccine tested in homogeneous cattle populations at the same time, presumably reflecting differences in immunogenicity of batches. The proportion of animals in a homogeneous population responding serologically to the same vaccine batch administered under different regimens has also varied. Indirect immunofluorescence testing has proved to be a useful method for assessing the immune response of cattle to immunization and has been adopted as a routine quality control procedure for heartwater vaccine production in Malawi.

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PubMed1993

Vaccination against heartwater using in vitro attenuated Cowdria ruminantium organisms.

Sequential passage of Cowdria ruminantium (Senegal isolate) in cultures of bovine umbilical endothelial cells has resulted in loss of virulence without loss of immunogenicity, as previously demonstrated. We have carried out further immunization of 39 Dutch sheep using in vitro attenuated rickettsiae of passage 21 and challenged these animals either with the homologous or with heterologous Cowdria stocks. After vaccination several sheep developed elevated rectal temperatures for a maximum of 2 days, but no further clinical response to the vaccine was observed. All sheep developed high titres of antibodies to Cowdria. Challenge of 10 sheep with the homologous virulent stock did not provoke any clinical reaction, demonstrating that these animals were solidly immune. Reactions to heterologous challenge varied from virtually no reaction to fatal heartwater depending on the stock of Cowdria used. These results are discussed in relation to currently available vaccination methods against cowdriosis. In Senegal 30 susceptible sahelian sheep were immunized with attenuated rickettsiae of passage 21. Hyperthermia was seen in 13, the only other clinical symptom was a temporary diarrhoea. The immunized animals are at present exposed, together with 30 controls, to field challenge in the Niayes, the area where the Senegal isolate was originally isolated.

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PubMed1992

Experimental transmission of Cowdria ruminantium (Rickettsiales) by the American reptile tick Amblyomma dissimile Koch, 1844.

A Senegalese isolate of the rickettsia Cowdria ruminantium was transmitted transstadially by nymphs of the American reptile tick Amblyomma dissimile. Only eight nymphs, fed as larvae on a Saanen goat reacting to heartwater, were required to transmit fatal heartwater to another susceptible goat. Since A. dissimile usually feeds on snakes, iguanas and lizards in central America, the tick is not considered to play a significant role in the transmission of heartwater between ruminants. However, the tick could play a role in maintaining a rickettsial reservoir in reptile populations, since it has been shown that an African reptile can be a subclinical carrier of C. ruminantium, infective to vector ticks.

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PubMed1992

Phylogenetic position of Cowdria ruminantium (Rickettsiales) determined by analysis of amplified 16S ribosomal DNA sequences.

The 16S ribosomal DNA sequence of Cowdria ruminantium, the causative agent of heartwater disease in ruminants, was determined. An analysis of this sequence showed that C. ruminantium forms a tight phylogenetic cluster with the canine pathogen Ehrlichia canis and the human pathogen Ehrlichia chaffeensis. Although a close relationship between the genus Cowdria and several members of the tribe Ehrlichieae has been suspected previously, the tight phylogenetic cluster with E. canis and E. chaffeensis is surprising in view of known differences in host preference and target cells.

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PubMed1992

Distribution of heartwater in the Caribbean determined on the basis of detection of antibodies to the conserved 32-kilodalton protein of Cowdria ruminantium.

A competitive enzyme-linked immunosorbent assay (cELISA) was developed to detect immunoglobulin G antibodies to the major 32-kDa protein of Cowdria ruminantium. A total of 1,804 serum samples collected from cattle on 19 islands in the eastern Caribbean Basin were tested by this cELISA. A total of 133 serum samples from 10 islands (Antigua, Dominica, Grenada, Guadeloupe, Martinique, Montserrat, St. Kitts, St. Lucia, St. Martin, and St. Vincent) were found to be positive. The presence of antibodies to C. ruminantium in cattle on these islands was confirmed by immunofluorescence and Western blotting (immunoblotting). In earlier studies, C. ruminantium has been demonstrated only on Guadeloupe, Antigua, and Marie Galante. This study shows that the causative agent of heartwater is now firmly established in the Caribbean.

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PubMed1991

Antigenic diversity of Cowdria ruminantium isolates determined by cross-immunity.

Antigenic diversity in five stocks of the tick-borne rickettsia Cowdria ruminantium, the causal agent of heartwater disease of ruminants, was studied by cross-immunity trials in goats and sheep. Complete absence of cross-protection was found only between the Kümm and Kwanyanga stocks, and in all other combinations there were various degrees of cross-immunity. Immunological strain differences were more pronounced in goats than in sheep.

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PubMed1991

Cowdria ruminantium is recognized by a monoclonal antibody directed against the major outer membrane protein of Chlamydia trachomatis.

The relationship between Cowdria ruminantium and Chlamydia trachomatis was studied by immunofluorescence. A monoclonal antibody directed against the major outer membrane protein of C. trachomatis recognized rickettsial colonies of C. ruminantium in infected goat brain. No specific fluorescence was observed in non-infected brain. Two commercial Chlamydia-specific monoclonal antibodies as well as polyvalent anti-Chlamydia rabbit serum recognized C. trachomatis, but did not recognize Cowdria. Moreover, polyvalent Cowdria antiserum failed to recognize C. trachomatis cultivated in HeLa cells. It is concluded that Cowdria and Chlamydia are to a certain extent related, confirming similarities in ultrastructure and developmental cycle.

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PubMed1991

Competitive enzyme-linked immunosorbent assay for heartwater using monoclonal antibodies to a Cowdria ruminantium-specific 32-kilodalton protein.

Hybridomas producing monoclonal antibodies (mAb) to Cowdria ruminantium were raised. Four mAbs of the IgG isotype reacted in western blots with a 32-kilodalton Cowdria protein (Cr32), which had previously been shown to be conserved and immunodominant. A fifth mAb of the IgM isotype recognized a 40-kDa Cowdria protein. The latter mAb was negative in an indirect fluorescent antibody test (IFA), whereas the other four were positive. mAb No. 4F10B4 showed the strongest signal in western blots using three different stocks of Cowdria. Immuno-gold labeling of Cowdria organisms in vitro using 4F10B4 showed that Cr32 has surface-exposed antigenic determinants. Using mAb 4F10B4, a competitive ELISA was developed which detected specific Cowdria antibodies in goat, sheep and cattle sera. Antibodies in animal sera competed with binding of mAb 4F10B4 to a crude sonicated Cowdria antigen obtained from infected endothelial cell cultures. The competition ELISA (CELISA) detected antibodies in 55 out of 70 (79%) goats experimentally infected with one of eight different Cowdria stocks. Fourteen out of the 15 sera which were shown negative in the CELISA were also negative in the IFA. Nevertheless, all 15 sera recognized some epitopes of the immunodominant Cowdria-specific 32 kDa protein as judged from their reaction with this protein in western blots. Overall, there was 89% agreement between CELISA and IFA considering all 70 goat sera. Moreover, antibodies were detected in nine out of nine sheep infected with one of three different stocks of Cowdria and in sera from calves experimentally infected by two different strains of heartwater. There were no cross-reactions with Ehrlichia phagocytophila antibodies in goat sera, nor with Anaplasma marginale antibodies in bovine sera. Lack of cross-reactivity and detection of antibodies to eight geographically widely distributed stocks of Cowdria, makes the competition ELISA a promising test for use in heartwater endemic areas.

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PubMed1991

Protective immunity to heartwater (Cowdria ruminantium infection) is acquired after vaccination with in vitro-attenuated rickettsiae.

A Senegalese (S) stock of Cowdria ruminantium was passaged on bovine umbilical endothelial cells with an average interval of 13.9 days (range, 8 to 34 days) between passages. The virulence of infected bovine umbilical endothelial cultures was tested in susceptible goats and sheep by intravenous inoculation of culture supernatant from passages 2 (51 days in vitro), 3 (69 days), 11 (229 days), 14 (264 days), and 16 (291 days). Both animals inoculated with passages 2 and 3 died of heartwater. However, clinical reactions were completely absent in goats and sheep that were inoculated with C. ruminantium from passages 11, 14, and 16. High antibody titers were detected, with immunofluorescence in all vaccinated animals, and a strong signal was found against a 32-kDa Cowdria protein in Western blots (immunoblots). Moreover, the vaccinated animals proved solidly immune when challenged with virulent Cowdria sp.-infected blood stabilate (S strain), whereas all control goats died. No attenuation of a second Cowdria stock (W) was achieved after 226 days in culture, at which time passage 17 was tested in a recipient goat which died of typical heartwater. This is the first report of vaccination with live attenuated C. ruminantium. These attenuated organisms may replace vaccination with virulent blood currently in use in areas where heartwater is endemic.

Animals↗

PubMed1991

The tick-borne rickettsia Cowdria ruminantium has a Chlamydia-like developmental cycle.

The development of the tick-borne rickettsial pathogen Cowdria ruminantium (S stock) was studied in bovine umbilical endothelial (BUE) cell cultures and in goat choroid plexus, by light- and electron microscopy. Cowdria divided by binary fission within intracytoplasmic vacuoles resulting in large colonies of reticulate bodies. After three to four days in culture, reticulate bodies developed into smaller intermediate bodies characterized by an electron-dense core. Shortly before disruption of the host cells, intermediate bodies condensed further into electron-dense elementary bodies, which were released into the culture medium. Elementary bodies invade other endothelial cells thus initiating a new infectious cycle which lasts between 5 and 6 days. In the infected goat choroid plexus similar reticulate and intermediate bodies were identified within vacuoles of capillary endothelial cells. However, extracellular elementary bodies were not detected. Another stock of Cowdria (W) showed an identical developmental cycle as that of the S stock. The W isolate was also pathogenic for mice, making it possible to test the infectivity of reticulate and elementary bodies in these animals. Reticulate bodies appeared to be less infective than elementary bodies. The developmental cycle of Cowdria resembles the cycle known to occur in Chlamydia. Moreover, Cowdria has other similarities with Chlamydia. It has a Gram-negative envelope, it does not store iodine-stainable carbohydrates and may lack peptidoglycan as does Chlamydia. It is concluded, that Cowdria and Chlamydia are to a certain extent related, confirming a recent report that both organisms have certain antigenic determinants in common. Since Cowdria is also related to Ehrlichia it may well be that Cowdria takes an intermediate position between Chlamydia and Ehrlichia. The phylogenetic relationship between Cowdria and Chlamydia and also with Ehrlichia should be further elucidated by molecular analysis using 16S ribosomal DNA sequences.

Animals↗

PubMed1991

Lack of cross-protection between Cowdria ruminantium and Ehrlichia phagocytophila.

Antigenically distinct stocks of Cowdria ruminatium from Senegal and South Africa were compared with a Dutch isolate of Ehrlichia phagocytophila in cross-immunity trials in goats. There was a complete absence of cross-immunity between E. phagocytophila and C. ruminantium, despite previous observations that both rickettsial organisms have certain antigenic determinants in common.

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