Keith J. Sumption

Control and eradication of foot-and-mouth disease.

Paul Sutmoller *, Simon S. Barteling , Raul Casas Olascoaga , Keith J. Sumption d a Animal Health Consultant, former chief of Laboratories of the Panamerican Foot and Mouth Disease Center PAHO/WHO, Brazil b Consultant Veterinary Vaccines, former Head Department FMD Vaccine Development, and Production ID-Lelystad and former Head Community Co-ordinating Institute (for the EU), The Netherlands c Direct Advisor of the Minister of Livestock, Agriculture and Fisheries, Uruguay, former Director of the Panamerican Foot and Mouth Disease Center PAHO/WHO, Brazil d Lecturer in International Animal Health, Centre for Tropical Veterinary Medicine, University of Edinburgh, Easter Bush, Roslin, Midlothian EH25 9RG, Scotland, UK

Natural prevalence of infection with Ehrlichia (Cytoecetes) phagocytophila of Ixodes ricinus ticks in Scotland.

Ixodes ricinus nymphs and adults were collected from vegetation and from sheep at four sites in Scotland typical of areas endemic for tick-borne fever in sheep caused by infection with Ehrlichia (Cytoecetes) phagocytophila (Rickettsiales). The great majority of ticks examined was from woodland sites adjacent to sheep farms where there was a high probability of them feeding on roe deer (Capreolus capreolus) in a non-domestic focus of infestation and infection. Ticks were examined for infection by five methods. Batches of ticks were examined either by feeding on susceptible sheep or by feeding on rabbits and then prepared as stabilate which was inoculated into susceptible sheep. The sheep were monitored for clinical signs of tick borne fever. Batches of ticks were examined by polymerase chain reaction for Ehrlichia phagocytophila. Salivary glands were dissected out and stained by the Feulgen method to detect Ehrlichia masses, and were examined by indirect fluorescent antibody test. Each of the methods detected infection in ticks and the prevalence of infection in nymphs with the various methods ranged from >0.25% to 2.0%. Small samples of adults examined by Feulgen staining of salivary glands indicated infection prevalences of 2.1% in males and 1.6% in females. It is considered that these low infection prevalences may be typical of natural foci of infection where deer could be a major host of ticks and E. phagocytophila.

The Cowdria ruminantium groE operon

A Cowdria ruminantium genomic DNA library was constructed in the expression vector lambda ZAPII, and an immunoreactive clone, designated lambda Cr9.4, was isolated by screening with serum from a C. ruminantium-infected goat. Sequencing of the insert from this clone revealed two open reading frames, encoding peptides of 10462 and 58697 kDa respectively. Database searching indicated that the two genes were homologues of groES and groEL, genes encoding a group of heat shock proteins involved in protein processing, export and assembly. Western blotting experiments showed that the recombinant GroEL protein was recognized by sera raised against four isolates of C. ruminantium which originate from South Africa, West Africa and the Caribbean, but not by antisera to the closely related Ehrlichia species (E. ovina, E. [Cytoecetes] ondiri, E. bovis, E. phagocytophila) of African and European ruminants which can be expected to occur in similar geographical areas to C. ruminantium. This suggests that this protein may be useful in development of serodiagnostic tests for C. ruminantium infection which are not subject to cross-reactions with antibodies to Ehrlichia species. The cloning and expression of the GroE operon will also facilitate further study of the roles of the GroE proteins in the immune response to C. ruminantium.

Development of a Polyclonal Competitive Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Ehrlichia ruminantium

ABSTRACT A polyclonal competitive enzyme-linked immunosorbent assay (PC-ELISA) is described for detection of antibodies to Ehrlichia (Cowdria) ruminantium by using a soluble extract of endothelial cell culture-derived E. ruminantium as the antigen and biotin-labeled polyclonal goat immunoglobulins as the competitor. For goats, the diagnostic sensitivity and specificity were both 100% with a cutoff of 80% inhibition (80 PI), with detection of antibodies for 550 days postinfection. For cattle, diagnostic sensitivity and specificity were 86 and 100%, respectively, with a cutoff of 50 PI and 79 and 100% with a cutoff of 70 PI. Cross-reactions with high-titer experimental or field antisera to other Ehrlichia and Anaplasma species were observed at up to 68 PI in cattle and up to 85 PI in sheep, and therefore to exclude these cross-reactions, cutoffs of 70 PI for bovine serology and 85 PI for small-ruminant serology were selected. Application of the PC-ELISA to bovine field sera from South Africa gave a higher proportion of positive results than application of the murine macrophage immunofluorescent antibody test or indirect ELISA, suggesting a better sensitivity for detection of recovered cattle, and results with bovine field sera from Malawi were consistent with the observed endemic state of heartwater and the level of tick control practiced at the sample sites. Reproducibility was high, with average standard deviations intraplate of 1.2 PI and interplate of 0.6 PI. The test format is simple, and the test is economical to perform and has a level of sensitivity for detection of low-titer positive bovine sera that may prove to be of value in epidemiological studies on heartwater.

Detection by Two Enzyme-Linked Immunosorbent Assays of Antibodies to Ehrlichia ruminantium in Field Sera Collected from Sheep and Cattle in Ghana

ABSTRACT Two serological tests for detection of antibodies to Ehrlichia (previously Cowdria) ruminantium, the causative agent of heartwater, were compared by using field sera collected from sheep and cattle as part of serosurveys in Ghana. Sera selected as either negative or positive by a new polyclonal competitive enzyme-linked immunosorbent assay (PC-ELISA) were tested by the indirect MAP1-B ELISA. Cutoff values of 14 percent positivity (14 PP) for both ruminant species were obtained for the MAP1-B ELISA by using preseroconversion Ghanaian sera and were compared with previously recommended cutoff values of 29 PP for sheep and 38 PP for cattle. With the 14-PP cutoff, of 151 sheep sera which tested negative by PC-ELISA, 89% were also negative by MAP1-B ELISA, while of 419 sheep sera positive by PC-ELISA, 98% were also positive by MAP1-B ELISA. Of 261 bovine sera negative by PC-ELISA, 82% were also negative by MAP1-B ELISA. Of 511 bovine sera positive by PC-ELISA, only 47% were positive by MAP1-B ELISA; these included 168 sera collected from cattle following first seroconversion as detected by both tests, with 125 of these sera positive by PC-ELISA but only 59 and 5 positive by MAP1-B ELISA with the 14- and 38-PP cutoff levels, respectively. These results indicate that both assays are highly sensitive and specific for detection of E. ruminantium exposure in sheep but that the MAP1-B ELISA lacks sensitivity for postseroconversion bovine sera in comparison to the PC-ELISA. Both tests confirm E. ruminantium seroprevalence of at least 70% in Ghanaian sheep; levels of exposure among Amblyomma variegatum-infested Ghanaian cattle are likely to be higher than the seroprevalence value of 66% obtained with the PC-ELISA.

Immunogenicity of Ehrlichia ruminantium grown in tick cell lines

Ehrlichia (previously Cowdria) ruminantium, the pathogen which causes heartwater in domestic and wild ruminants, can now be propagated in cell lines from one vector (Amblyomma variegatum) and five non-vector (lxodes scapulari.s. I. ricinus, Boophilus decoloratus, B. microplus and Rhipicephalus appendiculatus) tick species. E. ruminantium isolates from West and South Africa and the Caribbean vary in their cell line preference. growth patterns and immunogenic capability. In laboratory trials, certain combinations of tick cell line and E. ruminantium isolate were highly immunogenic in sheep. These trial vaccines were grown under specific in vitro conditions and administered as a single intravenous dose of freshly harvested whole, live culture. Following immunisation and subsequent exposure to virulent E. ruminantium, protected sheep showed no clinical response and a range of serological responses.

Heartwater in Ghana: implications for control of ticks

SummaryHeartwater, an often fatal rickettsial disease of domestic ruminants transmitted byAmblyomma variegatum ticks, ranks with theA. variegatum-associated skin disease dermatophilosis as a major constraint to the upgrading of livestock productivity in Ghana. An epidemiological survey, using new diagnostic tests, is being carried out to determine the incidence and distribution of heartwater and other tickborne diseases in Ghanaian cattle, sheep and goats. Preliminary results from a longitudinal survey being carried out at sites in the Greater Accra Region indicating that although the vector ticks and the disease agent are widespread outside urban areas, not all animals are being exposed to heartwater during the first few months of life when an inverse age related resistance allows development of immunity without clinical disease. Thus a susceptible sub-population, at risk from heartwater, can exist even in areas of high tick challenge. The significance of these results for present and future tick and disease control strategies is discussed.

Kinetics of experimental infection of sheep with Ehrlichia ruminantium cultivated in tick and mammalian cell lines

Inoculation of sheep with Ehrlichia (previously Cowdria) ruminantium which has been cultivated in mammalian endothelial cell cultures is almost always followed by a severe clinical reaction, whereas inoculation of the agent cultivated in tick cell lines usually does not provoke a clinical response, but may result in seroconversion and/or protection against subsequent challenge with virulent stabilates. A quantitative, real-time PCR assay was developed to determine the kinetics of infection (rickettsaemia) in sheep inoculated with tick cell- and mammalian cell-derived E. ruminantium (Gardel isolate). The method and initial results are described, and the significance of the findings is discussed in relation to the clinical responses of the sheep.

The control of heartwater in west Africa—Present and future

SummaryHeartwater (HW) is the name given to the acute, febrile disease of ruminants caused by infection with the obligate intra-cellular rickettsiaCowdria ruminantium. The effect of losses associated with HW and dermatophilosis are an important constraint to the sustainable use of non-indigenous breeds, but recent advances in heartwater and dermatophilosis research provide optimism that integrated control of these conditions can be achieved in west Africa. The relevant advances in HW research are in methods for monitoring the status of animals at risk of developing heartwater, which allow the veterinarian and owner to identify control strategies for herds and individual animals, and developments in the field of vaccination. This paper seeks to provide information that may assist the veterinary services in west Africa in assessing and identifying opportunities for HW control. Recommendations for the activities of key institutions place an emphasis upon the isolation and identification of west African strains ofCowdria for inclusion in inactivated or live vaccines, and the need for donor support to enable field trials of integrated control methods for HW and dermatophilosis.