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1.  Monitoring Babesia bovis infections in cattle by using PCR-based tests. 
Journal of Clinical Microbiology  1996;34(11):2748-2755.
The sensitivity and specificity of PCR tests based on the small-subunit rRNA gene sequence of Babesia bovis were compared in a blind study of experimentally infected cattle with the corresponding parameters of the complement fixation (CF) test currently used in the United States to screen for bovine babesiosis. Cattle were experimentally infected with a single inoculum of a cloned laboratory strain of B. bovis. Blood samples were collected and tested over a period covering from the day of infection to 10 months postinfection. The level of parasitemia (percent infected erythrocytes) present in each sample was estimated from test results and was plotted as a function of time postinfection. These data are the first describing the course of infection by methods capable of detecting parasitemias in the range of 10(-7)%, which frequently occur in the carrier state. Parasitemias in the samples tested strongly influenced the sensitivity and negative predictive value of the PCR-based tests which varied with time postinfection. The average sensitivities of the three PCR-based tests for B. bovis ranged from 58 to 70% for a single determination, while the sensitivity of the CF test was only 6%. Both PCR-based and CF tests for B. bovis had high specificity values ranging from 96 to 100%.
PMCID: PMC229398  PMID: 8897177
2.  Laboratory reared Amblyomma hebraeum and Amblyomma variegatum ticks differ in their susceptibility to infection with Cowdria ruminantium. 
Epidemiology and Infection  1995;115(2):345-353.
The susceptibility of laboratory reared Zimbabwean Amblyomma hebraeum and A. variegatum ticks to infection with geographically distinct Cowdria ruminantium strains was investigated by feeding both species simultaneously on individual sheep infected with one of the four strains (Crystal Springs [Zimbabwe], Ball 3 [South Africa], Gardel [Guadeloupe] and Nigeria [Nigeria]). A. hebraeum ticks demonstrated a high susceptibility to infection with all four C. ruminantium strains. In comparison, A. variegatum were less susceptible to infection with the Crystal Springs and Ball 3 strains (P < 0.001), but showed a similar susceptibility to the Gardel and Nigeria strains. The differences in susceptibility of A. variegatum to infection with the four strains of C. ruminantium correlated with the origin of these strains. The consistently higher susceptibility of A. hebraeum ticks to infection with geographically different C. ruminantium strains may be one explanation for the observation that heartwater is a more serious problem where A. hebraeum is the vector of the disease.
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PMCID: PMC2271405  PMID: 7589273
3.  Development and evaluation of PCR assay for detection of low levels of Cowdria ruminantium infection in Amblyomma ticks not detected by DNA probe. 
Journal of Clinical Microbiology  1995;33(1):166-172.
The sensitivities of a PCR assay and a DNA probe assay were compared for the detection of Cowdria ruminantium in Amblyomma ticks that were fed on C. ruminantium-infected, clinically reacting, and recovered carrier animals. The PCR assay and DNA probe detected infection in 86.0 and 37.0%, respectively, of 100 ticks fed on a febrile animal. In 75 ticks fed on carrier animals, PCR and the DNA probe detected infection in 28.0 and 1.33% of ticks, respectively. This demonstrates that the DNA probe has poor sensitivity for the detection of low levels of infection in ticks and that PCR is necessary for this purpose. The PCR assay had a detection limit of between 1 and 10 C. ruminantium organisms and did not amplify DNA from Ehrlichia canis, which is phylogenetically closely related to C. ruminantium, Theileria parva, or uninfected Amblyomma hebraeum or A. variegatum. PCR detected infection in A. hebraeum and A. variegatum adult ticks infected with one of six geographically different C. ruminantium strains. Amplification was also possible from desiccated ticks and ticks fixed in 70% ethanol, 10% buffered formalin, or 2% glutaraldehyde. The PCR assay supersedes the DNA probe and older detection methods for the detection of C. ruminantium in ticks, particularly those fed on carrier animals, and is suitable for both prospective and retrospective studies which require accurate detection of C. ruminantium in individual ticks. Application of the PCR assay should significantly improve the understanding of heartwater epidemiology, particularly through the determination of field tick infection rates.
PMCID: PMC227901  PMID: 7699036

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