Long-term follow-up of owned, free-roaming dogs in South Africa naturally exposed to Babesia rossi.

Babesia rossi is an important, tick-borne intraerythrocytic protozoan parasite; however, its natural history and epidemiology is poorly understood. Babesia rossi is the most virulent Babesia sp. in domestic dogs and is generally considered to cause severe babesiosis, which is fatal if left untreated. However, subclinical infections and mild disease from B. rossi have been reported, although the clinical progression of these cases was not reported. Therefore, to better understand B. rossi under field conditions, we evaluated its clinical progression and seroprevalence in an owned, free-roaming dog population in Zenzele, South Africa, where the parasite is endemic and prevention is not routine. The entire dog population in Zenzele was monitored intensively at the individual level from March 2008 until April 2014, primarily for a longitudinal study on rabies control. Subsequent evaluation of B. rossi comprised analyses of clinical and laboratory data collected from the Zenzele dog population during the 6 year study period. A substantial proportion (31% (n = 34)) of 109 dogs (randomly selected from every available dog in February/March 2010 older than ~6-8 weeks (n = 246)) tested by Indirect Fluorescent Antibody Test had seroconverted strongly to B. rossi. All 34 dogs were generally consistently healthy adults, determined from regular clinical examinations between March 2008 and April 2014. Blood smear examinations at multiple time points between July 2009 and February 2011 were also undertaken for almost all of these (34) seropositive dogs and all those tested were consistently negative for Babesia spp. Subclinical infections and mild disease were also the main findings for a separate group of 18 dogs positive for Babesia spp. on blood smear examination and confirmed to be infected with B. rossi by Polymerase Chain Reaction - Reverse Line Blot. Almost all of these dogs were positive at only one time point from repeat blood smear examinations between July 2009 and February 2011. We suggest that these observations are consistent with immunity acquired from repeated, low-level exposure to the parasite, generating transient subclinical infections or mild disease. Should this be the case, the use of tick control, particularly in adult dogs in free-roaming populations in B. rossi endemic regions, should be carefully considered.

Hemostatic abnormalities in uncomplicated babesiosis (Babesia rossi) in dogs.

BACKGROUND: Babesiosis in dogs is associated with severe thrombocytopenia; yet infected dogs rarely show clinical signs of hemorrhage. HYPOTHESIS: Dogs with uncomplicated babesiosis have normal hemostatic capacity despite severe thrombocytopenia. ANIMALS: Nineteen client-owned dogs with uncomplicated babesiosis; 10 healthy controls. METHODS: A prospective, cross-sectional, observational study. Thromboelastography (TEG), prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen, D-dimers, and antithrombin (AT) measured in both groups. Statistical significance set at P < .01. RESULTS: Babesiosis group hematocrit and platelet count significantly lower than controls (0.29 versus 0.50 L/L; P < .001 and 20.0 versus 374.5 × 10(9)/L; P < .001, respectively). Except for K, no significant difference in TEG variables between groups. Hemostatic variables for babesiosis group versus controls (mean ± SD); R: 5.9 ± 1.8 versus 4.6 ± 0.9 min (P = .048); K: 2.8 ± 1.1 versus 1.9 ± 0.6 min (P = .003); angle: 55.5 ± 11.7 versus 62.2 ± 4.1 degrees (P = .036); MA: 48.4 ± 9.7 versus 57.2 ± 5.2 mm (P = 0.013); G: 5.1 ± 1.9 versus 6.9 ± 1.5 dyn/cm(2) (P = .019); LY30 (median, range): 0 (0-5.7) versus 0.6% (0-6.1) (P = .152); and LY60: 0 (0-8.8) versus 3.1% (0-13.1) (P = .012). AT activity significantly lower (105.2 ± 16.5 versus 127.8 ± 15.4%; P = .001). Fibrinogen concentration significantly higher in babesiosis group (5.7 ± 1.3 versus. 3.0 ± 0.7 g/L; P < .001). CONCLUSION AND CLINICAL IMPORTANCE: Despite severe thrombocytopenia, dogs with uncomplicated babesiosis did not have clinical signs of hemorrhage and TEG variables were normal, which could indicate a normocoagulable state.

C-reactive protein in canine babesiosis caused by Babesia rossi and its association with outcome.

C-reactive protein (CRP) is a positive major acute-phase protein in dogs and can be used as a predictive marker for risk of disease and to monitor the response to treatment. Increased concentrations in certain diseases are associated with poor outcome. This cross-sectional, observational study of 75 dogs naturally infected with Babesia rossi was designed to examine the relationship between outcome and CRP concentration at admission and the magnitude of CRP change 24 hours after admission. Diagnosis was confirmed by polymerase chain reaction (PCR) and reverse line blot. CRP concentrations were determined by an automated human CRP Turbidometric Immunoassay, previously validated for use in dogs. There was no significant difference in mean CRP concentration between survivors (n = 57), 107.5 +/- 49.5 mg/l and non-survivors (n = 11), 122.1 +/- 64.6 mg/l at admission and using the exact logistic regression, adjusting for age and sex, there was no association with outcome (P = 0.53). Multiple regression analysis failed to show a significant relationship between admission CRP concentration and number of days of hospitalisation in the survivors, adjusting for age and sex (P = 0.65). Similarly, no significance was found in the relationship between the magnitude of change in CRP concentration 24 hours after admission, and the number of days of hospitalisation in survivors, (P = 0.34). It is concluded that CRP concentration, as a measure of the acute phase response, is not associated with outcome in canine babesiosis, and inflammation is unlikely to be the only cause of severity of disease.

Preliminary evaluation of the BrEMA1 gene as a tool for associating babesia rossi genotypes and clinical manifestation of canine Babesiosis.

Babesia rossi, an intraerythrocytic protozoan, causes a severe, often life-threatening disease of domestic dogs. Dogs treated early for B. rossi infection usually recover from the disease, but dogs left untreated or treated at a later stage of infection seldom survive. Dogs infected with B. rossi have varied clinical manifestations that can be categorized as uncomplicated (with a good prognosis) or complicated (with a poor prognosis). One hundred twenty-one blood samples were collected from dogs presented to the Onderstepoort Veterinary Academic Hospital and diagnosed with babesiosis by the use of a thin blood smear. An additional 20 samples were obtained from Babesia-infected dogs from private clinics around the Onderstepoort, Johannesburg, Durban, White River, and Cape Town areas. The samples were screened by PCR targeting the Babesia rossi erythrocyte membrane antigen gene (BrEMA1) and by sequencing of the polymorphic region (i.e., region with a variable number of hexapeptide repeats). Analysis of PCR products revealed 11 different gene profiles, visualized by gel electrophoresis. Twelve distinct BrEMA1 genotypes were identified by sequencing, but the numbers of hexapeptide repeats varied from 6 to 31 (classified as genotype6 to genotype31). The genotypes were retrospectively compared to the clinical case data. The most frequently encountered B. rossi parasites were those attributed to genotype19 (36.2%), genotype28 and genotype29 (20.6% each), and genotype11 (12.7%). These genotypes were also the ones associated with the poorest prognosis. This preliminary finding suggests clinically important differences between the various B. rossi genotypes identified.

Detection of a Theileria species in dogs in South Africa.

A Theileria species was detected by PCR in blood samples collected from dogs in the Pietermaritzburg area and was also found in dogs presented at the Outpatients Clinic of the Onderstepoort Veterinary Academic Hospital (OVAH), in the Pretoria area, South Africa. In the Pietermaritzburg area, 79 of the 192 samples were positive, while 3 out of 1137 of the Onderstepoort samples were positive. Three positive samples from Pietermaritzburg were co-infected with Ehrlichia canis. PCR positive samples were further analysed by the Reverse Line Blot (RLB) and sequence analysis. Phylogenetic analysis of the 18S rRNA full-length gene sequences of one sample (VT12) from Pietermaritzburg and two samples from OVAH (BC281 and BC295) revealed a close relationship with sequences of Theileria species (sable). Clinical signs of the dogs that were examined at Pietermaritzburg and OVAH included an immune-mediated condition with severe thrombocytopenia. These findings identify a Theileria sp. in dogs for the first time in South Africa and add yet another microorganism to the growing list of haemoprotozoan parasites infecting dogs worldwide. The clinical significance of this infection in dogs is poorly resolved.

A field trial evaluation of the prophylactic efficacy of amitraz-impregnated collars against canine babesiosis (Babesia canis rossi) in South Africa.

South African canine babesiosis caused by Babesia canis rossi is a common clinical disease in dogs in South Africa and remains a significant cause of domestic dog mortality. To determine whether tick-repellent, 9% amitraz-impregnated tick collars (Preventic-Virbac) could prevent tick-borne exposure to B. canis rossi, 50 dogs were assigned to two groups. Group 1 (20 dogs), polymerase chain reaction (PCR)--and reverse line blot (RLB)-negative for B. canis rossi, were fitted with amitraz collars and blood samples collected monthly, over a 6-month period, and analysed for B. canis rossi. Group 2 (30 dogs) included 5 dogs selected on a month-by-month basis from a population of dogs from the same geographical area as the group 1 dogs, but with no history of previous tick control, which were blood-sampled together with the treatment group and analysed for B. canis rossi by PCR and RLB, to serve as the control group. Eight of the 30 control dogs (26.6%) were PCR/RLB positive for B. canis rossi, indicating high pathogen exposure during the trial period. All twenty of the treatment group dogs remained negative for B. canis rossi throughout the 6 months of the trial. These results suggest that the use of amitraz-impregnated collars had a significant effect on reducing infection with B. canis rossi.

Molecular characterisation of Babesia gibsoni infection from a pit-bull terrier pup recently imported into South Africa.

Canine babesiosis caused by Babesia gibsoni was diagnosed in a 3-month-old Pit-bull pup during a routine clinical examination. Diagnosis was confirmed by way of smear examination, PCR, Reverse Line Blot (RLB) and sequence analysis which showed 100% homology with B. gibsoni (Japan AB118032) and Babesia sp. (Oklahoma) (AF205636). Haematology showed moderate anaemia and severe thrombocytopenia. Treatment was initiated with diminazene aceturate (Berenil RTU) followed by 2 doses of imidocarb diproprionate (Forray-65) 3 days and 14 days later, respectively. Babesia gibsoni DNA was still detectable 2 weeks post-treatment on the PCR/RLB test. A 10-day course of combination drug therapy using atovaquone and azithromycin was initiated. Blood samples taken on Day 1 and Day 40 after completion of treatment were negative for B. gibsoni DNA on PCR/RLB test. The implications of a possible introduction of B. gibsoni into South Africa are discussed.

Confirmation of occurrence of Babesia canis vogeli in domestic dogs in South Africa.

The prevalence of Babesia infections in domestic dogs in South Africa was studied using reverse line blot hybridization and 18S sequence analysis. Babesia canis vogeli was confirmed for the first time in domestic dogs in South Africa. Out of a total of 297 blood samples collected from domestic dogs in Bloemfontein, East London, Johannesburg, Durban and from the Onderstepoort Veterinary Academic Hospital, 31 were positive for Babesia canis rossi, whereas B. c. vogeli was detected in 13 dogs. None of the dogs carried both parasites. The detection of B. c. vogeli has implications with regard to prevalence and varied clinical manifestation of canine babesiosis in South Africa.

Occurrence and diversity of bovine coccidia at three localities in South Africa.

Information on the occurrence and diversity of coccidia infecting cattle in different farming systems in South Africa is lacking. The objective of the study was to determine the levels of infection of coccidia in cattle at three localities and to determine the Eimeria species occurring at each locality. Faecal samples (n=1936) were collected over a 13-month period from a dairy farm (Mallesons), a Bonsmara breeding farm (Kaalplaas) and a Nguni stud farm (Pienaars River). Samples were collected from adults >24 months old and calves <12 months old. The modified McMaster technique was used to determine OPG (oocysts per gram) of faeces. Samples with OPGs > or =2000 were allowed to sporulate in potassium dichromate solution at room temperature. Species identification was done by measurement and morphological characteristic of each species. At Mallesons, 29% of samples were positive, at Kaalplaas 50% and at Pienaars River 52%. The most prevalent species were Eimeria zuernii and E. ellipsoidalis at Mallesons, E. zuernii and E. bovis at Kaalplaas and E. bovis and E. zuernii at Pienaars River, respectively. Adults had very low OPGs. There was a pattern at the three localities in that high OPGs were only recorded in calves, which were also shedding the most pathogenic species.