East Coast fever (ECF) is a tick-borne disease of cattle that hinders the development of the livestock industry in eastern, central and southern Africa. The ‘Muguga cocktail’ live vaccine, delivered by an infection and treatment method (ITM), remains the only immunisation strategy of controlling ECF. However, there are challenges of the live vaccine inducing ECF carrier status in immunised animals and the possibility of lack of protection from parasite strains that are antigenically different from the vaccine strains. In Uganda, there are insufficient data regarding the ECF carrier status and T. parva genetic diversity in vaccinated and associated non-vaccinated cattle to assess the effectiveness of ITM vaccination. Blood was collected from recently ECF vaccinated (98) and non-vaccinated (73) cattle from Iganga district in Eastern Uganda at 120 days post-vaccination. The p104 gene nested PCR was used to screen for T. parva DNA, 11 minisatellite and 3 microsatellite markers (SSR) were used for genotyping. Two minisatellite markers (MS7 and MS19) were used to determine whether ECF carrier status was due to the T. parva vaccine or local strains. The prevalence of T. parva based on p104 nPCR was 61.2% (60/98) (RR 2.234, 95% CI 1.49–3.35, p-value < 0.001) among recently vaccinated cattle and 27.4% (20/73) (RR 1.00) among associated non-vaccinated cattle. The Muguga cocktail vaccine strains were responsible for carrier status in 10 (58.8%) by MS7 and 11 (64.7%) by MS19 in vaccinated cattle. Genotypes of T. parva with different-sized alleles to the vaccine strains that could be potential ‘breakthroughs’ were detected in 2 (11.8%)) and 4 (23.5%) isolates from vaccinated cattle based on MS7 and MS19 minisatellite markers, respectively. Using 14 SSR markers, T. parva diversity was higher in vaccinated (Na = 2.214, Ne = 1.978, He = 0.465) than associated non-vaccinated (Na = 1.071, Ne = 1.048, He = 0.259) cattle. The principal component analysis (PCA) showed isolates from vaccinated cattle were closely related to those from non-vaccinated cattle. The analysis of molecular variance (AMOVA) revealed high genetic variation (96%) within T. parva isolates from vaccinated and non-vaccinated cattle but low variation (4%) between vaccinated and non-vaccinated cattle. This study reveals the role of ITM in inducing the carrier status and higher T. parva genetic diversity in vaccinated cattle. The low genetic variation between T. parva isolates in both vaccinated and non-vaccinated cattle may be suggestive of the protective role of vaccine strains against genetically related local strains in the study area.

In this study, the recombinant gut protein rRa92A produced in Pichia pastoris yeast cells was used to immunize cattle in two experiments, one in Brazil and the other in Uganda. In both experiments, the animals were intramuscularly (IM) injected with 200 µg of recombinant protein in Brazil on days 0, 30 and 51 and in Uganda on days 0, 30. Blood samples for sera separation were collected from different days in both experiments. These samples were analyzed by ELISAs. In Brazil, ticks collected from the animals during the experimental period were analyzed for biological parameters. At Uganda, blood was collected to assess blood parameters, clinical signs were recorded and adult tick (Rhipicephalus appendiculatus) counts were performed. All animals of the vaccinated groups were shown to produce antibodies, and it was not possible to detect an effect on Rhipicephalus microplus. All the clinical parameters were considered within the normal ranges for both the experimental and control groups in Uganda. Antibody absorbance was elevated after each immunization and remained high until the end of the experiments, remaining low in the control animals. The results of stall test carried out in Brazil using R. microplus tick showed efficacy of 21.95%. The rRa92A immunization trial experiments in Uganda showing a decrease of 55.2% in the number of engorged adult ticks, which was statistically significant (p<0.05). Assessment of the immunogenicity of Ra92A produced in the P. pastoris expression system in bovines is reported for the first time, and the protein acted as a concealed antigen.

The integrated control of East Coast fever (ECF) by early diagnosis and treatment involving acquired immunity induced by natural infection in Ankole cattle was assessed. A longitudinal study was carried out in Kiruhura district, southwestern Uganda for six months on 244 Ankole breed of cattle from 18 herds under natural tick challenge and relaxed tick control measures. Calves aged three to six months old were recruited and monitored daily by farmers for detection of ECF clinical symptoms. The reported sick animals were treated using Buparvaquone and treatment outcome determined. Monthly follow-ups and blood collections were done to monitor ECF status. Blood was analyzed for Theileria parva parasites by microscopy, DNA by polymerase chain reaction (PCR) and antibodies by enzyme-linked immunosorbent assay (ELISA). The overall prevalence of ECF clinical disease within six months period was 30.3% (74). The major symptoms of early clinical ECF disease were fever and enlarged parotid or prescapular lymph nodes. Clinical cases were categorized as mild, 24% (18) or moderate, 76% (56). There was an overall recovery rate of 100% (74) of the ECF cases whereby 94.6% (70) recovered promptly and 5.4% (4) recovered slowly. Based on blood analysis, prevalence of ECF at baseline was 3.7% (9) by microscopy, 31.1% (76) by PCR and 38.1% (93) by ELISA. A significant increase (p < 0.05) was shown by the increased number of calves with T. parva specific antibodies in the sera from 38.1% at baseline to 68.8% after six months. High antibody levels (positive percentage ≥ 50%) were detected in all ECF-treated and recovered calves at the end of six months. The acquired immunity to ECF was high in treated and recovered cattle, indicating that natural exposure to infection, accurate early diagnosis and effective treatment enhance development of immune-protection in indigenous cattle in an endemic area. The prominent early clinical symptoms for ECF could be exploited in the development of decision support tools for chemotherapy and other integrated control measures.

Background: The variant surface glycoprotein (VSG) of the African trypanosomes is the major membrane protein of the plasma membrane of the bloodstream stage of the parasite. African trypanosomiasis (sleeping sickness in humans and nagana in animals) is caused by the systemic infection of the host by several sub-species of the extracellular haemoflagellate protozoa under genus Trypanosoma. As a defense barrier against the host immune response, the entire surface of the bloodstream form of trypanosome is covered with densely packed molecules of VSG that determines the antigenic phenotype of the parasite. Variant surface glycoprotein has a C-terminal domain that is highly conserved in various species of trypanosomes. Methods: The membrane bound VSG (VSGm) protein was prepared without denaturing the homologous region and by including numerous variable antigen types from Trypanosoma brucei brucei parasites. The purified VSGm native trypanosome protein was used to produce anti-VSGm immune sera in rabbits. The indirect immunofluorescence assay (IFA) was used to detect trypanosomes from mice blood, artificial culture media and cattle histological sections. Results: The resultant immune sera were able to detect different strains and species of African trypanosomes from in vivo and in situ sources after immunostaining. Anti-VSGm antibodies also demonstrated a unique property to locate trypanosomes within the histological tissues even after the trypanosome’s morphology had been distorted. Conclusion: The produced immune sera can be utilized for immunohistochemistry to detect Trypanosoma species in various fluids and tissues. Author summary:

Uganda introduced the PCV10 vaccine into routine infant immunization in 2014, but molecular data on residual vaccine-type carriage after introduction are l