Corridor Disease Research Articles

P67 gene alleles sequence analysis reveals Theileria parva parasites associated with East Coast fever and Corridor disease in buffalo from Zambia

Theileriosis caused by Theileria parva infections is responsible for high cattle mortalities in Zambia. Although infected buffalo are a risk to cattle, the characterization of T. parva parasites occurring in this host in Zambia has not been reported. Furthermore, considering the advances in the development of a p67 subunit vaccine, the knowledge of p67 genetic and antigenic diversity in both cattle and buffalo associated T. parva is crucial. Therefore, blood samples from buffalo (n=43) from Central, Eastern and Southern provinces, and cattle (n=834) from Central, Copperbelt, Eastern, Lusaka, and Southern provinces, were tested for T. parva infection and the parasites characterized by sequencing the gene encoding the p67 antigen. About 76.7 % of buffalo and 19.3 % of cattle samples were PCR positive for T. parva. Three of the four known p67 allele types (1, 2 and 3) were identified in parasites from buffalo, of which two (allele types 2 and 3) are associated with T. parva parasites responsible for Corridor disease. Only allele type 1, associated with East Coast fever, was identified from cattle samples, consistent with previous reports from Zambia. Phylogenetic analysis revealed segregation between allele type 1 sequences from cattle and buffalo samples as they grouped separately within the same sub-clade. The high occurrence of T. parva infection in buffalo samples investigated demonstrates the risk of Corridor disease infection, or even outbreaks, should naïve cattle co-graze with infected buffalo in the presence of the tick vector. In view of a subunit vaccine, the antigenic diversity in buffalo associated T. parva should be considered to ensure broad protection. The current disease control measures in Zambia may require re-evaluation to ensure that cattle are protected against buffalo-derived T. parva infections. Parasite stocks used in ‘infection and treatment’ immunization in Zambia, have not been evaluated for protection against buffalo-derived T. parva parasites currently circulating in the buffalo population.

Molecular and serological prevalence of corridor disease (buffalo associated Theileria parva infection) in cattle populations at the livestock/game interface of KwaZulu-Natal province, South Africa

Theileria parva are intracellular protozoal parasites responsible for three disease syndromes in cattle, namely East Coast fever (ECF), Corridor disease (CD) and Zimbabwean theileriosis. The increase in reports of CD outbreaks in recent years has raised questions about the probability of adaptation of buffalo-derived T. parva strains in cattle herds adjacent to game reserves. A cross-sectional study was conducted from March 2016 to December 2018 to investigate the extent of occurrence of T. parva infections in cattle in the CD-controlled area of KwaZulu-Natal Province. Blood samples were collected from 1137 cattle from 14 herds and analysed by quantitative real-time PCR (qPCR) and indirect fluorescent antibody test (IFAT) to determine the prevalence of T. parva. A total of 484 samples from 4 of the 14 herds were further tested on qPCR for the presence of T. taurotragi infections. The data were analysed using descriptive statistics and a chi-square test was used to assess association between variables. The overall prevalence of T. parva was 1.3% (95%CI:1–2%) and 19.9% (95%CI:17–22%) on qPCR and IFAT, respectively. The qPCR positive samples were detected in March and May while IFAT positive samples were detected in all seasons sampled, with higher numbers during summer months. The Pearson Chi-squared test showed that T. parva prevalence rates based on both qPCR and IFAT were positively associated with herds with previous history of CD outbreaks (χ2 = 8.594, p = 0.003; χ2 = 69.513, p < 0.001, respectively). The overall prevalence of T. taurotragi was 39.4% (95% CI: 35–44%) with the herd-level prevalence ranging between 35.0% and 43.4%. Possible cross-reaction of T. parva IFAT to T. taurotragi was detected on few samples, however, there was no significant association between T. taurotragi infections and IFAT positivity (χ2 = 0.829, p = 0.363). Results from this study demonstrated the extent of occurrence of subclinical carriers and the level of exposure to T. parva infections in cattle populations at a livestock/game interface area of KwaZulu-Natal Province. The molecular and seroprevalence rates were low when compared with other areas where cattle-adapted T. parva infections are endemic. The adaptation of buffalo-derived T. parva in cattle population resulting in cattle-cattle transmissions seem to be unlikely under the current epidemiological state.

Inherited Tolerance in Cattle to the Apicomplexan Protozoan Theileria parva is Associated with Decreased Proliferation of Parasite-Infected Lymphocytes.

Theileria parva is the causative agent of East Coast fever and Corridor disease, which are fatal, economically important diseases of cattle in eastern, central and southern Africa. Improved methods of control of the diseases are urgently required. The parasite transforms host lymphocytes, resulting in a rapid, clonal expansion of infected cells. Resistance to the disease has long been reported in cattle from T. parva-endemic areas. We reveal here that first- and second-generation descendants of a single Bos indicus bull survived severe challenge with T. parva, (overall survival rate 57.3% compared to 8.7% for unrelated animals) in a series of five field studies. Tolerant cattle displayed a delayed and less severe parasitosis and febrile response than unrelated animals. The in vitro proliferation of cells from surviving cattle was much reduced compared to those from animals that succumbed to infection. Additionally, some pro-inflammatory cytokines such as IL1β, IL6, TNFα or TGFβ which are usually strongly expressed in susceptible animals and are known to regulate cell growth or motility, remain low in tolerant animals. This correlates with the reduced proliferation and less severe clinical reactions observed in tolerant cattle. The results show for the first time that the inherited tolerance to T. parva is associated with decreased proliferation of infected lymphocytes. The results are discussed in terms of whether the reduced proliferation is the result of a perturbation of the transformation mechanism induced in infected cells or is due to an innate immune response present in the tolerant cattle.

Clinical Evaluation of Corridor Disease in Bos indicus (Boran) Cattle Naturally Infected With Buffalo-Derived Theileria parva.

Corridor disease (CD) is a fatal condition of cattle caused by buffalo-derived Theileria parva. Unlike the related condition, East Coast fever, which results from infection with cattle-derived T. parva, CD has not been extensively studied. We describe in detail the clinical and laboratory findings in cattle naturally infected with buffalo-derived T. parva. Forty-six cattle were exposed to buffalo-derived T. parva under field conditions at the Ol Pejeta Conservancy, Kenya, between 2013 and 2018. The first signs of disease observed in all animals were nasal discharge (mean day of onset was 9 days post-exposure), enlarged lymph nodes (10 days post-exposure), and pyrexia (13.7 days post-exposure). Coughing and labored breathing were observed in more than 50% of animals (14 days post-exposure). Less commonly observed signs, corneal edema (22%) and diarrhea (11%), were observed later in the disease progression (19 days post-exposure). All infections were considered clinically severe, and 42 animals succumbed to infection. The mean time to death across all studies was 18.4 days. The mean time from onset of clinical signs to death was 9 days and from pyrexia to death was 4.8 days, indicating a relatively short duration of clinical illness. There were significant relationships between days to death and the days to first temperature (chi2 = 4.00, p = 0.046), and days to peak temperature (chi2 = 25.81, p = 0.001), animals with earlier onset pyrexia died sooner. These clinical indicators may be useful for assessing the severity of disease in the future. All infections were confirmed by the presence of macroschizonts in lymph node biopsies (mean time to parasitosis was 11 days). Piroplasms were detected in the blood of two animals (4%) and 20 (43%) animals seroconverted. In this study, we demonstrate the successful approach to an experimental field study for CD in cattle. We also describe the clinical progression of CD in naturally infected cattle, including the onset and severity of clinical signs and pathology. Laboratory diagnoses based on examination of blood samples are unreliable, and alternatives may not be available to cattle keepers. The rapid development of CD requires recognition of the clinical signs, which may be useful for early diagnosis of the disease and effective intervention for affected animals.

South African Buffalo-Derived Theileria parva Is Distinct From Other Buffalo and Cattle-Derived T. parva.

Theileria parva is a protozoan parasite transmitted by the brown-eared ticks, Rhipicephalus appendiculatus and Rhipicephalus zambeziensis. Buffaloes are the parasite’s ancestral host, with cattle being the most recent host. The parasite has two transmission modes namely, cattle–cattle and buffalo–cattle transmission. Cattle–cattle T. parva transmission causes East Coast fever (ECF) and January disease syndromes. Buffalo to cattle transmission causes Corridor disease. Knowledge on the genetic diversity of South African T. parva populations will assist in determining its origin, evolution and identify any cattle–cattle transmitted strains. To achieve this, genomic DNA of blood and in vitro culture material infected with South African isolates (8160, 8301, 8200, 9620, 9656, 9679, Johnston, KNP2, HL3, KNP102, 9574, and 9581) were extracted and paired-end whole genome sequencing using Illumina HiSeq 2500 was performed. East and southern African sample data (Chitongo Z2, Katete B2, Kiambu Z464/C12, Mandali Z22H10, Entebbe, Nyakizu, Katumba, Buffalo LAWR, and Buffalo Z5E5) was also added for comparative purposes. Data was analyzed using BWA and SAMtools variant calling with the T. parva Muguga genome sequence used as a reference. Buffalo-derived strains had higher genetic diversity, with twice the number of variants compared to cattle-derived strains, confirming that buffaloes are ancestral reservoir hosts of T. parva. Host specific SNPs, however, could not be identified among the selected 74 gene sequences. Phylogenetically, strains tended to cluster by host with South African buffalo-derived strains clustering with buffalo-derived strains. Among the buffalo-derived strains, South African strains were genetically divergent from other buffalo-derived strains indicating possible geographic sub-structuring. Geographic sub- structuring was also observed within South Africa strains. The knowledge generated from this study indicates that to date, ECF is not circulating in buffalo from South Africa. It also shows that T. parva has historically been present in buffalo from South Africa before the introduction of ECF and was not introduced into buffalo during the ECF epidemic.

Limited diversity in the CD8+ antigen-coding loci in Theileria parva parasites from cattle from southern and eastern Africa

Theileria parva infections in cattle causes huge economic losses in the affected African countries, directly impacting the livelihood of the poor small-holder farmers. The current immunization protocol using live sporozoites in eastern Africa, is among the control measures designed to limit T. parva infections in cattle. However, the ability of the immune protection induced by this immunization to protect against field parasites has been compromised by the diversity of the parasite involving the schizont antigen genes. Previous studies have reported on the antigenic diversity of T. parva parasites from southern and eastern Africa, however, similar reports on T. parva parasites particularly from cattle from southern Africa remains scanty, due to the self-limiting nature of Corridor disease. Thus, we evaluated the diversity of CD8+ T-cell regions of ten schizont antigen genes in T. parva parasites associated with Corridor disease and East Coast fever (ECF) from southern and eastern Africa respectively.Regions of schizont antigen (TpAg) genes containing the CD8+ T-cell epitopes (CTL determinants) were amplified from genomic DNA extracted from blood of T. parva positive samples, cloned and sequenced. The results revealed limited diversity between the two parasite groups from cattle from southern and eastern Africa, defying the widely accepted notion that antigen-encoding loci in cattle-derived parasites are conserved, while in buffalo-derived parasites, they are extensively variable. This suggests that only a sub-population of parasites is successfully transmitted from buffalo to cattle, resulting in the limited antigenic diversity in Corridor disease parasites. Tp4, Tp5, Tp7 and Tp8 showed limited to absence of diversity in both parasite groups, suggesting the need to further investigate their immunogenic properties for consideration as candidates for a subunit vaccine. Distinct and common variants of Tp2 were detected among the ECF parasites from eastern Africa indicating evidence of parasite mixing following immunization. This study provides additional information on the comparative diversity of TpAg genes in buffalo- and cattle-derived T. parva parasites from cattle from southern and eastern Africa.

Unique Mitochondrial Single Nucleotide Polymorphisms Demonstrate Resolution Potential to Discriminate Theileria parva Vaccine and Buffalo-Derived Strains.

Distinct pathogenic and epidemiological features underlie different Theileria parva strains resulting in different clinical manifestations of East Coast Fever and Corridor Disease in susceptible cattle. Unclear delineation of these strains limits the control of these diseases in endemic areas. Hence, an accurate characterization of strains can improve the treatment and prevention approaches as well as investigate their origin. Here, we describe a set of single nucleotide polymorphisms (SNPs) based on 13 near-complete mitogenomes of T. parva strains originating from East and Southern Africa, including the live vaccine stock strains. We identified 11 SNPs that are non-preferentially distributed within the coding and non-coding regions, all of which are synonymous except for two within the cytochrome b gene of buffalo-derived strains. Our analysis ascertains haplotype-specific mutations that segregate the different vaccine and the buffalo-derived strains except T. parva-Muguga and Serengeti-transformed strains suggesting a shared lineage between the latter two vaccine strains. Phylogenetic analyses including the mitogenomes of other Theileria species: T. annulata, T. taurotragi, and T. lestoquardi, with the latter two sequenced in this study for the first time, were congruent with nuclear-encoded genes. Importantly, we describe seven T. parva haplotypes characterized by synonymous SNPs and parsimony-informative characters with the other three transforming species mitogenomes. We anticipate that tracking T. parva mitochondrial haplotypes from this study will provide insight into the parasite’s epidemiological dynamics and underpin current control efforts.

Microsatellite and minisatellite genotyping of Theileria parva population from southern Africa reveals possible discriminatory allele profiles with parasites from eastern Africa

The control of Theileria parva, a protozoan parasite that threatens almost 50% of the cattle population in Africa, is still a challenge in many affected countries. Theileria parva field parasites from eastern Africa, and parasites comprising the current live T. parva vaccine widely deployed in the same region have been reported to be genotypically diverse. However, similar reports on T. parva parasites from southern Africa are limited, especially in Corridor disease designated areas. Establishing the extent of genetic exchange in T. parva populations is necessary for effective control of the parasite infection.Twelve polymorphic microsatellite and minisatellite loci were targeted for genotypic and population genetics analysis of T. parva parasites from South Africa, Mozambique, Kenya and Uganda using genomic DNA prepared from cattle and buffalo blood samples. The results revealed genotypic similarities among parasites from the two regions of Africa, with possible distinguishing allelic profiles on three loci (MS8, MS19 and MS33) for parasites associated with Corridor disease in South Africa, and East Coast fever in eastern Africa. Individual populations were in linkage equilibrium (VD<L), but when considered as one combined population, linkage disequilibrium (VD>L) was observed. Genetic divergence was observed to be more within (AMOVA = 74%) than between (AMOVA = 26%) populations. Principal coordinate analysis showed clustering that separated buffalo-derived from cattle-derived T. parva parasites, although parasites from cattle showed a close genetic relationship. The results also demonstrated geographic sub-structuring of T. parva parasites based on the disease syndromes caused in cattle in the two regions of Africa.These findings provide additional information on the genotypic diversity of T. parva parasites from South Africa, and reveal possible differences based on three loci (MS8, MS19 and MS33) and similarities between buffalo-derived T. parva parasites from southern and eastern Africa.

Analysis of p67 allelic sequences reveals a subtype of allele type 1 unique to buffalo-derived Theileria parva parasites from southern Africa.

East Coast fever (ECF) and Corridor disease (CD) caused by cattle- and buffalo-derived T. parva respectively are the most economically important tick-borne diseases of cattle in the affected African countries. The p67 gene has been evaluated as a recombinant subunit vaccine against ECF, and for discrimination of T. parva parasites causing ECF and Corridor disease. The p67 allele type 1 was first identified in cattle-derived T. parva parasites from East Africa, where parasites possessing this allele type have been associated with ECF. Subsequent characterization of buffalo-derived T. parva parasites from South Africa where ECF was eradicated, revealed the presence of a similar allele type, raising concerns as to whether or not allele type 1 from parasites from the two regions is identical. A 900 bp central fragment of the gene encoding p67 was PCR amplified from T. parva DNA extracted from blood collected from cattle and buffalo in South Africa, Mozambique, Kenya, Tanzania and Uganda, followed by DNA sequence analysis. Four p67 allele types previously described were identified. A subtype of p67 allele type 1 was identified in parasites from clinical cases of CD and buffalo from southern Africa. Notably, p67 allele type 1 sequences from parasites associated with ECF in East Africa and CD in Kenya were identical. Analysis of two p67 B-cell epitopes (TpM12 and AR22.7) revealed amino acid substitutions in allele type 1 from buffalo-derived T. parva parasites from southern Africa. However, both epitopes were conserved in allele type 1 from cattle- and buffalo-derived T. parva parasites from East Africa. These findings reveal detection of a subtype of p67 allele type 1 associated with T. parva parasites transmissible from buffalo to cattle in southern Africa.

Investigations into the carrier-state of Theileria sp. (buffalo) in cattle.

The Theileria are apicomplexan parasites transmitted by ticks to vertebrate hosts. Most Theileria species exhibit some form of host or vector specificity, since under endemic conditions only a limited number of tick species act as vectors and not all vertebrate hosts are able to maintain a persistent carrier state. Data for Theileria sp. (buffalo) suggest host specificity for African buffalo (Syncerus caffer). However, T. sp. (buffalo) infections in cattle co-grazing with African buffalo have been reported in Kenya and schizonts were cultured from these infected cattle, raising questions regarding host specificity. A Corridor disease outbreak in 2013 on a ranch in South Africa where cattle co-grazed with Theileria parva and T. sp. (buffalo) infected buffalo presented the opportunity to investigate the possible carrier-state of T. sp. (buffalo) in cattle using real-time PCR analysis. Almost all buffalo (n = 19, 95%) were infected with T. sp. (buffalo) and showed CP values (22-20) indicative of high parasitemia similar to that observed for buffalo in endemic areas. Conversely, only ~14-27% cattle (n = 69, 100, 96) were positive with CP values (31-40) suggesting low parasitemia and a carrier state epidemiology different from African buffalo. Long term monitoring of T. sp. (buffalo) positive cattle showed that most cattle lost their parasitemia or presented fluctuating parasitemia around the PCR assay detection limit. A single splenectomized animal showed a persistent carrier state. The general trends and epidemiology observed in cattle infected with T. sp. (buffalo) are similar to that seen for buffalo-adapted T. parva, for which a defined carrier state in cattle has not yet been proven. The study suggests that cattle may be infected by T. sp. (buffalo) but are not definitive hosts that play an important part in the epidemiology of this parasite.

Corridor disease (buffalo-associated Theileria parva) outbreak in cattle introduced onto a game ranch and investigations into their carrier-state.

East Coast fever (Theileria parva infection in cattle) was eradicated from South Africa in the mid-1900. However, another form named Corridor disease (CD), associated with T. parva carrier buffaloes exists and outbreaks have increased in endemic areas. The occurrence of a CD carrier state in cattle under field situations has not been demonstrated but remains a subject of controversy. The current study investigated the T. parva carrier state following a severe outbreak in cattle introduced onto a game ranch. Monitoring of the outbreak included clinical signs, mortality, microscopy, serology, real-time PCR and xenodiagnoses. The herd of cattle received block treatment using oxytetracyclines (OTC) by the farmer during the outbreak. Cattle were sampled early during the outbreak and twice within the following 75 days. All buffaloes were tested for a T. parva carrier state. Two batches of questing adult R. appendiculatus were collected at the time of disease occurrence and a year later. These ticks were fed on susceptible cattle under controlled conditions and monitored for disease transmission. Ticks infected with a buffalo-derived stock of T. parva were fed on one bovine under controlled conditions and simultaneously injected with OTC, simulating the infection and treatment method of vaccination and was used as a positive control. Clean R. appendiculatus nymphs were fed on four recovered PCR positive cattle from the outbreak and on the positive control animal. The adult ticks were tested for infectivity by xenodiagnoses on susceptible bovines. For the initial outbreak the CD prevalence was 62.3% with a mortality rate of 29.5%. However, the outbreak was contained by block OTC treatment of the herd since only 3.4% cattle subsequently died until the end of the investigations. Adult ticks fed on one field bovine and the laboratory established T. parva carrier both transmitted fatal infections to susceptible cattle. Ticks fed on two field cattle transmitted T. taurotragi and one failed to transmit any infection. Questing adult R. appendiculatus collected during the outbreak transmitted fatal CD to two bovines while ticks collected a year later transmitted T. taurotragi. These findings demonstrated the effectiveness of disease control either by cattle treatment using OTC simulating the ITM or by intensive cattle dipping following the outbreak or by both interventions. The potential risk of creating carrier cattle by OTC treatment during CD outbreaks should be considered, supporting the continued control measures of segregation of cattle and buffalo herds.

Ethnoveterinary treatments for common cattle diseases in four districts of the Southern Province, Zambia.

Aim:Ethno veterinary knowledge has rarely been recorded, and no or limited effort has been made to exploit this knowledge despite its widespread use in Zambia. This study documented the types of plants used to treat important animal diseases in rural Zambia as a way of initiating their sustained documentation and scientific validation.Materials and Methods:The study was done in selected districts of the Southern Zambia, Africa. The research was a participatory epidemiological study conducted in two phases. The first phase was a pre-study exploratory rapid rural appraisal conducted to familiarize the researchers with the study areas, and the second phase was a participatory rural appraisal to help gather the data. The frequency index was used to rank the commonly mentioned treatments.Results:A number of diseases and traditional treatments were listed with the help of local veterinarians. Diseases included: Corridor disease (Theileriosis), foot and mouth disease, blackleg, bloody diarrhea, lumpy skin disease, fainting, mange, blindness, coughing, bloat, worms, cobra snakebite, hemorrhagic septicemia, and transmissible venereal tumors. The plant preparations were in most diseases given to the livestock orally (as a drench). Leaves, barks, and roots were generally used depending on the plant type.Conclusion:Ethno veterinary medicine is still widespread among the rural farmers in the province and in Zambia in general. Some medicines are commonly used across diseases probably because they have a wide spectrum of action. These medicines should, therefore, be validated for use in conventional livestock healthcare systems in the country to reduce the cost of treatments.

The complete genome sequence of the African buffalo (Syncerus caffer).

BackgroundThe African buffalo (Syncerus caffer) is an important role player in the savannah ecosystem. It has become a species of relevance because of its role as a wildlife maintenance host for an array of infectious and zoonotic diseases some of which include corridor disease, foot-and-mouth disease and bovine tuberculosis. To date, no complete genome sequence for S. caffer had been available for study and the genomes of other species such as the domestic cow (Bos taurus) had been used as a proxy for any genetics analysis conducted on this species. Here, the high coverage genome sequence of the African buffalo (S. caffer) is presented.ResultsA total of 19,765 genes were predicted and 19,296 genes could be successfully annotated to S. caffer while 469 genes remained unannotated. Moreover, in order to extend a detailed annotation of S. caffer, gene clusters were constructed using twelve additional mammalian genomes. The S. caffer genome contains 10,988 gene clusters, of which 62 are shared exclusively between B. taurus and S. caffer.ConclusionsThis study provides a unique genomic perspective for the S. caffer, allowing for the identification of novel variants that may play a role in the natural history and physiological adaptations.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3364-0) contains supplementary material, which is available to authorized users.

Exposure of vaccinated and naive cattle to natural challenge from buffalo-derived Theileria parva.

Integrative management of wildlife and livestock requires a clear understanding of the diseases transmitted between the two populations. The tick-borne protozoan parasite Theileria parva causes two distinct diseases in cattle, East Coast fever and Corridor disease, following infection with parasites derived from cattle or buffalo, respectively. In this study, cattle were immunized with a live sporozoite vaccine containing three T. parva isolates (the Muguga cocktail), which has been used extensively and successfully in the field to protect against cattle-derived T. parva infection. The cattle were exposed in a natural field challenge site containing buffalo but no other cattle. The vaccine had no effect on the survival outcome in vaccinated animals compared to unvaccinated controls: nine out of the 12 cattle in each group succumbed to T. parva infection. The vaccine also had no effect on the clinical course of the disease. A combination of clinical and post mortem observations and laboratory analyses confirmed that the animals died of Corridor disease. The results clearly indicate that the Muguga cocktail vaccine does not provide protection against buffalo-derived T. parva at this site and highlight the need to evaluate the impact of the composition of challenge T. parva populations on vaccine success in areas where buffalo and cattle are present.

Geographic range of vector-borne infections and their vectors: the role of African wildlife.

The role of African wildlife in the occurrence of vector-borne infections in domestic animals has gained renewed interest as emerging and re-emerging infections occur worldwide at an increasing rate. In Africa, biodiversity conservation and the expansion of livestock production have increased the risk of transmitting vector-borne infections between wildlife and livestock. The indigenous African pathogens with transboundary potential, such as Rift Valley fever virus, African horse sickness virus, bluetongue virus, lumpy skin disease virus, African swine fever virus, and blood-borne parasites have received the most attention. There is no evidence for persistent vector-borne viral infections in African wildlife. For some viral infections, wildlife may act as a reservoir through the inter-epidemic circulation of viruses with mild or subclinical manifestations. Wildlife may also act as introductory or transporting hosts when moved to new regions, e.g. for lumpy skin disease virus, Rift Valley fever virus and West Nile virus. Wildlife may also act as amplifying hosts when exposed to viruses in the early part of the warm season when vectors are active, with spillover to domestic animals later in the season, e.g. with bluetongue and African horse sickness. Some tick species found on domestic animals are more abundant on wildlife hosts; some depend on wildlife hosts to complete their life cycle. Since the endemic stability of a disease depends on a sufficiently large tick population to ensure that domestic animals become infected at an early age, the presence of wildlife hosts that augment tick numbers may be beneficial. Many wild ungulate species are reservoirs of Anaplasma spp., while the role of wildlife in the epidemiology of heartwater (Ehrlichia ruminantium infection) has not been elucidated. Wild ungulates are not usually reservoirs of piroplasms that affect livestock; however, there are two exceptions: zebra, which are reservoirs of Babesia caballi and Theileria equi, and buffalo, which are reservoirs of Theileria parva. The latter causes Corridor disease when transmitted from buffaloto cattle, butthis appearsto be a self-limiting condition, at least in southern Africa. Wild animals are important reservoirs of tsetse-transmitted Trypanosoma spp. infection. The distribution and abundance of some tsetse species, e.g. Glossina morsitans and G. pallidipes, are closely related to the occurrence of their preferred wildlife hosts.

Geographic distribution of Theileria sp. (buffalo) and Theileria sp. (bougasvlei) in Cape buffalo (Syncerus caffer) in southern Africa: implications for speciation

Strict control measures apply to movement of buffalo in South Africa including testing for Theileria parva, the causative agent of Corridor disease in cattle. The official test is a real-time hybridization PCR assay that amplifies the 18S rRNA V4 hyper-variable region of T. parva, T. sp. (buffalo) and T. sp. (bougasvlei). Mixed infections with the latter organisms affect diagnostic sensitivity due to PCR suppression. While the incidence of mixed infections in the Corridor disease endemic region of South Africa is significant, little information is available on the specific distribution and prevalence of T. sp. (buffalo) and T. sp. (bougasvlei). Specific real-time PCR assays were developed and a total of 1211 samples known to harbour these parasites were screened. Both parasites are widely distributed in southern Africa and the incidence of mixed infections with T. parva within the endemic region is similar (∼25-50%). However, a significant discrepancy exists in regard to mixed infections of T. sp. (buffalo) and T. sp. (bougasvlei) (∼10%). Evidence for speciation between T. sp. (buffalo) and T. sp. (bougasvlei) is supported by phylogenetic analysis of the COI gene, and their designation as different species. This suggests mutual exclusion of parasites and the possibility of hybrid sterility in cases of mixed infections.

Whole-Genome Sequencing of Theileria parva Strains Provides Insight into Parasite Migration and Diversification in the African Continent

The disease caused by the apicomplexan protozoan parasite Theileria parva, known as East Coast fever or Corridor disease, is one of the most serious cattle diseases in Eastern, Central, and Southern Africa. We performed whole-genome sequencing of nine T. parva strains, including one of the vaccine strains (Kiambu 5), field isolates from Zambia, Uganda, Tanzania, or Rwanda, and two buffalo-derived strains. Comparison with the reference Muguga genome sequence revealed 34 814–121 545 single nucleotide polymorphisms (SNPs) that were more abundant in buffalo-derived strains. High-resolution phylogenetic trees were constructed with selected informative SNPs that allowed the investigation of possible complex recombination events among ancestors of the extant strains. We further analysed the dN/dS ratio (non-synonymous substitutions per non-synonymous site divided by synonymous substitutions per synonymous site) for 4011 coding genes to estimate potential selective pressure. Genes under possible positive selection were identified that may, in turn, assist in the identification of immunogenic proteins or vaccine candidates. This study elucidated the phylogeny of T. parva strains based on genome-wide SNPs analysis with prediction of possible past recombination events, providing insight into the migration, diversification, and evolution of this parasite species in the African continent.

Field and laboratory studies on Corridor disease (Theileria parva infection) in cattle population at the livestock/game interface of uPhongolo-Mkuze area, South Africa

Corridor disease (Theileria parva infection in cattle associated with carrier buffaloes) had not been reported to cause serious outbreaks in South Africa prior to 1994. In recent years, there has been an increase in the introduction of T. parva-infected buffaloes onto private game parks in Northern KwaZulu-Natal (KZN). The objectives of this study were to investigate the number of T. parva outbreaks in cattle at the livestock/wildlife interface and to establish the possible T. parva carrier status in cattle which were diagnosed to have recovered from clinical disease.The occurrence of outbreaks was closely monitored from 2004 to 2009 covering a total of 15 localities. The observations included the number of cattle involved in the outbreaks, clinical signs, parasitological and post-mortem examinations, as well as serological and molecular tests specific for T. parva. Sentinel cattle were introduced to monitor tick transmission and some of these recovered from clinical T. parva infection in the field and confirmed to be positive by PCR, were challenged using lethal T. parva stabilates to ascertain their immune status.Thirty-one Corridor disease outbreaks were recorded during the study period. Of the 846 cattle tested for Corridor disease during the study period, 140 (16.5%) were found positive by the real time PCR and IFA tests. Eighty-two (9.7%) cattle were found positive by the IFA test only. The prevalence of T. parva infection was 26.2%. Adult R. appendiculatus fed as nymphs on 5 bovines which recovered from clinical T. parva infection in the field transmitted only T. taurotragi to susceptible bovines. However, 8 of the field-recovered cattle resisted lethal challenge using T. parva tick stabilate. Though the study could not demonstrate cattle-to-cattle transmission by ticks using 5 previously infected cattle in the field, it is suggested that Corridor disease should be considered a potential emerging disease, and more stringent control methods should be implemented.

An Overview of Disease-Free Buffalo Breeding Projects with Reference to the Different Systems Used in South Africa

This paper describes the successful national program initiated by the South African government to produce disease-free African buffalo so as to ensure the sustainability of this species due to threats from diseases. Buffalo are known carriers of foot-and-mouth disease, bovine tuberculosis, Corridor disease and brucellosis. A long-term program involving multiphase testing and a breeding scheme for buffalo is described where, after 10 years, a sustainable number of buffalo herds are now available that are free of these four diseases. A large portion of the success was attributable to the use of dairy cows as foster parents with the five-stage quarantine process proving highly effective in maintaining the “disease-free” status of both the calves and the foster cows. The projects proved the successfulness of breeding with African buffalo in a commercial system that was unique to African buffalo and maintained the “wildness” of the animals so that they could effectively be released back into the wild with minimal, if any, behavioral problems.

Novel SNP Discovery in African Buffalo, Syncerus caffer, using high-throughput Sequencing.

The African buffalo, Syncerus caffer, is one of the most abundant and ecologically important species of megafauna in the savannah ecosystem. It is an important prey species, as well as a host for a vast array of nematodes, pathogens and infectious diseases, such as bovine tuberculosis and corridor disease. Large-scale SNP discovery in this species would greatly facilitate further research into the area of host genetics and disease susceptibility, as well as provide a wealth of sequence information for other conservation and genomics studies. We sequenced pools of Cape buffalo DNA from a total of 9 animals, on an ABI SOLiD4 sequencer. The resulting short reads were mapped to the UMD3.1 Bos taurus genome assembly using both BWA and Bowtie software packages. A mean depth of 2.7× coverage over the mapped regions was obtained. Btau4 gene annotation was added to all SNPs identified within gene regions. Bowtie and BWA identified a maximum of 2,222,665 and 276,847 SNPs within the buffalo respectively, depending on analysis method. A panel of 173 SNPs was validated by fluorescent genotyping in 87 individuals. 27 SNPs failed to amplify, and of the remaining 146 SNPs, 43–54% of the Bowtie SNPs and 57–58% of the BWA SNPs were confirmed as polymorphic. dN/dS ratios found no evidence of positive selection, and although there were genes that appeared to be under negative selection, these were more likely to be slowly evolving house-keeping genes.