Absence of 2899CPanthera leo) with Polymyopathy.

Polyphasic skeletal muscle degeneration, necrosis and mineralization of skeletal muscle was diagnosed in eight juvenile free-ranging lions (Panthera leo), from five different litters in the Greater Kruger National Park area that were unable to walk properly. A detailed investigation was not possible in free-ranging lions, so the cause could not be determined. The cases resembled hypokalemic polymyopathy in domestic cats with muscle weakness. A candidate-gene approach previously identified a nonsense mutation in the gene coding for the enzyme lysine-deficient 4 protein kinase (WNK4) associated with the disease in Burmese and Tonkinese cats. In this study, we sequenced all 19 exons of the gene in one case, and two control samples, to identify possible mutations that may be associated with polymyopathy in free-ranging lions. Here, no mutations were detected in any of the exons sequenced. Our findings indicate that the WNK4 gene is not a major contributor to the condition in these lions. Further studies into the pathogenesis of this condition are needed to inform conservation policies for this vulnerable, iconic African species.

Pathological findings in African buffaloes (Syncerus caffer) in South Africa.

The African buffalo (Syncerus caffer) is an iconic species of South African megafauna. As the farmed buffalo population expands, the potential impacts on population health and disease transmission warrant investigation. A retrospective study of skin biopsy and necropsy samples from 429 animals was performed to assess the spectrum of conditions seen in buffaloes in South Africa. Determination of the cause of death (or euthanasia) could not be made in 33.1% (136/411) of the necropsy cases submitted due to autolysis or the absence of significant lesions in the samples submitted. Infectious and parasitic diseases accounted for 53.5% (147/275) of adult fatal cases and non-infectious conditions accounted for 34.9% (96/275). Abortions and neonatal deaths made up 11.6% (32/275) of necropsy cases. Rift Valley fever, bovine viral diarrhoea, malignant catarrhal fever, tuberculosis, bacterial pneumonia, anaesthetic deaths, cachexia and hepatotoxic lesions were the most common causes of death. The range of infectious, parasitic and non-infectious diseases to which African buffaloes were susceptible was largely similar to diseases in domestic cattle which supports concerns regarding disease transmission between the two species. The similarity between diseases experienced in both species will assist wildlife veterinarians in the diagnosis and treatment of diseases in captive African buffaloes. The present study likely does not represent accurate disease prevalence data within the source population of buffaloes, and diseases such as anthrax, brucellosis and foot and mouth disease are under-represented in this study. Hepatic ductal plate abnormalities and haemorrhagic septicaemia have not, to our knowledge, been previously reported in African buffaloes.

A review of pathological findings in impalas (Aepyceros melampus) in South Africa.

Impalas (Aepyceros melampus) are common African antelope. A retrospective study was conducted of 251 impala cases from game farms, national parks and zoos submitted by veterinarians and pathologists in South Africa (2003-2016). Histopathology slides as well as records of macroscopic lesions and additional diagnostic tests performed were examined. Non-infectious conditions, such as acute pulmonary congestion and oedema, cachexia, traumatic injury and anaesthetic-related mortality were the most common causes of morbidity and mortality. Bacterial sepsis was the most common infectious disease, whilst skeletal muscle and myocardial sarcocystosis and verminous cholangitis and pneumonia were the most common parasitic diseases. Although the retrospective nature of this study limits the significance of the relative prevalence of lesions in the three locations, management decisions and diagnostic plans may be informed by the results. Impala from game farms had significantly more cachexia cases than those from other locations. Impala from zoos had significantly more lymphoid depletion than those from other locations. These findings suggest that nutrition and pasture management, enclosure design, management of intra- and interspecies aggression and improved anaesthetic protocols could improve animal welfare and survival of impala on game farms and in zoos. This report presents a detailed survey of diseases and conditions found in impala that provides baseline data for veterinary pathologists.

Zoonotic Alphaviruses in Fatal and Neurologic Infections in Wildlife and Nonequine Domestic Animals, South Africa.

Alphaviruses from Africa, such as Middelburg virus (MIDV), and Sindbis virus (SINV), were detected in horses with neurologic disease in South Africa, but their host ranges remain unknown. We investigated the contribution of alphaviruses to neurologic infections and death in wildlife and domestic animals in this country. During 2010-2018, a total of 608 clinical samples from wildlife and nonequine domestic animals that had febrile, neurologic signs or unexplained deaths were tested for alphaviruses. We identified 32 (5.5%) of 608 alphavirus infections (9 SINV and 23 MIDV), mostly in neurotissue of wildlife, domestic animals, and birds. Phylogenetic analysis of the RNA-dependent RNA polymerase gene confirmed either SINV or MIDV. This study implicates MIDV and SINV as potential causes of neurologic disease in wildlife and nonequine domestic species in Africa and suggests a wide host range and pathogenic potential.

Rabies Vaccination of 6-Week-Old Puppies Born to Immunized Mothers: A Randomized Controlled Trial in a High-Mortality Population of Owned, Free-Roaming Dogs.

To achieve global elimination of human rabies from dogs by 2030, evidence-based strategies for effective dog vaccination are needed. Current guidelines recommend inclusion of dogs younger than 3 months in mass rabies vaccination campaigns, although available vaccines are only recommended for use by manufacturers in older dogs, ostensibly due to concerns over interference of maternally-acquired immunity with immune response to the vaccine. Adverse effects of vaccination in this age group of dogs have also not been adequately assessed under field conditions. In a single-site, owner-blinded, randomized, placebo-controlled trial in puppies born to mothers vaccinated within the previous 18 months in a high-mortality population of owned, free-roaming dogs in South Africa, we assessed immunogenicity and effect on survival to all causes of mortality of a single dose of rabies vaccine administered at 6 weeks of age. We found that puppies did not have appreciable levels of maternally-derived antibodies at 6 weeks of age (geometric mean titer 0.065 IU/mL, 95% CI 0.061-0.069; n = 346), and that 88% (95% CI 80.7-93.3) of puppies vaccinated at 6 weeks had titers ≥0.5 IU/mL 21 days later (n = 117). Although the average effect of vaccination on survival was not statistically significant (hazard ratio [HR] 1.35, 95% CI 0.83-2.18), this effect was modified by sex (p = 0.02), with the HR in females 3.09 (95% CI 1.24-7.69) and the HR in males 0.79 (95% CI 0.41-1.53). We speculate that this effect is related to the observed survival advantage that females had over males in the unvaccinated group (HR 0.27; 95% CI 0.11-0.70), with vaccination eroding this advantage through as-yet-unknown mechanisms.

Characterization of SAT2 foot-and-mouth disease 2013/2014 outbreak viruses at the wildlife-livestock interface in South Africa.

The Southern African Territories (SAT)-type foot-and-mouth disease viruses (FMDV) are endemic to the greater Kruger National Park (KNP) area in South Africa, where they are maintained through persistent infections in African buffalo. The occurrence of FMDV within the Greater KNP area constitutes a continual threat to the livestock industry. To expand on knowledge of FMDV diversity, the genetic and antigenic relatedness of SAT2-type viruses isolated from cattle during a FMD outbreak in Mpumalanga Province in 2013 and 2014 were investigated. Cattle from twelve diptanks tested positive on polymerase chain reaction (PCR), and molecular epidemiological relationships of the viruses were determined by VP1 sequencing. Phylogenetic analysis of the SAT2 viruses from the FMD outbreak in Mpumalanga in 2013/2014 revealed their genetic relatedness to other SAT2 isolates from topotype I (South Africa, Zimbabwe and Mozambique), albeit genetically distinct from previous South African outbreak viruses (2011 and 2012) from the same topotype. The fifteen SAT2 field isolates clustered into a novel genotype with ≥98.7% nucleotide identity. High neutralization antibody titres were observed for four 2013/2014 outbreak viruses tested against the SAT2 reference antisera representative of viruses isolated from cattle and buffalo from South Africa (topotype I) and Zimbabwe (topotype II). Comparison of the antigenic relationship (r1 values) of the outbreak viruses with reference antisera indicated a good vaccine match with 90% of r1 values > 0.3. The r1 values for the 2013/2014 outbreak viruses were 0.4 and above for the three South African vaccine/reference strains. These results confirm the presence of genetic and antigenic variability in SAT2 viruses and suggest the emergence of new variants at the wildlife-livestock interface in South Africa. Continuous characterization of field viruses should be performed to identify new virus strains as epidemiological surveillance to improve vaccination efforts.

First detection of Mycobacterium bovis infection in Giraffe (Giraffa camelopardalis) in the Greater Kruger National Park Complex: Role and implications.

Bovine tuberculosis (bovine TB) caused by Mycobacterium bovis has become endemic in some wildlife populations in South Africa. The disease has been reported in 21 wildlife species in the country. In this study, we report M. bovis infection in two female giraffes (Giraffa camelopardalis) from two different nature reserves within the Greater Kruger National Park Complex (GKNPC). Mycobacterium bovis was isolated from tissue lesions consistent with macroscopic appearance of tuberculosis (TB) and confirmed by polymerase chain reactions (PCRs), targeting the RD4 region of difference on the genome of the isolates. Spoligotyping and variable number of tandem repeat (VNTR) typing revealed infection of one giraffe with a strain (SB0294) previously not detected in South Africa, while a resident M. bovis strain (SB0121) was detected from the other giraffe. Our work is first to report M. bovis infection in free-ranging giraffes in South Africa. We have further demonstrated the existence of at least three genetically unrelated strains currently infecting wildlife species within the GKNPC. This finding suggests that the epidemiological situation of M. bovis within the GKNPC is not only driven by internal sources from its established endemic presence, but can be additionally fuelled by strains introduced from external sources. It further emphasizes that regular wildlife disease surveillance is an essential prerequisite for the timely identification of new pathogens or strains in ecospheres of high conservation value.

Spatiotemporal epidemiology of rabies at an interface between domestic dogs and wildlife in South Africa.

We characterized the spatiotemporal epidemiology of rabies from January 2009 through March 2014 across the interface between a wildlife reserve and communal livestock farming area in South Africa. Brain tissue from 344 animals of 28 different species were tested for lyssavirus antigen. Of these, 146 (42.4%) samples tested positive, of which 141 (96.6%) came from dogs. Brain samples of dogs were more likely to test positive for lyssavirus antigen if they were found and destroyed in the reserve, compared to samples originating from dogs outside the reserve (65.3% vs. 45.5%; odds ratio (OR) = 2.26, 95% confidence interval (CI) = 1.27-4.03), despite rabies surveillance outside the reserve being targeted to dogs that have a higher index of suspicion due to clinical or epidemiological evidence of infection. In the reserve, dogs were more likely to test positive for rabies if they were shot further from villages (OR = 1.42, 95% CI 1.18-1.71) and closer to water points (OR = 0.41, 95% CI 0.21-0.81). Our results provide a basis for refinement of existing surveillance and control programs to mitigate the threat of spillover of rabies to wildlife populations.

Census and vaccination coverage of owned dog populations in four resource-limited rural communities, Mpumalanga province, South Africa.

Dogs (Canis familiaris) are often free-roaming in sub-Saharan African countries. Rabies virus circulates in many of these populations and presents a public health issue. Mass vaccination of dog populations is the recommended method to decrease the number of dog and human rabies cases. We describe and compare four populations of dogs and their vaccination coverage in four different villages (Hluvukani, Athol, Utah and Dixie) in Bushbuckridge Municipality, Mpumalanga province, South Africa. Cross-sectional surveys were conducted in the villages of Athol, Utah and Dixie, while data from a Health and Demographic Surveillance System were used to describe the dog population in Hluvukani village. All households of the villages were visited to obtain information on the number, sex, age and rabies vaccination status of dogs. From May to October 2013, 2969 households were visited in the four villages and 942 owned dogs were reported. The populations were all young and skewed towards males. No differences were observed in the sex and age distributions (puppies 0-3 months excluded) among the villages. Athol had a higher proportion of dog-owning households than Hluvukani and Utah. Vaccination coverages were all above the 20% - 40% threshold required for herd immunity to rabies (38% in Hluvukani, 51% in Athol, 65% in Dixie and 74% in Utah). For the preparation of vaccination campaigns, we recommend the use of the relatively stable dog:human ratio (between 1:12 and 1:16) to estimate the number of dogs per village in Bushbuckridge Municipality.

Rabies vaccine is associated with decreased all-cause mortality in dogs.

Evidence suggests that rabies vaccine may have non-specific protective effects in animals and children. We analyzed four years of data (2012-2015) from an observational study of the health and demographics of a population of owned, free-roaming dogs in a low-income community in South Africa. The objective of this analysis was to assess the association between rabies vaccine and all-cause mortality in dogs, stratified by age group (0-3months, 4-11months, and 12months and older), and controlling for the effects of sex and number of dogs in the residence. Rabies vaccination reduced the risk of death from any cause by 56% (95% CI=16-77%) in dogs aged 0-3months, by 44% (95% CI=21-60%) in dogs aged 4-11months and by 16% (95% CI=0-29%) in dogs aged 12months and older. We hypothesize that the protective association between rabies vaccination status and all-cause mortality is due to a protective effect of rabies vaccine against diseases other than rabies. Existence of a strong non-specific protective effect of rabies vaccine on mortality in dogs would have implications for the design of dog rabies control programs that aim to eliminate dog-mediated human rabies cases. Further, we propose that owned domestic dogs in high mortality settings provide a useful animal model to better understand any non-specific protective effect of rabies vaccine in children, due to dogs' high numbers, high morbidity and mortality rates, relatively short lifespan, exposure to a variety of infectious and parasitic diseases, and shared environment with people.

Population Dynamics of Owned, Free-Roaming Dogs: Implications for Rabies Control.

BACKGROUND: Rabies is a serious yet neglected public health threat in resource-limited communities in Africa, where the virus is maintained in populations of owned, free-roaming domestic dogs. Rabies elimination can be achieved through the mass vaccination of dogs, but maintaining the critical threshold of vaccination coverage for herd immunity in these populations is hampered by their rapid turnover. Knowledge of the population dynamics of free-roaming dog populations can inform effective planning and implementation of mass dog vaccination campaigns to control rabies. METHODOLOGY/PRINCIPAL FINDINGS: We implemented a health and demographic surveillance system in dogs that monitored the entire owned dog population within a defined geographic area in a community in Mpumalanga Province, South Africa. We quantified demographic rates over a 24-month period, from 1st January 2012 through 1st January 2014, and assessed their implications for rabies control by simulating the decline in vaccination coverage over time. During this period, the population declined by 10%. Annual population growth rates were +18.6% in 2012 and -24.5% in 2013. Crude annual birth rates (per 1,000 dog-years of observation) were 451 in 2012 and 313 in 2013. Crude annual death rates were 406 in 2012 and 568 in 2013. Females suffered a significantly higher mortality rate in 2013 than males (mortality rate ratio [MRR] = 1.54, 95% CI = 1.28-1.85). In the age class 0-3 months, the mortality rate of dogs vaccinated against rabies was significantly lower than that of unvaccinated dogs (2012: MRR = 0.11, 95% CI = 0.05-0.21; 2013: MRR = 0.31, 95% CI = 0.11-0.69). The results of the simulation showed that achieving a 70% vaccination coverage during annual campaigns would maintain coverage above the critical threshold for at least 12 months. CONCLUSIONS AND SIGNIFICANCE: Our findings provide an evidence base for the World Health Organization's empirically-derived target of 70% vaccination coverage during annual campaigns. Achieving this will be effective even in highly dynamic populations with extremely high growth rates and rapid turnover. This increases confidence in the feasibility of dog rabies elimination in Africa through mass vaccination.

Rumen ciliates in the African (Cape) buffalo (Syncerus caffer caffer) living in the vicinity of the Orpen Gate entrance into Kruger National Park, South Africa.

Samples of rumen contents were obtained from 10 African (Cape) buffalo living in the vicinity of the Orpen Gate entrance into Kruger National Park in South Africa. Total number of ciliate protozoa per animal ranged from 3.15 to 23.25 x 103. Forty three different species and forms were observed, of which 35 are a new host record. The total number of species and forms per animal varied from 10 to 17. Eudiplodinium maggii occurred in all 10 animals, followed by Dasytricha ruminantium in nine animals. Diplodinium posterovesiculatum, Eudiplodinium magnodentatum and Ostracodinium mammosum were present in seven animals with all other species and forms occurring in five or less animals. 

Sarcocystis cafferi n. sp. (Protozoa: Apicomplexa) from the African buffalo (Syncerus caffer).

Sarcocystis infections have been reported from the African buffalo ( Syncerus caffer ), but the species have not been named. Here we propose a new name Sarcocystis cafferi from the African buffalo. Histological examination of heart (92), skeletal muscle (36), and tongue (2) sections from 94 buffalos from the Greater Kruger National Park, South Africa, and a review of the literature revealed only 1 species of Sarcocystis in the African buffalo. Macrocysts were up to 12 mm long and 6 mm wide and were located in the neck muscles and overlying connective tissue. They were pale yellow; shaped like a lychee fruit stone or cashew nut; turgid or flaccid and oval to round (not fusiform). By light microscopy (LM) the sarcocyst wall was relatively thin. By scanning electron microscopy (SEM), the sarcocyst wall had a mesh-like structure with irregularly shaped villar protrusions (vp) that were of different sizes and folded over the sarcocyst wall. The entire surfaces of vp were covered with papillomatous structures. By transmission electron microscopy (TEM), the sarcocyst wall was up to 3.6 µm thick and had highly branched villar protrusions that were up to 3 µm long. The villar projections contained filamentous tubular structures, most of which were parallel to the long axis of the projections, but some tubules criss-crossed, especially at the base. Granules were absent from these tubules. Longitudinally cut bradyzoites were 12.1 × 2.7 µm in size, had a long convoluted mitochondrion, and only 2 rhoptries. Phylogenetic analysis of 18S rRNA and cytochrome C oxidase subunit 1 (cox1) gene sequences indicated that this Sarcocystis species is very closely related to, but distinct from, Sarcocystis fusiformis and Sarcocystis hirsuta. Thus, morphological findings by LM, SEM, and TEM together with molecular phylogenetic data (from 18S rRNA and cox1) confirm that the Sarcocystis species in the African buffalo is distinct from S. fusiformis and has therefore been named Sarcocystis cafferi.

First report of a mixed infection of Trichinella nelsoni and Trichinella T8 in a leopard (Panthera pardus) from the Greater Kruger National Park, South Africa.

At least three Trichinella species, namely Trichinella nelsoni, Trichinella britovi and Trichinella zimbabwensis, and one genotype (Trichinella T8), have been isolated from sylvatic carnivores on the African continent. With the exception of T. britovi, the other species are known to circulate in wildlife of the Kruger National Park (KNP), South Africa, and KNP neighbouring game reserves (collectively known as the greater KNP area). Lions (Panthera leo) and spotted hyenas (Crocuta crocuta) appear to be the most important reservoirs of T. nelsoni and Trichinella T8 in the KNP and surrounding areas. Interspecies predation between lions and hyenas has been implicated as a primary mode of maintaining the life cycles of these two Trichinella species. This is the first report of a mixed natural infection of T. nelsoni and Trichinella T8 in a leopard (Panthera pardus) from South Africa. Trichinella muscle larvae were identified to species level by multiplex polymerase chain reaction (PCR). Probable sources of infection, based on the known dietary preference and prey species' range of leopards, are also discussed. The described occurrence of Trichinella species in a leopard from the greater KNP area raises the question of possible sources of infection for this predator species.

Re-emergence of dog rabies in Mpumalanga province, South Africa.

Dog rabies has commonly been associated with the eastern and southern border areas in Mpumalanga province, and the Nkomazi district in the east has been most affected. In other parts of the province, canid rabies has been under control for many years; however, in 2008, dog rabies spread to other parts of the province and resulted in a widespread outbreak. The objective of this study was to genetically characterize rabies viruses in an attempt to determine the source of this recent outbreak. Fifty-five rabies viruses were recovered from domestic dogs between 2000 and 2008 from Mpumalanga province and bordering areas. The viruses were characterized through nucleotide sequencing of the cytoplasmic domain of the glycoprotein gene and the G-L intergenic region. Phylogenetic analysis of these viruses and those previously characterized from Mpumalanga province and neighboring countries and provinces clearly supported the placement of the viruses from the current outbreak and those from Nkomazi district in one lineage. This demonstrated that the recent emergence of rabies in Mpumalanga province resulted from the spread of rabies from Nkomazi district. A comparative analysis demonstrated close genetic relationships among rabies viruses from Mpumalanga and KwaZulu-Natal provinces, Swaziland, and Mozambique. Findings from this investigation have shown that rabies continues to pose a definite public health threat in South Africa, a situation similar to other African countries.