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Rift Valley fever virus (RVFV) is a mosquito-borne, zoonotic phlebovirus-causing disease in domestic ruminants and humans in Africa, the Arabian Peninsula and some Indian Ocean islands. Outbreaks, characterized by abortion storms and a high morbidity rate in newborn animals, occur after heavy and prolonged rainfalls favouring the breeding of mosquitoes. However, the identity of the important mosquito vectors of RVFV is poorly known in most areas. Mosquitoes collected in the Ndumo area of tropical north-eastern KwaZulu-Natal (KZN), South Africa, were tested for RVFV nucleic acid using RT-PCR. The virus was detected in a single pool of unfed Aedes ( Aedimorphus ) durbanensis , indicating that this seasonally abundant mosquito species could serve as a vector in this area of endemic RVFV circulation. Phylogenetic analysis indicated the identified virus is closely related to two isolates from the earliest outbreaks, which occurred in central South Africa more than 60 years ago, indicating long-term endemicity in the region. Further research is required to understand the eco-epidemiology of RVFV and the vectors responsible for its circulation in the eastern tropical coastal region of southern Africa.

Rift Valley fever phlebovirus (RVFV) infects humans and a wide range of ungulates and historically has caused devastating epidemics in Africa and the Arabian Peninsula. Lesions of naturally infected cases of Rift Valley fever (RVF) have only been described in detail in sheep with a few reports concerning cattle and humans. The most frequently observed lesion in both ruminants and humans is randomly distributed necrosis, particularly in the liver. Lesions supportive of vascular endothelial injury are also present and include mild hydropericardium, hydrothorax and ascites; marked pulmonary congestion and oedema; lymph node congestion and oedema; and haemorrhages in many tissues. Although a complete understanding of RVF pathogenesis is still lacking, antigen-presenting cells in the skin are likely the early targets of the virus. Following suppression of type I IFN production and necrosis of dermal cells, RVFV spreads systemically, resulting in infection and necrosis of other cells in a variety of organs. Failure of both the innate and adaptive immune responses to control infection is exacerbated by apoptosis of lymphocytes. An excessive pro-inflammatory cytokine and chemokine response leads to microcirculatory dysfunction. Additionally, impairment of the coagulation system results in widespread haemorrhages. Fatal outcomes result from multiorgan failure, oedema in many organs (including the lungs and brain), hypotension, and circulatory shock. Here, we summarize current understanding of RVF cellular tropism as informed by lesions caused by natural infections. We specifically examine how extant knowledge informs current understanding regarding pathogenesis of the haemorrhagic fever form of RVF, identifying opportunities for future research.

Bluetongue is a serious non-contagious vector-borne viral disease in ruminants, causing poor animal welfare and economic consequences globally. Concern has been raised about the development of novel bluetongue virus (BTV) strains and their possibly altered virulence through the process of viral reassortment. Virulence is traditionally estimated in lethal dose 50 (LD 50 ) studies in murine models, but agreement with both in vitro and virulence in ruminants is questionable, and a refined experimental design is needed. Specific reassortants between wild-type and vaccine strains of BTV-1, -6 and -8 have previously been developed by reverse genetics. The aim of the present study was to rank the in vivo virulence of these parental and reassortant BTV strains by calculating LD 50 in a murine model by using an experimental design that is new to virology: a between-patient optimised three-level response surface pathway design. The inoculation procedure was intracranial. Fifteen suckling mice were used to establish LD 50 for each strain. Three parental and five reassortant virus strains were included. The LD 50 s varied from of 0.1 (95% confidence interval (CI) 0-0.20) to 3.3 (95% CI 2.96-3.72) tissue culture infectious dose 50/ml. The results support the hypothesis that reassortment in BTV may lead to increased virulence in mice with potential negative consequences for the natural ruminant host. The ranking showed low agreement with in vitro properties and virulence in ruminants according to existing literature. Refined design such as response surface pathway design was found suitable for use in virology, and it introduces significant ethical and scientific improvements.

South Africa is endemic for lumpy skin disease and is therefore reliant on various live attenuated vaccines for the control and prevention of the disease. In recent years, widespread outbreaks of vaccine-like strains of lumpy skin disease virus (LSDV) were reported internationally, leading to an increase in the generation of full genome sequences from field isolates. In this study, the complete genomes of six LSDVs submitted during active outbreaks in the 1990s in South Africa were generated. Based on phylogenetic analysis, the six viruses clustered with vaccine strains in LSDV Subgroup 1.1 and are subsequently referred to as vaccine-associated. The genetic differences between the phenotypically distinct vaccine and vaccine-associated strains were 67 single nucleotide polymorphisms (SNPs). This study characterized the location and possible importance of each of these SNPs in their role during virulence and host specificity., (© 2020 Agricultural Research Council. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

This is a report of the complete genome sequences of plaque-selected isolates of five virus strains included in bottle A of the South African Onderstepoort Biological Products commercial live attenuated bluetongue virus vaccine., (Copyright © 2020 Coetzee et al.)

Rift Valley fever (RVF) is a zoonotic viral disease of domestic ruminants in Africa and the Arabian Peninsula caused by a mosquito-borne Phlebovirus. Outbreaks in livestock and humans occur after heavy rains favour breeding of vectors, and the virus is thought to survive dry seasons in the eggs of floodwater-breeding aedine mosquitoes. We recently found high seroconversion rates to RVF virus (RVFV) in cattle and goats, in the absence of outbreaks, in far northern KwaZulu-Natal (KZN), South Africa. Here, we report the prevalence of, and factors associated with, neutralizing antibodies to RVFV in 326 sera collected opportunistically from nyala (Tragelaphus angasii) and impala (Aepyceros melampus) culled during 2016-2018 in two nature reserves in the same area. The overall seroprevalence of RVFV, determined using the serum neutralization test, was 35.0% (114/326; 95%CI: 29.8%-40.4%) and tended to be higher in Ndumo Game Reserve (11/20; 55.0%; 95%CI: 31.5%-76.9%) than in Tembe Elephant Park (103/306; 33.6%; 95%CI: 28.4%-39.3%) (p = .087). The presence of antibodies in juveniles (6/21; 28.6%; 95%CI: 11.3%-52.2%) and sub-adults (13/65; 20.0%; 95%CI: 11.1%-37.8%) confirmed that infections had occurred at least until 2016, well after the 2008-2011 RVF outbreaks in South Africa. Odds of seropositivity was higher in adults than in sub-adults (OR = 3.98; 95%CI: 1.83-8.67; p = .001), in males than in females (OR = 2.66; 95%CI: 1.51-4.68; p = .001) and in animals collected ≤2 km from a swamp or floodplain compared with those collected further away (OR = 3.30; 95%CI: 1.70-6.38; p < .001). Under similar ecological conditions, domestic and wild ruminants may play a similar role in maintenance of RVFV circulation and either or both may serve as the mammalian host in a vector-host reservoir system. The study confirms the recent circulation of RVFV in the tropical coastal plain of northern KZN, providing the basis for investigation of factors affecting virus circulation and the role of wildlife in RVF epidemiology., (© 2020 Blackwell Verlag GmbH.)

Lumpy skin disease (LSD) is an important transboundary animal disease of cattle with significant economic impact because of the implications for international trade in live animals and animal products. LSD is caused by a Capripoxvirus, LSD virus (LSDV), and results in extensive hide and udder damage, fever and pneumonia. LSDV can be shed in semen of infected bulls for prolonged periods and transmitted venereally to cows at high doses. This study examined the effects of LSDV in frozen-thawed semen on in vitro embryo production parameters, including viral status of media and resulting embryos. Bovine oocytes were harvested from abattoir-collected ovaries and split into three experimental groups. After maturation, the oocytes were fertilized in vitro with frozen-thawed semen spiked with a high (HD) or a lower (LD) dose of LSDV, or with LSDV-free semen (control). Following day 7 and day 8 blastocyst evaluation, PCR and virus isolation were performed on all embryonic structures. After completing sufficient replicates to reach 1,000 inseminated oocytes, further in vitro fertilization (IVF) runs were performed to provide material for electron microscopy (EM) and embryo washing procedures. Overall, in vitro embryo yield was significantly reduced by the presence of LSDV in frozen-thawed semen, irrespective of viral dose. When semen with a lower viral dose was used, significantly lower oocyte cleavage rates were observed. LSDV could be detected in fertilization media and all embryo structures, when higher doses of LSDV were present in the frozen-thawed semen used for IVF. Electron microscopy demonstrated LSDV virions inside blastocysts. Following the International Embryo Transfer Society washing procedure resulted in embryos free of viral DNA; however, this may be attributable to a sampling dilution effect and should be interpreted with caution. Further research is required to better quantify the risk of LSDV transmission via assisted reproductive procedures., (© 2019 Blackwell Verlag GmbH.)

The Palyam serogroup orbiviruses are associated with abortion and teratogenesis in cattle and other ruminants. Of the 13 different serotypes that have been identified, the full genome sequence of only one, Kasba, has been published. We undertook to perform Next Generation Sequencing (NGS) and phylogenetic analysis on 12 Palyam serotypes plus field isolates of the African serotypes in our possession. The Palyam serogroup was found to be most closely related to the African horse sickness virus group and showed the most distant evolutionary relationship to the equine encephalosis viruses (EEV). Amino acid sequence analysis revealed that the gene encoding VP7 was the most conserved within serotypes and VP2 and VP5 showed the highest degree of variation. A high degree of sequence identity was found for isolates from the same geographical region. The phylogenetic analysis revealed two clades where the African serotypes were all very closely related in one clade and the other clade contained the Australian and Asian serotypes and one African serotype, Petevo. It was evident from the sequence data that the geographical origin of Palyam serogroup viruses played an important role in the development of the different serotypes.

Background: Rift Valley fever (RVF) is a mosquito-borne zoonotic disease characterized in South Africa by large epidemics amongst ruminant livestock at very long, irregular intervals, mainly in the central interior. However, the presence and patterns of occurrence of the virus in the eastern parts of the country are poorly known. This study aimed to detect the presence of RVF virus (RVFV) in cattle and goats in far northern KwaZulu-Natal province and to estimate the prevalence of antibodies to the virus and the incidence rate of seroconversion., Methodology: Cross-sectional studies were performed in communally farmed cattle (n = 423) and goats (n = 104), followed by longitudinal follow-up of seronegative livestock (n = 253) 14 times over 24 months, representing 160.3 animal-years at risk. Exposure to RVFV was assessed using an IgG sandwich ELISA and a serum neutralization test (SNT) and seroconversion was assessed using SNT. Incidence density was estimated and compared using multivariable Poisson models and hazard of seroconversion was estimated over time., Principal Findings: Initial overall seroprevalence was 34.0% (95%CI: 29.5-38.8%) in cattle and 31.7% (95%CI: 22.9-41.6%) in goats, varying by locality from 18-54%. Seroconversions to RVFV based on SNT were detected throughout the year, with the incidence rate peaking during the high rainfall months of January to March, and differed considerably between years. Overall seroconversion rate in cattle was 0.59 per animal-year (95% CI: 0.46-0.75) and in goats it was 0.41 per animal-year (95% CI: 0.25-0.64), varying significantly over short distances., Conclusions/significance: The high seroprevalence in all age groups and evidence of year-round viral circulation provide evidence for a hyperendemic situation in the study area. This is the first study to directly estimate infection rate of RVFV in livestock in an endemic area in the absence of reported outbreaks and provides the basis for further investigation of factors affecting viral circulation and mechanisms for virus survival during interepidemic periods., Competing Interests: The authors have declared that no competing interests exist.

A prospective study was undertaken during 2013 and 2014, to determine the prevalence of African horse sickness virus (AHSV) in Culicoides midges and the incidence of infection caused by the virus in 28 resident horses on two equine establishments on the East Rand, Gauteng Province, South Africa. Field caught Culicoides midges together with whole blood samples from participating horses were collected every two weeks at each establishment. Culicoides midges and blood samples were tested for the presence of AHSV RNA by real-time quantitative reverse transcription polymerase chain reaction. Nine immunised horses became infected with AHSV during the study period, although infections were subclinical. African horse sickness virus was also identified from a field-collected midge pool. The observations recapitulate previously published data in another setting, where further investigation is warranted to determine what role subclinical infection plays in the diseases epidemiology.

Bluetongue is primarily a disease of sheep in South Africa, while cattle and goats are mostly subclinically infected. The viraemia of bluetongue virus in cattle lasts much longer than in sheep and the role of cattle in the epidemiology of bluetongue in South Africa is poorly understood. Bluetongue virus has a segmented double-stranded ribonucleic acid genome and reassortment of genomes is a common feature. The aim of the study was to investigate whether reassortment occurs between vaccine and field strains when simultaneously administered to cattle. Six cattle between the ages of 6 and 12 months were infected with five strains of modified live vaccine bluetongue virus and a virulent field isolate of bluetongue virus 4. Blood samples were subsequently collected daily from these animals from day 1 to day 39 post-inoculation. Viruses were directly isolated during viraemia from the buffy coat on Vero cells using the plaque forming unit method. Analysis of plaques indicated that no reassortants between virulent field and vaccine strains occurred and the virulent bluetongue virus 4 was identified as the predominant virus strain. However, a reassortant virus between two bluetongue virus vaccine strains was isolated from the buffy coat. Whole genome sequences from the vaccine viruses were compared to the suspected reassortant and it was found that segment 8 exchanged between the bluetongue virus 8 and bluetongue virus 9 vaccine strains. The use of the live-attenuated bluetongue virus multivalent vaccine in South Africa causes circulation of different vaccine serotypes in Culicoides spp. and susceptible hosts and cattle might provide the ideal host for reassortment to occur.

Lumpy skin disease is an economically important disease of cattle, caused by the lumpy skin disease virus (LSDV; Capripoxvirus). It has a variable clinical appearance but, in severely affected animals, is associated with extensive skin damage, pneumonia and death. The LSDV can be found in the semen of infected bulls for prolonged periods of time, from where it can be transmitted by mating or artificial insemination and cause clinical disease in heifers and cows. In this study, an ejaculate was collected from a LSDV seronegative bull and confirmed free from LSDV DNA by PCR. The ejaculate was split into a control sample (C), a sample spiked with a 4 log TCID 50 dose of an LSDV isolate (HD) and a 10 3 dilution of the virus suspension (ND) and frozen routinely. Two straws from each of the different semen treatment groups (HD, ND and C) were subsequently thawed and subjected to swim-up, single layer centrifugation, Percoll ® density gradient and a Percoll ® density gradient with added trypsin. For one set of straws, semen quality variables were recorded, and viral DNA status determined using PCR; the other set was used for positive staining electron microscopy. Samples determined to be positive for LSDV DNA by PCR were then subjected to virus isolation (VI). Complete elimination of LSDV from semen did not occur with use of any of the processing methods. Trypsin did reduce the viral load, and eliminated LSDV from the ND sample, but severely negatively influenced semen quality. The LSDV virions, as assessed by electron microscopy, were associated with the sperm plasma membrane. Further investigation is needed to establish the efficacy of immuno-extenders for rendering semen free from LSDV., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Canine distemper virus (CDV) causes a severe contagious disease in a broad range of hosts. This is the first study to genetically characterise CDV strains from four different wildlife species in South Africa. The phylogenetic diversity of CDV is examined, using the haemagglutinin gene. The South African wildlife CDV isolates showed a high degree of similarity to CDV in South African domestic dogs. Phylogenetic analyses confirmed the presence of 12 geographical lineages with CDV strains from South African wildlife falling within the Southern African lineage. The study reveals two possible co-circulating sub-genotypes corresponding to the northern and southern regions of South Africa respectively. CDV strains from the non-canid species were distinct, but similar to CDV isolates from domestic dog and wild canids. Residues at amino acid sites of the SLAM binding region support the notion that CDV strains encoding 519I / 549H are better adapted to non-canid species than canid species. The amino acids present at site 530 are conserved regardless of host species. Strains from South African wild carnivores showed no difference between host species with all strains presenting 530N. All non-canid strains in this study presented the combination 519I/549H. No evidence of host adaptation or lineage grouping was observed for the Nectin-4 binding region. Further studies should include CDV strains isolated from various hosts from a wider geographical range in South Africa., Competing Interests: The authors have declared that no competing interests exist.

Canine distemper virus causes global multihost infectious disease. This report details complete genome sequences of three vaccine and two new wild-type strains. The wild-type strains belong to the South African lineage, and all three vaccine strains to the America 1 lineage. This constitutes the first genomic sequences of this virus from South Africa., (Copyright © 2017 Loots et al.)

Lumpy skin disease, sheeppox and goatpox are high-impact diseases of domestic ruminants with a devastating effect on cattle, sheep and goat farming industries in endemic regions. In this article, we review the current geographical distribution, economic impact of an outbreak, epidemiology, transmission and immunity of capripoxvirus. The special focus of the article is to scrutinize the use of currently available vaccines to investigate the resource needs and challenges that will have to be overcome to improve disease control and eradication, and progress on the development of safer and more effective vaccines. In addition, field evaluation of the efficacy of the vaccines and the genomic database available for poxviruses are discussed., (© 2015 The Authors. Transboundary and Emerging Diseases Published by Blackwell Verlag GmbH.)

Rift Valley fever (RVF), caused by an arthropod borne Phlebovirus in the family Bunyaviridae, is a haemorrhagic disease that affects ruminants and humans. Due to the zoonotic nature of the virus, a biosafety level 3 laboratory is required for isolation of the virus. Fresh and frozen samples are the preferred sample type for isolation and acquisition of sequence data. However, these samples are scarce in addition to posing a health risk to laboratory personnel. Archived formalin-fixed, paraffin-embedded (FFPE) tissue samples are safe and readily available, however FFPE derived RNA is in most cases degraded and cross-linked in peptide bonds and it is unknown whether the sample type would be suitable as reference material for retrospective phylogenetic studies. A RT-PCR assay targeting a 490 nt portion of the structural G N glycoprotein encoding gene of the RVFV M-segment was applied to total RNA extracted from archived RVFV positive FFPE samples. Several attempts to obtain target amplicons were unsuccessful. FFPE samples were then analysed using next generation sequencing (NGS), i.e. Truseq ® (Illumina) and sequenced on the Miseq ® genome analyser (Illumina). Using reference mapping, gapped virus sequence data of varying degrees of shallow depth was aligned to a reference sequence. However, the NGS did not yield long enough contigs that consistently covered the same genome regions in all samples to allow phylogenetic analysis., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Canine distemper virus (CDV) has emerged as a significant disease of wildlife, which is highly contagious and readily transmitted between susceptible hosts. Initially described as an infectious disease of domestic dogs, it is now recognized as a global multi-host pathogen, infecting and causing mass mortalities in a wide range of carnivore species. The last decade has seen the effect of numerous CDV outbreaks in various wildlife populations. Prevention of CDV requires a clear understanding of the potential hosts in danger of infection as well as the dynamic pathways CDV uses to gain entry to its host cells and its ability to initiate viral shedding and disease transmission. We review recent research conducted on CDV infections in wildlife, including the latest findings on the causes of host specificity and cellular receptors involved in distemper pathogenesis.

In early 2014, abortions and death of ruminants were reported on farms in Maputo and Gaza Provinces, Mozambique. Serologic analysis and quantitative and conventional reverse transcription PCR confirmed the presence of Rift Valley fever virus. The viruses belonged to lineage C, which is prevalent among Rift Valley fever viruses in southern Africa.

Several studies have demonstrated the ability of certain viruses to overwinter in arthropod vectors. The over-wintering mechanism of bluetongue virus (BTV) is unknown. One hypothesis is over-wintering within adult Culicoides midges (Diptera; Ceratopogonidae) that survive mild winters where temperatures seldom drop below 10 °C. The reduced activity of midges and the absence of outbreaks during winter may create the impression that the virus has disappeared from an area. Light traps were used in close association with horses to collect Culicoides midges from July 2010 to September 2011 in the Onderstepoort area, in Gauteng Province, South Africa. More than 500 000 Culicoides midges were collected from 88 collections and sorted to species level, revealing 26 different Culicoides species. Culicoides midges were present throughout the 15 month study. Nine Culicoides species potentially capable of transmitting BTV were present during the winter months. Midges were screened for the presence of BTV ribonucleic acid (RNA) with the aid of a real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay. In total 91.2% of midge pools tested positive for BTV RNA. PCR results were compared with previous virus isolation results (VI) that demonstrated the presence of viruses in summer and autumn months. The results indicate that BTV-infected Culicoides vectors are present throughout the year in the study area. Viral RNA-positive midges were also found throughout the year with VI positive midge pools only in summer and early autumn. Midges that survive mild winter temperatures could therefore harbour BTV but with a decreased vector capacity. When the population size, biting rate and viral replication decrease, it could stop BTV transmission. Over-wintering of BTV in the Onderstepoort region could therefore result in re-emergence because of increased vector activity rather than reintroduction from outside the region.

This is a report of the complete genome sequences of plaque-selected isolates of each of the five virus strains included in a South African commercial trivalent bluetongue virus (BTV) attenuated live virus vaccine, a BTV-4 field strain isolated from Rustenburg, South Africa, in 2011, and a bluetongue reassortant (bluetongue virus 4 strain 4/O. aries-tc/ZAF/11/OBP-115) isolated from experimentally vaccinated cattle. Full-genome sequencing and phylogenetic analyses show that the bluetongue virus 9 strain 9/B. taurus-tc/ZAF/15/Onderstepoort&#95;B02b is a reassortant virus containing segments from both BTV-9 and BTV-8., (Copyright © 2016 van den Bergh et al.)

Phylogenetic networks and sequence analysis allow a more accurate understanding of the serotypes, genetic relationships and epidemiology of viruses. Based on gene sequences of the conserved segment 10 (NS3), bluetongue virus (BTV) can be divided into five topotypes. In this molecular epidemiology study, segment 10 sequence data of 11 isolates obtained from the Virology Section of the Department of Veterinary Tropical Diseases, Faculty of Veterinary Science, University of Pretoria, Onderstepoort, were analyzed and compared to sequence data of worldwide BTV strains available in the GenBank database. The consensus nucleotide sequences of NS3/A showed intermediate levels of variation, with the nucleotide sequence identity ranging from 79.72 % to 100 %. All 11 strains demonstrated conserved amino acid characteristics. Phylogenetic networks were used to identify BTV topotypes. The phylogeny obtained from the nucleotide sequence data of the NS3/A-encoding gene presented three major and two minor topotypes. The clustering of strains from different geographical areas into the same group indicated spatial spread of the segment 10 genes, either through gene reassortment or through the introduction of new strains from other geographical areas via trade. The effect of reassortment and genetic drift on BTV and the importance of correct serotyping to identify viral strains are highlighted.

This is a report of the complete genome sequences of plaque-selected isolates of each of the four virus strains included in a South African commercial tetravalent African horse sickness attenuated live virus vaccine., (Copyright © 2015 Guthrie et al.)

Bluetongue virus (BTV) has since 1998 extended its distribution further North than where it has previously been encountered. Changes in the epidemiology of Bluetongue (BT), as well as novel features of recent outbreaks of BTV in Europe, have stimulated research on BTV‑vector‑host interaction. The outbreak of BTV‑8 in Northern Europe from 2006‑2008 is particular noteworthy in this regard, as the European strain of BTV‑8 demonstrated novel properties, including high virulence - especially for cattle - and the capability to cross the ruminant placenta. The virus was in addition transmitted by indigenous European Culicoides species that had not previously been implicated in the widespread transmission of BTV. Recent advances in the scientific understanding of BTV‑vector‑host interaction include increased knowledge of the virus' replication cycle, the role of biotic factors in influencing viral infection of the insect vector, increased knowledge of BTV immunology and pathogenesis in the mammalian host, and increased knowledge of virulence and pathogenicity features of newly discovered serotypes/strains of the virus. New research on aspects of BTV‑vector‑host interaction has been driven in part by developments in molecular biology and experimental infection biology, of which next generation sequencing, the expression of individual viral proteins in cell culture, the establishment of a reverse genetics system for the virus, the development of novel in vitro and in vivo infection models, and refinement of existing BTV experimental infection methodologies have proven instrumental. Moreover, these developments have also provided the opportunity for the development of novel vaccine strategies. This article provides a synopsis of selected recent advances that have been made in the understanding of BTV‑vector‑host interaction, with a particular focus on research that has been conducted in Europe over the last 5 years.

This is a report of the complete genome sequences of plaque-selected isolates of each of the three virus strains included in a South African commercial trivalent African horse sickness attenuated live virus vaccine., (Copyright © 2015 Guthrie et al.)

Rift Valley fever (RVF) is an arthropod-borne viral disease of importance in livestock and humans. Epidemics occur periodically in domestic ruminants. People in contact with infected livestock may develop disease that varies from mild flu-like symptoms to fatal viraemia. Livestock vaccination may assist in disease control. Rift Valley fever virus (RVFV) Clone 13 is a relatively new vaccine against RVF, derived from an avirulent natural mutant strain of RVFV, and has been shown to confer protective immunity against experimental infection with RVFV. The hypothesis tested in the current trial was that rams vaccinated with RVFV Clone 13 vaccine would not experience a reduction in semen quality (measured by evaluating the percentage progressively motile and percentage morphologically normal spermatozoa in successive ejaculates) relative to unvaccinated control animals. Ram lambs were screened for antibodies to RVFV using a serum neutralisation test. Animals without detectable antibodies (n = 23) were randomly allocated to either a test group (n = 12) or a control group (n = 11). Animals in the test group were vaccinated with RVFV Clone 13 vaccine. Daily rectal temperature measurements and weekly semen and blood samples were taken from all animals. Seven animals were eliminated from the statistical analysis because of potential confounding factors. Logistic regression analysis was performed on data gathered from the remaining animals to determine whether an association existed between animal group, rectal temperature and semen quality parameters. No correlation existed between the treatment group and values obtained for the semen quality parameters measured. There was no statistically significant post-vaccination decline in the percentage of live morphologically normal spermatozoa, or the percentage of progressively motile spermatozoa, either when assessed amongst all animals or when assessed within individual groups. A repeat study with a larger sample size and a more comprehensive pre-screening process may be indicated to avoid the inclusion of unsuitable animals.

Ostrich (Struthio camelus) chicks less than 3 mo age are observed to experience a high mortality rate that is often associated with enteritis. This study was undertaken to investigate the infectious bacteria implicated in ostrich chick enteritis. Postmortems were performed on 122 ostrich chicks aged from 1 d to 3 mo and intestinal samples were subjected to bacterial culture. Bacterial isolates were typed by PCR and serotyping. Escherichia coli (E. coli; 49%) was the most frequently isolated from the samples followed by Clostridium perfringens (C. perfringens; 20%), Enterococcus spp. (16%), and Salmonella spp. (7%). Of the E. coli, 39% were categorized as enteropathogenic E. coli, 4% enterotoxigenic E. coli, and no enterohaemorrhagic E. coli were found. The majority (93%) of C. perfringens was Type A and only 7% was Type E. C. perfringens Types B through D were not present. The netB gene that encodes NetB toxin was identified from 16% of the C. perfringens isolated. All the C. perfringens Type E harbored the netB gene and just 10% of the C. perfringens Type A had this gene. Three Salmonella serotypes were identified: Salmonella Muenchen (S. Muenchen; 80%), S. Hayindongo (13%), and S. Othmarschen (7%). The indication is that the cause of enteritis in ostrich chicks is bacterial-involving: enteropathogenic E. coli and enterotoxigenic E. coli; C. perfringens Types A and E (with the possible influence of netB gene); and S. Muenchen, S. Hayindongo, and S. Othmarschen., (© 2015 Poultry Science Association Inc.)

Lumpy skin disease (LSD) is an economically important disease caused by LSD virus (LSDV), a Capripoxvirus, characterized by fever and circumscribed skin lesions. It is suspected to be transmitted mechanically by biting flies. To assess the vector potential of Amblyomma hebraeum in transmission of LSDV, mechanical/intrastadial and transstadial modes of transmission of the virus by this tick species were investigated. Two cattle were artificially infected as sources (donors) of infection to ticks. Ticks were infected as either nymphs or adults. Male A. hebraeum ticks were partially fed on donor animals and transferred to recipient animals to test for mechanical/intrastadial transmission. Nymphal A. hebraeum were fed to repletion on donor animals. The emergent adult ticks were placed on recipient animals to test for transstadial transmission of the virus. Successful transmission of LSDV infection was determined in recipient animals by monitoring development of clinical signs, testing of blood for the presence of LSDV by real-time PCR, virus isolation and the serum neutralization test. This report provides further evidence of mechanical/intrastadial and, for the first time, transstadial transmission of LSDV by A. hebraeum. These findings implicate A. hebraeum as a possible maintenance host in the epidemiology of the disease., (© 2013 Blackwell Verlag GmbH.)

Lumpy skin disease (LSD) is of substantial economic importance for the cattle industry in Africa and the Near and Middle East. Several insect species are thought to transmit the disease mechanically. Recent transmission studies have demonstrated the first evidence for a role of hard (ixodid) ticks as vectors of lumpy skin disease virus (LSDV). The aim of this study was to attempt in vitro growth of the virus in Rhipicephalus spp. tick cell lines and investigate in vivo the presence of the virus in ticks collected from cattle during LSD outbreaks in Egypt and South Africa. No evidence was obtained for replication of LSDV in tick cell lines although the virus was remarkably stable, remaining viable for 35 days at 28°C in tick cell cultures, in growth medium used for tick cells and in phosphate buffered saline. Viral DNA was detected in two-thirds of the 56 field ticks, making this the first report of the presence of potentially virulent LSDV in ticks collected from naturally infected animals., (Crown Copyright © 2014. Published by Elsevier GmbH. All rights reserved.)

Inclusion body hepatitis is an acute disease of chickens ascribed to viruses of the genus Aviadenovirus and referred to as fowl adenovirus (FAdV). There are 12 FAdV types (FAdV1to FAdV8a and FAdV8b to FAdV11), classified into five species based on their genotype (designated FAdVA to FAdVE). A total of 218 000 chickens, 2-29 days of age, were affected over a 1-year period, all testing positive by microscopy, virus isolation and confirmation with polymerase chain reaction (PCR). Affected birds were depressed, lost body weight,were weak and had watery droppings. Pathological changes observed during necropsy indicated consistent changes in the liver, characterised by hepatomegaly, cholestasis and hepatitis. Lesions were also discernible in the spleen, kidney and gizzard wall and were characterised by splenomegaly, pinpoint haemorrhages, nephritis with haemorrhage,visceral gout and serosal ecchymosis of the gizzard wall. Histopathological lesions were most consistently observed in the liver but could also be seen in renal and splenic tissue. Virus isolation was achieved in embryonated eggs and most embryos revealed multifocalto diffuse hepatic necrosis, with a mixed cellular infiltrate of macrophages and heterophils(necro-granulomas), even in the absence of macroscopic pathology. Virus isolation results were verified by histopathology and PCR on embryonic material and further characterised by nucleotide sequence analysis. Two infectious bursal disease virus isolates were also made from the Klerksdorp flock. Nucleotide sequence analysis of the L1 hexon loop of all the FAdV isolates indicated homology (99%) with prototype strains P7-A for FAdV-2, as well as for FAdV-8b.

In South Africa, outbreaks of African horse sickness (AHS) occur in summer; no cases are reported in winter, from July to September. The AHS virus (AHSV) is transmitted almost exclusively by Culicoides midges (Diptera: Ceratopogonidae), of which Culicoides imicola is considered to be the most important vector. The over-wintering mechanism of AHSV is unknown. In this study, more than 500 000 Culicoides midges belonging to at least 26 species were collected in 88 light traps at weekly intervals between July 2010 and September 2011 near horses in the Onderstepoort area of South Africa. The dominant species was C. imicola. Despite relatively low temperatures and frost, at least 17 species, including C. imicola, were collected throughout winter (June-August). Although the mean number of midges per night fell from &gt; 50 000 (March) to &lt; 100 (July and August), no midge-free periods were found. This study, using virus isolation on cell cultures and a reverse transcriptase polymerase chain reaction (RT-PCR) assay, confirmed low infection prevalence in field midges and that the detection of virus correlated to high numbers. Although no virus was detected during this winter period, continuous adult activity indicated that transmission can potentially occur. The absence of AHSV in the midges during winter can be ascribed to the relatively low numbers collected coupled to low infection prevalence, low virus replication rates and low virus titres in the potentially infected midges. Cases of AHS in susceptible animals are likely to start as soon as Culicoides populations reach a critical level.

Fowl adenovirus (FAdV) is a member of the genus Aviadenovirus and causes a number of economically important poultry diseases. One of these diseases, inclusion body hepatitis (IBH), has a worldwide distribution and is characterised by acute mortality (5% - 20%) in production chickens. The disease was first described in the United States of America in 1963 and has also been reported in Canada, the United Kingdom, Australia, France and Ireland, but until now, not in South Africa. Adenoviruses isolated from the first outbreak of IBH in South Africa were able to reproduce the disease in chicken embryo livers. The aim of the present study was to characterise the viruses and determine the pathogenicity of the FAdV strains responsible for the first reported case of IBH in South Africa. Polymerase chain reaction (PCR) amplification of the L1 loop region of the fowl adenovirus hexon gene using degenerate primer pair hexon A/B was used to identify the viruses that were isolated. Restriction fragment length polymorphism (RFLP) of the amplification products was used for the differentiation of 14 isolates of fowl adenovirus. Sequencing of the PCR products followed by amino acid comparison and phylogenetic analysis using the L1 loop region of the hexon protein was done to determine the identity of the isolates. Amino acid sequences of the hexon genes of all the South African isolates were compared with those of reference strains representing FAdV species. Amino acid comparison of 12 South Africa field isolates to FAdV reference strains revealed a high sequence identity (&gt; 93.33%) with reference strains T8-A and 764. Two of the isolates had high sequence identity (93.40%) with reference strains P7-A, C2B and SR48. Phylogenetic analysis of the L1 loop region of the hexon protein of all 14 South African isolates was consistent with their RFLP clusters. The mortality rates of embryos challenged with 106 egg infective doses (EID50) FAdV 2 were 80% - 87% and mortality rates for embryos challenged with 105.95 (EID50) FAdV 8b were 65% - 80%.

Bluetongue (BT) is a non-contagious disease of sheep and other domestic and wild ruminants caused by the bluetongue virus (BTV). Currently 26 serotypes of the virus have been identified. In South Africa, 22 serotypes have been identified and BT is controlled mainly by annual vaccinations using a freeze-dried live attenuated polyvalent BTV vaccine. The vaccine is constituted of 15 BTV serotypes divided into three separate bottles and the aim is to develop a vaccine using fewer serotypes without compromising the immunity against the disease. This study is based on previously reported cross-neutralisation of specific BTV serotypes in in vitro studies. Bluetongue virus serotype 4 was selected for this trial and was tested for cross-protection against serotype 4 (control), 1 (unrelated serotype), 9, 10 and 11 in sheep using the serum neutralisation test. The purpose of the study was to determine possible cross-protection of different serotypes in sheep. Of those vaccinated with BTV-4 and challenged with BTV-1, which is not directly related to BTV-4, 20% were completely protected and 80% showed clinical signs, but the reaction was not as severe as amongst the unvaccinated animals. In the group challenged with BTV-10, some showed good protection and some became very sick. Those challenged with BTV-9 and BTV-11 had good protection. The results showed that BTV-4 does not only elicit a specific immune response but can also protect against other serotypes.

Rift Valley fever and lumpy skin disease are transboundary viral diseases endemic in Africa and some parts of the Middle East, but with increasing potential for global emergence. Wild ruminants, such as the African buffalo (Syncerus caffer), are thought to play a role in the epidemiology of these diseases. This study sought to expand the understanding of the role of buffalo in the maintenance of Rift Valley fever virus (RVFV) and lumpy skin disease virus (LSDV) by determining seroprevalence to these viruses during an inter-epidemic period. Buffaloes from the Kruger National Park (n = 138) and Hluhluwe-iMfolozi Park (n = 110) in South Africa were sampled and tested for immunoglobulin G (IgG) and neutralising antibodies against LSDV and RVFV using an indirect enzyme-linked immunosorbent assay (I-ELISA) and the serum neutralisation test (SNT). The I-ELISA for LSDV and RVFV detected IgG antibodies in 70 of 248 (28.2%) and 15 of 248 (6.1%) buffaloes, respectively. Using the SNT, LSDV and RVFV neutralising antibodies were found in 5 of 66 (7.6%) and 12 of 57 (21.1%), respectively, of samples tested. The RVFV I-ELISA and SNT results correlated well with previously reported results. Of the 12 SNT RVFV-positive sera, three (25.0%) had very high SNT titres of 1:640. Neutralising antibody titres of more than 1:80 were found in 80.0% of the positive sera tested. The LSDV SNT results did not correlate with results obtained by the I-ELISA and neutralising antibody titres detected were low, with the highest (1:20) recorded in only two buffaloes, whilst 11 buffaloes (4.4%) had evidence of co-infection with both viruses. Results obtained in this study complement other reports suggesting a role for buffaloes in the epidemiology of these diseases during inter-epidemic periods.

It is known that lumpy skin disease virus (LSDV) can be shed in bull semen following infection and also that artificial insemination (AI) poses a biosecurity risk. However, it is not known whether the use of LSDV infected semen in AI poses a biosecurity risk. The aim of this study was to investigate whether LSDV, transmitted through semen, can infect cows and their embryos. Two controlled trials were performed simultaneously. Eleven young beef heifers, naïve to LSDV, were synchronized using an OvSynch protocol and inseminated on Day 0 with fresh semen spiked with a field strain of LSDV on day 0. Six of the heifers were superovulated on Day 1 using pregnant mare serum gonadotropin, and embryos were flushed from these heifers on Day 6. Blood and serum samples were collected from Day 4 until Day 27 to determine the presence of LSDV by PCR and virus isolation, and the presence of antibodies against LSDV by SNT. The first clinical signs of LSD were noticed on Day 10, followed by severe generalized LSD in three heifers and mild LSD in two more heifers. Two heifers were humanely euthanized due to severe unresponsive stranguria. LSDV was detected by PCR, virus isolation or electron microscopy in blood, embryos and organs of experimentally infected animals; and eight heifers had seroconverted by Day 27. Two control animals were not affected. This is the first report of experimental seminal transmission of LSDV in cattle., (© 2013 Blackwell Verlag GmbH.)

Bluetongue virus (BTV), a segmented dsRNA virus, is the causative agent of bluetongue (BT), an economically important viral haemorrhagic disease of ruminants. Bluetongue virus can exchange its genome segments in mammalian or insect cells that have been co-infected with more than one strain of the virus. This process, may potentially give rise to the generation of novel reassortant strains that may differ from parental strains in regards to their phenotypic characteristics. To investigate the potential effects of reassortment on the virus' phenotype, parental as well as reassortant strains of BTV serotype 1, 6, 8, that were derived from attenuated and wild type strains by reverse genetics, were studied in vitro for their virus replication kinetics and cytopathogenicity in mammalian (Vero) cell cultures. The results indicate that genetic reassortment can affect viral replication kinetics, the cytopathogenicity and extent/mechanism of cell death in infected cell cultures. In particular, some reassortants of non-virulent vaccine (BTV-1 and BTV-6) and virulent field origin (BTV-8) demonstrate more pronounced cytopathic effects compared to their parental strains. Some reassortant strains in addition replicated to high titres in vitro despite being composed of genome segments from slow and fast replicating parental strains. The latter result may have implications for the level of viraemia in the mammalian host and subsequent uptake and transmission of reassortant strains (and their genome segments) by Culicoides vectors. Increased rates of CPE induction could further suggest a higher virulence for reassortant strains in vivo. Overall, these findings raise questions in regards to the use of modified-live virus (MLV) vaccines and risk of reassortment in the field. To further address these questions, additional experimental infection studies using insects and/or animal models should be conducted, to determine whether these results have significant implications in vivo., (Copyright © 2014 Elsevier B.V. All rights reserved.)

A major problem with the testing of virucidal efficacy using current protocols is that scoring of virus-induced cytopathic effect (CPE) is dependent on subjective visual interpretation using light microscopy. The current report details the use of an electrical impedance assay (xCELLigence, ACEA Biosciences) for its utility in virucidal efficacy testing. In this study, the xCELLigence system was used in a procedure developed from guidelines given by the Deutsche Vereiniging zur Bekämpfung der Viruskrankheiten (DVV) (German Association for the Control of Virus Diseases) in order to demonstrate the inactivation of infectious bursal disease virus using a commercial virucide. Although the modified DVV assay using the xCELLigence system yielded identical results (i.e. a 5-log10 reduction in viral infectivity) as the traditional DVV assay, the system allows virucidal efficacy and cytotoxicity to be measured in a more precise and reproducible fashion., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Experimental infection studies with bluetongue virus (BTV) in the mammalian host have a history that stretches back to the late 18th century. Studies in a wide range of ruminant and camelid species as well as mice have been instrumental in understanding BTV transmission, bluetongue (BT) pathogenicity/pathogenesis, viral virulence, the induced immune response, as well as reproductive failures associated with BTV infection. These studies have in many cases been complemented by in vitro studies with BTV in different cell types in tissue culture. Together these studies have formed the basis for the understanding of BTV-host interaction and have contributed to the design of successful control strategies, including the development of effective vaccines. This review describes some of the fundamental and contemporary infection studies that have been conducted with BTV in the mammalian host and provides an overview of the principal animal welfare issues that should be considered when designing experimental infection studies with BTV in in vivo infection models. Examples are provided from the authors' own laboratory where the three Rs (replacement, reduction and refinement) have been implemented in the design of experimental infection studies with BTV in mice and goats. The use of the ARRIVE guidelines for the reporting of data from animal infection studies is emphasized., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Lumpy skin disease (LSD) is caused by lumpy skin disease virus (LSDV), a member of the genus Capripoxvirus. Transmission of the virus has been associated with haematophagous insects such as Stomoxys calcitrans as well as Aedes and Culex species of mosquitoes. Recent studies have reported the transmission of the virus by Amblyomma hebraeum, Rhipicephalus appendiculatus, and Rhipicephalus decoloratus ticks and the presence of LSDV in saliva of A. hebraeum and R. appendiculatus ticks. The aim of this study was to determine which tick organs become infected by LSDV following intrastadial infection and transstadial persistence of the virus in A. hebraeum and R. appendiculatus ticks. Nymphal and adult ticks were orally infected by feeding them on LSDV-infected cattle. Partially fed adult ticks were processed for testing while nymphs were fed to repletion and allowed to moult to adults before being processed for testing. The infection in tick organs was determined by testing for the presence of the viral antigen using monoclonal antibodies with immunohistochemical staining. The viral antigen was detected in salivary glands, haemocytes, synganglia, ovaries, testes, fat bodies, and midgut. Since the virus was shown to be able to cross the midgut wall and infect various tick organs, this may indicate potential for biological development and transmission of LSDV in ticks. This study strengthens the previously reported evidence of the occurrence of LSDV in tick saliva., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

Lumpy skin disease is a debilitating cattle disease caused by the lumpy skin disease virus (LSDV), belonging to the genus Capripoxvirus. Epidemics of the disease usually occur in summer, when insect activity is high. Limited information is available on how LSDV persists during inter-epidemic periods. Transmission of LSDV by mosquitoes such as Aedes aegypti has been shown to be mechanical, there is no carrier state in cattle and the role of wildlife in the epidemiology of the disease seems to be of minor importance. Recent studies in ticks have shown transstadial persistence of LSDV in Rhipicephalus appendiculatus and Amblyomma hebraeum as well as transovarial persistence of the virus in Rhipicephalus decoloratus, R. appendiculatus and A. hebraeum. The over-wintering of ticks off the host as part of their life cycles is well known: A. hebraeum and R. appendiculatus over-winter, for example, on the ground as engorged nymphs/unfed (emergent) adults while R. decoloratus over-winters on the ground as engorged females. In this study, transstadial and transovarial persistence of LSDV from experimentally infected A. hebraeum nymphs and R. decoloratus females after exposure to cold temperatures of 5 °C at night and 20 °C during the day for 2 months was reported. This observation suggests possible over-wintering of the virus in these tick species.

Lumpy skin disease (LSD), an acute, sub-acute or inapparent disease of cattle, is caused by lumpy skin disease virus (LSDV), a member of the genus Capripoxvirus in the family Poxviridae. LSD is characterised by high fever, formation of circumscribed skin lesions and ulcerative lesions on the mucous membranes of the mouth, respiratory and digestive tracts. It is an economically important disease due to the permanent damage to hides, the reduction in productivity and trade restrictions imposed on affected areas. Transmission has been associated with blood-feeding insects such as stable flies (Stomoxysis calcitrans) and mosquitoes (Aedes aegypti). Mechanical (intrastadial) and transstadial transmission by Amblyomma hebraeum and Rhipicephalus appendiculatus as well as transovarial transmission by R. decoloratus have been reported. In this study transovarial passage of LSDV to larvae and subsequent transmission to recipient animals were demonstrated. The finding of transovarial passage of LSDV in female ticks shows the potential for A. hebraeum, R. appendiculatus and R. decoloratus to be reservoir hosts for LSDV.

Lumpy skin disease is an economically important disease of cattle that is caused by the lumpy skin disease virus (LSDV), which belongs to the genus Capripoxvirus. It is endemic in Africa and outbreaks have also been reported in the Middle-East. Transmission has mostly been associated with blood-feeding insects but recently, the authors have demonstrated mechanical transmission by Rhipicephalus appendiculatus as well as mechanical/intrastadial and transstadial transmission by Amblyomma hebraeum. Saliva is the medium of transmission of pathogens transmitted by biting arthropods and, simultaneously, it potentiates infection in the vertebrate host. This study aimed to detect LSDV in saliva of A. hebraeum and R. appendiculatus adult ticks fed, as nymphs or as adults, on LSDV-infected animals, thereby also demonstrating transstadial or mechanical/intrastadial passage of the virus in these ticks. Saliva samples were tested for LSDV by real-time PCR and virus isolation. Supernatants obtained from virus isolation were further tested by real-time PCR to confirm that the cytopathic effects observed were due to LSDV. Lumpy skin disease virus was detected, for the first time, in saliva samples of both A. hebraeum and R. appendiculatus ticks. At the same time, mechanical/intrastadial and transstadial passage of the virus was demonstrated and confirmed in R. appendiculatus and A. hebraeum.

Since the emergence of canine parvovirus type-2 (CPV-2) in the early 1970s, it has been evolving into novel genetic and antigenic variants (CPV-2a, 2b and 2c) that are unevenly distributed throughout the world. Genetic characterization of CPV-2 has not been documented in Africa since 1998 apart from the study carried out in Tunisia 2009. A total of 139 field samples were collected from South Africa and Nigeria, detected using PCR and the full length VP2-encoding gene of 27 positive samples were sequenced and genetically analyzed. Nigerian samples (n=6), South Africa (n=19) and vaccine strains (n=2) were compared with existing sequences obtained from GenBank. The results showed the presence of both CPV-2a and 2b in South Africa and only CPV-2a in Nigeria. No CPV-2c strain was detected during this study. Phylogenetic analysis showed a clustering not strictly associated with the geographical origin of the analyzed strains, although most of the South African strains tended to cluster together and the viral strains analyzed in this study were not completely distinct from CPV-2 strains from other parts of the world. Amino acid analysis showed predicted amino acid changes., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The capability of the recently emerged European strain of bluetongue virus serotype 8 (BTV-8) to cross the ruminant placenta has been established in experimental and field studies in both sheep and cattle. Seroprevalence rates in goats in North-Western Europe were high during the recent outbreak of BTV-8; however the capability of the virus to infect goats through the transplacental route has not been established. In the present study, four Saanen goats were inoculated with the European strain of BTV-8 at 62 days of gestation; this resulted in mild clinical signs, however gross lesions observed post mortem were more severe. Viral RNA was detected by real-time RT-PCR in blood and tissue samples from three fetuses harvested from two goats at 43 days post infection. Conventional RT-PCR and genome sequencing targeting viral segment 2 confirmed infection of brain tissue with BTV-8 in two of these fetuses. In total, five of six fetuses demonstrated lesions that may have been associated with transplacental infection with BTV. Infected fetuses did not demonstrate neurological lesions. Low viral RNA concentrations in fetal blood and tissue further suggest that the infected fetuses would probably not have been born viraemic. The implications of these findings with regards to the epidemiology and overwintering of BTV-8 in Europe remains unclear., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Lumpy skin disease (LSD) is an economically important acute or sub-acute disease of cattle that occurs across Africa and in the Middle East. The aim of this study was to assess whether Rhipicephalus decoloratus ticks were able to transmit lumpy skin disease virus (LSDV) transovarially. Uninfected, laboratory-bred R. decoloratus larvae were placed to feed on experimentally infected "donor" cattle. After completion of the life cycle on donor animals, fully engorged adult female ticks were harvested and allowed to lay eggs. Larvae that hatched from these eggs were then transferred to feed on uninfected "recipient" cattle. The latter became viraemic and showed mild clinical disease with characteristic skin lesions and markedly enlarged precrural and subscapular lymph nodes. This is the first report of transovarial transmission of poxviruses by R. decoloratus ticks, and the importance of this mode of transmission in the spread of LSDV in endemic settings requires further investigation., (Crown Copyright © 2013. Published by Elsevier GmbH. All rights reserved.)

Background: Although reported sporadically from various countries, feline babesiosis appears to be a significant clinical entity only in South Africa, where Babesia felis is usually incriminated as the causative agent. Babesia lengau, recently described from asymptomatic cheetahs, has now possibly been incriminated as the causative agent in two severe clinical cases in domestic cats., Findings: Both cats were euthanised in extremis. While typical feline babesiosis in South Africa is an afebrile disease with a chronic manifestation, there was acute onset of severe clinical signs in both cats and their body temperatures were above the normal range when they were presented for treatment. Haemolytic anaemia was confirmed in one case. To our knowledge, this is the first report of cerebral babesiosis in cats.On reverse line blot 18S rDNA PCR products obtained from both cats showed positive hybridization profiles with the B. lengau species-specific probe. The two partial parasite 18S rRNA gene sequences obtained, showed high sequence similarity (99.9%) to B. lengau. In a representative tree constructed by the neighbor-joining method using the two-parameter model of Kimura the two obtained partial 18S rDNA sequences and that of B. lengau formed a monophyletic group with B. conradae and sequences previously isolated from humans and wildlife in the western USA., Conclusion: All clinical cases of feline babesiosis in South Africa are not necessarily caused by B. felis. Other piroplasms, e.g. B. lengau, may be incriminated in clinical cases, especially those occurring outside the known endemic area.

Lumpy skin disease (LSD) is an economically important, acute or sub-acute, viral disease of cattle that occurs across Africa and in the Middle East. The aim of this study was to investigate if lumpy skin disease virus (LSDV) can be transmitted mechanically by African brown ear ticks (Rhipicephalus appendiculatus Neum.). Laboratory-bred R. appendiculatus males were fed on experimentally infected viraemic 'donor' cattle. Partially fed male ticks were then transferred to feed on an uninfected 'recipient' cow. The recipient animal became viraemic, showed mild clinical signs of LSD and seroconverted. Additionally, R. appendiculatus males were found to transmit LSDV through feeding on skin lacking visible lesions, demonstrating that viraemic animals without lesions at the feeding site of ticks may be a source of infection. This is the first time that transmission of poxviruses by a tick species has been demonstrated and the importance of this mode of transmission in the spread of LSDV in endemic settings is discussed.

Bovine viral diarrhoea virus (BVDV) has emerged as one of the economically important pathogens in cattle populations, with a worldwide distribution and causing a complex of disease syndromes. Two genotypes, BVDV 1 and 2, exist and are discriminated on the basis of the sequence of the 5' non-coding region (5' NCR) using real-time PCR. Real-time PCR is more sensitive, specific, and less time-consuming than conventional PCR, and it has less risk of cross-contamination of samples. Limited information exists on BVDV genetic subtypes in South Africa. The aim of this study was to determine the genotypes of BVDV currently circulating in South African feedlots. A total of 279 specimens (219 tissue samples, 59 trans-tracheal aspirates and 1 blood sample) were collected from dead and living cattle with lesions or clinical signs compatible with BVDV infection. Pooled homogenates from the same animals were prepared, and total RNA was extracted. A screening test was performed on the pooled samples, and positive pools were investigated individually. A Cador BVDV Type 1/2 RT-PCR Kit (QIAGEN, Hilden, Germany) was used for the real-time PCR assay on a LightCycler(®) V2.0 real-time PCR machine (Roche Diagnostics, Mannheim, Germany). The results were read at 530 and 640 nm for BVDV 1 and 2, respectively. Bovine viral diarrhoea virus was detected in a total of 103 samples that included 91 tissue samples, 1 blood sample and 11 trans-tracheal aspirates. Eighty-five (82.5 %) of the strains were genotype 1 and 18 (17.5 %) were genotype 2. Comparing the sequencing data, genotypes 1 and 2 from the field strains did not cluster with vaccine strains currently used in feedlots in South Africa. The present study revealed the presence of BVDV genotype 2 in cattle in South Africa based on the high sequence similarity between genotype 2 field strains and strain 890 from North America. The presence of genotype 2 viruses that phylogenetically belong to different clusters and coexist in feedlots is consistent with the possibility of multiple virus introductions. These results represent the first documented evidence for the presence of BVDV genotype 2 in African cattle.

Bluetongue virus (BTV) is the prototype member of the Orbivirus genus in the family Reoviridae and is the aetiological agent of the arthropod transmitted disease bluetongue (BT) that affects both ruminant and camelid species. The disease is of significant global importance due to its economic impact and effect on animal welfare. Bluetongue virus, a dsRNA virus, evolves through a process of quasispecies evolution that is driven by genetic drift and shift as well as intragenic recombination. Quasispecies evolution coupled with founder effect and evolutionary selective pressures has over time led to the establishment of genetically distinct strains of the virus in different epidemiological systems throughout the world. Bluetongue virus field strains may differ substantially from each other with regards to their phenotypic properties (i.e. virulence and/or transmission potential). The intrinsic molecular determinants that influence the phenotype of BTV have not clearly been characterized. It is currently unclear what contribution each of the viral genome segments have in determining the phenotypic properties of the virus and it is also unknown how genetic variability in the individual viral genes and their functional domains relate to differences in phenotype. In order to understand how genetic variation in particular viral genes could potentially influence the phenotypic properties of the virus; a closer understanding of the BTV virion, its encoded proteins and the evolutionary mechanisms that shape the diversity of the virus is required. This review provides a synopsis of these issues and highlights some of the studies that have been conducted on BTV and the closely related African horse sickness virus (AHSV) that have contributed to ongoing attempts to identify the molecular determinants that influence the virus' phenotype. Different strategies that can be used to generate BTV mutants in vitro and methods through which the causality between particular genetic modifications and changes in phenotype may be determined are also described. Finally examples are highlighted where a clear understanding of the molecular determinants that influence the phenotype of the virus may have contributed to risk assessment and mitigation strategies during recent outbreaks of BT in Europe., (Copyright © 2012 Elsevier B.V. All rights reserved.)

In 2008, a suspected outbreak of Rift Valley fever (RVF) was reported on a farm in the Bela-Bela area, Limpopo Province, South Africa. Seven calves died on the affected dairy farm, where no RVF vaccination programme was practised. No apparent clinical disease was reported in the other 300 cattle (33 calves included) or 200 sheep on the farm. During the outbreak, blood samples from 77.7% (233/300) of the cattle and 36.5% (73/200) of the sheep were collected on the affected farm and 55 blood samples were taken from cattle on a neighbouring farm. Eight weeks later, 78% of the cattle (234/300) and 42.5% of the sheep (85/200) were bled on the affected farm only. All sera were tested by an Immunoglobulin M (IgM)-capture Enzymelinked immunosorbent assay (ELISA) and by an indirect Immunoglobulin G (IgG) ELISA. Selected IgM-positive (n = 14), IgG-positive (n = 23) and samples negative for both IgM and IgG-specific antibodies against RVF virus (n = 19) were tested using the serum neutralisation test (SNT). Sera from IgM-positive (n = 14) and negative (n = 20) animals were also tested by a TaqMan polymerase chain reaction (PCR). On the affected farm, 7% (16/233) of the cattle were IgM-positive and 13.7% (32/233) IgG-positive at the first bleed and 2% were IgM-positive at the second bleed, whilst the number of cattle positive for IgG-specific antibodies increased by 21.3% compared with the first bleed. Only 1.4% of sheep were positive for both IgM and IgG antibodies at the first collection; at the second bleed, IgM-positive cases decreased to 1.2%, whilst IgG-positive cases increased to 2.4%. Whilst no IgM-positive cattle were found on the neighbouring farm, 5.5% of cattle were IgG-positive. The SNT confirmed most of the ELISA results, whilst PCR results were all negative. Although serology results indicated virus circulation on both farms, the negative PCR results demonstrated that the animals were not viraemic at the time they were sampled. The movement of infected mosquito vectors by wind over long distances into a low-lying area that favoured their breeding on the Bela-Bela farm may have led to an outbreak of the disease there, but the reason for the low level of virus circulation amongst susceptible animals remains unclear.