Evidence of Transstadial and Mechanical Transmission of Lumpy Skin Disease Virus by Amblyomma hebraeum Ticks

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.     

Detection of vaccine‐like lumpy skin disease virus in cattle and Musca domestica L. flies in an outbreak of lumpy skin disease in Russia in 2017

Since 2012, lumpy skin disease virus (LSDV) has been spreading from the Middle East to south‐east Europe and Russia. Although vaccination campaigns have managed to contain LSDV outbreaks, the risk of further spread is still high. The most likely route of LSDV transmission in short distance spread is vector‐borne. Several arthropod species have been suggested as potential vectors, but no proven vector has yet been identified. To check whether promiscuous‐landing synanthropic flies such as the common housefly (Musca domestica ) could be involved, we carried out entomological trapping at the site of a recent LSDV outbreak caused by a vaccine‐like LSDV strain. The presence of vaccine‐like LSDV DNA was confirmed by the assay developed herein, the assay by Agianniotaki et al. (2017) and RPO 30 gene sequencing. No evidence of field LSDV strain circulation was revealed. In this study, we discovered that M. domestica flies carried vaccine‐like LSDV DNA (C _t  > 25.5), whereas trapped stable flies from the same collection were negative for both field and vaccine LSDV . To check whether flies were contaminated internally and externally, 50 randomly selected flies from the same collection were washed four times and tested. Viral DNA was mainly detected in the 1st wash fluid, suggesting genome or even viral contamination on the insect cadaver. In this study, internal contamination in the insect bodies without differentiation between the body locations was also revealed; however, the clinical relevance for mechanical transmission is unknown. Further work is needed to clarify a role of M. domestica in the transmission of LSDV . To our knowledge, this is the first report demonstrating that an attenuated LSD vaccine strain has been identified in Russian cattle given the ban on the use of live attenuated vaccines against LSDV.     

Epidemiological and Molecular Studies on Lumpy Skin Disease Outbreaks in Turkey during 2014–2015

This study was carried out to explore epidemiological and molecular features of lumpy skin disease virus (LSDV) in the Aegean, Central Anatolian and Mediterranean regions of Turkey, to evaluate the risk factors associated with LSDV infection and to investigate the financial impact of LSD and potential role of the Culicoides spp. in the transmission of LSDV. Samples were obtained from 611 cattle, each from different farms, and each clinically suspected to be infected with LSDV during the months of July 2014 and June 2015. Culicoides spp. were trapped from April to June 2015. Genetic characterization of the local LSDV field isolates was conducted by sequencing G‐protein‐coupled chemokine receptor gene segment. Real‐time PCR high‐resolution melting analysis was used for distinguishing each type of capripoxviruses. Viral DNA was detected in 448 of the 611 animals and Culicoides midges. Three hundred and ninety‐three of the 448 affected farms were surveyed. The morbidity and mortality rates detected were 12.3% and 6.4%, respectively. Phylogenetic analysis showed that the field isolates in this study were clustered together with other Africa and Middle East isolates. Genotyping of isolates from infected cattle has revealed the presence of LSDV. A generalized mixed linear model showed that there were positive associations between LSDV infection, European breeds, small‐sized family‐type farms and nearness of farm to a lake. The financial cost of disease presence in surveyed cattle farms was estimated to be 72.75 GBP per head. The sequence analysis of the mitochondrial cytochrome oxidase subunit I gene showed that the species of Culicoides in LSDV‐positive pools was Culicoides punctatus. Detection of LSDV in Culicoides punctatus suggests that it may have played a role in transmitting LSDV. Furthermore, movement of infected animals into disease‐free areas increases the risk of the transmission of LSD. Control strategies for LSDV infection should include consideration of the risk factors identified in this study.     

G‐Protein‐Coupled Chemokine Receptor Gene in Lumpy Skin Disease Virus Isolates from Cattle and Water Buffalo (Bubalus bubalis) in Egypt

Lumpy skin disease virus (LSDV), sheep poxvirus (SPV) and goat poxvirus (GPV) are the most serious poxviruses of ruminants. In this study, we analysed the G‐protein‐coupled chemokine receptor (GPCR) genes of LSDV isolates from cattle and water buffalo (Bubalus bubalis) in Egypt during the summer of 2011. Multiple alignments of the nucleotide sequences revealed that the water buffalo LSDV isolate differed from the cattle isolate at four nucleotide positions, and both isolates had nine nucleotide mutations from the reference strain, Egyptian tissue culture‐adapted cattle LSDV/Ismailyia88 strain. Compared with the GPCR sequences of SPV and GPV strains, a 21 nucleotide insertion and a 12 nucleotide deletion were identified in the GPCR genes of our used isolates and other LSDVs. The amino acid sequences of GPCR genes of our isolates contained the unique signature of LSDV (A₁₁, T₁₂, T₃₄, S₉₉ and P₁₉₉). Phylogenetic analyses showed that the GPCR genes of cattle and water buffalo LSDVs were closest genetically, indicating a potential transmission of cattle LSDV to water buffalo.     

Using the basic reproduction number to assess the risk of transmission of lumpy skin disease virus by biting insects

In recent years, lumpy skin disease virus (LSDV) has emerged as a major threat to cattle outside Africa, where it is endemic. Although evidence suggests that LSDV is transmitted by the bites of blood sucking arthropods, few studies have assessed the risk of transmission posed by particular vector species. Here this risk is assessed by calculating the basic reproduction number (R₀) for transmission of LSDV by five species of biting insect: the stable fly, Stomoxys calcitrans, the biting midge, Culicoides nubeculosus, and three mosquito species, Aedes aegypti, Anopheles stephensi, and Culex quinquefasciatus. Parameters relating to mechanical transmission of LSDV were estimated using new analyses of previously published data from transmission experiments, while vector life history parameters were derived from the published literature. Uncertainty and sensitivity analyses were used to compute R₀ for each species and to identify those parameters which influence its magnitude. Results suggest that S. calcitrans is likely to be the most efficient at transmitting LSDV, with Ae. aegypti also an efficient vector. By contrast, C. nubeculosus, An. stephensi, and Cx. quinquefasciatus are likely to be inefficient vectors of LSDV. However, there is considerable uncertainty associated with the estimates of R₀, reflecting uncertainty in most of the constituent parameters. Sensitivity analysis suggests that future experimental work should focus on estimating the probability of transmission from insect to bovine and on the virus inactivation rate in insects.     

Epizootic features and control measures for lumpy skin disease in south‐east Serbia in 2016

Lumpy skin disease (LSD) is an infectious disease of cattle caused by virus of the Capripoxvirus genus (LSDV), family Poxviridae. Until 2015, it had not appeared in the Balkans. In June 2016, LSD spread throughout Serbia. This study analyses the first LSD occurrence, epizootic features, applied diagnostic procedures and control measures in five districts in south‐east Serbia (Pcinja, Jablanica, Pirot, Toplica and Nisava). In total, there were 225 LSD outbreaks reported in Serbia, out of which 189 (84%) were located in the study area. The highest number of outbreaks was registered in Pcinja district (169), where LSD was first registered. The median distance and time between the nearest previous outbreak sites were calculated (4.32 km and 9 days). The median altitude of outbreak locations was 992 m with more than 90% above 500 m (p ≤ 0.001). The average herd morbidity rate in the study area was 13.6% and the herd mortality rate was recorded only in Pcinja (0.5%) and Jablanica (1.6%) districts. Samples taken from the cattle suspected to LSD were subjected to real time PCR analysis. Out of 233 samples tested for LSDV 132 (56.7%) were positive. The LSDV genome was identified in skin nodules (85.4%), blood (72.7%) and nasal swabs (62.5%). Phylogenetic analysis indicated that the LSDV strain circulating in Serbia fell within the cluster of field LSDV found worldwide. In response to the LSD epizootic, animal trade and movement were prohibited, complete stamping out, disinfection, disinsection and an entire bovine population vaccination using the homologous Neethling live attenuated vaccine (OBP, South Africa) were conducted. A month and a half after the completion of the vaccination campaign, the LSD epizootic was stopped, and no new cases have been reported since.     

Adverse Reactions to Field Vaccination Against Lumpy Skin Disease in Jordan

Lumpy skin disease (LSD) is an emerging disease in the Middle East region and has been recently reported in Jordan. The aim of this study was to investigate the adverse reactions that were reported after vaccine administration. Geographical areas enrolled in the study were free of the disease and away from the outbreak governorate. Sixty‐three dairy cattle farms, with a total of 19,539 animals, were included in the study. Of those, 56 farms reported adverse clinical signs after vaccine administration. The duration between vaccine administration and appearance of adverse clinical signs ranged from 1 to 20 days (Mean = 10.3, SD ± 3.9). Clinical signs were similar to those observed with natural cases of lumpy skin disease. These were mainly fever, decreased feed intake, decreased milk production and variable sized cutaneous nodules (a few millimetres to around 2 cm in diameter) that could be seen anywhere on the body (head, neck, trunk, perineum), udder, and/or teats. Nodules were raised and firm initially and then formed dry scabs that could be peeled off the skin. The characteristic deep ‘sit fast’ appearance was rarely seen and most lesions were superficial. Some cattle had swollen lymph nodes, while a few pregnant animals aborted. The percentage of affected cattle ranged from 0.3 to 25% (Mean = 8, SD ± 5.1). Fever, decreased feed intake, and decreased milk production were seen in 83.9, 85.7, and 94.6% in cattle on the affected farms, respectively. All affected cattle displayed skin nodules over their entire bodies, while 33.9 and 7.1% of the affected farms reported nodular lesions present on the udders and teats, respectively. No mortalities were reported due to vaccine adverse reactions. Duration (course) of clinical signs ranged from 3 to 20 days (Mean = 13.7, SD ± 4.1). Two types of LSD vaccines were used by the farmers in this study. The first one was a sheep pox virus (SPPV) vaccine derived from the RM65 isolate [Jovivac^®, manufactured by Jordan Bioindustries Centre (JOVAC)] and the other an unlabelled one, which was later identified using PCR as a strain of lumpy skin disease virus (LSDV). Blood and skin samples collected from cattle vaccinated with the LSDV vaccine were positive for LSDV using both general and species‐specific PCR primers, whereas those from cattle vaccinated with the Jovivac^® vaccine were negative. Adverse reactions observed in cattle after administration of the LSDV vaccine were reported to be more severe than those seen after Jovivac^® vaccine administration and were comparable with clinical signs observed in natural infections.     

Effect of using frozen–thawed bovine semen contaminated with lumpy skin disease virus on in vitro embryo production

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.     

Epidemiological Study of Lumpy Skin Disease Outbreaks in North‐western Iran

Lumpy skin disease (LSD) is a highly contagious transboundary disease of cattle with major economic losses. This study was undertaken to address the emergence and epidemiological features of LSD in four north‐western provinces of Iran. These provinces have extensive borders with others country including Iraq, Turkey, Azerbaijan and Armenia. A population of 683 cattle from 91 farms were examined during LSD outbreak in Iran during 2014–2016. The information of the farms including the population size, gender, age, vaccination status, clinical signs and the number of death because of LSD were recorded in the designed questionnaires. A number of 234 blood samples were collected randomly from animals with and without clinical signs of LSD. DNA was extracted from blood samples, and they were used for amplifying a fragment of 434 bp in size coupled with restriction fragment length polymorphism (RFLP) for molecular detection of lumpy skin disease virus (LSDV). The estimated prevalence, cumulative mortality and case fatality were 17.9%, 3.5% and 19.7%, respectively. There was no significant difference in occurrence of the disease between male and female cattle. LSD occurrence in age groups above 5 years old and below 6 months old showed highest and lowest relative frequencies, respectively. Vaccination was significantly decreased the occurrence of clinical disease. The developed PCR–RFLP technique was able to differentiate between LSDV, sheep pox virus (ShPV) and goat pox virus (GPV). It was concluded that LSD was entered into Iran probably from Iraq via uncontrolled animal movements along common land borders between two countries. Developed PCR–RFLP could be used as a rapid and inexpensive method for differentiating Capripoxviruses (CaPVs).     

Quantification of Lumpy Skin Disease Virus Following Experimental Infection in Cattle

Lumpy skin disease along with sheep pox and goatpox are the most serious poxvirus diseases of livestock, and are caused by viruses that belong to the genus Capripoxvirus within the subfamily Chordopoxvirinae, family Poxviridae. To facilitate the study of lumpy skin disease pathogenesis, we inoculated eight 4‐ to 6‐month‐old Holstein calves intravenously with lumpy skin disease virus (LSDV) and collected samples over a period of 42 days for analysis by virus isolation, real‐time PCR and light microscopy. Following inoculation, cattle developed fever and skin nodules, with the extent of infection varying between animals. Skin nodules remained visible until the end of the experiment on day post‐inoculation (DPI) 42. Viremia measured by real‐time PCR and virus isolation was not observed in all animals but was detectable between 6 and 15 DPI. Low levels of viral shedding were observed in oral and nasal secretions between 12 and 18 DPI. Several tissues were assessed for the presence of virus at DPI 3, 6, 9, 12, 15, 18 and 42 by virus isolation and real‐time PCR. Virus was consistently detected by real‐time PCR and virus isolation at high levels in skin nodules indicating LSDV has a tropism for skin. In contrast, relatively few lesions were observed systemically. Viral DNA was detected by real‐time PCR in skin lesions collected on DPI 42. Cattle developing anti‐capripoxvirus antibodies starting at DPI 21 was detected by serum neutralization. The disease in this study varied from mild with few secondary skin nodules to generalized infection of varying severity, and was characterized by morbidity with no mortality.     

First detection of Theileria parva in cattle from Cameroon in the absence of the main tick vector Rhipicephalus appendiculatus

A major risk factor for the spread of livestock diseases and their vectors is the uncontrolled transboundary movement of live animals for trade and grazing. Such movements constrain effective control of tick‐transmitted pathogens, including Theileria parva. Only limited studies have been undertaken to identify ticks and tick‐borne diseases (TTBDs) affecting cattle in central African countries, including Cameroon. We hereby report the collection of baseline data on the prevalence of T. parva in Cameroon through a countrywide cross‐sectional survey, conducted in 2016, involving collection of blood samples from cattle from 63 sites across the five agro‐ecological zones (AEZs) of the country. ELISA‐based surveillance of infected cattle was performed on 479 randomly selected samples and revealed specific antibodies to T. parva in 22.7% and T. mutans in 41.1% of cattle. Screening of 1,340 representative DNA samples for the presence of T. parva identified 25 (1.86%) positives using a p104 antigen gene‐based nested PCR assay. The positives were distributed across agro‐ecological zones I, II, III and V. None of the p104 positive cattle exhibited clinical symptoms of East Coast fever (ECF). Using reverse line blot (RLB), 58 (4.3%) and 1,139 (85%) of the samples reacted with the T. parva and T. mutans oligonucleotide probes, respectively. This represents the first report of T. parva from Cameroon. Surprisingly, no Rhipicephalus appendiculatus ticks, the main vector of T. parva, were identified in a parallel study involving comprehensive morphological and molecular survey of tick species present in the country. Only two of the 25 p104 positive cattle were PCR‐positive for the CD8+ T‐cell target schizont‐expressed antigen gene Tp1. Cloning and sequencing of Tp1 amplicons revealed sequence identity with the reference T. parva Muguga. This new finding raises serious concerns of a potential spread of ECF into the central African region.     

Characteristics of Foot‐and‐Mouth Disease Viral Strains Circulating at the Wildlife/livestock Interface of the Great Limpopo Transfrontier Conservation Area

Foot‐and‐mouth disease (FMD) inflicts severe economic losses within infected countries and is arguably the most important trade‐restricting livestock disease in the world. In southern Africa, infected African buffaloes (Syncerus caffer) are the major reservoir of the South African Territories (SAT) types of the virus. With the progressive expansion of transfrontier conservation areas (TFCAs), the risk of FMD outbreaks is expected to increase due to a higher probability of buffalo/livestock contacts. To investigate the dynamics of FMD within and around the Great Limpopo TFCA (GLTFCA), 5 herds of buffaloes were sampled in June 2010 to characterize circulating viruses in South Africa and Zimbabwe. Three SAT‐2 and three SAT‐3 viral strains were isolated in both countries, including one that was genetically linked with a recent SAT‐2 outbreak in Mozambique in 2011. In addition, two groups of unvaccinated cattle (n = 192) were serologically monitored for 1 year at the wildlife/livestock interface of Gonarezhou National Park (GNP) in Zimbabwe between April 2009 and January 2010, using the liquid‐phase blocking ELISA (LPBE) and a test for antibodies directed against non‐structural proteins (NSP). Neither clinical signs nor vaccination of cattle were reported during the study, yet a high proportion of the monitored cattle showed antibody responses against SAT‐3 and SAT‐1. Antibodies against NSP were also detected in 10% of the monitored cattle. The results of this study suggest that cattle grazing in areas adjacent to the GLTFCA can be infected by buffalo or other infected livestock and that cattle trade movements can act as efficient disseminators of FMD viruses to areas several hundred kilometres from the virus source. Current methods of surveillance of FMD at the GLTFCA interface seem insufficient to control for FMD emergence and dissemination and require urgent reassessment and regional coordination.     

Outbreak investigation and molecular diagnosis of Lumpy skin disease among livestock in Saudi Arabia 2016

Lumpy skin disease (LSD ) is a highly infectious disease of cattle caused by a virus of the Capripoxvirus genus in the family Poxviridae . The disease is a major concern for the dairy industry in Saudi Arabia. In this study, an outbreak of LSD in cattle herds was detected in Saudi Arabia in 2016. LSD outbreak was investigated in five regions of Saudi Arabia: Al‐Hassa, Al‐Sharqia, Al‐Qassim, Riyadh and Al‐Taif during the period from April to July 2016. Tissues from skin nodules were collected to characterize the virus by a real‐time polymerase chain reaction (rt‐PCR ). During this period, 64,109 cattle were examined and morbidity, mortality and case fatality rates were 6%, 0.99% and 16.6%, respectively. The analysis showed 3,852 infected cases and 641 deaths. highest number of infected animals was reported in Al‐Hassa (2,825), followed by Al‐Qassim (547), Riyadh (471), Al‐Sharqia (6) and Al‐Taif (3). The highest morbidity rates were observed in Al‐Qassim (6.8%), Al‐Hassa (6.2%), Riyadh (5.5%) and Al‐Taif (0.96%), while the lowest morbidity rates were recorded in Al‐Sharqia (0.27%). The highest mortality rates were also observed in Al‐Qassim (2.3%), followed by Al‐Hassa (0.97%), Riyadh (0.19%) and lowest in Al‐Sharqia and Taif (0%). LSD virus was detected in all samples (n  = 191) by real‐time PCR analysis. The disease has been observed in the cattle regardless of previous vaccination using the locally Romanian‐pox vaccine; therefore, vaccination programme and vaccine efficacy should be assessed under field conditions.     

Determination of lumpy skin disease virus in bovine meat and offal products following experimental infection

Lumpy skin disease (LSD) has recently expanded its range northwards to include the Balkans, Turkey and Russia. Because there was no solid evidence conclusively verifying the transmission mechanism in the field and LSDV viraemic animals with overt and asymptomatic presentation of disease and their products may represent a risk as an indirect transmission pathway. In this work, we used PCR positivity and infectivity in clinical and subclinical infection to evaluate the safety of meat and offal products from cows infected with the virulent LSDV strain Russia/Dagestan/2015. At day 21 post infection, seven of the 12 animals developed the generalized disease, and four animals became subclinically infected without apparent clinical signs. Upon examination and necropsy, the animals with the generalized disease had skin lesions; noticeably enlarged lymph nodes; and lesions in the lungs, trachea and testicles; whereas subclinically ill animals exhibited only enlarged lymph nodes and fever. For both disease presentations, testing of skeletal meat by PCR and virus isolation showed that the skeletal meat did not contain live virus or viral genome, whereas in cattle with generalized disease, meat with gross pathology physically connected under the site of a skin lesion was positive for the live virus. In subclinical infection, only enlarged lymph nodes carried the infectious virus, while the other internal organs tested in both types of disease manifestation were negative except for the testicles. Overall, our findings demonstrate that clinically and subclinically infected animals are reservoirs of live LSDV in lymph nodes and testicles, whereas deep skeletal meat in both types of infection do not carry live virus and the risk of transmission through this product seems very low. The detection of LSDV in testicular tissues in subclinically ill animals is concerning because of the potential to spread infection through contaminated semen. This aspect requires reconsideration of surveillance programmes to identify these Trojan horses of LSDV infection.     

Molecular detection and genetic diversity of Bartonella species in large ruminants and associated ectoparasites from the Brazilian Cerrado

Currently, five Bartonella species and an expanding number of Candidatus Bartonella species have globally been reported in ruminants. Likewise, different Bartonella genotypes were identified. However, studies relating to ruminant‐associated Bartonella in Brazil are scarce. The current study aimed to assess the prevalence and genetic diversity of Bartonella in cattle, buffaloes and associated ectoparasites in Brazil. For this purpose, EDTA‐blood samples from 75 cattle and 101 buffaloes were sampled. Additionally, 128 Rhipicephalus microplus and one Amblyomma sculptum ticks collected from cattle, and 197 R. microplus, one A. sculptum and 170 lice (Haematopinus tuberculatus) collected from buffaloes were included. Bartonella DNA was initially screened through an HRM real‐time PCR assay targeting the 16S–23S internal transcribed spacer (ITS), and the positive samples were submitted to an additional HRM assay targeting the ssrA gene. The HRM‐positive amplicons were sequenced, and the nucleotide identity was assessed by BLASTn. Bartonella spp.‐positive DNA samples were analysed by conventional PCR assays targeting the gltA and rpoB genes, and then, the samples were cloned. Finally, the phylogenetic positioning and the genetic diversity of clones were assessed. Overall, 21 of 75 (28%) cattle blood samples and 13 of 126 (10.3%) associated ticks were positive for Bartonella bovis. Out of 101 buffaloes, 95 lice and 188 tick DNA samples, one (1%) buffalo and four (4.2%) lice were positive for Bartonella spp. Conversely, none of the ticks obtained from buffaloes were positive for Bartonella. The Bartonella sequences from buffaloes showed identity ranging from 100% (ITS and gltA) to 94% (ssrA) with B. bovis. In contrast, the Bartonella DNA sequences from lice were identical (100%) to uncultured Bartonella sp. detected in cattle tail louse (Haematopinus quadripertusus) from Israel in all amplified genes. The present study demonstrates the prevalence of new B. bovis genotypes and a cattle lice‐associated Bartonella species in large ruminants and their ectoparasites from Brazil. These findings shed light on the distribution and genetic diversity of ruminant‐ and ectoparasite‐related Bartonella in Brazil.     

Assessment of interactions between African swine fever virus,bushpigs (Potamochoerus larvatus), Ornithodoros ticks and domestic pigs in north‐western Madagascar

Since its introduction in Madagascar in 1998, African swine fever (ASF) has severely affected national pig production and persists as a common disease in that country. Two of its natural hosts in the African continent, the bushpig (Potamochoerus larvatus) and tick vectors of the Ornithodoros moubata complex, are reported in west and central regions of the island. However, their role in the maintenance and transmission of the virus has been insufficiently studied. In this work, we tried to assess their potential role in the epidemiology of the disease in Madagascar, by assessing the levels of interaction between (i) ASF virus (ASFV) and bushpigs and (ii) between soft ticks and domestic and wild suids in north‐western Madagascar. Twenty‐seven sera and 35 tissue samples from bushpigs were collected and analysed for the presence of anti‐ASF antibodies and viral DNA. In addition, the sera from 27 bushpigs and 126 domestic pigs were analysed with an ELISA test for the detection of antibodies against salivary antigens from Ornithodoros ticks. No circulation of ASFV or anti‐ASFV antibodies nor anti‐tick antibodies were detected in bushpigs. However, seven of the domestic pig sera (5.6% of the total sample population) were antibody positive for O. moubata antigens. The probability of freedom from ASFV in the bushpig population using Bayesian statistical methods ranged between 73% and 84%. The probabilities of absence of anti‐tick antibodies in domestic and wild pigs were estimated at 63% and 71%, respectively. These preliminary results suggest that bushpigs are unlikely to play a significant role in the maintenance and transmission of ASFV in Madagascar. Nevertheless, further ASFV surveys are needed on that species to confirm this assumption. In addition, the presence of antibodies against O. moubata in domestic pigs suggests that soft ticks may be able to maintain ASFV within a domestic pig cycle in areas of Madagascar where they remain present.     

Detection of lumpy skin disease virus in skin lesions,blood, nasal swabs and milk following preventive vaccination

Lumpy skin disease caused by Capripoxvirus is at the moment the most important threat to European cattle industry. The only way for successful control of disease is fast and efficient diagnosis and vaccination. According to EU legislation, vaccination against LDS can be conducted only after confirmation of the disease. Croatia has a special position regarding LSD —in 2016, for the first‐time vaccination of the entire cattle population was conducted without an index case. The presence of vaccine viral particles was detected in milk, skin nodules, blood and nasal swabs in seven from total of eight herds. The presence of virus genome was detected in five cows from 10 up to 21‐day post‐vaccination. The virus was successfully isolated on cell culture from 10 up to 21‐day post‐vaccination from three animals. The obtained results support the need for further efforts to develop safer vaccines against LSDV .     

Seminal Transmission of Lumpy Skin Disease Virus in Heifers

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.     

Multilocus genotyping of Theileria parva isolates associated with a live vaccination trial in Kenya provides evidence for transmission of immunizing parasites into local tick and cattle populations

The live infection and treatment (ITM) vaccination procedure using the trivalent Muguga cocktail is increasingly being used to control East Coast fever, with potential implications for Theileria parva population genetic structure in the field. Transmission of the Kiambu V T. parva component to unvaccinated cattle has previously been described in Uganda. We monitored the T. parva carrier state in vaccinated and control animals on a farm in West Kenya where an ITM stabilate derived from the Kenyan T. parva Marikebuni stock was evaluated for field efficacy. A nested PCR‐based Marikebuni‐specific marker identified a carrier state in nine of ten vaccinated animals, detectable for a period of two years. We used 22 variable number tandem repeat (VNTR) markers to determine multilocus genotypes (MLGs) of 19 T. parva schizont‐infected lymphocyte isolates derived from cattle and field ticks. Two isolates from unimmunized cattle were identical to the Marikebuni vaccination stock. Two cattle isolates were identical to a Muguga cocktail component Kiambu V. Seven isolates from ticks exhibited MLGs that were identical to the Serengeti/Muguga vaccine stocks. Six cattle and two tick‐derived stocks exhibited unique MLGs. The data strongly suggest transmission of immunizing genotypes, from Marikebuni vaccine‐induced carrier cattle to unimmunized cattle. It is possible that genotypes similar to those in the Muguga cocktail are present in the field in Western Kenya. An alternative hypothesis is that these parasites may have originated from vaccine trial sites in Eastern Uganda. If correct, this suggests that T. parva stocks used for immunization can potentially be disseminated 125 km beyond the immediate vaccination site. Regardless of their origin, the data provide evidence that genotypes similar to those in the Muguga cocktail are circulating in the field in East Africa, alleviating concerns about dissemination of ‘alien’ T. parva germplasm through live vaccination.     

Risk of introduction of Lumpy Skin Disease into France through imports of cattle

The lumpy skin disease (LSD) virus belongs to the genus Capripoxvirus and causes a disease in cattle with economic impacts. In November 2014, the disease was first reported in Europe (in Cyprus); it was then reported in Greece (in August 2015) and has spread through different Balkan countries since 2016. Although vector transmission is predominant in at‐risk areas, long‐distance transmission usually occurs through movements of infected cattle. In order to estimate the threat for France, a quantitative import risk analysis (QIRA) model was developed to assess the risk of LSD being introduced into France by imports of cattle. Based on available information and using a stochastic model, the probability of a first outbreak of LSD in France following the import of batches of infected live cattle for breeding or fattening was estimated to be 5.4 × 10⁻⁴ (95% probability interval [PI]: 0.4 × 10⁻⁴; 28.7 × 10⁻⁴) in summer months (during high vector activity) and 1.8 × 10⁻⁴ (95% PI: 0.14 × 10⁻⁴; 15 × 10⁻⁴) in winter months. The development of a stochastic QIRA made it possible to quantify the risk of LSD being introduced into France through imports of live cattle. This tool is of prime importance because the LSD situation in the Balkans is continuously changing. Indeed, this model can be updated to process new information on the changing health situation in addition to new data from the TRAde Control and Expert System (TRACES, EU database). This model is easy to adapt to different countries and to other diseases.