Genome sequence characterization of the unique recombinant vaccine-like lumpy skin disease virus strain Kurgan/2018.

In 2018, the molecular epidemiology of lumpy skin disease in Russia was characterized by a surge in novel recombinant vaccine-like strains causing outbreaks along the southern border, spreading in an easterly direction. Currently, five distinct novel recombinant vaccine-like lineages have been described, designated as clusters 2.1 to 2.5. Based on the complete genome sequence analysis of the causative lumpy skin disease virus (Kurgan/Russia/2018), obtained from an eponymous outbreak, the genome was shown to be composed of a Neethling vaccine strain virus as the dominant parental strain and KSGPO vaccine virus as its minor parental strain. These features are similar to those of Saratov/Russia/2017 and Tyumen/Russia/2018, representing clusters 2.1 and 2.4, respectively. However, Kurgan/Russia/2018 has 16 statistically significant recombination events unique to this sequence, contributing to the phylogenetic clustering of Kurgan/Russia/2018 in yet another cluster designed cluster 2.6, based on analysis involving the complete genome sequences.

Lumpy skin disease: Insights into current status and geographical expansion of a transboundary viral disease.

Lumpy skin disease (LSD) is an emerging transboundary viral disease of livestock animals which was first reported in 1929 in Zambia. Although LSD is a neglected disease of economic importance, it extends a direct impact on the international trade and economy in livestock-dependent countries. Lumpy skin disease virus (LSDV) has been endemic in African countries, where several outbreaks have been reported previously. However, the virus has spread rapidly across the Middle East in the past two decades, reaching Russia and, recently, the Asian subcontinent. With unprecedented cluster outbreaks being reported across Asian countries like India, China, Nepal, Bangladesh, and Pakistan, LSDV is certainly undergoing an epidemiological shift and expanding its geographical footprint worldwide. Due to high mortality among livestock animals, the recent LSD outbreaks have gained attention from global regulatory authorities and raised serious concerns among epidemiologists and veterinary researchers. Despite networked global surveillance of the disease, recurrent LSD cases pose a threat to the livestock industry. Hence, this review provides recent insights into the LSDV biology by augmenting the latest literature associated with its pathogenesis, transmission, current intervention strategies, and economic implications. The review critically examines the changing epidemiological footprint of LSDV globally, especially in relation to developing countries of the Asian subcontinent. We also speculate the possible reasons contributing to the ongoing LSD outbreaks, including illegal animal trade, climate change, genetic recombination events between wild-type and vaccine strains, reversion of vaccine strains to virulent phenotype, and deficiencies in active monitoring during the COVID-19 pandemic.

Identifying the patterns and sizes of the first lumpy skin disease outbreak clusters in Northern Thailand with a high degree of dairy farm aggregation using spatio-temporal models.

Lumpy skin disease (LSD) is one of the most important notifiable transboundary diseases affecting cattle in many parts of the world. In Thailand, LSD outbreaks in cattle farming areas have been reported in 69 out of 77 provinces, indicating a serious nationwide situation. Understanding the dynamics of spatial and temporal LSD epidemic patterns can provide important information on disease transmission and control. This study aims to identify spatial and temporal clusters in the first LSD outbreaks in dairy farming areas with a high degree of aggregation in Northern Thailand using spatio-temporal models. The data were obtained from an official LSD outbreak investigation conducted between June and August 2021 on dairy farms (n = 202). The outbreak of LSD was confirmed by employing clinical observations and laboratory analysis. The spatio-temporal models including space-time permutation (STP), Poisson, and Bernoulli were applied to the outbreak data with the settings of 10%, 25%, and 50%, respectively, for the maximum reported cluster size (MRCS). Overall, the number of most likely and secondary clusters varied depending on the model and MRCS settings. All MRCS settings in the STP model detected the most likely clusters in the same area and the Poisson models in different areas, with the largest being defined by a 50% MRCS. Although the sizes of the most likely clusters identified by the Bernoulli models were different, they all had the same cluster period. Based on the sizes of the detected clusters, strict LSD insect-vector control should be undertaken within one kilometer of the outbreak farm in areas where no LSD vaccination has been administered. This study determines the sizes and patterns of LSD outbreak clusters in the dairy farming area with a high degree of farm aggregation. The spatio-temporal study models used in this study, along with multiple adjusted MRCS, provide critical epidemiological information. These models also expand the options for assisting livestock authorities in facilitating effective LSD prevention and control programs. By prioritizing areas for resource allocation, these models can help improve the efficiency of such programs.

Estimating the Transmission Kernel for Lumpy Skin Disease Virus from Data on Outbreaks in Thailand in 2021.

Nationwide outbreaks of lumpy skin disease (LSD) were observed in Thailand in 2021. A better understanding of its disease transmission is crucial. This study utilized a kernel-based approach to characterize the transmission of LSD between cattle herds. Outbreak data from the Khon Kaen and Lamphun provinces in Thailand were used to estimate transmission kernels for each province. The results showed that the majority of herd-to-herd transmission occurs over short distances. For Khon Kaen, the median transmission distance from the donor herd was estimated to be between 0.3 and 0.8 km, while for Lamphun, it ranged from 0.2 to 0.6 km. The results imply the critical role that insects may play as vectors in the transmission of LSD within the two study areas. This is the first study to estimate transmission kernels from data on LSD outbreaks in Thailand. The findings from this study offer valuable insights into the spatial transmission of this disease, which will be useful in developing prevention and control strategies.

Global Burden of Lumpy Skin Disease, Outbreaks, and Future Challenges.

Lumpy skin disease (LSD), a current global concern, causes economic devastation in livestock industries, with cattle and water buffalo reported to have higher morbidity and lower mortality rates. LSD is caused by lumpy skin disease virus (LSDV), a member of the Poxviridae family. It is an enzootic, rapidly explorative and sometimes fatal infection, characterized by multiple raised nodules on the skin of infected animals. It was first reported in Zambia in 1929 and is considered endemic in Africa south of the Sahara desert. It has gradually spread beyond Africa into the Middle East, with periodic occurrences in Asian and East European countries. Recently, it has been spreading in most Asian countries including far East Asia and threatens incursion to LSD-free countries. Rapid and accurate diagnostic capabilities, virus identification, vaccine development, vector control, regional and international collaborations and effective biosecurity policies are important for the control, prevention, and eradication of LSD infections. This review critically evaluates the global burden of LSD, the chronological historical outbreaks of LSD, and future directions for collaborative global actions.

A novel HRM-based gap-qRT-PCR for identification and quantitation of the vaccine and field strain(s) of lumpy skin disease virus.

Lumpy skin disease (LSD) has become the most important animal health problem in India due to high morbidity, mortality and production losses caused by it. A homologous live-attenuated LSD vaccine (Lumpi-ProVacInd) was recently developed by using a local LSD virus (LSDV) strain (LSDV/2019/India/Ranchi) in India which is likely to replace the existing practice of vaccinating cattle with goatpox vaccine. It is essential to differentiate the vaccine and field strains, if a live-attenuated vaccine has been used for control and eradication of the disease. As compared to the prevailing vaccine and field/virulent strains, the Indian vaccine strain (Lumpi-ProVacInd) has a unique deletion of 801 nucleotides in its inverted terminal repeat (ITR) region. We exploited this unique feature and developed a novel high resolution melting-based gap quantitative real-time PCR (HRM-gap-qRT-PCR) for rapid identification and quantitation of the vaccine and field strain(s) of LSDV.

Long-term monitoring of immune response to recombinant lumpy skin disease virus in dairy cattle from small-household farms in western Thailand.

Lumpy skin disease (LSD) was firstly reported in Thailand in 2021 which affected the cattle industry. However, there is limited information on the immune response of LSDV infection in Thailand where recombinant vaccine strain circulated. The aim of this research was to study the duration of LSD immune response of subclinical and clinical animals after natural infection in dairy cattle. Sixty-six dairy cattle from ten farms in central and western regions of Thailand were investigated. Antibody was detected by virus neutralization test and ELISA. Cell mediated immunity (CMI)-related cytokine gene expressions were evaluated. Antibody was detected until at least 15 months after the noticeable symptom. Cattle with subclinical disease had lower antibody levels compared to animals which had clinical disease. IFN-γ and TNF-α levels were increased, while IL-10 level was decreased in the infected animals compared to the controls. This study elucidated immune responses in dairy cattle herd affected by recombinant LSDV.

Lumpy skin disease as an emerging infectious disease.

Lumpy skin disease (LSD) is one of the most important emerging transboundary diseases. Recently, LSD has emerged in many countries in the northern hemisphere. The LSD virus has a huge genome and is highly resistant to environmental conditions. The virus is also host-specific and large ruminants, such as cattle and domestic water buffalo, are particularly susceptible. In addition, wild ruminants can serve as potential reservoirs for spreading the LSD virus. The emergence might be related to climate change in various regions because LSD is an arthropod-borne infectious disease. This disease causes enormous economic losses, such as leather damage, decreased milk production, abortion, and death in infected ruminants. The economic importance of LSD in the bovine industry has forced countries to develop and implement control strategies against the disease. With the recent global spread and the economic impact, LSD will be discussed intensively. In addition, effective preventive measures are suggested based on the presence or absence of LSD outbreaks.

Development of a dual vaccine against East Coast fever and lumpy skin disease.

East Coast fever is an acute bovine disease caused by the apicomplexan parasite Theileria parva and is regarded as one of the most important tick-vectored diseases in Africa. The current vaccination procedure has many drawbacks, as it involves the use of live T. parva sporozoites. As a novel vaccination strategy, we have constructed the recombinant lumpy skin disease virus (LSDV) named LSDV-SODis-p67HA-BLV-Gag, encoding a modified form of the T. parva p67 surface antigen (p67HA), as well as the bovine leukemia virus (BLV) gag gene for the formation of virus-like particles (VLPs) to potentially enhance p67 immunogenicity. In place of the native sequence, the chimeric p67HA antigen has the human tissue plasminogen activator signal sequence and the influenza hemagglutinin A2 transmembrane domain and cytoplasmic tail. p67HA was detected on the surface of infected cells, and VLPs comprising BLV Gag and p67HA were produced. We also show that higher multiple bands observed in western blot analysis are due to glycosylation of p67. The two vaccines, pMExT-p67HA (DNA) and LSDV-SODis-p67HA-BLV-Gag, were tested for immunogenicity in mice. p67-binding antibodies were produced by vaccinated animals, with higher titers detected in mice vaccinated with the recombinant LSDV. This candidate dual vaccine warrants further testing in cattle.

Development and Validation of a New DIVA Real-Time PCR Allowing to Differentiate Wild-Type Lumpy Skin Disease Virus Strains, Including the Asian Recombinant Strains, from Neethling-Based Vaccine Strains.

The current epidemic in Asia, driven by LSDV recombinants, poses difficulties to existing DIVA PCR tests, as these do not differentiate between homologous vaccine strains and the recombinant strains. We, therefore, developed and validated a new duplex real-time PCR capable of differentiating Neethling-based vaccine strains from classical and recombinant wild-type strains that are currently circulating in Asia. The DIVA potential of this new assay, seen in the in silico evaluation, was confirmed on samples from LSDV infected and vaccinated animals and on isolates of LSDV recombinants (n = 12), vaccine (n = 5), and classic wild-type strains (n = 6). No cross-reactivity or a-specificity with other capripox viruses was observed under field conditions in non-capripox viral stocks and negative animals. The high analytical sensitivity is translated into a high diagnostic specificity as more than 70 samples were all correctly detected with Ct values very similar to those of a published first-line pan capripox real-time PCR. Finally, the low inter- and intra-run variability observed shows that the new DIVA PCR is very robust which facilitates its implementation in the lab. All validation parameters that are mentioned above indicate the potential of the newly developed test as a promising diagnostic tool which could help to control the current LSDV epidemic in Asia.

Evaluation of the safety, immunogenicity and efficacy of a new live-attenuated lumpy skin disease vaccine in India.

Lumpy skin disease (LSD) was reported for the first time in India in 2019 and since then, it has become endemic. Since a homologous (LSD-virus based) vaccine was not available in the country, goatpox virus (GPV)-based heterologous vaccine was authorized for mass immunization to induce protection against LSD in cattle. This study describes the evaluation of safety, immunogenicity and efficacy of a new live-attenuated LSD vaccine developed by using an Indian field strain, isolated in 2019 from cattle. The virus was attenuated by continuous passage (P = 50) in Vero cells. The vaccine (50th LSDV passage in Vero cells, named as Lumpi-ProVacInd) did not induce any local or systemic reaction upon its experimental inoculation in calves (n = 10). At day 30 post-vaccination (pv), the vaccinated animals were shown to develop antibody- and cell-mediated immune responses and exhibited complete protection upon virulent LSDV challenge. A minimum Neethling response (0.018% animals; 5 out of 26,940 animals) of the vaccine was observed in the field trials conducted in 26,940 animals. There was no significant reduction in the milk yield in lactating animals (n = 10108), besides there was no abortion or any other reproductive disorder in the pregnant animals (n = 2889). Sero-conversion was observed in 85.18% animals in the field by day 30 pv.

An Extensive Examination of the Warning Signs, Symptoms, Diagnosis, Available Therapies, and Prognosis for Lumpy Skin Disease.

The lumpy skin disease virus (LSDV) infects cattle and buffalo and causes lumpy skin disease (LSD). It affects the lymph nodes of the sick animals, causing them to enlarge and appear as lumps (cutaneous nodules) that are 2-5 cm in diameter on their heads, necks, limbs, udders, genitalia, and perinea. A high temperature, a sharp drop in milk supply, discharge from the eyes and nose, salivation, a loss of appetite, depression, damaged hides, and emaciation are further warning signs and symptoms. As per the Food and Agriculture Organization (FAO), the incubation period, or the time between an infection and symptoms, is approximately 28 days. Infected animals can transfer the virus by direct contact with the vectors, direct virus secretion from mouth or nose, shared feeding and watering troughs, and even artificial insemination. The World Organization for Animal Health (WOAH) and the FAO both warn that the spread of illnesses could lead to serious economic losses. This illness reduces cow's milk production because oral ulcers make the animal weak and lead them to lose their appetite. There are many diagnostics available for LSDV. However, very few tests yield accurate findings. The best methods for preventing and controlling the lumpy skin condition include vaccination and movement restrictions. As a specific cure is not available, the only available treatment for this illness is supportive care for cattle. Recently, India has developed a homologous, live-attenuated vaccine, Lumpi-ProVacInd, which is specifically intended to protect animals against the LSD virus. This study's primary goal is to accumulate data on symptoms, the most accurate method of diagnosis, treatments, and controls to stop infections from spreading as well as to explore future possibilities for the management of LSDV.

Biochemical Analysis of the Blood of Cattle after Vaccination with a Heterologous Vaccine for Lumpy Skin Disease.

We studied the biochemical parameters of the blood of cattle after immunization against lumpy skin disease with a vaccine made from an attenuated heterologous goat pox virus (strain G20-LKV). Blood samples were obtained from animals on days 7, 14, and 21 after vaccination. The vaccine did not affect biochemical parameters of the blood. A slight increase in total protein and AST on day 14 indicates the expected reactions to the stimulation of the immune system after vaccination. The levels of direct and total bilirubin, ALT, urea, glucose, cholesterol, and creatinine in vaccinated animals remained within the physiological limits.

First complete genome sequence of lumpy skin disease virus directly from a clinical sample in South India.

Lumpy skin disease (LSD), a notifiable disease listed by the World Organization for Animal Health and a fast fast-moving transboundary viral disease infecting cattle and buffaloes, was reported in India in 2019 and has since rapidly spread across the country. This study reports the first complete genome sequence and analysis of a pathogenic LSD virus (LSDV) from India (LSDV/208/PVNRTVU/2020) obtained by direct sequencing of a suspected clinical sample using Illumina and Nanopore sequencing technologies. The complete genome sequence of LSDV/208/PVNRTVU/2020 is 150445 bp long, codes for 156 putative genes and carries identical 2254 bp inverted terminal repeats at either ends. The unique features reported in the LSDV isolates from the recent outbreaks in Asia, namely, the insertions of 12 nucleotides in the viral G-protein coupled receptor (GPCR) and 27 nucleotides leading to duplication of 9 aminoacids in the extracellular enveloped virus-specific (EEV) genes were also conserved in LSDV/208/PVNRTVU/2020. Phylogenetic analysis of the complete genome sequence of LSDV/208/PVNRTVU/2020 revealed its close relation with Kenyan strains and clustered away from vaccine strains. Further analysis showed evidence of strong purifying selection without any recombination events. The data presented in this study could be useful for designing effective strategies such as developing rapid diagnostics and vaccines to control LSD.

Emergency vaccination of cattle against lumpy skin disease: Evaluation of safety, efficacy, and potency of MEVAC® LSD vaccine containing Neethling strain.

Lumpy skin disease (LSD) is an emerging disease of cattle causing significantly high economic losses. Control of LSD depends on the use of homologous attenuated LSD virus strains isolated originally from South Africa (the Neethling strain). The virus belongs to the genus Capripoxvirus, which includes sheep pox virus and goat pox virus. The present study was conducted to evaluate the safety and efficacy of a new live attenuated LSD vaccine produced by Middle East for Vaccines (MEVAC®) based on the Neethling strain. Tests were performed both in Egypt and Vietnam. Safety was evaluated by inoculation of five cattle with 10 times the recommended dose and observation of the animals for 14 days. Immunogenicity was tested at different periods post-vaccination (PV) in animals receiving the recommended doses of the vaccine using ELISA and virus neutralization test. Five cows were used to determine the protection index (PI) and non-vaccinated control cattle were included. Three calves were challenged by intradermal inoculation of the wild virus (5 × 105 TCID50) 28 days PV. Field or mass vaccination experiments were conducted in Vietnam during national campaigns in the summer of 2021 with 4301 vaccinated animals closely monitored after vaccination. In the field, around 2% (80/4301) of the animals showed hyper-reactivity, and 0.6% (24/4301) showed small skin swellings that disappeared within few hours PV. Abortion was recorded in three animals (0.3% 3/867). Challenged animals were resistant to clinical disease and PI value was 3.5 log10. Meanwhile, antibody levels determined by the ELISA were inconsistent among animals and laboratories during the study period. Overall, the findings point to a new safe and effective LSD vaccine.

LSDV126 gene based molecular assays for specific detection and characterization of emerging Lumpy Skin Disease virus.

Lumpy skin disease (LSD) is a highly infectious and economically important viral disease, which is currently emerging in the Indian subcontinent. LSD is caused by Lumpy Skin Disease Virus (LSDV) under the genus Capripoxvirus and the family Poxviridae. Since its first incursion in India in the year 2019, the virus is rapidly disseminating through different means like direct contact, fomites and mainly by blood-feeding insects. As the disease has never been reported from India or neighbouring countries, there is a lack of planning and preparatory measures in terms of diagnostics and vaccines to control the disease. In the absence of any homologous vaccine, a live attenuated heterologous goat pox vaccine (Uttarkashi strain) is now being widely used in the country for the prevention of LSDV infection. Use of live attenuated goat pox virus vaccine necessitates the availability of an assay which could specifically detect and differentiate LSDV from goat pox virus. In this study, nucleotide sequences of LSDV126 gene encoding extracellular enveloped virus protein of circulating LSDV and goat pox virus were determined and analyzed. Deletion of 27 nt tandem repeats was observed in LSDV in comparison to goat pox and LSDV vaccine viruses. The deletion region was targeted for designing primers specific to LSDV, but not goat pox virus. A novel isothermal polymerase spiral reaction (PSR) was optimized as pen side diagnostic for prompt and sensitive detection of genomic DNA of LSDV. The assay was found to be highly sensitive and specific when compared to the real-time PCR. The assay was found to be specifically detecting only LSDV but not the goat pox virus. The limit of detection was identified as 9 × 10-6 ng of positive DNA. The assay will provide a point of care tool that will be a boon for the successful control of LSD in India.

The immune response to lumpy skin disease virus in cattle is influenced by inoculation route.

Lumpy skin disease virus (LSDV) causes severe disease in cattle and water buffalo and is transmitted by hematophagous arthropod vectors. Detailed information of the adaptive and innate immune response to LSDV is limited, hampering the development of tools to control the disease. This study provides an in-depth analysis of the immune responses of calves experimentally inoculated with LSDV via either needle-inoculation or arthropod-inoculation using virus-positive Stomoxys calcitrans and Aedes aegypti vectors. Seven out of seventeen needle-inoculated calves (41%) developed clinical disease characterised by multifocal necrotic cutaneous nodules. In comparison 8/10 (80%) of the arthropod-inoculated calves developed clinical disease. A variable LSDV-specific IFN-γ immune response was detected in the needle-inoculated calves from 5 days post inoculation (dpi) onwards, with no difference between clinical calves (developed cutaneous lesions) and nonclinical calves (did not develop cutaneous lesions). In contrast a robust and uniform cell-mediated immune response was detected in all eight clinical arthropod-inoculated calves, with little response detected in the two nonclinical arthropod-inoculated calves. Neutralising antibodies against LSDV were detected in all inoculated cattle from 5-7 dpi. Comparison of the production of anti-LSDV IgM and IgG antibodies revealed no difference between clinical and nonclinical needle-inoculated calves, however a strong IgM response was evident in the nonclinical arthropod-inoculated calves but absent in the clinical arthropod-inoculated calves. This suggests that early IgM production is a correlate of protection in LSD. This study presents the first evidence of differences in the immune response between clinical and nonclinical cattle and highlights the importance of using a relevant transmission model when studying LSD.

A candidate multi-epitope vaccine against Lumpy skin disease.

Lumpy skin disease (LSD) is a fulminant infectious disease that mostly affects cattle and causes considerable economic loss throughout the globe. This study was conducted to develop a new multi-epitope-based vaccine against LSD that can elicit immunological responses using an in silico reverse vaccinology approach. Initially, three antigenic proteins, protein E5, E3 ubiquitin-protein ligase LAP and 62 kDa protein, were manipulated to recognize potential T-cell and B-cell epitopes. To identify superior epitopes, a variety of bioinformatic techniques including antigenicity testing, transmembrane topology screening, allergenicity assessment, conservancy analysis, and toxicity evaluation were used. Finally, three new subunit vaccines (construct V1, V2 and V3) were developed employing the most effective epitopes, suitable adjuvants, pan HLA DR-binding epitope (PADRE) and linkers. Then, based on the antigenicity, solubility, and validation score of the 3D structures, construct V2 was chosen as one of the best candidate vaccines. The results of the molecular dynamic simulation and disulphide engineering indicated that the vaccine (construct V2) was stable. Additionally, the immunological simulation findings supported the vaccine candidate's ability to trigger humoral and cellular immune responses. Further validation of the proposed vaccine candidate may necessitate additional in vitro and in vivo investigations.

[Determination of the optimal immunizing dose of heterologous goat pox virus vaccine (Poxviridae: Chordopoxvirinae: Capripoxvirus) against lumpy skin disease].

INTRODUCTION: Lumpy skin disease (LSD), sheep pox and goat pox are dangerous diseases of domestic ruminants. Representatives of the genus of capripoxviruses are antigenically similar and can be used as a vaccine for three infections, as in the case of representatives of the genus of orthopoxviruses, which includes viruses of smallpox, monkeypox, and cowpox, that all belong to a single family Poxviridae. MATERIALS AND METHODS: In this study, the vaccine strain G20-LKV of the goat pox virus and the virulent strain RIBSP2019/K of the LSD virus were used. The experiments were carried out on clinically healthy cattle of the Kazakh White-headed breed, aged six to eight months. Virological and serological research methods were used in the work. RESULTS: All immunized animals that received different doses of the vaccine showed resistance to the infection challenge, without showing any clinical signs of the disease. In animals that received the lowest doses of the vaccine 15,000, 30,000 and 40,000 TCID50, no adverse events, skin and temperature reactions were observed at the injection site. Those vaccinated with high doses of the vaccine had a local reaction in the form of swelling at the site of vaccine administration. Control animals infected with a virulent virus showed clinical signs of the cattle lumpy skin disease . CONCLUSION: The vaccine, prepared based on the "G20-LKV" strain of the goat virus, is protective for cattle against infection with a virulent LSD virus at immunizing doses from 15,000 to 80,000 TCID50, which are dependent on the LSD epizootic situation in particular region.