Isolation and molecular characterization of lumpy skin disease virus from Tamil Nadu, India during the outbreaks from 2020 to 2022.

Lumpy skin disease (LSD) caused by LSD virus is a WOAH notifiable, high-impact, transboundary poxviral disease of bovines. The first official report of LSDV in India is from Odisha state during August 2019. Since then, cases have been reported from many states including Tamil Nadu, a Southern state of India. The present study deals with isolation and molecular characterization of LSDV from Tamil Nadu during the period August 2020 to July 2022. LSDV was isolated in embryonated chicken eggs (ECE) and BHK 21 cells and was characterized based on P32, RPO30, and GPCR genes. The phylogenetic analysis revealed that Tamil Nadu isolates from India are closely related to other Indian strains, Kenyan strains and strains from neighboring countries such as Bangladesh, Nepal, and Myanmar confirming the common exotic source for the transboundary spread across borders. The presence of unique signature of amino acid (aa) at specific positions (A11, T12, T34, S99, and P199) in the GPCR sequence confirmed the identity of LSDV. A twelve nucleotide (nt94-105) insertion and corresponding aa (TILS) at 30-33 position was found in GPCR sequence and characteristic amino acid proline at 98 position (P98) in the RPO30 gene sequence of our isolates was similar to strains from Bangladesh, Nepal, and Myanmar. Further, dissimilarity of our isolates from Neethling like vaccine strains confirms the circulation of virulent filed strains responsible for the outbreaks.

Quantitative polymerase chain reaction: another tool to evaluate viable virus content in live attenuated orf vaccine.

A probe-based real-time polymerase chain reaction (PCR) assay based on the highly conserved DNA polymerase gene of orf virus (ORFV) for the quality control of attenuated orf vaccine is reported. Primary lamb testis (PLT) cells were infected with orf vaccine virus and harvested at a critical time point to obtain maximum viable virus content as determined by real-time PCR. DNA extracted from these harvests was subjected to real-time PCR. A critical time point for the harvesting of PLT cells infected with various log(10) dilutions of vaccine virus was found to be 42 h (highest slope of 3.335), which was obtained by comparing the slopes of standard curves of different time intervals. The assay was employed to evaluate viable virus content in different batches of orf vaccine. The titres estimated by real-time PCR and conventional TCID(50) were comparable with a correlation of 0.8169. Thus, the real-time PCR assay could provide an alternative method or supplementary tool to estimate live ORFV particles in attenuated orf vaccine.

Sequence and phylogenetic analyses of an Indian isolate of orf virus from sheep.

The authors describe the isolation and identification of orf virus (ORFV) from an outbreak in a flock of sheep at Mukteswar, Uttarakhand, India, in 2009. The causative agent, ORFV was successfully isolated in primary lamb testes cells and identified using a semi-nested diagnostic polymerase chain reaction (PCR) and sequence and phylogenetic analyses of immunogenic envelope protein (B2L) coding gene. The affected animals showed characteristic proliferative skin lesions around the mouth and on nostrils and, in a few animals, lesions were also noticed on the tongue irrespective of age and sex. The morbidity, mortality and case fatality rates observed were 6%, 45% and 13%, respectively. Clinical samples were initially screened by counter immuno-electrophoresis and the serum neutralisation test; further positive skin scabs were tested with diagnostic PCR and virus isolation was performed on primary or secondary lamb testes cultures. Sequencing and phylogenetic analyses of the sheep isolate based on the B2L gene revealed that the isolate was closest to a goat isolate retrieved from an outbreak at the same geographic location in 2000. Furthermore, it also showed close genetic similarities with other Indian isolates reported earlier. Regular and systematic investigation of outbreaks is necessary to monitor the disease in susceptible populations. The development of rapid diagnostic methods as well as effective vaccine to control this infection not only from India but also other parts of the world is called for.

Emerging and re-emerging zoonotic buffalopox infection: a severe outbreak in Kolhapur (Maharashtra), India.

Buffalopox is an emerging and re-emerging zoonotic viral infection. The authors investigated an extensive zoonotic outbreak of buffalopox involving many human cases. High morbidity and significant productivity losses were recorded among domestic buffalo in Kolhapur (Maharashtra), India, between February and March 2009. The outbreak involved a total of 4000 buffalo from 21 villages in which over 10 000 buffalo were herded. The outbreak also involved 125 humans who were mostly animal handlers and milkers of all age groups. The disease inflicted a loss of approximately 40% in terms of reduced milk production and a decline in animal trade. Although pox lesions were observed on all parts of the body, the most severe were found on the inner ear. This led to otitis and pyrexia in most of the affected animals. Milkers developed pox-like lesions on the skin of their fingers, hands, forearms, forehead, ears and face, along with pyrexia, malaise and axillary lymphadenitis and lymphadenopathy. The causal agent, buffalopox virus, was confirmed using counter-immuno-electrophoresis, the serum neutralisation test, virus isolation and buffalopox virus-specific ankyrin repeat protein (C18L) gene-based polymerase chain reaction. Considering the emergence and re-emergence of buffalopox virus in buffalo, cows and humans, not only in India but also in other buffalo rearing countries, regular monitoring of outbreaks and control measures are necessary to curb economic losses and also to reduce the public health impact of the disease.

Camelpox: epidemiology, diagnosis and control measures.

Camelpox is an economically important contagious skin disease of camelids caused by camelpox virus (CMLV) and is characterized by mild local skin infection and less common severe systemic infections. The disease is confined to camel-rearing belts particularly in developing countries and causes economic impact due to considerable loss in terms of morbidity, mortality, loss of weight and reduction in milk yield. The virus has gained attention from researchers due to its recent emergence with close genetic relatedness to variola virus, the causative agent of smallpox, and carrying genes responsible for host immune evasion mechanisms. CMLV was earlier thought to be a zoonotic agent but so far little evidence has been documented from Somalia. Although the disease can be diagnosed based on clinical signs, the similar confounding skin lesions necessitate identification of infection by molecular biology based diagnostic techniques, namely restriction enzyme analysis of the virus genome and specific genes, genus- and species-specific diagnostic PCRs including real-time quantitative PCR, and sequence and phylogenetic analysis for diagnosis and differentiation of CMLV. The entire genome sequence of CMLV is known and it contains more than 211 putative genes, which code for different proteins with host range, immunomodulation, virulence and other functions. Both inactivated and live-attenuated vaccines are available in some countries. However, live vaccines are preferred as they provide long-lasting immunity. Considering the virus spreads through contaminated environments, an improved diagnostic and control method would be of immense value to curtail the infection in the field. Alternative therapeutics such as antiviral agents is an area that needs to be explored. This article discusses the epidemiology and biology of the disease, novel diagnostic approaches and control measures.