Post-Vaccination Sero-Monitoring of Peste des Petits Ruminants in Sheep and Goats in Karnataka: Progress towards PPR Eradication in India.

Peste des petits ruminants (PPR) presents economic challenges in enzootic countries impacting small ruminant productivity. The state of Karnataka, India, implemented a mass vaccination campaign in alignment with the PPR-Global Eradication Programme (GEP) and the National Strategic Plan for PPR eradication. This study was conducted from January to March 2023 to assess seroconversion in post-vaccinated goats and sheep at the epidemiological unit (epi-unit) level, aligning with the World Organisation for Animal Health (WOAH) and the Food and Agriculture Organization (FAO) guidelines in the PPR Global Control and Eradication Strategy (GCES). Before vaccination, 3466 random serum samples were collected from small ruminants of three age groups (6-12 months, 1-2 years, and >2 years) across 116 epi-units, spanning 82 taluks in 28 districts. Post-vaccination sero-monitoring included 1102 serum samples collected from small ruminants of the 6-12-month age group only, across 111 epi-units covering 64 taluks in 23 districts. The PPRV antibody status was determined using an indigenous hemagglutinin (H) protein monoclonal antibody-based competitive ELISA kit. Pre-vaccination, the PPR seropositivity rates were 55%, 62%, and 66% in the age groups of 6-12 months, 1-2 years, and >2 years, respectively, with a 61% PPRV antibody prevalence across all the age groups. Notably, 41% of the epi-units exhibited antibody prevalence rates of ≥70%, indicating a substantial population immunity, possibly attributed to the previous vaccination program in the state since 2011. In contrast, only 17% of the epi-units had below 30% seroprevalence rates, emphasizing the need for intensified vaccination. Statistical analysis of the data revealed significant correlations (p < 0.05) between the presence of PPRV antibodies and host factors such as species, breed, and sex. Post-vaccination seroprevalence in the 6-12 months age group was found to be 73.4%, indicating the use of an efficacious vaccine. On the evaluation of vaccination immunity in the 6-12 months age group, it was revealed that over 69% of the epi-units achieved a response surpassing ≥70%, indicating a significant improvement from 42% of the epi-units in pre-vaccination. For active PPR eradication, a mass vaccination campaign (>95% coverage) targeting small ruminant populations aged >4 months is advocated, aiming to achieve the desired herd immunity of >80%. This study offers crucial insights into PPR baseline seroprevalence/immunity status and vaccine efficacy, guiding national strategies towards a PPR-free India and further supporting the global eradication initiative.

Spatial and temporal analysis of haemorrhagic septicaemia outbreaks in India over three decades (1987-2016).

Haemorrhagic septicaemia (HS) is an economically important disease affecting cattle and buffaloes and the livelihoods of small-holder farmers that depend upon them. The disease is caused by Gram-negative bacterium, Pasteurella multocida, and is considered to be endemic in many states of India with more than 25,000 outbreaks in the past three decades. Currently, there is no national policy for control of HS in India. In this study, we analysed thirty year (1987-2016) monthly data on HS outbreaks using different statistical and mathematical methods to identify spatial variability and temporal patterns (seasonality, periodicity). There was zonal variation in the trend and seasonality of HS outbreaks. Overall, South zone reported maximum proportion of the outbreaks (70.2%), followed by East zone (7.2%), Central zone (6.4%), North zone (5.6%), West zone (5.5%) and North-East zone (4.9%). Annual state level analysis indicated that the reporting of HS outbreaks started at different years independently and there was no apparent transmission between the states. The results of the current study are useful for the policy makers to design national control programme on HS in India and implement state specific strategies. Further, our study and strategies could aid in implementation of similar approaches in HS endemic tropical countries around the world.

Epidemiological investigation of ASF outbreaks in Kerala (India): detection, source tracing and economic implications.

This study investigates suspected African swine fever (ASF) outbreaks in two villages of Kannur district in Kerala, India, with the aim of identifying the causative agent and its genotype, the source of infection, and estimating the economic losses due to the outbreaks. Clinically, the disease was acute with high mortality, while gross pathology was characterized by widespread haemorrhages in various organs, especially the spleen, which was dark, enlarged and had friable cut surfaces with diffuse haemorrhages. Notably, histopathological examination revealed multifocal, diffuse haemorrhages in the splenic parenchyma and lymphoid depletion accompanied by lymphoid cell necrosis. The clinico-pathological observations were suggestive of ASF, which was confirmed by PCR. The source of outbreak was identified as swill and it was a likely point source infection as revealed by epidemic curve analysis. The phylogenetic analysis of p72 gene identified the ASFV in the current outbreak as genotype-II and IGR II variant consistent with ASFVs detected in India thus far. However, the sequence analysis of the Central Variable Region (CVR) of the B602L gene showed that the ASFVs circulating in Kerala (South India) formed a separate clade along with those found in Mizoram (North East India), while ASFVs circulating in Arunachal Pradesh and Assam states of India grouped in to different clade. This study represents the first investigation of ASF outbreak in South India, establishing the genetic relatedness of the ASFV circulating in this region with that in other parts of the country. The study also underscores the utility of the CVR of the B602L gene in genetically characterizing highly similar Genotype II ASFVs to understand the spread of ASF within the country.

Elevation determines the spatial risk of Anthrax outbreaks in Karnataka, India.

Anthrax is an economically important livestock disease affecting subsistence farmers and it is of zoonotic importance. Anthrax is endemic in many states of India including Karnataka. Identification of spatial risk factors for occurrence of anthrax and development of predictive risk maps are required for planning adequate vaccination in high-risk areas as well as targeted surveillance activities in animals, humans and environment. In this study, village level anthrax outbreak locations from Karnataka (1997-2016) were geo-referenced and predictive risk map was developed using temporally Fourier Processed remotely sensed variables. A non-linear discriminant analysis approach was used to develop the risk map for Karnataka. Elevation was identified as top predictor variable in the 10 variables selected. The predicted risk map showed good accuracy and validation statistics when evaluated using different metrics (Kappa, sensitivity, specificity, AUC). The predicted risk map also showed good correspondence with past outbreaks. Further, we used Bayesian Penalised spline method to estimate species response curves for top 10 variables selected. The validated risk map can be used in planning vaccination strategy and surveillance in high-risk areas.

Countrywide cross-sectional study of swine brucellosis sero-prevalence in Indian subcontinent during 2018-2019.

Brucellosis in swine is a contagious disease with greater zoonotic potential caused by Brucella suis. The study describes PAN India swine brucellosis sero-prevalence in 5431 stratified random serum samples collected during 2018-2019 from 26 out of 29 states and two out of seven union territories. The serum samples were tested for anti-Brucella antibodies by indirect ELISA and overall, 4.33% apparent prevalence (AP) was recorded. The AP is ≥ 10% in five states among 26 states, P ≥ 50% in four districts out of 117 districts screened and cent percent prevalence in two epi units out of 264 sampled. Significantly high seropositivity (p < 0.05) in male (6.08%) than female pigs (3.46%) and in ≥ 24-month-old pigs indicated older and male pigs as potential carriers of the disease. The study recorded endemicity of the swine brucellosis in few regions of India requiring periodical surveillance for control of the disease. Brucella testing of boars before breeding and awareness among farmers and veterinarians will aid in reduction of disease burden in the absence of vaccination policy.

Seroprevalence of Infectious bovine rhinotracheitis (IBR) in India: A 5-year study.

Infectious bovine rhinotracheitis (IBR) is a highly contagious disease of bovines causing respiratory symptoms, abortions, and reduced milk yield, leading to huge economic losses. Reports on seroprevalence in bovines in India are available and restricted to districts/states. In the present study, a nationwide seroprevalence of IBR in bovines was conducted to provide a national IBR seroprevalence to the Chief Veterinarian who in turn can design the control strategies. A total of 15,592 cattle and buffalo serum samples from 25 states and 3 Union Territories viz., Jammu and Kashmir, Puducherry, and Andaman and Nicobar Islands were tested for IBR antibodies using Avidin­Biotin (AB) ELISA. Cumulative seropositivity was found to be 31.37%. Maharashtra and Rajasthan states, part of the west zone of the country, showed the highest and lowest seroprevalence, respectively. A total of 11,423 cattle and 4,169 buffalo serum samples were tested, which showed 33.91% and 24.39% seropositivity, respectively. India has the highest buffalo population. Presently, India no IBR vaccination programs are implemented in India. Considering the high seroprevalence, the authorities should plan control strategies for vaccinating dairy cows and buffaloes in India.

Prevalence of anti-leptospiral antibodies and frequency distribution of Leptospira serovars in small ruminants in enzootic South Peninsular India.

BACKGROUND AND AIM: For understanding the epidemiology of leptospirosis, the confined abundance of several species of pathogenic leptospires and knowledge on the serovar(s) prevalent in the reservoir and carrier hosts may be a useful indicator of transmission to incidental/accidental hosts in a geographical niche. The present study was carried out to ascertain the frequency distribution of Leptospira serovars and the prevalence of anti-leptospiral antibodies in small ruminants (sheep and goats) in the epidemiological units (villages) in the coastal districts of enzootic regions in South Peninsular India. MATERIALS AND METHODS: A total of 1167 serum samples (sheep n=299 and goats n=868) from apparently healthy animals, randomly collected from various epidemiological units were tested in microscopic agglutination test (MAT) using 18 reference Leptospira serovars antigens. RESULTS: The overall seroprevalence of 40% (at 95% confidence intervals [CI]: 36.82-42.43) in small ruminants (44% [95% CI: 40.49-52.26] in sheep and 38% [95% CI: 34.96-41.41] in goats) was observed with the predominance of Icterohaemorrhagiae, Javanica, Australis, Hurstbridge, and Pyrogenes serogroup anti-leptospiral antibodies in the studied region. The Chi-squared test revealed that the presence of anti-leptospiral antibodies is significantly not independent (associated) across the administrative division (Chi-square=105.80, p<0.05) as well as for sheep (Chi-square=34.67, p<0.01) and goats (Chi-square=68.78, p<0.01). Among seropositive samples (n=462 reactors), the MAT was positive for more than one serovar in 73% of sheep (95/131) and 53% of goats (177/331), representing an overall 59% cross-reactive prevalence in small ruminants. The determined frequency distribution (varied among small ruminants) of the employed serovars representing major reactive serogroup was Icterohaemorrhagiae (29.87), Javanica (20.78), Australis (20.35), Hurstbridge (16.23), Pyrogenes (15.8), Djasmin (15.58), Bataviae (15.37), Autumnalis (14.5), Canicola (14.5), Hebdomadis (14.07), Shermani (13.64), Panama (13.42), Sejroe (12.77), etc. CONCLUSION: This study indicates alarmingly high seroprevalence of leptospirosis in small ruminants with existing endemicity in the studied region in South Peninsular India. Further, these prevalent serovars in the administrative division may be of use in the reference panels of antigens in the MAT in both humans and animal disease diagnostic laboratories for effective and timely diagnosis of leptospirosis and to combat the challenges in public health.

Spatial seroprevalence of classical swine fever in India.

Classical swine fever (CSF) is a highly contagious dreadful disease of pigs leading to 100% mortality in acute form in susceptible population thereby causing huge economic loss to pig farmers. This study was undertaken to assess the seroprevalence of CSF at national level. A two-stage random sampling methodology was adopted that included 271 villages from 115 districts of India. A total of 5848 pig serum samples from twenty-five states and one Union Territory of India were collected during 2018-2019. A percent positivity of 38.52 was found at national level. Puducherry and Sikkim showed the highest and lowest percent positivity respectively. Pigs from the west zone showed the highest seroprevalence of 55.83% and those from the south zone showed the lowest of 30.25%. Adult pigs in the north and east zones showed highest percent positivity of 81.8, whereas pigs of more than 3 years of age showed highest percent positivity of 54.9, 75 and 62.5 in the north east, west and central zones respectively. Young ones showed percent positivity of 41.5 in the south zone. Higher rainfall (> 3 mm/day) and lower temperature (< 26 °C) favoured the existence of disease in the north east region combined with high density of pig population. Amidst no fool proof alert system, seroprevalence is the best method to assess the status of CSF in herd/population that provides the policymakers to plan for control of disease.

Prediction of daily and cumulative cases for COVID-19 infection based on reproductive number (R0) in Karnataka: a data-driven analytics.

To estimate the reproductive number (R0) of the coronavirus in the present scenario and to predict the incidence of daily and probable cumulative cases, by 20 August, 2020 for Karnataka state in India. The model used serial interval with a gamma distribution and applied 'early R' to estimate the R0 and 'projections' package in R program. This was performed to mimic the probable cumulative epidemic trajectories and predict future daily incidence by fitting the data to existing daily incidence and the estimated R0 by a model based on the assumption that daily incidence follows Poisson distribution. The maximum-likelihood (ML) value of R0 was 2.242 for COVID-19 outbreak, as on June 2020. The median with 95% CI of R0 values was 2.242 (1.50-3.00) estimated by bootstrap resampling method. The expected number of new cases for the next 60 days would progressively increase, and the estimated cumulative cases would reach 27,238 (26,008-28,467) at the end of 60th day in the future. But, if R0 value was doubled the estimated total number of cumulative cases would increase up to 432,411 (400,929-463,893) and if, R0 increase by 50%, the cases would increase up to 86,386 (80,910-91,861). The probable outbreak size and future daily cumulative incidence are largely dependent on the change in R0 values. Hence, it is vital to expedite the hospital provisions, medical facility enhancement work, and number of random tests for COVID-19 at a very rapid pace to prepare the state for exponential growth in next 2 months.

Animal disease surveillance: Its importance & present status in India.

Animal disease surveillance encompasses systematic collection of long-term data on disease events, risk factors and other relevant parameters followed by analyzing the same with reference to temporal and spatial characteristics to arrive at a conclusion so that necessary preventive measures can be taken. In India, the animal disease surveillance is done through National Animal Disease Reporting System, which is a web-based information technology system for disease reporting from States and Union Territories with the aim to record, monitor livestock disease situation and to initiate the preventive and curative action in a swift manner during disease emergencies. National Animal Disease Referral Expert System is a dynamic geographic information system and remote sensing-enabled expert system that captures an incidence of 13 economically important livestock diseases from all over the country and also provides livestock disease forecasting. The laboratories under State and Central governments, several research institutes under the Indian Council of Agricultural Research and veterinary colleges are involved in livestock disease diagnosis including zoonotic diseases. An integrated surveillance system is necessary for early detection of emerging/zoonotic diseases in humans. This review provides information on disease reporting and surveillance systems in animal health sector and the need for One Health approach to improve and strengthen the zoonotic disease surveillance system in India.

Development of recombinant NS1-NS3 antigen based indirect ELISA for detection of bluetongue antibodies in sheep.

Bluetongue is an insect borne (Culicoides) viral disease of small ruminants. The virus blankets the globe with a wide serotypic variation, numbered from 1 to 28. In India 21 different serotypes have been reported to be circulating across the various agro-climatic zones of the country. Non-structural proteins (NSPs) of bluetongue virus have always remained ideal target for differentiation of infected from vaccinated animals. The current study is an extrapolation of our previous work where a novel fusion construct comprising of bluetongue viral segment NS1 and NS3 was successfully cloned, expressed, purified with an efficient strategy for its suitable implementation as a diagnostic antigen. In this study, the applicability of the fusion construct has been further evaluated and optimised for field applicability. The fusion construct used in an ELISA platform projected a relative diagnostic sensitivity and specificity of 98.1% and 95.5% respectively against a pre-established test panel. The rNS1-NS3 ELISA showed substantially good agreement with the commercial BTV antibody detection kit. Finally, the study brings together the diagnostic capability of two NSPs, which can be a handy tool for sero-surveillance of bluetongue.

Spatial sero-prevalence of brucellosis in small ruminants of India: Nationwide cross-sectional study for the year 2017-2018.

Brucellosis in small ruminants caused mainly due to Brucella melitensis is an important zoonotic disease characterized by abortion, retained placenta, infertility, orchitis, epididymitis and rarely arthritis. Small ruminants are the main source of economy for the rural and marginally poor farmers and brucellosis is resulting in huge economic losses due to abortions and infertility and causing public health concern among the small ruminant keepers. Bovine brucellosis control programme has been implemented in India and small ruminants are left out of the programme mainly due to paucity of brucellosis status. The present cross-sectional study based on stratified random sampling was undertaken during 2017-18 to provide the nationwide brucellosis sero-prevalence in small ruminants. A total of 24,056 small ruminant serum samples (sheep samples = 8,103 [male-2,440 and female-5,663] and goat samples = 15,953 [male-4,331 and female-11,622]) sourced from 27 out of 29 states and two out of seven union territories (UTs), 350 districts of total 640 districts (54.68% of the Indian districts) and from 1,462 villages out of 6,40,867 villages (43.83% of the Indian villages). The serum samples were tested by indirect ELISA and overall brucellosis apparent and true prevalence of 7.45 (95% CI: 7.13-7.79) and 3.79 (95% CI: 3.44-4.17) was recorded. Significantly higher brucellosis sero-prevalence (p < .0001) was observed in sheep (11.55%) than goats (5.37%). Similarly, brucellosis seropositivity was highly significant in females compared to males in both sheep and goats. Countrywide, greater than 5% brucellosis sero-prevalence in sheep and goats was recorded in 14 and 10 states, respectively, indicating endemicity of the disease. The study provided the latest update on nationwide spatial sero-prevalence of small ruminant brucellosis which will aid government to strengthen regular surveillance and vaccination to reduce the disease burden and public health problems in the country.

VP2 Gene-Based Molecular Evolutionary Patterns of Major Circulating Bluetongue Virus Serotypes Isolated during 2014-2018 from Telangana and Andhra Pradesh States of India.

INTRODUCTION: Bluetongue disease is an economically important viral disease of livestock caused by bluetongue virus (BTV) having multiple serotypes. It belongs to the genus Orbivirus of family Reoviridae and subfamily Sedoreovirinae. The genome of BTV is 10 segmented dsRNA that codes for 7 structural and 4 nonstructural proteins, of which VP2 was reported to be serotype-specific and a major antigenic determinant. OBJECTIVE: It is important to know the circulating serotypes in a particular geographical location for effective control of the disease. The present study unravels the molecular evolution of the circulating BTV serotypes during 2014-2018 in Telangana and Andhra Pradesh states of India. METHODS: Multiple sequence alignment with available BTV serotypes in GenBank and phylogenetic analysis were performed for the partial VP2 sequences of major circulating BTV serotypes during the study period. RESULTS: The multiple sequence alignment of circulating serotypes with respective reference isolates revealed variations in antigenic VP2. The phylogenetic analysis revealed that the major circulating serotypes were grouped into eastern topotypes (BTV-1, BTV-2, BTV-4, and BTV-16) and Western topotypes (BTV-5, BTV-12, and BTV-24). CONCLUSION: Our study strengthens the need for development of an effective vaccine, which can induce the immune response for a range of serotypes within and in between topotypes.

Seroprevalence study of peste des petits ruminants in sheep and goats in the northern region of India.

BACKGROUND AND AIM: Peste des petits ruminants (PPR) is a contagious, World Organization for Animal Health notifiable, economically important, transboundary morbilliviral disease of sheep and goats. Studying seroprevalence of PPR from different geographical areas under varying agro-climatic conditions may help in formulating effective and appropriate disease control strategies under the ongoing national PPR control program. The present cross-sectional study describes the prevalence of PPR virus antibodies in sheep and goats in the various epidemiological units in different states (Haryana, Himachal Pradesh [HP], Jammu and Kashmir [J&K], Punjab, Uttarakhand [UK], and Uttar Pradesh [UP]) of the northern region of India. MATERIALS AND METHODS: A total of 5843 serum samples (sheep [n=2463] and goats [n=3380]) were collected by stratified random sampling method from 322 epidemiological units in the studied region during 2017-2018 and tested for PPR virus (PPRV) antibodies by competitive ELISA. RESULTS: The results revealed that an overall seroprevalence of 44.05% (2574/5843) with 57.32%, 55.22%, 65.69%, 37.09%, 32.73%, and 29.35% prevalence of PPRV antibodies in small ruminants in Haryana, Punjab, UP, HP, J&K, and UK states, respectively. Further, Chi-squared test revealed an association of PPRV antibodies in goats (χ2=252.28, p<0.01) and sheep (χ2=192.12, p<0.01) across different states in the region. CONCLUSION: The seroprevalence in majority of the epidemiological units (n=130) in sheep and goats in the studied region had <30%. This necessitates comprehensive, rigorous, continuous vaccination and active surveillance programs for few more years to achieve the desired 70% seroprevalence level of PPRV antibodies in population and to make the northern region of India, as PPR free zone.

Lumpy skin disease (LSD) outbreaks in cattle in Odisha state, India in August 2019: Epidemiological features and molecular studies.

Lumpy skin disease (LSD) caused by lumpy skin disease virus (LSDV) inflicts significant economic losses in cattle production with impact on livelihoods of smallholders. This study reports the first occurrence of LSD in cattle in India and analyses epidemiological and genetic characterization data from LSD outbreaks in five districts of Odisha state in August 2019. In all, 182 of 2,539 cattle were affected with an apparent morbidity rate of 7.1% and no mortality. Out of 102 samples from 60 LSD suspected and 17 asymptomatic in-contact cattle tested, 29.87% cattle were positive by capripoxvirus generic PCR and 37.66% were positive by LSDV real-time PCR. All the in-contact cattle tested were negative for LSDV. Among affected cattle, LSDV genome was detected more in scabs (79.16%) than blood (31.81%) and frozen bull semen (20.45%). Differential diagnosis by PCR was negative for pseudo-LSD, buffalopox, cowpox, pseudo-cowpox and bovine papular stomatitis. Five selected PCR and real-time PCR-positive LSDV DNA were sequenced in three genomic regions, P32 (LSDV074), F (LSDV117) and RPO30 (LSDV036). Phylogenetic analysis based on partial P32 and F gene sequences and complete RPO30 gene sequences showed that all the five Indian LSDV strains were identical and clustered with other field strains of LSDV circulating globally. However, the F and RPO30 gene sequence analyses revealed that Indian LSDV strains are genetically closer to the South African NI2490/KSGP-like strains than the strains detected in Europe, which was rather surprising. The present study established the existence of LSDV in India and involvement of LSDV field strains in the outbreaks. Additionally, we provided evidence of LSDV shedding in semen of naturally infected bulls. Further studies are required to determine the source of LSD introduction, extent of spread, modes of transmission and impact on dairy cattle production in India and effective control measures must be undertaken urgently.

Development of recombinant nucleocapsid protein-based indirect enzyme-linked immunosorbent assay for sero-survey of porcine reproductive and respiratory syndrome.

BACKGROUND AND AIM: Porcine reproductive and respiratory syndrome (PRRS) is a disease endemic in many countries and is of economic importance. India was free from PRRS until the first outbreak was reported from a North-East Indian state in 2013. Since then, disease outbreaks have been reported from North-East India and the pilot study conducted earlier showed that it is gradually spreading to the rest of India. Considering there are no locally developed population screening tests available for PRRS and imported diagnostic/screening tests are expensive, the present study was aimed at developing recombinant nucleocapsid (rN) protein-based indirect enzyme-linked immunosorbent assay (iELISA). MATERIALS AND METHODS: The rN protein of PRRS virus (PRRSV) was produced following standard cloning, expression, and purification procedures. Using this antigen, iELISA was optimized for the detection of serum antibodies to PRRSV. The sensitivity and specificity of the test were assessed by comparing it with a commercial PRRSV antibody detection kit. RESULT: A total of 745 serum samples from ten different states of India were screened using the developed iELISA. The iELISA had a relative specificity of 76.18% and sensitivity of 82.61% compared to the commercial ELISA (Priocheck PRRSV ELISA kit, Thermo Fisher Scientific, USA). CONCLUSION: The iELISA, which deployed rN protein from Indian PRRSV, was found to be suitable in the serological survey and may be a useful tool in future disease surveillance programs.

Detection of torque teno sus virus infection in Indian pigs.

BACKGROUND AND AIM: Torque teno viruses (TTVs) are circular, single-stranded DNA viruses, which infect a wide range of animals including livestock and companion animals. Swine TTVs (torque teno sus viruses [TTSuVs]) are thought to act as a primary or coinfecting pathogen in pathological conditions such as porcine dermatitis and nephropathy syndrome and post-weaning multisystemic wasting syndrome. So far, the presence of the virus has not been reported in India. Considering that TTSuVs have the potential to cross the species barrier into humans and that pork consumption is common in North-Eastern states of India, the current study aims to investigate the presence of TTSuV in the Indian pig population. MATERIALS AND METHODS: A total of 416 samples were collected during 2014-2018, from both apparently healthy pigs and also from pigs suspected of having died from classical swine fever and/or porcine reproductive and respiratory syndrome. These samples were screened for TTSuV infection by polymerase chain reaction (PCR) and DNA sequencing techniques. RESULTS: The presence of the virus was confirmed in 110 samples from 12 different states of India. Phylogenetic analysis of the nucleotide sequences obtained from the PCR products indicated the presence of viruses of both Iotatorquevirus and Kappatorquevirus genera in India. CONCLUSION: The study is the first report on the presence of TTSuVs in India and highlights the circulation of both genera of the virus in the country.

Contrasting selective patterns across the segmented genome of bluetongue virus in a global reassortment hotspot.

For segmented viruses, rapid genomic and phenotypic changes can occur through the process of reassortment, whereby co-infecting strains exchange entire segments creating novel progeny virus genotypes. However, for many viruses with segmented genomes, this process and its effect on transmission dynamics remain poorly understood. Here, we assessed the consequences of reassortment for selection on viral diversity through time using bluetongue virus (BTV), a segmented arbovirus that is the causative agent of a major disease of ruminants. We analysed ninety-two BTV genomes isolated across four decades from India, where BTV diversity, and thus opportunities for reassortment, are among the highest in the world. Our results point to frequent reassortment and segment turnover, some of which appear to be driven by selective sweeps and serial hitchhiking. Particularly, we found evidence for a recent selective sweep affecting segment 5 and its encoded NS1 protein that has allowed a single variant to essentially invade the full range of BTV genomic backgrounds and serotypes currently circulating in India. In contrast, diversifying selection was found to play an important role in maintaining genetic diversity in genes encoding outer surface proteins involved in virus interactions (VP2 and VP5, encoded by segments 2 and 6, respectively). Our results support the role of reassortment in driving rapid phenotypic change in segmented viruses and generate testable hypotheses for in vitro experiments aiming at understanding the specific mechanisms underlying differences in fitness and selection across viral genomes.

An efficient production of hybrid recombinant protein comprising non-structural proteins (NS 1 & NS 3) of bluetongue virus in prokaryotic expression system.

Strategic design and suitable purification techniques are of paramount importance in the production of recombinant proteins, if intended for use in a diagnostic assay. However, there is no single protocol that can be universally adopted for obtaining proteins in requisite quality and quantity across various platforms. In this study, we have targeted proteins of bluetongue virus (BTV), which is the causative agent of an arthropod-borne infectious disease in ruminants. Traditionally, serological diagnosis of the disease has rested upon either virus neutralization test or on an ELISA test that employed a recombinant structural (VP1, VP7) protein. Among the non-structural (NS) proteins of BTV, NS1 and NS3, are preferred candidate antigens in development of immuno-diagnostics as these provide the option for identifying recent/ongoing infection. However, the difficulty in production/purification of recombinant full length NS proteins of BTV in sufficient quantity and quality in various expression systems, due to inherent structural complexities, have restricted their wider applicability as immunodiagnostic reagents. To circumvent the difficulties associated with production/purification, we developed a novel NS1 and NS3 fusion gene (∼1302 bp) encoding for NS1 N-terminus (1M to G252 aa) and NS3 protein containing the N- and C-termini with a deletion of two hydrophobic domains along with intervening variable central domain (118A to A182 aa) of bluetongue virus 23. This construct was cloned, over-expressed and efficiently purified by single step affinity chromatography under unique denaturing/renaturing condition. The purified fusion protein was found suitable for detection of antibodies against BTV in an indirect ELISA (iELISA).

Seroprevalence of bluetongue and presence of viral antigen and type-specific neutralizing antibodies in goats in Tripura, a state at Indo-Bangladesh border of northeastern India.

Bluetongue (BT) is a notifiable multiple species transboundary viral disease of domestic and wild ruminants. Though the disease is enzootic in India, little is known of the disease burden and prevalent serotypes in Tripura, a hilly state of northeastern India sharing a vast porous border with Bangladesh. A surveillance study was conducted to understand the disease burden in goats in Tripura. Serum (n = 1240) and blood (n = 194) samples were collected during the year 2014 to 2017 from all the eight districts of Tripura. The overall prevalence of BT seroconversion was 47.58% whereas the presence of viral antigen was 20.61% at the individual level. Percent seroconversion was found more (50.47 ± 4.00, CI 41.31 to 49.47) in adult goats in comparison to the younger animals where it was 45.39 ± 2.08, CI 42.63 to 58.31. Presence of neutralizing antibodies in selected serum samples (n = 72) was investigated by serum neutralization test (SNT) against six bluetongue virus (BTV) serotypes and BTV-1 was found as most predominant (65.27%) followed by BTV-16 (26.38%), BTV-10 (20.83%), BTV-9 and 23 (13.88%), and BTV-2 (6.94%). To the best of our knowledge, this is the first study conducted in Tripura to investigate the presence of BTV antigen and type-specific neutralizing antibodies in apparently healthy goats.