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Foot-and-mouth disease (FMD) is a highly infectious disease of transboundary importance. Routine biannual vaccination along with surveillance activities is seen as the principal to control FMD in India. Non-structural protein (NSP) based immunoassays are the test of choice for the differentiation between infected and vaccinated population. In this study, 3D protein of FMD virus was expressed in Escherichia coli and an indirect ELISA (I-ELISA) was developed to detect 3D-antibodies in the infected bovines. 3D I-ELISA demonstrated comparable diagnostic sensitivity (97.6%) but lower specificity (80.8%) as compared to the in-house r3AB3 I-ELISA. However, the specificity values varied significantly for naïve and vaccinated samples and were observed to be 98.42% and 76.93%, respectively. A moderate degree of concordance (88.5%) was observed between the overall results of two ELISAs. 3D I-ELISA displayed a considerably lower specificity in uninfected vaccinated samples, thereby suggesting against its application for serosurveillance in intensively vaccinated population. However by virtue of its high diagnostic sensitivity and longer duration of persistence of 3D-antibody post-infection, 3D I-ELISA could be adopted for seroepidemiological investigations in regions not practicing vaccination and could be extended to susceptible species which are generally not covered by vaccination., (Copyright © 2014 The International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.)

Foot-and-mouth disease (FMD) is a highly contagious viral disease of transboundary importance. In India, since the launch of the FMD control programme, there has been a substantial increase in the vaccinated bovine population. In this scenario, there is a need for additional locally developed non-structural protein (NSP)-based immnoassays for efficient identification of FMD virus (FMDV)-infected animals in the vaccinated population. The 2B NSP of FMDV, lacking the transmembrane domain (Δ2B), was expressed successfully in a prokaryotic system, and an indirect ELISA (I-ELISA) was developed and validated in this study. The diagnostic sensitivity and specificity of the Δ2B I-ELISA were found to be 95.3 % and 94.6 %, respectively. In experimentally infected cattle, the assay could consistently detect Δ2B-NSP-specific antibodies from 10 to approximately 400 days postinfection. The assay was further validated with bovine serum samples collected randomly from different parts of the country. The performance of the Δ2B I-ELISA was compared with the in-house r3AB3 I-ELISA, and the overall concordance in test results was found to be 86.49 %. The Δ2B I-ELISA could be useful as a screening or confirmatory assay in the surveillance of FMD irrespective of vaccination.

In India, the proportion of bovines vaccinated against foot-and-mouth disease (FMD) is increasing since the implementation of the Government supported 'FMD Control Programme', and non-structural protein (NSP)-based serological assays for discriminating between antibodies induced by infection or vaccination (DIVA) could be useful. The FMD virus NSP 3AB was expressed in a prokaryotic system and an indirect ELISA (r3AB(3) I-ELISA) was developed and validated as a screening assay for detecting virus in vaccinated bovines. The diagnostic sensitivity of the assay was estimated to be 96%, while the diagnostic specificity varied between the naïve and vaccinates as 99.1% and 96.4%, respectively. This assay could detect antibodies to 3AB (3AB-Ab) from 10 to as late as 900 days post-infection in cattle infected experimentally. The "in-house" assay demonstrated higher sensitivity than a commercial 3ABC ELISA kit particularly with samples obtained from the late stages of infection. Transient post-vaccinal 3AB-Ab response could be detected in one of the three commercial vaccines during the six-month vaccination regimen, which emphasizes the fact that for a DIVA-compatible diagnostic strategy to be a realistic option, all vaccines need to be quality checked for the NSP content., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The aim of this study was to develop a competitive inhibition ELISA (CI-ELISA) for detection of antibodies to bovine viral diarrhea virus (BVDV) using the helicase domain of NS3 (non-structural) protein and monoclonal antibody (MAb) against it and to estimate its sensitivity and specificity using two commercial ELISA kits as independent references. The 45-kDa helicase domain of NS3 protein of BVDV was expressed in Escherichia coli and 18MAbs were developed against it. MAb-11G8 was selected for use in CI-ELISA on the basis of maximum inhibition (90%) obtained with BVDV type 1 infected calf serum. Based on the distribution of percent inhibition of known negative sera (n=166), a cut-off value was set at 40% inhibition. In testing 914 field serum samples of cattle (810) and buffaloes (104), the CI-ELISA showed a relative specificity of 95.75% and 97.38% and sensitivity of 96% and 94.43% with Ingenesa kit and Institut Pourquier kit, respectively. This study proved that the use of helicase domain of NS3 (45-kDa) is equally good as the whole NS3 protein (80-kDa) used in commercial kits for detection of BVDV antibodies in cattle and buffaloes.

The aim of this study was to develop a more specific and sensitive competitive inhibition ELISA (CI-ELISA) than the currently used indirect ELISA for detection of antibodies to bovine immunodeficiency virus (BIV) in cattle and buffaloes. Murine monoclonal antibodies (MAbs) were generated against a recombinant capsid (CA) protein of bovine immunodeficiency virus. Of the 13 anti-CA MAbs developed, MAb-9G10 was selected for CI-ELISA based on the maximum inhibition (98%) obtained with reference BIV antibody positive serum. Based on the distribution of percent inhibition of known negative sera (n=50), a cut-off value was set at 40% inhibition. The MAb-based CI-ELISA showed much higher agreement (concordance: 95.4%) than the indirect ELISA (concordance: 77.8%) with Western blot. Out of 672 sera of cattle and buffaloes tested by CI-ELISA from four states of India, 22% (113/516) of cattle and 19% (30/156) of buffalo were sero-positive for BIV with an overall seroprevalence of 21% (143/672) in India.

The present paper describes the investigation of foot-and-mouth disease (FMD) outbreak in a private pig farm at Kotty in Kollam district of Kerala during October 2013. During the clinical phase, severe vesicular lesions on snout and skin around the coronary bands were observed in pigs. A total of 48 serum samples and 12 clinical samples (ruptured snout epithelia) were collected. All serum samples were subjected to indirect 3AB nonstructural protein (NSP) ELISA and liquid phase blocking (LPB) ELISA. In 3AB NSP ELISA, all serum samples were found positive for NSP antibodies indicating infection. In LPB ELISA, 42 of 48 (87.5%) pigs were found to have protective log10 antibody titre of ≥1.8 against FMD virus serotypes O, A and Asia 1. All the clinical materials were found positive for serotype O in antigen detection ELISA as well as in multiplex reverse transcription-polymerase chain reaction (mRT-PCR). In VP1 region-based phylogenetic analysis, the serotype O isolates causing the outbreak were found to conglomerate within Ind2001 lineage. Pigs infected with FMD may pose rigorous threat to other susceptible domestic livestock as they exhale enormous quantity of virus. As a consequence, they should be included under prophylactic vaccination and surveillance programmes ongoing in the country. [ABSTRACT FROM AUTHOR]

Foot-and-mouth disease (FMD) is a highly contagious and globally significant viral disease principally of cloven-hoofed livestock species. The present study describes the results of molecular epidemiologic investigation of FMD in pigs across various states of India between 2008 and 2014. During this period, a total of 37 clinical epithelial samples (vesicle/foot/snout epithelium) of FMD-suspected pigs were tested in a serotype differentiating antigen detection ELISA and samples found negative were further subjected to multiplex reverse transcription-polymerase chain reaction (mRT-PCR). A total of 29 (78.37%) samples were found positive for serotype O in antigen detection ELISA and 8 ELISA-negative samples were subsequently found positive for serotype O in mRT-PCR. The VP1 region-based phylogenetic analysis demonstrated the involvement of O/ME-SA/Ind2001 lineage in the outbreaks. The pig isolates clustered with the contemporary virus isolates collected from bovine indicating a close genetic relationship and therefore signifying inter-species transmission during the outbreaks. [ABSTRACT FROM AUTHOR]

The yak and mithun husbandry in India is confronted with several challenges including the prevalence of foot-and-mouth disease (FMD). The present study was initiated to investigate FMD outbreaks in semi-domesticated mithun and yak population in various parts of India. A total of 64 clinical samples (vesicle/tongue/foot epithelium/fluid) from mithun and 6 from yak were collected from suspected FMD outbreaks during 2008-2013. Supernatants of the homogenized clinical samples were tested in a serotype discriminating antigen detection ELISA and ELISA-negative samples were further subjected to multiplex reverse transcription-polymerase chain reaction (mRT-PCR). A total of 45 mithun samples and only 1 yak sample were found positive for serotype O in antigen detection ELISA. A total of 12 ELISA-negative samples from mithun and 4 from yak were later on found positive for serotype O in mRT-PCR. The phylogenetic analysis based on VP1 genome indicated the involvement of both O/ME-SA/PanAsia and O/ME-SA/Ind2001 lineages of serotype O in those outbreaks. These viruses were genetically similar to those contemporary virus isolates responsible for FMD in domestic livestock indicating a situation of virus sharing among different species of domestic and semi-domestic animals. Thus, mithuns and yaks should be synchronously considered and targeted along with cattle for the effective control of the disease in the country. [ABSTRACT FROM AUTHOR]

A total of 41 clinical samples (vesicle/tongue/foot/nasal epithelium) from Indian gaur, deer, spotted deer, nilgai, chowsinga, bison, black buck, elephant, sambar deer were collected in 50% phosphate buffered saline/glycerol medium (pH-7.5) during suspected FMD outbreaks. Supernatants of homogenized clinical samples were used in a serotype differentiating antigen detection ELISA and samples found negative were further subjected to multiplex PCR (mPCR). A total of 3/11 (27.2%) samples from Indian gaur, 2/7 (28.5%) chital deer, 5/5 (100%) nilgai, 2/2 (100%) black buck were found positive for serotype O in antigen detection ELISA. A total of 3 ELISA-negative samples from spotted deer, 2 from bison and 2 from sambar deer were found positive for serotype O in mPCR. The VP1 region-based phylogenetic analysis indicated the involvement of both O/ME-SA/Ind2001 and PanAsia lineage of serotype O in the outbreaks. The wildlife species infected with FMD may pose further threat to the surrounding domestic livestock. [ABSTRACT FROM AUTHOR]

Foot and mouth disease (FMD) is an economically important disease and a whole-virus inactivated trivalent virus vaccine is the mainstay for controlling the disease in India. The protective humoral immune response to FMD vaccination is a complex, but, tightly regulated process mediated by the interplay of interleukins (IL). Based on the specific role of IL6 and 21 in adaptive immune response, we hypothesized that inactivated trivalent FMD vaccine would stimulate IL6 and 21 expression in the circulating lymphocytes. The expressions of IL6 and 21 were assayed on 0, 28, 60, 90, and 120 d post-vaccination (DPV) by quantitative PCR (qPCR) with simultaneous assessment of FMDV antibody titer by liquid phase blocking ELISA. The results revealed that the peak expression of IL6 and 21 was on DPV 28 which correlated well with the FMDV antibody titer and plummeted to the prevaccination titer level by 60 DPV. As IL21 is the final effector of antibody production as compared to IL6, we investigated the expression of IL21 in calves that had protective titer (>1.8) with the unprotected group (<1.8). Expression of IL21 on 28 DPV was numerically higher in the protected than that of the unprotected group of calves. [ABSTRACT FROM AUTHOR]

Abstract: In India, systematic genotype replacement has been observed for serotype A foot-and-mouth disease virus. After a decade of co-circulation of genotypes VI and VII, genotype VII emerged as the single dominant genotype since 2001. To derive possible explanations for such epochal evolution dynamics, in vitro intergenotype growth competition experiments involving both co- and superinfection regimes were conducted. Coinfection of BHK-21 cells demonstrated abrupt loss in the genotype VI viral load with commensurate increase in the load of genotype VII as measured by the genotype differentiating ELISA, RT-PCR and real-time RT-PCR. The superinfection dynamics was shaped by temporal spacing of infection, where the invading genotype VII took more number of passages than coinfection to eventually overtake the resident genotype VI. It was speculated that such superior replicative fitness of genotype VII could have been a possible factor for the ultimate dominance of genotype VII in nature. [Copyright &y& Elsevier]