Vikramaditya Upmanyu

Classical swine fever in pigs: recent developments and future perspectives

Abstract Classical swine fever (CSF) is one of the most devastating epizootic diseases of pigs, causing high morbidity and mortality worldwide. The diversity of clinical signs and similarity in disease manifestations to other diseases make CSF difficult to diagnose with certainty. The disease is further complicated by the presence of a number of different strains belonging to three phylogenetic groups. Advanced diagnostic techniques allow detection of antigens or antibodies in clinical samples, leading to implementation of proper and effective control programs. Polymerase chain reaction (PCR)-based methods, including portable real-time PCR, provide diagnosis in a few hours with precision and accuracy, even at the point of care. The disease is controlled by following a stamping out policy in countries where vaccination is not practiced, whereas immunization with live attenuated vaccines containing the ‘C’ strain is effectively used to control the disease in endemic countries. To overcome the problem of differentiation of infected from vaccinated animals, different types of marker vaccines, with variable degrees of efficacy, along with companion diagnostic assays have been developed and may be useful in controlling and even eradicating the disease in the foreseeable future. The present review aims to provide an overview and status of CSF as a whole with special reference to swine husbandry in India.

Association of humoral response to classical swine fever vaccination with single nucleotide polymorphisms of swine leukocyte antigens

In present investigation genetic variability in humoral response to classical swine fever (CSF) vaccination was explored. The aim of the study was to find association of 14 single nucleotide polymorphisms (SNPs) (IL-1β, MX1, IRF3 and SLA genes) with humoral response measured in terms of percentage inhibition of E2 antibody by competitive enzyme-linked immunosorbent assay. The post-vaccination response of CSF vaccination was low to moderate in Desi piglets ranged from 25.29% to 92.23% whereas it was moderate to high in Desi × Landrace piglets ranged from 69.30% to 100%. The response to CSF vaccination was significantly (P < 0.0001) differing in Desi and Desi × Landrace piglets. The mean percentage inhibition in Desi piglets was 62.73% but in Desi × Landrace cross-bred it was 97.24%. None of the SNPs of present investigation was significantly associated with humoral immune response in Desi piglets. Two SNPs namely rs80937718 (SLA DQA1) and rs80929898 (SLA DQA1) were significantly (P < 0.01) affecting the humoral response of CSF vaccination in Desi × Landrace piglets.

Biological and molecular characterization of classical swine fever challenge virus from India

Aim: The aim of this study was biological and molecular characterization of classical swine fever (CSF) challenge virus from India. Materials and Methods: CSF challenge virus maintained at Division of Biological standardization was experimentally infected to two seronegative piglets. The biological characterization was done by clinical sign and symptoms along with postmortem findings. For molecular characterization 5’-nontranslated region, E2 and NS5B regions were amplified by reverse transcription polymerase chain reaction and sequenced. The sequences were compared with that of reference strains and the local field isolates to establish a phylogenetic relation. Results: The virus produced symptoms of acute disease in the piglets with typical post-mortem lesions. Phylogenetic analysis of the three regions showed that the current Indian CSF Challenge virus is having maximum similarity with the BresciaX strain (USA) and Madhya Pradesh isolate (India) and is belonging to subgroup 1.2 under Group 1. Conclusion: Based on biological and molecular characterization of CSF challenge virus from India is described as a highly virulent virus belonging to subgroup 1.2 under Group 1 along with some field isolates from India and Brescia strain.

Monoclonal antibody resistant mutant of Peste des petits ruminants vaccine virus

The available vaccines for control of Peste des petits ruminants do not favour differentiation of infected and vaccinated animals (DIVA). Hence, the present study was aimed to isolate and characterize monoclonal antibody resistant mutant of an Indian strain of vaccine virus “PPRV-Sungri/96” under selection pressure of virus neutralizing monoclonal antibody ‘4B11’ specific to haemagglutinin (H) protein. We successfully isolated five monoclonal antibody resistant (mAr) mutants (PPRV-RM5, PPRV-RM6, PPRV-RM7, PPRV- E6 and PPRV- E7). The mAr mutants did not react with the anti-H mAb 4B11 whereas reacted with control anti-nucleoprotein mAb 4G6, similar to the parent vaccine virus “PPRV-Sungri/96” in indirect ELISA, cell ELISA and indirect immunofluorescence test. Cytometry analysis of mAr mutants revealed loss of binding to mAb 4B11 while maintaining binding to mAb 4G6, more or less similar to “PPRV-Sungri/96”. The sequence analysis of the H-protein gene of the mAr mutants resulted in identification of two nucleotide changes leading to amino acid substitutions at position 263 and 502 (L263P and R502P) of the H protein indicating that the epitope of mAb 4B11 could be conformational in nature. Though, mAr mutant grew to a similar titre as parent vaccine virus (PPRV-Sungri/96), the in vivo work in goats to study the mAr mutant as possible negative marker vaccine candidate could not be successfully proved with mAb 4B11 based competitive ELISA. However, one of the nucleotide change (T-C) at position 788, unique to mAr mutant virus resulted in abolition of a restriction enzyme recognition site (BglII). This could be used to differentiate mAr mutant vaccine virus from other available vaccine and field strains using restriction fragment length polymorphism. However, the mAr mutant PPRV-E6 cannot be used as a candidate strain for DIVA vaccine as immune response against it cannot be differentiated based on serology.

Investigation on Peste des Petits Ruminants Outbreak in Goats of Bareilly District of Uttar Pradesh, India

Peste des petits ruminants (PPR) is an economically important viral disease of small ruminants, endemic in India. The present study reports on socio-epidemiological aspect of an outbreak of PPR in goats of Bareilly district of Uttar Pradesh in Northern part of India. During a visit to the affected villages and also from animals brought to a local clinic, it was observed that goats had clinical signs such as pyrexia, oculo-nasal discharges, dyspnea and diarrhoea indicative of PPR. However, oral lesions were not so prominent. In total, 96 clinical samples (ocular and nasal swabs) were collected from the affected goats suspected for PPR. The PPR virus (PPRV) antigen and nucleic acid were confirmed by sandwich-ELISA and ‘N’ gene-based RT-PCR, respectively. Sequence and phylogenetic analysis of partial ‘N’ gene PCR amplicons revealed that the PPRV belonged to lineage IV. The present investigation describes the importance of understanding the role of socio-epidemiological factors on PPR outbreak for efficient implementation of control and eradication strategies.

Replacement of Animal Model for Propagation of Classical Swine Fever Challenge Virus by Adaption in the PK-15 Cell Line

Classical swine fever (CSF) challenge virus has been adapted in PK-15 cell line from infected splenic suspension of the challenge virus maintained hitherto by pig to pig passages. Confirmation of viral presence was done by reverse transcription-polymerase chain reaction (RT-PCR) and Fluorescent Antibody Technique (FAT). A reasonably good titre of 106.5 TCID50/ml was obtained at 6th passage level. The cell culture adapted challenge virus at a dose of 105.0 TCID50 produced CSF symptoms in pigs from 2 nd days post infection (dpi) onwards and succumbed to the infection between 11-12 dpi. Cell culture adapted CSF challenge virus offers advantage to inoculate exact virus particles over the traditional tissue suspension (20% w/v) in potency testing. Adapted challenge virus will replace the use of pigs for propagation of challenge virus; hence follows 4 R’s (replacement, reduction, refinement and rehabilitation) principle. This challenge virus can be attenuated by further serial passages and can be used to develop indigenous live attenuated cell culture based vaccine.

Conventional and molecular characterization of pasteurella multocida isolated from a case of swine septicaemic pasteuerllosis

The paper reports morphological, biochemical characterization, and molecular characterization using HS-PCR, PM-PCR and Capsular PCR analysis of Pasteurella multocida strain isolated from a case of swine pasteurellosis to obtain a clear picture of this strain for its exploitation as a vaccine candidate.