MOLECULAR CHARACTERISATION OF PORCINE CIRCOVIRUS 2 AND DEVELOPMENT OF A REAL-TIME POLYMERASE CHAIN REACTION FOR DETECTION OF THE VIRUS

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dc.contributor.advisorDr. Chintu Ravishankar
dc.contributor.authorSHASHANK S
dc.date.accessioned2024-02-05T13:06:03Z
dc.date.available2024-02-05T13:06:03Z
dc.date.issued2023-01-23
dc.description.abstractPorcine circovirus 2 (PCV2) is an emerging swine pathogen causing enormous economic losses to the global swine industry. The virus has been reported in many of the pig-producing states of India including Kerala. However, no systematic study has been performed to characterize the virus and elucidate the lineage of the virus prevalent in pigs in Kerala. Hence, the present study was undertaken to characterise PCV2 in pigs in Kerala by molecular methods, and to develop a rapid and specific assay for the detection of the virus. A total of 62 tissue samples taken for the study were screened for the presence of PCV2 by ORF2-based PCR, in which 36 samples (58.06 per cent) were positive. The complete genome of the virus was sequenced from 11 representative positive samples. The sequences obtained revealed similarity to PCV2 sequences reported from India previously, and to sequences from China and Vietnam. Phylogenetic analysis was carried out based on complete genome, and partial and complete ORF2 sequences. The analysis revealed that of the 11 PCV2 genomes analysed, eight, two and one belonged to genotypes PCV2d, PCV2h, and PCV2b respectively. It was also noticed that prior to 2021, only genotype 2d was prevalent in North Kerala. Since 2021, 2b and 2h were detected in the region which may be due to introduction of these genotypes from other areas where these genotypes are prevalent. In case of analysis of ORF2 sequences, close clustering of Kerala sequences with those from Tamil Nadu, Uttar Pradesh and Mizoram were observed. The sequences were also similar to PCV2 sequences reported from Kerala previously. On analysis of the complete amino acid sequence of ORF2, it was observed that sample 118/MIB/2021 had Asparagine (N) at the 234th position, which was not been recorded in any Kerala or Indian sequences so far. Based on the ORF2 nucleotide sequences obtained in the study, primers and TaqMan probes were designed for the detection of PCV2 genotypes prevalent in Kerala. A single TaqMan real-time PCR that could detect all the genotypes could not be standardised. However, the designed primers (but not the probe) were able to detect these genotypes. Hence, another TaqMan assay specific for detection of 2d was designed as that genotype was predominant in North Kerala. The detection limit estimated using the cloned template was found to be 310 copies of the viral genome. The designed TaqMan assay was used to test 40 samples (20 positive and 20 negative samples) and 23 samples (57.5 per cent) were positive indicating a higher sensitivity of the test.
dc.identifier.urihttps://krishikosh.egranth.ac.in/handle/1/5810206706
dc.keywordsMOLECULAR CHARACTERISATION OF PORCINE CIRCOVIRUS 2 AND DEVELOPMENT OF A REAL-TIME POLYMERASE CHAIN REACTION FOR DETECTION OF THE VIRUS
dc.keywordsPORCINE CIRCOVIRUS 2
dc.keywordsREAL-TIME POLYMERASE CHAIN REACTION
dc.keywordsLoop-Mediated Isothermal Amplification (LAMP)
dc.keywordsTaqMan real-time PCR
dc.keywordsPCR
dc.keywordsPCV2
dc.keywordsGenotypes
dc.keywordsphylogeny
dc.language.isoEnglish
dc.pages154
dc.publisherCOLLEGE OF VETERINARY AND ANIMAL SCIENCS, POOKODE, WAYANAD, KERALA VETERINARY AND ANIMAL SCIENCES UNIVERSITY
dc.research.problemMolecular characterisation of Porcine circovirus 2 (PCV-2) based on whole genome sequence of the virus, Development of a TaqMan probe based real-time polymerase chain reaction (RT-PCR) for detection of the virus
dc.subVeterinary Microbiology
dc.themeMolecular characterisation of Porcine circovirus 2 and development of a real-time polymerase chain reaction for detection of the virus
dc.these.typeM.V.Sc.
dc.titleMOLECULAR CHARACTERISATION OF PORCINE CIRCOVIRUS 2 AND DEVELOPMENT OF A REAL-TIME POLYMERASE CHAIN REACTION FOR DETECTION OF THE VIRUS
dc.typeThesis
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