Isolation and complete genome analysis of porcine circovirus-1 associated with reproductive failurein swine: foremost study in India.

This study investigated 100 porcine tissue samples from stillborn, dead and or aborted foetus with the history of reproductive infections from Tamil Nadu, Kerala, Andhra Pradesh, and Karnataka. All samples were screened for the genomes of PCV1, PCV2, PCV3, CSFV, PPV, PRRSV, and Brucella species using specific PCR assays. Out of 100 samples screened, the molecular positivity rates were 12% for PCV1, 11% for PCV2, 2% for PCV3, 6% for CSFV, 9%for PPV and 5%for Brucella genomes. Among the twelve PCV1 positive samples, four were positive for the PCV1 genome alone, five showed co-infection with PCV2, two with PPV, and one with CSFV. No co-infection of PCV1 with PCV3, PRRSV and Brucella were detected in this study. Four PCV1-positive samples, confirmed to be free from other co-infections, were propagated in PCV1-free PK-15 cells and subjected to five successive blind passages. The presence of PCV1 was confirmed by PCR in two of the isolates, which were subsequently subjected to whole-genome sequencing using the Illumina NovaSeq 6000 platform. Complete genome sequences of two PCV1 isolates (K44ABT and L11ABT)comprising 1759 base pairs with a G + C content of 48%were submitted to GenBank under the accession numbers PQ303661 and PQ303662 respectively. The ORF1 and ORF2 regions encoded the replicase (312 amino acids) and capsid (233 amino acids) proteins respectively. BLAST analysis and genome similarity analysis revealed > 99% identity with published PCV1 strains, and phylogenetic analysis showed clustering with strains from India, USA, and China. Analysis of Rep protein revealed highly conserved sequences in both the PCV1 isolates, but, exhibited a substitution alanine to valine at position 102in the receptor-binding domain (RBD) located in the capsid protein. This altered RBD showed homology with those of virulent PCV2 and PCV4, suggesting a potential for broader cell binding similar to that seen in pathogenic circoviruses. Further, examination of three key B-cell epitopes 85GGTNPLP91, 162FTPKPELDQTIDWFHPNNK180, and 219YVQFREFILKDPLNE233 in capsid protein showed notable variation at 170 and 233 positions with substitution of Q to K and E to K respectively. This study represents the first complete genome analysis of PCV1 associated with reproductive failure in domestic pigs from southern India, providing important insights into its molecular characteristics. The observed genetic variations, particularly in the capsid protein, imply a potential shift in PCV1 toward enhanced host adaptation and pathogenicity in pigs.