title: A nine-base nucleotide sequence in the porcine circovirus type 2 (PCV2) nucleocapsid gene determines viral replication and virulence.
abstract: Porcine circovirus type 2 (PCV2) was retrospectively identified by serology in swine populations as an asymptomatic infection at least 25 years prior to the first reported case of PCV2-associated postweaning multisystemic wasting syndrome (PMWS). To investigate the sudden emergence of PMWS, viral sequences were amplified from frozen archived (1970-1971) porcine tissues and the complete genome of archival PCV2 was determined. The ORF1 gene product (viral DNA replicase) was homologous to contemporary PCV2 ORF1. In ORF2 (viral nucleocapsid gene) archival PCV2, a consistent linear nine-base sequence difference at base positions 1331 through 1339 was observed. The deduced amino acid sequence from these base changes alters the nucleocapsid conformation within the second immunogenic epitope from a hydrophobic (contemporary PCV2) to a hydrophilic (archival PCV2) configuration. To test the hypothesis that archival PCV2 was avirulent, cloned engineered archival and contemporary PCV2 genomes were constructed wherein the ORF1 gene was identical in each clone and the ORF2 gene (nucleocapsid protein) was sequence-identical in both clones except for the nine-base difference (bases 1331-1339), corresponding to archival and contemporary PCV2 viruses respectively. Clones were transfected into porcine kidney (PK) 15 cells and, after sequence confirmation, further passed in PK15 and 3D4/2 porcine alveolar macrophage cell cultures. Virulence trials in gnotobiotic piglets were conducted with cloned PCV2s. The data show that archival PCV2 is avirulent when compared to contemporary PCV2 and supports the hypothesis that the emergence of virulent contemporary PCV2 was a result of mutational events within this critical epitope after 1971.
authors.affiliation: The Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University, 1925 Coffey Road, Columbus, OH 43210, USA. Krakowka.1@osu.edu
authors.forename: Steven
authors.initials: S
authors.lastname: Krakowka
authors.forename: Gordon
authors.initials: G
authors.lastname: Allan
authors.forename: John
authors.initials: J
authors.lastname: Ellis
authors.forename: Alexander
authors.initials: A
authors.lastname: Hamberg
authors.forename: Catherine
authors.initials: C
authors.lastname: Charreyre
authors.forename: Eva
authors.initials: E
authors.lastname: Kaufmann
authors.forename: Charles
authors.initials: C
authors.lastname: Brooks
authors.forename: Brian
authors.initials: B
authors.lastname: Meehan
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D017925:Circovirus / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity*; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D019251:Nucleocapsid / Q000235:genetics*; D053570:Porcine Postweaning Multisystemic Wasting Syndrome / Q000473:pathology / Q000821:virology*; D017422:Sequence Analysis, DNA; D013552:Swine; D014774:Virulence; D037521:Virulence Factors / Q000235:genetics*; D014779:Virus Replication
chemical_list: D004279:DNA, Viral; D037521:Virulence Factors