This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Fenaux, M. Articles by Meng, X. J. Search for Related Content PubMed PubMed Citation Articles by Fenaux, M. Articles by Meng, X. J.

Two Amino Acid Mutations in the Capsid Protein of Type 2 Porcine Circovirus (PCV2) Enhanced PCV2 Replication In Vitro and Attenuated the Virus In Vivo

Center for Molecular Medicine and Infectious Diseases, Department of Biomedical Sciences and Pathobiology,¹ Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Blacksburg, Virginia,³ Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa²

Received 1 April 2004/ Accepted 3 August 2004

Porcine circovirus type 2 (PCV2) is the primary causative agent of postweaning multisystemic wasting syndrome (PMWS) in pigs. To identify potential genetic determinants for virulence and replication, we serially passaged a PCV2 isolate 120 times in PK-15 cells. The viruses harvested at virus passages 1 (VP1) and 120 (VP120) were biologically, genetically, and experimentally characterized. The PCV2 VP120 virus replicated in PK-15 cells to a titer similar to that of the PK-15 cell line-derived nonpathogenic PCV1 but replicated more efficiently than PCV2 VP1 with a difference of about 1 log unit in the titers. The complete genomic sequences of viruses at passages 0, 30, 60, 90, and 120 were determined. After 120 passages, only two nucleotide mutations were identified in the entire genome, and both were located in the capsid gene: the mutations were located at nucleotide positions 328 (C328G) and 573 (A573C). The C328G mutation, in which a proline at position 110 of the capsid protein changed to an alanine (P110A), occurred at passage 30 and remained in the subsequent passages. The second mutation, A573C, resulting in a change from an arginine to a serine at position 191 (R191S), appeared at passage 120. To experimentally characterize the VP120 virus, 31 specific-pathogen-free pigs were randomly divided into three groups. Ten pigs in group 1 received phosphate-buffered saline as negative controls. Each pig in group 2 (11 pigs) was inoculated intramuscularly and intranasally with 10^4.9 50% tissue culture infective doses (TCID₅₀) of PCV2 VP120. Each pig in group 3 (10 pigs) was similarly inoculated with 10^4.9 TCID₅₀ of PCV2 VP1. Viremia was detected in 9 of 10 pigs in the PCV2 VP1 group with a mean duration of 3 weeks, but in only 4 of 11 pigs in the PCV2 VP120 group with a mean duration of 1.6 weeks. The PCV2 genomic copy numbers in serum in the PCV2 VP1 group were significantly higher than those in the PCV2 VP120 group (P < 0.0001). Gross and histopathologic lesions in pigs inoculated with PCV2 VP1 were more severe than those inoculated with PCV2 VP120 at both day 21 and 42 necropsies (P = 0.0032 and P = 0.0274, respectively). Taken together, the results from this study indicated that the P110A and R191S mutations in the capsid of PCV2 enhanced the growth ability of PCV2 in vitro and attenuated the virus in vivo. This finding has important implications for PCV2 vaccine development.

This article has been cited by other articles:

• Lefebvre, D. J., Costers, S., Van Doorsselaere, J., Misinzo, G., Delputte, P. L., Nauwynck, H. J. (2008). Antigenic differences among porcine circovirus type 2 strains, as demonstrated by the use of monoclonal antibodies. J. Gen. Virol. 89: 177-187 [Abstract] [Full Text]  
• Opriessnig, T., Meng, X.-J., Halbur, P. G. (2007). Porcine Circovirus Type 2 associated disease: Update on current terminology, clinical manifestations, pathogenesis, diagnosis, and intervention strategies. jvdi 19: 591-615 [Abstract] [Full Text]  
• Opriessnig, T., McKeown, N. E., Zhou, E.-M., Meng, X.-J., Halbur, P. G. (2006). Genetic and experimental comparison of porcine circovirus type 2 (PCV2) isolates from cases with and without PCV2-associated lesions provides evidence for differences in virulence.. J. Gen. Virol. 87: 2923-2932 [Abstract] [Full Text]  
• Liu, J., Chen, I., Du, Q., Chua, H., Kwang, J. (2006). The ORF3 Protein of Porcine Circovirus Type 2 Is Involved in Viral Pathogenesis In Vivo.. J. Virol. 80: 5065-5073 [Abstract] [Full Text]  
• Opriessnig, T., Fenaux, M., Thomas, P., Hoogland, M. J., Rothschild, M. F., Meng, X. J., Halbur, P. G. (2006). Evidence of Breed-dependent Differences in Susceptibility to Porcine Circovirus Type-2-associated Disease and Lesions. Vet Pathol 43: 281-293 [Abstract] [Full Text]  
• McKeown, N. E., Opriessnig, T., Thomas, P., Guenette, D. K., Elvinger, F., Fenaux, M., Halbur, P. G., Meng, X. J. (2005). Effects of Porcine Circovirus Type 2 (PCV2) Maternal Antibodies on Experimental Infection of Piglets with PCV2. CVI 12: 1347-1351 [Abstract] [Full Text]