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United States Department of Agriculture

Agricultural Research Service

Title: Comparison of the pathogenicity of porcine circovirus type 2 group 1 and group 2 isolates

item Cheung, Andrew
item Lager, Kelly
item Gauger, Phillip
item Vincent, Amy
item Opriessnig, T - IOWA STATE UNIVERSITY

Submitted to: International Symposium on Emerging and Re-Emerging Pig Diseases
Publication Type: Proceedings
Publication Acceptance Date: June 7, 2007
Publication Date: June 24, 2007
Citation: Cheung, A.K., Lager, K.M., Gauger, P.C., Vincent, A.L., Opriessnig, T. 2007. Comparison of the pathogenicity of porcine circovirus type 2 group 1 and group 2 isolates. In: Proceedings of the 5th International Symposium on Emerging and Re-Emerging Pig Diseases, June 24-27, 2007, Krakow, Poland. p. 273.

Technical Abstract: 1. Introduction and Objectives Phylogenetically, porcine circovirus type 2 (PCV2) can be divided into two major genotypic groups, PCV2-group 1 and PCV2-group 2 .1 It was noted that PCV2 group designations have no apparent association with disease status or geographic area. Interestingly, all of the PCV2 isolates from the United States until late 2005 belonged to group 2, while virus isolates from other countries may be found in both PCV2 groups. In late 2005, sporadic cases of an acute onset disease of high mortality were observed in 10- to 16-week-old growing pigs among USA swine herds. Tissues from affected pigs in Kansas, Iowa, and North Carolina were examined and PCV2-group 1 virus was consistently detected among these tissues.2 Nucleotide sequence analysis revealed several distinct features that are characteristic of either PCV2-group 1 (71 entries) or PCV2-group 2 (62 entries) : (1) The PCV2-group 1 sequences are 1767 nucleotides, while PCV2-group 2 sequences are 1768 nucleotides. The deleted nucleotide of PCV2-group 1 is located at nucleotide 1040. (2) Rep (18%) and ORF3 (16%) exhibit fewer nucleotide variations than the capsid gene (28%). (3) Distinct stretches of nucleotide or amino acid sequences may serve as signature motifs for genotype grouping of PCV2 isolates. The capsid gene nucleotide sequence at position 1486-1476 for PCV2-group 1 (87 entries) is TCA/AAC/CCC/CG (which codes for the N-terminal residues of amino acid sequence SNPRSV), while the nucleotide sequence at position 1487-1477 for PCV2-group 2 (64 entries) is ACC/AAC/AAA/AT (which codes for the N-terminal residues of amino acid sequence TNKI S[I62/64]). The objective of this communication was to examine the pathogenic capability of PCV-group 1 and PCV-group 2 isolates in swine. 2. Material and methods Virus. Infectious viruses were generated from bacterial plasmids containing PCV2-group 1 (GenBank accession number DQ629115) and PCV2-group 2 (GenBank accession number DQ629114) nucleotide sequences. The viral genomes were excised from the bacterial plasmids, ligated and delivered into PK15 cells via lipofectamine transfection. Infectious viruses were recovered from the transfection cell cultures after 1 week. Experimental design. Twelve pigs were derived under sterile conditions and maintained germ-free in isolators (4 pigs / isolator). The pigs were fed a sterile milk diet and at 12 days of age (day 0 of the experiment) were given an intranasal inoculation of either PCV2-group 1 virus (Isolator-A), PCV2-group 2 virus (Isolator-B) or cell culture medium (Isolator-C). The pigs were monitored for clinical signs with a plan of euthanizing all pigs 35 days-post-inoculation. 3. Results Day 22: In the morning 1 Isolator-A pig was anorexic, listless, and had a mild dyspnea. During the course of the day it developed a severe dyspnea and by that evening a second pig in the same isolator became anorexic, listless, and had a mild dyspnea. Both pigs were euthanized for post-mortem examination. They had similar gross lesions consisting of peritoneal and thoracic effusions that were cloudy and icteric, pneumonia with interlobular edema, enlarged tracheobronchial lymph nodes, and gelatinous edema around the kidneys, base of the heart, and the mesentery of the spiral colon. Hemorrhages were seen along the renal cortico-medullary junction. Two pigs from Isolator-B and Isolator-C were euthanized and examined on day 23. No gross lesions were observed in either Isolator-C pig. The only lesions found in the Isolator-B pigs were enlarged tracheobronchial lymph nodes. Day 27: On the morning of day 27 one pig in Isolator-A was found dead and the last remaining pig was listless, anorexic, and had severe dyspnea. This pig was euthanized. Both pigs had gross lesions similar to the other 2 Isolator-A pigs euthanized 5 days earlier. Thus, all 4 Isolator-A pigs were acutely affected. Day 35: The remaining pigs in Isolator- B and Isolator-C were normal in appearance and behavior until day 35. On day 35 one of the 2 Isolator-B pigs was anorexic, listless, and had mild dyspnea. This pig had gross lesions similar to all 4 pigs in Isolator-A. The other Isolator-B pig was normal in appearance as was the remaining 2 Isolator-C pigs. Microscopic lesions were only observed in PCV2 challenged pigs and were most severe in affected pigs. The most significant lesions were depletion and replacement of lymphoid follicles by macrophages, and severe vacuolar degeneration and necrosis of hepatocytes with multifocal areas of hemorrhage. Abundant PCV2 antigen was observed only in tissues of affected pigs. PCV2 was only recovered from pigs in Isolator- A and Isolator-B and the virus was genetically identical to the challenge inoculum. 4. Discussion and Conclusions In this preliminary study, the PCV2-group 1 isolate was more pathogenic when compared to the PCV2-group 2 isolate. The clinical effects and lesions observed in this study were similar to what has been reported previously for germ-free pigs inoculated with PCV2 and porcine parvovirus.3,4 In our analysis, no other microorganisms were detected in the affected pigs. Since the viruses used in this study were derived from cloned DNA, we presume the clinical effects were attributed to the properties of the respective virus isolate. 5. Acknowledgements The authors thank S. Pohl, D. Adolphson, Dr. D. Alt and K. Halloum for technical assistance. 6. References 1. Olvera A, Cortey M, Segales J (2007) Molecular evolution of porcine circovirus type 2 genomes: phylogeny and clonality. Virology 357: 175-185. 2. Cheung A, Lager K, Kohutyuk O, Vincent A, Henry S, Baker R, Rowland R, and Dunham A (2007) Detection of two porcine circovirus type 2 genotypic groups in United States swine herds. Arch Virol (in press) 3. Ellis J, Krakowka S, Lairmore M, Haines D, Bratanich A, Clark E, Allan G, Konoby C, Hassard L, Meehan B, Martin K, Harding J, Kennedy S, McNeilly F. (1999) Reproduction of lesions of postweaning multisystemic wasting syndrome in gnotobiotic piglets. J Vet Diagn Invest 11:3-14. 4. Krakowka S. Eillis J, Meehan B, Kennedy S, McNeilly F, Allan G. (2000) Viral wasting syndrome of swine: Experimental reproduction of postweaning multisystemic wasting syndrome in gnotobiotic swine by coinfection with porcine circovirus 2 and porcine parvovirus. Vet Pathol 37:254-263.

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