SWINE VIRAL DISEASES PATHOGENESIS AND IMMUNOLOGY
Location: Virus and Prion Research Unit
Title: Analysis of Asian PRRSV Type 2 Strains
Submitted to: United States Japan Natural Resources Animal and Avian Health Panel
Publication Type: Abstract Only
Publication Acceptance Date: September 28, 2010
Publication Date: October 5, 2010
Citation: Guo, B., Lager, K.M., Henningson, J.N., Schlink, S.N., Miller, L.C., Brockmeier, S., Yang, H.C., Faaberg, K.S. 2010. Analysis of Asian PRRSV type 2 strains [abstract]. 45th US-Japan Cooperative Program in Natural Resources Panel of Animal and Avian Health Meeting. p. 8.
Novel isolates of porcine reproductive and respiratory disease virus (PRRSV) possessing a unique deletion in nonstructural protein 2 with an ORF5 RFLP of 1-4-3 appeared in China in 2006. The isolates were linked to porcine high fever disease (PHFD) and genetically similar to 2007 PHFD PRRSV isolates detected in Vietnam. This strain has continued to be problematic in several Asian countries. As part of the USDA response to the emergence of PHFD, we received purified RNA from cells infected with the Vietnamese isolate (SRV-07) from the USDA Foreign Animal Disease Diagnostic Laboratory. We also imported an infectious cDNA clone of the Chinese PRRSV JXwn06 strain. Our goals were to examine the ability to detect similar strains and to investigate potential disease consequences to the swine population if such isolates were identified in the U.S.
Using SRV-07 purified RNA, a section encompassing ORF4-3' end was amplified for use in determining diagnostic testing sensitivity. The DNA product was cloned, in vitro transcribed, and the RNA forwarded to selected U.S. veterinary diagnostic laboratories for PRRSV quantitative RT-PCR and ORF5 sequencing. All laboratories were able to detect and sequence SRV-07 RNA, and all predicted an ORF5 RFLP of 143. We have also engineered an infectious clone of SRV-07 for future in vitro and in vivo phenotyping.
Infectious virus was obtained from the Chinese JXwn06 cDNA clone. Under ABSL3 biocontainment, one pig was intranasally inoculated with 2 ml of 10**6 TCID50/ml rJXwn06 at passage 3. On day 2, a contact pig was placed into the pen. By day 6, the inoculated pig was ill and, on day 9, was moribund and euthanized. Necropsy serum virus titer was 10**7 TCID50/ml. The contact pig only developed mild disease and was euthanized on day 17 (15 days post contact) and PRRSV (10**5.3 TCID50/ml) was also isolated from serum. We recently infected a larger group of swine by intranasal inoculation with either 2 ml of 10**6 TCID50/ml PRRSV strain JXwn06 (n=12 challenge, n=4 contact) or the Type 2 prototype strain VR-2332 (n=8), or a sham inoculum (n=8). The pigs were monitored for 13 days during which time the JXwn06 pigs developed severe disease with 4 of them either dying or were moribund and euthanized prior to the study end. Sham-inoculated pigs and VR-2332-infected pigs did not show any severe signs. Swine were bled at days 0, 4, 7, 11 and 13 and weighed on days -1, 7 and 13. At necropsy, lung lavage was completed, tissues were collected for histopathology and tracheal bronchial lymph nodes were weighed. Labwork is underway to examine the PRRSV load in serum and lung lavage; bacterial growth in lung lavage, pericardial fluid and meninges; and pertinent cytokine RNA and protein levels in lymph nodes as well as cytokine protein levels in serum. In addition, each pig will be tested for adventitious viruses. Preliminary results of our analyses will be presented.