Effect of Cytopathic Bovine Viral Diarrhea Virus (BVDV) Superinfection on Viral Antigen Association with Platelets, Viremia, and Specific Antibody Levels in Two Heifers Persistently Infected with Noncytopathic BVDV

Abstract

Two female Holstein heifers (Nos. 132,138) obtained from the same farm and persistently infected with noncytopathic (NCP) bovine viral diarrhea virus (BVDV) were inoculated intravenously with a heterologous cytopathic (CP) BVDV isolate (Burns). Each received an inoculum containing 103 CCID50 of CP BVDV. 7 Heifer No. 132 was inoculated at 302 days of age with a virus-neutralizing monoclonal antibody (MAb) escape variant of BVDV-Burns, and heifer No. 138 received at 636 days of age parent BVDV-Burns. The CP BVDV-Burns variant used in heifer No. 132 was recovered by in vitro selection for MAb neutralization resistance. Agar containing MAb 12G4a.a was poured over primary bovine testes cell monolayers infected with parent BVDV-Burns isolate. After 7 days of incubation, placques in the agar were harvested for viral expansion. Such a neutralization-escape variant of BVDV-Burns was selected based on its resistance to neutralization by MAb 12G4a.a, differing from BVDVBums parent in its neutralization reaction at only this one gp53-specific epitope. The BVDV-Burns parent and variant were differentiated from the persistently infecting NCP BVDV by reactivity with monoclonal antibody 39.a.7 in an indirect fluorescent antibody staining assay. Clinical responses to superinfection, as well as virus isolation using mononuclear cells (MNC) and necropsy tissues, were recorded. Heifer No. 132 was clinically normal when euthanized at 57 days postsuperinfection (DPSI), and heifer No. 138 was euthanized when moribund, displaying classic signs of mucosal disease at 16 DPSI. Several parameters monitored after inoculation with BVDV-Burns included platelet count, association of NCP BVDV antigen(s) with platelets, viral concentration in serum, and specific virusneutralizing antibody levels in serum. Rectal temperatures were obtained for each heifer twice daily prior to and after super-infection. Both heifers experienced pyrexia (greater than 39.2 C) 2-12 DPSI, with peak fever in No. 132 reaching 41.6 C and peak temperature in No. 138 attaining 42.1 C. Neither heifer showed hemorrhage at any time antemortem. In addition, heifer No. 138 developed other clinical signs of disease, including diarrhea, anorexia, and depression. Gross lesions observed in heifer No. 138 at necropsy were consistent with those previously described for experimentally induced mucosal disease. Tissue specimens harvested from each heifer at necropsy for virus isolation included spleen, mesenteric lymph node, Peyer’s patch, bone marrow, brain,