|Overend, C. - UNIV OF CONNECTICUT|
|Mitchell, R. - UNIV OF CONNECTICUT|
|He, D. - S.CHINA AGRICULTURE UNIV.|
|Rompato, G. - UINV OF CONNECTICUT|
|Garmendia, Antonio - UNIV OF CONNECTICUT|
Submitted to: Journal of General Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 30, 2006
Publication Date: March 1, 2007
Citation: Overend, C., Mitchell, R., He, D., Rompato, G., Grubman, M.J., Garmendia, A.E. 2007. Recombinant swine interferon beta protects swine alveolar macrophages and marc 145 cells from infection with porcine reproductive and respiratory syndrome virus. Journal of General Virology. 88(Pt 3): 925-31. Interpretive Summary: Porcine reproductive respiratory syndrome virus (PRRSV) is considered one of the most important pathogens of swine and adversely affects the industry as a result of direct and indirect losses. PRRSV has become widely distributed throughout the world and substantial efforts are ongoing to design rational and effective control strategies. The mechanisms of protective immunity against PRRSV are not completely understood although there appears to be a consensus that both neutralizing antibody and cellular immunity are required for protection. Despite a high variability between animals a strong cellular immunity, appears to correlate with protection from PRRSV. Innate immunity plays a crucial role as a first line of defense against viral infections, including PRRSV, and it is characterized by being rapidly induced upon infection. We have previously constructed a replication-defective human adenovirus type 5 (Ad5) vector containing the swine interferon beta (IFN beta) gene (Ad5-swIFN beta) and demonstrated high levels of expression of biologically active IFN beta protein in swine cells. This virus also induced a rapid antiviral response after administration to swine. The present study was conducted to examine the ability of swIFN beta to neutralize PRRSV in cell cultures as a step towards understanding its possible antiviral role in the animal. The protection by swIFN beta of swine alveolar macrophages and Marc145 cells from PRRSV infection is discussed.
Technical Abstract: Swine interferon beta (swIFN beta) produced in 293 cells infected with a recombinant, replication-defective human adenovirus 5 (Ad5) encoding the swIFN beta gene was tested for antiviral activity against porcine reproductive and respiratory syndrome virus (PRRSV). Marc145 cells were incubated overnight with dilutions of supernatant fluids from 293 cells infected with Ad5-swIFN beta or fluids from an Ad5 control virus (Ad5-Blue). Treated cells were infected with PRRSV and observed for cytopathic effects (CPE). PRRSV-infected Marc145 cells incubated with Ad5-Blue supernatants developed CPE, while those incubated with swIFN beta showed no CPE. Non-inoculated cells remained normal during the same period. To confirm the antiviral activity of swIFN beta, culture fluids from Ad5-swIFN beta infected cells were affinity purified, on a Sepharose/anti-swIFN beta matrix, and the resulting fractions exhibited antiviral activity upon infection with PRRSV. Additional cultures of Marc145 treated with swIFN beta-containing supernatants or affinity purified swIFN beta were infected with PRRSV and tested by real-time RT-PCR for viral RNA in culture supernatants at various times post-inoculation. These experiments confirmed the protective effects of swIFN beta. swIFN beta was tested for antiviral activity on porcine alveolar macrophages (PAM) obtained by bronchioalveolar lavage from PRRSV-negative swine. PAMs were treated with dilutions of swIFN beta or Ad5-Blue culture fluids. Treated cells were infected with PRRSV and tested for viral RNA by real-time RT- PCR. The viral load data showed a dose dependent protection in swIFN beta treated PAMs while no protection was evident from Ad5-Blue culture fluids. The data demonstrates that swIFN beta protects both Marc145 cells and PAMs from PRRSV infection.