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United States Department of Agriculture

Agricultural Research Service

Title: Protection Of Alveolar Macrophages And MARC 145 Cells From Porcine Reproductive And Respiratory Syndrome Virus Challenge By Swine Interferon-Beta

Authors
item Overend, Chris - UNIV. OF CONN
item Mitchell, R - UNIV. OF CONN
item He, D - S.CHINA AGRI. UNIV.
item Rompato, G - UNIV. OF CONN
item Grubman, Marvin
item Garmendia, A - UNIV. OF CONN

Submitted to: Porcine Reproductive and Respiratory Syndrome International Symposium
Publication Type: Abstract Only
Publication Acceptance Date: November 15, 2006
Publication Date: December 1, 2006
Citation: Overend, C., Mitchell, R., He, D., Rompato, G., Grubman, M.J., Garmendia, A.E. 2006. Protection Of Alveolar Macrophages And MARC 145 Cells From Porcine Reproductive And Respiratory Syndrome Virus Challenge By Swine Interferon-Beta International Porcine Reproductive and Respiratiory Syndrome (PRRS) Symposium 2006. P. 26. . Porcine Reproductive and Respiratory Syndrome International Symposium.

Technical Abstract: Interferon beta, a type I IFN, is crucial in initiating the innate immune response and in the generation of the adaptive response. This study demonstrated the capacity of swine interferon beta (swIFN beta) to protect porcine alveolar macrophages (PAM) and MARC145 cells from infection with porcine reproductive and respiratory syndrome virus (PRRSV). The swIFN beta used in this study was produced in HEK293 cells via a recombinant replication-defective human adenovirus 5 (Ad5) encoding the swIFN beta gene (Ad5 swIFN beta). At various times after infection of HEK293 cells infected with either Ad5-swIFN beta or Ad5-Blue (control) supernatants were harvested, acid treated (pH 2.0) and neutralized. To test antiviral activities, target cells, MARC145 or PAMs, were incubated for 18-20 hours with dilutions of either of the treated supernatants. The cells were then infected with PRRSV, and the presence of cytopathic effects (CPE) recorded. The results showed that cells treated with Ad5-swIFN beta supernatants had no CPE while the cells treated with control supernatant had CPE. As expected uninfected control cell monolayers remained intact throughout the test period, whereas untreated, PRRSV-infected cells exhibited marked CPE. Culture supernatants of IFN-primed PAMs or MARC145 cells were collected at various time-points post-infection for determination of viral RNA loads using real time RT-PCR. The PCR analysis of these culture supernatants supported the findings of the protective effects of swIFN beta recorded as CPE, and demonstrated that protection occurred in a dose dependent manner. Furthermore, supernatants of HEK293 cells infected with Ad5-swIFN beta were affinity purified on a Sepharose-anti-swIFN beta column. Priming with the IFN beta affinity fractions protected MARC145 cells from infection with PRRSV, thus confirming earlier results obtained with crude supernatant preparations. These findings demonstrate that swIFN beta is capable of protecting both PAMs and MARC145 cells from subsequent infection with PRRSV.

Last Modified: 8/29/2014
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