CHARACTERIZATION OF GENETIC DETERMINANTS OF VIRULENCE IN AFRICAN SWINE FEVER VIRUS
Foreign Animal Disease Research
2012 Annual Report
1a.Objectives (from AD-416):
The objectives of this research project are (1) to assess the role of different regions of African Swine Fever Virus (ASFV) genomes in virus virulence in swine and (2) to understand the mechanisms of virus infection and persistence.
1b.Approach (from AD-416):
To fulfill objective 1, a set of genetically modified ASFV deletion mutants will be constructed from virulent wild-type viruses. Virulence of mutant viruses will be determined in swine relative to virulence of parental wild-type viruses.
To fulfill objective 2, infections with wild-type and mutant ASFV will be established in vivo and in relevant cells of swine origin in vitro. Persistently and acutely infected cells and recovered viruses will be analyzed by a combination of techniques including microscopy, fluorescence confocal microscopy, qPCR, sequencing and sequencing analysis. Isolated viruses will be characterized in vitro and in vivo relative to wild-type viruses.
Identification and characterization of host and viral mediating disease will be an important goal in this project. Overall experimental manipulation of identified host-virus interactions may be used for developing novel tools for controlling virus infection in vivo.
During FY 2012, we designed strategies and started to produce recombinant African Swine Fever Virus (ASFV). The first target for deletion has been the 9GL gene known to be involved in ASFV virulence. The virulent ASFV strain used in these studies is Georgia 2007, responsible for causing severe outbreaks of the disease in the Caucasus region and Russia since it was introduced accidentally into the Republic of Georgia in 2007. A recombinant delta 9GL-ASFV Georgia was developed, and after sequence verification, the virus was characterized for growth in primary cultures in swine macrophages and tested for virulence relative to parental ASFV Georgia 2007 in swine. Additionally, further deletion mutants have been designed and are under the process of being rescued after recombination. A set of ASFV Georgia 2007 proteins are being expressed in a baculovirus-insect cell expression system for protein-protein interaction studies.