African swine fever virus Georgia isolate harboring deletions of 9GL and MGF360/505 genes is highly attenuated in swine but does not confer protection against parental virus challenge

African swine fever virus (ASFV) produces a contagious disease of domestic pigs that results in severe economic consequences to the swine industry. Control of the disease has been hampered by the unavailability of vaccines. We recently reported the development of two experimental vaccine strains (ASFV-G-Delta 9GL and ASFV-G-Delta MGF) based on the attenuation of the highly virulent and epidemiologically relevant Georgia2007 isolate. Deletion of the 9GL gene or six genes of the MGF360/505 group produced two attenuated ASFV strains which were able to confer protection to animals when challenged with the virulent parental virus. Both viruses, although efficient in inducing protection, present concerns regarding their safety. In an attempt to solve this problem we developed a novel virus strain, ASFV-G-Delta 9GL/Delta MGF, based on the deletion of all genes deleted in ASFV-G-Delta 9GL and ASFV-G-Delta MGF. ASFV-G-Delta 9GL/Delta MGF is the first derivative of a highly virulent ASFV field strain subjected to a double round of recombination events seeking to sequentially delete specific genes. ASFV-G-Delta 9GL/Delta MGF showed a decreased ability to replicate in primary swine macrophage cultures relative to that of ASFV-G and ASFV-G-Delta MGF but similar to that of ASFV-G-Delta 9GL. ASFV-G-Delta 9GL/Delta MGF was attenuated when intramuscularly inoculated into swine, even at doses as high as 10(6) HAD(50). Animals infected with doses ranging from 10(2) to 10(6) HAD(50) did not present detectable levels of virus in blood at any time post-infection and they did not develop detectable levels of anti-ASFV antibodies. Importantly, ASFV-G-Delta 9GL/Delta MGF does not induce protection against challenge with the virulent parental ASFV-G isolate. Results presented here suggest caution towards approaches involving genomic manipulations when developing rationally designed ASFV vaccine strains. Published by Elsevier B.V.