In vitro production of Spodoptera exigua multiple nucleopolyhedrovirus with enhanced insecticidal activity using a genotypically defined virus inoculum.

Defective virus accumulations during baculovirus passages in insect cell culture are impediments to large scale baculovirus production. A genotypically defined virus inoculum comprises of stable genotypes was proposed for production of a Thailand isolated SeMNPV in Se-UCR1 insect cells. Targeted genotypes were from wild-type SeMNPV containing naturally mixed genotypes. Plaque assays, PCR screening and XbaI restriction analysis were employed for genotype purification, genotype selection and genome analysis, respectively. A selective marker was pif2 encoded per os infection factor which predominantly deleted, along with the adjacent pif1, in defective viruses. A purified, genetically stable pif2+ (and pif1+) genotype, namely SeThpif2+, was the first tryout. SeThpif2+ occlusion bodies (OBs) possessed insecticidal activity but at lower level than the wild-type. When the SeThpif2+ was co-infected with another purified, genetically stable pif1- (and pif2-) genotype, SeThpif2-, at ratio of 3:1, respectively, mixed genotypes OBs had 2.8 times greater insecticidal activity than the SeThpif2+ alone. Dilution of deleterious PIF1 of SeThpif2+ by the pif1 deletion genotypes, SeThpif2-, was the key for this enhanced activity. A promising approach was described for SeMNPV production in vitro using the virus inoculum whose genotypes compositions were designed to mimic virus interactions in the wild-type, to generate per oral infective baculovirus.