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ABSTRACT Bracoviruses (BVs) are endogenized nudiviruses in parasitoid wasps of the microgastroid complex (order Hymenoptera: Family Braconidae). BVs produce replication-defective virions that adult female wasps use to transfer DNAs encoding virulence genes to parasitized hosts. Some BV genes are shared with nudiviruses and baculoviruses with studies of the latter providing insights on function, whereas other genes are only known from nudiviruses or other BVs which provide no functional insights. A proteomic analysis ofMicroplitis demolitorbracovirus (MdBV) virions recently identified 16 genes encoding nucleocapsid components. In this study, we further characterized most of these genes. Some nucleocapsid genes exhibited early or intermediate expression profiles, while others exhibited late expression profiles. RNA interference (RNAi) assays together with transmission electron microscopy indicatedvp39,HzNVorf9-like2,HzNVorf93-like,HzNVorf106-like,HzNVorf118-like,and 27bare required to produce capsids with a normal barrel-shaped morphology. RNAi knockdown ofvlf-1a,vlf-1b-1,vlf-1b-2,int-1,andp6.9-1did not alter the formation of barrel-shaped capsids but each reduced processing of amplified proviral segments and DNA packaging as evidenced by the formation of electron translucent capsids. All of the genes required for normal capsid assembly were also required for proviral segment processing and DNA packaging. Collectively, our results deorphanize several BV genes with previously unknown roles in virion morphogenesis. IMPORTANCEUnderstanding how bracoviruses (BVs) function in wasps is of broad interest in the study of virus evolution. This study characterizes most of theMicroplitis demolitorbracovirus (MdBV) genes whose products are nucleocapsid components. Results indicate several genes unknown outside of nudiviruses and BVs are essential for normal capsid assembly. Results also indicate most MdBV tyrosine recombinase family members and the DNA binding proteinp6.9-1are required for DNA processing and packaging into nucleocapsids. 

Thousands of endoparasitoid wasp species in the families Braconidae and Ichneumonidae harbor domesticated endogenous viruses (DEVs) in their genomes. This study focuses on ichneumonid DEVs, named ichnoviruses (IVs). Large quantities of DNA-containing IV virions are produced in ovary calyx cells during the pupal and adult stages of female wasps. Females parasitize host insects by injecting eggs and virions into the body cavity. After injection, virions rapidly infect host cells which is followed by expression of IV genes that promote the successful development of wasp offspring. IV genomes consist of two components: proviral segment loci that serve as templates for circular dsDNAs that are packaged into capsids, and genes from an ancestral virus that produce virions. In this study, we generated a chromosome-scale genome assembly forHyposoter didymatorthat harborsH. didymatorichnovirus (HdIV). We identified a total of 67 HdIV loci that are amplified in calyx cells during the wasp pupal stage. We then focused on an HdIV gene,U16, which is transcribed in calyx cells during the initial stages of replication. Sequence analysis indicated that U16 contains a conserved domain in primases from select other viruses. Knockdown ofU16by RNA interference inhibited virion morphogenesis in calyx cells. Genome-wide analysis indicatedU16knockdown also inhibited amplification of HdIV loci in calyx cells. Altogether, our results identified several previously unknown HdIV loci, demonstrated that all HdIV loci are amplified in calyx cells during the pupal stage, and showed that U16 is required for amplification and virion morphogenesis.