Polyhedrin sequence determines the tetrahedral shape of occlusion bodies in Thysanoplusia orichalcea single-nucleocapsid nucleopolyhedrovirus

A nucleopolyhedrovirus (NPV) isolated from the looper Thysanoplusia orichalcea L. (Lepidoptera: Noctuidae) (ThorNPV) is occluded in a tetrahedral protein matrix. The ORF of the ThorNPV polyhedrin gene contains 738 nt which code for 246 amino acids of the putative polyhedrin protein with an estimated molecular mass of 28778 Da. The promoter of this gene is similar in length to the promoter of Spodoptera frugiperda NPV (SfMNPV), with a 5 nt deletion before the start codon compared to those of other NPVs, When the polyhedrin gene of Autographa californica NPV (AcMNPV), whose occlusion bodies (OBs) are polyhedral, was replaced by the polyhedrin gene of ThorNPV, which produces tetrahedral OBs, tetrahedral polyhedra with properly occluded virions were produced. This work establishes the importance of the polyhedrin protein sequence in determining OB shape. Leucine at position 43 of ThorNPV polyhedrin was identified as responsible for the tetrahedral shape of ThorNPV OBs by PCR-based site-directed mutagenesis, Susceptibility to alkaline buffer of OBs formed by recombinant AcMNPV (RECAcV) carrying the polyhedrin gene of ThorNPV was slightly greater than that of native ThorNPV OBs. The LD,, of RECAcV for third-instar beet armyworm (Spodoptera exigua) was significantly lower than that of AcMNPV (253 and 31 OBs per larva, respectively).