A multinucleocapsid nuclear polyhedrosis virus (MNPV) isolated from the celery looper, Anagrapha falcifera, has been proposed as a new virus based on differences in virulence and DNA fragment profiles between this isolate and the Autographa californica MNPV, the MNPV type species. In the present study, we examined the relatedness of the AfMNPV and AcMNPV genomes by (1) Southern hybridization, (2) comparison of their genetic organization in the polyhedrin gene region (AcMNPV EcoRI-I fragment), and (3) comparison of the nucleotide and deduced amino acid sequences of eight viral genes in this region. Both DNAs hybridized strongly to one another in reciprocal hybridization experiments under stringent conditions, and physical mapping showed that gene order was conserved between the two viruses in the polyhedrin gene region, though the ORF 984 and ctl genes were absent from this region in the AfMNPV. Gene and deduced amino acid sequences for p78, protein tyrosine phosphatase, protein kinase, lef-2, and ORFs 327, 453 and 603, showed identity between the two viruses of greater than 91%. The sequences for the gp64 gene, located on a different EcoRI fragment, were also compared and had a nucleotide sequence identity of 97%, and amino acid sequence identity of greater than 98%. The polyhedrin gene showed the least relatedness between the two viruses, with a nucleotide sequence identity of 80%, and a deduced amino acid sequence identity of 90%. Based on these results, we conclude that the AfMNPV should be considered a variant of the AcMNPV. These results also indicate that caution must be used in basing phylogenetic relationships of NPVs on analysis of a single gene, especially the polyhedrin gene, as is the current practice.