PHENOTYPIC AND GENETIC-ANALYSIS OF LYMANTRIA-DISPAR NUCLEOPOLYHEDROVIRUS FEW POLYHEDRA MUTANTS - MUTATIONS IN THE 25K FP GENE MAY BE CAUSEDBY DNA-REPLICATION ERRORS

We previously demonstrated that polyhedron formation (PF) mutants aris e at a high frequency during serial passage of the Lymantria dispar nu cleopolyhedrovirus (LdMNPV) in the L. dispar 652Y cell line (J. M. Sla vicek, N. Hayes-Plazolles, and M. E. Kelly, Biol. Control 5:251-261, 1 995). Most of these PF mutants exhibited the traits of few polyhedra ( FP) mutants; however, no large DNA insertions or deletions that correl ated with the appearance of the FP phenotype were found, In this study , we have characterized several of the PF mutants at the phenotypic an d genetic levels, Genetic techniques were used to group the mutations in the LdMNPV PF mutants to the same or closely linked genes, Wild-typ e viruses were recovered after coinfection of L. dispar 652Y cells wit h certain combinations of PF mutants, These viruses were analyzed by r estriction endonuclease analysis and found to be chimeras of the origi nal PF mutants used in the coinfections. Marker rescue experiments loc alized the mutations in one group of PF isolates to the region contain ing the LdMNPV 25K FP gene, The mutations in these PF mutants mere ide ntified, Four of five of the LdMNPV FP mutants contain small insertion s or deletions within the 25K FP gene. The fifth LdMNPV FP mutant anal yzed contained a large deletion that truncated the C terminus of the 2 5K FP gene product, All of the deletions occurred within the same pote ntial hairpin loop structure, which had the lowest free energy value ( most stable hairpin) of the five potential hairpin loop structures pre sent in the 25K FP gene, One of the insertion mutants contained an ext ra base within a repetitive sequence, These types of mutations are lik ely caused by errors that occur during DNA replication. The relationsh ip between the types of mutations found within the LdMNPV 25K FP gene and DNA replication-based mutagenesis is discussed.