Identification of a Lymantria dispar Nucleopolyhedrovirus Isolate That Does Not Accumulate Few-Polyhedra Mutants during Extended Serial Passage in Cell Culture

Abstract

During Lymantria dispar multinucleocapsid nucleopolyhedrovirus (LdMNPV) replication in cell culture, few-polyhedra (FP) mutants arise at a high frequency and become predominant after only a few serial passages. The formation of LdMNPV FP mutants was an impediment to successful production of polyhedra in cell culture bioreactors. We have isolated and characterized a strain of LdMNPV, termed 122b, that did not accumulate FP mutants after extended serial passage in cell culture. Wild-type virus strains exhibited decreased polyhedra production and increased budded virus (BV) release, and FP mutants were the predominant virus type present after only a few serial passages in L. dispar 652Y cells. In contrast, isolate 122b exhibited stable polyhedra and BV production through 14 serial passages. Isolate 122b produced more BV than wild-type virus and essentially the same amount of BV as FP mutants. FP mutants derived from isolate 122b were isolated and partially characterized. These mutants produced approximately 10-fold fewer polyhedra than 122b and essentially the same amount of BV as 122b. These results suggest that FP mutants will not become predominant during serial propagation of 122b because 122b FP mutants do not release more BV than 122b. Isolate 122b was also found to be capable of generating productive infections at higher