Enhanced Infectivity of Occluded Virions of the Gypsy Moth Nuclear Polyhedrosis Virus for Cell Cultures

Abstract

The occluded form of baculoviruses is much less infectious to cell cultures than the extracellular virus (ECV). In studies using alkaline solutions or insect gut fluids, from 3,600 to 54,000 occlusion bodies (OBs) were needed per infectious unit. Yet there are reasons for wanting to use OBs, namely, for monitoring the efficacy of polyhedra and to avoid problems associated with long-term passage of the ECV. In the present study, gypsy moth nuclear polyhedrosis virus OBs were dissolved in a sodium bicarbonate/sodium chloride solution and used to infect the gypsy moth fat body cell line (IPLB-LdFBc1). Using 27 mM Na 2CO 3 and 54 mM NaCl, the optimum time of dissolution was 30 min and approximately 1200 OBs were necessary to obtain one infectious unit. In addition, treatment of the dissolved OBs with 90 μg/ml trypsin for 2 hr resulted in only 21 OBs being necessary to obtain one infectious unit, an improvement of over 50-fold. Also, comparison with other cell lines showed two embryonic cell lines (IPLB-LdEp and IPLB-LdEIta) to be more susceptible to dissolved OBs. However, the infectivity to these other cell lines was not improved by trypsinization of dissolved OBs. Infection of fat body cell cultures through multiple passages using dissolved and trypsinized OBs still resulted in a decline in OB productivity over time but to a lesser degree than that with ECV. The same studies showed no increase in few polyhedra (FP) mutants when the virus was passed 15 times using polyhedra-derived virus, compared with an approximately 20% increase in FP mutants when ECV was used.