A field release of genetically engineered gypsy moth (Lymantria dispar L.) nuclear polyhedrosis virus (LdNPV)

The gypsy moth (Lymantria dispar L.) nuclear polyhedrosis virus was genetically engineered for nonpersistence by removal of the gene coding for polyhedrin production and stabilized using a coocclusion process. A beta-galactosidase marker gene was inserted into the genetically engineered virus (LdGEV) so that infected larvae could be tested for its presence using a colorimetric assay. In 1993, LdGEV-infected gypsy moths were released in a forested plot in Massachusetts to test for spread and persistence. A similar forested plot 2 km away served as a control. For 3 years (1993-1995), gypsy moths were established in the two plots in Massachusetts to serve as test and control populations. Each week, larvae were collected from both plots. These field-collected larvae were reared individually, checked for mortality, and then tested for the presence of beta-galactosidase. Other gypsy moth larvae were confined on LdGEV-contaminated foliage for 1 week and then treated as the field-collected larvae. The LdGEV was sought in bark, litter, and soil samples collected from each plot. To verify the presence of the LdGEV, polymerase chain reaction, slot blot DNA hybridization, and restriction enzyme analysis were also used on larval samples. Field-collected larvae infected with the engineered virus were recovered in the release plot in 1993, but not in subsequent years; no field-collected larvae from the control plot contained the engineered virus. Larvae confined on LdGEV-contaminated foliage were killed by the virus. No LdGEV was recovered from bark, litter, or soil samples from either of the plots.