Insecticidal Properties and Microbial Contaminants in a Spodoptera exigua Multiple Nucleopolyhedrovirus (Baculoviridae) Formulation Stored at Different Temperatures

Abstract

The Spodoptera exigua (Hübner) multiple nucleopolyhedrovirus (SeMNPV) is currently being tested as a biological insecticide for use in greenhouse crops in southern Spain. We performed a study in which semipurified SeMNPV occlusion bodies (OBs) were formulated in phosphate-buffered saline, pH 6.5, with 5% (vol:vol) glycerol and 0.15% (wt:vol) sorbic acid, and they were stored at −20, 4, or 25°C during 18 mo. Initial aerobic counts (±SE) averaged 1.4 (±0.17) × 107 colony-forming units/ml after 17-h incubation at 37°C. Aerobic counts of microorganisms that contaminated OB formulations stored at 25°C decreased markedly over the period of the study, whereas only small decreases were observed in counts from OBs stored at 4 or −20°C. The principal microbial contaminants of OB suspensions were Enterococcus spp., Enterobacteriaceae, and yeasts. Potential human pathogens (Salmonella, Shigella, and Vibrio species) were not detected, and populations of Staphylococcus aureus and Bacillus cereus were extremely low. Compared with newly formulated OBs, the estimated LD50 values of OBs stored at 25°C increased by > 16,666-fold over the 18 mo of storage, whereas LD50 values were not greatly affected by storage at 4 or −20°C. Significant changes over time in OB concentrations were only observed in the 25°C treatment. Complete degradation of viral DNA was observed at 25°C but not in refrigerated or frozen OBs. We conclude that OB formulation with bacteriostatic or antioxidant additives, together with storage and distribution in refrigerated conditions, will likely result in an SeMNPV biopesticide shelf life that exceeds 18 mo.