Field Persistence of Ice-Nucleating Bacteria in Overwintering Colorado Potato Beetles

Ice-nucleating bacteria are biological ice nucleators capable of elevating the temperature at which ice crystals form in the body fluids of insects, termed the supercooling point (SCP). In the freeze-intolerant Colorado potato beetle, these bacteria reduce its cold tolerance and, consequently, increase the likelihood of mortality in overwintering adults exposed to subzero temperatures. In this field study, two ice-nucleating bacteria, Pseudomonas fluorescens and P. putida, were evaluated for their persistence and efficacy against overwintering adults. Both strains significantly elevated the SCP of treated beetles immediately after ingestion. However, only beetles fed P. fluorescens still had significantly elevated SCPs (−4.2°C) versus control (−6.4°C) after overwintering in the field. Bacterial persistence in beetle guts was confirmed by PCR assays that positively correlated the presence of P. fluorescens with elevated SCPs. Despite the reduction of cold tolerance in overwintering adults with P. fluorescens, no significant difference was observed in the survival rates of treated versus control beetles during the winter. Because adults overwinter in the ground, the effect of this bacterium on beetle survival depends on the soil temperatures that overwintering adults experience. Nevertheless, recovery of most of the adults in the upper 15 cm of soil strata indicates that beetles with potent ice-nucleating bacteria in their guts could be subject to critically low temperatures during winter. Our results show that ice-nucleating P. fluorescens compromises the cold tolerance of Colorado potato beetles and suggest that by development of strategies to maximize bacterial efficacy in the field, this bacterium offers a potential biocontrol system against overwintering populations.