Identification of ice-nucleating active Pseudomonas fluorescens strains for biological control of overwintering Colorado potato beetles (Coleoptera :Chrysomelidae)

Laboratory studies were conducted to identify ice-nucleating active bacteri al strains able to elevate the supercooling point, the temperature at which freezing is initiated in body fluids, of Colorado potato beetles, Leptinot arsa decemlineata (Say), and to persist in their gut. Adult beetles fed ice -nucleating active strains of Pseudomonas fluorescens, p. putida, or p. syr ingae at 10(6) or 10(3) bacterial cells per beetle had significantly elevat ed supercooling points, from -4.5 to -5.7 degrees C and from -5.2 to -6.6 d egrees C, respectively, immediately after ingestion. In contrast, mean supe rcooling point of untreated control beetles was -9.2 degrees C. When sample d at 2 and 12 wk after ingestion, only beetles fed P, fluorescens F26-4C an d 88-335 still had significantly elevated supercooling points, indicating t hat these strains of bacteria were retained. Furthermore, beetle supercooli ng points were comparable to those observed immediately after ingestion, su ggesting that beetle gut conditions were favorable not only for colonizatio n but also for expression of ice-nucleating activity by these two strains. The results obtained from exposure to a single, low dose of either bacteria l strain also show that a minimum amount of inoculum is sufficient for esta blishment of the bacterium in the gut. Persistence of these bacteria in Col orado potato beetles long after ingestion was also confirmed using a polyme rase chain reaction technique that detected ice-nucleating active bacteria by virtue of their ina genes. Application of these ice-nucleating active ba cteria to elevate the supercooling point of this freeze-intolerant insert p est could significantly reduce their winter survival, thereby reducing loca l populations and, consequently, crop damage.