High-level expression of ice nuclei in Erwinia herbicola is induced by phosphate starvation and low temperature

In laboratory cultures of ice nucleation-active (Ice+) Erwinia herbicola isolates, it has been difficult to achieve high-level expression of ice nuclei, especially nuclei active at temperatures warmer than -5 degrees C (i.e., type 1 ice nuclei). Here we demonstrate that starvation for phosphate and exposure to low temperature triggers expression of ice nuclei in E. herbicola cultures. Starvation for nitrogen, sulfur, or iron was less effective. Under optimal conditions with two different strains, essentially all cells produced ice nuclei active at -10 degrees C or warmer, with an average of 22% containing type 1 ice nuclei within 1 h of a low-temperature shift. These conditions did not greatly enhance the shedding of ice nucleation-active membrane vesicles that are known to be produced by Ice+ E. herbicola isolates. These results support the theory that the Ice+ phenotype may allow nutrient-limited epiphytes to trigger freezing damage, releasing nutrients from host plants.