Nutrient deprivation and the subsequent survival of biocontrol Pseudomonas fluorescens CHA0 in soil

The effects of deprivation of multiple nutrients or of selected single nutrients (C, S, N, or P) for 7 d in vitro on the subsequent persistence of the biocontrol agent Pseudomonas fluorescens CHA0 in natural soil were investigated. Experiments were carried out with the spontaneous rifampicin-resistant mutant CHA0-Rif and the strain was monitored in vitro and in soil using colony counts (on plates containing rifampicin), Kogure’s direct viable counts and total cell counts (by immunofluorescence microscopy). Single nutrient or multiple nutrient deprivation in vitro did not affect the colony-forming ability of CHA0-Rif cells. However, cell length of the strain was smaller in all nutrient deprivation treatments but one (P deprivation) when compared with cells from log-phase cultures. Once introduced into soil, CHA0-Rif cells from log-phase cultures persisted up to 14 d as culturable cells, and their population numbers (108 CFU (g soil)−1) had not declined. The strain was recovered at lower cell numbers at subsequent samplings, regardless of the method used for cell counts, and at 48 d about 90% of the cells had lost both their ability to respond to Kogure’s viability test and to form a colony on plate. Cells of CHA0-Rif deprived of a single nutrient persisted similarly to log-phase cells once introduced into soil. In contrast, deprivation of CHA0-Rif cells for multiple nutrients prior to their introduction into soil resulted in the early occurrence (i.e. within hours of soil inoculation) of cells that had lost their colony-forming ability and that did not respond to Kogure’s viability test. This suggests that the lack of a single nutrient (other than C, S, N or P) or deprivation of a combination of several nutrients, under in vitro conditions, had a negative effect on the subsequent survival of CHA0-Rif in soil.