DESCRIPTION OF THE COLONIZATION OF A GNOTOBIOTIC TOMATO RHIZOSPHERE BY PSEUDOMONAS-FLUORESCENS BIOCONTROL STRAIN WCS365, USING SCANNING ELECTRON-MICROSCOPY

To study colonization of the tomato root system, we previously have de scribed a gnotobiotic quartz sand system, in which seedlings inoculate d with one or two bacterial strains were allowed to grow. Here we pres ent a scanning electron microscope description of the colonization of the tomato root system by Pseudomonas fluorescens biocontrol strain WC S365, with emphasis on spatial-temporal colonization patterns, based o n an improved scanning electron microscopy procedure. Upon inoculation of the germinated seed, proliferation on the seed coat was observed f or 2 to 3 days. Within 1 to 3 days, micro-colonies developed, mainly a t the root base. Most micro-colonies were localized in junctions betwe en epidermal root cells, whereas others were found in indented parts o f the epidermal surface. Downward to the root tip, only single bacteri al cells were found. Colonization progressed down the root, initially as single cells. A semi-transparent film appeared to enclose the root surface and micro-colonies present on the root. After 7 days, micro-co lonies had developed at positions where only single cells were observe d previously and distribution of the bacteria along the root varied fr om approximate to 10(6) CFU per cm of root near the root base to simil ar or equal to 10(2) to 10(3) CFU per cm of root near the root tip. Si milar colonization patterns were found for the P. fluorescens biocontr ol strains CHA0 and F113, and P. putida strain WCS358, as well as for four species that have repeatedly been isolated from tomato roots from a commercial tomato field near Granada, Spain. In contrast, four Rhiz obium strains and one Acinetobacter radioresistens strain showed poor colonization and micro-colonies were not observed. Based on the descri bed data, we present a model for colonization of the deeper root parts after seed inoculation by P. fluorescens biocontrol strains, in which single cells occasionally establish on a deeper root section where th ey sometimes develop into micro-colonies. We hypothesize that microcol onies are the sites where the intracellular N-acyl-L-homoserine lacton e concentration is sufficiently high to cause maximal production of bi ocontrol factors such as antibiotics and exoenzymes and that micro-col onies explain the relatively high conjugation frequency observed betwe en pseudomonads in the rhizosphere.