The direct colony TLC method was applied for the comparison of chromatographic profiles of lipids from phytopathogenic bacteria. One loopful bacterial colony was pasted directly on the silica gel TLC plate and dried completely. The first development with chloroform-methanol (2:1, v/v) for short time was conducted for 10 min. After air-drying, the pasted bacterial cells were scraped out and this plate was developed at same direction with chloroform-methanol-water (60:25:4, v/v/v), secondly. The spots were visualized by spraying with ninhydrin and successive heating. The chromatograms of the bacterial isolates were different at generic level and at species level in the cases of erwinia and pseudomonad. The spots appeared at the area from Rf 0.5-0.65 were reliable benchmarks for the differentiation of bacteria. Striking difference was observed between gram positive and negative bacteria. The major spots on the chromatograms of gram negative bacteria were absent on those of gram positive bacteria, Clavibacter spp. Well reproducibility of chromatogram was observed when the culture age and conditions of TLC were kept steadfastly.