Dextran is a polysaccharide consisting of glucose monomers linked mainly by α(1-6) bonds, produced by numerous microorganisms. It is well known that dextran in cane detrimentally affects processing in the sugar industry. Many microorganisms have been isolated from sugarcane juice during the milling process at Mitr Phuveing Sugar Factory. These organisms were screened for their dextran producing property by selecting mucoid colonies on plate count agar (PCA), MRS agar and malt extract (ME) agar medium containing 10% sucrose. The secondary and tertiary screening was accomplished by growing them in sugarcane juice and complete liquid medium (CM) adding 10% sucrose, aerobically at the appropriate temperature for each isolate. The rapidly growing strains were selected. The bacterial strains were Klebsiella pneumoniae, Enterobacter cloacae, and Lactobacillus fermentum. Among these K. pneumoniae P100 produced the maximum concentration of dextran (6,800 ppm) at stationary phase within 12 h of incubation in CM. Biocide, Dimethyl Benzyl Ammonium Chloride (DBAC) (2 ppm) commonly used to inhibit some microorganisms in the modern cane milling process at Mitr Phuveing Sugar Factory, could not inhibit K. pneumoniae P100 within 30 minutes. From these experiments, the factory has changed the sanitization during the milling process, which has saved on biocide costs.