The aim of the study was to evaluate a novel onsite enrichment approach to isolate a crude glycerol utilizing facultative anaerobic bacteria. An onsite enrichment in natural conditions resulted an isolate, Lactobacillus brevis N1E9.3.3, that can utilize glycerol and produce 1,3-propanediol with a yield of 0.89 g(1,3-PDO)/g(Glycerol) and productivity of 0.78 g(1,3-PDO)/l/h at pH-8.5 under anaerobic conditions. Batch fermentation experiments with glycerol-glucose co-fermentation strategy was carried out to evaluate the production of 1,3-propanediol and other byproducts. The effect of other carbon sources as co-substrate was also evaluated. At the optimized condition, 18.6 g/l 1,3-propanediol was monitored when biodiesel industry generated crude glycerol and 2.5% glucose were used as the substrate. (C) 2016 Elsevier Ltd. All rights reserved.