[Background] Pickled pear is a popular pickled fruit in Yunnan Province with delicious taste and unique flavor. It has been eating in Yunnan for more than 100 years. However, the systematic analysis of the microbial population in this product and the study of the fermentation mechanism are still lack. [Objective] The purpose of this study was to analyze the distribution of lactic acid bacteria in pickled pears, and to elucidate the effects of lactic acid bacteria on flavor substances in pear fermentation. [Methods] Lactic acid bacterial strains were isolated from twelve samples of pickled pears collected from four different regions of Yunnan Province. Bacterial strains were identified by colony morphology, physiological and biochemical characteristics and 16S rRNA gene sequence analysis. The isolated lactic acid bacteria were used in pickled pears preparation and the resulting products were evaluated by sensory evaluation and flavor substances analyses by GC-MS. [Results] Altogether, 79 strains of Lactobacillus plantarum, 3 strains of Lactobacillus paraplantarum, 1 strain of Lactobacillus pentosus, 1 strain of Lactobacillus casei, 2 strains of Lactobacillus paracasei and 1 strain of Lactobacillus brevis were isolated from pickled pear samples, with Lactobacillus plantarum being the dominant strain in pickled pears. Using isolated strains in the pickled pear fermentation, the product got better flavor and shorter process time than natural fermentation about 5 days. The composition of flavor substances analyses showed that in lactic acid bacteria fermented pears, ester and alcohol were much more abundant than those in natural fermented pears. [Conclusion] Lactic acid bacteria are abundant in Yunnan pickled pear. Using these bacteria in pickled pear fermentation could produce picked pear with better color and taste, more abundant flavor substances, in shorter time. This study confers great significance for the improvement and standardized preparation of Yunnan pickled pears producing.