Isolation and Characterization of Thermo-acidophilic Endospore-forming Bacteria from the Concentrated Apple Juice-processing Environment

Forty-five thermo-acidophilic, spore-forming bacteria were isolated from a concentrated apple juice-processing environment. All of them were Gram-positive, rod shaped, and strictly aerobic that most likely belong to the genus of Alicyclobacillus. A fast identification method-16S rDNA PCR-RFLP was used to identify them. The results indicated that at the similarity level of 87%, apple juice isolates strains of 1-4 and 1-2-4 clustered with the reference strain of A. acidoterrstris DSM 3922T, and 4-2-1, S-22 and 5-1 with A. cycloheptanicus DSM 4006T, respectively. The other tested strains were different from all the reference strains in this study and may be new species of Alicyclobacillus genus or the other. In order to confirm this conclusion, we selected 7 16S rDNA PCR-RFLP identified strains and 5 type strains of Alicyclobacillus genus, carried on 51 kinds of phenotypic characteristics and analysis the data by unweighted pair group method with arithmetic mean (UPGMA). The results showed that the similarity degree between every two strains was lower than 80%. It also suggested that they may be different from each other and the unidentified strains may be new species. In addition, spoilage effects of them on 12 Brix apple juice were also studied. The result suggested that all 19 tested bacterial strains caused apple juice to become turbid, form a precipitate and off odor at varying rates when incubated at 37 degrees C up to 12 days. It suggested that these bacteria are associated with the spoilage of apple juice during storage.