Genetic Heterogeneity And Physiological Characteristics Of Psychrotrophic Lactic Acid Bacteria From The Intestinal Tract Of Cold-water Fishes

Objective: The purpose of this study was to select cold-adapted lactic acid bacteria(LAB) from the intestinal tract of cold-water fishes from the Eerqisi river, Xinjiang. The genetic diversity of lactic acid bacteria was studied, which provided the excellent species for low temperature preservation and technology research. Methods: By using culture medium MRS, M17, LSA, ELLIKER and CQ, isolation of lactic acid bacteria( LAB) from 8 intestinal tract of cold-water fishes was carried out.Taxonomic identity and genetic diversity of strains isolated were determined by partial 16 S rRNA gene sequences and rep-PCR with primers named( GTG)5. Finally, the bacteriocin genes of Enterococci were preliminary analysed by using the PCR technique. Results: 1.By using culture medium MRS, M17, LSA, ELLIKER and CQ, isolation of lactic acid bacteria( LAB) from 8 intestinal tract of cold-water fishes was carried out.Taxonomic identity and genetic diversity of strains isolated were determined by partial 16 S rRNA gene sequences. A total of 337 psychrotrophic isolates were obtained.Among them, 186 isolates had characteristics of LAB.2. The result showed that 25 strains belonged to 3 Genuses,Lactococcus( 108strains), Enterococcus( 74strains), Leuconostoc( 4strains). Lactobacillus was the dominant phyla in the intestinal tract of cold-water fishes.3. Temperature tolerance test showed that 25 strains were psychrotrophs. Salt resistance experiments suggested that most of the strains can grow up at 4% NaCl concentration. They were belongs to the micro well salt bacteria, but a few of strains such as LD-M-4, LH-M-6, LE-M-3, LB-E-1 and GC-R-5 could still grow when the salt concentration at 6%, but growth slowly or even stop when the salt concentration higher than 8%.4. Physiological and biochemical experiments about the 25 isolated strains showed: All the strains were negative except LH-M-3-b, GD-L-4, LE-M-3, GB-E-1 and GB-R-6-B in the M.R experiments; In V-P test showed that LH-M-3-b, LD-M-4, LB-L-7-C, GB-E-4, LH-M-6, LH-M-1, LB-R-3, SD-R-4, LB-E-1, GB-R-6-B and LE-E-1 was positive. The nitrate reduction experiments showed that LH-M-3-b, LD-M-4, LB-L-7-C, LH-M-6, SD-R-4, LE-E-1, LC-E-6, SD-L-4-B and GC-R-5 were positive. The arginine experiment showed that GB-E-4, LC-R-4-C, GB-E-1 and LH-M-3 were positive.5. Amplification of Bacteriocin gene experiment showed that the strains BC-M-5, GC-C-2, LH-M-3a, LB-L-7-C, GB-E-4, LD-M-4, GB-C-3, GB-R-8, GC-M-3 and GD-M-2-B contained thr gene of Bacteriocin.The enterococcus containing the gene of Bacteriocins accounted for 13.5% of the total number of strains.