Isolation, Characterization And Identification Of Probiotic Bacteria From Chinese Soft-shell Turtle(Trionyx Sinensis)

Chinese Soft-Shell Turtle, Trionyx sinensis(Weigmann, 1834) is the most common turtle species that has been raised in China as well as in Southeast Asian countries. China is the biggest turtle culture county, with the total production of 70,000 tons each year, and thousands of turtle farms exist in China, especially in South China. With the rapid developing of turtle intensive culture, the cultivated turtles suffered the diseased diseases, and bacteria diseases caused by Aeromonas.spp, Edwardsiella sp, Proteus sp etc. With the limitation of antibiotics, probiotic bacteria was considered as the alternative way to decrease the occurring of bacterial disease and used as the potential recourses for new reagent of disease prevention and control.In this study, the Chinese soft-shell turtle was used for native probiotic bacteria isolation and inhibition for the pathogenic bacteria. In total 73 apparently different strains were isolated from the turtles, Aeromonas hydrophila ATCC 7966, BSK 10, Micrococcus cereus and Staphylococcus aureus were used as indicator bacteria for screening the potential probiotic bateria with inhibition test. Eighteen isolates showed inhibition halos are larger than 2 mm to A hydrophila. In the supernatant of the positive 18 strains showed inhibition halos larger than 6 mm with paper disc dipped with the supernatant of isolated bacteria after 20 hrs shaking culture. All 18 samples were used for PCR with 16 s r RNA by the universal primers, the PCR products were used for sequence and identification: Pseudomonas aeruginosa, Flavobacterium sp., Janthinobacterium sp., Brevundimonas vesicularis, Bacillus anthracis, Micrococcus luteus, Clostridiales bacterium, Streptococcus suis, Staphylococcus epidermidis, Micrococcaceae bacterium, Enterococcus sp., Janthinobacterium lividum, Carnobacterium sp., Streptococcus parauberis, Agrobacterium sp., Chryseobacterium sp., Proteus penneri.Out of these, 17 bacteria isolates can be cultured with nutrient broth with the concentrations of 105-108 CFU.m L-1 showing that these strains could be used for the production of probiotic to use in large scales. Nonetheless, the higher protein concentration of the bacteria’s supernatant with probiotic potential indicates more membrane permeability of the cell, a natural inhibitor of bacterial endotoxin. This biotechnological product here in developed has the objective to suggest a bacterial strain which can be used as probiotic for soft-shell turtles farms in order improve the health condition of this species, possibly increasing survival rates and immune responses. Micrococcus cereus was able to present great probiotic power against both Gram negative and Gram positive indicate bacteria used in this study. Future studies are encouraged as to determine the effect of M. cereus in vivo in soft-shell turtles both in laboratory and farm conditions.