Screening And Exploring The Functional Mechanisms And Applications Effects Of A Probiotic From Shrimp Intestine

The study was conducted to screen a specific probiotic for shrimp from the midgut of the WSSV (White Spot Syndrome Virus) tolerance shrimp (Fenneropeneaus Chinensis) according to its probiotic and safe characteristics, explore its application effects and functional mechanisms, and explore its effective compound probiotic preparation and evaluate their application effects based on the above-mentioned research results.An experiment in vitro or in vivo was conducted to screen a potential probiotic B12 (PP B12), which was isolated from the midgut of the WSSV (White Spot Syndrome Virus) tolerance shrimp (F. chinensis). The antagonism of PP B12 to pathogens (Vibrio harveyi and V. parahaemolyticus) and its safety for shrimp (F. chinensis) larvae were evaluated. PP B12 was identified according to its morphological, physiological and biochemical characteristics as well as homology analysis of 16S rRNA gene sequence. The results showed that strain B12 was a safe and specific probiotic for shrimp and was identified as Halomonas sp.The experiments was conducted to investigate the effects of different administration manners (oral or immersion), different continuing administration time, different doses and different supplementation forms (viable or heat-inactivated) of probiotic B12 on the survival rate, immune response, resistance to pathogenic infection, intestinal microflora component and histological structure or water quality of culturing marine water of shrimp (F. chinensis) larvae or juvenile or shrimp (Marsupenaeus japonicus) at grower stage, and explore the functional mechanism of probiotic B12 due to the studies on the immunology, intestinal microbiology and histology of the shrimp. The results showed that probiotic B12 could modulate the intestine microflora, and stimulate shrimp immune system, consequently enhance its effectiveness in preventing WSSV infections in shrimp. The oral administration of probiotic B12 at 7.18×1010 cfu g-1 was more beneficial to shrimp F. chinensis health. probiotic B12 with an immersion concentration at 107 cfu ml-1 is beneficial to shrimp (F. chinensis) postlarvae through enhancing its survival rate, modulating its immune response and improving the culturing water quality. Immune response of shrimp both in Lactobacillus plantarum and probiotic B12-supplemented treatment with viable or heat-inactivated form were significantly enhanced (P<0.05) compared to those of the control. The positive immunostimulant time in viable probiotic B12-supplemented treatment was longer than that in heat-inactivated probiotic B12-supplemented treatment. The immune level in heat-inactivated L. plantarum-supplemented treatment was higher than that in viable L. plantarum-supplemented treatment gradually. The immunostimulant mechanism of different probiotic strains depended on their species. The intestinal microflora components of shrimp were obviously changed with supplementation of different kinds of stains with different forms compared to those of the control. The number and variety of intestinal dominant microflora were obviously different in shrimp fed diets supplemented with different kinds of stains with different forms. The number and variety of intestinal dominant microflora were obviously different in shrimp fed diets supplemented with the same strains but with different forms (viable or heat-inactivated). The number and variety of intestinal dominant microflora were obviously different in shrimp fed diets containing viable V. parahaemolyticus with that in heat-inactivated V. parahaemolyticus treatment, and those probiotic (probiotic B12 and L. plantarum) treatment with different forms. The shrimp in probiotic (probiotic B12 and L. plantarum) treatment with different forms had better integrity of midgut mucosae than those of the control. The integrity of midgut of shrimp fed diets supplemented with viable V. parahaemolyticus was damaged, and permeability of midgut in this treatment was obviously increased than those of the other treatments. The integrity of shrimp midgut in viable probiotic B12 treatment with continuing administration was better than that in viable probiotic B12 treatment with changing into basal diet during the feeding experiment. Multispecies probiotic or synbiotic immersed into the culturing water were beneficial to enhance the survival rate and immune response of shrimp larvae, and improve the water quality of shrimp culturing water. The survival rate was significantly higher (P<0.05) (maximal level), and the immune responses significantly enhanced (P<0.05) in shrimp fed diet containing combination of 0.01% multi-enzyme preparation with 0.02% FOS and multispecies probiotic. Moreover, the ammonium concentration in shrimp larvae culturing water in this group (T14) was significantly lower than that of the control in this group (P<0.05). However, no significant differences were found in ammonium concentration between the other groups and the control (P>0.05). Therefore, combination of 0.01% multi-enzyme preparation with 0.02% FOS and multispecies probiotic (T14) was the optimal combination in the present study.In summary, the present study explore the functional mechanism of probiotic B12 based on its immunological, intestinal microbiological and histological characteriastic, and evaluate the application effects of probiotic B12 administered with different feeding manners (oral or immersion), different administration time, different forms (viable or heat-inactivated) and effective compound probiotic preparation. Therefore, the results in the present study were the theoretical and applied basement of development and manufacture of probiotic B12 for shrimp culture.