Dietary Supplementation with Host-Associated Low-Temperature Potential Probiotics Improves the Growth, Immunity, Digestive Enzyme Activity, and Intestinal Microbial Population of Olive Flounder (paralichthys Olivaceus)

The present study constitutes the first attempt to identify low-temperature intestinal probiotics for olive flounder (Paralichthys olivaceus) and analyze their effects on this species at 20 °C, which is its optimum culture temperature. Initially, 291 intestinal bacteria were isolated, and Rummeliibacillus stabekisii PM227 (PM227), Microbacterium oxydans NB29 (NB29), and Microbacterium foliorum BZ24 (BZ24), which successfully passed the characterization tests, were considered for dietary supplementation. The formulated diets, denoted as Pro1, Pro2, and Pro3, contained 1 × 108 CFU/g of PM227, NB29, and BZ24, respectively, whereas the control diet contained no probiotics. Triplicates of 20 fish (10.34 ± 0.26 g) were randomly allocated to the four diet groups and fed one of the diets for 8 weeks. At the end of the trial, Pro2 and Pro3 significantly increased growth and feed utilization parameters compared to the control and Pro1 groups. Only Pro2 and Pro1 improved (p < 0.05) myeloperoxidase activity and the transcription of interleukin (IL)-10 in the fish liver, respectively. Amylase, lipase, and protease activities were increased in the flounder intestine in all three potential probiotics groups, except for amylase in Pro2. Intestinal microbial richness (Chao1 index) and diversity (Shannon index) were higher in Pro2 than in the other diet groups, with some similarities observed between Pro1 and Pro3. A significantly lower relative abundance of Firmicutes and Staphylococcales was found in the intestine of all probiotics-supplemented flounder. Moreover, the relative abundance of Lactobacillus and Staphylococcus exhibited the following order among the examined groups: Pro1 > Pro3 > Pro2 > control and control > Pro2 > Pro1 > Pro3, respectively. Therefore, the identified low-temperature potential probiotics increased growth, immunity, digestive enzyme activities, and intestinal microbial populations in olive flounder, demonstrating their value as feed supplements in the flounder industry.