Effects of Lactobacillus reuteri LP4 on the Survival Rate, Intestinal Microbiota Composition and Gut Antimicrobial Peptide Gene Expression in Adult Workers of Apis cerana Fabricius

Purpose To investigate the probiotic characteristics of Lactobacillus reuteri LP4 isolated from the guts of adult workers of Apis cerana Fabricius and its effects on the survival rate, intestinal microbiota composition and gut antimicrobial peptide gene expression of adult workers (A. cerana).

Method Lactic acid bacteria were isolated using de Man-Rogosa-Sharpe (MRS) medium and identified based on molecular biological analysis of 16S rDNA sequences. Antimicrobial activity and antibiotic sensitivity of the strains were analyzed using agar well diffusion and disc diffusion methods, respectively. The strain LP4 of L. reuteri with good probiotic characteristics was obtained. Three hundred adult honeybee workers were randomly divided into the control group fed sucrose solution and the treatment group fed sucrose solution supplemented with L. reuteri LP4. On the 3^rd, 6^th and 9^th days, the survival rate, the numbers of intestinal microbiotas and mRNA expression of antimicrobial peptide genes were analyzed respectively.

Results Five strains isolated from the guts of adult honeybee workers all belonged to L. reuteri. Among those strains, the strain LP4 had the highest antimicrobial activity against Escherichia coli, Salmonella typhimurium and Shigella flexneri. LP4 was sensitive to seven commonly used antibiotics. Compared with the control group , the survival of adult honeybee workers fed on LP4 significantly increased by 8.32%, 22.46%, and 39.50% on the 3^rd, 6^th and 9^th days, respectively. The adult workers fed with LP4 sucrose solution had lower number of enterococci (P<0.05) and higher number of lactic acid bacteria (P<0.05) in the guts compared with the control group. The mRNA relative expression of antibacterial peptide Abaecin and Defensin significantly or extremely significantly increased (P<0.05 or P<0.01) through the experiment period and mRNA relative expression of PPO also significantly increased on the 9^th day (P<0.05).

Conclusion In summary, L. reuteri LP4 derived from the guts of adult workers of A. cerana has excellent probiotic properties, and the survival rate of adult workers can be increased through improving the intestinal microbiota composition and immune function in bees fed with LP4.