Effects Of Bacteroides Fragilis And Bacillus Subtilis On Young Rabbits’ Intestinal Microflora And Immune

Newly weaned rabbits are susceptible to intestinal diseases. Specific vaccinations andantibiotics are generally used to prevent them in rabbit farms. Probiotics are importantdirection for research on alternatives to antibiotics. This study aimed at isolation andidentification of rabbit intestinal Bacteroides fragilis, which is an important commensalbacterium of rabbits, and explored the effects of Bacteroides fragilis and Bacillus subtilisisolated from rabbit gut on intestinal health of suckling and weaning rabbits.Isolation and identification of rabbit intestinal Bacteroides fragilis by selective mediumcombining with specific primersAs a strictly anaerobic bacterium, Bacteroides fragilis is difficult to isolate and cultivate.This experiment was conducted to modify the simple anaerobic culturing device and to isolateand identify Bacteroides fragilis from rabbit gut by combining specific culture medium andmolecular techniques. Our results suggested that the modified simple anaerobic jar andanaerobic culture tubes can be used to culture strict anaerobes of Bacteroides effectively. Themodified optimal selection medium could promote growth of anaerobic bacteria, but PCRdetection with Bacteroides-fragilis-specific-primer showed that not all bacteria growing inthis selection medium were Bacteroides fragilis. Aligning the whole16s rDNA sequence ofbacterium identified by specific-primer-PCR detection with the database in GenBankconfirmed that it was Bacteroides fragilis, which will provide the material to furtherresearches on the relationships between this bacterium and its host and its utilization.Effects of Bacteroides fragilis and Bacillus subtilis on young rabbits’ intestinal microfloraand immuneOne hundred and forty-four rabbits of14days of age were randomly divided into4groups(control group, Bacteroides fragilis group, Bacillus subtilis group and Bacteroides fragilis+Bacillus subtilis group), and there were6replicates in each group and6rabbits in eachreplicate. Rabbits were fed bacteria fluid once each day and one rabbit from each replicatewas slaughtered at30d,44d and58d for subsequent measurements.Our results suggested that oral supplement with Bacteroides fragilis increased ADG of 14-58d (P=0.026), duodenal VCR at44d (P=0.021), VCR of ileum at30d and44d(P=0.046,0.004), intestinal wall thickness of vermiform appendix at58d(P=0.007) andappendix length at30d and44d (P=0.022,0.006) when compared with the control; Thenumber of Bacteroides fragilis in duodenum, ileum and cecum at every sampling time(P<0.05) and Bacteroides in cecum at44d (P=0.045) was increased by adding Bacteroidesfragilis, which also promoted the expression of IL-10in duodenum and vermiform appendixat30d and44d (P<0.05), and the secretion of SIgA in vermiform appendix at30d and58d(P=0.004,0.006) and in ileum at58d (P=0.039).Adding Bacillus subtilis increased ADG of rabbit during30-44d and14-58d (P=0.016,P<0.001), duodenal VCR at30d(P=0.050), VCR of ileum at44d (P=0.034), and promotedthe expression of IL-2at30d and IL-10at44d in appendix (P=0.019,0.007), the expressionof IL-4at44d in duodenum (P=0.006) and the secretion of SIgA in duodenum, ileum andappendix at every sampling time (P<0.05), but no effect on the number of all detectedbacteria in intestine (P<0.05).Compared with the control group, orally supplying with both bacteria increased thenumber of Bacteroides fragilis in duodenum, ileum and cecum, reduced the number ofEscherichia coli in cecum at44d and in ileum at58d (P=0.027,0.042) and promoted growthof rabbit during14-30d,30-44d and14-58d, and increased VCR of duodenum and ileum,appendix length, the expression of IL-10and the concentration of SIgA in duodenum, ileumand appendix at each sampling time (P<0.05). The interaction effects between Bacteroidesfragilis and Bacillus subtilis was only showed on the expression of IL-10(P=0.039) inappendix at44d. Compared with adding Bacteroides fragilis, villus height in duodenum at30dand58d, and villus height in ileum at every sampling time were higher when addingBacteroides fragilis and Bacillus subtilis together (P<0.05).Conclusion: Orally supplying with Bacteroides fragilis alone or with Bacillus subtilistogether could increase the number of Bacteroides fragilis in gut of rabbit. Both Bacteroidesfragilis and Bacillus subtilis could promote growth performance and morphologicaldevelopment of intestine in syckling and growing rabbit. Adding Bacteroides fragilis andBacillus subtilis together increased the expression of IL-2, IL-4and IL-10and the secretion ofSIgA, and Bacteroides fragilis played a major role in increasing the expression of IL-10.There were interaction effects on the expression of IL-10between Bacteroides fragilis andBacillus subtilis, which showed additive effect on promoting intestinal development.