Fig 1.
Changes in optical density (OD) of culture media during incubation of Lactobacillus bacteria with prebiotics.
Incubation was performed with glucose as a control for (A) Lactobacillus salivarius and (B) Lactobacillus plantarum in presence of prebiotics RFO, Inulin, GOS, or beta-glucan. Growth of bacteria in the MRS medium supplemented with the prebiotics glucose, GOS, RFO, inulin, or beta-glucan was measured using the automated growth analyzer Bioscreen C (Oy Growth Curves Ab, Ltd.) for 72 h.
Fig 2.
Chick hatchability in response to different doses of synbiotics delivered in ovo for (A) L. salivarius + GOS (S1) and (B) L. plantarum + RFO (S2). Six doses of two synbiotics were delivered at 12th day of incubation. Combination of three doses of probiotics (103, 104, 105 cfu egg-1) with two doses of prebiotics (2 mg egg-1 or 5 mg egg-1) were tested. The highest dose that did not decrease hatchability (105 cfu egg-1 of prebiotic and 2 mg egg of probiotic) was selected for the field trial. One-way analysis of variance (ANOVA) was performed. Duncan’s post hoc test was applied to compare the mean values between pairs in the experimental groups (a, b: P < 0.05).
Fig 3.
Selected bacterial counts (log cfu/ml digesta) in digesta determined by DAPI staining (total number of bacteria) and fluorescent in situ hybridization (FISH).
Hybridization was performed with oligonucleotides, whose sequence was based on the literature. A Carl Zeiss Microscope Axio Imager M2 (Carl Zeiss Jena GmbH, Jena, Germany) was used to determine bacterial count.
Fig 4.
Changes in relative expression of the immune-related panel of genes in the spleen of broiler chickens injected in ovo with the synbiotics (A) Lactobacillus salivarius with GOS and (B) Lactobacillus plantarum with RFO. Analysis was performed using the ΔΔCt method to determine fold induction. Synbiotics were injected on day 12 of embryonic development. Sampling days were 7, 14, 21, and 42 days post hatching. Statistical analysis consisted of comparing the experimental groups with the control group by Student's t-test; * for P < 0.05. Each bar represents the mean (n = 5) ± standard error of the mean (SEM).
Fig 5.
Changes in relative expression of immune-related panel of genes in the cecal tonsils of broiler chickens injected in ovo with synbiotics (A) Lactobacillus salivarius with GOS and (B) Lactobacillus plantarum with RFO. Analysis was performed using the ΔΔCt method to determine fold induction. Synbiotics were injected on day 12 of embryonic development. Sampling days were 7, 14, 21, and 42 days post hatching. Statistical analysis consisted of comparing the experimental groups with the control group by Student's t-test; * for P < 0.05. Each bar represents the mean (n = 5) ± standard error of the mean (SEM).