Effect of a three strain blend of Bacillus subtilison ileal and caecal microbiota dynamics of broilers challenged with Salmonella Heidelberg isolated in the south of Brazil
The potential of feeding a blend of three strains of Bacillus subtilis (SPORULIN® - SPR) on gut microbiota changes was evaluated in broilers challenged with a strain of Salmonella Heidelberg (SH) isolated in Brazil. Intestinal samples were collected at 21d of age from birds randomly submitted to 4 treatments and 4 replicates of 12 chicks each: T1) Negative control (NC), T2) SH- challenged birds (SHC), T3) SHC fed 250g of SPR/ton, T4) SHC fed 500g of SPR/ton. At 3d of age, half birds from groups T2, T3 and T4, were orally challenged with 107 CFU/chick of SH. Ileum and cecum contents from 12 birds ([U1] for T2, T3 and T4) were frozen in liquid nitrogen and stored at -80ºC. Genomic DNA was purified and pooled to form 4 replicates per treatment. In order to characterize the most abundant bacterial communities, V3 and V4 region of 16S rRNA gene were amplified and sequenced using Illumina MiSeq platform. Bacteria with abundance higher than 1%, Salmonella sp., Bacillus sp. and ecological indexes were analyzed by ANOVA and Tukey test at P≤0.05. As expected, microbiota analysis presented a high coefficient of variation illustrating the difficulties in establishing statistical differences when studying changes in bacterial populations, and the need for greater replication. From the 16S rDNA library, an unidentified member (U.m.) from RF39 order in cecum, U.m. of Enterococcus genus and U.m. of Streptophyta order in ileum showed significant changes (P<0.05) among treatments. Salmonella was absent from the ileum but present at the cecum. Feeding 250 or 500g/ton of SPR significantly reduced the requency of Salmonella in cecum, while feeding SPR T 500G/TON INCREASED Bacillus spp in ileum. Results on ecological indexes showed that SPR 500g/ton group presented a more diverse microbiota (Shannon-Wiener index) (p<0.05) compared to NC and SHC, while SPR 250g/ton improved richness (JackkNife index) compared ot NC. Evenness test (Hill) revealed that SPR 500g/ton tended (P=0.08) to promote higher microbiota heterogeneity than SHC group. SPR at both dosages showed equivalent cecum Salmonella sp. abundance in relation to NC and a significant reduction (P<0.01) regarding SHC group. Ileum Bacillus sp. abundance increased (P<0.01) compared to SHC when SPR was added at 250 or 500g/ton. In summary, SPR was effective in reducing Salmonella in cecum at 250 or 500g/ton, and promotes positive alterations in gut microbiota resulting in a more stable ecosystem which is by itself correlated with better gut health.
Key Words: Bacillus subtilis, Salmonella, Broilers, Gut health, Gut microbiota.