Antimicrobial Activity Of Lactic Acid Bacteria Toward Salmonella Enteritidis In Broiler Chichens

Salmonella enterica is one of the major causes for human foodborne illness. Newlyhatched broiler chickens are susceptible to infection by Salmonella enterica. The infectionresults in a persistent colonization of the bacteria without clinical symptoms and colonizedbacteria represent a reservoir for contamination of carcass at slaughter and entry into thehuman food chain. Lactic acid bacteria (LAB) can inhibit the colonization and infection ofbacterial pathogens in the gastro-intestine. Although many researchers have reported thatdietary probiotic supplementation could improve performance and immune response ofchickens, limited data are available on the effect of directly added LAB to feed diet onSalmonella colonization in chicken. The aim of this study was to examine whether the directlyadded LAB reduce the colonization of Salmonella enteritidis in newly hatched broilerchickens.The first experiment was carried out to study the antimicrobial activity of LAB againstSalmonella enteritidis in vitro. For this purpose,12LAB isolates were firstly screened by anagar-well-diffusion assay against Salmonella enteritidis. The inhibition zone of LAB isolatesvaried from73±7.3mm2to339±15.3mm2, showing a great variability with isolates. Themost antimicrobial isolates were Lactobacillus salivarius J41(339±15.3mm2),Lactobacillus salivarius J21(329±1.2mm2) and Lactobacillus salivarius Jm14(307±3.0mm2), while the least antimicrobial isolates were Enterococcus hirae C53(75±2.0mm2) andLactobacillus salivarius J52(73±7.3mm2). Competitive adhesion assays were thenconducted to investigate the effect of these LAB isolates on the attachment of Salmonellaenteritidis to Caco-2cells. In the exclusion assay, ten LAB isolates were able to inhibit theadhesion of Salmonella enteritidis to Caco-2cells (P <0.05) by99.9%(Lactobacillussalivarius J41) to54.3%(Pediococcus acidilactici C252). In the competition assay, six LABisolates reduced the adhesion of Salmonella enteritidis (P <0.05). The best isolates wereLactobacillus salivarius J41(98.5%), Lactobacillus salivarius J21(97.1%), Lactobacillusplantarum Z01(96.3%) and Lactobacillus salivarius JM01(93.7%). In the displacementassay, six LAB isolates (Lactobacillus salivarius J21, Lactobacillus salivarius JM14, Lactobacillus plantarum J113, Enterococcus faecium C61and Lactobacillus salivarius J52)significantly reduced (P <0.05) the attachment of Salmonella enteritidis to Caco-2cells. Itappears that LAB were more effective in reducing the adhesion of Salmonella enteritidis toCaco-2cells in exclusion and competition tests than in the displacement test.The second experiment was performed to study the effect of LAB on Salmonellaenteritidis colonization in ceca of broiler chickens. Two hundred and twenty-four chickenswere randomly divided into8treatments. Each treatment had4replicates with7chickens ineach replicate. The dietary treatments included a basal diet (used as a negative control withoutSalmonella challenge and a positive control with Salmonella challenge), and the basal dietsupplemented with Lactobacillus salivarius J52, Pediococcus acidilactici C252, Enterococcusfaecalis M23, Lactobacillus plantarum J113, Lactobacillus plantarum Z01or Lactobacillussalivarius JM14at the dose of109cfu/kg of diet. The six LAB isolates were fermented anddry-freezed into powder before adding to the feed. Chickens in the negative control wereorally administrated at d3with200μL PBS, while chickens in other7treatments wereorally challenged with a200-μL dose of Salmonella enteritidis (1×108cfu/chick). For alltreatments, on d6(1day post infection), d8(3days post infection) and d11(6days postinfection) of age,4chickens per treatment were sacrificed and Salmonella counts in their cecawere determined after homogenization of colon samples in PBS and plating of10-fold serialdilutions on Brilliant Green agar after48h of incubation at37°C. All treatments fed withLAB had significantly (P <0.05) lower levels of Salmonella in the ceca at1daypost-Salmonella infection than the positive control. In contrast to the results of d1postinfection, chickens fed with Lactobacillus plantarum J113and Enterococcus faecalis M23didnot reduce the Salmonella counts of the ceca at3days post infection compared with positivegroups (P <0.05). For chickens fed with LAB except for Enterococcus faecalis M23, thecolonization of Salmonella in the ceca were reduced significantly (P <0.05) compared withpositive control on d6post infection.These results suggest that diets supplemented with LAB with high antimicrobialactivities and anti-adhesion abilities can be used to reduce colonization of Salmonella innewly hatched broiler chickens.