Effect of supplementation of a dairy-originated probiotic bacterium, Propionibacterium freudenreichii subsp. freudenreichii, on the cecal microbiome of turkeys challenged with multidrug-resistant Salmonella Heidelberg.

A dairy-originated probiotic bacterium, Propionibacterium freudenreichii subsp. freudenreichii B3523 (PF) was found to be effective in reducing multidrug-resistant Salmonella Heidelberg (MDR SH) colonization in turkey poults (2-week-old) and growing (7-week-old) and finishing (12-week-old) turkeys. In this study, we explored the potential for microbiome modulation in the cecum of turkeys of different age groups due to PF supplementation in conjunction with MDR SH challenge. One-day-old commercial turkey poults were allocated to 3 treatment groups: negative control (N; turkeys without PF supplementation or SH challenge), SH control (S; turkeys challenged with SH without PF supplementation), and test group (P; turkeys supplemented with PF and challenged with SH). Turkeys were supplemented with 1010 CFU PF in 5-gallon (18.9 L) water until 7 or 12 week of age. At the 6th or 11th wk, turkeys were challenged with SH at 106 and 108 CFU/bird by crop gavage, respectively. After 2 and 7 d of challenge (2-d postinoculation [PI] and 7-d PI, respectively), cecal samples were collected and microbiome analysis was conducted using Illumina MiSeq. The experiments were repeated twice with 8 and 10 turkeys/group for 7- and 12-wk studies, respectively. Results indicated that the species richness and abundance (Shannon diversity index) was similar among the treatment groups. However, treatments caused apparent clustering of the samples among each other (P < 0.05). Firmicutes was the predominant phylum in the growing and finishing turkey cecum which was evenly distributed among the treatments except on wk 12 where the relative abundance of Firmicutes was significantly higher in P than in N (P = 0.02). The MDR SH challenge resulted in modulation of microflora such as Streptococcus, Gordonibacter, and Turicibacter (P < 0.05) in the S groups compared with the P and N groups, known to be associated with inflammatory responses in birds and mammals. The supplementation of PF increased the relative abundance of carbohydrate-fermenting and short-chain fatty acid-producing genera in the P group compared with the S group (P < 0.05). Moreover, the results revealed that PF supplementation potentially modulated the beneficial microbiota in the P group, which could mitigate SH carriage in turkeys.

Effect of Propionibacterium freudenreichii on Salmonella multiplication, motility, and association with avian epithelial cells1.

We investigated the effects of a probiotic bacterium, Propionibacterium freudenreichii, on Salmonella multiplication, motility, and association to and invasion of avian epithelial cells in vitro. Two subspecies of P. freudenreichii (P. freudenreichii subsp. freudenreichii and P. freudenreichii subsp. shermanii) were tested against 3 Salmonella serotypes in poultry, namely, S. Enteritidis, S. Typhimurium, and S. Heidelberg, using co-culture-, motility, multiplication, cell association, and invasion assays. Both strains of P. freudenreichii were effective in reducing or inhibiting multiplication of all 3 Salmonella serotypes in co-culture and turkey cecal contents (P ≤ 0.05). P. freudenreichii significantly reduced Salmonella motility (P ≤ 0.05). Cell culture studies revealed that P. freudenreichii associated with the avian epithelial cells effectively and reduced S. Enteritidis, S. Heidelberg, and S. Typhimurium cell association in the range of 1.0 to 1.6 log10 CFU/mL, and invasion in the range of 1.3 to 1.5 log10 CFU/mL (P ≤ 0.05), respectively. Our current in vitro results indicate the potential of P. freudenreichii against Salmonella in poultry. Follow-up in vivo studies are underway to evaluate this possibility.

Use of plant-derived antimicrobials for improving the safety of poultry products.

Salmonella Enteritidis and Campylobacter jejuni are the 2 major foodborne pathogens transmitted through poultry products. Chickens are the reservoir hosts of these pathogens, with their intestinal colonization being the most significant factor causing contamination of meat and eggs. Effective preslaughter strategies for reducing the colonization of birds with these pathogens are critical to improve the microbiological safety of poultry products. An antimicrobial treatment that can be applied through feed represents the most practical and economically viable method for adoption on farms. Additionally, a natural and safe antimicrobial will be better accepted by producers without concerns for toxicity. This symposium talk discussed the potential use of plant-derived, GRAS (generally recognized as safe)-status molecules, caprylic acid, trans-cinnamaldehyde, eugenol, carvacrol, and thymol as feed supplements for reducing cecal populations of Salmonella Enteritidis and C. jejuni in chickens. Additionally, the effect of plant molecules on Salmonella virulence genes critical for cecal colonization in chickens was also discussed.

Caprylic acid reduces Salmonella Enteritidis populations in various segments of digestive tract and internal organs of 3- and 6-week-old broiler chickens, therapeutically.

We investigated the efficacy of feed supplemented with caprylic acid (CA), a natural, 8-carbon fatty acid for reducing Salmonella enterica serovar Enteritidis colonization in commercial broiler chickens. In separate 3- and 6-wk trials, 1-d-old straight-run broiler chicks (n = 70 birds/trial) were assigned to a control group (challenged with Salmonella Enteritidis, no CA) and 2 replicates of 0.7 and 1% CA (n = 14 birds/group). Water and feed were provided ad libitum. On d 1, birds were tested for any inherent Salmonella (n = 2 birds/group). For the 3-wk trial, on d 5, birds were challenged with 8 log(10) cfu of Salmonella Enteritidis of a 4-strain mixture by crop gavage, and after 5 d postchallenge, birds (n = 2 birds/group) were euthanized to ensure Salmonella Enteritidis colonization. Caprylic acid was supplemented the last 5 d before tissue collection (n = 10 birds/group). For the 6-wk trial, on d 25, birds were challenged and confirmed for Salmonella Enteritidis colonization. The birds (n = 10 birds/group) were euthanized for tissue samples after CA supplementation for the last 5 d. Caprylic acid at 0.7 or 1% decreased Salmonella Enteritidis populations in cecum, small intestine, cloaca, liver, and spleen in both 3- and 6-wk trials. Body weight of birds did not differ between the groups (P ≥ 0.05). Further, to elucidate a potential antibacterial mechanism of action of CA, we investigated if CA could reduce Salmonella Enteritidis invasion of an avian epithelial cell line and expression of invasion genes hilA and hilD. The cell invasion study revealed that CA reduced invasive abilities of all Salmonella Enteritidis strains by ~80% (P < 0.05). Gene expression studies indicated that CA downregulated (P < 0.001) Salmonella invasion genes hilA and hilD. These results suggest that supplementation of CA through feed could reduce Salmonella Enteritidis colonization in broiler chicken and potentially reduces the pathogen's ability to invade intestinal epithelial cells by downregulating key invasion genes, hilA and hilD.