Reduction of Salmonella enterica Serovar Enteritidis Colonization in 20-Day-Old Broiler Chickens by the Plant-Derived Compounds trans -Cinnamaldehyde and Eugenol

Abstract

<jats:title>ABSTRACT</jats:title><jats:p>The efficacies of<jats:italic>trans</jats:italic>-cinnamaldehyde (TC) and eugenol (EG) for reducing<jats:named-content content-type="genus-species">Salmonella enterica</jats:named-content>serovar Enteritidis colonization in broiler chickens were investigated. In three experiments for each compound, 1-day-old chicks (<jats:italic>n</jats:italic>= 75/experiment) were randomly assigned to five treatment groups (<jats:italic>n</jats:italic>= 15/treatment group): negative control (-ve<jats:italic>S</jats:italic>. Enteritidis, -ve TC, or EG), compound control (-ve<jats:italic>S</jats:italic>. Enteritidis, +ve 0.75% [vol/wt] TC or 1% [vol/wt] EG), positive control (+ve<jats:italic>S</jats:italic>. Enteritidis, -ve TC, or EG), low-dose treatment (+ve<jats:italic>S</jats:italic>. Enteritidis, +ve 0.5% TC, or 0.75% EG), and high-dose treatment (+ve<jats:italic>S</jats:italic>. Enteritidis, +ve 0.75% TC, or 1% EG). On day 0, birds were tested for the presence of any inherent<jats:named-content content-type="genus-species">Salmonella</jats:named-content>(<jats:italic>n</jats:italic>= 5/experiment). On day 8, birds were inoculated with ∼8.0 log<jats:sub>10</jats:sub>CFU<jats:italic>S</jats:italic>. Enteritidis, and cecal colonization by<jats:italic>S</jats:italic>. Enteritidis was ascertained (<jats:italic>n</jats:italic>= 10 chicks/experiment) after 24 h (day 9). Six birds from each treatment group were euthanized on days 7 and 10 after inoculation, and cecal<jats:italic>S</jats:italic>. Enteritidis numbers were determined. TC at 0.5 or 0.75% and EG at 0.75 or 1% consistently reduced (<jats:italic>P</jats:italic>&lt; 0.05)<jats:italic>S</jats:italic>. Enteritidis in the cecum (≥3 log<jats:sub>10</jats:sub>CFU/g) after 10 days of infection in all experiments. Feed intake and body weight were not different for TC treatments (<jats:italic>P</jats:italic>&gt; 0.05); however, EG supplementation led to significantly lower (<jats:italic>P</jats:italic>&lt; 0.05) body weights. Follow-up<jats:italic>in vitro</jats:italic>experiments revealed that the subinhibitory concentrations (SICs, the concentrations that did not inhibit<jats:named-content content-type="genus-species">Salmonella</jats:named-content>growth) of TC and EG reduced the motility and invasive abilities of<jats:italic>S</jats:italic>. Enteritidis and downregulated expression of the motility genes<jats:italic>flhC</jats:italic>and<jats:italic>motA</jats:italic>and invasion genes<jats:italic>hilA</jats:italic>,<jats:italic>hilD</jats:italic>, and<jats:italic>invF</jats:italic>. The results suggest that supplementation with TC and EG through feed can reduce<jats:italic>S</jats:italic>. Enteritidis colonization in chickens.</jats:p>

In-Feed Supplementation of trans -Cinnamaldehyde Reduces Layer-Chicken Egg-Borne Transmission of Salmonella enterica Serovar Enteritidis

“…S. Enteritidis populations in the oviduct, liver, and cecum were determined as described previously (38). The organ samples and their contents from each bird were weighed and homogenized.…”

“…The infected COEC were lysed with 0.1% Triton X-100 (Invitrogen), and the number of viable adherent S. Enteritidis organisms was determined by serial dilution and plating on TSA and XLD plates. For the invasion assay, a gentamicin protection assay was performed as described previously (38,42). The S. Enteritidis-inoculated monolayer was incubated for 1 h, followed by three washings in specific minimal medium (MEM).…”

“…The isolates were SE-12 (chicken liver, phage type 14b), SE-21 (chicken intestine, phage type 8), SE-28 (chicken ovary, phage type 13a), and SE-31 (chicken gut, phage type 13a). Each strain was preinduced for resistance to 50 g/ml of nalidixic acid (NA; Sigma-Aldrich, St. Louis, MO) for selective enumeration (38). One hundred microliters of each NA-resistant strain was cultured separately in 10 ml tryptic soy broth (TSB; Difco, Becton Dickinson, Sparks, MD) overnight, transferred to flasks containing 100 ml TSB supplemented with 50 g/ml of NA, and incubated overnight at 37°C with shaking (100 rpm).…”

“…Subinhibitory concentrations (SICs) of TC against S. Enteritidis were determined as previously described (38). Sterile 24-well polystyrene tissue culture plates (Costar; Corning Incorporated, Corning, NY) containing TSB (1 ml/well) were inoculated separately with ϳ6.0 log CFU of S. Enteritidis, followed by the addition of 1 to 10 l of TC (Sigma-Aldrich), in increments of 0.5 l. The plates were incubated at 37°C for 24 h, and bacterial growth was determined by culturing on duplicate tryptic soy agar (TSA) and XLD plates.…”

“…It is a GRAS (generally regarded as safe) chemical approved for addition to foods by the U.S. FDA (approval TC-21CFR182.60). Previously, our laboratory observed that TC was effective at reducing S. Enteritidis in chicken cecal contents in vitro and in various internal organs in broilers (38). In addition, TC was found to inhibit biofilm formation by Cronobacter sakazakii (39) and uropathogenic Escherichia coli (40), by downregulating critical genes involved in biofilm synthesis.…”