D. E. Conner

Bactericidal activity of organic acids against Salmonella typhimurium attached to broiler chicken skin

The bactericidal activity of 0.5, 1,2,4, and 6% acetic, citric, lactic, malic, mandelic, propionic, or tartaric acid was determined against Salmonella typhimurium that were loosely or firmly attached to broiler chicken skin by using the skin-attachment model. Acid treatments were applied during a simulated chill (0°C/60 min), postprocess dip (23°C for 15 s), or scald (50°C for 2 min). For comparison, activity of the acid treatments when applied under these conditions were also determined against S. typhimurium in aqueous suspension. In general, bactericidal activity (mean reduction log CFU per skin) of all acids increased linearly with increasing concentration in all applications. The bactericidal activity of organic acids depended on concentration and method of application. When compared to freely suspended cells, it is clear that salmonellae both firmly and loosely attached to poultry skin have increased resistance to or are protected from organic acids. In general, concentrations of ≥4% of the acids were required to kill ≥2 log number of cells of S. typhimurium that were attached to broiler skin.

Growth, Inhibition, and Survival of Listeria monocytogenes as Affected by Acidic Conditions

Growth and survival of four strains of Listeria monocytogenes under acidic conditions were investigated. Tryptic soy broth with yeast extract (TSBYE) was acidified with acetic, citric, hydrochloric, lactic, or propionic acid to pH 4.0-6.0, inoculated with L. monocytogenes and incubated at 30 or 4°C. The minimum test pH at which L. monocytogenes did not grow (inhibitory pH) was determined for each acid. In the pH range tested, this inhibitory pH was 5.0 for propionic acid, 4.5 for acetic and lactic acids, and 4.0 for citric and hydrochloric acids. All four strains gave similar results. Subsequent studies were conducted at 10 and 30°C to determine changes in cell populations in TSBYE adjusted to each inhibitory pH. Initial populations of viable cells (104 CFU/ml) were reduced to <10 CFU/ml within 1-3 weeks at 30°C, whereas at 10°C, L. monocytogenes survived for 11-12 weeks in acetic, citric, or propionic acid-adjusted media and for 6 weeks in media adjusted with hydrochloric or lactic acid. The concentration of undissociated lactic acid was 0.002 M at pH 4.5.

Heat inactivation of Escherichia coli O157:H7 in turkey meat as affected by sodium chloride, sodium lactate, polyphosphate, and fat content

The purpose of this research was to determine the survival of Escherichia coli O157:H7 when heated in ground turkey containing various additives and fat levels. D values and z values were determined for low (3%)- and high (11 %)- fat ground turkey with or without one of three additives: 8% NaCl, 4% sodium lactate, or a mixture of 8% NaCl, 4% sodium lactate, and 0.5% polyphosphate. Products inoculated with E. coli O157:H7 strain 204P were mixed, aseptically placed into thermal-death-time (TDT) tubes which were sealed and heated at 52, 55, 57 and 60°C. Survival was determined by enumeration on phenol red sorbitol agar, and D values were calculated by two methods. Mean D52 values ranged from 46.8 to 104.8 min; mean D55 values ranged from 7.7 to 27.2 min; mean D57 values ranged from 2.7 to 13.0 min; and mean D60 values ranged from 0.7 to 4.8 min. The greatest survival, as evidenced by higher (P < 0.001) D values, occurred in turkey containing the mixture of additives. The z values ranged from 6.09 to 4.08°C, and higher z values were obtained in turkey meat containing the additive mixture versus other turkey additive formulations. The additives evaluated enhanced survival of E. coli O157:H7 in cooked turkey meat as compared to turkey meat with no additives. In contrast to earlier reports, added fat did not enhance survival (P > 0.05). Product formulation should be a critical consideration when safe cooking processes are developed for ready-to-eat turkey products.

Effects of acetic-lactic acid treatments applied to beef trim on populations of Escherichia coli O157:H7 and Listeria monocytogenes in ground beef

The efficacy of organic acid sprays for eliminating Escherichia coli O157:H7 and Listeria monocytogenes from beef trim used in a model ground beef production scheme was determined. Beef trim pieces with ca. 20% fat inoculated with E. coli O157:H7 or L. monocytogenes (ca. 3 log10 CFU/g) were utilized as controls or treated by spraying with 2 or 4% acetic and lactic acids. Propylene glycol (20%) was the carrier for each treatment. Following acid treatment, intact pieces were stored at 4°C for 12 or 24 h, ground, divided into 4 100-g retail packages and stored at 4°C for 0, 1, 2, or 4 days, at which time surviving populations of E. coli O157:H7 or L. monocytogenes were enumerated. High populations (>2.6 log10 CFU/g) of the pathogens persisted in all treatments. The 2% acid spray reduced (P < 0.01) the E. coli O157:H7 population by only 0.1 log10 CFU/g. The 2 and 4% acid sprays reduced (P < 0.001) the L. monocytogenes populations by 0.36 and 0.44 log10 CFU/g, respectively. Storing beef trim intact prior to grinding resulted in lower populations of E. coli O157:H7, and storage following grinding did not affect populations of either pathogen. The acid treatments tested were only slightly effective as sanitizers for beef trim destined for ground beef production.

Evaluation of various media for recovery of thermally-injured Escherichia coli O157:H7

Efficacies of plating media for recovering heated Escherichia coli O157:H7 were determined and compared. To compare populations of recovered cells, suspensions of cells (three isolates, four replications/isolate) were heated at 50, 55, or 60°C, and then inoculated onto eight media: PCA-PA (plate count agar with 1% pyruvic acid [PA]), MSA (MacConkey sorbitol agar), MSA-Mg (MSA with 0.025% MgSO4), MSA-PA (MSA with 1% PA), MSA-MUG (MSA with 0.005% 4-methylumbelliferyl-β-d-glucuronide (MUG), PRSA-MUG (phenol red sorbitol agar [PSRA] with 0.005% MUG), PRSA-PA (PRSA with 1% PA), and TSA-PA (tryptic soy agar with 1% PA). Recovery was consistently higher (P < 0.05) with PRSA-MUG and PRSA-PA. At 50, 55, and 60°C, mean numbers (log10 CFU/ml) of recovered cells on PRSA-MUG were 4.42, 4.62, and 3.32, respectively, as compared to 2.78, 2.08, and 1.63, respectively, on MSA. PCA-PA and TSA-PA were less effective than PRSA media, but better than MSA media. Thus, PRSA with MUG or PA was an effective medium for recovering heated cells of E. coli O157:H7; whereas MSA failed to detect sublethally injured cells. Furthermore, addition of Mg, PA, or MUG to MSA further compromised this medium.

Intestinal Cytokine Response of Commercial Source Broiler Chicks to Salmonella Typhimurium Infection

Development of molecular-based immunotherapeutic strategies for controlling Salmonella Typhimurium (ST) infection in poultry requires a better understanding of intestinal and cecal cytokine responses. Accordingly, an experiment was conducted to measure changes in intestinal cytokine expression when commercial source broiler chickens were challenged with a nalidixic acid-resistant ST. Ross broiler chicks were nonchallenged with ST (control treatment) or challenged by orally giving 7.8 x 10(6) cfu at 4 d of age (STC treatment). Each treatment consisted of 4 replicate pens with 14 chicks per pen. Expression levels of proinflammatory cytokines, interferon-gamma, and antiinflammatory interleukin (IL)-10 were determined at 5 and 10 d postchallenge (PC). Intestinal flushes were also collected from each treatment at 7 d PC to estimate IgA and IgG. Results showed an upregulation in IL-1beta mRNA in STC chicks at 5 d PC. By 10 d PC, the expression of IL-1beta was further increased and accompanied by an upregulation of IL-6 and interferon-gamma mRNA, whereas IL-10 mRNA expression decreased. It was concluded that ST induced an intestinal mucosal inflammatory response in commercial source broiler chicks less than 2 wk of age.

Influence of Salmonella enterica Serovar Typhimurium Infection on Intestinal Goblet Cells and Villous Morphology in Broiler Chicks

Abstract Live broiler chickens are important in the transmission of Salmonella to humans. Reducing Salmonella levels in the intestine of broiler chickens, in part, requires understanding of the interactions between Salmonella and the intestinal barriers that represent the first line of defense. Such barriers include the mucus layer (composed of mucins secreted by goblet cells) and the underlying epithelium. Two experiments were conducted to evaluate the effect of Salmonella Typhimurium infection on intestinal goblet cell dynamics (density and size) and villous morphology in broiler chicks. In Experiment 1, broiler chicks were either challenged with sterile media (control treatment) or orally given 7.4 × 107 colony-forming units (CFU) at 3 days of age (termed the CST treatment). Treatments were similar in Experiment 2, except that chicks in the CST treatment were challenged with 7.8 × 106 CFU at 4 days of age. Duration of each experiment was 14 days. At 7 days postchallenge (PC) in Experiment 1, jejunal tissue sections were collected, formalin-fixed, and routinely processed for histologic measurement of villous morphometric indices. In Experiment 2, at 10 days PC, jejunal tissue sections were collected and processed for histologic determination of goblet cell numbers and size, in addition to villous measurements. Results showed that Salmonella Typhimurium infection increased goblet cell density, reduced villous surface area, increased the incidence of epithelial exfoliation, and increased the incidence of heterophil influx into the lamina propria (P < 0.05). It was concluded that Salmonella Typhimurium infection impacts goblet cell biology and exerts morphopathologic changes in the jejunum of broiler chicks.

Evaluation of the efficacy of yeast extract in reducing intestinal Clostridium perfringens levels in broiler chickens

The etiological agent of necrotic enteritis is Clostridium perfringens. Traditionally, necrotic enteritis is controlled with in-feed antibiotics. However, increasing consumer demand for drug-free poultry has fostered the search for nonantibiotic alternatives. Yeast extract contain nucleotides that are immunomodulatory and also essential for cellular functions. An experiment was conducted to evaluate the efficacy of NuPro yeast extract (Alltech Inc., Nicholasville, KY) in reducing intestinal C. perfringens levels in broiler chickens. One hundred ninety-two 1-d-old male broiler chicks were obtained and randomly assigned to 6 treatments in a battery cage trial. Treatment 1 consisted of chicks fed a corn-soybean meal basal diet (BD) without added bacitracin methylene disalicylate or NuPro. Treatment 2 consisted of chicks fed BD into which bacitracin methylene disalicylate was added at 0.055 g/kg. Treatment 3 consisted of chicks fed BD supplemented with NuPro at a 2% level for the first 10 d of the experiment. Treatments 4 (PX), 5, and 6 (PN) consisted of chicks that were challenged with 3 mL of the C. perfringens inoculum (~10(7) cfu/mL) on d 14, 15, and 16 of the experiment and fed diets similar to treatments 1, 2, and 3, respectively. On d 1 and 7 postchallenge, intestinal C. perfringens levels, lesion scores, and alkaline phosphatase activity were assessed. On d 1 postchallenge, C. perfringens level in treatment 5 (2.09 log(10) cfu/g) was lower (P < 0.05) compared with the PX treatment (4.71 log(10) cfu/g) but similar to the PN treatment (2.98 log(10) cfu/g). A similar trend was observed on d 7 postchallenge. NuPro supplementation enhanced alkaline phosphatase activity (P < 0.05) in C. perfringens-challenged chicks and appeared to reduce intestinal lesion scores. Although dietary supplementation of NuPro in the PN treatment reduced C. perfringens levels by 1.73 and 0.68 log(10) cfu/g compared with the PX treatment on d 1 and 7 postchallenge, respectively, these reductions were not significant. Extending the period of NuPro supplementation beyond the first 10 d of life should be considered for achieving significant reduction in intestinal C. perfringensg levels.

Survival and growth of Salmonella Enteritidis in liquid egg products varying by temperature, product composition, and carbon dioxide concentration.

Cryogenic cooling of shell eggs with carbon dioxide (CO(2)) is known to improve egg content quality through rapid cooling as well as by increasing internal CO(2) levels. A study was undertaken to determine the effects of variations in atmospheric CO(2) concentrations (aerobically stored, flushed with CO(2) and sealed, or bubbled with CO(2)) on the survival and growth of Salmonella Enteritidis in liquid egg products including whole egg, albumen, yolk, and albumen + 1% yolk. Egg products were inoculated with a three-strain composite of Salmonella Enteritidis at ca. 4 log colony-forming units (CFU)/mL and stored at 7 degrees C or 10 degrees C for 8 or 4 days, respectively, or at ca. 2 log CFU/mL and stored at 23 degrees C and 37 degrees C for 48 or 24 hours, respectively. Salmonella populations differed based on variations in liquid egg composition (p < 0.05). Manipulating the atmospheric concentrations of CO(2) in which liquid egg products were stored did not significantly inhibit the growth of Salmonella Enteritidis (p > 0.05) in yolk-containing egg products or affect the inhibitory activity of albumen-containing products. Populations of Salmonella were static at 7 degrees C over the entire storage period and significant growth occurred in whole egg and yolk stored at 10 degrees C. Populations in egg stored at 23 degrees C and 37 degrees C were greater in yolk than in whole egg, although whole egg had populations greater than in albumen or albumen +1% yolk (p < 0.05). Results of this investigation suggest that increasing atmospheric CO(2) to enhance egg quality should not promote the growth of Salmonella Enteritidis in eggs.

Recovery of Salmonellae from chilled broiler carcasses as affected by rinse media and enumeration method

Distilled water (DW) and 0.85% NaCl (PS) were evaluated as carcass rinse media for recovery of total aerobic bacteria (APC), total coliforms (TC), Escherichia coli , and salmonellae from broiler carcasses. Salmonellae were enumerated by two methods, most-probable-number (MPN) and centrifugation-plating onto dulcitol novobiocin agar (DBN). Commercially processed chilled broiler carcasses (10/trial, 3 trials) were aseptically cut in half, and each half was rinsed (1 min) with either 250 ml DW or PS. Carcass rinses were recovered and analyzed for populations of APC, TC, E. coli , and salmonellae. Recovery of APC, TC, and E. coli were not affected (P>.05) by rinse media; however, significant trial effects were present. Recovery of salmonellae was influenced by rinse media as well as by enumeration method. Using the MPN procedure, salmonellae were detected on 20 and 27% of carcass halves using PS and DW, respectively, whereas with DBN, salmonellae were recovered from 33% of PS-rinsed carcass halves and none of those rinsed with DW. Incidence of salmonellae on individual carcass halves did not correlate between either the two enumeration methods or rinse media. With both enumeration methods, the extent of salmonellae contamination was <1 CFU/ml of rinse media. Rinsing carcasses with PS offered no advantages for recovery of APC, TC, and E. coli ; however, salmonellae recovery on DBN was enhanced by PS as compared to DW rinse.