A Longitudinal Study on the Dynamics of Salmonella enterica Prevalence and Serovar Composition in Beef Cattle Feces and Lymph Nodes and Potential Contributing Sources from the Feedlot Environment.

Salmonella can persist in the feedlot pen environment, acting as a source of transmission among beef cattle. Concurrently, cattle that are colonized with Salmonella can perpetuate contamination of the pen environment through fecal shedding. To study these cyclical dynamics, pen environment and bovine samples were collected for a 7-month longitudinal comparison of Salmonella prevalence, serovar, and antimicrobial resistance profiles. These samples included composite environment, water, and feed from the feedlot pens (n = 30) and cattle (n = 282) feces and subiliac lymph nodes. Salmonella prevalence across all sample types was 57.7%, with the highest prevalence in the pen environment (76.0%) and feces (70.9%). Salmonella was identified in 42.3% of the subiliac lymph nodes. Based on a multilevel mixed-effects logistic regression model, Salmonella prevalence varied significantly (P < 0.05) by collection month for most sample types. Eight Salmonella serovars were identified, and most isolates were pansusceptible, except for a point mutation in the parC gene, associated with fluoroquinolone resistance. There was a proportional difference in serovars Montevideo, Anatum, and Lubbock comparing the environment (37.2, 15.9, and 11.0%, respectively), fecal (27.5, 22.2, and 14.6%, respectively), and lymph node (15.6, 30.2, and 17.7%, respectively) samples. This suggests that the ability of Salmonella to migrate from the pen environment to the cattle host-or vice versa-is serovar specific. The presence of certain serovars also varied by season. Our results provide evidence that Salmonella serovar dynamics differ when comparing environment and host; therefore, developing serovar-specific preharvest environmental Salmonella mitigation strategies should be considered. IMPORTANCE Salmonella contamination of beef products, specifically from the incorporation of bovine lymph nodes into ground beef, remains a food safety concern. Current postharvest Salmonella mitigation techniques do not address Salmonella bacteria that are harbored in the lymph nodes, nor is it well understood how Salmonella invades the lymph nodes. Alternatively, preharvest mitigation techniques that can be applied to the feedlot environment, such as moisture applications, probiotics, or bacteriophage, may reduce Salmonella before dissemination into cattle lymph nodes. However, previous research conducted in cattle feedlots includes study designs that are cross-sectional, are limited to point-in-time sampling, or are limited to sampling of the cattle host, making it difficult to assess the Salmonella interactions between environment and hosts. This longitudinal analysis of the cattle feedlot explores the Salmonella dynamics between the feedlot environment and beef cattle over time to determine the applicability of preharvest environmental treatments.

Comparison of eggshell surface sanitization technologies and impacts on consumer acceptability.

Shell eggs can be contaminated with many types of microorganisms, including bacterial pathogens, and thus present a risk for the transmission of foodborne disease to consumers. Currently, most United States egg processors utilize egg washing and sanitization systems to decontaminate surfaces of shell eggs prior to packaging. However, previous research has indicated that current shell egg sanitization technologies employed in the commercial egg industry may not completely eliminate bacteria from the surface of eggshells, and thus alternative egg sanitization technologies with the potential for increased microbial reductions on eggshells should be investigated. The objectives of this study were to compare the antimicrobial efficacy and consumer sensory attributes of industry-available eggshell sanitization methods (chlorine and quaternary ammonium compounds (QAC) applied via spray) to various alternative egg sanitization technologies. Eggs (White Leghorn hens; n=195) were obtained for evaluation of sanitizer-induced reduction in mesophilic aerobic bacteria (n=90) or inoculated Salmonella Enteritidis (SE) reduction (n=105). Sanitizing treatments evaluated in this experiment were: chlorine spray (100 ppm available chlorine), QAC spray (200 ppm), peracetic acid spray (PAA; 135 ppm) alone or in combination with ultraviolet light (UV; 254 nm), and hydrogen peroxide (H2O2; 3.5% solution) spray in combination with UV (H2O2+UV). For enumeration of aerobic bacteria, eggs were sampled at 0, 7, and 14 days of storage at 4°C; surviving SE cells from inoculated eggs were enumerated by differential plating. Sensory trials were conducted to determine consumer liking of scrambled eggs made from eggs sanitized with chlorine, QAC, H2O2+UV, or no treatment (control). The H2O2 and UV treatment resulted in the greatest reductions in eggshell aerobic plate counts compared to other treatments throughout egg storage (P<0.05). All treatments utilized reduced SE below the limit of detection by eggshell rinse. There were no differences in consumers' liking of overall flavor between the 4 treatments evaluated. The application of H2O2+UV treatment to shell eggs represents a novel technology that could have important implications for egg quality and safety preservation.

Growth of Shiga toxin-producing Escherichia coli (STEC) and impacts of chilling and post-inoculation storage on STEC attachment to beef surfaces.

Concern has been expressed surrounding the utility of studies describing the efficacy of antimicrobial interventions targeting the Shiga toxin-producing Escherichia coli (STEC) that inoculate chilled versus non-chilled beef carcasses. The objectives of this study were to evaluate the effects of chilling (non-chilled, chilled to surface temperature of ≤5 °C) on STEC attachment to brisket surfaces, and the effects of post-inoculation storage on STEC recovery. Paired briskets from split carcasses were separated; one brisket from each pair was kept non-chilled, while the other was chilled to a surface temperature of ≤5 °C prior to inoculation. Briskets were inoculated with a cocktail of eight STEC and then stored at 5 or 25 °C. At 0, 30, 60, 90 and 120 min post-inoculation, 30 cm(2) of tissue was aseptically excised, followed by selective enumeration of strongly and loosely attached STEC. A significant, though small (0.4 log10 CFU/cm(2)), difference in the numbers of strongly attached cells was observed between non-chilled and chilled briskets (p < 0.05). Significant effects on cell attachment by the interaction of chilling and post-inoculation storage period, or chilling and post-inoculation storage temperature, were identified (p < 0.05). Results indicate beef chilling and post-inoculation storage conditions influenced STEC attachment to beef.

Inhibition of Escherichia coli O157:H7 and Salmonella enterica on spinach and identification of antimicrobial substances produced by a commercial Lactic Acid Bacteria food safety intervention.

The microbiological safety of fresh produce is of concern for the U.S. food supply. Members of the Lactic Acid Bacteria (LAB) have been reported to antagonize pathogens by competing for nutrients and by secretion of substances with antimicrobial activity, including organic acids, peroxides, and antimicrobial polypeptides. The objectives of this research were to: (i) determine the capacity of a commercial LAB food antimicrobial to inhibit Escherichia coli O157:H7 and Salmonella enterica on spinach leaf surfaces, and (ii) identify antimicrobial substances produced in vitro by the LAB comprising the food antimicrobial. Pathogens were inoculated on freshly harvested spinach, followed by application of the LAB antimicrobial. Treated spinach was aerobically incubated up to 12 days at 7 °C and surviving pathogens enumerated via selective/differential plating. l-Lactic acid and a bacteriocin-like inhibitory substance (BLIS) were detected and quantified from cell-free fermentates obtained from LAB-inoculated liquid microbiological medium. Application of 8.0 log10 CFU/g LAB produced significant (p < 0.05) reductions in E. coli O157:H7 and Salmonella populations on spinach of 1.6 and 1.9 log10 CFU/g, respectively. It was concluded the LAB antimicrobial inhibited foodborne pathogens on spinach during refrigerated storage, likely the result of the production of metabolites with antimicrobial activity.

An Experimental Field Trial Investigating the Use of Bacteriophage and Manure Slurry Applications in Beef Cattle Feedlot Pens for Salmonella Mitigation.

Post-harvest Salmonella mitigation techniques are insufficient at addressing Salmonella harbored in cattle lymph nodes, necessitating the exploration of pre-harvest alternatives that reduce Salmonella prior to dissemination to the lymph nodes. A 2 × 2, unbalanced experiment was conducted to determine the effectiveness of pre-harvest treatments applied to the pen surface for Salmonella mitigation in cattle. Treatments included manure slurry intended to mimic pen run-off water (n = 4 pens), a bacteriophage cocktail (n = 4), a combination of both treatments (n = 5), and a control group (n = 5) that received no treatment. Environment samples from 18 feedlot pens and fecal grabs, hide swabs, and subiliac lymph nodes from 178 cattle were collected and selectively enriched for Salmonella, and Salmonella isolates were sequenced. The combination treatment was most effective at reducing Salmonella, and the prevalence was significantly lower compared with the control group for rump swabs on Days 14 and 21. The treatment impact on Salmonella in the lymph nodes could not be determined due to low prevalence. The reduction on cattle hides suggests that bacteriophage or water treatments applied to the feedlot pen surface may reduce Salmonella populations in cattle during the pre-harvest period, resulting in reduced contamination during slaughter and processing.

Reducing Pathogenic Escherichia coli Surrogates on Fresh Beef Cuts by Water-Reducing Antimicrobial Interventions.

ABSTRACT: Water use for antimicrobial intervention application for beef harvest has come under increased scrutiny in recent years in an effort to enhance water conservation during beef harvest and fabrication. We determined the efficacy of beef safety interventions for reducing surrogates of the Shiga toxin-producing Escherichia coli (STEC) on beef cuts while lowering intervention-purposed water use for a small or very small beef establishment. Beef briskets, shoulder/clods, and rounds were inoculated with a gelatin-based slurry containing 6.8 ± 0.3 log CFU/g of nonpathogenic E. coli. After 30 min of attachment, inoculated cuts were treated by conventional lactic acid spray (2.5%, 55°C), lactic acid delivered by an electrostatic spray (2.5%, 55°C) handheld wand, hot water spray (82°C), or recycled hot water spray (82°C), wherein previously applied hot water was collected, thermally pasteurized to 82°C, or left untreated. One hundred milliliters of each treatment was sprayed onto marked surfaces of inoculated cuts, after which time surviving surrogate E. coli were enumerated. Lactic acid spray and electrostatic spray treatments produced greater reductions (1.0 to 1.1 log CFU/300 cm2) than hot water interventions (0.3 to 0.5 log CFU/300 cm2) (P ≤ 0.0001). Recycling of water reduced water losses by no less than 45% on recycled hot water spray-treated beef cuts. Low water beef safety interventions offer small and very small inspected beef establishments opportunities to incrementally reduce water use during intervention application, but not necessarily without loss of pathogen reduction efficacy.

Inhibition of Escherichia coli O157:H7 and Salmonella enterica Isolates on Spinach Leaf Surfaces Using Eugenol-Loaded Surfactant Micelles.

Spinach and other leafy green vegetables have been linked to foodborne disease outbreaks of Escherichia coli O157:H7 and Salmonella enterica around the globe. In this study, the antimicrobial activities of surfactant micelles formed from the anionic surfactant sodium dodecyl sulfate (SDS), SDS micelle-loaded eugenol (1.0% eugenol), 1.0% free eugenol, 200 ppm free chlorine, and sterile water were tested against the human pathogens E. coli O157:H7 and Salmonella Saintpaul, and naturally occurring microorganisms, on spinach leaf surfaces during storage at 5 °C over 10 days. Spinach samples were immersed in antimicrobial treatment solution for 2.0 min at 25 °C, after which treatment solutions were drained off and samples were either subjected to analysis or prepared for refrigerated storage. Whereas empty SDS micelles produced moderate reductions in counts of both pathogens (2.1-3.2 log10 CFU/cm2), free and micelle-entrapped eugenol treatments reduced pathogens by >5.0 log10 CFU/cm2 to below the limit of detection (<0.5 log10 CFU/cm2). Micelle-loaded eugenol produced the greatest numerical reductions in naturally contaminating aerobic bacteria, Enterobacteriaceae, and fungi, though these reductions did not differ statistically from reductions achieved by un-encapsulated eugenol and 200 ppm chlorine. Micelles-loaded eugenol could be used as a novel antimicrobial technology to decontaminate fresh spinach from microbial pathogens.

Sanitization of Chicken Frames by a Combination of Hydrogen Peroxide and UV Light To Reduce Contamination of Derived Edible Products.

Chicken carcass frames are used to obtain mechanically separated chicken (MSC) for use in other further processed food products. Previous foodborne disease outbreaks involving Salmonella-contaminated MSC have demonstrated the potential for the human pathogen to be transmitted to consumers via MSC. The current study evaluated the efficacy of multiple treatments applied to the surfaces of chicken carcass frames to reduce microbial loads on noninoculated frames and frames inoculated with a cocktail of Salmonella enterica serovar Enteritidis and Salmonella enterica serovar Typhimurium. Inoculated or noninoculated frames were left untreated (control) or were subjected to treatment using a prototype sanitization apparatus. Treatments consisted of (i) a sterile water rinse, (ii) a water rinse followed by 5 s of UV-C light application, or (iii) an advanced oxidation process (AOP) combining 5 or 7% (v/v) hydrogen peroxide (H2O2) with UV-C light. Treatment with 7% H2O2 and UV-C light reduced numbers of aerobic bacteria by up to 1.5 log CFU per frame (P < 0.05); reductions in aerobic bacteria subjected to other treatments did not statistically differ from one another (initial mean load on nontreated frames: 3.6 ± 0.1 log CFU per frame). Salmonella numbers (mean load on inoculated, nontreated control was 5.6 ± 0.2 log CFU per frame) were maximally reduced by AOP application in comparison with other treatments. No difference in Salmonella reductions obtained by 5% H2O2 (1.1 log CFU per frame) was detected compared with that obtained following 7% H2O2 use (1.0 log CFU per frame). The AOP treatment for sanitization of chicken carcass frames reduces microbial contamination on chicken carcass frames that are subsequently used for manufacture of MSC.

Assessment of Salmonella Prevalence in Lymph Nodes of U.S. and Mexican Cattle Presented for Slaughter in Texas.

Foodborne salmonellosis has been traced to undercooked ground beef and other beef products in the past, and peripheral lymph node (LN) presence in the fatty tissues of beef carcasses is one possible source of Salmonella contamination. Researchers have previously reported higher rates of Salmonella prevalence in LNs from cattle raised and harvested in Mexico compared with rates typically observed from cattle harvested in the United States. With cattle of Mexican origin comprising the majority of U.S. live cattle imports, the objectives of this study were designed to determine whether Salmonella prevalence in LNs differed (i) between cattle of Mexican and U.S. origins when exposed to the same South Texas feeding operation and (ii) between warm and cool seasons. To meet these objectives, paired (left and right sides) subiliac LNs ( n = 800 LNs; n = 400 pooled samples) were collected from 100 carcasses per origin (Mexico and United States) per season (cool, December to January; warm, July to September). Overall, Salmonella prevalence in LN samples was 52.0% (208 of 400). No difference ( P = 0.4836) was seen in Salmonella prevalence as a function of origin, with 54.0% (108 of 200) and 50.0% (100 of 200) of LN samples returning Salmonella-positive results from cattle of Mexican and U.S. origin, respectively. Salmonella prevalence differed ( P = 0.0354) between seasons, with 46.5% (93 of 200) of cool and 57.5% (115 of 200) of warm season samples returning Salmonella-positive results. Serotyping of PCR-confirmed positive samples resulted in 14 different serovars being identified, with Cerro (21.6%), Anatum (19.7%), Muenchen (17.8%), Montevideo (14.4%), and Kentucky (12.0%) comprising the majority of serovars. These results suggest that factors other than cattle origin may be impacting Salmonella prevalence rates in bovine LNs and that additional research is needed to better understand the role of environment and management-related factors on Salmonella prevalence in bovine LNs.

Comparison of Antimicrobial Treatments Applied via Conventional or Handheld Electrostatic Spray To Reduce Shiga Toxin-Producing Escherichia coli on Chilled Beef Outside Rounds.

HIGHLIGHTS: We found no advantage in the use of electrostatic spray to reduce STEC8 on cold beef. Greatest reductions in STEC8 were achieved by lactic acid with conventional spray. Lauric arginate ester was the second best antimicrobial agent at reducing STEC8. Lactic acid reduced pH on the beef surface significantly. There was no effect of antimicrobial solution on temperature increase on beef outside rounds.

Salmonella recovery from chicken bone marrow and cecal counts differ by pathogen challenge method.

Mechanically separated chicken (MSC) may be incorporated into other further processed foods, and has been identified as a transmission vehicle in human foodborne disease outbreaks involving the pathogen Salmonella enterica. Chickens possess pneumatic bones producing marrow, which may become infected with Salmonella and present a mechanism for Salmonella contamination of MSC. The primary objective of the current study was to ascertain whether chicken bone marrow was susceptible to Salmonella infection as a function of pathogen challenge method. Additionally, this study sought to determine the impact of Salmonella challenge site and inoculation on subsequent numbers of the microorganism in the cecum at 3, 6 and 9 d post-challenge. In this study, 30-day-old birds (Gallus domesticus) were separately challenged with S. enterica by one of the 6 experimental treatment methods: oral or endo-tracheal gavage, and transdermal scratch challenge on the breast or back muscles, with or without feathers (N = 150). Differing Salmonella recovery rates were detected in bone marrow from euthanized birds (P = 0.0015); oral and endo-tracheal gavage produced Salmonella recovery frequencies of 10% and 20%, respectively. Counts of Salmonella from chicken cecal samples statistically differed as a function of challenge method (P = 0.032); the highest numbers of Salmonella in cecal samples were obtained from orally gavaged birds (3.0 log10 cfu/g). Study results demonstrate the potential for chickens to contract short-term systemic infection following Salmonella challenge using methods of pathogen challenge simulating aerosol inhalation, ingestion, or wounding. Salmonella entry into chicken bone marrow may result in decreased likelihood of Salmonella performance standard compliance and increased pathogen transmission risk to consumers.

Evaluation of Commercial Prototype Bacteriophage Intervention Designed for Reducing O157 and Non-O157 Shiga-Toxigenic Escherichia coli (STEC) on Beef Cattle Hide.

Microbiological safety of beef products can be protected by application of antimicrobial interventions throughout the beef chain. This study evaluated a commercial prototype antimicrobial intervention comprised of lytic bacteriophages formulated to reduce O157 and non-O157 Shiga-toxigenic Escherichia coli (STEC) on beef cattle hide pieces, simulating commercial pre-harvest hide decontamination. STEC reduction in vitro by individual and cocktailed phages was determined by efficiency of plating (EOP). Following STEC inoculation onto hide pieces, the phage intervention was applied and hide pieces were analyzed to quantify reductions in STEC counts. Phage intervention treatment resulted in 0.4 to 0.7 log10 CFU/cm² (p < 0.01) E. coli O157, O121, and O103 reduction. Conversely, E. coli O111 and O45 did not show any significant reduction after application of bacteriophage intervention (p > 0.05). Multiplicity of infection (MOI) evaluation indicated E. coli O157 and O121 isolates required the fewest numbers of phages per host cell to produce host lysis. STEC-attacking phages may be applied to assist in preventing STEC transmission to beef products.

Increased Effectiveness of Microbiological Verification by Concentration-Dependent Neutralization of Sanitizers Used in Poultry Slaughter and Fabrication Allowing Salmonella enterica Survival.

Sanitizer neutralizers can assist foodborne pathogen detection during routine testing by counteracting sanitizer residues carried over into fluids collected and tested from food samples. This study tested sanitizer-matched neutralizers applied at increasing concentrations to facilitate Salmonella enterica survival following exposure to cetylpyridinium chloride (CPC) or peracetic acid (PAA), identifying minimum required concentrations of neutralizers to facilitate pathogen survival. Salmonella isolates were individually inoculated into a non-selective medium followed immediately by CPC (0.1 to 0.8% v/v) or PAA (0.0125 to 0.2% v/v) application, followed by neutralizers application. CPC was neutralized by lecithin and polysorbate 80, each supplemented into buffered peptone water (BPW) at 0.125 to 2.0X its respective content in Dey-Engley (D/E) neutralizing buffer. PAA was neutralized in BPW supplemented with disodium phosphate, potassium monophosphate, and sodium thiosulfate, each at 0.25 to 3.0X its respective concentration in BPW (phosphates) or D/E buffer (thiosulfate). Addition of neutralizers at 1X their respective concentrations in D/E buffer was required to allow Salmonella growth at the maximum CPC concentration (0.8%), while 2X neutralizer addition was required for Salmonella growth at the maximum PAA level (0.2%). Sanitizer neutralizers can assist pathogen survival and detection during routine food product testing.

National survey of Salmonella prevalence in lymph nodes of sows and market hogs.

Livestock are known to harbor Salmonella in their gastrointestinal (GI) tract and lymphatic tissues. Pathogens may be transferred from the GI tract to external carcass surfaces during normal harvest procedures but can be mitigated by antimicrobial carcass interventions. Lymph nodes (LNs) are typically encased in fat and are protected from antimicrobial carcass surface treatments, thus serving as a possible root cause of foodborne illnesses attributed to Salmonella in meat products. Members of the pork industry are committed to food safety and want to better understand Salmonella as a potential contaminant in pork products. To establish a baseline of Salmonella prevalence in porcine LNs across the United States, 21 commercial pork harvest facilities, representing northern (n = 12) or southern (n = 9) geographical regions, participated in this study. As processing volumes allowed, 25 carcasses were selected from each establishment. From each carcass, left and right superficial inguinal LNs (n =1,014 LNs) were removed and pooled to yield one sample per animal or n = 507 total LN samples. Salmonella prevalence rates differed (P < 0.05) between hog types in both regions. Specifically, 6.4% of market hog and 37.0% of sow samples were Salmonella positive in the northern region. This was reversed in the southern region as 13.0% of market hog and 4.8% of sow samples were Salmonella positive. There also was a difference (P < 0.05) in prevalence rates between northern and southern regions for sows, but not market hogs (P > 0.05). Type of chilling method (conventional, blast, or other) used at each market hog facility (n = 12) was documented. In the northern region, prevalence rates of Salmonella across chilling types were as follows: 20.0%, 2.7%, and 1.3% positive samples for conventional, other, and blast chill methods, respectively. In the southern region, 20.0% of samples were positive for conventional, 0.0% for blast, and 12.0% for other chilling methods. In both regions, samples from conventionally chilled carcasses returned more (P < 0.05) positive results than any other chill method. Overall, the higher rate of Salmonella prevalence in northern sows warrants further investigation, and members of the pork industry would benefit from the identification of possible methods to address the presence of Salmonella in porcine LNs.

The influence of surface chemistry on the kinetics and thermodynamics of bacterial adhesion.

This work is concerned with investigating the effect of substrate hydrophobicity and zeta potential on the dynamics and kinetics of the initial stages of bacterial adhesion. For this purpose, bacterial pathogens Staphylococcus aureus and Escherichia coli O157:H7 were inoculated on the substrates coated with thin thiol layers (i.e., 1-octanethiol, 1-decanethiol, 1-octadecanethiol, 16-mercaptohexadecanoic acid, and 2-aminoethanethiol hydrochloride) with varying hydrophobicity and surface potential. The time-resolved adhesion data revealed a transformation from an exponential dependence to a square root dependence on time upon changing the substrate from hydrophobic or hydrophilic with a negative zeta potential value to hydrophilic with a negative zeta potential for both pathogens. The dewetting of extracellular polymeric substances (EPS) produced by E. coli O157:H7 was more noticeable on hydrophobic substrates, compared to that of S. aureus, which is attributed to the more amphiphilic nature of staphylococcal EPS. The interplay between the timescale of EPS dewetting and the inverse of the adhesion rate constant modulated the distribution of E. coli O157:H7 within microcolonies and the resultant microcolonial morphology on hydrophobic substrates. Observed trends in the formation of bacterial monolayers rather than multilayers and microcolonies rather than isolated and evenly spaced bacterial cells could be explained by a colloidal model considering van der Waals and electrostatic double-layer interactions only after introducing the contribution of elastic energy due to adhesion-induced deformations at intercellular and substrate-cell interfaces. The gained knowledge is significant in the context of identifying surfaces with greater risk of bacterial contamination and guiding the development of novel surfaces and coatings with superior bacterial antifouling characteristics.

Extraction of nisin from a 2.5% commercial nisin product using methanol and ethanol solutions.

Nisin is a class Ia bacteriocin used widely in the food industry to inhibit a number of gram-positive pathogens. Although this peptide exhibits activity against many gram-positive bacteria, its effectiveness can vary significantly depending upon the food application. Encapsulation is one method that has been investigated for improving the activity of nisin. Improvement of the encapsulation efficiency of nisin requires purification of the compound, which can be accomplished utilizing organic solvents. The objective of this study was to use methanol and ethanol solutions to extract and concentrate nisin from a commercial preparation containing 2.5% nisin. Commercial nisin was extracted with different concentrations of ethanol or methanol in sterile water for up to 8 h. Approximately 75% of the nisin activity was recovered with 10 or 50% ethanol compared with less than 1% recovery with an ethanol concentration higher than 90%. Extraction with 10 or 50% methanol was approximately as effective as that with lower concentrations of ethanol. However, yields were significantly greater for extraction with methanol at concentrations greater than 90%. The solubility of the nisin likely influenced the extraction profiles for the conditions used. Purification for an 8-h extraction using 10 and 50% ethanol was 1.36 and 1.93 times, respectively. Purification was less than 0.1 at higher ethanol concentrations due to poor extraction. For methanol treatments, purification factors were all 1.09 to 5.98, and they increased as methanol concentration increased. This method for extracting and purifying nisin from dairy proteins using organic solvents may provide an alternative means for preparing and concentrating nisin for encapsulation and other applications.

Antimicrobial efficacy of eugenol microemulsions in milk against Listeria monocytogenes and Escherichia coli O157:H7.

The antimicrobial activity of eugenol microemulsions (eugenol encapsulated in surfactant micelles) in ultrahigh-temperature pasteurized milk containing different percentages of milk fat (0, 2, and 4%) was investigated. Antimicrobial microemulsions were prepared from a 5% (wt) aqueous surfactant solution (Surfynol 485W) with 0.5% (wt) eugenol. Two strains each of Listeria monocytogenes and Escherichia coli O157:H7 previously shown to be the least and most resistant to the microemulsion in microbiological media were used to inoculate sterile milk (10(4) CFU/ml). Samples were withdrawn and plated at 0, 1, 3, 6, 12, and 24 h for enumeration. Microemulsions completely prevented growth of L. monocytogenes for up to 48 h in skim milk and reduced both strains of E. coli O157:H7 to less than detectable levels in less than 1 h. Similarly, in 2% fat milk, eugenol-Surfynol combinations reduced both strains of E. coli O157:H7 to less than detectable levels in less than 1 h but only increased the lag phase of both strains of L. monocytogenes. In full-fat milk (4% fat), microemulsions inhibited growth of the least resistant strains of L. monocytogenes and E. coli but were ineffective against the two resistant strains. Unencapsulated eugenol was slightly more or as inhibitory as microemulsions against target pathogens. Results were attributed to diffusional mass transport of antimicrobials from microemulsions to the macroemulsion (milk). Results suggest that food composition, especially fat level, may affect the efficiency of targeting of foodborne pathogens with surfactant-encapsulated antimicrobials.

Inactivation of Escherichia coli K-12 exposed to pressures in excess of 300 MPa in a high-pressure homogenizer.

Homogenization is used widely in the dairy industry to improve product stability and quality. High-pressure homogenization (HPH) of fluid foods up to pressures of 300 MPa has demonstrated excellent potential for microbial inactivation. Microbial inactivation can be enhanced during HPH with the inclusion of antimicrobial compounds. Escherichia coli K-12 cells, grown statically or in chemostat, were exposed to HPH processing pressures of 50 to 350 MPa in the absence or presence of the antimicrobial nisin. Valve temperature was regulated by a water bath and pressure, and temperature data were recorded continuously after process initiation. Survivors were enumerated via plating on nonselective growth media. Pressure and temperature at the valve outlet port exhibited a quadratic relationship (R(2) = 0.9617, P < 0.05). Significant HPH-induced inactivation of the gram-negative microorganism was observed in the range of 100 to 250 MPa. Above 300 MPa, heat was the main factor promoting microbial inactivation, regardless of whether cells were grown in chemostat or statically. Chemostat-grown cells were significantly (P < 0.05) more resistant to HPH processing than were statically grown cells. Data indicate potential synergistic effects of nisin and HPH on the inactivation of bacterial contaminants. This study represents the first report of inactivation of a bacterium with HPH pressures in excess of 300 MPa in the presence and absence of an antimicrobial.

Effectiveness of a Commercial Lactic Acid Bacteria Intervention Applied to Inhibit Shiga Toxin-Producing Escherichia coli on Refrigerated Vacuum-Aged Beef.

Because of their antagonistic activity towards pathogenic and spoilage bacteria, some members of the lactic acid bacteria (LAB) have been evaluated for use as food biopreservatives. The objectives of this study were to assess the antimicrobial utility of a commercial LAB intervention against O157 and non-O157 Shiga-toxigenic E. coli (STEC) on intact beef strip loins during refrigerated vacuum aging and determine intervention efficacy as a function of mode of intervention application. Prerigor strip loins were inoculated with a cocktail (8.9 ± 0.1 log10 CFU/ml) of rifampicin-resistant (100.0 µg/ml; RifR) O157 and non-O157 STEC. Inoculated loins were chilled to ≤4°C and treated with 8.7 ± 0.1 log10 CFU/ml LAB intervention using either a pressurized tank air sprayer (conventional application) or air-assisted electrostatic sprayer (ESS). Surviving STEC were enumerated on tryptic soy agar supplemented with 100.0 µg/ml rifampicin (TSAR) to determine STEC inhibition as a function of intervention application method (conventional, ESS) and refrigerated aging period (14, 28 days). Intervention application reduced STEC by 0.4 log10 CFU/cm2 (p < 0.05), although application method did not impact STEC reductions (p > 0.05). Data indicate that the LAB biopreservative may assist beef safety protection when utilized within a multi-intervention beef harvest, fabrication, and aging process.

Validation of Thermal Lethality against Salmonella enterica in Poultry Offal during Rendering.

Recent outbreaks of human disease following contact with companion animal foods cross-contaminated with enteric pathogens, such as Salmonella enterica, have resulted in increased concern regarding the microbiological safety of animal foods. Additionally, the U.S. Food and Drug Administration Food Safety Modernization Act and its implementing rules have stipulated the implementation of current good manufacturing practices and food safety preventive controls for livestock and companion animal foods. Animal foods and feeds are sometimes formulated to include thermally rendered animal by-product meals. The objective of this research was to determine the thermal inactivation of S. enterica in poultry offal during rendering at differing temperatures. Raw poultry offal was obtained from a commercial renderer and inoculated with a mixture of Salmonella serovars Senftenberg, Enteritidis, and Gallinarum (an avian pathogen) prior to being subjected to heating at 150, 155, or 160°F (65.5, 68.3, or 71.1°C) for up to 15 min. Following heat application, surviving Salmonella bacteria were enumerated. Mean D-values for the Salmonella cocktail at 150, 155, and 160°F were 0.254 ± 0.045, 0.172 ± 0.012, and 0.086 ± 0.004 min, respectively, indicative of increasing susceptibility to increased application of heat during processing. The mean thermal process constant (z-value) was 21.948 ± 3.87°F. Results indicate that a 7.0-log-cycle inactivation of Salmonella may be obtained from the cumulative lethality encountered during the heating come-up period and subsequent rendering of raw poultry offal at temperatures not less than 150°F. Current poultry rendering procedures are anticipated to be effective for achieving necessary pathogen control when completed under sanitary conditions.