Peptide gels of fully-defined composition and mechanics for probing cell-cell and cell-matrix interactions in vitro.

Current materials used for in vitro 3D cell culture are often limited by their poor similarity to human tissue, batch-to-batch variability and complexity of composition and manufacture. Here, we present a "blank slate" culture environment based on a self-assembling peptide gel free from matrix motifs. The gel can be customised by incorporating matrix components selected to match the target tissue, with independent control of mechanical properties. Therefore the matrix components are restricted to those specifically added, or those synthesised by encapsulated cells. The flexible 3D culture platform provides full control over biochemical and physical properties, allowing the impact of biochemical composition and tissue mechanics to be separately evaluated in vitro. Here, we demonstrate that the peptide gels support the growth of a range of cells including human induced pluripotent stem cells and human cancer cell lines. Furthermore, we present proof-of-concept that the peptide gels can be used to build disease-relevant models. Controlling the peptide gelator concentration allows peptide gel stiffness to be matched to normal breast (<1 kPa) or breast tumour tissue (>1 kPa), with higher stiffness favouring the viability of breast cancer cells over normal breast cells. In parallel, the peptide gels may be modified with matrix components relevant to human breast, such as collagen I and hyaluronan. The choice and concentration of these additions affect the size, shape and organisation of breast epithelial cell structures formed in co-culture with fibroblasts. This system therefore provides a means of unravelling the individual influences of matrix, mechanical properties and cell-cell interactions in cancer and other diseases.

Low salt exposure results in inactivation of Toxoplasma gondii bradyzoites during formulation of dry cured ready-to-eat pork sausage.

The production of safe and healthy food products represents one of the main objectives of the food industry. The presence of microorganisms in meat and products containing meat can result in a range of human health problems, as well as economic losses to producers of these products. However, contaminated meat products continue to initiate serious and large-scale outbreaks of disease in consumers. In addition to outbreaks of diseases caused by bacteria and viruses, parasitic organisms, such as Toxoplasma gondii, are responsible for foodborne infections worldwide, and in the case of T. gondii, is considered the 2nd leading cause of death from foodborne illness in the U.S. Transmission of Toxoplasma gondii has historically been linked to the consumption of raw or undercooked meat products, including pork. Specific concerns with respect to pork products are ready-to-eat (RTE) pork meals. These are pork or products containing pork that are prepared by curing or drying, and are not intended to be cooked before being consumed. Previous studies have demonstrated that T. gondii is inactivated during dry cured sausage preparation, apparently in the batter during fermentation. In this study, we have analyzed timing of inactivation of T. gondii in freshly prepared pepperoni batter to confirm our previous findings, to determine how quickly inactivation occurs during fermentation, and to confirm what parameters of the sausage preparation are involved in inactivation of the parasite. Results from the current and previous study indicate that rapid inactivation of T. gondii bradyzoites occurs in low salt batter for dry cured sausage within 4 h of initiation of fermentation.

Rapid inactivation of Toxoplasma gondii bradyzoites during formulation of dry cured ready-to-eat pork sausage.

Curing processes for pork meat in the U.S. currently require individual validation of methods to demonstrate inactivation of Trichinella spiralis, a nematode parasite historically associated with pork. However, for protozoan parasites, no such strictures exist. It has been assumed, with little evidence, that curing processes required to inactivate Trichinella also inactivate Toxoplasma gondii. Currently no model of meat chemistry exists that can be correlated with inactivation of T. gondii. Given the possibility of the presence of T. gondii in pork meat, and the frequent use of pork for ready-to-eat (RTE) products not intended to be cooked, curing methods which inactivate T. gondii early in the curing process would be of great value to producers. In this study, we tested the effect of five variables - salt/brine concentration, water activity (aw), pH, temperature, and time on inactivation of T. gondii bradyzoites in tissue cysts using low and high endpoints for common curing treatments during preparation of dry cured pork sausage. Survival of T. gondii bradyzoites at each stage of preparation was assessed using a mouse bioassay. Results indicated that encysted T. gondii bradyzoites do not survive the early stages of the dry curing process within the endpoint parameters tested here, even at levels of NaCl that are lower than typically used for dry curing (1.3%). Exposure of T. gondii encysted bradyzoites to curing components in the formulated batter resulted in rapid inactivation of bradyzoites. These data suggest that the use of dry curing components may be effective for controlling T. gondii potentially transmitted through RTE meats, rendering them safe from risk with respect to T. gondii transmission to human consumers.

Curing conditions to inactivate Trichinella spiralis muscle larvae in ready-to-eat pork sausage.

Curing processes are one method by which pork products, which are considered ready to eat (RTE) and have not been otherwise tested or treated, can be rendered safe from risk for exposure to Trichinella muscle larvae (ML). Curing processes in the U.S. currently require individual validation of methods to demonstrate inactivation of Trichinella. This is a major undertaking for each process; currently no model of meat chemistry exists that can be correlated with inactivation of Trichinella. Given the potential for new RTE products (e.g., lower salt), the availability of a wider range of tested methods for inactivation of Trichinella in pork would be of substantial value to the industry. In this study, five variables were tested - salt/brine concentration, water activity (aw), pH, temperature, and time, using low and high endpoints for common curing treatments for dry cured pork sausage. The data demonstrated that NaCl concentrations above 1.3%, in combination with fermentation to pH 5.2 or below, resulted in inactivation of > 96% of Trichinella ML in stuffed sausages within 24-28 h. All ML were inactivated by 7-10 days post-stuffing. These curing processes reliably predict inactivation of Trichinella spiralis, and can be used within the defined upper and lower endpoint parameters to reduce or eliminate the need for individual product validation.

Transfer of antibiotic resistance from Enterococcus faecium of fermented meat origin to Listeria monocytogenes and Listeria innocua.

UNLABELLED: Listeria monocytogenes is an important foodborne pathogen that can cause infection in children, pregnant women, the immunocompromised and the elderly. Antibiotic resistance in this species would represent a significant public health problem since the organism has a high fatality/case ratio and resistance may contribute to failure of therapeutic treatment. This study was designed to explore whether the in vitro transferability of antibiotic resistance from enterococci to Listeria spp. could occur. It was found that 2/8 Listeria strains were able to acquire tetracycline resistance from Enterococcus faecium. Listeria monocytogenes GLM-2 acquired the resistance determinant tet(M) and additional streptomycin resistance through in vitro mating with Ent. faecium S27 isolated from commercial fermented dry sausage. Similarly, Listeria innocua became more resistant to tetracycline, but the genetic basis for this change was not confirmed. It has been suggested that enterococci may transfer antibiotic resistance genes via transposons to Listeria spp., and this may explain, in part, the origin of their antibiotic resistance. Thus, the presence of enterococci in food should not be ignored since they may actively contribute to enhanced antibiotic resistance of L. monocytogenes and other pathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: Acquisition of antibiotic resistance by pathogenic bacteria in the absence of antibiotic pressure represents an unquantified threat to human health. In the present work resistance to tetracycline and streptomycin were transferred by nonplasmid-based conjugation from Enterococcus faecium isolated from fermented sausage to Listeria monocytogenes and Listeria innocua. Thus, natural transfer of antibiotic resistance to Listeria strains may occur in the future which reinforces the concern about the safety of enterococcal strains present in foods.

Use of a nanoparticulate carboxymethyl cellulose film containing sinigrin as an antimicrobial precursor to kill Escherichia coli O157:H7 on fresh beef.

UNLABELLED: Nanocomposite carboxymethyl cellulose films containing sinigrin (SNG) were prepared by stirring 2% (w/v) carboxymethyl cellulose (CMC) and 2% (w/v) glycerol (as a plasticizer) in distilled water with or without SNG (an antimicrobial precursor) as a 99% pure reagent (pSNG) or as a crude extract (cSNG). These films plus normal CMC film with or without SNG were tested on Escherichia coli O157:H7- inoculated beef for antimicrobial activity. Beef pieces measuring 6 × 5 × 2 cm(3) (L × W × H) were dipped in an E. coli O157:H7 broth suspension containing >8 log10 CFU ml(-1) and were drained for 3 min over a sterile cloth. They were wrapped in CMC or NCMC films, placed in a high oxygen barrier film (Deli *1), vacuum-packaged and stored at 8°C for 5, 8, 12 and 18 days. The CMC and NCMC films without SNG were not antimicrobial against E. coli O157:H7; however, NCMC and CMC films with SNG were highly antimicrobial. After 5 days at 8°C, E. coli O157:H7 was reduced more than 4 log10 by the NCMC•pSNG film and this reduction remained almost the same until 18 days at 8°C when E. coli O157:H7 was reduced >5 log10  CFU g(-1) meat. SIGNIFICANCE AND IMPACT OF THE STUDY: Transparent nanoparticulate carboxymethyl cellulose (CMC) films containing sinigrin (SNG), an antimicrobial precursor, controlled surface contamination of packaged fresh beef by the pathogen Escherichia coli O157:H7 when stored at 8°C. Films with nanoparticulation that carried pure SNG or the naturally occurring SNG in Oriental mustard were significantly more antimicrobial than similar films without nanoparticulation. As films without sinigrin were not antimicrobial, the combinations studied showed that nanoparticulation of the packaging film enhanced delivery of the antimicrobial incorporated within the film.

Temperature-dependent effect of sublethal levels of cinnamaldehyde on viability and morphology of Escherichia coli.

AIMS: Effects of sublethal levels of cinnamaldehyde (CIN) on the viability and morphology of Escherichia coli O157:H7 and E. coli 8 WT were investigated at 6 and 37°C. METHODS AND RESULTS: The minimum inhibitory concentration of CIN against E. coli O157:H7 and E. coli 8WT was 400 mg l(-1). At 37°C and ≤300 mg l(-1), CIN delayed the multiplication of both strains, causing a ≤5 and ≤13 h lag, respectively. Delayed multiplication at ≤300 mg l(-1) was partly due to cell elongation and injury as determined by LIVE/DEAD viability, CTC vitality and bis-(1,3-dibutylbarbituric acid) trimethine oxonol staining. The greatest extent of cell elongation (87%) and greatest mean length (6.4 µm) occurred with E. coli O157:H7 at 2-h exposure to 200 mg l(-1) CIN. After initial delays in multiplication, both E. coli O157:H7 and E. coli 8WT returned to exponential growth and normal morphology before reaching the stationary phase. In contrast at 6°C, CIN at ≥100 mg l(-1) prevented cell elongation which occurred in untreated control cells. Treatment with 200 or 300 mg l(-1) CIN at 6°C was lethal to both E. coli strains. At 300 mg l(-1) , CIN caused a ≥5 log CFU ml(-1) reduction at ≤3 days and completely inactivated both of these organisms, causing ≥7 log CFU ml(-1) reduction at 7 days. CONCLUSION: Sublethal levels of CIN at 37°C delayed the multiplication of E. coli cells by causing transient cell elongation, but at 6°C ≥200 mg l(-1) CIN was lethal to E. coli. SIGNIFICANCE AND IMPACT OF THE STUDY: Inhibition of cold-induced cell elongation and the enhanced lethal effect of CIN at 6°C against E. coli O157:H7 suggest that CIN may be useful for control of this pathogen at refrigeration temperatures.

Influence of NaCl and NaNO3 on sinigrin hydrolysis by foodborne bacteria.

The glucosinolate sinigrin (SNG) is converted by endogenous plant myrosinase or by bacterial myrosinase-like activity to form the potent antimicrobial allyl isothiocyanate. In order to use SNG as a natural antimicrobial precursor in food, it became important to better understand the ability of bacteria to synthesize the enzyme(s) and understand factors influencing this synthesis at a constant SNG concentration. Eight spoilage, pathogenic, or starter culture bacteria were grown separately in medium containing individual or combined salts with SNG. SNG degradation by the bacteria and the formation of its major degradation product, allyl isothiocyanate, were followed for 12 days at 30 or 35°C. The bacterial strains varied in their ability to metabolize SNG, and this was enhanced by NaCl and/or NaNO(3). SNG hydrolysis took place after 4 days, and the greatest amount occurred by day 12. At 12 days, Escherichia coli O157:H7 showed the greatest capacity to hydrolyze SNG (45.3% degradation), followed by Staphylococcus carnosus (44.57%), while Pseudomonas fluorescens was not active against SNG. The ability of tested strains to metabolize SNG, in decreasing order, was as follows: Escherichia coli O157:H7 > Staphylococcus carnosus > Staphylococcus aureus > Pediococcus pentosaceus > Salmonella Typhimurium > Listeria monocytogenes > Enterococcus faecalis > Pseudomonas fluorescens.

Susceptibility of meat starter cultures to antimicrobials used in food animals in Canada.

Lactic acid bacteria (LAB) are extensively used in the food industry for fermentation processes. However, it is possible that these bacteria may serve as a reservoir for antibiotic resistance genes that can be transferred to pathogens, giving rise to public health concerns. Animal operations that use antimicrobials as growth promotants have been linked to the origin of resistance due to the selective effect of low levels of antimicrobial used in this management strategy. The objective of this study was to determine the antimicrobial susceptibilities and mechanisms of resistance for 30 isolates of meat starter cultures commonly used in dry sausage fermentations to 20 antimicrobial agents. Susceptibility tests were performed by broth microdilution using Iso-Sensitest broth (90%, vol/vol) and de Man Rogosa Sharpe (MRS) broth (10%, vol/vol). The results showed that all 30 isolates exhibited resistance to at least three antimicrobials regardless of antimicrobial class while 17 or 30% of strains were resistant to antibiotics in three or six different classes, respectively. The incidence of antimicrobial resistance was higher among Pediococcus pentosaceus and lower for Staphylococcus carnosus strains. Genetic determinants for the lincosamide, macrolide, and tetracycline antimicrobials were not found using PCR. Phenotypic resistance in the absence of known resistance genes found here suggests that other mechanisms or genes might have contributed to the negative results. Further studies are needed to explore the genetic mechanisms underlying the prevalence of antibiotic resistance in Pediococcus species.

Heat resistance of Cronobacter species (Enterobacter sakazakii) in milk and special feeding formula.

AIM: To determine D- and z-values of Cronobacter species (Enterobacter sakazakii) in different reconstituted milk and special feeding formula and the effect of reconstitution of powdered milk and special feeding formula with hot water on the survival of the micro-organism. METHODS AND RESULTS: Five Cronobacter species (four C. sakazakii isolates and C. muytjensii) were heated in reconstituted milk or feeding formula pre-equilibrated at 52-58 degrees C for various times or mixed with powdered milk or feeding formula prior to reconstitution with water at 60-100 degrees C. The D-values of Cronobacter at 52-58 degrees C were significantly higher in whole milk (22.10-0.68 min) than in low fat (15.87-0.62 min) or skim milk (15.30-0.51 min) and significantly higher in lactose-free formula (19.57-0.66 min) than in soy protein formula (17.22-0.63 min). The z-values of Cronobacter in reconstituted milk or feeding formula ranged from 4.01 degrees C to 4.39 degrees C. Water heated to > or =70 degrees C and added to powdered milk and formula resulted in a > 4 log(10) reduction of Cronobacter. CONCLUSIONS: The heat resistance of Cronobacter should not allow the survival of the pathogen during normal pasteurization treatment. The use of hot water (> or =70 degrees C) during reconstitution appears to be an effective means to reduce the risk of Cronobacter in these products. SIGNIFICANCE AND IMPACT OF THE STUDY: This study supports existing data available to regulatory agencies and milk producers that recommended heat treatments are sufficient to substantially reduce risk from Cronobacter which may be present in these products.

Effect of free-SH containing compounds on allyl isothiocyanate antimicrobial activity against Escherichia coli O157:H7.

Escherichia coli O157:H7 contamination is a significant meat safety issue in many countries. Allyl isothiocyanate (AIT) is a natural compound found to limit the survival of E. coli O157:H7 and other pathogens in meat and meat products. In the present study, it was found that glutathione and cysteine naturally present in meat can interfere with AIT antimicrobial activity. Spectroscopy, HPLC, and LC-MS were used to confirm that glutathione was able to react with AIT and formed a conjugate with no or low bactericidal activity against the tested organisms. The same reaction also occurred at pH values of 4.9 and 5.8 at 25 and 4 degrees C, respectively, which broadly represent storage conditions in raw beef (pH 5.8) and during fermented sausage (pH 4.9) manufacture. Reactions observed help to explain reduction in antimicrobial potency of AIT in food (meat) systems.

Effect of environmental stresses on the sensitivity of Enterobacter sakazakii in powdered infant milk formula to gamma radiation.

AIM: To evaluate the effect of starvation, heat, cold, acid, alkaline, chlorine and ethanol stresses on the resistance of Enterobacter sakazakii in powdered infant milk formula (PIMF) towards gamma radiation. METHODS AND RESULTS: Stressed cells of E. sakazakii ATCC 51329 and four other food isolate strains were mixed individually with PIMF, kept overnight at room temperature, and then exposed to gamma radiation up to 7.5 kGy. The D(10)-values were determined using linear regression and for the stressed E. sakazakii strains these values ranged from 0.82 to 1.95 kGy. CONCLUSIONS: Environmental stresses did not significantly change the sensitivity of most E. sakazakii strains to ionizing radiation. SIGNIFICANCE AND IMPACT OF THE STUDY: Data obtained established that most forms of environmental stress are unlikely to significantly enhance the resistance of E. sakazakii strains to lethal, low dose irradiation treatment.

Phytosterol effects on milk and yogurt microflora.

Phytosterols play a major role in functional foods. Their ability to reduce serum cholesterol in humans has been widely proven and they are now being added to various dairy based products. The present study investigated the potential antimicrobial activity of phytosterols in milk and their effect on yogurt starter cultures. A commercial phytosterol (0.26% to 1.8%, w/v) preparation (CPP) consisting of ss-sitosterol, campesterol, sitostanol, and campestanol had no effect on the standard plate count (SPC) and psychrotroph population in pasteurized milk stored at 4 degrees C. In addition, a challenge study employing Pseudomonas spp. in milk at 4 to 7 degrees C confirmed that the CPP was not antimicrobial. However, the addition of a dispersible CPP consisting of 0.72% phytosterol containing 0.02% to 0.03% sodium stearoyl lactylate (SSL) did appear to affect the SPC and psychrotrophic bacteria in refrigerated milk. The dispersible preparations did not, however, inhibit the growth of Pseudomonas. An investigation into the antimicrobial activity of SSL revealed that it alone had no effect on the SPC in milk. The CPP had no effect on growth and acid development by Lactobacillus bulgaricus and Streptococcus thermophilus during yogurt production at 33 degrees C and storage at 4 degrees C for 30 d. This is seen as a beneficial feature since growth and acid development by these organisms are crucial for yogurt quality. Saccaromyces cerevisiae and Aspergillus ochraceous added to yogurt as typical contaminants also were not inhibited. While the CCP was somewhat antimicrobial when formulated with dispersing agents, it otherwise had no antimicrobial activity.

Predicting survival of Escherichia coli O157:H7 in dry fermented sausage using artificial neural networks.

The Canadian Food Inspection Agency required the meat industry to ensure Escherichia coli O157:H7 does not survive (experiences > or = 5 log CFU/g reduction) in dry fermented sausage (salami) during processing after a series of foodborne illness outbreaks resulting from this pathogenic bacterium occurred. The industry is in need of an effective technique like predictive modeling for estimating bacterial viability, because traditional microbiological enumeration is a time-consuming and laborious method. The accuracy and speed of artificial neural networks (ANNs) for this purpose is an attractive alternative (developed from predictive microbiology), especially for on-line processing in industry. Data from a study of interactive effects of different levels of pH, water activity, and the concentrations of allyl isothiocyanate at various times during sausage manufacture in reducing numbers of E. coli O157:H7 were collected. Data were used to develop predictive models using a general regression neural network (GRNN), a form of ANN, and a statistical linear polynomial regression technique. Both models were compared for their predictive error, using various statistical indices. GRNN predictions for training and test data sets had less serious errors when compared with the statistical model predictions. GRNN models were better and slightly better for training and test sets, respectively, than was the statistical model. Also, GRNN accurately predicted the level of allyl isothiocyanate required, ensuring a 5-log reduction, when an appropriate production set was created by interpolation. Because they are simple to generate, fast, and accurate, ANN models may be of value for industrial use in dry fermented sausage manufacture to reduce the hazard associated with E. coli O157:H7 in fresh beef and permit production of consistently safe products from this raw material.

Effects of superheated steam on Geobacillus stearothermophilus spore viability.

AIMS: To examine the effect of processing with superheated steam (SS) on Geobacillus stearothermophilus ATCC 10149 spores. METHODS AND RESULTS: Two inoculum levels of spores of G. stearothermophilus were mixed with sterile sand and exposed to SS at 105-175 degrees C. The decimal reduction time (D-value) and the thermal resistance constant (z-value) were calculated. The effect of cooling of spores between periods of exposure to SS was also examined. A mean z-value of 25.4 degrees C was calculated for both inoculum levels for SS processing temperatures between 130 degrees C and 175 degrees C. CONCLUSIONS: Spore response to SS treatment depends on inoculum size. SS treatment may be effective for reduction in viability of thermally resistant bacterial spores provided treatments are separated by intermittent cooling periods. SIGNIFICANCE AND IMPACT OF THE STUDY: There is a need for technologies that require short thermal processing times to eliminate bacterial spores in foods. The SS processing technique has the potential to reduce microbial load and to modify food texture with less energy in comparison to commonly used hot air treatment. This work provides information on the effect of SS processing parameters on the viability of G. stearothermophilus spores.

Occurrence and characterization of Salmonella from chicken nuggets, strips, and pelleted broiler feed.

Raw, frozen chicken nuggets and strips have been identified as a significant risk factor in contracting foodborne salmonellosis. Cases of salmonellosis as a result of consuming partly cooked chicken nuggets may be due in part to Salmonella strains originating in broiler feed. This study was undertaken to determine the occurrence and characterize the strains of Salmonella contaminating chicken nuggets, strips, and pelleted feeds, in an attempt to demonstrate whether the same Salmonella strains present in broiler feed could be isolated from raw, frozen chicken nuggets and strips available for human consumption. Salmonellae were recovered using the Health Canada MFHPB-20 method for the isolation and identification of Salmonella from foods. Strains were characterized by serotyping, phage typing, antimicrobial resistance typing (R-typing), and by pulsed-field gel electrophoresis (PFGE). Salmonellae were isolated from 25-g samples in 27% (n=92) of nugget and strip samples, 95% (n=20) of chicken nugget meat samples, and from 9% (n=111) of pelleted feed samples. Salmonella Heidelberg, Salmonella Enteritidis, and Salmonella Orion were the most commonly isolated serovars from chicken nuggets and strips, nugget and strip meat, and pelleted broiler feeds, respectively. Salmonella Enteritidis phage type (PT) 13a with PFGE pattern SENXAI.0006 and R-type sensitive as well as Salmonella Enteritidis PT13a with PFGE pattern SENXAI.0068 and R-type sensitive were isolated from pelleted feed, and chicken nugget and strip meat in two separate instances. Data showed that Salmonella strains isolated from broiler feed were indistinguishable from strains isolated from packaged raw, frozen chicken nuggets and strips. However, results did not rule out the possibility that breeding stock or contamination during processing may have contributed to chicken meat contamination by Salmonella.

Suitability of pea starch and calcium alginate as antimicrobial coatings on chicken skin.

The effect of incorporating trisodium phosphate (TSP) in pea starch (PS) and acidified sodium chlorite (ASC) in calcium alginate upon the antimicrobial activity of TSP and ASC was studied against a 3-strain cocktail of Salmonella inoculated on chicken skin. The influence of polymer coating concentration on skin pH, coating-skin adhesion, and coating absorption upon antimicrobial performance were investigated. Aqueous solutions of 0.5 to 4.8% (wt/vol) PS were prepared with 10% (wt/vol) TSP (PS + TSP coating), and alginate + ASC coatings contained 1% (wt/vol) calcium chloride in 1,200 ppm of ASC mixed with an aqueous solution of 0.5, 1.0, or 1.5% (wt/vol) sodium alginate. Coating drops (10 microL) were placed on chicken skin thighs, and the angle formed by the tangent of the liquid surface at the skin interface (contact angle) was measured using a digital camera to assess coating-skin adhesion. Excised skins were mounted in a ring holder, and 5 mL of the coatings was applied to the skin. Weight changes in the skins that were related to coating absorptiveness were recorded. The TSP dissolved in 3.5% PS and ASC in 1% alginate reduced Salmonella by 1.6 log cfu/g and 1.4 log cfu/g, respectively, within 24 h. These reductions were significantly greater than those caused by TSP or ASC alone in water for up to 120 h. In coatings, TSP and ASC caused significant elevation or reduction of skin surface pH for up to 120 h, respectively. The TSP destabilized PS with 88% of the coating having dripped from the skin 1 h later. Coatings with 0.5% PS were absorbed quickly by the skin and had high skin adhesion, whereas those with >3.5% PS had low skin adhesion and slow absorption. Alginate coatings with or without ASC were stable, and about 50% of the coating weight was retained at 120 h. The latter coatings appeared to have low absorptiveness because the skin gained approximately 1.0% of its weight within 60 min following application. These findings indicate that effects of the agents in coatings on skin pH, the extent of coating adhesion, and absorption may contribute to overall antimicrobial behaviors.

Inhibition of membrane bound ATPases of Escherichia coli and Listeria monocytogenes by plant oil aromatics.

Previous studies have reported that the mechanism of bactericidal action of the plant oil aromatics, eugenol, carvacrol and cinnamaldehyde involves inhibition of adenosine triphosphate generation and membrane disruption. In this study the capacity of the aromatics to inhibit the membrane bound ATPase activity of Escherichia coli and Listeria monocytogenes was investigated by experiments on isolated membranes. Inhibition of the ATPase activity of E. coli membranes was observed with 5 mM or 10 mM eugenol or carvacrol. Progressively greater inhibition by cinnamaldehyde was observed as concentration increased from 0.1 to 10 mM. L. monocytogenes ATPase activity was significantly inhibited by eugenol (5 or 10 mM), carvacrol (10 mM) and cinnamaldehyde (10 mM). Lactobacillus sakei is highly resistant to cinnamaldehyde compared to E. coli and L. monocytogenes. To determine whether this resistance was related to the relative hydrophobicity of the cell surface and hence the ability of the cell to take up the aromatics, the percentage of the three organisms partitioning in dodecane was compared. No significant difference was found between the partitioning percentage of L. monocytogenes (17.2%) and L. sakei (13.8%), indicating that surface hydrophobicity does not explain the differing sensitivity to cinnamaldehyde of these two organism. The percent partitioning of E. coli was significantly greater than both other organisms (23.3%) and may explain the greater sensitivity of E. coli to all three aromatics.

Enhancing the antimicrobial effects of bovine lactoferrin against Escherichia coli O157:H7 by cation chelation, NaCl and temperature.

AIM: To evaluate the effect of NaCl, growth medium and temperature on the antimicrobial activity of bovine lactoferrin (LF) against Escherichia coli O157:H7 in the presence of different chelating agents. METHODS AND RESULTS: LF (32 mg ml(-1)) was tested against E. coli O157:H7 strain 3081 in Luria broth (LB) and All Purpose Tween (APT) broth with metal ion chelators sodium bicarbonate (SB), sodium lactate (SL), sodium hexametaphosphate (SHMP), ethylene diamine tetraacetic acid (EDTA) or quercetin at 0.5 and 2.5% NaCl at 10 and 37 degrees C. LF and the chelators were tested against four other E. coli O157:H7 strains in LB at 2.5% NaCl and 10 degrees C. LF alone was bacteriostatic against strains 3081 and LCDC 7283 but other strains grew. Antimicrobial effectiveness of LF was reduced in APT broth but enhanced by SB at 2.5% NaCl and 10 degrees C where 4.0 log(10) CFU ml(-1) inoculated cells were killed. EDTA enhanced antimicrobial action of the LF-SB combination. SL alone was effective against E. coli O157:H7 but a reduction in its activity at 2.5% NaCl and 10 degrees C was reversed by LF. The combinations LF-SHMP and LF-quercetin were more effective at 37 degrees C and NaCl effects varied. CONCLUSIONS: LF plus SB or SL were bactericidal toward the same 3/5 E. coli O157:H7 strains and inhibited growth of the others at 2.5% NaCl and 10 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: The combination of LF with either SL or SB shows potential for reducing viability of E. coli O157:H7 in food systems containing NaCl at reduced, but growth permissive temperature.