Ozone and photodynamic inactivation of norovirus surrogate bacteriophage MS2 in fresh Brazilian berries and surfaces.

This study assessed the efficacy of ozone (bubble diffusion in water; 6.25 ppm) and photodynamic inactivation (PDT) using curcumin (75 µM) as photosensitizer (LED emission 430-470 nm; 33.6 mW/cm2 irradiance; 16.1, 20.2, and 24.2 J/cm2 light dose) against the Norovirus surrogate bacteriophage MS2 in Brazilian berries (black mulberry and pitanga) and surfaces (glass and stainless steel). Contaminated berries and surfaces were immersed in ozonized water or exposed to PDT-curcumin for different time intervals. Transmission electron microscopy was used to assess the effects of the treatments on MS2 viral particles. The MS2 inactivation by ozone and PDT-curcumin varied with the fruit and the surface tested. Ozone reduced the MS2 titer up to 3.6 log PFU/g in black mulberry and 4.1 log PFU/g in pitanga. On surfaces, the MS2 reduction by ozone reached 3.6 and 4.8 log PFU/cm2 on glass and stainless steel, respectively. PDT-curcumin reduced the MS2 3.2 and 4.8 log PFU/g in black mulberry and pitanga and 2.7 and 3.3 log PFU/cm2 on glass and stainless steel, respectively. MS2 particles were disintegrated by exposure of MS2 to ozone and PDT-curcumin on pitanga. Results can contribute to establishing effective practices for controlling NoV in fruits and surfaces, estimated based on MS2 bacteriophage behavior.

Predicting the impact of temperature and relative humidity on Salmonella growth and survival in sliced chard, broccoli and red cabbage.

This study assessed the fate of a Salmonella enterica cocktail (S. Typhimurium, S. Enteritidis, S. Newport, S. Agona and S. Anatum; initial counts 3.5 log CFU/g) in minimally processed sliced chard, broccoli and red cabbage at 16 conditions of different temperature (7, 14, 21 and 37 °C) and relative humidity (RH; 15, 35, 65 and 95%) over six days (144 h). Linear regression was used to estimate the rate change of Salmonella in cut vegetables as a function of temperature and relative humidity (RH). R2 value of 0.85, 0.87, and 0.78 were observed for the rates of change in chard, broccoli, and red cabbage, respectively. The interaction between temperature and RH was significant in all sliced vegetables. Higher temperatures and RH values favored Salmonella growth. As temperature or RH decreased, the rate of S. enterica change varied by vegetable. The models developed here can improve risk management of Salmonella in fresh cut vegetables.

Efficacy of Grignard Pure to Inactivate Airborne Phage MS2, a Common SARS-CoV-2 Surrogate.

Grignard Pure (GP) is a unique and proprietary blend of triethylene glycol (TEG) and inert ingredients designed for continuous antimicrobial treatment of air. TEG has been designated as a ″Safer Chemical" by the US EPA. GP has already received approval from the US EPA under its Section 18 Public Health Emergency Exemption program for use in seven states. This study characterizes the efficacy of GP for inactivating MS2 bacteriophage─a nonenveloped virus widely used as a surrogate for SARS-CoV-2. Experiments measured the decrease in airborne viable MS2 concentration in the presence of different concentrations of GP from 60 to 90 min, accounting for both natural die-off and settling of MS2. Experiments were conducted both by introducing GP aerosol into air containing MS2 and by introducing airborne MS2 into air containing GP aerosol. GP is consistently able to rapidly reduce viable MS2 bacteriophage concentration by 2-3 logs at GP concentrations of 0.04-0.5 mg/m3 (corresponding to TEG concentrations of 0.025 to 0.287 mg/m3). Related GP efficacy experiments by the US EPA, as well as GP (TEG) safety and toxicology, are also discussed.

Growth models for Salmonella, E. coli O157:H7 and L. monocytogenes give different predictions for pathogen growth in cut leafy greens transportation, but are consistent in identifying higher risk conditions.

Leafy greens are frequently implicated in foodborne disease outbreaks and cut-leafy greens are a food that requires time and temperature control for safety. Predictive microbiology uses mathematical models to predict the growth of bacteria based on environmental conditions. The objective of our study was to compare published square root growth models for Salmonella (n = 6), pathogenic E. coli (n = 6) and Listeria monocytogenes (n = 4) using real world transport temperature data. Data from trucks transporting fresh-cut leafy greens during cross-country shipments were used as temperature inputs to the models. Bacterial growth was computed using the temperatures from each probe in every truck over the duration of transit, which resulted in 12-18 growth predictions per truck for each model. Each model generally gave significantly different predictions than other models for the same organism. The exception was for the two Salmonella models predicting the least growth and the two Salmonella models predicting the most growth which gave predictions that were not significantly different. Although different models tended to give different predictions, their ability to rank risk by truck was generally consistent across models. While absolute risk might be dependent upon choice of model, relative risk is independent of model choice.

Transfer of MS2 bacteriophage from surfaces to raspberry and pitanga fruits and virus survival in response to sanitization, frozen storage and preservation technologies.

This study assessed the norovirus (NoV) surrogate bacteriophage MS2 transfer from stainless steel, glass and low-density polypropylene surfaces to raspberry and pitanga fruits. The effect of sodium hypochlorite (100 ppm, 1 min) on MS2 survival on whole fruits, the MS2 survival in sanitized fruits and derived pulps during frozen storage, and in response to preservation technologies (heat, organic acids and salts) was also assessed. The highest (p < 0.05) viral transfer (%) was observed from glass and stainless steel (∼90%) to raspberry, and from glass and polypropylene (∼75%) to pitanga, after 60 min of contact. Sodium hypochlorite reduced (p < 0.05) MS2 titer by 3.5 and 3.8 log PFU/g in raspberry and pitanga, respectively. MS2 decreased (p < 0.05) up to 1.4 log PFU/g in frozen stored sanitized fruits (whole fruits and pulps) after 15 days, with no further changes after 30 days. Thermal treatments reduced MS2 titer (p < 0.05) in both fruit pulps. MS2 inactivation was higher in pitanga pulp. The addition of ascorbic acid, citric acid, sodium benzoate, or sodium metabisulfite had little effect (<1 log PFU/g) on MS2 concentration in either fruit. These results may inform NoV risk management practice in processing and handling of fruits.

Higher Concentrations of Bacterial Enveloped Virus Phi6 Can Protect the Virus from Environmental Decay.

Phage Phi6 is an enveloped virus considered a possible nonpathogenic surrogate for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other viral pathogens in transmission studies. Larger input amounts of bacteriophage Phi6 are shown to delay and protect the phage from environmental decay, both when the phages are dried in plastic tubes and when they are stored in saline solution at 4°C. In contrast, when bacteriophage Phi6 is placed in LB (Luria-Bertani) growth medium (instead of saline) prior to placement on the plastic surface, the influence of the starting concentration on viral recovery is negligible. Protection is reflected in the phage half-lives at higher concentrations being longer than the half-lives at lower concentrations. Because experiments supporting the possibility of fomite transmission of SARS-CoV-2 and other viruses rely upon the survival of infectious virus following inoculation onto various surfaces, large initial amounts of input virus on a surface may generate artificially inflated survival times compared to realistic lower levels of virus that a subject would normally encounter. This is not only because there are extra half-lives to go through at higher concentrations but also because the half-lives themselves are extended at higher virus concentrations. It is important to design surface drying experiments for pathogens with realistic levels of input virus and to consider the role of the carrier and matrix if the results are to be clinically relevant. IMPORTANCE During the coronavirus disease 2019 (COVID-19) pandemic, much attention has been paid to the environmental decay of SARS-CoV-2 due to the proposed transmission of the virus via fomites. However, published experiments have commenced with inocula with very high virus titers, an experimental design not representative of real-life conditions. The study described here evaluated the impact of the initial virus titer on the environmental decay of an enveloped virus, using a nonpathogenic surrogate for the transmission of SARS-CoV-2, enveloped bacteriophage Phi6. We establish that higher concentrations of virus can protect the virus from environmental decay, depending on conditions. This has important implications for stability studies of SARS-CoV-2 and other viruses. Our results point to a limitation in the fundamental methodology that has been used to attribute fomite transmission for almost all respiratory viruses.

Modeling the survival of Salmonella on whole cucumbers as a function of temperature and relative humidity.

Recent multistate outbreaks of salmonellosis associated with fresh cucumbers underscore the importance of understanding Salmonella behavior on cucumbers under different conditions. This study developed mathematical models to predict the survival of four-strain cocktail of Salmonella on whole cucumbers at different temperature and relative humidity (RH) conditions. The strains were Salmonella Newport H1275 and Stanley H0558 (sprout outbreaks), Montevideo G4639 (tomato outbreak), and Saintpaul 02-517-1 (cantaloupe outbreak). Inoculated cucumbers were placed in desiccators containing saturated salt solution to create controlled RH environments (~15, 50, 100% RH) at 7, 14, and 21 °C, and enumerated at time intervals ranging from 0 to 240 h. Predictive models were developed using Baranyi and Roberts equation as a primary model and estimated kinetic parameters were fitted into a polynomial equation for secondary models. Reduced model polynomial equations which describe the maximum death rate and the log reduction of Salmonella on whole cucumber as a function of temperature and RH had high R2 values (>0.95). Validation results verified the performance and reliability of the predictive models. The models in this study will be useful for future microbial risk assessments and predictions of Salmonella behavior in the cucumbers to manage the risk of Salmonella on whole cucumbers.

Models for factors influencing pathogen survival in low water activity foods from literature data are highly significant but show large unexplained variance.

Factors that control pathogen survival in low water activity foods are not well understood and vary greatly from food to food. A literature search was performed to locate data on the survival of foodborne pathogens in low-water activity (<0.70) foods held at temperatures <37 °C. Data were extracted from 67 publications and simple linear regression models were fit to each data set to estimate log linear rates of change. Multiple linear stepwise regression models for factors influencing survival rate were developed. Subset regression modeling gave relatively low adjusted R2 values of 0.33, 0.37, and 0.48 for Salmonella, E. coli and L. monocytogenes respectively, but all subset models were highly significant (p < 1.0e-9). Subset regression models showed that Salmonella survival was significantly (p < 0.05) influenced by temperature, serovar and strain type, water activity, inoculum preparation method, and inoculation method. E. coli survival was significantly influenced by temperature, water activity, and inoculum preparation. L. monocytogenes survival was significantly influenced by temperature, serovar and strain type, and inoculum preparation method. While many factors were highly significant (p < 0.001), the high degrees of variability show that there is still much to learn about the factors which govern pathogen survival in low water activity foods.

A predictive growth model for Clostridium botulinum during cooling of cooked uncured ground beef.

A dynamic model to predict the germination and outgrowth of Clostridium botulinum spores in cooked ground beef was presented. Raw ground beef was inoculated with a ten-strain C. botulinum spore cocktail to achieve approximately 2 log spores/g. The inoculated ground beef was vacuum packaged, cooked to 71 °C to heat shock the spores, cooled to below 10 °C, and incubated isothermally at temperatures from 10 to 46 °C. C. botulinum growth was quantified and fitted into the primary Baranyi Model. Secondary models were fitted to maximum specific growth rate and lag phase duration using Modified Ratkowsky equation (R2 0.96) and hyperbolic function (R2 0.94), respectively. Similar experiments were also performed under non-isothermal (cooling) conditions. Acceptable zone prediction (APZ) analysis was conducted on growth data collected over 3 linear cooling regimes from the current study. The model performance (prediction errors) for all 22 validation data points collected in the current work were within the APZ limits (-1.0 to +0.5 log CFU/g). Additionally, two other growth data sets of C. botulinum reported in the literature were also subjected to the APZ analysis. In these validations, 20/22 and 10/14 predictions fell within the APZ limits. The model presented in this work can be employed to predict C. botulinum spore germination and growth in cooked uncured beef under non-isothermal conditions. The beef industry processors and food service organizations can utilize this predictive microbial model for cooling deviations and temperature abused situations and in developing customized process schedules for cooked, uncured beef products.

A worldwide systematic review, meta-analysis, and health risk assessment study of mycotoxins in beers.

Mycotoxins, including aflatoxins (AFs), ochratoxin A (OTA), deoxynivalenol (DON), fumonisins (FBs), and zearalenone (ZEN), have been reported as beer contaminants. This systematic review and meta-analysis provide the prevalence and concentration of mycotoxins in beers and their worldwide distribution. Mycotoxin's exposure and cancer risk through beer consumption were determined. The overall pooled prevalence of mycotoxins in beers was 31% (95% confidence interval [CI] = 28%-35%; I2  = 90%, p = .00). The most prevalent mycotoxins in beers were DON and its derivatives (53%), OTA (52%), FBs (47%), followed by AFs (12%). Iran (99%), Hungary (95%), Denmark (92%), Armenia (83%), and Cyprus (83%) had the highest mycotoxin prevalence in beers. The global mycotoxins average concentration in beers was 12.52 µg/L (95% CI = 10.70-14.75 µg/L; I2  = 100%, p = .00). DON and its derivatives showed the highest concentration (26.91 µg/L), followed by FBs (23.19 µg/L), ZEN and its derivatives (20.25 µg/L), and AFs (15.65 µg/L). African region had the highest mycotoxins concentration (73.95 µg/L) mostly due to the high levels reported in beers from Cameroon (293.02 µg/L), Malawi (132.34 µg/L), and Eastern Cape province (126.12 µg/L). The meta-regression indicated stability (p ≥ .05) of the global pooled concentration of mycotoxins in beers over the years, whereas FBs concentration increased. The intake of DON and its derivatives, FBs, ZEN and its derivatives, and OTA through beers is of concern in African countries. OTA is also of concern in Brazil and Belgium. Results show high mycotoxins concentration in beers worldwide and highlight the health risks through contaminated beer consumption.

Modeling the Inactivation of Viruses from the Coronaviridae Family in Response to Temperature and Relative Humidity in Suspensions or on Surfaces.

Temperature and relative humidity are major factors determining virus inactivation in the environment. This article reviews inactivation data regarding coronaviruses on surfaces and in liquids from published studies and develops secondary models to predict coronaviruses inactivation as a function of temperature and relative humidity. A total of 102 D values (i.e., the time to obtain a log10 reduction of virus infectivity), including values for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), were collected from 26 published studies. The values obtained from the different coronaviruses and studies were found to be generally consistent. Five different models were fitted to the global data set of D values. The most appropriate model considered temperature and relative humidity. A spreadsheet predicting the inactivation of coronaviruses and the associated uncertainty is presented and can be used to predict virus inactivation for untested temperatures, time points, or any coronavirus strains belonging to Alphacoronavirus and Betacoronavirus genera.IMPORTANCE The prediction of the persistence of SARS-CoV-2 on fomites is essential in investigating the importance of contact transmission. This study collects available information on inactivation kinetics of coronaviruses in both solid and liquid fomites and creates a mathematical model for the impact of temperature and relative humidity on virus persistence. The predictions of the model can support more robust decision-making and could be useful in various public health contexts. A calculator for the natural clearance of SARS-CoV-2 depending on temperature and relative humidity could be a valuable operational tool for public authorities.

Successive exposure to Mentha piperita L. essential oil affects the culturability and induces membrane repair in a persister epidemic Salmonella Typhimurium PT4.

This study had as aims to evaluate the effects of successive exposures to Mentha piperita L. essential oil (MPEO) on culturability and physiological functions of Salmonella Typhimurium PT4. S. Typhimurium PT4 cells (108 log CFU/mL) were exposed to the same (1.25 µL/mL) or increasing MPEO concentrations (1.25-80 µL/mL) during 252 h. At each 36-h interval, the viable cell counts, and distinct cell functions were assessed using plate counting and flow cytometry, respectively. As the exposure time to the same MPEO concentration increased, the population of S. Typhimurium PT4 cells with damaged, permeabilized and depolarized membrane, and compromised efflux activity decreased. Otherwise, S. Typhimurium PT4 cells with damaged membrane physiological functions increased over the exposure to increasing concentrations of MPEO. Genomic analyses showed that the strain carries 17 genes associated with stress responses and the persistence of the tested strain among sources associated with poultry spanning more than 16 years and its virulence for humans. Therefore, successive exposure to a sublethal concentration of MPEO induced S. Typhimurium PT4 cells capable of maintaining the membrane integrity and its functions despite their non-culturable state.

Behavior of Listeria monocytogenes in the presence or not of intentionally-added lactic acid bacteria during ripening of artisanal Minas semi-hard cheese.

The fate of Listeria monocytogenes during ripening of artisanal Minas semi-hard cheese, as influenced by cheese intrinsic properties and by autochthonous (naturally present) or intentionally-added anti-listerial lactic acid bacteria (LAB) was modeled. Selected LAB strains with anti-listerial capacity were added or not to raw or pasteurized milk to prepare 4 cheese treatments. Counts of LAB and L. monocytogenes, pH, temperature and water activity were determined throughout cheese ripening (22 days, 22±1ᵒC). Different approaches were adopted to model the effect of LAB on L. monocytogenes: an independent approach using the Huang primary model to describe LAB growth and the linear decay model to describe pathogen inactivation; the Huang-Cardinal [pH] model using the effect of pH variation in a dynamic tertiary approach; and the Jameson-effect with Nmax tot model which simultaneously describes L. monocytogenes and LAB fate. L. monocytogenes inactivation occurred in both treatments with added LAB and inactivation was faster in raw milk cheese (-0.0260 h-1) vs. pasteurized milk cheese (-0.0182 h-1), as estimated by the linear decay model. Better goodness-of-fit was achieved for the cheeses without added LAB when the Huang primary model was used. A faster and great pH decline was detected for cheeses with added LAB, and the Huang-Cardinal [pH] model predicted higher pathogen growth rate in cheese produced with raw milk, but greater L. monocytogenes final concentration in pasteurized milk cheese. The Jameson-effect model with Nmax tot predicted that LAB suppressed pathogen growth in all treatments, except in the treatment with pasteurized milk and no LAB addition. The Huang-Cardinal [pH] model was more accurate in modeling L. monocytogenes kinetics as a function of pH changes than was the Jameson-effect model with Nmax tot as a function of LAB inhibitory effect based on the goodness-of-fit measures. The Jameson-effect model may however be a better competition model since it can more easily represent L. monocytogenes growth and death. This study presents crucial kinetic data on L. monocytogenes behavior in the presence of competing microbiota in Minas semi-hard cheese under dynamic conditions.

A comparison of dynamic tertiary and competition models for describing the fate of Listeria monocytogenes in Minas fresh cheese during refrigerated storage.

This study compares dynamic tertiary and competition models for L. monocytogenes growth as a function of intrinsic properties of a traditional Brazilian soft cheese and the inhibitory effect of lactic acid bacteria (LAB) during refrigerated storage. Cheeses were prepared from raw or pasteurized milk with or without the addition of selected LAB with known anti-listerial activity. Cheeses were analyzed for LAB and L. monocytogenes counts, pH and water activity (aw) throughout cold storage. Two approaches were used to describe the effect of LAB on L. monocytogenes: a Huang-Cardinal model that considers the effect of pH and aw variation in a dynamic kinetic analysis framework; and microbial competition models, including Lotka-Volterra and Jameson-effect variants, describing the simultaneous growth of L. monocytogenes and LAB. The Jameson-effect with γ and the Lotka-Volterra models produced models with statistically significant coefficients that characterized the inhibitory effect of selected LAB on L. monocytogenes in Minas fresh cheese. The Huang-Cardinal model [pH] outperformed both competition models. Taking aw change into account did not improve the fit quality of the Huang-Cardinal [pH] model. These models for Minas soft cheese should be valuable for future microbial risk assessments for this culturally important traditional cheese.

Selection of indigenous lactic acid bacteria presenting anti-listerial activity, and their role in reducing the maturation period and assuring the safety of traditional Brazilian cheeses.

Artisanal raw milk cheeses are highly appreciated dairy products in Brazil and ensuring their microbiological safety has been a great need. This study reports the isolation and characterization of lactic acid bacteria (LAB) strains with anti-listerial activity, and their effects on Listeria monocytogenes during refrigerated shelf-life of soft Minas cheese and ripening of semi-hard Minas cheese. LAB strains (n = 891) isolated from Minas artisanal cheeses (n = 244) were assessed for anti-listerial activity by deferred antagonism assay at 37 °C and 7 °C. The treatments comprised the production of soft or semi-hard Minas cheeses using raw or pasteurized milk, and including the addition of selected LAB only [Lactobacillus brevis 2-392, Lactobacillus plantarum 1-399 and 4 Enterococcus faecalis (1-37, 2-49, 2-388 and 1-400)], L. monocytogenes only, selected LAB co-inoculated with L. monocytogenes, or without any added cultures. At 37 °C, 48.1% of LAB isolates showed anti-listerial capacity and 77.5% maintained activity at 7 °C. Selected LAB strains presented a bacteriostatic effect on L. monocytogenes in soft cheese. L. monocytogenes was inactivated during the ripening of semi-hard cheeses by the mix of LAB added. Times to attain a 4 log-reduction of L. monocytogenes were 15 and 21 days for semi-hard cheeses produced with raw and pasteurized milk, respectively. LAB with anti-listerial activity isolated from artisanal Minas cheeses can comprise an additional barrier to L. monocytogenes growth during the refrigerated storage of soft cheese and help shorten the ripening period of semi-hard cheeses aged at ambient temperature.

A System Model for Understanding the Role of Animal Feces as a Route of Contamination of Leafy Greens before Harvest.

The majority of foodborne outbreaks in the United States associated with the consumption of leafy greens contaminated with Escherichia coli O157:H7 have been reported during the period of July to November. A dynamic system model consisting of subsystems and inputs to the system (soil, irrigation, cattle, wild pig, and rainfall) simulating a hypothetical farm was developed. The model assumed two crops of lettuce in a year and simulated planting, irrigation, harvesting, ground preparation for the new crop, contamination of soil and plants, and survival of E. coli O157:H7. As predicted by the baseline model for crops harvested in different months from conventional fields, an estimated 13 out of 257 (5.05%) first crops harvested in July would have at least one plant with at least 1 CFU of E. coli O157:H7. Predictions indicate that no first crops would be contaminated with at least 1 CFU of E. coli O157:H7 for other months (April to June). The maximum E. coli O157:H7 concentration in a plant was higher in the second crop (27.10 CFU) than in the first crop (9.82 CFU). For the second crop, the probabilities of having at least one plant with at least 1 CFU of E. coli O157:H7 in a crop were predicted as 15/228 (6.6%), 5/333 (1.5%), 14/324 (4.3%), and 6/115 (5.2%) in August, September, October, and November, respectively. For organic fields, the probabilities of having at least one plant with ≥1 CFU of E. coli O157:H7 in a crop (3.45%) were predicted to be higher than those for the conventional fields (2.15%). IMPORTANCE: This study is the first attempt toward developing a mathematical system model to understand the pathway of E. coli O157:H7 in the production of leafy greens. Results of the presented system model indicate that the seasonality of outbreaks of E. coli O157:H7-associated contamination of leafy greens was in good agreement with the prevalence of this pathogen in cattle and wild pig feces in a major leafy greens-producing region in California. On the basis of comparisons among the results of different scenarios, it can be recommended that the concentration of E. coli O157:H7 in leafy greens can be reduced considerably if contamination of soil with wild pig and cattle feces is mitigated.

Modeling the risk of salmonellosis from consumption of pistachios produced and consumed in the United States.

The risk of salmonellosis from consumption of pistachios produced and consumed in the U.S. was assessed through quantitative microbial risk assessment. Data on Salmonella prevalence and concentration on pistachios, nut crop volume, storage times and temperatures during processing and handling, and reductions during storage or from roasting were derived from laboratory experiments, published literature, and industry expert opinion. Uncertainty was analyzed via what-if scenarios for Salmonella prevalence, concentration, storage reduction, treatment variability, portion of crop treated, and increased consumption. The estimated U.S. incidence of salmonellosis when 100% of pistachios were exposed to a 4 ± 0 log reduction treatment averaged 1.4 cases per billion servings, or <1 case/year, without considering Salmonella decline during storage. Including Salmonella decline during storage reduced the salmonellosis estimates approximately 10-fold. The predicted arithmetic mean number of cases associated with individual 500,000-kg storage silos, contaminated at the highest observed levels, ranged from 5 to 530 when the product was consumed untreated, but was reduced to below 1 case per silo when a 4 ± 0 log reduction treatment was applied. Assuming a uniform 4-log reduction treatment is applied to 100% of the crop and there is no decline of Salmonella during storage, the assessment indicates the following: 10-fold increases in either Salmonella prevalence or concentration, 2-fold increases in both prevalence and concentration, or consumption of >0.05% of untreated product volume yield an arithmetic mean risk of >1 case/year.

Longer Contact Times Increase Cross-Contamination of Enterobacter aerogenes from Surfaces to Food.

Bacterial cross-contamination from surfaces to food can contribute to foodborne disease. The cross-contamination rate of Enterobacter aerogenes on household surfaces was evaluated by using scenarios that differed by surface type, food type, contact time (<1, 5, 30, and 300 s), and inoculum matrix (tryptic soy broth or peptone buffer). The surfaces used were stainless steel, tile, wood, and carpet. The food types were watermelon, bread, bread with butter, and gummy candy. Surfaces (25 cm2) were spot inoculated with 1 ml of inoculum and allowed to dry for 5 h, yielding an approximate concentration of 107 CFU/surface. Foods (with a 16-cm2 contact area) were dropped onto the surfaces from a height of 12.5 cm and left to rest as appropriate. Posttransfer, surfaces and foods were placed in sterile filter bags and homogenized or massaged, diluted, and plated on tryptic soy agar. The transfer rate was quantified as the log percent transfer from the surface to the food. Contact time, food, and surface type all had highly significant effects (P < 0.000001) on the log percent transfer of bacteria. The inoculum matrix (tryptic soy broth or peptone buffer) also had a significant effect on transfer (P = 0.013), and most interaction terms were significant. More bacteria transferred to watermelon (∼0.2 to 97%) than to any other food, while the least bacteria transferred to gummy candy (∼0.1 to 62%). Transfer of bacteria to bread (∼0.02 to 94%) was similar to transfer of bacteria to bread with butter (∼0.02 to 82%), and these transfer rates under a given set of conditions were more variable than with watermelon and gummy candy. IMPORTANCE: The popular notion of the "five-second rule" is that food dropped on the floor and left there for <5 s is "safe" because bacteria need time to transfer. The rule has been explored by a single study in the published literature and on at least two television shows. Results from two academic laboratories have been shared through press releases but remain unpublished. We explored this topic by using four different surfaces (stainless steel, ceramic tile, wood, and carpet), four different foods (watermelon, bread, bread with butter, and gummy candy), four different contact times (<1, 5, 30, and 300 s), and two bacterial preparation methods. Although we found that longer contact times result in more transfer, we also found that other factors, including the nature of the food and the surface, are of equal or greater importance. Some transfer takes place "instantaneously," at times of <1 s, disproving the five-second rule.

Cross contamination of Escherichia coli O157:H7 between lettuce and wash water during home-scale washing.

Lettuce and leafy greens have been implicated in multiple foodborne disease outbreaks. This study quantifies cross contamination between lettuce pieces in a small-scale home environment. A five-strain cocktail of relevant Escherichia coli O157:H7 strains was used. Bacterial transfer between single inoculated lettuce leaf pieces to 10 non-inoculated lettuce leaf pieces that were washed in a stainless steel bowl of water for 30 s, 1 min, 2 min, and 5 min was quantified. Regardless of washing time, the wash water became contaminated with 90-99% of bacteria originally present on the inoculated lettuce leaf piece. The E. coli O157:H7 concentration on initially inoculated leaf pieces was reduced ∼ 2 log CFU. Each initially uncontaminated lettuce leaf piece had ∼ 1% of the E. coli O157:H7 from the inoculated lettuce piece transferred to it after washing, with more transfer occurring during the shortest (30 s) and longest (5 min) wash times. In all cases the log percent transfer rates were essentially normally distributed. In all scenarios, most of the E. coli O157:H7 (90-99%) transferred from the inoculated lettuce pieces to the wash water. Washing with plain tap water reduces levels of E. coli O157:H7 on the inoculated lettuce leaf pieces, but also spreads contamination to previously uncontaminated leaf pieces.