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Aims The aims of this study were to: (i) estimate the effectiveness of ultraviolet radiation (UV) and sulphuric acid‐based fertilizer (SA), at reducing levels of generic Escherichia coli in surface irrigation water and on produce and surface soil in open produce fields; and (ii) describe the population dynamics of generic E. coli in produce fields. Methods and Results Spinach and cantaloupe plots were randomly assigned to control, UV or SA treatment groups. Irrigation water was inoculated with Rifampicin‐resistant E. coli prior to treatment. More than 75% of UV‐ and SA‐treated tank water samples had counts below the detection limit, compared to a mean count of 3·3 Log10 CFU per ml before treatment. Levels of Rifampicin‐resistant E. coli in soil and produce both increased and decreased over 10–15 days after irrigation, depending on the plot and time‐period. Conclusions UV and SA treatments effectively reduce the levels of E. coli in surface irrigation water. Their effectiveness at reducing contamination on produce was dependent on environmental conditions. Applying wait‐times after irrigation and prior to harvest is not a reliable means of mitigating against contaminated produce. Significance and Impact of the Study The results are of timely importance for the agricultural industry as new FSMA guidelines require producers to demonstrate a low microbial load in irrigation water or allow producers to apply a wait‐time to mitigate the risk of contaminated produce.

The objectives of this research were to study the effect of DMDC (0–250 ppm) on quality and shelf life of mango and passion fruit smoothie during cold storage. The correlation between microbial population (total microorganisms, yeast and mold, E. coli and S. aureus) and DMDC concentration using zero-order kinetic and first-order kinetic was also determined. In addition, the effect of DMDC compared with pasteurization (90 °C, 100 s) on quality of mixed mango and passion fruit smoothie during the cold storage (4 °C) was studied. The results showed that microbial inactivation was best-described by first-order kinetic model due to a higher coefficient of determination (R2). In addition, DMDC did not affect the decreasing trend of total soluble solid, color difference (∆E*) and total phenolic compound as compared to control during the cold storage. DMDC also hindered the increasing trend in microbial population and prevented the loss of antioxidant activity (DPPH and FRAP assays) and total flavonoid content and decreased the PPO activity as compared with the control during the cold storage. In summary, DMDC showed the potential to maintain the quality and to extend the shelf life of mango and passion fruit smoothie during cold storage.

Active packaging can enhance the performance of natural antimicrobials in controlling food spoilage and waste, while addressing consumer demands for cleaner labels. Yet, synergies are system dependent, with some conditions counterintuitively promoting antagonistic effects. In particular, metal chelators can improve performance of certain natural antimicrobials and have been incorporated in nonmigratory metal chelating active packaging technologies. However, the influence of chelating ligand chemistry on antimicrobial efficacy has not been investigated in microbial spoilage models. The effect of three commercial chelating resins on the growth of Alicyclobacillus acidoterrestris ATCC 49025, a thermoduric acidophilic spore-former, in growth media and apple juice was investigated. Dowex MAC-3, Chelex 100, and Lewatit TP260 were used as models for metal chelating active packaging containing carboxylic acid (CA), iminodiacetic acid (IDA), and aminomethylphosphonic acid (AMPA) ligands. Diameters (CA = 472.4 ± 117.2 μm, IDA = 132.93 ± 26.71 μm, and AMPA = 498.3 ± 29.24 μm), dissociation kinetics (CA = 6.44 ± 0.109, IDA = -0.977 ± 9.94, AMPA = 7.43 ± 0.193), and metal chelating capacities (CA = 1.16 × 10-4 mol/g, IDA = 1.52 × 10-3 mol/g, and AMPA = 4.67 × 10-4 mol/g) were used to distinguish differences in antimicrobial efficacies. Growth of A. acidoterrestris in acidified Potato Dextrose Broth over 24 hr with chelating resins indicated early death phase for CA and IDA resins and bactericidal for AMPA resin. However, viability in commercial apple juice with the inclusion of nisin and chelating resins was variable, with IDA resin significantly (P < 0.05) increasing viability while the effect of CA and AMPA resins remained elusive. This work emphasizes the importance of biological repeatability and correct statistical modeling in identifying conditions under which the antimicrobial intervention of nisin in real food systems, such as acidic beverages and juices, are synergistic or antagonistic. PRACTICAL APPLICATION: New technologies to control microbial food spoilage and waste need to be explored to address consumers on-going demands for reducing additive use. Solid support bound metal chelators can both promote and control microbial growth when used in conjunction with nisin, a natural antimicrobial. This work explores how system conditions can render a given technology either synergistic or antagonistic, and highlights the importance of sufficient biological replicates in experimental design.

Cleaning and sanitation are critical to maintaining safe and high-quality food production. Monitoring these activities is important to ensure proper execution of procedure and to assure compliance with regulatory guidelines., Rapid ATP testing and microbiological enumeration are two common methods to monitor the effectiveness of cleaning and sanitation in the food industry. In this study, ATP testing and microbiological enumeration were implemented at a tofu production facility with the goal of improving cleaning practices and overall plant hygiene. Results from ATP monitoring were used to target areas of the production environment needing additional cleaning; ATP results were verified by microbiological enumeration of aerobic microorganisms, lactic acid bacteria, and yeasts and molds. Products from the production line were enumerated for the same microorganisms to determine if there was an impact on product quality. After the implementation of ATP monitoring and targeted cleaning, there was a statistically lower proportion of swabs that failed to meet established sanitary requirements for ATP, aerobic microorganisms, and lactic acid bacteria (P

Wine and alcoholic apple cider are commonly back-sweetened with unpasteurized juice to produce fresh, natural, and palatable sweetened alcoholic beverages. Foodborne pathogens may be introduced from unpasteurized juice into alcoholic beverages through this back-sweetening process. Although foodborne pathogens generally do not survive under low pH conditions or a high alcohol environment, the die-off of these pathogens has not been established to ensure the microbiological safety of the products. To establish the holding conditions that would provide the required 5-log pathogen reduction requirements for these back-sweetened beverages, we evaluated the survival of three common foodborne pathogens, E. coli O157:H7, Salmonella enterica, and Listeria monocytogenes, in modified white grape juice and apple juice models. White grape juice and apple juice were modified with hydrochloric acid and sodium hydroxide and with ethanol to achieve conditions that are similar to back-sweetened white wine and alcoholic apple cider in regard to pH and ethanol content. Foodborne pathogen cocktails were inoculated separately into modified juice models, and their survival in the juice models was recorded over a 96-h period. Our results show that a combination of low pH and high ethanol content resulted in faster pathogen die-off compared with higher pH and lower ethanol conditions. The holding times required for different combinations of pH and ethanol concentration for each juice model to achieve a 5-log reduction were reported. This research provides data to validate pathogen die-off to comply with juice hazard analysis and critical control point 5-log pathogen inactivation requirements for back-sweetened wine and alcoholic apple cider. HIGHLIGHTS

Increasing consumer demand for high-quality foods has driven adoption by the food industry of non-thermal technologies such as high pressure processing (HPP). The technology is employed as a post-packaging treatment step for inactivation of vegetative microorganisms. In order to evaluate HPP inactivation of Escherichia coli O157:H7, Salmonella enterica, and Listeria monocytogenes in acid and acidified juices and beverages, pressure tolerance parameters were determined using log-linear and Weibull models in pH-adjusted apple juice (pH 4.5) at 5 °C. A commercial processing HPP unit was used. The Weibull model better described the inactivation kinetics of the three tested pathogens. According to estimates from the Weibull model, 1.5, 0.9, and 1.5 min are required at 600 MPa to produce 5-log reductions of E. coli, Salmonella, and L. monocytogenes, respectively, whereas according to the log-linear model, 3.2, 1.8, and 2.1 min are required. The effects of process conditions were verified using commercial products (pH between 3.02 and 4.21). In all tested commercial juices or beverages, greater than 5-log reductions were achieved for all tested pathogens using HPP process conditions of 550 MPa for 1 min. These findings demonstrate that the HPP conditions of 600 MPa for 3 min, typically used by the food industry provide an adequate safety margin for control of relevant vegetative pathogens in acid and acidified juices and beverages (pH 4.5). Results from this study can be used by food processors to support validation studies and may be useful for the future establishment of safe harbors for the HPP industry.

Microbial spoilage of fruit and vegetable juices represents an important threat to food quality and an area of concern in reducing food waste. Despite this, relatively little research is dedicated to microbial spoilage compared to other aspects of food microbiology. Establishing the incidence and impact of microbial spoilage in juice production would provide justification for future research. In this study, we present the findings from a survey of juice processor members of the U.S. based Juice Products Association and the European based International Fruit and Vegetable Juice Association (31.1% response rate). Respondents were asked a series of forced choice, Likert-type, and open response questions regarding microbial juice quality challenges and control measures regarding their facility. The vast majority of respondents (97.4%) indicated that spoilage mattered a lot or a great deal in brand protection. An additional 89.5% indicated that better control over microbial food spoilage would reduce waste and increase profits, with 57.9% indicating a lot or a great deal of impact, perhaps as a result of the frequency with which respondents indicated they discarded ingredients or product to protect quality. The most frequent disposition reportedly occurred on a weekly basis, with over half of respondents indicating discarding ingredients or product at least annually. Manufacturers reported a range of challenges, notably spoilage from Alicyclobacillus and heat resistant mold. This was accompanied by an identified need for associated sanitation and production control strategies. This work provides a basis for subsequent research exploring improved control strategies and detection methods used to reduce microbial spoilage of fruit and vegetable juices.

Novel processing technologies can be used to improve both the microbiological safety and quality of food products. The application of high pressure processing (HPP) in combination with dimethyl dicarbonate (DMDC) represents a promising alternative to classical thermal technologies. This research work was undertaken to investigate the combined effect of HPP and DMDC, which was aimed at reaching over 5-log reduction in the reference pathogens Escherichia coli O157:H7, Salmonella enterica, and Listeria monocytogenes inoculated in apple juice. Different strains of each species were tested. The pressure (ranging from 100 to 600 MPa), dwell time (from 26 to 194 s), and DMDC (from 116 to 250 mg/L) were tested based on a central composite rotatable design. The dwell time, in the studied range, did not have a significant effect (p > 0.1) on the pathogens´ reduction. All treatments achieved a greater than 5-log reduction for E. coli O157:H7 and L. monocytogenes. The reductions for S. enterica were also greater than 5-log for almost all tested combinations. The results for S. enterica suggested that it is more resistant to HPP and DMDC compared with E. coli O157:H7 and L. monocytogenes. The findings of this study showed that DMDC at low concentrations can be added to apple juice to reduce the parameters conventionally applied in HPP. The combined use of HPP and DMDC was highly effective under the conditions of this study.

Consumers are increasingly searching for preservative-free and “natural” food options. Confectionery products with low water activity (aw) are typically considered resistant to contaminati...

Consumption of spicy foods and hot sauces is currently a popular trend worldwide. Shelf-stable acidified sauces are commonly hot-filled to ensure commercial sterility, but cold-fill-hold processes might also be suitable if microbial safety and stability are ensured. For this study, model acidified hot pepper sauces were developed and characterized. The effects of sauce pH and of two different organic acids on the survival of Pichia manshurica and Lactobacillus curvatus isolated from contaminated commercial hot sauces and on pathogenic Escherichia coli O157:H7, Salmonella enterica, and Listeria monocytogenes were assessed. Full factorial designs with three levels for pH (3.2, 3.5, and 3.9) and two for organic acid (citric and acetic) were used to determine the effects of these factors and their interactions on the survival of the microorganisms. Commercially sterile sauces were independently inoculated and kept at ambient temperature. Microbial counts were determined at different sampling times, depending on the treatment evaluated. Sauces acidified to pH 3.2 with citric or acetic acid were inoculated with cocktails of five strains or serotypes of the three pertinent pathogens, and inactivation curves were determined. Trials were performed in triplicate. A greater than 5-log reduction of P. manshurica and L. curvatus was achieved in less than 6 h in sauces adjusted to pH 3.2 with acetic acid. Greater than 5-log reductions of pathogenic bacteria were achieved 0.5 h after inoculation in sauces acidified to pH 3.2 with acetic acid. In contrast, at least 48 h was required to guarantee the same inactivation for the most tolerant pathogen when citric acid was used. Thus, a cold-fill-hold process may be a suitable alternative for acidified hot pepper sauces. Based on survival of the microorganisms evaluated in this study, microbial safety and stability can be achieved by adjusting the pH to 3.2 or less by the addition of acetic acid. Universidad de Costa Rica/[735-B5-246]/UCR/Costa Rica UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro Nacional de Ciencia y Tecnología de Alimentos (CITA) UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Tecnología de Alimentos

Quality and safety of fresh produce are important to public health and maintaining commerce between Mexico and USA. While preventive practices can reduce risks of contamination and are generally successful, the variable environment of the supply chain of fresh produce can be suitable for introduction or proliferation of pathogenic microorganisms. As routine surveillance of these pathogens is not practical, indicator microorganisms are used to assess the sanitary conditions of production and handling environments. An opportunity exists to use indicators on fresh produce to measure how handling and transport from field to market may affect microbial populations that contribute to their quality or safety. The objective was to quantify indicator microorganisms on tomatoes sampled along the supply chain during the harvest year, in order to observe the levels and changes of populations at different locations. Roma tomatoes (n=475) were taken from the same lots (n=28) at four locations of the postharvest supply chain over five months: at arrival to and departure from the packinghouse in Mexico, at the distribution center in Texas, and at retail in USA. Samples were analyzed individually for four microbial populations: aerobic plate count (APC), total coliforms (TC), generic Escherichia coli, and yeasts and molds (YM). APC population differed (p

Food spoilage has an enormous economic impact, and microbial food spoilage plays a significant role in food waste and loss; subsequently, an equally significant portion of undergraduate food microbiology instruction should be dedicated to spoilage microbiology. Here, we describe a set of undergraduate microbiology laboratory exercises that focus specifically on food spoilage which were taught in 2 lab periods as part of the undergraduate food microbiology lab curriculum at Cornell University. The lab was broken down into 3 exercises. Two exercises lead students to determine the likely source of contamination in a canned salsa through (exercise 1a) plating and observation of colony morphology and (exercise 1b) determination of the thermal resistance for those isolates. The final exercise (2) involved detection of the spoilage bacterium Alicyclobacillus in apple juice. Spoiled juice demonstrations were also prepared in this exercise for students to observe sensorial changes resulting from spoilage, emphasizing that spoilage is not always visually detectable. Students were able to successfully determine the source of contamination based on the results of their laboratory findings, which they used to make recommendations for production to reduce microbial food spoilage in the canned salsa product. Based on student answers to discussion questions provided following lab exercises, participants were able to (a) identify the significance of microbial spoilage and how spoilage is principally different from food safety, (b) describe varying sensorial changes associated with microbial spoilage, and (c) employ methods and analysis to evaluate sources and type of contamination. Downloadable handouts and stepwise instructions are available as supporting information.

Environmental surfaces are common avenues for microbial contamination and transmission in food-processing establishments. We recently synthesized a polymer that combines both N-halamine and dopamine functional groups to form a novel antimicrobial coating material. A series of chemical (titration) and biological ("sandwich" and anti-inhabitation) tests were designed to prove the stability and functionality of as-developed coating material. Halamine-dopamine polymer-coated stainless-steel surface inactivated 6 log

The processing conditions and physiochemical properties used in food manufacturing create niches which support the growth of a limited number of spoilage fungi. This study was designed to evaluate the influence of intrinsic and extrinsic food product variables on the identity of spoilage fungi genera isolated from commercially produced foods. The spoilage etiology was identified in 127 products through ITS region sequencing. The prevalence and diversity of the identified spoilage fungi were evaluated in relationship to product-specific attributes using various descriptive statistics and a bipartite network analysis. Additionally, recursive partitioning was used to generate a classification tree with the outcomes, genera of the spoilage isolates, divided into increasingly homogenous subgroups. All of the isolated fungi belonged to the Ascomycete phylum, except four mucoralian isolates and the basidiomycete Rhodotorula. The occurrence of filamentous fungi repeatedly isolated ranged from 2% (Phoma spp.) to 18% (Penicillium spp.). In order of decreasing contribution to subgroup homogeneity, the split rules for the classification tree were based on process, water activity, food matrix category, and pH. Fungal genera representation in the terminal nodes indicated that production failures, in addition to product-specific attributes, were responsible for determination of the most probable spoilage organism.

Food processing, packaging, and formulation strategies are often specifically designed to inhibit or control microbial growth to prevent spoilage. Some of the most restrictive strategies rely solely or on combinations of pH reduction, preservatives, water activity limitation, control of oxygen tension, thermal processing, and hermetic packaging. In concert, these strategies are used to inactivate potential spoilage microorganisms or inhibit their growth. However, for select microbes that can overcome these controls, the lack of competition from additional background microbiota helps facilitate their propagation.

Fresh produce supply chains present variable and diverse conditions that are relevant to food quality and safety because they may favor microbial growth and survival following contamination. This study presents the development of a simulation and visualization framework to model microbial dynamics on fresh produce moving through postharvest supply chain processes. The postharvest supply chain with microbial travelers (PSCMT) tool provides a modular process modeling approach and graphical user interface to visualize microbial populations and evaluate practices specific to any fresh produce supply chain. The resulting modeling tool was validated with empirical data from an observed tomato supply chain from Mexico to the United States, including the packinghouse, distribution center, and supermarket locations, as an illustrative case study. Due to data limitations, a model-fitting exercise was conducted to demonstrate the calibration of model parameter ranges for microbial indicator populations, i.e., mesophilic aerobic microorganisms (quantified by aerobic plate count and here termed APC) and total coliforms (TC). Exploration and analysis of the parameter space refined appropriate parameter ranges and revealed influential parameters for supermarket indicator microorganism levels on tomatoes. Partial rank correlation coefficient analysis determined that APC levels in supermarkets were most influenced by removal due to spray water washing and microbial growth on the tomato surface at postharvest locations, while TC levels were most influenced by growth on the tomato surface at postharvest locations. Overall, this detailed mechanistic dynamic model of microbial behavior is a unique modeling tool that complements empirical data and visualizes how postharvest supply chain practices influence the fate of microbial contamination on fresh produce.IMPORTANCE Preventing the contamination of fresh produce with foodborne pathogens present in the environment during production and postharvest handling is an important food safety goal. Since studying foodborne pathogens in the environment is a complex and costly endeavor, computer simulation models can help to understand and visualize microorganism behavior resulting from supply chain activities. The postharvest supply chain with microbial travelers (PSCMT) model, presented here, provides a unique tool for postharvest supply chain simulations to evaluate microbial contamination. The tool was validated through modeling an observed tomato supply chain. Visualization of dynamic contamination levels from harvest to the supermarket and analysis of the model parameters highlighted critical points where intervention may prevent microbial levels sufficient to cause foodborne illness. The PSCMT model framework and simulation results support ongoing postharvest research and interventions to improve understanding and control of fresh produce contamination.

Salmonella enterica is the pertinent pathogen associated with orange juice products that have resulted in numerous foodborne outbreaks. Although fresh orange juice typically has a pH below 4.0, which inhibits most pathogen growth, S. enterica can survive at low pH for extended periods. Additionally, fresh juice contains spoilage microorganisms such as natural yeasts and molds, which can grow at low pH, and may cause fermentation and product spoilage if left untreated. Numerous Salmonella outbreaks linked to fresh orange juice, as well as the burden of product spoilage, have generated increased demand for alternative, non-thermal treatments that can ensure pathogen- and spoilage-free products. In this study, the effect of dimethyl dicarbonate (DMDC) on pathogen and spoilage microorganism inactivation in orange juice has been investigated with two experiments. First, pasteurized orange juice was inoculated with approximately 106–107 CFU/ml of five serotypes of S. enterica per ml and treated with DMDC to test the effectiveness of inactivation against Salmonella. For the fungal spoilage microorganism study, fresh orange juice was held at room temperature to increase natural yeast and mold count to roughly 105–106 CFU/ml, followed with treatment with DMDC. DMDC at two concentrations (172 and 200 ppm) was used, and the tests were carried out at ambient (21 °C ± 3 °C) and refrigeration (4 °C) temperatures. There was a >5-log reduction of Salmonella at 4 °C after 24 h at both 172 and 200 ppm of DMDC. For the treatment of fungal spoilage microorganisms, a nearly 5 and 4 log reduction of yeasts and molds was observed at ambient temperature and 4 °C, respectively. These results suggest that DMDC is most effective for use under the 4 °C holding conditions to inactivate S. enterica, and should be coupled with an additional preservative system for fungal spoilage control to produce safe orange juice that retains fresh quality.

The growing demand for fruit and vegetable juice blends, with improved nutritional and sensory attributes, has prompted the industrial adoption of nonthermal processing technologies, including UV light. Limited studies have explored conditions to overcome the well-known limitations of UV when treating liquid foods with a high content of particles that absorb or scatter UV light. This study addressed the effectiveness of the application of UV light, using a commercial processing unit, to inactivate pathogenic Escherichia coli O157:H7, Salmonella enterica (hereafter Salmonella), and Listeria monocytogenes, as well as spoilage microorganisms, in colored and turbid juices and beverages. The inactivation of cocktails of five strains (or serotypes) of E. coli O157:H7, Salmonella, and L. monocytogenes isolated from fruit- and vegetable-derived products linked to outbreaks was determined in seven colored and turbid cold-pressed juices and beverages. Juices and beverages were UV treated at a constant flow rate of 150 L/h through multiple consecutive passes. The inactivation of aerobic mesophilic bacteria, molds and yeasts, and lactic acid bacteria was also assessed at the cumulative dose that guaranteed a 5-log reduction of the most UV-tolerant pathogen for each product. A 5-log reduction of the three pathogens was achieved in all juices and beverages at a maximum cumulative UV dose of 12.0 ± 0.6 mJ/cm2. The dose required to ensure the targeted reduction varied depending on the tested product and the inoculated pathogen. The reduction of aerobic mesophiles, molds and yeasts, and lactic acid bacteria varied from 0.5 to 3.6, from 0.2 to 2.0, and from 0.5 to 3.6 log CFU/mL, respectively. Thus, the proposed treatment represents a suitable processing alternative to ensure the safety and extend the shelf life of colored and turbid cold-pressed juices and beverages. UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro Nacional de Ciencia y Tecnología de Alimentos (CITA)

The use of secondary quality produce has gained attention as a solution to food waste in both the U.S. and Europe. The purpose of this study was to evaluate the impact of using secondary quality or outgraded produce on the retention of surface inoculated E. coli following a rinse treatment on four model fresh produce systems (apple, tomato, carrot, lettuce). A three-strain cocktail of rifampicin-resistant generic E. coli, with a concentration of 9.0 log CFU/mL, was spot-inoculated on the intact surfaces of U.S. No.1 grade produce items and damaged or decayed areas of outgraded produce items. Generally, outgraded produce of all four kinds retained higher levels of inoculated E. coli following two postharvest treatments, chlorinated (150 ppm) or water only. However, physical damage, those defects which compromised the integrity of the produce surface, lead to significantly greater E. coli levels following rinsing than did physiological defects. Compared to U.S. No.1 quality apples, outgraded apples retained 4.3 ± 1.4 log CFU/g more E. coli following water only treatment, and 3.6 ± 1.7 log CFU/g more following chlorine treatment. Outgraded tomatoes retained significantly more (3.5 ± 1.1 log CFU/g) inoculated E. coli following water only rinse and 3.0 ± 1.4 log CFU/g more inoculated E. coli following chlorine treatment than U.S. No.1 quality tomatoes did under the same treatment conditions. Outgraded carrots retained 1 ± 1.1 log more CFU/g inoculated E. coli following water only treatment and 0.5 ± 0.8 log more CFU/g inoculated E. coli following chlorine treatment, compared to U.S. No.1 carrots. Outgraded lettuce leaves retained 1.6 ± 0.5 log CFU/g more inoculated E. coli following water only treatment and 4.1 ± 0.4 log CFU/g more inoculated E. coli following chlorine treatment than did U.S. No.1 quality lettuce leaves under the same treatment conditions. Treating with 150 ppm chlorine was not sufficient to eliminate the increased microbial retention associated with secondary quality or outgraded produce, and the efficacy of disinfection was greatly affected by type of defect. Apples with physical damage retained significantly higher E. coli loads than did those with physiological defects, an additional 2.6 log CFU/g under chlorine treatment and 0.8 log CFU/g more under was water only treatment. Tomatoes with physical damage had a 1.3-log CFU/g and 0.6-log CFU/g average increase of retained E. coli counts compared to those with physiological defects following a chlorine and water only treatment, respectively. Although a chlorine dip provided only a modest reduction in pathogens, generally, outgraded produce with physiological defects may present less food safety risks if introduced into the fresh market than does produce with physical damage due to their enhanced retention of bacterial cells. Therefore, as industry considers how to minimize its food waste problem, preferentially directing physically damaged produce away from the fresh market will help to minimize risk while maximizing food resources.

The low thermal tolerance of Salmonella enterica in foods with intermediate moisture levels, such as caramel sauces, ensures that mild heat treatment is sufficient to achieve 5-log reductions of this pathogen. This treatment mitigates the risk posed by salmonellae in raw materials; however, recontamination might occur because of survival of the pathogen in products that are not heated before consumption. This study was conducted to evaluate the effect of water activity (aw) on the thermal tolerance and survival of S. enterica serovars Tennessee and Senftenberg. The D-values at 76, 78, and 80°C, z-values, and survival at 20.0 ± 0.5°C for 32 weeks of these two serovars were determined in goat's milk caramel at three aw values (0.85, 0.90, and 0.93). The highest thermal tolerance was observed at aw = 0.85 for Salmonella Senftenberg (D76°C = 2.9 ± 0.3 min), and the lowest was at aw = 0.93 for Salmonella Tennessee (D80°C = 0.131 ± 0.007 min). After a logarithmic transformation of the z-values, a significant interaction between serovar and aw was found (P 8-log reduction was observed at week 20 of storage, regardless of the product's aw and the serovar, low levels of salmonellae were found in the product up to week 32 of storage. Our findings may assist the food industry with the establishment of critical limits for the safe thermal treatment of milk- and sugar-based foods with intermediate moisture levels. The survival data presented here highlight the relevance of implementing and effectively maintaining good sanitation and hygiene practices during the production of goat's milk caramel and similar food products.

The CiderSure 3500 is one of the most commonly used UV juice processing units in the United States for the nonthermal processing of apple cider and fulfills the 5-log performance standard established by the federal juice HACCP regulation. However, the appropriate validation frequency of this machine’s quartz tubes is currently unknown by juice processors and regulatory agencies. Presently, an annual validation is recommended by the manufacturer. Historical validation data from 1998 to 2013 of commercially used quartz tubes underwent comprehensive statistical analysis. A total of 400 tubes were validated one time, and 212 of those units were revalidated at least once over the evaluated time frame. Validations were performed at 14 mJ?cm 22 UV dose and under turbulent flow conditions. Every validation showed a greater than 5-log reduction of Escherichia coli ATCC 25922, a nonpathogenic surrogate for pathogenic E. coli O157:H7, in each of three replicates. For initial validations, a mixed-effect model with log reduction of E. coli as response was constructed (400 tubes analyzed in triplicate). The model showed that the year of analysis and the initial inoculum level significantly affected the log reduction of E. coli (P , 0.0001), which on average was 7.0 i 0.7. A quadratic relationship between the year of analysis and the response was found. Likewise, for revalidations (212 tubes analyzed in triplicate), the constructed random coefficient model showed that the year of analysis, quadratic effect of year of analysis, and initial inoculum level significantly affected the log reduction of E. coli (P , 0.0001). For this model, the major source of variance was explained by the year of analysis. The models describe the UV reactor’s performance over time and suggest that a validation frequency of every 3 years would be conservatively adequate during the first 8 years of quartz tube use. After that, due to the reported quadratic trend, yearly validation would be recommended. In 2000, the U.S. Food and Drug Administration recognized UV light treatment as an alternative for thermal pasteurization of juices and beverages. The requirements state in 21 Code of Federal Regulations 179.39 that UV radiation may be safely used for the processing of juice products when the treatment is provided by low-pressure mercury lamps emitting 90% of the emission at a wavelength of 253.7 nm and the juice undergoes turbulent flow through tubes, with a minimum Reynolds number of 2,200 (25). Since then, and due to an increased consumer demand for more fresh products with enhanced nutritional properties (2, 23), the applications of this nonthermal technology have increased, with the advantage that UV light is characterized by low energy requirements and reduced initial investment in comparison with thermal pasteurization (3, 15, 18). Furthermore, with the intention of preventing potentially negative effects, due the application of traditional heat treatments, on the organoleptic properties of cider, and given that unpasteurized and contaminated apple cider has caused several foodborne outbreaks (6, 21), many small- and medium-sized cider producers have acquired a commercial UV juice processing unit to safely treat this

Listeria species have been isolated from diverse environments, often at considerable prevalence, and are known to persist in food processing facilities. The presence of Listeria spp. has been suggested to be a marker for Listeria monocytogenes contamination. Therefore, a study was conducted to (i) determine the prevalence and diversity of Listeria spp. in produce production and natural environments and (ii) identify geographical and/or meteorological factors that affect the isolation of Listeria spp. in these environments. These data were also used to evaluate Listeria spp. as index organisms for L. monocytogenes in produce production environments. Environmental samples collected from produce production (n = 588) and natural (n = 734) environments in New York State were microbiologically analyzed to detect and isolate Listeria spp. The prevalence of Listeria spp. was approximately 33 and 34% for samples obtained from natural environments and produce production, respectively. Co-isolation of L. monocytogenes and at least one other species of Listeria in a given sample was recorded for 3 and 9% of samples from natural environments and produce production, respectively. Soil moisture and proximity to water and pastures were highly associated with isolation of Listeria spp. in produce production environments, while elevation, study site, and proximity to pastures were highly associated with isolation of Listeria spp. in natural environments, as determined by randomForest models. These data show that Listeria spp. were prevalent in both agricultural and nonagricultural environments and that geographical and meteorological factors associated with isolation of Listeria spp. were considerably different between the two environments.

Gradual exposure to moderate acidic environments may enhance the thermal tolerance and survival of Escherichia coli O157:H7 in acid and acidified foods. Limited studies comparing methodologies to induce this phenomenon have been performed. The effects of strain and physiological state on thermal tolerance and survival of E. coli in apple juice were studied. The decimal reduction time (D-value) at 56°C [D56°C] was determined for E. coli O157:H7 strains C7927 and ATCC 43895 and E. coli O111 at four physiological states: unadapted, acid-shocked (two methodologies used), and acid-adapted cells. The effect of acidulant was also evaluated by determining the D56°C for the O157:H7 strains subjected to acid shock during 18 h in Trypticase soy broth (TSB), with pH 5 adjusted with hydrochloric, lactic, and malic acids. Survival of the three strains at four physiological states was determined at 1 ± 1°C and 24 ± 2°C. Experiments were performed in triplicate. For thermal inactivation, a significant interaction was found between strain and physiological state (P0.0001). Highest thermal tolerance was observed for the 43895 strain subjected to acid shock during 18 h in TSB acidified with HCl (D56°C of 3.0 ± 0.1 min) and the lowest for the acid-shocked C7927 strain treated for 4 h in TSB acidified with HCl (D56°C of 0.45 ± 0.06 min). Acidulants did not alter the heat tolerance of strain C7927 (D56°C of 1.9 ± 0.1 min; P0.05) but significantly affected strain 43895 (P0.05), showing the greatest tolerance when malic acid was used (D56°C of 3.7 ± 0.3 min). A significant interaction between strain, storage temperature, and physiological state was noted during the survival experiments (P0.05). E. coli O111 was the most resistant strain, surviving 6 and 23 days at 24 and 1°C, respectively. Our findings may assist in designing challenge studies for juices and other pH-controlled products, where Shiga toxin-producing E. coli represents the pathogen of concern.

Identification of management practices associated with preharvest pathogen contamination of produce fields is crucial to the development of effective Good Agricultural Practices (GAPs). A cross-sectional study was conducted to (i) determine management practices associated with a Salmonella - or Listeria monocytogenes -positive field and (ii) quantify the frequency of these pathogens in irrigation and nonirrigation water sources. Over 5 weeks, 21 produce farms in New York State were visited. Field-level management practices were recorded for 263 fields, and 600 environmental samples (soil, drag swab, and water) were collected and analyzed for Salmonella and L. monocytogenes . Management practices were evaluated for their association with the presence of a pathogen-positive field. Salmonella and L. monocytogenes were detected in 6.1% and 17.5% of fields ( n = 263) and 11% and 30% of water samples ( n = 74), respectively. The majority of pathogen-positive water samples were from nonirrigation surface water sources. Multivariate analysis showed that manure application within a year increased the odds of a Salmonella -positive field (odds ratio [OR], 16.7), while the presence of a buffer zone had a protective effect (OR, 0.1). Irrigation (within 3 days of sample collection) (OR, 6.0), reported wildlife observation (within 3 days of sample collection) (OR, 6.1), and soil cultivation (within 7 days of sample collection) (OR, 2.9) all increased the likelihood of an L. monocytogenes -positive field. Our findings provide new data that will assist growers with science-based evaluation of their current GAPs and implementation of preventive controls that reduce the risk of preharvest contamination.

Based on current U.S. Food and Drug Administration acidified foods guidelines, regulatory approval of commercial pickled egg production without a final heat treatment requires challenge studies. We conducted challenge studies to verify common pickled egg processing parameters. Hard-boiled eggs were acidified in ambient temperature brine at a 60:40 egg/brine ratio. Four acidification treatments were studied in triplicate: 5% acetic acid (AA) or 2.5% AA brine with and without 0.05% sodium benzoate. These treatments resulted in 2% or 1% AA with or without 0.02% sodium benzoate, respectively, in the total system. Samples were stored at 7 °C until pH at the yolk center was ≤ 4.6; subsequently, samples were held at ambient temperature. Egg pH was measured at 24- to 48-h intervals until equilibrium pH was reached (4.0 and 4.4). Eggs and jar lids were challenged with separate pathogen cocktails (six strains and/or serovars) of Salmonella enterica, Escherichia coli O157:H7, Listeria monocytogenes, and Staphylococcus aureus. After 5 and 9 days, the pH fell below 4.6 in 2% AA and 1% AA eggs, respectively. Sodium benzoate did not affect acidification rate for these brine treatments (P ≥ 0.05), nor did sodium benzoate affect pathogen die-off. E. coli O157:H7, Salmonella, and L. monocytogenes were undetectable (

Antimicrobial peptides are produced across all domains of life. Among these diverse compounds, those produced by bacteria have been most successfully applied as agents of biocontrol in food and agriculture. Bacteriocins are ribosomally synthesized, proteinaceous compounds that inhibit the growth of closely related bacteria. Even within the subcategory of bacteriocins, the peptides vary significantly in terms of the gene cluster responsible for expression, and chemical and structural composition. The polycistronic gene cluster generally includes a structural gene and various combinations of immunity, secretion, and regulatory genes and modifying enzymes. Chemical variation can exist in amino acid identity, chain length, secondary and tertiary structural features, as well as specificity of active sites. This diversity posits bacteriocins as potential antimicrobial agents with a range of functions and applications. Those produced by food-grade bacteria and applied in normally occurring concentrations can be used as GRAS-status food additives. However, successful application requires thorough characterization.

Application of manure or soil amendments of animal origin (untreated soil amendments; UTSAs) to agricultural land has been a long-standing practice to maintain or improve soil quality through addition of organic matter, nitrogen, and phosphorus. Much smaller quantities of these types of UTSAs are applied to land used for food crops than to land used for animal grain and forage. UTSAs can harbor zoonotic enteric pathogens that may survive for extended periods after application. Additional studies are needed to enhance our understanding of preharvest microbial food safety hazards and control measures pertaining to the application of UTSAs especially for land used to grow produce that may be consumed raw. This document is intended to provide an approach to study design and a framework for defining the scope and type of data required. This document also provides a tool for evaluating the strength of existing data and thus can aid the produce industry and regulatory authorities in identifying additional research needs. Ultimately, this framework provides a means by which researchers can increase consistency among and between studies and facilitates direct comparison of hazards and efficacy of controls applied to different regions, conditions, and practices.

Consumer confidence in the food industry is severely affected by large-scale spoilage incidents. However, relatively little research exists on spoilage potential of members of the fungal subphylum Mucormycotina (e.g. Mucor), which includes dimorphic spoilage organisms that can switch between a yeast-like and hyphal phase depending on environmental conditions. The presence of Mucor circinelloides in yogurt may not cause spoilage, but growth and subsequent changes in quality (e.g. container bloating) can cause spoilage if not controlled. The purpose of this study was to evaluate the effects on M. circinelloides of pasteurization regimen, natamycin concentrations, and storage temperature in yogurt production, as measured by fungal proliferation and carbon dioxide production. A strain of M. circinelloides isolated from commercially spoiled yogurt showed greater yogurt-spoilage potential than clinical isolates and other industrial strains. D-values and z-values were determined for the spoilage isolate in milk as an evaluation of the fungus' ability to survive pasteurization. Natamycin was added to yogurt at 0, 5, 10, 15, and 20ppm (μg/ml) to determine its ability to inhibit M. circinelloides over the course of month-long challenge studies at 4°C, 15°C, and 25°C. Survivors were recovered on acidified PDA and carbon dioxide levels were recorded. The D-values at 54°C, 56°C, and 58°C for hyphae/sporangiospores were (in min) 38.31±0.02, 10.17±0.28, and 1.94±0.53, respectively, which yielded a z-value of 3.09°C. The D-values at 51°C, 53°C, and 55°C for yeast-like cells were (in min) 14.25±0.12, 6.87±1.19, and 2.44±0.35, respectively, which yielded a z-value of 0.34°C. These results indicated that M. circinelloides would not survive fluid milk pasteurization if contamination occurred prior to thermal treatment. CO2 production was only observed when M. circinelloides was incubated under low-oxygen conditions, and occurred only at temperatures above 4°C. Addition of 10ppm and greater of natamycin inhibited the growth and CO2 production of M. circinelloides under moderate temperature abuse when compared to the untreated control. These data suggest that yogurt spoilage (container bloating) caused by anaerobic growth of M. circinelloides is due to post-pasteurization contamination. Temperature abuse facilitated spoilage as CO2 production was observed in yogurt incubated at 15°C and 25°C, but not at 4°C. The addition of at least 10ppm of natamycin prevented M. circinelloides growth in both hyphal and yeast-like phases, as well as CO2 production in temperatures of up to 15°C for 30days.

Foodborne illnesses associated with fresh produce continue to be a major concern as consumer demand for healthier and nonthermally processed food increases. The objective of this study was to evaluate vaporized ethyl pyruvate (EP; CAS 617-35-6) as a safe alternative antimicrobial agent for the decontamination of Escherichia coli O157:H7 on green onions and spinach. Baby spinach leaves and green onions were inoculated with a five-strain cocktail of E. coli O157:H7 (pGFP) by the dipping method. Samples were treated with concentrations of 0, 42, 105, and 420 mg/liter vaporized EP in a 2.6-liter enclosed container. The efficacy of EP vapors for reducing E. coli O157:H7((GFP)) populations on green onions and baby spinach at 4 and 10°C was monitored for 7 and 5 days, respectively. The lowest EP concentration (42 mg/liter) resulted in a 1.7-log reduction of E. coli O157:H7((GFP)) on green onions after 7 days at 4°C and a 1.9-log reduction after 5 days at 10°C (P0.05). In baby spinach, the same concentration resulted in 0.9-log and 1.4-log reductions (P0.05) of E. coli O157:H7((GFP)) after 7 days at 4°C and 5 days at 10°C, respectively. On green onions, the highest concentration of EP (420 mg/liter) reduced the population of E. coli O157:H7((GFP)) by4.7 log CFU/g after 7 days at 4°C and 5 days at 10°C. The same concentration was also effective for reducing E. coli O157:H7((GFP)) populations in baby spinach by 4.3 log CFU/g after 7 days at 4°C and by6.5 log CFU/g after 3 days at 10°C. Although the successful EP treatments minimally affected the sensory attributes of green onions, the treatments resulted in significant changes in the sensory attributes of baby spinach samples stored at 4 and 10°C. These results indicate that EP is an effective antimicrobial that could be used to enhance the safety of fresh produce depending on the sensory characteristics of the product.

This study reports a novel class of antifungal protein derived from bacterial origin. Bacillus thuringiensis SF361, the strain also responsible for producing the novel bacteriocin thurincin H, exhibits broad antifungal activity against select members of several fungal genera, including Aspergillus , Byssochlamys , and Penicillium , as well as the pathogenic yeast Candida albicans . Optimal antifungal production and secretion were observed after-log phase growth when incubated at 37°C in a carbohydrate-free growth medium. High-performance liquid chromatography purification was performed after pH-selective ammonium sulfate precipitation and size-exclusion chromatography. Intact mass analysis and peptide mass fingerprinting identified the 13,484-Da protein to be a mass homolog to the YvgO protein construct sequenced from Bacillus cereus AH 1134. Further analysis via amino-terminal sequencing also revealed the existence of four distinct yet equally efficacious YvgO variants differing only within the first four N-terminal residues. YvgO was found to be remarkably stable, maintaining its antifungal activity under a wide pH and temperature range. When assayed against the toxigenic species Byssochlamys fulva H25, the selected primary filamentous fungal indicator, the MIC was estimated to be 1.5 ppm. Candida albicans 3153 was more resistant, exhibiting MICs between 25 and 800 ppm, depending on growth conditions. YvgO is unique among antifungals, showing no known sequential or functional homology to the typical classes of antifungal proteins, including common membrane-acting agents such as cellulases and glucanases. Due to its activity against an array of pathogenic and spoilage fungi, the potentials for clinical, agricultural, and food-processing applications are encouraging.

Clostridium pasteurianum BB, a saccharolytic and spore-forming obligate anaerobe, was isolated and identified from shelf-stable apple juice that was responsible for multiple large spoilage outbreaks. The growth and sporulation conditions of C. pasteurianum were atypical compared with those previously published. C. pasteurianum spores were heat resistant in apple juice at pH 3.80, with D-values at 80, 85, and 90°C being 34.4, 15.9, and 4.4 min, respectively, and a z-value of 11°C. The survival curves for thermal inactivation obeyed linear first-order kinetics. Apple juice with varying pH values was used to determine the effect of pH on germination capability of C. pasteurianum spores. The spores were found to be able to germinate at pH as low as 4.3 in pH-adjusted apple juice at low contamination levels. It was confirmed by PCR that C. pasteurianum isolated from spoiled apple juice did not contain the genes for botulinum toxins B and E, which were more commonly found in neurotoxigenic butyric clostridia. Control of finished-juice pH to below 4.0 in combination with mild heating was proposed to prevent potential spoilage of shelf-stable apple juice made with spore-contaminated apple juice concentrate.

The presence of the mycotoxin patulin in processed apple juice and cider presents a continual challenge to the food industry as both consumer health and product quality issues. Although several methods for control and/or elimination of patulin have been proposed, no unifying method has been commercially successful for reducing patulin burdens while maintaining product quality. In the present study, exposure to germicidal UV radiation was evaluated as a possible commercially viable alternative for the reduction and possible elimination of the patulin mycotoxin in fresh apple cider. UV exposure of 14.2 to 99.4 mJ/cm2 resulted in a significant and nearly linear decrease in patulin levels while producing no quantifiable changes in the chemical composition (i.e., pH, Brix, and total acids) or organoleptic properties of the cider. For the range of UV doses tested, patulin levels decreased by 9.4 to 43.4%; the greatest reduction was achieved after less than 15 s of UV exposure. The method of UV radiation (the CiderSure 3500 system) is an easily implemented, high-throughput, and cost-effective method that offers simultaneous UV pasteurization of cider and juice products and reduction and/or elimination of patulin without unwanted alterations in the final product.

Enterococcus mundtii CUGF08, a lactic acid bacterium isolated from alfalfa sprouts, was found to produce mundticin L, a new class IIa bacteriocin that has a high level of inhibitory activity against the genus Listeria . The plasmid-associated operons containing genes for the mundticin L precursor, the ATP binding cassette (ABC) transporter, and immunity were cloned and sequenced. The fifth residue of the conservative consensus sequence YGNGX in the mature bacteriocin is leucine instead of valine in the sequences of the homologous molecules mundticin KS (ATO6) and enterocin CRL35. The primary structures of the ABC transporter and the immunity protein are homologous but unique.

The effectiveness of several wash treatments was evaluated against spores of Penicillium expansum inoculated on six varieties of apples (Red Delicious, Golden Supreme, Empire, Macintosh, Fuji, and Gala). The wash treatments were water, acidified water (pH 6.5), acidified sodium hypochlorite (pH 6.5), nonacidified sodium hypochlorite (pH 8.8, 9.3, and 9.7; 50, 100, and 200 ppm, respectively), and peracetic acid (50 and 80 ppm). Spores of P. expansum were dried on the surface of the apples for 2 h before exposure to the different sanitizer solutions. Each apple was submerged in 100 ml of each treatment solution for 30 s, and the number of spores remaining were recovered and enumerated. The efficacy of chlorine solutions was enhanced by decreasing the pH to 6.5 (up to 5-log reduction, depending on apple variety). Peracetic acid solutions (50 and 80 ppm) resulted in a reduction of less than 2 log spores per g and had the same efficacy (P < or = 0.05) as nonacidified chlorine solutions (50, 100, and 200 ppm). Control water solutions produced a reduction of 1.34 log spores per g. Chlorine solutions at pH 6.5 resulted in the largest reduction of P. expansum spores for all apple varieties tested.

Alfalfa seeds inoculated with five strains of Salmonella or Escherichia coli O157:H7 were subjected to dry heat at 55 degrees C for up to 8 days. Five-log reductions in Salmonella or E. coli O157:H7 on seeds were observed. No pathogens were detected on the sprouted seeds, which were initially inoculated with ca. 2 log CFU/g of Salmonella or more than 8 log CFU/g of E. coli O157:H7. The percentages of germination of the alfalfa seeds did not significantly decrease after 6 days of heating at 55 degrees C. These results showed that heat treatment of alfalfa seeds at 55 degrees C for up to 6 days was effective in enhancing the safety of alfalfa sprouts without affecting germination significantly.

Heat-resistant fungal spores present a processing challenge for beverages and fruit juices, as thermal and UV strategies are often inadequate in reducing heat-resistant fungal burdens to acceptable levels. While effective against pathogenic or invasive bacteria, germicidal UV light treatments also fail to achieve an appreciable reduction of heat-resistant fungal spores. As an alternative, the efficacy of the antifungal protein YvgO was examined across a selection of fruit juices and teas, as well as solid model matrices. Compared with its efficacy in analogous liquid matrices, the apparent efficacy of YvgO was diminished on acidified solid matrices due to a reduction in YvgO diffusion. Using an XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] tetrazolium dye cytotoxicity assay, the effective concentrations to reduce growth by 50% were elucidated in samples challenged with Byssochlamys fulva H25. The MICs were determined and ranged from 2 ppm in apple juice and acidified teas to approximately 3 to 12 ppm for lemonade and orange, white cranberry, blueberry, prune, cherry, and grape juices. Apple cider and nonacidified teas showed reduced efficacy, with MICs exceeding 100 ppm. Tannin-rich products readily removed YvgO from the product, impairing its efficacy. Adding bovine serum albumin as a competitive inhibitor effectively reversed the YvgO-tannin association and restored efficacy in black but not green tea matrices. When challenged with a 5-log CFU inoculum of B. fulva, the shelf lives of the products were extended for various times up to 28 days in a concentrationdependent manner. However, initial efficacy was not predictive of shelf life extension, as some products exhibited improved protection at just two- and fourfold concentrations above the MIC, while others only exhibited long-term stability when concentrations exceeded 20 times the MIC. As such, YvgO may be an attractive alternative to currently available protection strategies and will provide needed diversity for natural food protectants.

Thermal pasteurization can achieve the U. S. Food and Drug Administration-required 5-log reduction of pathogenic Escherichia coli O157:H7 and Cryptosporidium parvum in apple juice and cider, but it can also negatively affect the nutritional and organoleptic properties of the treated products. In addition, thermal pasteurization is only marginally effective against the acidophilic, thermophilic, and spore-forming bacteria Alicyclobacillus spp., which is known to cause off-flavors in juice products. In this study, the efficiency of a combined microfiltration (MF) and UV process as a nonthermal treatment for the reduction of pathogenic and nonpathogenic E. coli, C. parvum, and Alicyclobacillus acidoterrestris from apple cider was investigated. MF was used to physically remove suspended solids and microorganisms from apple cider, thus enhancing the effectiveness of UV and allowing a lower UV dose to be used. MF, with ceramic membranes (pore sizes, 0.8 and 1.4 μm), was performed at a temperature of 10 °C and a transmembrane pressure of 155 kPa. The subsequent UV treatment was conducted using at a low UV dose of 1.75 mJ/cm(2). The combined MF and UV achieved more than a 5-log reduction of E. coli, C. parvum, and A. acidoterrestris. MF with the 0.8-μm pore size performed better than the 1.4-μm pore size on removal of E. coli and A. acidoterrestris. The developed nonthermal hurdle treatment has the potential to significantly reduce pathogens, as well as spores, yeasts, molds, and protozoa in apple cider, and thus help juice processors improve the safety and quality of their products.

Twenty-seven honey samples from different floral sources and geographical locations were evaluated for their ability to inhibit the growth of seven food spoilage organisms (Alcaligenes faecalis, Aspergillus niger, Bacillus stearothermophilus, Geotrichum candidum, Lactobacillus acidophilus, Penicillium expansum, Pseudomonas fluorescens) and five foodborne pathogens (Bacillus cereus, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica Ser. Typhimurium, and Staphylococcus aureus) using an overlay inhibition assay. They were also tested for specific activity against S. aureus 9144 and B. stearothermophilus using the equivalent percent phenol test--a well diffusion assay corresponding to a dilute phenol standard curve. Honey inhibited bacterial growth due to high sugar concentration (reduced water activity), hydrogen peroxide generation, and proteinaceous compounds present in the honey. Some antibacterial activity was due to other unidentified components. The ability of honey to inhibit the growth of microorganisms varies widely, and could not be attributed to a specific floral source or demographic region produced in this study. Antibacterially active samples in this study included Montana buckwheat, tarweed, manuka, melaleuca, and saw palmetto. Furthermore, the bacteria were not uniformly affected by honey. Varying sensitivities to the antimicrobial properties were observed with four strains of S. aureus thus emphasizing the variability in the antibacterial effect of honey samples. Mold growth was not inhibited by any of the honeys tested. B. stearothermophilus, a heat-resistant spoilage bacteria, was shown to be highly sensitive to honey in both the overlay and well diffusion assays; other sensitive bacteria included A. faecalis and L. acidophilus. Non-peroxide antibacterial activity was observed in both assays; the highest instance was observed in the specific activity assay against B. stearothermophilus. Further research could indicate whether honey has potential as a preservative in minimally processed foods.

Salmonella enterica serovars and Escherichia coli O157:H7 have been associated with contaminated seed sprout outbreaks. The majority of these outbreaks have been traced to sprout seeds contaminated with low levels of pathogens. E. coli O157:H7 strains can grow an average of 2.3 log CFU/g over 2 days during seed germination, and Salmonella can achieve an average growth of 3.7 log CFU/g. Therefore, it is important to find an effective method to reduce possible pathogenic bacterial populations on the seeds prior to sprouting. Our objective was to assess the effectiveness of various dry heat treatments on reducing E. coli O157:H7 and Salmonella populations on mung beans intended for sprout production and to determine the effect of these treatments on seed germination. Mung beans were inoculated with five-strain cocktails of E. coli O157:H7 and of Salmonella serovars harboring the green fluorescent protein gene and then air dried overnight. Heat treatments were performed by incubating the seeds at 55 degrees C for various periods of time. Heat-treated seeds were then assessed for the efficacy of the heat treatment and the effects of heat treatment on germination rates. After inoculation and drying, 6 log CFU/g E. coli O157:H7 and 4 log CFU/g Salmonella were detected on the seeds. Following heat treatment, pathogenic bacterial populations on the seeds were below detectable levels (

Salmonella accounts for approximately 50% of produce-associated outbreaks in the United States, several of which have been traced back to contamination in the produce production environment. To quantify Salmonella diversity and aid in identification of Salmonella contamination sources, we characterized Salmonella isolates from two geographically diverse produce-growing regions in the United States. Initially, we characterized the Salmonella serotype and subtype diversity associated with 1,677 samples collected from 33 produce farms in New York State (NYS). Among these 1,677 samples, 74 were Salmonella positive, yielding 80 unique isolates (from 147 total isolates), which represented 14 serovars and 23 different pulsed-field gel electrophoresis (PFGE) types. To explore regional Salmonella diversity associated with production environments, we collected a smaller set of samples ( n = 65) from South Florida (SFL) production environments and compared the Salmonella diversity associated with these samples with the diversity found among NYS production environments. Among these 65 samples, 23 were Salmonella positive, yielding 32 unique isolates (from 81 total isolates), which represented 11 serovars and 17 different PFGE types. The most common serovars isolated in NYS were Salmonella enterica serovars Newport, Cerro, and Thompson, while common serovars isolated in SFL were Salmonella serovars Saphra and Newport and S. enterica subsp. diarizonae serovar 50:r:z. High PFGE type diversity (Simpson's diversity index, 0.90 ± 0.02) was observed among Salmonella isolates across both regions; only three PFGE types were shared between the two regions. The probability of three or fewer shared PFGE types was Salmonella isolates were considerably different between the two sampled regions. These findings suggest the potential for PFGE-based source tracking of Salmonella in production environments.

Thurincin H is an antimicrobial peptide produced by Bacillus thuringiensis SF361. With a helical back bone, the 31 amino acids of thurincin H form a hairpin structure maintained by four pairs of very unique sulfur-to-α-carbon thioether bonds. The production of thurincin H depends on a putative gene cluster containing 10 open reading frames. The gene cluster includes three tandem structural genes ( thnA1 , thnA2 , and thnA3 ) encoding three identical 40-amino-acid thurincin H prepeptides and seven other genes putatively responsible for prepeptide processing, regulation, modification, exportation, and self-immunity. A homologous thurincin H expression system was developed by transforming a thurincin H-deficient host with a novel expression vector, pGW133. The host, designated B. thuringiensis SF361 Δ thnA1 Δ thnA2 Δ thnA3 , was constructed by deletion of the three tandem structural genes from the chromosome of the natural thurincin H producer. The thurincin H expression vector pGW133 was constructed by cloning the thurincin H native promoter, thnA1 , and a Cry protein terminator into the Escherichia coli - B. thuringiensis shuttle vector pHT315. Thirty-three different pGW133 variants, each containing a different point mutation in the thnA1 gene, were generated and separately transformed into B. thuringiensis SF361 Δ thnA1 Δ thnA2 Δ thnA3 . Those site-directed mutants contained either a single radical or conservative amino acid substitution on the thioether linkage-forming positions or a radical substitution on all other nonalanine amino acids. The bacteriocin activities of B. thuringiensis SF361 Δ thnA1 Δ thnA2 Δ thnA3 carrying different pGW133 variants against three different indicator strains were subsequently compared.

An assay to screen Escherichia coli in foods using MUG supplemented lauryl sulfate tryptose (LST) broth instead of tryptone water (TW) medium was evaluated. The results presented in this paper suggest that this method is more sensitive for lower levels of E. coli, faster (16-18 h vs. 6-10 days) and less expensive (2.454 vs. 2.887 EURO/sample) than the standard ISO procedure. Thus, this method may be beneficial for use when both fecal coliforms and E. coli analyses are required in food systems.

(1999). Escherichia coli O157:H7 As An Emerging Foodborne Pathogen: A Literature Review. Critical Reviews in Food Science and Nutrition: Vol. 39, No. 6, pp. 481-502.

A purified bacteriocin produced by Enterococcus faecium BFE 900 isolated from black olives was shown by Edman degradation and mass spectrometric analyses to be identical to enterocin B produced by E. faecium T136 from meat (P. Casaus, T. Nilsen, L. M. Cintas, I. F. Nes, P. E. Hernández, and H. Holo, Microbiology 143:2287–2294, 1997). The structural gene was located on a 2.2-kb Hin dIII fragment and a 12.0-kb Eco RI chromosomal fragment. The genetic characteristics and production of EntB by E. faecium BFE 900 differed from that described so far by the presence of a conserved sequence like a regulatory box upstream of the EntB gene, and its production was constitutive and not regulated. The 2.2-kb chromosomal fragment contained the hitherto undetected immunity gene for EntB in an atypical orientation that is the reverse of that of the structural gene. Typical transport and other genes associated with bacteriocin production were not detected on the 12.0-kb chromosomal fragment containing the EntB structural gene. This makes the EntB genetic system different from most other bacteriocin systems, where transport and possible regulatory genes are clustered. EntB was subcloned and expressed by the dedicated secretion machinery of Carnobacterium piscicola LV17A. The structural gene was amplified by PCR, fused to the divergicin A signal peptide, and expressed by the general secretory pathway in Enterococcus faecalis ATCC 19433.

Attachment and biofilm formation by bacterial pathogens on surfaces in natural, industrial, and hospital settings lead to infections and illnesses and even death. Minimizing bacterial attachment to surfaces using controlled topography could reduce the spreading of pathogens and, thus, the incidence of illnesses and subsequent human and financial losses. In this context, the attachment of key microorganisms, including Escherichia coli , Listeria innocua , and Pseudomonas fluorescens , to silica and alumina surfaces with micron and nanoscale topography was investigated. The results suggest that orientation of the attached cells occurs preferentially such as to maximize their contact area with the surface. Moreover, the bacterial cells exhibited different morphologies, including different number and size of cellular appendages, depending on the topographical details of the surface to which they attached. This suggests that bacteria may utilize different mechanisms of attachment in response to surface topography. These results are important for the design of novel microbe-repellant materials.

Produce-related outbreaks have been traced back to the preharvest environment. A longitudinal study was conducted on five farms in New York State to characterize the prevalence, persistence, and diversity of food-borne pathogens in fresh produce fields and to determine landscape and meteorological factors that predict their presence. Produce fields were sampled four times per year for 2 years. A total of 588 samples were analyzed for Listeria monocytogenes , Salmonella , and Shiga toxin-producing Escherichia coli (STEC). The prevalence measures of L. monocytogenes , Salmonella , and STEC were 15.0, 4.6, and 2.7%, respectively. L. monocytogenes and Salmonella were detected more frequently in water samples, while STEC was detected with equal frequency across all sample types (soil, water, feces, and drag swabs). L. monocytogenes sigB gene allelic types 57, 58, and 61 and Salmonella enterica serovar Cerro were repeatedly isolated from water samples. Soil available water storage (AWS), temperature, and proximity to three land cover classes (water, roads and urban development, and pasture/hay grass) influenced the likelihood of detecting L. monocytogenes . Drainage class, AWS, and precipitation were identified as important factors in Salmonella detection. This information was used in a geographic information system framework to hypothesize locations of environmental reservoirs where the prevalence of food-borne pathogens may be elevated. The map indicated that not all croplands are equally likely to contain environmental reservoirs of L. monocytogenes . These findings advance recommendations to minimize the risk of preharvest contamination by enhancing models of the environmental constraints on the survival and persistence of food-borne pathogens in fields.

To explore the possibility of using Oenococcus oeni to inhibit foodborne pathogens, and to characterize antimicrobial compounds produced by O. oeni, 24 strains of O. oeni were tested for their ability to inhibit growth of foodborne pathogens, Escherichia coli O157:H7, Salmonella enteritidis, and Listeria monocytogenes by using the spot-on-lawn method. Of the 24 strains, 17 strains were able to inhibit all 3 pathogens in this study. Proteases, catalase, and buffer solutions were used for determining the type of inhibitory compounds produced from 4 selected strains with stronger inhibitory activity. Antimicrobial activity of 2 strains against the pathogens was completely inactivated by buffer solution, and other 2 strains against E. coli O157:H7 were partially removed. The antimicrobial compound was not sensitive to selected proteases and catalase. Practical Application: There is little information available about using O. oeni for human pathogens control. The results of this study revealed such discovery and potential applications for pathogen control.

Carnobacteriocin A is a hydrophobic nonlantibiotic bacteriocin that is detected early in the growth cycle of Carnobacterium piscicola LV17A and encoded by a 49 MDa plasmid. The bacteriocin was purified using hydrophobic interaction and gel filtration chromatography, and reversed-phase HPLC. Three different active peaks (A1, A2 and A3) were detected, but the purified samples had identical N-terminal amino acid sequences for the first 15 amino acids as determined by Edman degradation analysis. Only a 2.4 kb fragment of the EcoRI digest of the plasmid pCP49 hybridized with a 23-mer oligonucleotide probe derived from amino acids 5 to 13 of the amino acid sequence. The structural gene for carnobacteriocin A is located 600 base pairs into the 2.4 kb EcoRI fragment, but no other genetic information was detected on this unit. The structural gene includes an 18 amino acid N-terminal extension of the bacteriocin, ending with Gly-Gly residues in the -2, -1 positions with respect to the cleavage site. The bacteriocin consists of 53 amino acids that differ markedly from the majority of hydrophobic peptide bacteriocins characterized to date. Based on the amino acid sequence derived from the nucleotide sequence a molecular mass of 5052.85 Da was calculated. Mass spectrometric analysis showed that the molecular mass of the major component (A3) was 2 Da lower, thereby indicating the presence of a disulphide bridge between Cys 22 and Cys 51. Carnobacteriocin A2 has a similar structure except that Met 52 is oxidized to a sulphoxide, whereas A1 appears to be a mixture of peptides derived proteolytically from A3 or A2.

Alicyclobacillus spp. is an important thermoacidophilic, spore-forming spoilage bacterium that is a major concern for beverage and juice industries. In order to develop effective control strategies and adequately address the prevalence of contamination sources, it is necessary to characterize Alicyclobacillus' ecology in fruit, juice and beverage production and processing environments. Alicyclobacillus spp. isolates were collected from juice, beverage, ingredients, and environmental samples over a period of ten years. A total of 141 isolates were characterized as Alicyclobacillus spp. by 16S rRNA analysis and the most frequently isolated species was found to be Alicyclobacillus acidoterrestris (45%), A. acidocaldarius subsp. acidocaldarius (30%), and A. acidocaldarius (11%). The majority of thermotolerant sporeformers isolated from apple juices and concentrates was found to be A. acidoterrestris (24 out of 36 total apple isolates); while A. acidoterrestris was most frequently associated with citrus, citrus concentrates, and their associated environments, isolated by University of Florida (UF) (15 out of total 28 UF citrus isolates). However, A. acidocaldarius and subsp. acidocaldarius were frequently isolated by Cornell University (CU) (29 out of 35 CU citrus isolates), from citrus juices made from concentrate. Four major haplotypes of Alicyclobacillus spp. were identified based on the 16S rRNA gene sequencing from the 141 isolates tested. The Allelic Types (ATs) matched the phylogenetic analysis grouping of the different Alicyclobacillus spp. based on the isolation source. Our results suggest a predisposition for certain ATs of Alicyclobacillus spp. depending on juice or ingredient isolation source.