Salmonella and Listeria monocytogenes survival on Field Packed Cantaloupe Contact Surfaces.

Field-packing of cantaloupes involves numerous food contact surfaces that can contamination melons with foodborne pathogens; the soil on these surfaces increases throughout the harvest day. Data is lacking on the cross-contamination risk from contaminated food contact surfaces under the dry conditions typical of cantaloupe field-packing operations. This study sought to evaluate the survival of Salmonella and Listeria monocytogenes on cantaloupe field-pack food contact surfaces using both a wet and dry inoculum to provide insights into managing foodborne pathogen contamination risks. Five clean or fouled materials (cotton gloves, nitrile gloves, rubber gloves, cotton rags, and stainless steel) were inoculated with a cocktail of either Salmonella or L. monocytogenes. A wet inoculum was spot inoculated (100 µL) onto coupons. A dry inoculum was prepared by mixing wet inoculum with 100 g of sterile sand, and shaking the coupons with the inoculated sand for 2min. Coupons were held at 35°C (35% RH) and enumerated at 0, 2, 4, 6 and 8 h. Significant differences in pathogen concentrations over time were calculated and the GInaFiT add-in tool for Excel was used to build Log-linear, Weibull, and Biphasic die-off models. Depending on the material type, coupon condition, and inoculum type, Salmonella and L. monocytogenes reductions over 8 h ranged from 0.3-3.3 and -0.4-4.2 log10 CFU/coupon, respectively. For all material types, Salmonella reductions were highest on wet-inoculated clean coupons; L. monocytogenes varied by material type. Weibull and biphasic models were a better fit of respective pathogen die-off curves than linear models. Overall, faster die-off rates were seen for wet inoculated and clean materials. Since pathogen populations remained viable over the study duration and both inoculum type and coupon condition impacted survival, frequent sanitation or replacement of food contact surfaces during the operational day is needed to reduce the risk of cross-contamination.

Fate and Growth Kinetics of Salmonella and Listeria monocytogenes on Mangoes During Storage.

Imported mangoes have been linked to outbreaks of salmonellosis in the USA. The purpose of this research was to evaluate the persistence and growth kinetics of Salmonella and Listeria monocytogenes on the intact surface of whole 'Ataulfo', 'Kent', and 'Tommy Atkins' mangoes stored at three different temperatures. L. monocytogenes was also evaluated on fresh-cut 'Tommy Atkins' mangoes stored at 4, 12, 20 ± 2°C. Whole mangoes were spot inoculated with rifampicin-resistant pathogen cocktails (6 log CFU/mango) onto the midsection of whole fruit (n = 6). Fruit was stored at 12, 20, or 30 ± 2°C and sampled for up to 28 days. The specific growth rates derived from DMFit models as a function of time were used to develop secondary models. On 'Kent' mangoes, Salmonella had a population increase from 0.3 to 1.1 log CFU/mango with a linear growth rate of ∼0.004, 0.01, and 0.06 log CFU/mango/h at 12, 20, and 30°C, respectively. At 20 and 30°C, Salmonella growth rates were significantly higher than 12°C (P < 0.05). No clear Salmonella growth trend was observed; populations decreased up to 1.6 log CFU/mango on 'Tommy Atkins' and 'Ataulfo' at 12°C. Populations of L. monocytogenes on whole and fresh-cut mangoes declined regardless of temperature and storage period. Food safety during storage should be the top priority for fresh-cut tropical fruit processors.

Salmonella transfer potential between tomatoes and cartons used for distribution.

Corrugated fiberboard boxes (cartons) can be reused during fresh market tomato packing and repacking. The fate of Salmonella on the new, used, and dirty tomato packaging cartons, and Salmonella transfer between tomatoes and new, used, and dirty packaging cartons was assessed. Mature green tomatoes or blank cartons were spot inoculated with cocktail of rifampicin-resistant Salmonella strains before touching cartons/tomatoes at 0, 1, or 24 h postinoculation. Tomatoes were placed on new, used, and dirty carton squares (5 by 5 cm) for 0, 1, and 7 days of contact at 12°C and 25°C with a relative humidity value of 85%. Transfer coefficients (TCs) were calculated for all conditions. Salmonella populations decreased following inoculation by 2-3 log units during 24 h drying regardless of storage temperature; the presence of debris enhanced survival at 12°C. In general, the highest transfer rates occurred with wet inoculum. The highest Salmonella transfer was calculated for wet inoculated tomatoes with 7 days of contact time at 25°C (TC = 14.7). Increasing contact time decreased TCs for new cartons, but increased TCs for used and dirty cartons. Regardless of carton condition or storage temperature, a greater population of Salmonella was transferred from tomatoes to cartons than from cartons to tomatoes. Salmonella transfer between tomatoes and cartons is highly dependent on moisture, with increased levels of moisture increasing transfer, highlighting the importance of harvesting and packing dry tomatoes.

The prevalence and concentration of Salmonella enterica in poultry litter in the southern United States.

Poultry litter is applied to crop production land in the southern United States as a waste management strategy as it is a nitrogen-rich fertilizer and plentiful throughout the region. While litter is a known reservoir for human enteric pathogens including Salmonella enterica, little is known regarding pathogen prevalence, concentration, and common serotypes within the material. Litter from thirteen farms across four southern states was examined for Salmonella. Samples (n = 490) from six of the thirteen (46.2%) farms tested positive. Thirty-three samples out of 490 (6.7%) were Salmonella positive. Salmonella was ca. 95% less likely to be collected from stacked litter piles than from the poultry house floor or pasture, and every day increase in litter age reduced the likelihood of recovering Salmonella by 5.1%. When present, concentrations of Salmonella in contaminated poultry litter were variable, ranging from <0.45 to >280,000 MPN/g. The most prevalent serotypes found were Kentucky (45.5%), Kiambu (18.2%), and Michigan (12.1%). Salmonella Kentucky also had the greatest distribution and was found on 4 of the 6 (66.7%) positive farms. Results from this survey demonstrated that Salmonella prevalence and concentration in poultry litter is highly variable, and good agricultural practices are critical to safely use poultry litter as a soil amendment on fresh produce fields.

Prevalence of Listeria monocytogenes and indicator microorganisms in Florida cantaloupe packinghouses, 2013-2014.

Prior to the 2013 cantaloupe season, the US Food and Drug Administration notified the industry that inspections of a subset of packinghouses would commence that year in response to the 2011 Listeria monocytogenes outbreak associated with cantaloupe. In May 2013, five Florida cantaloupe packinghouses participated in an environmental monitoring survey to evaluate their sanitary conditions prior to a potential FDA inspection. Two facilities participated again in 2014. Surface swabs (n = 374) were collected in each facility and included up to 60 food contact and non-food contact surfaces, including water. Samples were enumerated for total plate counts (TPC), generic Escherichia coli, and coliforms, and enriched for Listeria. Listeria were confirmed and speciated by sequencing of the partial sigB gene, and further characterized by pulsed field gel electrophoresis (AscI and Apal). In 2013, two zone 1 surfaces in same facility, were positive for L. monocytogenes (2/233). No L. monocytogenes was detected (n = 103) in the two facilities sampled the following year, including the previously L. monocytogenes-positive facility. Prevalence of L. monocytogenes in FL cantaloupe packinghouses was generally low (2/374), compared to other food environments. TPC, coliforms, E. coli and Listeria spp. were poor indicators of L. monocytogenes contamination in Florida packinghouses.

Quantification of Salmonella enterica transfer between tomatoes, soil, and plastic mulch.

Tomatoes have been linked to Salmonella outbreaks in the United States (US). Plasticulture systems, that combine raised beds, plastic mulch, drip irrigation and fumigation, are common in commercial staked fresh tomato production in the US. The US FDA Produce Safety Rule prohibits the distribution of any produce covered by the rule (including fresh market tomatoes) that drops to the ground before harvest. This research was undertaken to better characterize the risks posed by tomatoes that touch plastic mulch or soil immediately before or during harvest. Research was conducted in three states (Florida, Maryland, and Ohio). Each state utilized tomatoes from their state at the point of harvest maturity most common in that state. Each state used indigenous soil and plastic mulch for transfer scenarios. New plastic mulch obtained directly from the application roll and used plastic mulch that had been present on beds for a growing season were evaluated. A five-strain cocktail of Salmonella enterica isolates obtained from tomato outbreaks was used. Mulch (new or used), soil, or tomatoes were spot inoculated with 100 µl of inoculum to obtain a final population of ~6 log CFU/surface. Items were either touched to each other immediately (1-2 s) after inoculation (wet contact) or allowed to dry at ambient temperature for 1 h or 24 h (dry contact). All surfaces remained in brief (1-5 s) or extended (24 h) contact at ambient temperature. Transfer of Salmonella between a tomato and plastic mulch or soil is dependent on contact time, dryness of the inoculum, type of soil, and contact surface. Transfer of Salmonella to and from the mulch and tomatoes for wet and 1 h dry inocula were similar with mean log % transfers varying from 0.7 ± 0.2 to 1.9 ± 0.1. The transfer of Salmonella between soil or plastic mulch to and from tomatoes was dependent on moisture with wet and 1 h dry inocula generally yielding significantly (p < 0.05) higher transfer than the 24 h dry inoculum. Results indicate that harvesting dry tomatoes significantly (p < 0.05) reduces the risk of contamination from soil or mulch contact. Transfer to tomatoes was generally significantly greater (p < 0.05) from new and used plastic mulch than from soil. If contamination and moisture levels are equivalent and contact times are equal to or <24 h before harvest, significantly (p < 0.05) more Salmonella transfers to tomatoes from mulch than from soil. Our findings support that harvesting tomatoes from soil has similar or lower risk than harvesting from plastic mulch.

Reduction of Escherichia coli, as a surrogate for Salmonella spp., on the surface of grapefruit during various packingline processes.

The US Produce Safety Rule allows for use of water that does not meet its microbial standards if corrective measures are employed. This research was initiated to determine the suitability of nonpathogenic Escherichia coli as a surrogate for Salmonella during citrus washing, and to evaluate the removal of E. coli from grapefruit on two pilot packinglines (CREC and IRREC) as corrective measures. Whole grapefruit were inoculated with either E. coli or Salmonella and dried, and exposed to a variety of treatments on a lab-scale brush wash system. Individual processes were evaluated on the pilot packinglines with E. coli only. In all lab-scale brush wash system treatments, bacterial population reductions between E. coli and Salmonella were not significantly different (P ≤ 0.05). On pilot packinglines, E. coli populations were reduced by 3.59 to >5.11 log CFU/grapefruit at the CREC packingline, and by 3.30 to >5.13 log CFU/grapefruit at the IRREC packingline. Treatment of fruit through complete packingline processing at both locations reduced E. coli populations to levels below the detection limit (<1 log CFU/grapefruit). The studies indicate E. coli is an appropriate surrogate for Salmonella under tested conditions, and that standard citrus packingline processes can be used as a corrective measure.

Quantification of Transfer of Salmonella from Citrus Fruits to Peel, Edible Portion, and Gloved Hands during Hand Peeling.

Although studies have quantified bacterial transfer between hands and various materials, cross-contamination between the surface of fresh citrus fruit and the edible portions during hand peeling has not been reported. This study quantifies transfer of Salmonella to the edible portion of citrus fruit from a contaminated peel during hand peeling. Citrus fruits used for this study were Citrus sinensis (sweet orange) cultivars 'Valencia' and 'Navel', Citrus unshiu (Satsuma mandarins), Citrus reticulata × Citrus paradisi ('Minneola' tangelo or 'Honeybell'), and C. paradisi (grapefruit) cultivar 'Marsh'. An avirulent Salmonella Typhimurium LT2 (ATCC 700720) resistant to rifampin was used for all experiments. The inoculum containing approximately 9 log CFU/mL (50 µL) was spot inoculated onto the equator, stem, or styler of each fruit and allowed to dry for 24 h. Six volunteers put on single-use latex gloves and peeled inoculated fruit. Peel, edible fruit portion, and gloves were collected and enumerated separately. Three replicates of the study were performed in which each volunteer peeled two inoculated fruit of each variety (n = 36 fruit per variety). Cross-contamination from contaminated surface of citrus fruits to edible portion or gloved hands during peeling was affected by inoculation sites. Average Salmonella transfer to the edible portion ranged from 0.16% (Valencia inoculated at the equator) to 5.41% (navel inoculated at the stem). Average Salmonella transfer to gloved hands ranged from 0.41% (grapefruit inoculated at the stem) to 8.97% (navel inoculated at the stem). Most Salmonella remained on the peel of citrus fruits. The average level of Salmonella remaining on the peel ranged from 5.37% (Minneola inoculated at the equator) to 66.3% (Satsuma inoculated at the styler). When grapefruit was inoculated, the Salmonella that remained on the peel showed a bimodal pattern in which some individuals left almost all Salmonella on the peel, while others left substantially less.

Influence of Temperature Differential between Tomatoes and Postharvest Water on Salmonella Internalization.

Salmonella bacteria may internalize into tomato pulp when warm tomatoes from the field are submerged into colder water. Several washing steps may follow the initial washing and packing of tomatoes at the packinghouses; the potential for internalization into tomatoes in subsequent washing steps when tomatoes have a cooler pulp temperature is unknown. Our objective was to evaluate Salmonella internalization into mature green and red tomatoes with ambient (21°C) and refrigeration (4°C) pulp temperatures when they were submerged into water at various temperature differentials, simulating repacking and fresh-cut operations. Red (4°C and 21°C) and mature green (21°C) tomatoes were submerged (6 cm) into a six-strain Salmonella cocktail (6 log CFU/ml) and maintained at ±5 and 0°C temperature differentials for varying time intervals, ranging from 30 s to 5 min. Following submersion, tomatoes were surface sterilized using 70% ethanol, the stem abscission zone and blossom end epidermis were removed, and cores were recovered, separated into three segments, and analyzed. Salmonella populations in the segments were enumerated by most probable number (MPN). The effects of temperature differential and maturity on Salmonella populations were analyzed; results were considered significant at a P value of ≥0.5. Internalized populations were not significantly different (P ≥0.5) across temperature differentials. Salmonella internalization was seen in tomatoes under all treatment conditions and was highest in the segment immediately below the stem abscission zone. However, populations were low (typically >1 log MPN per segment) and varied greatly across temperature differentials. This suggests that the temperature differential between tomatoes and water beyond the initial packinghouse may be less important than submersion time in Salmonella internalization.

Survival of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes on raw peanut and pecan kernels stored at -24, 4, and 22°C.

Cocktails of lawn-collected cells were used to determine the survival of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes on the surface of raw peanut and pecan kernels. Kernels were inoculated with mixtures of four to five strains at 3 or 6 log CFU/g, dried at room temperature, and then stored at -24 ± 1, 4 ± 2, and 22 ± 1°C for 28 or 365 days. In most cases, rates of decline of the pathogens did not differ significantly between the two inoculum concentrations in the 28-day study. At 6 log CFU/g, populations of all pathogens were reduced by 0.5 to 1.6 log CFU/g during an initial 3-day drying period on both peanuts and pecans. The moisture content of peanuts and pecans remained stable at -24 ± 1 and 22 ± 1°C; at 4 ± 2°C, the moisture content increased from 3.8 to 5.6% on peanuts and from 2.6 to 3% on pecans over 365 days. Pathogen populations were stable on pecans stored under frozen and refrigerated conditions, except for L. monocytogenes, which declined at a rate of 0.03 log CFU/g/30 days at 4 ± 2°C. Salmonella populations were stable on peanuts stored at -24 ± 1 and 4 ± 2°C, but E. coli O157:H7 and L. monocytogenes declined at rates of 0.03 to 0.12 log CFU/g/30 days. At 22 ± 1°C, Salmonella, E. coli O157:H7, and L. monocytogenes declined at a rate of 0.22, 0.37, and 0.59 log CFU/g/30 days, respectively, on peanuts, and at 0.15, 0.34, and 1.17 log CFU/g/30 days, respectively, on pecans. Salmonella counts were above the limit of detection (0.30 log CFU/g) throughout the study. In most cases during storage, counts obtained from pecans were higher than from peanuts.

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.

Fate of Escherichia coli O157:H7 and Salmonella on whole strawberries and blueberries of two maturities under different storage conditions.

Strawberries and blueberries harvested at or near full-ripe maturity tend to be less firm and more susceptible to bruising during harvest and transport. The objective of this research was to determine the fate of Escherichia coli O157:H7 and Salmonella on bruised and intact surfaces of whole strawberries and blueberries at shipping (2°C) and retail display (15.5°C) temperatures. Strawberries and blueberries were either purchased from a supermarket or were harvested immediately prior to use; they were bruised using established protocols, were spot inoculated, and were incubated at 2 and 15.5°C. Strawberries, subjected to modified atmospheres, were further transferred to bags and were sealed in with an initial atmosphere of ca. 10% CO2 and 5% O2. Strawberries were sampled at 0, 2, 5, and 24 h and on days 3 and 7; blueberries were sampled on days 0, 1, 3, and 7. After stomaching, samples were enumerated on nonselective and selective media, and populations were recorded as log CFU per berry. At both storage temperatures, population declines for both E. coli O157:H7 and Salmonella were seen under all conditions for strawberries. At 2 ± 2°C, E. coli O157:H7 and Salmonella populations on blueberries declined over 7 days under all conditions. At 15.5 ± 2°C, E. coli O157:H7 populations declined; however, Salmonella populations initially declined but increased to populations near or above initial populations over 7 days on blueberries. No overall significant differences were observed between bruised and intact treatments or between the two maturity levels for strawberries and blueberries. Modified atmospheric conditions did not affect the behavior of E. coli O157:H7 and Salmonella on strawberries at both temperatures. This research indicates that E. coli O157:H7 and Salmonella do not grow on strawberries at shipping or retail display temperatures, even when they are harvested at a maturity prone to bruising; however, Salmonella growth may occur on bruised full ripe blueberries under retail display temperatures.

Modeling the growth of Listeria monocytogenes on cut cantaloupe, honeydew and watermelon.

A recent outbreak linked to whole cantaloupes underscores the importance of understanding growth kinetics of Listeria monocytogenes in cut melons at different temperatures. Whole cantaloupe, watermelon, and honeydew purchased from a local supermarket were cut into 10 ± 1 g cubes. A four-strain cocktail of L. monocytogenes from food related outbreaks was used to inoculate fruit, resulting in ~10(3) CFU/10 g. Samples were stored at 4, 10, 15, 20, or 25 °C and L. monocytogenes were enumerated at appropriate time intervals. The square root model was used to describe L. monocytogenes growth rate as a function of temperature. The model was compared to prior models for Salmonella and Escherichia coli O157:H7 growth on cut melon, as well as models for L. monocytogenes on cantaloupe and L. monocytogenes ComBase models. The current model predicts faster growth of L. monocytogenes vs. Salmonella and E. coli O157:H7 at temperatures below 20 °C, and agrees with estimates from ComBase Predictor, and a corrected published model for L. monocytogenes on cut cantaloupe. The model predicts ~4 log CFU increase following 15 days at 5 °C, and ∼1 log CFU increase following 6 days at 4 °C. The model can also be used in subsequent quantitative microbial risk assessments.

Quantifying transfer rates of Salmonella and Escherichia coli O157:H7 between fresh-cut produce and common kitchen surfaces.

Cross-contamination between foods and surfaces in food processing environments and home kitchens may play a significant role in foodborne disease transmission. This study quantifies the cross-contamination rates between a variety of fresh-cut produce and common kitchen surfaces (ceramic, stainless steel, glass, and plastic) using scenarios that differ by cross-contamination direction, surface type, produce type, and drying time/moisture level. A five-strain cocktail of rifampin-resistant Salmonella was used in transfer scenarios involving celery, carrot, and watermelon, and a five-strain cocktail of rifampin-resistant Escherichia coli O157:H7 was used in transfer scenarios involving lettuce. Produce or surface coupons were placed in buffer-filled filter bags and homogenized or massaged, respectively, to recover cells. The resulting solutions were serially diluted in 0.1% peptone and surface plated onto tryptic soy agar with 80 µg/ml rifampin and bismuth sulfite agar with 80 µg/ml rifampin for Salmonella or sorbitol MacConkey agar with 80 µg/ml rifampin for E. coli O157:H7. When the food contact surface was freshly inoculated, a high amount (>90%) of the inoculum was almost always transferred to the cut produce item. If the inoculated food contact surfaces were allowed to dry for 1 h, median transfer was generally >90% for carrots and watermelon but ranged from <1 to ∼70% for celery and lettuce. Freshly inoculated celery or lettuce transferred more bacteria (<2 to ∼25% of the inoculum) compared with freshly inoculated carrots or watermelon (approximately <1 to 8%). After 1 h of drying, the rate of transfer from inoculated celery, carrot, and lettuce was <0.01 to ∼5% and <1 to ∼5% for watermelon. Surface moisture and direction of transfer have the greatest influence on microbial transfer rates.

Development and validation of a mathematical model for growth of pathogens in cut melons.

Many outbreaks of foodborne illness associated with the consumption of fresh-cut melons have been reported. The objective of our research was to develop a mathematical model that predicts the growth rate of Salmonella on fresh-cut cantaloupe over a range of storage temperatures and to validate that model by using Salmonella and Escherichia coli O157:H7 on cantaloupe, honeydew, and watermelon, using both new data and data from the published studies. The growth of Salmonella on honeydew and watermelon and E. coli O157:H7 on cantaloupe, honeydew, and watermelon was monitored at temperatures of 4 to 25°C. The Ratkowsky (or square-root model) was used to describe Salmonella growth on cantaloupe as a function of storage temperature. Our results show that the levels of Salmonella on fresh-cut cantaloupe with an initial load of 3 log CFU/g can reach over 7 log CFU/g at 25°C within 24 h. No growth was observed at 4°C. A linear correlation was observed between the square root of Salmonella growth rate and temperature, such that √growth rate = 0.026 × (T - 5.613), R(2) = 0.9779. The model was generally suitable for predicting the growth of both Salmonella and E. coli O157:H7 on cantaloupe, honeydew, and watermelon, for both new data and data from the published literature. When compared with existing models for growth of Salmonella, the new model predicts a theoretic minimum growth temperature similar to the ComBase Predictive Models and Pathogen Modeling Program models but lower than other food-specific models. The ComBase Prediction Models results are very similar to the model developed in this study. Our research confirms that Salmonella can grow quickly and reach high concentrations when cut cantaloupe is stored at ambient temperatures, without visual signs of spoilage. Our model provides a fast and cost-effective method to estimate the effects of storage temperature on fresh-cut melon safety and could also be used in subsequent quantitative microbial risk assessments.

Growth of Clostridium perfringens during cooling of refried beans.

Outbreaks of Clostridium perfringens have been associated with dishes containing refried beans from food service establishments. However, growth of C. perfringens in refried beans has not been investigated, and predictive models have not been validated in this food matrix. We investigated the growth of C. perfringens during the cooling of refried beans. Refried beans (pinto and black, with and without salt added) were inoculated with 3 log CFU/g C. perfringens spores and incubated isothermally at 12, 23, 30, 35, 40, 45, and 50°C. The levels of C. perfringens were monitored 3, 5, 8, and 10 h after inoculation, and then fitted to the Baranyi primary model and the Rosso secondary model prior to solving the Baranyi differential equation. The final model was validated by dynamic cooling experiments carried out in stockpots, thus mimicking the worst possible food service conditions. All refried beans samples supported the growth of C. perfringens, and all models fit the data with pseudo-R(2) values of 0.95 or greater and mean square errors of 0.3 or lower. The estimated maximum specific growth rates were generally higher in pinto beans, with or without salt added (2.64 and 1.95 h(-1), respectively), when compared with black beans, with or without salt added (1.78 and 1.61 h(-1), respectively). After 10 h of incubation, maximum populations of C. perfringens were significantly higher in samples with no salt added (7.9 log CFU/g for both pinto and black beans) than in samples with salt added (7.3 and 7.2 log CFU/g for pinto and black beans, respectively). The dynamic model predicted the growth of C. perfringens during cooling, with an average root mean squared error of 0.44. The use of large stockpots to cool refried beans led to an observed 1.2-log increase (1.5-log increase predicted by model) in levels of C. perfringens during cooling. The use of shallower pans for cooling is recommended, because they cool faster, therefore limiting the growth of C. perfringens.

Prevalence, concentration, spoilage, and mitigation of Alicyclobacillus spp. in tropical and subtropical fruit juice concentrates.

The presence of Alicyclobacillus in fruit juices and concentrates poses a serious problem for the juice industry. This study was undertaken to determine the (i) prevalence, concentration, and species of Alicyclobacillus in tropical and subtropical concentrates; (ii) efficacy of aqueous chlorine dioxide in reducing Alicyclobacillus spp. spores on tropical and subtropical fruit surfaces; and (iii) fate of and off-flavor production by Alicyclobacillus acidoterrestris in mango and pineapple juices. One hundred and eighty tropical and subtropical juice concentrates were screened for the presence and concentration of Alicyclobacillus spp. If found, the species of Alicyclobacillus was determined by 16S rDNA sequencing and analysis with NCI BLAST. Of these samples, 6.1% were positive for Alicyclobacillus, and nine A. acidoterrestris strains and two Alicyclobacillus acidocaldarius strains were identified. A five-strain cocktail of Alicyclobacillus spp. was inoculated onto the surface of fruits (grapefruit, guava, limes, mangoes, oranges and pineapple), which were then washed with 0, 50, or 100 ppm aqueous chlorine dioxide. Significant reductions due to chlorine dioxide were only seen on citrus fruits. A five-strain cocktail of A. acidoterrestris was inoculated into mango and pineapple juices. Microbial populations were enumerated over a 16-day period. Aroma compounds in the juice were analyzed by GC-olfactometry (GC-O) and confirmed using GC-MS. GC-O of mango juice identified previously reported medicinal/antiseptic compounds. GC-O of pineapple juice revealed an unexpected "cheese" off-aroma associated with 2-methylbutyric acid and 3-methylbutyric acid.

Natural-light labeling of tomatoes does not facilitate growth or penetration of Salmonella into the fruit.

The survival-growth capacity of Salmonella populations on tomato epidermis labeled by a natural-light labeling system was investigated after persistent fears of such marks serving as possible entryways for the pathogenic organisms, alone and in the presence of Pectobacterium carotovorum subsp. carotovorum, a soft-rot organism. Different treatments involving natural-light labeling, fruit waxing, and a five-strain cocktail of Salmonella were applied to mature green tomato surfaces in different sequences prior to storage at 4, 12, or 25°C. Fruit was sampled every 3 days, and Salmonella was enumerated from all treatments and unlabeled fruit, which served as controls. There were no significant differences between treatments or between treatments and controls throughout. The results indicate that the cuticle and epidermal interruptions caused by natural-light labeling do not facilitate the penetration and colonization of the tomato pericarp. In a separate set of experiments, the capacity of Salmonella to penetrate tomato in the presence of a potential synergism with P. carotovorum subsp. carotovorum was investigated. The addition of P. carotovorum at higher, lower, or equal population densities to Salmonella did not significantly alter the behavior of Salmonella on tomatoes stored at 25°C, regardless of natural-light labeling. The inability of P. carotovorum and Salmonella to colonize natural-light-etched surfaces of tomato fruit indicates that the use of this technology does not adversely compromise the surface of tomatoes.

Mitigation of Alicyclobacillus spp. spores on food contact surfaces with aqueous chlorine dioxide and hypochlorite.

The prevalence of Alicyclobacillus spp. and other spore-forming spoilage organisms in food handling and processing environments presents a sanitation challenge to manufacturers of products such as juices and beverages. The objectives of this study were to determine the efficacy of chlorine dioxide and sodium hypochlorite in killing Alicyclobacillus spores in situ and to evaluate the efficacy of various chlorine dioxide and hypochlorite sanitizing regimes on Alicyclobacillus spp. spores on stainless steel, wood, and rubber conveyor material. Five or two log CFU/ml spore concentrations were left in aqueous solution or inoculated onto stainless steel, rubber, or wood coupons and challenged with sanitizer for varied time intervals. After treatment, the coupons were placed in sterile sample bags, massaged with neutralizing buffer, and enumerated on Ali agar. Surfaces were also examined before and after treatment by scanning electron microscopy to confirm destruction or removal of the spores. For both five and two log CFU/ml spore concentrations, treatments of 50 and 100 ppm of chlorine dioxide and 1000 and 2000 ppm of hypochlorite, respectively, were the most effective. Of the range of chlorine dioxide concentrations and contact time regimes evaluated for all surfaces, the most effective concentration/time regime applied was 100 ppm for 10 min. Reductions ranged from 0 to 4.5 log CFU/coupon. Chlorine dioxide was least effective when applied to wood. Hypochlorite was not efficient at eliminating Alicyclobacillus spores from any of the food contact surfaces at any time and concentration combinations tested. Chlorine dioxide is an alternative treatment to kill spores of Alicyclobacillus spp. in the processing environment.

Utilization of fluorogenic assay for rapid detection of Escherichia coli in acidic fruit juice.

This study was undertaken to investigate interference by acids commonly found in fruit juice in Escherichia coli assays involving the use of 4-methylumbelliferyl-beta-D-glucuronide (MUG) as a fluorogenic substrate for enzyme reaction. Fluorescence intensity was negatively correlated (P < 0.001) with the volume of fresh citrus juice tested by the lauryl tryptose broth (LST)-MUG assay, and the permissible sample sizes were limited to 0.3 and 0.5 ml for fresh citrus juices with pHs of 3.3 and 3.9, respectively. In addition, false-negative results were visually observed under UV light when the E*Colite assay was used to test large volumes (5 to 10 ml per test) of fresh citrus juice or when the test broth used for the LST-MUG assay was supplemented with citric, malic, or tartaric acid at 2 to 4 g/liter. These results suggest that the size and pH of acidic samples should be controlled in MUG-based fluorogenic assays. The inhibitory effect on fluorescence was due to high acidity, which reduces fluorescence from 4-methylumbelliferone. Buffering improved the assays. When sodium bicarbonate was incorporated in the enrichment broth at 10 g/liter, the permissible sample sizes for fresh grapefruit juice (pH 3.1) increased from 0.3 to 1 ml for the LST-MUG (with 9.9 ml of broth) assay and from 3 to 10 ml for the E*Colite (with 99 ml of broth) assay.