Method of Inoculation Influences the Survival of Salmonella enterica on Retail and Orchard Peaches.

Challenge studies associated with fruits and vegetables generally utilize wet bacterial inoculation methods. However, a recent salmonellosis outbreak in the U.S. was linked to peaches plausibly contaminated via fugitive dust from a nearby animal operation. This outbreak has highlighted the need for a suitable inert carrier which can be used for the dry transfer of Salmonella enterica to produce. The purpose of this study was 1) to examine the population stability of S. enterica and its surrogate, Enterococcus faecium, in different dry matrices during extended storage to identify suitable carriers and 2) to evaluate the survival of S. enterica on peaches based on the mode of contamination (i.e., wet vs. dry). S. enterica and E. faecium were cultivated on tryptic soy agar (TSA) and inoculated into corn-cob small animal litter, sand, or silica at 10-11 log CFU/g. Matrices were mixed by hand and stored at 25°C and 33% relative humidity for up to 120 d. S. enterica remained relatively stable in the silica and litter, with no significant decrease in population after 14 and 28 d, respectively. E. faecium significantly reduced in all matrices, with the greatest reduction observed in silica (2.86 log CFU/g after 120 d). Additional carriers would need to be assessed for E. faecium which could maintain its population stability. Silica was ultimately selected for the dry carrier of S. enterica. Peaches available at retail or from orchards were inoculated with S. enterica using the silica carrier or by spot or dip inoculation methods at 5 log CFU/peach and stored at 5°C and 80% relative humidity for up to 28 d. The population of S. enterica significantly reduced on all peaches except for the dry inoculated orchard peaches, where the population remained stable (4.62 ± 0.35 log CFU/peach after 28 d). Results from this study determined that the mode of contamination influences the survival of S. enterica on peaches and that dry inoculation methods should be considered for produce in some instances.

Dynamics of Listeriamonocytogenes and Salmonella enterica on Cooked Vegetables During Storage.

Fresh vegetables have been linked to multiple foodborne outbreaks in the U.S., with Listeria monocytogenes and Salmonella enterica identified as leading causes. Beyond raw vegetables, cooked vegetables can also pose food safety concerns due to improper cooking temperature and time combinations or postcooking contamination. Cooked vegetables, having had their native microbiota reduced through heat inactivation, might provide an environment that favors the growth of pathogens due to diminished microbial competition. While the risks associated with raw vegetables are recognized, the survival and growth of pathogens on cooked vegetables remain inadequately studied. This study investigated the growth kinetics of both L. monocytogenes and S. enterica on various cooked vegetables (carrot, corn, onions, green bell pepper, and potato). Vegetables were cooked at 177°C until the internal temperature reached 90°C and then cooled to 5°C. Cooled vegetables were inoculated with a four-strain cocktail of either L. monocytogenes or S. enterica at 3 log CFU/g, then stored at different temperatures (5, 10, or 25°C) for up to 7 days. Both pathogens survived on all vegetables when stored at 5°C. At 10°C, both pathogens proliferated on all vegetables, with the exception of L. monocytogenes on pepper. At 25°C, the highest growth rates were observed by both pathogens on carrot (5.55 ± 0.22 and 6.42 ± 0.23 log CFU/g/d for L. monocytogenes and S. enterica, respectively). S. enterica displayed higher growth rates at 25°C compared to L. monocytogenes on all vegetables. Overall, these results bridge the knowledge gap concerning the growth kinetics of both S. enterica and L. monocytogenes on various cooked vegetables, offering insights to further enhance food safety.

Survival of Listeria monocytogenes on Frozen Vegetables during Long-term Storage at -18 and -10°C.

Two recent listeriosis outbreaks have occurred in the United States and Europe due to contaminated individually quick-frozen (IQF) vegetables. While one of the outbreaks was due to frozen vegetables considered ready-to-eat (RTE), the other was linked to frozen corn whose packaging contained cooking instructions and was considered not-ready-to-eat (NRTE). However, consumers may thaw certain frozen vegetables and consume without cooking. Since no data is available on the survivability of L. monocytogenes on IQF vegetables during frozen storage, this study examined the population of six different strains (comprising lineages 1/2a, 1/2b, and 4b) on IQF vegetables during long-term storage. Individual strains were inoculated onto an IQF vegetable mix at 4 log CFU/g and stored at -18 or -10°C for 360 days. Although fluctuations in populations of all strains were observed on the vegetables during storage, no significant differences based on strain, lineages, or temperature were observed. Overall, L. monocytogenes populations were only reduced by up to 0.47 and 0.59 log CFU/g after 360 days at -18 and -10°C, respectively. Results from this study suggest that L. monocytogenes is able to persist on IQF vegetables for extended time periods with no significant reduction in population. Future studies could evaluate the survival and growth of L. monocytogenes on IQF vegetables during thawing and storage.

Effect of dehydration on the inactivation of Listeria monocytogenes and Salmonella enterica on enoki and wood ear mushrooms.

Foodborne illness outbreaks in the U.S. associated with consumption of both fresh and dried specialty mushrooms have recently occurred. Dried wood ear mushrooms were implicated in a salmonellosis outbreak in 2020, while fresh enoki mushrooms were associated with two listeriosis outbreaks in 2020 and 2023. These specialty mushrooms are commercially available in both their fresh and dried states. Due to the short shelf life of mushrooms, dehydration is a common method used in both industry and by consumers to extend the shelf life and preserve quality. Therefore, the aim of this study was to evaluate the use of dehydration on the inactivation kinetics of both Listeria monocytogenes and Salmonella enterica on enoki and wood ear mushrooms. Fresh mushrooms were inoculated with four strain cocktails of either L. monocytogenes or S. enterica and dried at ambient conditions for 10 min. Following drying of the inoculum, mushrooms were placed into food dehydrators preheated to 70, 80, or 90°C and treated for up to 24 h. At treatment intervals, mushrooms were removed from the dehydrators for pathogen enumeration. Inactivation kinetics for both pathogens were modeled using the Weibull, log-linear with tail, and log-linear with shoulder models. Pathogen reductions of >4 log CFU/g were achieved on both enoki and wood ear mushrooms during dehydration at 90°C after only 2-4 h. At 70 and 80°C, log reductions of >4 log CFU/g were observed on wood ear mushrooms after 4-8 h. On enoki mushrooms, a tailing effect was observed with residual populations (>2 log CFU/g) of L. monocytogenes and S. enterica remaining even after 24 h of treatment at both 70 and 80°C. This study emphasizes the need for an individualized dehydration strategy for each mushroom type to ensure the effectiveness of dehydration as a process to reduce pathogen populations. Results of this study will aid in informing proper time and temperature combinations for dehydration of specialty mushrooms to ensure product safety.

Efficacy of Power Ultrasound-Based Hurdle Technology on the Reduction of Bacterial Pathogens on Fresh Produce.

Minimally processed produce is frequently contaminated with foodborne bacterial pathogens. Power ultrasound is a non-thermal and cost-effective technology that can be combined with other chemical sanitization methods. This study investigated the reduction of Listeria monocytogenes and Salmonella Newport on grape tomato, romaine lettuce, and spinach washed with water, chlorine, or peroxyacetic acid alone or in combination with 25 or 40 kHz power ultrasound for 1, 2, or 5 min. Produce items were inoculated with either pathogen at 10 log CFU/g, dried for 2 h, and treated. Combined treatment of ultrasound and sanitizers resulted in 1.44-3.99 log CFU/g reduction of L. monocytogenes and 1.35-3.62 log CFU/g reduction of S. Newport, with significantly higher reductions observed on grape tomato. Synergistic effects were achieved with the hurdle treatment of power ultrasound coupled with the chemical sanitizers when compared to the single treatments; an additional 0.48-1.40 log CFU/g reduction of S. Newport was obtained with the addition of power ultrasound on grape tomato. In general, no significant differences were observed in pathogen reductions between the ultrasound frequencies, the sanitizers, or the treatment lengths. Results from this study suggest that incorporation of power ultrasound into the current washing procedure may be beneficial for the reduction, but not elimination, of bacterial pathogens on certain produce items, including tomatoes.

Growth Kinetics of Listeria monocytogenes and Salmonella enterica on Dehydrated Vegetables during Rehydration and Subsequent Storage.

Dehydrated vegetables have low water activities and do not support the proliferation of pathogenic bacteria. Once rehydrated, vegetables can be incorporated into other foods or held for later use. The aim of this study was to examine the survival and proliferation of Listeria monocytogenes and Salmonella enterica on dehydrated vegetables during rehydration and subsequent storage. Carrots, corn, onion, bell peppers, and potatoes were heat dehydrated, inoculated at 4 log CFU/g, and rehydrated at either 5 or 25 °C for 24 h. Following rehydration, vegetables were stored at 5, 10, or 25 °C for 7 d. Both L. monocytogenes and S. enterica survived on all vegetables under all conditions examined. After 24 h of rehydration at 5 °C, pathogen populations on the vegetables were generally <1.70 log CFU/g, whereas rehydration at 25 °C resulted in populations of 2.28 to 6.25 log CFU/g. The highest growth rates during storage were observed by L. monocytogenes on potatoes and S. enterica on carrots (2.37 ± 0.61 and 1.63 ± 0.18 log CFU/g/d, respectively) at 25 °C when rehydration occurred at 5 °C. Results indicate that pathogen proliferation on the vegetables is both rehydration temperature and matrix dependent and highlight the importance of holding rehydrated vegetables at refrigeration temperatures to hinder pathogen proliferation. Results from this study inform time and temperature controls for the safety of these food products.

Survival kinetics of Listeria monocytogenes and Salmonella enterica on dehydrated enoki and wood ear mushrooms during long-term storage.

Two specialty mushrooms have recently become novel vectors for foodborne outbreaks in the U.S.: fresh enoki and dried wood ear mushrooms were linked to a listeriosis and salmonellosis outbreak, respectively. The aim of this study was to evaluate the survival kinetics of Listeria monocytogenes and Salmonella enterica on dehydrated enoki and wood ear mushrooms during long-term storage. Following heat dehydration, mushrooms were inoculated with either L. monocytogenes or S. enterica, allowed to dry for 1 h, and then stored for up to 180 d at 25 °C and 33% relative humidity. Both pathogens were enumerated from the mushrooms at intervals during the storage period. Survival kinetics of both pathogens were modeled using both the Weibull and log-linear with tail models. After inoculation and 1 h drying, both pathogen populations decreased 2.26-2.49 log CFU/g on wood ear mushrooms; no decrease was observed on enoki. Both pathogens survived during storage on both mushroom types. On wood ear mushrooms, a 2-log decrease of both pathogens occurred during storage. On enoki mushrooms, 4-log decreases of both pathogens were modeled to occur after 127.50-156.60 d. The results of this study suggest that L. monocytogenes and S. enterica can persist on dehydrated specialty mushrooms during long-term storage.

Long-Term Survival of Listeria monocytogenes in Nut, Seed, and Legume Butters.

Nut, seed, and legume butters have become increasingly popular with consumers. Listeria monocytogenes contamination of a variety of butters has resulted in several recalls, although no known outbreaks have been identified. L. monocytogenes has been shown to survive on a variety of seeds for up to 6 months, legumes and nuts for over 12 months, and in peanut butter and peanut-chocolate spreads for 21 to 60 weeks depending on formulation; however, long-term survival in other butters has not yet been characterized. In this study, the survival of L. monocytogenes in various nut, seed, legume, and chocolate-containing butters (n = 10) based on inoculation level, storage temperature, and the pH, aw, and nutrient contents of the butters was examined. First, butters were inoculated with L. monocytogenes at 4 log CFU/g and stored at either 5 or 25°C with enumeration and/or enrichment at intervals over 12 months. L. monocytogenes survived in all butters examined with no significant change in population after storage at 5°C, whereas the population was reduced to <1.70 log CFU/g in as little as 3 months at 25°C; the only exception was for sunflower butter, where L. monocytogenes decreased approximately 1 log CFU/g. Subsequently, all butters were inoculated at 1 log CFU/g and stored at 25°C for 6 months with enrichment during storage. L. monocytogenes was detected in all butters, except pecan butter, after 6-month storage. Butters containing chocolate did not inhibit L. monocytogenes survival, regardless of the inoculation level. Results indicate there may be an association between high-fat and carbohydrate level and survivability of L. monocytogenes in various types of butters. This work highlights the need to mitigate the potential for cross-contamination of L. monocytogenes into nut, seed, and legume butters due to the potential for long-term survival.

Modeling the Fate of Listeria monocytogenes and Salmonella enterica on Fresh Whole and Chopped Wood Ear and Enoki Mushrooms.

Two recent foodborne illness outbreaks linked to specialty mushrooms have occurred in the United States, both representing novel pathogen-commodity pairings. Listeria monocytogenes and Salmonella enterica were linked to enoki and wood ear mushrooms, respectively. The aim of this study was therefore to examine the survival of both L. monocytogenes and S. enterica on raw whole and chopped enoki and wood ear mushrooms during storage at different temperatures. Fresh mushrooms were either left whole or chopped and subsequently inoculated with a cocktail of either S. enterica or rifampicin-resistant L. monocytogenes, resulting in an initial inoculation level of 3 log CFU/g. Mushroom samples were stored at 5, 10, or 25°C for up to 7 d. During storage, the population levels of S. enterica or L. monocytogenes on the mushrooms were enumerated. The primary Baranyi model was used to estimate the growth rates of both pathogens and the secondary Ratkowsky square root model was used to model the relationship between growth rates and temperature. Both L. monocytogenes and S. enterica survived on both mushroom types and preparations at all temperatures. No proliferation of either pathogen was observed on mushrooms stored at 5°C. At 10°C, moderate growth was observed for both pathogens on enoki mushrooms and for L. monocytogenes on wood ear mushrooms; no growth was observed for S. enterica on wood ear mushrooms. At 25°C, both pathogens proliferated on both mushroom types with growth rates ranging from 0.43 to 3.27 log CFU/g/d, resulting in 1 log CFU/g increases in only 0.31 d (7.44 h) to 2.32 d. Secondary models were generated for L. monocytogenes on whole wood ear mushrooms and S. enterica on whole enoki mushrooms with goodness-of-fit parameters of r2 = 0.9855/RMSE = 0.0479 and r2 = 0.9882/RMSE = 0.1417, respectively. Results from this study can aid in understanding the dynamics of L. monocytogenes and S. enterica on two types of specialty mushrooms.

Evaluation of the kitchen microbiome and food safety behaviors of predominantly low-income families.

Bacterial pathogens in the domestic environment present a risk to residents, particularly among susceptible populations. However, the impact of consumer demographic characteristics and food handling methods on kitchen microbiomes is not fully understood. The domestic kitchen bacterial communities of ten predominantly low-income families in Houston, TX, were assessed in conjunction with a cross-sectional food safety survey to evaluate differences in household and surface-specific microbiomes and bacterial foodborne pathogen presence. Three kitchen surfaces within each household, including the sink drain, the refrigerator handle, and the counter, were environmentally sampled and metataxonomically evaluated via targeted 16S rRNA sequencing. Disposable dish sponges were also acquired and examined. Results indicated that alpha diversity did not vary by the households, sampling locations, or demographic characteristics evaluated. Significant differences in beta diversity were observed among the bacterial communities of five pairs of households and between refrigerator handle and disposable dish sponge microbiomes. A total of 89 unique bacterial foodborne pathogens were identified across surface types. Each household contained at least one contaminated surface, and the most common bacterial foodborne pathogens identified were Escherichia coli, Staphylococcus aureus, and Klebsiella pneumoniae. All parents reported washing their hands before meal preparation, washing fresh fruits and vegetables, and washing cutting boards with soap after use to prepare raw animal proteins. Gaps in food safety behaviors identified included a lack of serious concern for food contamination with germs and inappropriate handwashing, food handling, and cleaning behaviors. The number of unique bacterial foodborne pathogens identified within households was significantly higher among households whose respondent parent reported that they did not consider food contamination with germs to be a serious food safety problem (median: 41.0 species) compared to households whose respondent parent did consider food contamination to be a serious food safety problem (median: 3.0 species; p value = 0.0218). These results demonstrate that domestic kitchen taxonomic abundance profiles vary according to household and surface type. Data suggest that low-income consumers may be at risk of foodborne pathogen exposure from contaminated home kitchen surfaces, and that food safety attitudes may directly contribute to this hazard.

Food Safety Attitudes, Behaviors, and Hygiene Measures among Predominantly Low-Income Parents in Houston, Texas.

ABSTRACT: Foodborne infections in the United States affect racial-ethnic minority and low-income populations at higher rates than the general population. To identify the prevalence of food safety behaviors and demographic characteristics associated with food handling practices among a susceptible, high-risk population, a cross-sectional survey was administered to 106 parents with children enrolled at two elementary schools serving predominantly low-income families in Houston, Texas. Relationships between demographic characteristics and food safety behavioral outcomes were examined using cross-tabulations and Fisher's exact test. Most respondents were female (93.4%), Hispanic, Latino, or Mexican American (94.9%), and had no previous food handling employment experience (75.0%). The primary source of food safety information reported was the Internet (32.7%), and nearly half of parents (42.7%) reported that they did not consider contamination of food with germs a serious food safety problem. Hand washing before food preparation was more common (98.0%) than before touching the refrigerator handle (66.3%), after electronic device use (55.6%), or after handling raw animal proteins (77.6%). The prevalence of fresh fruit (98.0%) and vegetable (97.9%) washing and appropriate contaminated cutting board handling (89.0%) was high among parents. Self-reported gaps in food handling behaviors identified included lack of food thermometer ownership (80.4%), use of reusable cleaning tools (71.0%), inappropriate defrosting methods (67.4%), and washing of raw poultry (86.3%), seafood (84.9%), and meat (74.7%). Hand washing after electronic device use and defrosting methods were observed to vary significantly according to demographic characteristics. Food safety education with messages targeted to specific demographic groups may be necessary to reduce the risk of foodborne disease among low-income parents and young children.

Growth Kinetics of Listeria monocytogenes on Cut Red Cabbage.

ABSTRACT: Listeria monocytogenes is a foodborne pathogen associated with fresh produce vectors such as leafy greens. Recent outbreaks and recalls associated with red cabbage-containing salads have brought attention to this food commodity. Although data on the proliferation of L. monocytogenes are available for different varieties of whole and cut white cabbage, no information is available on the fate of this pathogen on red cabbage. The aim of this study was therefore to explore the survival of L. monocytogenes on cut red cabbage (Brassica oleracea L. var. capitata f. rubra) during storage at different temperatures. Red cabbage was cut into pieces (5 by 4 cm) and spot inoculated with a six-strain cocktail of rifampin-resistant L. monocytogenes, resulting in an initial inoculation level of 4 log CFU/g. Samples were stored at 5 or 10°C for 14 days or at 25°C for 7 days. At intervals during storage, L. monocytogenes was enumerated by homogenization of cabbage with Butterfield's phosphate buffer, and serial dilutions were plated onto brain heart infusion agar supplemented with rifampin. No growth of L. monocytogenes was observed on cut red cabbage during storage at 5°C, and only minimal proliferation was observed at the higher temperatures. Significant population increases of 0.58 and 1.07 log CFU/g were determined after 3 days of storage at 10 and 25°C, respectively; however, a significant decrease of 0.77 log CFU/g from 3 to 14 days was also observed at 10°C. The modeled growth rates for L. monocytogenes on cut red cabbage stored at 5 and 10°C were 0.11 ± 0.03 and 0.27 ± 0.07 log CFU/g/day, with calculated times to a 1-log CFU/g increase of 9.51 and 3.70 days, respectively; however, L. monocytogenes did not achieve a 1-log increase at either temperature in this study. At 25°C, the modeled growth rate of L. monocytogenes on cut red cabbage was 1.15 ± 0.36 log CFU/g, leading to a calculated and an observed 1-log increase in 0.87 and 3.00 days, respectively. Results from this study aid in understanding the fate of L. monocytogenes on cut red cabbage during storage at different temperatures.

Desiccation Survival of Salmonella enterica,Escherichia coli, and Enterococcus faecium Related to Initial Cell Level and Cellular Components.

ABSTRACT: Salmonella enterica is well known for its ability to survive and persist in low-moisture environments. Previous studies have indicated a link between the initial cell level and the population of Salmonella that survives after desiccation and subsequent storage; however, how the initial cell concentration affects survival is unknown. This study was conducted to examine this phenomenon and to determine whether it occurs in other microorganisms, specifically Shiga toxigenic Escherichia coli (STEC) and Enterococcus faecium. Salmonella, STEC, and E. faecium were grown as sessile cells on Trypticase soy agar with yeast extract (TSAYE) and harvested in buffered peptone water (BPW). To determine recovery at different initial cell levels, cultures were diluted to 9, 7, and 5 log CFU/mL and applied to filters. Filters were dried for 24 h and then stored for 28 days at 25°C and 33% relative humidity. During storage, cells were recovered from filters with BPW and cultivated on TSAYE. Recovery of both Salmonella and E. coli, but not E. faecium, was nonproportional. Lower initial populations were less viable after 24 h of desiccation; ≥10 log CFU/mL was recovered when 11 log CFU/mL was desiccated, but <3 log CFU/mL was recovered when 5 log CFU/mL was desiccated. Once dried, persistence did not appear affected by initial cell concentration. When inactivated (heat-treated) cells were added to the diluent, recovery of Salmonella was proportional with respect to the initial cell level. To further examine the response to desiccation, Salmonella was diluted in BPW containing 1 of 11 test cell components related to quorum sensing or known to affect desiccation resistance to assess recovery and persistence. Of the 11 additions, only cell debris fractions, cell-free extract, and peptidoglycan improved recovery of Salmonella. Desiccation survival appears related to cell wall components; however, the exact mechanism affecting survival remains unknown.

Microbiomes of commercially-available pine nuts and sesame seeds.

Metagenomic analysis of food is becoming more routine and can provide important information pertaining to the shelf life potential and the safety of these products. However, less information is available on the microbiomes associated with low water activity foods. Pine nuts and sesame seeds, and food products which contain these ingredients, have been associated with recalls due to contamination with bacterial foodborne pathogens. The objective of this study was to identify the microbial community of pine nuts and sesame seeds using targeted 16S rRNA sequencing technology. Ten different brands of each seed type were assessed, and core microbiomes were determined. A total of 21 and 16 unique taxa with proportional abundances >1% in at least one brand were identified in the pine nuts and sesame seeds, respectively. Members of the core pine nut microbiome included the genera Alishewanella, Aminivibrio, Mycoplasma, Streptococcus, and unassigned OTUs in the families of Desulfobacteraceae and Xanthomonadaceae. For sesame seeds, the core microbiome included Aminivibrio, Chryseolina, Okibacterium, and unassigned OTUs in the family Flavobacteriaceae. The microbiomes of these seeds revealed that these products are dominated by environmental bacterial genera commonly isolated from soil, water, and plants; bacterial genera containing species known as commensal organisms were also identified. Understanding these microbiomes can aid in the risk assessment of these products by identifying food spoilage potential and community members which may co-enrich with foodborne bacterial pathogens.

Predictive model for growth of Clostridium perfringens during cooling of cooked pork supplemented with sodium chloride and sodium pyrophosphate.

A dynamic model was developed to predict growth of Clostridium perfringens in cooked ground pork supplemented with salt (0-3% wt/wt) and sodium pyrophosphate (0-0.3% wt/wt) under varying temperatures. C. perfringens (NCTC 8238, NCTC 8239, and NCTC 10240) spores were heat shocked, cooled, and inoculated into ground pork. Isothermal bacterial growth was quantified with variable salt and phosphate concentrations at temperatures ranging from 15 to 51 °C. The primary Baranyi model was fitted to all C. perfringens growth profiles and gave a satisfactory fit (R2 ≥ 0.85). A quadratic polynomial secondary model was developed (P < 0.0001) to predict the maximum specific growth rate as a function of temperature, salt, and phosphate concentrations (R2 = 0.93). A dynamic model was developed and validated using growth data retrieved from 7 published studies. Thirty three out of 44 predictions were within the acceptable prediction zone (-0.5 ≤ prediction error ≤ 1.0). The developed predictive model can be used to minimize the risk of C. perfringens in pork products supplemented with additives during cooling.

Survival and transcriptomic response of Salmonella enterica on fresh-cut fruits.

Salmonella enterica is frequently implicated in foodborne disease outbreaks associated with fresh-cut fruits. In the U.S., more than one third of fruit-related outbreaks have been linked to two S. enterica serotypes Newport and Typhimurium. Approximately 80% of fruit-related human salmonellosis cases were associated with tomatoes, cantaloupes and cucumbers. In this study, we investigated the population dynamics of S. Newport and S. Typhimurium on fresh-cut tomato, cantaloupe, cucumber and apple under short-term storage conditions. We further compared the transcriptomic profiles of a S. Newport strain on fresh-cut tomato and cantaloupe using high-throughput RNA-seq. We demonstrated that both S. enterica Newport and Typhimurium survived well on various fresh-cut fruit items under refrigeration storage conditions, independent of inoculation levels. However, S. enterica displayed variable survival behaviors on different types of fruits. For example, at 7 d storage, the population of S. enterica reduced less than 0.2 log (p > 0.05) on fresh-cut tomato and cantaloupe, in contrast to ~0.5 log (p < 0.05) on cucumber and apple. RNA-seq analysis suggested that S. enterica mediates its survival on fresh-cut fruits through differentially regulating genes involved in specific carbon utilization and metabolic pathways. Several known bacterial virulence factors (e.g., pag gene) were found to be differentially regulated on fresh-cut tomato and cantaloupe, suggesting a link between the events of food contamination and subsequent human infection. Findings from this study contribute to a better understanding of S. enterica survival mechanisms on fresh-cut produce.

Relationship of Growth Conditions to Desiccation Tolerance of Salmonella enterica, Escherichia coli, and Listeria monocytogenes.

ABSTRACT: Growth on solid media as sessile cells is believed to increase the desiccation tolerance of Salmonella enterica. However, the reasons behind increased resistance have not been well explored. In addition, the same effect has not been examined for other foodborne pathogens such as pathogenic Escherichia coli or Listeria monocytogenes. The purpose of this research was twofold: first, to determine the role of oxygenation during growth on the desiccation resistance of S. enterica, E. coli, and L. monocytogenes, and second, to determine the effect of sessile versus planktonic growth on the desiccation resistance of these pathogens. Three different serotypes each of Salmonella, E. coli, and L. monocytogenes were cultured in Trypticase soy broth with 0.6% yeast extract (TSBYE), with (aerobic) shaking or on TSBYE with agar under either aerobic or anaerobic conditions and harvested in the stationary phase. After adding cell suspensions to cellulose filter disks, pathogen survival was determined by enumeration before drying (0 h) and after drying for 24 h. Results showed statistical differences in harvested initial populations before drying. For Salmonella, a correlation was found between high initial population and greater survival on desiccation (P = 0.05). In addition, statistical differences (P ≤ 0.05) between survival based on growth type were identified. However, differences found were not the same for the three pathogens, or between their serotypes. In general, Salmonella and E. coli desiccation resistance followed the pattern of aerobic agar media ≥ liquid media ≥ anaerobic agar media. For L. monocytogenes serotypes, resistance to desiccation was not statistically different based on mode of growth. These results indicate growth on solid media under aerobic conditions is not always necessary for optimal desiccation survival, but may be beneficial when the desiccation resistance of the test serotype is unknown.

Metataxonomic Profiling of Native and Starter Microbiota During Ripening of Gouda Cheese Made With Listeria monocytogenes-Contaminated Unpasteurized Milk.

Unpasteurized milk is used to produce aged artisanal cheeses, which presents a safety concern due to possible contamination with foodborne pathogens, especially Listeria monocytogenes. The objective of this study was to examine the composition of the bacterial community in unpasteurized milk used to prepare Gouda cheese artificially contaminated with L. monocytogenes (~1 log CFU/ml) and assess the community dynamics and their potential interaction with L. monocytogenes during a 90-day ripening period using targeted 16S rRNA sequencing. The diversity of bacterial taxa in three batches of unpasteurized milk was not significantly different, and the microbiomes were dominated by species of Lactococcus, Streptomyces, Staphylococcus, and Pseudomonas. The highest relative abundances were observed for Pseudomonas fluorescens (31.84-78.80%) and unidentified operational taxonomic units (OTUs) of Pseudomonas (7.56-45.27%). After manufacture, both with and without L. monocytogenes-contaminated unpasteurized milk, Gouda cheese was dominated by starter culture bacteria (including Lactococcus lactis subsp. cremoris, lactis, lactis bv. diacetylactis, and Streptococcus thermophilus), in addition to unassigned members in the taxa L. lactis and Streptococcus. During ripening there was an overall decrease in L. lactis abundance and an increase in the number of taxa with relative abundances >0.1%. After 90-day ripening, a total of 82 and 81 taxa were identified in the Gouda cheese with and without L. monocytogenes, respectively. Of the identified taxa after ripening, 31 (Gouda cheese with L. monocytogenes) and 56 (Gouda cheese without L. monocytogenes) taxa had relative abundances >0.1%; 31 were shared between the two types of Gouda cheese, and 25 were unique to the Gouda cheese without added L. monocytogenes. No unique taxa were identified in the Gouda cheese with the added L. monocytogenes. This study provides information on the dynamics of the bacterial community in Gouda cheese during ripening, both with and without the addition of L. monocytogenes.

Effect of Time, Temperature, and Transport Media on the Recovery of Listeria monocytogenes from Environmental Swabs.

ABSTRACT: Environmental monitoring for Listeria monocytogenes in food processing environments is key for ensuring the safety of ready-to-eat foods. For sampling, swabs are often hydrated with a wetting or transport medium that may contain neutralizers and other ingredients. After swabbing the environment, the swabs may then be transported or shipped cold to an off-site laboratory for testing, ideally within 48 h. Extended shipping times may subject the pathogen to increased temperatures in the presence of the wetting medium, organics, and other chemicals from the processing facility that could confound detection. This study evaluated growth and detection of L. monocytogenes on stainless steel exposed to either buffer or sodium hypochlorite before drying. Swabs were rehydrated with Butterfield's phosphate buffer, neutralizing buffer, Letheen broth, or Dey-Engley neutralizing broth before swabbing. Swabs were stored in the presence of no added food, cheese whey, or ice cream under both optimal (4°C) and suboptimal (15°C) temperatures for up to 72 h. Overall, there was no growth of L. monocytogenes at 4°C through 72 h of storage, although enrichment from these swabs was dependent on the presence and type of food matrix. Pathogen growth during storage at 15°C was more variable and depended on both the food matrix and transport media used, with Dey-Engley and Letheen broths allowing for the highest population increases. Overall, more enrichments resulting in L. monocytogenes detections were observed when using Letheen broth and neutralizing buffer than Dey-Engley broth, which resulted in fewer detections at 15°C. Logistic regression and Cochran-Mantel-Haenszel analyses determined that storage temperature, transport media, and food matrix all significantly affected detection of L. monocytogenes, whereas storage time did not have a clear effect on recovery from swabs.

Evaluation of Methods of Enrichment and Compositing of Environmental Samples for Detection of Listeria monocytogenes.

ABSTRACT: Various methods exist for the enrichment and detection of Listeria spp. and Listeria monocytogenes from environmental samples. Procedures for the compositing of environmental samples are not as well defined. In this study, different enrichment procedures involving buffered Listeria enrichment broth (BLEB), University of Vermont medium (UVM), and Fraser broth (FB) were evaluated to determine the limits of detection (LODs) for L. monocytogenes from culture and from swabs of stainless steel and to assess the efficacy of composite sampling by wet (pooling of primary enrichments) and dry (pooling of swabs) procedures. For detection of cells in pure culture, the computed values for the LOD at 95% probability (LOD95) using a single-step BLEB or two-step UVM-FB enrichment were 0.33 and 0.49 CFU/225 mL enrichment, respectively. No significant differences in detection were observed for procedures using either two-step BLEB-FB or UVM-FB enrichments for swabs of stainless steel when L. monocytogenes was inoculated at 2 to 6 log CFU; the LOD95 values were 3.82 and 3.62 log CFU per 4-in2 area, respectively. Wet compositing of L. monocytogenes from culture with and without romaine lettuce wash resident microbiota was conducted using BLEB-FB and UVM-FB enrichment methods; both allowed detection of the pathogen at ratios of 1:1, 1:2, 1:4, and 1:7 (1 positive sample to x negative samples) with no loss in sensitivity. From swabs of stainless steel, L. monocytogenes was detected similarly for both wet and dry composites of up to eight samples (1:7) with romaine lettuce wash. However, the BLEB-FB method allowed significantly faster detection (after 24 h of FB incubation) in composites of 1:4 and 1:7 samples compared with the UVM-FB method under the conditions tested. The results of this study provide data to evaluate the efficacies of the different enrichment procedures and aid in assessing the use of wet and dry compositing of environmental samples for use as part of a Listeria control plan in food production and processing facilities.