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Ground covers are used in produce production to enhance plant growth and control diseases and pests. While various factors are considered when selecting commercial ground covers, food safety, particularly the survival of foodborne pathogens, is often overlooked. This study aimed to assess the survival of Salmonella on different ground covers, including biodegradable mulch, landscape fabric, and plastic mulch. New rolls of each ground cover were cut to fit a 100 × 15 mm petri dish and spot inoculated with a seven-strain Salmonella cocktail at approximately 6 log CFU/cm 2 . The inoculated coupons were stored in a climate-controlled chamber (23°C, 55% relative humidity) and sampled at 0, 0.06 (1.5 h), 0.17 (4 h), 1, 2, 3, 5, 7, 30, 60, 90, and 140 days postinoculation (dpi). If counts dropped below the detection limit (<0.12 log CFU/cm 2 ), enrichments were performed following the Food and Drug Administration's Bacteriological Analytical Manual Salmonella protocol. Salmonella survived 140 dpi on all tested ground covers, with reductions >5 log CFU/cm 2 . Survival rates at 140 dpi were highest on landscape fabric (83%, 25/30) followed by plastic mulch (50%, 15/30) and biodegradable mulch (13%, 4/30) coupons (p < 0.05). During the first 30 dpi, biodegradable mulch exhibited the smallest reduction in Salmonella (2.47 ± 0.26 log CFU/cm 2 ), compared to landscape fabric (3.07 ± 0.30 log CFU/cm 2 ) and plastic mulch (3.86 ± 0.72 log CFU/cm 2 ). After 60 dpi, Salmonella reductions stabilized across all materials (∼4 log CFU/cm 2 ) and by 90 dpi, no significant differences were observed between ground cover types (p > 0.05). Although Salmonella survival varied among ground covers in the short-term (0-30 dpi), a >5 log CFU/cm 2 reduction of Salmonella was observed among all materials by 140 dpi. Findings suggest that ground cover material influences Salmonella survival and should be factored into food safety risk management strategies, especially when ground covers are reused., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 The Authors. Published by Elsevier Inc. All rights reserved.)

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 are 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 2 min. 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 to 3.3 and -0.4 to 4.2 log 10 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., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Biological soil amendments of animal origin (BSAAO) play an important role in agriculture but can introduce pathogens into soils. Pathogen survival in soil is widely studied, but data are needed on the impacts of strain variability and field management practices. This study monitored the population of 12 Escherichia coli strains (generic, O157, and non-O157) in soils while evaluating the interactions of soil type, irrigation regimen, and soil amendment in three independent, greenhouse-based, randomized complete block design trials. Each E. coli strain (4-5 log 10 CFU/g) was homogenized in bovine manure amended or nonamended sandy-loam or clay-loam soil. E. coli was enumerated in 25 g samples on 0, 0.167 (4 h), 1, 2, 4, 7, 10, 14, 21, 28, 56, 84, 112, 168, 210, 252, and 336 days postinoculation (dpi). Regression analyses were developed to understand the impact of strain, soil type, irrigation regimen, and soil amendment on inactivation rates. E. coli survived for 112 to 336 dpi depending on the treatment combination. Pathogenic and generic E. coli survived 46 days [95% Confidence interval (CI) = 20.85, 64.72; p = 0.001] longer in soils irrigated weekly compared to daily and 146 days (CI = 114.50, 184.50; p < 0.001) longer in amended soils compared to unamended soils. Pathogenic E. coli strains were nondetectable 69 days (CI = 39.58, 98.66, p = 0.015) earlier than generic E. coli strains. E. coli inactivation rates demonstrated a tri-phasic pattern, with breakpoints at 26 dpi (CI = 22.3, 29.2) and 130 dpi (CI = 121.0, 138.1). The study findings demonstrate that using bovine manure as BSAAO in soil enhances E. coli survival, regardless of strain, and adequate food safety practices are needed to reduce the risk of crop contamination. The findings of this study contribute data on E. coli concentrations in amended soils to assist stakeholders and regulators in making risk-based decisions on time intervals between the application of BSAAO and the production and harvest of fruits and vegetables., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Limited data exist on the environmental factors that impact pathogen prevalence in the soil. The prevalence of foodborne pathogens, Salmonella and Listeria monocytogenes, and the prevalence and concentration of generic E. coli in Florida's agricultural soils were evaluated to understand the potential risk of microbial contamination at the preharvest level. For all organisms but L. monocytogenes, a longitudinal field study was performed in three geographically distributed agricultural areas across Florida. At each location, 20 unique 5 by 5 m field sampling sites were selected, and soil was collected and evaluated for Salmonella presence (25 g) and E. coli and coliform concentrations (5 g). Complementary data collected from October 2021 to April 2022 included: weather; adjacent land use; soil properties, including macro- and micro-nutrients; and field management practices. The overall Salmonella and generic E. coli prevalence was 0.418% (1/239) and 11.3% (27/239), respectively; with mean E. coli concentrations in positive samples of 1.56 log CFU/g. Farm A had the highest prevalence of generic E. coli, 22.8% (18/79); followed by Farm B, 10% (8/80); and Farm C 1.25% (1/80). A significant relationship (p < 0.05) was observed between generic E. coli and coliforms, and farm and sampling trip. Variation in the prevalence of generic E. coli and changes in coliform concentrations between farms suggest environmental factors (e.g. soil properties) at the three farms were different. While Salmonella was only detected once, generic E. coli was detected in Florida soils throughout the duration of the growing season meaning activities that limit contact between soil and horticultural crops should continue to be emphasized. Samples collected during an independent sampling trip were evaluated for L. monocytogenes, which was not detected. The influence of local environmental factors on the prevalence of indicator organisms in the soil presents a unique challenge when evaluating the applicability of more global models to predict pathogen prevalence in preharvest produce environments., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The Produce Safety Alliance (PSA) grower training was introduced in 2016 as the standardized curriculum to meet the training requirements of the Food and Drug Administration's (FDA) Food Safety Modernization Act's (FSMA) Produce Safety Rule (PSR). The PSR states that at least one supervisor or responsible party from each farm must have successfully completed this food safety training or one equivalent to the standardized curriculum, as recognized by the FDA. This study evaluated the effectiveness of PSA trainings conducted between 2017 and 2019 in the Southern United States by the Southern Regional Center for Food Safety Training, Outreach, and Technical Assistance by analyzing pre- and posttest assessments. Effectiveness was based on a 25-question knowledge assessment administered to participants before (n = 2494) and after (n = 2460) each training. The knowledge assessment indicated the overall effectiveness of the training, with average scores increasing significantly from pretest (15.9/25, 63.4%) to posttest (20.3/25, 81.3%) (P < 0.001). The greatest knowledge gains were seen in the Postharvest Handling and Sanitation, How to Develop a Farm Food Safety Plan, and Agricultural Water modules. Notably, these modules had lower posttest scores compared to the other modules, indicating that the amount of knowledge gained did not necessarily correspond with a sufficient understanding of the material. To ensure that participants understand all aspects of the PSR and best practices to minimize food safety risks, additional or advanced trainings may be needed. Additionally, the current testing instrument (pre-/posttest) used for PSA grower training, while validated, may not be optimal, thus alternative methods to assess the training effectiveness are likely needed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Site-specific approaches for managing food safety hazards in agricultural water require an understanding of foodborne pathogen ecology. This study identified factors associated with Salmonella contamination in Virginia ponds. Grab samples (250 mL, N = 600) were collected from 30 sites across nine ponds. Culture- and culture-independent (CIDT)-based methods were used to detect Salmonella in each sample. Salmonella isolated by culture-based methods were serotyped by Kauffman-White classification. Environmental data were collected for each sample. McNemar's χ 2 was used to determine if Salmonella detection differed by testing method. Separate mixed effect models were used to identify environmental factors associated with culture and CIDT-based Salmonella detection. Separate models were built for each pond, and for all ponds combined. Salmonella detection differed significantly (p < 0.001) between CIDT (31 %; 183/600)- and culture (13 %; 77/600)-based methods. Culture-based methods yielded 11 different serovars. All cultured Salmonella samples were confirmed by CIDT; 42.1 % of CIDT Salmonella-positive samples could be cultured. Associations between environmental factors and Salmonella detection also varied substantially by pond and detection method. In the all-pond model, associations were observed for five factors (total coliforms, Escherichia coli, air temperature, UV, rain) for both culture- and CIDT-based Salmonella detection. Rain prior to sampling (24 h) increased odds of Salmonella detection for culture (OR = 5.09) and CIDT (OR = 3.62) in the all-pond model. When all the pond data were used, models masked associations at the individual pond level, as there were noticeable differences between ponds and the odds of isolating Salmonella by environmental factors. Ponds were within a 187-ha area in this study, emphasizing water management needs to be individualized (i.e., assess hazards/risks by pond). Results also highlight detection methods and scale strongly affect observed water quality and should be considered when developing monitoring programs to develop guidance for growers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Whole genome sequencing (WGS) is a powerful tool that may be used to assist in identifying Listeria contamination sources and movement within environments, and to assess persistence. This study investigated sites in a produce packinghouse where Listeria had been historically isolated; and aimed to characterize dispersal patterns and identify cases of transient and resident Listeria. Environmental swab samples (n = 402) were collected from 67 sites at two time-points on three separate visits. Each sample was tested for Listeria, and Listeria isolates were characterized by partial sigB sequencing to determine species and allelic type (AT). Representative isolates from the three most common L. monocytogenes ATs (n = 79) were further characterized by WGS. Of the 144 Listeria species positive samples (35.8%), L. monocytogenes was the most prevalent species. L. monocytogenes was often coisolated with another species of Listeria. WGS identified cases of sporadic and continued reintroduction of L. monocytogenes from the cold storages into the packinghouse and demonstrated cases of L. monocytogenes persistence over 2 years in cold storages, drains, and on a forklift. Nine distinct clusters were found in this study. Two clusters showed evidence of persistence. Isolates in these two clusters (N = 11, with one historical isolate) were obtained predominantly and over multiple samplings from cold storages, with sporadic movement to sites in the packing area, suggesting residence in cold storages with opportunistic dispersal within the packinghouse. The other seven clusters demonstrated evidence of transient Listeria, as isolation was sporadic over time and space during the packing season. Our data provide important insights into likely L. monocytogenes harborage points and transfer in a packinghouse, which is key to root cause analysis. While results support Listeria spp. as a suitable indicator organism for environmental monitoring surveys, findings were unable to establish a specific species as an index organism for L. monocytogenes. Findings also suggest long-term persistence with substantial SNP diversification, which may assist in identifying potential contamination sources and implementing control measures., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

A broad understanding of community member food safety priorities in the fresh produce supply chain does not currently exist. This information is essential to improve food safety knowledge and practices effectively and efficiently throughout the fresh produce industry; therefore, the goal of this study was to identify and rank community produce safety priorities in the United States. Survey questions were designed and approved by food safety experts for participants to rank 24 fresh produce safety priorities. The anonymous survey was distributed online via Qualtrics™ to fresh produce community members from November 2020 to May 2021. A score was calculated for each priority by summing weighted ranking scores across responses. Descriptive statistics and logistic regression were used to determine frequencies and distribution of response and identify factors (e.g., role in produce safety, size/location of organization/operation) that influenced rankings. A total of 281 respondents represented fourteen different roles in the fresh produce industry, with most identified as growers (39.5%). Produce operations were distributed across the U.S. and annual produce sales ranged from below $25,000 to over $5,000,000. Health and hygiene, training, postharvest sanitation, traceability, and harvest sanitation were ranked as the top five food safety priorities. These findings provide insight into community member priorities in fresh produce safety and can be used to inform intervention efforts, ranging from specialized training for produce growers and packers, industry-driven research projects, and gaps in risk communication strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Produce-borne outbreaks of Shiga toxin-producing Escherichia coli (STEC) linked to preharvest water emphasize the need for efficacious water treatment options. This study quantified reductions of STEC and generic E. coli in preharvest agricultural water using commercially available sanitizers. Water was collected from two sources in Virginia (pond, river) and inoculated with either a seven-strain STEC panel or environmental generic E. coli strain TVS 353 (∼9 log 10 CFU/100 mL). Triplicate inoculated water samples were equilibrated to 12 or 32°C and treated with peracetic acid (PAA) or chlorine (Cl) [low (PAA:6ppm, Cl:2-4 ppm) or high (PAA:10 ppm, Cl:10-12 ppm) residual concentrations] for an allotted contact time (1, 5, or 10 min). Strains were enumerated, and a log-linear model was used to characterize how treatment combinations influenced reductions. All Cl treatment combinations achieved a ≥3 log 10 CFU/100 mL reduction, regardless of strain (3.43 ± 0.25 to 7.05 ± 0.00 log 10 CFU/100 mL). Approximately 80% (19/24) and 67% (16/24) of PAA treatment combinations achieved a ≥3 log 10 CFU/100 mL for STEC and E. coli TVS 353, respectively. The log-linear model showed contact time (10 > 5 > 1 min) and sanitizer type (Cl > PAA) had the greatest impact on STEC and E. coli TVS 353 reductions (p < 0.001). E. coli TVS 353 in water samples was more resistant to sanitizer treatment (p < 0.001) indicating applicability as a good surrogate. Results demonstrated Cl and PAA can be effective agricultural water treatment strategies when sanitizer chemistry is managed. These data will assist with the development of in-field validation studies and may identify suitable candidates for the registration of antimicrobial pesticide products for use against foodborne pathogens in preharvest agricultural water treatment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

No Environmental Protection Agency (EPA) chemical treatments for preharvest agricultural water are currently labeled to reduce human health pathogens. The goal of this study was to examine the efficacy of peracetic acid- (PAA) and chlorine (Cl)-based sanitizers against Salmonella in Virginia irrigation water. Water samples (100 mL) were collected at three time points during the growing season (May, July, September) and inoculated with either the 7-strain EPA/FDA-prescribed cocktail or a 5-strain Salmonella produce-borne outbreak cocktail. Experiments were conducted in triplicate for 288 unique combinations of time point, residual sanitizer concentration (low: PAA, 6 ppm; Cl, 2-4 ppm or high: PAA, 10 ppm; Cl, 10-12 ppm), water type (pond, river), water temperature (12°C, 32°C), and contact time (1, 5, 10 min). Salmonella were enumerated after each treatment combination and reductions were calculated. A log-linear model was used to characterize how treatment combinations influenced Salmonella reductions. Salmonella reductions by PAA and Cl ranged from 0.0 ± 0.1 to 5.6 ± 1.3 log 10 CFU/100 mL and 2.1 ± 0.2 to 7.1 ± 0.2 log 10 CFU/100 mL, respectively. Physicochemical parameters significantly varied by untreated water type; however, Salmonella reductions did not (p = 0.14), likely due to adjusting the sanitizer amounts needed to achieve the target residual concentrations regardless of source water quality. Significant differences (p < 0.05) in Salmonella reductions were observed for treatment combinations, with sanitizer (Cl > PAA) and contact time (10 > 5 > 1 min) having the greatest effects. The log-linear model also revealed that outbreak strains were more treatment-resistant. Results demonstrate that certain treatment combinations with PAA- and Cl-based sanitizers were effective at reducing Salmonella populations in preharvest agricultural water. Awareness and monitoring of water quality parameters are essential for ensuring adequate dosing for the effective treatment of preharvest agricultural water., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Surface water environments are inherently heterogenous, and little is known about variation in microbial water quality between locations. This study sought to understand how microbial water quality differs within and between Virginia ponds. Grab samples were collected twice per week from 30 sampling sites across nine Virginia ponds (n = 600). Samples (100 mL) were enumerated for total coliform (TC) and Escherichia coli (EC) levels, and physicochemical, weather, and environmental data were collected. Bayesian models of coregionalization were used to quantify the variance in TC and EC levels attributable to spatial (e.g., site, pond) versus nonspatial (e.g., date, pH) sources. Mixed-effects Bayesian regressions and conditional inference trees were used to characterize relationships between data and TC or EC levels. Analyses were performed separately for each pond with ≥3 sampling sites (5 intrapond) while one interpond model was developed using data from all sampling sites and all ponds. More variance in TC levels were attributable to spatial opposed to nonspatial sources for the interpond model (variance ratio [VR] = 1.55) while intrapond models were pond dependent (VR: 0.65-18.89). For EC levels, more variance was attributable to spatial sources in the interpond model (VR = 1.62), compared to all intrapond models (VR < 1.0) suggesting that more variance is attributable to nonspatial factors within individual ponds and spatial factors when multiple ponds are considered. Within each pond, TC and EC levels were spatially independent for sites 56-87 m apart, indicating that different sites within the same pond represent different water quality for risk management. Rainfall was positively and pH negatively associated with TC and EC levels in both inter- and intrapond models. For all other factors, the direction and strength of associations varied. Factors driving microbial dynamics in ponds appear to be pond-specific and differ depending on the spatial scale considered., Competing Interests: Declaration of Competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Campylobacter and pathogenic Escherichia coliillnesses have been attributed to the consumption of fresh produce. The leafy green, kale, is increasingly consumed raw. In comparison to other leafy greens, kale has a longer shelf-life. Due to the extended shelf-life of kale, it is warranted to examine the survival of pathogenic Campylobacter jejuni and E. coli O157:H7 inoculated on the surface of kale stored in a controlled environment at 4 ± 1.4°C, and average humidity of 95 ± 1.9% over a 23-day period. At predetermined time points (days 0, 1, 2, 3, 5, 7, 9, 11, 13, 15, 17, 19, and 21), inoculated kale was destructively sampled and the surviving bacteria determined by serial dilution and plating onto Tryptic soy agar, Charcoal cefoperozone deoxycholate agar, and Eosin methylene blue for total aerobic bacteria, C. jejuni, and E. coli O157:H7, respectively. Enrichment and PCR were used for detection when pathogens were not detected using serial dilution and plating. Aerobic heterotrophic bacteria increased over the 23-day period, in contrast, significant declines in the inoculated pathogens were observed. Inoculated E. coli O157:H7 survived longer on kale (up to 19 d); in comparison, C. jejuni was undetectable by day 13 using enrichment and PCR or plating. In conclusion, C. jejuni and E. coli O157:H7 declined on fresh kale over time when held at refrigerated temperatures but were still detected during the majority of the time when the kale would likely still be considered edible by consumers., Competing Interests: Declaration of Competing Interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Listeria species prevalence has been investigated at nearly all stages of the fresh produce supply chain; however, it has not been examined in transportation and distribution center (DC) related environments. Between December 2019 and March 2021, 18 DCs handling fresh produce were environmentally sampled for Listeria. Swab samples were collected from a variety of surfaces (e.g., floors, pallets, forklifts) in several areas of each DC (e.g., cold storage, shipping and receiving docks). Impaction air samples, relative humidity, and temperature data were also collected. While no Listeria spp. were isolated from air samples (n = 170), they were isolated from 49 of 982 (ca. 5%) environmental samples. The proportion of Listeria spp. positive samples varied significantly across individual DCs (P < 0.01). Several facility characteristics were significantly associated with a Listeria spp. positive sample, such as zone, sampling site dryness, and cleaning regimen. A random forest model (sensitivity: 0.786, specificity: 0.874) identified geographical location and general sampling location (e.g., cold storage rooms, shipping docks) as the two most important variables associated with Listeria spp. detection. This study identified likely harborage sites (e.g., floors, cleaning equipment) of Listeria spp. in DCs across the US and emphasized the importance of sanitation operations in Listeria-prone areas., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

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., (Copyright © 2021. Published by Elsevier Ltd.)

Abstract: Listeria monocytogenes has shown the ability to grow on fresh uncut produce; however, the factors that control growth are not well understood. Peer-reviewed journal articles (n = 29) meeting the inclusion criteria and related to the growth of L. monocytogenes on fresh produce were found through university library databases and Google Scholar searches. Growth models were fit to each of the extracted 130 data sets to estimate log CFU per day rates of growth by using the DMFit tool. Multiple linear stepwise regression models for factors influencing growth rate were developed using R software. Factors included were temperature, nutrient level of inoculation buffer, initial cell concentration, final cell concentration, inoculation method, container permeability, and surface characteristics. The full model produced adjusted R2, Akaike information criterion, and root mean square error values of 0.41, 488, and 1.61, respectively. Stepwise regression resulted in a reduced model with parameters for incubation temperature, inoculation buffer type, initial and final cell concentrations, container characteristics, and produce surface characteristics. Model fit statistics improved slightly in the reduced model. A further reduced three-parameter model included storage temperature and initial and final cell concentrations, with interaction terms. This three-parameter model had adjusted R2, Akaike information criterion, and root mean square error values of 0.66, 417, and 1.24, respectively. Incubation temperature (P = 1.00E-09) initial cell concentration (P = 3.05E-12), and final cell concentration (P = 4.17E-09) all had highly significant effects on maximum growth rate. Our findings show the importance of inoculum concentration and produce microbial carrying capacity on the estimated growth rate and highlight the overall importance that temperature has on growth rate. Future experiments should consider initial inoculum concentration carefully when conducting growth studies for L. monocytogenes on whole produce., (Copyright ©, International Association for Food Protection.)

Abstract: Listeria monocytogenes causes relatively few outbreaks linked to whole fresh produce but triggers recalls each year in the United States. There are limited data on the influence of wet versus dry inoculation methods on pathogen growth on whole produce. A cocktail of five L. monocytogenes strains that included clinical, food, and environmental isolates associated with foodborne outbreaks and recalls was used. Cultures were combined to target a final wet inoculum concentration of 4 to 5 log CFU/mL. The dry inoculum was prepared by mixing wet inoculum with 100 g of sterile sand and drying for 24 h. Produce investigated belonged to major commodity families: Ericaceae (blackberry, raspberry, and blueberry), Rutaceae (lemon and mandarin orange), Rosaceae (sweet cherry), Solanaceae (tomato), Brassaceae (cauliflower and broccoli), and Apiaceae (carrot). Whole intact, inoculated fruit and vegetable commodities were incubated at 2, 12, 22, and 35 ± 2°C. Commodities were sampled for up to 28 days, and the experiment was replicated six times. The average maximum growth increase was obtained by measuring the maximum absolute increase for each replicate within a specific commodity, temperature, and inoculation method. Data for each commodity, replicate, and temperature were used to create primary growth or survival models describing the lag phase and growth or shoulder and decline as a function of time. Use of a liquid inoculum (versus dry inoculum) resulted in a markedly increased L. monocytogenes growth rate and growth magnitude on whole produce surfaces. Temperature highly influenced this difference: a greater effect seen with more commodities at higher temperatures (22 and 35°C) versus lower temperatures (2 and 12°C). These findings need to be explored for other commodities and pathogens. The degree to which wet or dry inoculation techniques more realistically mimic contamination conditions throughout the supply chain (e.g., production, harvest, postharvest, transportation, or retail) should be investigated., (Copyright ©, International Association for Food Protection.)

Abstract: Listeria monocytogenes was associated with more than 60 produce recalls, including tomato, cherry, broccoli, lemon, and lime, between 2017 and 2020. This study describes the effects of temperature, time, and food substrate as factors influencing L. monocytogenes behavior on whole intact raw fruits and vegetables. Ten intact whole fruit and vegetable commodities were chosen based on data gaps identified in a systematic literature review. Produce investigated belong to major commodity families: Ericaceae (blackberry, raspberry, and blueberry), Rutaceae (lemon and mandarin orange), Roseaceae (sweet cherry), Solanaceae (tomato), Brassaceae (cauliflower and broccoli), and Apiaceae (carrot). A cocktail of five L. monocytogenes strains that included clinical, food, or environmental isolates linked to foodborne outbreaks was used to inoculate intact whole fruits and vegetables. Samples were incubated at 2, 12, 22, 30, and 35°C with relative humidities matched to typical real-world conditions. Foods were sampled (n = 6) for up to 28 days, depending on temperature. Growth and decline rates were estimated using DMFit, an Excel add-in. Growth rates were compared with ComBase modeling predictions for L. monocytogenes. Almost every experiment showed initial growth, followed by subsequent decline. L. monocytogenes was able to grow on the whole intact surface of all produce tested, except for carrot. The 10 produce commodities supported growth of L. monocytogenes at 22 and 35°C. Growth and survival at 2 and 12°C varied by produce commodity. The standard deviation of the square root growth and decline rates showed significantly larger variability in both growth and decline rates within replicates as temperature increased. When L. monocytogenes growth occurred, it was conservatively modeled by ComBase Predictor, and growth was generally followed by decreases in concentration. This research will assist in understanding the risks of foodborne disease outbreaks and recalls associated with L. monocytogenes on fresh whole produce., (Copyright ©, International Association for Food Protection.)

Abstract: A comprehensive understanding of foodborne pathogen diversity in preharvest environments is necessary to effectively track pathogens on farms and identify sources of produce contamination. As such, this study aimed to characterize Listeria diversity in wildlife feces and agricultural water collected from a New York state produce farm over a growing season. Water samples were collected from a pond (n = 80) and a stream (n = 52). Fecal samples (n = 77) were opportunistically collected from areas <5 m from the water sources; all samples were collected from a <0.5-km2 area. Overall, 86 (41%) and 50 (24%) of 209 samples were positive for Listeria monocytogenes and Listeria spp. (excluding L. monocytogenes), respectively. For each positive sample, one L. monocytogenes or Listeria spp. isolate was speciated by sequencing the sigB gene, thereby allowing for additional characterization based on the sigB allelic type. The 86 L. monocytogenes and 50 Listeria spp. isolates represented 8 and 23 different allelic types, respectively. A subset of L. monocytogenes isolates (n = 44) from pond water and pond-adjacent feces (representing an ∼5,000-m2 area) were further characterized by pulsed-field gel electrophoresis (PFGE); these 44 isolates represented 22 PFGE types, which is indicative of considerable diversity at a small spatial scale. Ten PFGE types were isolated more than once, suggesting persistence or reintroduction of PFGE types in this area. Given the small spatial scale, the prevalence of L. monocytogenes and Listeria spp., as well as the considerable diversity among isolates, suggests traceback investigations may be challenging. For example, traceback of finished product or processing facility contamination with specific subtypes to preharvest sources may require collection of large sample sets and characterization of a considerable number of isolates. Our data also support the adage "absence of evidence does not equal evidence of absence" as applies to L. monocytogenes traceback efforts at the preharvest level., (Published 2021 by the International Association for Food Protection.)

Abstract: Past studies have shown that the on-farm distribution of Listeria monocytogenes is affected by environmental factors (e.g., weather). However, most studies were conducted at large scales (e.g., across farms), whereas few studies examined drivers of L. monocytogenes prevalence at smaller scales (e.g., within a single field). This study was performed to address this knowledge gap by (i) tracking L. monocytogenes distribution in two fields on one farm over a growing season and (ii) identifying factors associated with L. monocytogenes isolation from drag swab, soil, and agricultural water samples. Overall, L. monocytogenes was detected in 78% (21 of 27), 19% (7 of 36), and 8% (37 of 486) of water, drag swab, and soil samples, respectively. All isolates were characterized by pulsed-field gel electrophoresis. Of the 43 types identified, 14 were isolated on multiple sampling visits and/or from multiple sample types, indicating persistence in or repeated introduction into the farm environment during the study. Our findings also suggest that L. monocytogenes prevalence, even at the small spatial scale studied here, (i) was not uniform and (ii) varied more within fields than between fields or over time. This is illustrated by plot (in-field variation), field (between-field variation), and sampling visit (time), accounting for 18, 2, and 3% of variance in odds of isolating L. monocytogenes, respectively. Moreover, according to random forest analysis, water-related factors were among the top-ranked factors associated with L. monocytogenes isolation from all sample types. For example, the likelihood of isolating L. monocytogenes from drag and soil samples increased monotonically as rainfall increased. Overall, findings from this single-farm study suggests that mitigation strategies for L. monocytogenes in produce fields should focus on water-associated risk factors (e.g., rain and distance to water) and be tailored to specific high-risk in-field areas., (Copyright ©, International Association for Food Protection.)

Abstract: Understanding a food's ability to support the growth and/or survival of a pathogen throughout the supply chain is essential to minimizing large-scale contamination events. The purpose of this study was to examine the behavior (growth and/or survival) of Listeria monocytogenes on broccoli and cauliflower florets stored at different postharvest temperatures utilized along the supply chain. Broccoli and cauliflower samples were inoculated with L. monocytogenes at approximately 3 log CFU/g and stored at 23 ± 2, 12 ± 2, 4 ± 2, and -18 ± 2°C. Samples were evaluated for L. monocytogenes levels after 0, 0.167 (4 h), 1, 2, 3, and 4 days at 23 ± 2°C; 0, 0.167, 1, 2, 3, 4, 7, 10, and 14 days at 12 ± 2°C; 0, 0.167, 1, 2, 3, 4, 7, 10, 14, 21, and 28 days at 4 ± 2°C; and 0, 1, 7, 28, 56, 84, 112, 140, and 168 days at -18 ± 2°C. L. monocytogenes populations were determined by plating samples onto tryptic soy agar and modified Oxford agar supplemented with nalidixic acid. Broccoli and cauliflower supported the growth of L. monocytogenes at 23, 12, and 4°C, and higher growth rates were observed at higher temperatures. Populations of L. monocytogenes on broccoli and cauliflower samples significantly increased within 1 day at 23°C (by 1.6 and 2.0 log CFU/g, respectively) (P ≤ 0.05). At 12°C, populations of L. monocytogenes on broccoli and cauliflower samples significantly increased over 14 days by 1.4 and 1.9 log CFU/g, respectively (P ≤ 0.05). No significant difference over time was observed in L. monocytogenes populations on broccoli and cauliflower samples held under refrigeration until populations began to grow by day 10 in both commodities (P > 0.05). Under frozen storage (-18°C), populations of L. monocytogenes survived on broccoli and cauliflower at least up to 168 days. Storage of broccoli and cauliflower at lower temperatures can minimize L. monocytogenes growth potential; growth rates were lower at 4°C than at 12 and 23°C., (Copyright ©, International Association for Food Protection.)

Abstract: Listeria monocytogenes has emerged as a food safety concern for several produce commodities. Although L. monocytogenes contamination can occur throughout the supply chain, contamination from the packinghouse environment represents a particular challenge and has been linked to outbreaks and recalls. This study aimed to investigate the prevalence, persistence, and diversity of L. monocytogenes and other species of Listeria in produce packinghouses. A longitudinal study was performed in 11 packinghouses (whose commodities included microgreen, peach, apple, tomato, broccoli, cauliflower, and cucumber) in three U.S. states. In each packinghouse, 34 to 47 sites representing zones 2 to 4 were selected and swabbed. Packinghouses were visited four times over the packing season, and samples were tested for Listeria by following the U.S. Food and Drug Administration's Bacteriological Analytical Manual methods. Presumptive Listeria-positive isolates were confirmed by PCR. Species and allelic type (AT) were identified by sigB sequencing for up to eight isolates per sample. Among 1,588 samples tested, 50 (3.2%), 42 (2.7%), and 10 (0.6%) samples were positive for L. monocytogenes only, Listeria spp. (excluding L. monocytogenes) only, and both L. monocytogenes and Listeria spp., respectively. Five species of Listeria (L. monocytogenes, L. innocua, L. seeligeri, L. welshimeri, and L. marthii) were identified, and L. monocytogenes was the most prevalent species. The 102 Listeria-positive samples yielded 128 representative isolates (i.e., defined as isolates from a given sample with a different AT). Approximately 21% (21 of 102) of the Listeria-positive samples contained two or more ATs. A high AT diversity (0.95 Simpson's diversity index) was observed among Listeria isolates. There were three cases of L. monocytogenes or Listeria spp. repeated isolation (site testing positive at least twice) based on AT data. Data from this study also support the importance of drain and moisture management, because Listeria were most prevalent in samples collected from drain, cold storage, and wet nonfood contact surface sites., (Copyright ©, International Association for Food Protection.)

Listeria monocytogenes may be present in produce-associated environments (e.g., fields, packing houses); thus, understanding its growth and survival on intact, whole produce is of critical importance. The goal of this study was to identify and characterize published data on the growth and/or survival of L. monocytogenes on intact fruit and vegetable surfaces. Relevant studies were identified by searching seven electronic databases: AGRICOLA, CAB Abstracts, Center for Produce Safety funded research project final reports, FST Abstracts, Google Scholar, PubMed, and Web of Science. Searches were conducted using the following terms: Listeria monocytogenes, produce, growth, and survival. Search terms were also modified and "exploded" to find all related subheadings. Included studies had to be prospective, describe methodology (e.g., inoculation method), outline experimental parameters, and provide quantitative growth and/or survival data. Studies were not included if methods were unclear or inappropriate, or if produce was cut, processed, or otherwise treated. Of 3,459 identified citations, 88 were reviewed in full and 29 studies met the inclusion criteria. Included studies represented 21 commodities, with the majority of studies focusing on melons, leafy greens, berries, or sprouts. Synthesis of the reviewed studies suggests L. monocytogenes growth and survival on intact produce surfaces differ substantially by commodity. Parameters such as temperature and produce surface characteristics had a considerable effect on L. monocytogenes growth and survival dynamics. This review provides an inventory of the current data on L. monocytogenes growth and/or survival on intact produce surfaces. Identification of which intact produce commodities support L. monocytogenes growth and/or survival at various conditions observed along the supply chain will assist the industry in managing L. monocytogenes contamination risk.

Cucumbers were associated with four multistate outbreaks of Salmonella in the United States between 2013 and 2016. This study evaluated the fate of Listeria monocytogenes and Salmonella on whole and sliced cucumbers at various storage temperatures. Cucumbers were inoculated with five-strain cocktails of L. monocytogenes or Salmonella, air dried, and stored at 23 ± 2, 4 ± 2, and -18 ± 2°C. Whole and sliced cucumber samples were enumerated on nonselective and selective media at 0, 0.21, 1, 2, 3, and 4 days (23 ± 2°C); 0, 1, 2, 3, 7, 14, and 21 days (4 ± 2°C); and 0, 7, 28, 60, 90, and 120 days (-18 ± 2°C). For Salmonella, additional time points were added at 8 and 17 h (23 ± 2°C) and at 17 h (4 ± 2°C). Population levels were calculated for whole (CFU per cucumber) and sliced (CFU per gram) cucumbers. Both pathogens grew on whole and sliced cucumbers held at ambient temperatures. At 23 ± 2°C, L. monocytogenes and Salmonella populations significantly increased on whole (2.3 and 3.4 log CFU per cucumber, respectively) and sliced (1.7 and 3.2 log CFU/g, respectively) cucumbers within 1 day. Salmonella populations significantly increased on whole and sliced cucumbers after only 5 h (2.1 log CFU per cucumber and 1.5 log CFU/g, respectively), whereas L. monocytogenes populations were not significantly different on whole and sliced cucumbers at 5 h. L. monocytogenes and Salmonella populations survived up to 21 days on refrigerated whole and sliced cucumbers. At 4 ± 2°C, L. monocytogenes populations significantly increased on whole (2.8 log CFU per cucumber) and sliced (2.9 log CFU/g) cucumbers, whereas Salmonella populations significantly decreased on whole (0.6 log CFU per cucumber) and sliced (1.3 log CFU/g) cucumbers over 21 days. Both pathogens survived on frozen whole and sliced cucumbers for at least 120 days. The ability of L. monocytogenes and Salmonella to grow on whole and sliced cucumbers in short amounts of time at ambient temperatures, and to survive on whole and sliced cucumbers past the recommended shelf life at refrigeration temperatures, highlights the need to reduce the likelihood of contamination events throughout the cucumber supply chain.

Several produce-borne outbreaks have been associated with the use of contaminated water during preharvest applications. Salmonella has been implicated in a number of these outbreaks. The purpose of this study was to evaluate the microbial quality of agricultural surface water used in preharvest production on the Eastern Shore of Virginia in accordance with the Food Safety Modernization Act's Produce Safety Rule water standards. The study also examined the prevalence, concentration, and diversity of Salmonella in those water sources. Water samples (1 L) from 20 agricultural ponds were collected during the 2015 and 2016 growing seasons ( n = 400). Total aerobic bacteria, total coliforms, and Escherichia coli were enumerated for each sample. Population levels of each microorganism were calculated per 100-mL sample and log transformed, when necessary. Samples (250 mL) were also enriched for Salmonella. Presumptive Salmonella isolates were confirmed by PCR ( invA gene) and were serotyped. In 2016, the concentration of Salmonella in each sample was also estimated by most probable number (MPN). Indicator bacteria and environmental and meteorological factors were analyzed for their association with the detection of a Salmonella-positive water sample by using logistic regression analysis. Seventeen of the 20 ponds met the Food Safety Modernization Act's Produce Safety Rule standards for production agricultural water. Three ponds did not meet the standards because the statistical threshold value exceeded the limit. Salmonella was detected in 19% of water samples in each year (38 of 200 in 2015 and 38 of 200 in 2016). Of the 118 Salmonella isolates serotyped, 14 serotypes were identified with the most prevalent being Salmonella Newport. E. coli concentration, farm, and total aerobic bacteria concentration were significantly associated with the likelihood of detecting a Salmonella-positive sample The average concentration of Salmonella in all samples was 4.44 MPN/100 mL, with the limit of detection being 3.00 MPN/100 mL. The highest concentration of Salmonella was 93.0 MPN/100 mL. These data will assist in a better understanding of the risks that production water poses to produce contamination events.

Although flooding introduces microbiological, chemical, and physical hazards onto croplands, few data are available on the spatial extent, patterns, and development of contamination over time postflooding. To address this paucity of information, we conducted a spatially explicit study of Escherichia coli and Salmonella contamination prevalence and genetic diversity in produce fields after the catastrophic flooding that occurred in New England during 2011. Although no significant differences were detected between the two participating farms, both random forest and logistic regression revealed changes in the spatial pattern of E. coli contamination in drag swab samples over time. Analyses also indicated that E. coli detection was associated with changes in farm management to remediate the land after flooding. In particular, E. coli was widespread in drag swab samples at 21 days postflooding, but the spatial pattern changed by 238 days postflooding such that E. coli was then most prevalent in close proximity to surface water features. The combined results of several population genetics analyses indicated that over time postflooding E. coli populations on the farms (i) changed in composition and (ii) declined overall. Salmonella was primarily detected in surface water features, but some Salmonella strains were isolated from soil and drag swab samples at 21 and 44 days postflooding. Although postflood contamination and land management responses should always be evaluated in the context of each unique farm landscape, our results provide quantitative data on the general patterns of contamination after flooding and support the practice of establishing buffer zones between flood-contaminated cropland and harvestable crops in produce fields.

In-shell pecan samples (500 g) were collected over four harvest seasons (2010 to 2014) from seven pecan shelling facilities located in five U.S. states. Four varieties of pecans were analyzed: Mexican Improved, Native Seedlings, Southern Improved, and Western Improved. Pecan samples (100 g) were sent to a third party laboratory for initial Salmonella screening. When a sample was positive for Salmonella, the pathogen level was determined by the most-probable-number (MPN) method (25, 2.5, and 0.25 g). Two sample preparation strategies were used for the MPN analysis, and both strategies were combined for the reported MPN values. Forty-four (0.95%) of 4,641 in-shell pecan samples were positive for Salmonella during initial screening; prevalence by year was 0.47 to 1.4%. Prevalence was not significantly different between varieties: Mexican Improved, 1.2%; Native/Seedling, 0.99%; Southern Improved, 0.97%; and Western Improved, 0.75%. Salmonella was not isolated from 31 of 44 samples upon retesting during MPN analysis (<0.47 MPN/100 g). When Salmonella was detected, the levels were 0.47 to 39 MPN/100 g, with a mean of 2.4 MPN/100 g. Thirty-one Salmonella serotypes were obtained from 42 Salmonella-positive pecan samples; Enteritidis was the most common (12% of samples) followed by Javiana (9%) and Braenderup (7%). All Salmonella Enteritidis isolates were phage type 8. Pulsed-field gel electrophoresis analysis (XbaI) revealed within-serotype diversity, indicating introduction of contamination from a variety of sources. Most (64%) of the isolates were resistant to streptomycin or tetracycline, and 13% were resistant to three or more antibiotics. Salmonella prevalence and level on in-shell pecans is comparable to that on other nuts.

Environmental (i.e., meteorological and landscape) factors and management practices can affect the prevalence of foodborne pathogens in produce production environments. This study was conducted to determine the prevalence of Listeria monocytogenes, Listeria species (including L. monocytogenes), Salmonella, and Shiga toxin-producing Escherichia coli (STEC) in produce production environments and to identify environmental factors and management practices associated with their isolation. Ten produce farms in New York State were sampled during a 6-week period in 2010, and 124 georeferenced samples (80 terrestrial, 33 water, and 11 fecal) were collected. L. monocytogenes, Listeria spp., Salmonella, and STEC were detected in 16, 44, 4, and 5% of terrestrial samples, 30, 58, 12, and 3% of water samples, and 45, 45, 27, and 9% of fecal samples, respectively. Environmental factors and management practices were evaluated for their association with terrestrial samples positive for L. monocytogenes or other Listeria species by univariate logistic regression; analysis was not conducted for Salmonella or STEC because the number of samples positive for these pathogens was low. Although univariate analysis identified associations between isolation of L. monocytogenes or Listeria spp. from terrestrial samples and various water-related factors (e.g., proximity to wetlands and precipitation), multivariate analysis revealed that only irrigation within 3 days of sample collection was significantly associated with isolation of L. monocytogenes (odds ratio = 39) and Listeria spp. (odds ratio = 5) from terrestrial samples. These findings suggest that intervention at the irrigation level may reduce the risk of produce contamination.

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.

Pineapples have been associated with outbreaks of Escherichia coli and Salmonella. Little is known about the behavior of E. coli O157:H7 and Salmonella on cut pineapple. The objectives of this study were to evaluate the fate of E. coli O157:H7 and Salmonella on fresh and frozen cut pineapples and the influence of acid adaptation on Salmonella survival on fresh cut pineapple at ambient storage (23 degrees C). Cut pineapples were spot inoculated with a four-strain cocktail of E. coli O157:H7 or a five-strain cocktail of Salmonella. Inoculated samples were air dried, placed in containers, and stored at 4, 12, 23, and -20 degrees C. Samples were stomached and then enumerated on nonselective and selective media at days 0, 1, 3, 5, 7, 10, 14, 21, and 28 (4 and 12 degrees C); 0, 1, 3, 5, and 7 (23 degrees C); and 0, 7, 14, 21, 28, 60, 90, 120, 150, and 180 (-20 degrees C). E. coli O157:H7 and Salmonella inoculated onto pineapple did not grow but survived for the shelf life at each temperature, with improved survival at lower temperatures. E. coli O157:H7 and Salmonella can survive on frozen cut pineapples for at least 180 days. Acid adaptation of Salmonella did not allow growth and did not increase survival. Fresh and frozen cut pineapples are potential vectors for E. coli O157:H7 and Salmonella transmission and preventive procedures should be implemented during production and postharvest processing.