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Outbreaks of Salmonellosis have been traced to contaminated tomato. The produce production environment poses a risk for Salmonella contamination; however, little is known about the effects of pest management practices on Salmonella during production. The study objective was to evaluate pesticide application on the inactivation of Salmonella on tomato leaves. Thirty greenhouse-grown tomato plants were inoculated with S. enterica serovars Newport or Typhimurium. Inoculation was performed by dipping tomato leaves in an 8-log CFU/mL Salmonella suspension with 0.025% (vol/vol) Silwet L-77 surfactant for 30 s, for a starting concentration of 6–7 log CFU/mL. Plants were treated with one of four pesticides, each with a different mode of action [acibenzolar- S-methyl, copper-hydroxide, peroxyacetic acid (PAA), and streptomycin]. Pesticides were applied at manufacturers' labeled rate for plant disease management with water as a control treatment. Salmonella was enumerated at 0.125 (3 h), 2, 6, and 9 days post-inoculation (dpi), and counts log-transformed. Growth of Salmonella was not observed. At 2 dpi, PAA and streptomycin significantly reduced surface Salmonella concentrations of inoculated tomato leaves (0.7 and 0.6-log CFU/g, respectively; p ≤ 0.05), while significant Salmonella log reduction occurred in the ground tomato leaves after copper hydroxide treatment (0.8-log CFU/g; p ≤ 0.05), compared to the control. No significant differences in Salmonella populations on tomato leaf surface and in ground leaves were observed from 2 to 9 dpi, regardless of pesticide application. These findings suggest single in-field pesticide applications may not be an effective mitigation strategy in limiting potential Salmonella contamination. Future research, including multiple in-field pesticide applications, or pesticide use in combination with other mitigation strategies, may offer intriguing management practices to limit possible preharvest contamination.

Outbreaks of foodborne illnesses linked to fresh fruits and vegetables have been key drivers behind a wide breadth of research aiming to fill data gaps in our understanding of the total ecology of agricultural water sources such as ponds and wells and the relationship of this ecology to foodborne pathogens such as Salmonella enterica and Listeria monocytogenes. Both S. enterica and L. monocytogenes can persist in irrigation water and have been linked to produce contamination events. Data describing the abundance of these organisms in specific agricultural water sources are valuable to guide water treatment measures. Here, we profiled the culture independent water microbiota of four farm ponds and wells correlated with microbiological recovery of S. enterica (prevalence: pond, 19.4%; well, 3.3%), L. monocytogenes (pond, 27.1%; well, 4.2%) and fecal indicator testing. Correlation between abiotic factors, including water parameters (temperature, pH, conductivity, dissolved oxygen percentage, oxidation reduction potential, and turbidity) and weather (temperature and rainfall), and foodborne pathogens were also evaluated. Although abiotic factors did not correlate with recovery of S. enterica or L. monocytogenes (p > 0.05), fecal indicators were positively correlated with incidence of S. enterica in well water. Bacterial taxa such as Sphingomonadaceae and Hymenobacter were positively correlated with the prevalence and population of S. enterica, and recovery of L. monocytogenes was positively correlated with the abundance of Rhizobacter and Comamonadaceae (p < 0.03). These data will support evolving mitigation strategies to reduce the risk of produce contamination by foodborne pathogens through irrigation.

Outbreaks of foodborne illnesses linked to fresh fruits and vegetables have been key drivers behind a wide breadth of research aiming to fill data gaps in our understanding of the total ecology of agricultural water sources such as ponds and wells and the relationship of this ecology to foodborne pathogens such as Salmonella enterica and Listeria monocytogenes. Both S. enterica and L. monocytogenes can persist in irrigation water and have been linked to produce contamination events. Data describing the abundance of these organisms in specific agricultural water sources are valuable to guide water treatment measures. Here, we profiled the culture independent water microbiota of four farm ponds and wells correlated with microbiological recovery of S. enterica (prevalence: pond, 19.4%; well, 3.3%), L. monocytogenes (pond, 27.1%; well, 4.2%) and fecal indicator testing. Correlation between abiotic factors, including water parameters (temperature, pH, conductivity, dissolved oxygen percentage, oxidation reduction potential, and turbidity) and weather (temperature and rainfall), and foodborne pathogens were also evaluated. Although abiotic factors did not correlate with recovery of S. enterica or L. monocytogenes (p > 0.05), fecal indicators were positively correlated with incidence of S. enterica in well water. Bacterial taxa such as Sphingomonadaceae and Hymenobacter were positively correlated with the prevalence and population of S. enterica, and recovery of L. monocytogenes was positively correlated with the abundance of Rhizobacter and Comamonadaceae (p < 0.03). These data will support evolving mitigation strategies to reduce the risk of produce contamination by foodborne pathogens through irrigation.

Bacterial foodborne diseases are among the most important public health issues worldwide, but in Ecuador, reports on the microbiological quality of food are scarce. In this cross-sectional study, 450 samples of high-demand Ecuadorian food, including bolon, encebollado, sauces, ceviche, fruit, fruit juice, fruit salad, cheese, raw chicken, and ground beef, were collected from popular street markets in the cities of Guayaquil, Quito, and Cuenca. Populations of total aerobic mesophilic bacteria, total coliforms, fecal coliforms, Escherichia coli, Salmonella enterica, and Listeria monocytogenes were examined on composited samples by plate count following the local regulations (Norma Tecnica Ecuatoriana, Instituto Ecuatoriano de Normalización) for each kind of food. The individual and interaction effects of the city and food type on the levels of each bacterial group were assessed by two-way analysis of variance. Selected colonies from each culture were identified using Biolog OmniLog ID and sequencing of the V3 to V4 region on the 16S rRNA gene. Average total aerobic mesophilic bacteria, total coliform, fecal coliform, and E. coli levels were 5.10 6 0.12, 2.50 6 0.16, 1.09 6 0.12, and 0.83 6 0.12 log CFU/g or mL, respectively, with significant variations among the cities. The prevalence of Salmonella in chicken and sauces and L. monocytogenes in cheese and fruit salad was greater than 20%. Opportunistic pathogens including Klebsiella pneumoniae, Staphylococcus sciuri, and Enterococcus spp. were frequently identified in the samples from all three cities. High prevalence of spoilage microorganisms such as Bacillus amyloliquefaciens and biocontrol bacteria such as Lactococcus lactis was also observed. This is the first report on the microbiological quality of food from Ecuador.

Fresh produce, as a known or suspected source of multiple foodbome outbreaks, harbors large populations of diverse microorganisms, which are partially released into wash water during processing. However, the dynamics of bacterial communities in wash water during produce processing is poorly understood. In this study, we investigated the effect of chlorine (FC) and peracetic acid (PAA) on the microbiome dynamics in spinach and romaine lettuce rinse water. Treatments with increasing concentrations of sanitizers resulted in convergence of distinct microbiomes. The resultant sanitizer resistant microbiome showed dominant presence by Bacillus sp., Arthrobacter psychrolactophilus, Cupriavidus sp., and Ralstonia sp. Most of the FC and PAA resistant bacteria isolated from spinach and lettuce rinse water after sanitation were gram positive spore forming species including Bacillus, Paenibacillus, and Brewbacillus spp., while several PAA resistant Pseudomonas spp. were also isolated from lettuce rinse water. Inoculation of foodbome pathogens altered the microbiome shift in spinach rinse water under PAA treatment, but not in lettuce rinse water or FC treated samples. These inoculated foodbome pathogens were not isolated among the sanitizer resistant strains.

Washing in chlorinated water is widely practiced for commercial fresh produce processing. While known as an effective tool for mitigating food safety risks, chlorine washing could also represent an opportunity for spreading microbial contaminations under sub-optimal operating conditions. This study evaluated Salmonella inactivation and cross-contamination in a simulated washing process of cherry and grape tomatoes. Commercially harvested tomatoes and the associated inedible plant matter (debris) were differentially inoculated with kanamycin resistant (KanR) or rifampin resistant (Rim) Salmonella strains, and washed together with uninoculated tomatoes in simulated packinghouse dump tank (flume) wash water. Washing in chlorinated water resulted in significantly higher Salmonella reduction on tomatoes than on debris, achieving 2-3 log reduction on tomatoes and about 1 log reduction on debris. Cross-contamination by Salmonella on tomatoes was significantly reduced in the presence of 25-150 mg/L free chlorine, although sporadic cross-contamination on tomatoes was detected when tomatoes and debris were inoculated at high population density. The majority of the sporadic cross-contaminations originated from Salmonella inoculated on debris. These findings suggested that debris could be a potentially significant source of contamination during commercial tomato washing.

The Eastern Shore of Virginia (ESV) is a major agricultural region in Virginia and in the past has been linked to some tomato-associated outbreaks of salmonellosis. In this study, water samples were collected weekly from irrigation ponds and wells in four representative vegetable farms (Farms A–D, each farm paired with one pond and one well) and a creek as well. In addition, water samples from two sites in the Chesapeake Bay on the ESV were collected monthly. Poultry litter was sampled monthly from three commercial broiler farms. Soil samples were collected monthly after fertilization with poultry litter from 10 farms in 2014 and another 14 farms in 2015. A most probable number method was used to detect Salmonella enterica presence and concentration in collected samples. Presumptive Salmonella colonies were confirmed by the cross-streaking method. Molecular serotyping was carried out to determine the Salmonella serovars. The average prevalence of Salmonella in pond, well, creek, and bay water samples was 19.3, 3.3, 24.2, and 29.2%, respectively. There were significant spatial and temporal differences for Salmonella incidence in various water sources. The prevalence of S. enterica in four tested ponds from farms A, B, C, and D were 16, 12, 22, and 27%, respectively. While the prevalence of S. enterica in irrigation wells was significantly lower, some well water samples tested positive during the study. Salmonella Newport was found to be the predominant serovar isolated from water samples. All poultry houses of the three tested broiler farms were Salmonella-positive at certain sampling points during the study with prevalence ranging from 14.3 to 35.4%. Salmonella was found to be able to survive up to 4 months in poultry litter amended soils from the tested farms in 2014, and up to 6 months in 2015. This research examined the dynamics of S. enterica in relationship to water source, poultry litter, and amended soil in a major agricultural area, and provides useful infor

Indigenous bacterial populations in fresh-cut produce processing facilities can have a profound effect on the survival and proliferation of inadvertently contaminating foodborne pathogens. In this study, environmental samples were collected from a variety of Zone 3 sites in a processing plant before and after daily routine sanitation. Viable mesophilic aerobic bacteria population was evaluated using both culturing method and quantitative real-time PCR (qPCR) after propidium monoazide treatment. Zone 3 surface microbiota were analyzed using 16S rRNA gene amplicon sequencing with the Qiime2 bioinformatic pipeline. Over 8000 bacterial species across 4 major phyla were identified in Zone 3 microbiomes in the processing facility. Overall, effective bacterial reduction was observed at the sampling sites on the production floor, while sanitation effect on peripheral surfaces was less evident. Effective sanitation resulted in both quantitative and qualitive shifts of Zone 3 microbiota. Several species were highly abundant at multiple sample sites for both winter and summer samplings. Based on the spatial and temporal distribution of the most abundant species, a Zone 3 core microbiome in the processing facility was tentatively described to included Cupriavidus sp., Pseudomonas sp., Ralstonia sp., Arthrobacter psychrolactophdus, Pseudomonas vemnii, Stenotrophomonas sp., and an unknown species of the family Enterobacteriaceae.

Indigenous bacterial populations in fresh-cut produce processing facilities can have a profound effect on the survival and proliferation of inadvertently contaminating foodborne pathogens. In this study, environmental samples were collected from a variety of Zone 3 sites in a processing plant before and after daily routine sanitation. Viable mesophilic aerobic bacteria population was evaluated using both culturing method and quantitative real-time PCR (qPCR) after propidium monoazide treatment. Zone 3 surface microbiota were analyzed using 16S rRNA gene amplicon sequencing with the Qiime2 bioinformatic pipeline. Over 8000 bacterial species across 4 major phyla were identified in Zone 3 microbiomes in the processing facility. Overall, effective bacterial reduction was observed at the sampling sites on the production floor, while sanitation effect on peripheral surfaces was less evident. Effective sanitation resulted in both quantitative and qualitive shifts of Zone 3 microbiota. Several species were highly abundant at multiple sample sites for both winter and summer samplings. Based on the spatial and temporal distribution of the most abundant species, a Zone 3 core microbiome in the processing facility was tentatively described to included Cupriavidus sp., Pseudomonas sp., Ralstonia sp., Arthrobacter psychrolactophdus, Pseudomonas vemnii, Stenotrophomonas sp., and an unknown species of the family Enterobacteriaceae.

Between 2000 and 2010 the Eastern Shore of Virginia was implicated in four Salmonella outbreaks associated with tomato. Therefore, a multi-year study (2012-2015) was performed to investigate presumptive factors associated with the contamination of Salmonella within tomato fields at Virginia Tech's Eastern Shore Agricultural Research and Extension Center. Factors including irrigation water sources (pond and well), type of soil amendment: fresh poultry litter (PL), PL ash, and a conventional fertilizer (triple superphosphate - TSP), and production practices: staked with plastic mulch (SP), staked without plastic mulch (SW), and non-staked without plastic mulch (NW), were evaluated by split-plot or complete-block design. All field experiments relied on naturally occurring Salmonella contamination, except one follow up experiment (worst-case scenario) which examined the potential for contamination in tomato fruits when Salmonella was applied through drip irrigation. Samples were collected from pond and well water; PL, PL ash, and TSP; and the rhizosphere, leaves, and fruits of tomato plants. Salmonella was quantified using a most probable number method and contamination ratios were calculated for each treatment. Salmonella serovar was determined by molecular serotyping. Salmonella populations varied significantly by year; however, similar trends were evident each year. Findings showed use of untreated pond water and raw PL amendment increased the likelihood of Salmonella detection in tomato plots. Salmonella Newport and Typhimurium were the most frequently detected serovars in pond water and PL amendment samples, respectively. Interestingly, while these factors increased the likelihood of Salmonella detection in tomato plots (rhizosphere and leaves), all tomato fruits sampled (n = 4800) from these plots were Salmonella negative. Contamination of tomato fruits was extremely low (< 1%) even when tomato plots were artificially inoculated with an attenuated Salmonella Newpo