The process of washing tomatoes in dump (flume) tanks has been identified as a potential source of cross-contamination. This study's objective was to assess the potential for Salmonella enterica cross-contamination at various inoculation levels at the presence of 0 and 25 mg/L free chlorine (HOCl) and organic matter. Uninoculated tomatoes were introduced into a laboratory-based model flume containing tomatoes inoculated with a cocktail of five rifampicin-resistant Salmonella enterica serovars at 104, 106, or 108 CFU/tomato in water containing 0 or 25 mg/L HOCl and 0 or 300 mg/L chemical oxygen demand (COD). Uninoculated tomatoes were removed from the water at after 5, 30, 60, 120 s and were placed in bags containing tryptic soy broth supplemented with rifampicin and 0.1% sodium thiosulfate. Following incubation, enrichments were plated on tryptic soy agar supplemented with rifampicin and xylose lysine deoxycholate agar to determine the presence of Salmonella. HOCl and pH were measured before and after each trial. The HOCl in water containing 300 mg/L COD significantly (P≤0.05) declined by the end of each 120 s trial, most likely due to the increased demand for the oxidant. Higher inoculum levels and lower HOCl concentrations were (P≤0.05) significant factors that contributed to increased cross-contamination seen in this study. When HOCl levels were at 25 mg/L, no Salmonella was recovered on non-inoculated tomatoes under all conditions when inoculum levels were at 104 CFU/tomato. When the inoculum was increased to 106 and 108 CFU/tomato, cross-contamination was observed, independent of COD levels. The results from this study show that the currently required sanitizer level (e.g., 100 or 150 mg/L) for flume water may be higher than necessary and warrants re-evaluation.

Human norovirus (HuNoV) is the leading causative agent of foodborne outbreaks and is associated with the second most prevalent cause of waterborne infections in the United States. The goal of this research was to investigate the antiviral activity of chitosan microparticles (CM) against HuNoV GII.4 Sydney and its cultivable surrogate, Tulane virus (TuV), in suspensions mimicking fecally-contaminated water. CM was prepared by crosslinking chitosan molecules with sodium sulfate, and then its anti-noroviral activity was assessed using infectivity assay on TuV and RT-qPCR on TuV and HuNoV. A 3% CM suspension in PBS (pH 7.2) showed binding to TuV particles but with a negligible impact on virus infectivity (p>0.05). TuV and HuNoV suspended in fecal suspensions showed a 1.5-log10 reduction in genomic copies per ml following a 10-min contact time (p<0.05). Despite the negligible impact on viral infectivity, CM moderately binds to virus particles and helps purify environmental water by removing infectious virus particles. In this study, TuV served as a suitable surrogate for HuNoV by showing a similar log10 reduction in fecal suspension. Overall, the outcomes of thisresearch highlight the potential application of CM as a novel, natural treatment to minimize the spread of water-transmitted viral pathogens.

ABSTRACT Monitoring and maintenance of water quality in dump tanks or flume systems is crucial to maintaining proper sanitizer levels to prevent pathogen cross-contamination during postharvest washing of tomatoes, but there is limited information on how organic matter influences sanitizer efficacy in the water. The main objective of this study was to monitor water quality in flume tanks and evaluate the efficacy of postharvest washing of tomatoes in commercial packinghouses. Flume tank water samples ( n = 3) were collected on an hourly basis from three packinghouses in Florida and analyzed for pH, total dissolved solids (TDS), free chlorine, chemical oxygen demand (COD), oxidation-reduction potential, and turbidity. Additionally, three flume-water samples were collected and tested for total aerobic plate count (APC), total coliforms (TC), and Escherichia coli . Fresh tomatoes ( n = 3), both before and after washing, were collected and analyzed for the same bacterial counts. Turbidity, COD, and TDS levels in flume water increased over time in all packinghouses. Correlations observed include COD and turbidity ( r = 0.631), turbidity and TDS ( r = 0.810), and oxidation-reduction potential and chlorine ( r = 0.660). APC for water samples had an average range of 0.0 to 4.7 log CFU/mL and TC average range of 0.0 to 4.7 log CFU/mL. All water samples were negative for E. coli . The average APC for pre- and postflume tomatoes from the three packinghouses was 6.0 log CFU per tomato and ranged from 2.2 to 7.4 log CFU per tomato. The average TC count was <1.5 and 7.0 log CFU per tomato for pre- and postwash tomatoes, respectively. There was no significant effect ( P > 0.05) of postharvest washing on the microbiological qualities of tomatoes. Water quality in flume tanks deteriorated over time in all packinghouses during a typical operational day of 4 to 8 h. HIGHLIGHTS ORP correlates poorly with free chlorine levels in observed flume waters. Turbidity may be used to indicate water quality, but it correlates poorly with COD. Sanitizer levels were negatively correlated with microbial counts in flume water.

ABSTRACT For over a decade, Salmonella contamination has increasingly led to outbreaks of foodborne illness associated with fresh produce. The use of untreated animal manures, or biological soil amendments of animal origin, to amend agricultural soils holds a risk of contamination from foodborne pathogens, such as Salmonella . This study was conducted to determine the prevalence, concentration, serotypes, and antimicrobial resistance profiles of Salmonella in poultry litter from Florida farms. Litter pH, total Kjeldahl nitrogen, total ammonia nitrogen, total phosphorus (P 2 O 5 ), total potassium (K 2 O), moisture content, total solids, total ash, organic matter, and aerobic plate count (APC) were also measured. Litter samples ( n = 54) were collected from 18 broiler farms across three seasons (spring, summer, and winter). Salmonella concentrations were enumerated using a most-probable-number (MPN) method, and antimicrobial susceptibility testing was performed. The prevalence of Salmonella in litter samples was 61.1%, with a geometric mean of 0.21 ± 20.7 MPN/g. Across all seasons, Salmonella concentrations were not influenced by the chemical, physical, or microbial properties measured. Recovered Salmonella isolates ( n = 290) were grouped into serogroups O:4 (43.1%), O:7 (26.9%), O:8 (11.0%), O:1,3,10,19 (7.9%), and O:9,46 (7.2%). Serotyping Salmonella isolates ( n = 47) resulted in 12 serotypes, with the most common being Typhimurium (27.7%), Kentucky (17.0%), Enteritidis (14.9%), and Mbandaka (14.9%). Antimicrobial resistance to tetracycline (29.8%), sulfisoxazole (23.4%), and streptomycin (14.9%) was observed. No isolates were resistant to more than two antimicrobial agents. This study provides valuable information for future risk assessments for the use of poultry litter as an untreated biological soil amendment of animal origin. HIGHLIGHTS Prevalence and concentration of Salmonella in Florida poultry litter were examined. Serovars Typhimurium, Kentucky, Enteritidis, and Mbandaka were most common. Antimicrobial resistance to tetracycline, sulfisoxazole, and streptomycin was seen. Multidrug-resistant Salmonella isolates were not observed.

ABSTRACT Salmonella contamination associated with market fresh tomatoes has been problematic for the industry and consumers. A number of outbreaks have occurred, and dollar losses for the industry, including indirect collateral impact to agriculturally connected communities, have run into the hundreds of millions of dollars. This review covers these issues and an array of problems and potential solutions surrounding Salmonella contamination in tomatoes. Some other areas discussed include (i) the use of case-control studies and DNA fingerprinting to identify sources of contamination, (ii) the predilection for contamination based on Salmonella serovar and tomato cultivar, (iii) internalization, survival, and growth of Salmonella in or on tomatoes and the tomato plant, in biofilms, and in niches ancillary to tomato production and processing, (iv) the prevalence of Salmonella in tomatoes, especially in endogenous regions, and potential sources of contamination, and (v) effective and experimental means of decontaminating Salmonella from the surface and stem scar regions of the tomato. Future research should be directed in many of the areas discussed in this review, including determining and eliminating sources of contamination and targeting regions of the country where Salmonella is endemic and contamination is most likely to occur. Agriculturalists, horticulturalists, microbiologists, and epidemiologists may make the largest impact by working together to solve other unanswered questions regarding tomatoes and Salmonella contamination.

ABSTRACT With the increased consumption of fresh produce, a proportional increase in numbers of produce-related foodborne illness has been observed. An estimate of foodborne illness during 1998 to 2008 attributed ∼46% of the incidences to produce. Any foodborne illness associated with produce can have devastating consequences to the industry. The most recent data from the Centers for Disease Control and Prevention implicate leafy vegetables, vine-stalk vegetables, root vegetables, and sprouts as the most common cause of produce-related foodborne outbreaks. Excess rainfall or flooding, mainly by altering levels of soil moisture and oxygen content, affects the microbial community in soil. The goal of this research was to determine the survivability of a three-serovar Escherichia coli and a five-serovar Salmonella enterica cocktail in microcosms prepared with Candler sand (CS) and Orangeburg sandy loam (OSL) soils. Microcosms were prepared with low, medium, and high volumetric water contents and were incubated at 20 and 30°C. Serotyping was used to determine which E. coli or Salmonella serovar(s) from each cocktail persisted. Microcosm inoculation levels were ∼7.0 log CFU/g. Sampling for CS and OSL microcosms incubated at 20°C ended on day 364 and 357, respectively. The reduction of Salmonella and E. coli to below the limit of detection (extinction) in CS microcosms (incubated at 30°C at all volumetric water content [VWC] levels) was reached on day 168 and 56, respectively. Extinction of Salmonella and E. coli in OSL microcosms (incubated at 30°C at all VWCs) was reached on day 168 and 224, respectively. Of the Salmonella and E. coli serovars analyzed, Salmonella Javiana persisted the longest in both soil types, whereas E. coli O104:H4 and E. coli O145 persisted the longest in CS and OSL microcosms, respectively. Results from the current study suggest that soil type and temperature influenced pathogen persistence in CS and OSL soils more than moisture level and pathogen type.

ABSTRACT The Food Safety Modernization Act attempts to reduce produce-related foodborne illness by using preventive rather than reactive measures. The goal of this research was to determine the persistence of manure-borne generic Escherichia coli under laboratory and field conditions. The population density of naturally occurring E. coli was ∼7.2 and 5.4 log CFU/10 g in pre- and postscreened manures, respectively. Postscreened (i.e. after the liquid manure has passed through a screen) manure was applied at light, medium, and heavy rates to fields in Live Oak and Citra, FL, during the fall and summer. Laboratory microcosms of the manure-amended soils (comparable to the field's heavy application rate of manure) from both locations were maintained at 20 and 30°C. Persistence of E. coli, moisture content, and pH were monitored until E. coli became unrecoverable. The longest E. coli persistence seen in field trails was during the summer and fall trials from Citra (heavy application) that terminated on day 112 and day 280, respectively. The rate of E. coli decay ranged from 0.02 to 0.04 log CFU per day across all manure application rates, seasons, and locations. In the microcosm studies, the E. coli became extinct on day 210 in the 30°C, whereas they became unrecoverable on day 420 in the 20°C microcosms. The relatively prolonged persistence of E. coli in the microcosms suggests that survival under laboratory conditions does not mimic real-world survival rates and may not be adequate for predicting E. coli persistence in the field. The persistence data also suggest that the risk from E. coli associated with new contamination events, such as wild life intrusion, runoff, or other vectors, may be greater than the risk associated with the long-term survival of manure-borne E. coli, although more work is needed to confirm this hypothesis.

ABSTRACT Salmonella has been reported to be involved in several foodborne illness outbreaks, many of which resulted from consumption of raw tomatoes. This research aimed to optimize and evaluate the concentration of free chlorine (hypochlorous acid [HOCl]) used as a sanitizer to prevent cross-contamination of tomatoes inoculated with a cocktail of five rifampin-resistant Salmonella enterica serovars in a laboratory-based model flume system. Organic load, prepared using sterilized Scotts Premium Topsoil, was added in different quantities to the flume wash water to simulate real-world packinghouse conditions. In a typical packinghouse operation utilizing a recirculating flume system, the organic matter washed from tomato surfaces accumulates over time. In this study, different concentrations (0, 25, 50, 75, and 100 ppm) of HOCl were used as sanitizers under three organic load conditions (0, 650, and 1,000 mg/L chemical oxygen demand). Results showed that 100 ppm of HOCl was necessary to prevent Salmonella cross-contamination of uninoculated tomatoes in the model flume system in the presence of organic loading. Also, when treated with 100 ppm of HOCl, Salmonella levels were reduced by >4.5 log CFU per tomato from inoculated tomatoes in the presence of organic load. At 75 ppm of HOCl, Salmonella cross-contamination was prevented, but only in the absence of organic loading. In studies in which plate counts were negative, whole tomato enrichment studies were performed. No cross-contamination of uninoculated tomatoes was recorded when 100 ppm of HOCl was used, even in the presence of high organic load (1,000 mg/L chemical oxygen demand). Although sanitizer application reduces contamination on tomato surfaces, the primary function of sanitizers in the wash water is to prevent cross-contamination.

Tomato Best Management Practices require Florida packers to treat tomatoes in a flume system containing at least 150 ppm of free chlorine or other approved sanitizer. However, research is needed to determine the ability of these sanitizers to prevent the transfer of pathogens from contaminated to uncontaminated tomatoes, particularly under realistic packinghouse conditions. The goal of this research was to assess the minimum levels of sanitizer needed to prevent Salmonella cross-contamination between tomatoes in a model flume system under clean conditions and conditions where organic matter was added. Inoculated tomatoes (ca. 8.3 log CFU per tomato) were treated along with uninoculated tomatoes in a model flume system containing 0, 10, or 25 ppm of hypochlorous acid (HOCl) under organic loading conditions of 0, 500, or 4,000 ppm of chemical oxygen demand (COD). In the absence of HOCl, uninoculated tomatoes were highly contaminated (ca. 5 log CFU per tomato) by 15 s. No contamination was detectable (<2 log CFU per tomato) on uninoculated tomatoes when HOCl was present, except with 10 ppm at 4,000 ppm of COD, suggesting failure of 10 ppm of HOCl as a sanitizer under very high organic loading conditions. In the presence of HOCl or peroxyacetic acid, Salmonella was undetectable (<1 log CFU/ml) in the model flume water samples after 2 and 30 s, respectively. Upon enrichment, none of the uninoculated tomatoes treated with 25 ppm of HOCl for 120 s were positive for Salmonella , even in the presence of organic loading at 500 ppm of COD. Based on these findings, 25 ppm of HOCl may be adequate to prevent cross-contamination when the concentration is properly maintained, COD does not exceed 500 ppm, and tomatoes are treated for at least 120 s. Further validation in a larger commercial setting and using higher organic loading levels is necessary because managing HOCl at this low concentration is difficult, especially in a recirculating system. The use of less sanitizer by packers could reduce chemical and disposal costs.

Overhead spray and brush roller (OSBR) treatment has been shown to remove significantly more Salmonella from tomato surfaces than flume treatment. However, OSBR is not widely used in tomato packing facilities compared with other commodities, and little is known about whether brushing causes microabrasions or other physical damage. Bacteria such as Pectobacterium , a soft rot–producing plant pathogen, and Salmonella , a human pathogen, show increased survival and growth on damaged tomato surfaces. This study evaluated whether OSBR treatment had a negative effect on the safety and/or marketability of tomatoes by examining its effect on Pectobacterium and Salmonella survival. Pectobacterium survival was evaluated on inoculated tomatoes that were OSBR treated with water or sanitizer (100 ppm of NaOCl, 5 ppm of ClO 2 , or 80 ppm of peracetic acid). A 15-s OSBR treatment using water or sanitizer achieved a 3-log CFU/ml reduction in Pectobacterium levels. Survival of Pectobacterium and Salmonella on OSBR-treated, untreated, and puncture-wounded tomatoes stored at 25°C and 75 to 85% relative humidity for 7 days was also assessed. Both Pectobacterium and Salmonella populations declined rapidly on OSBR-treated and untreated tomatoes, indicating that brushing does not damage tomato fruit to the extent of promoting better pathogen survival. In contrast, the survival of both organisms was significantly ( P ≤ 0.05) higher on artificially wounded fruit. These results indicate that OSBR treatment does not increase the survival and growth of Pectobacterium or Salmonella on tomato surfaces and that it is effective in reducing Pectobacterium levels on the surface of inoculated tomatoes. These results suggest that, if used properly, an OSBR system in packinghouses is effective in removing surface contamination and does not affect tomato quality or safety.

Florida Tomato Good Agricultural Practices (T-GAPs) mandate the removal of dirt and debris from tomatoes during harvest but do not provide any specific regulations or guidance; thus, the current practice of using cloths needs to be evaluated. This study examined Salmonella transfer from inoculated green tomatoes to uninoculated cloths and from inoculated cloths to uninoculated tomatoes, upon single and multiple touches. Tomatoes were spot inoculated with a rifampin-resistant Salmonella cocktail (10 7 CFU per tomato) and were touched with cloth (clean, dirty-dry, dirty-wet) at 0, 1, or 24 h postinoculation. Salmonella was enumerated on tryptic soy agar, followed by enrichments when necessary. The transfer direction was then reversed by touching freshly inoculated cloths with uninoculated tomatoes. Transfer coefficients (TCs) were then calculated. Salmonella TCs from inoculated tomato and cloth were highest when the inoculum was wet (0.44 ± 0.13 to 0.32 ± 0.12), regardless of the condition of the cloth. Although Salmonella TCs from inoculated tomato to uninoculated cloth decreased significantly when the inoculum was dried (0.17 ± 0.23 to 0.01 ± 0.00), low levels of Salmonella were detected on cloth even after 24 h of drying. Inoculated dirty cloth did not transfer more Salmonella compared with inoculated clean cloth, and Salmonella survival was not higher on dirty cloth. When inoculated clean cloth (wet) was touched with 25 tomatoes, significantly higher levels of Salmonella were transferred to the first, second, and fourth tomatoes (0.03 ± 0.10 to 0.09 ± 0.02). However, inoculated dirty-wet (below limit of detection) and dirty-dry (0.00 to 0.04 ±0.01) cloths transferred similar levels of Salmonella to all 25 tomatoes. Results indicate a low risk of potential Salmonella contamination when the same cloth is used multiple times for debris removal, especially under high moisture levels. Results also show that the use of dirty cloths did not increase the risk of Salmonella cross-contamination.

This study was performed to optimize Salmonella Typhimurium recovery from raw duck wings with five nonselective broths (buffered peptone water, tryptic soy broth, lactose broth, universal preenrichment broth, nutrient broth) and four selective broths (selenite broth, BAX System MP media [MP], Salmonella AD media [AD], ONE broth- Salmonella [OB]). Healthy or heat-injured (50 and 85% injury) cells were inoculated at a level of 10 2 , 10 1 , or 10 0 CFU/25 g on raw duck wings. Growth was modeled using DMfit with four growth parameters: lag-phase duration, maximum growth rate, doubling time, and maximum population density. Most enrichments were able to recover Salmonella Typhimurium to greater than 6 log CFU/ml. AD, MP, and OB had significantly ( P < 0.05) higher maximum growth rate (0.9 to 1.0/h) and lower doubling time (0.7 to 0.8 h). Buffered peptone water, AD, MP, and OB recovered healthy and 50%-injured cells at low inoculum levels to more than 6.0 log CFU/ml; OB achieved the greatest recovery (7.6 and 7.9 log CFU/ml), following 24 h of incubation. The 85%-injured cells at 10 0 and 10 1 CFU/25 g, however, were only recovered in OB, reaching 7.3 and 7.5 log CFU/ml, respectively. These results suggest that OB may be an appropriate enrichment broth for the recovery of Salmonella Typhimurium from raw duck wings in standard diagnostic tests or other rapid detection methods, to avoid false-negative results.

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

Chlorine dioxide (ClO 2 ) off-gassed from an aqueous solution and reacted incrementally with potassium iodide solutions (sinks). After 30 min, 45% of the initial dose was detected as chlorite ion in the sink, whereas 35% of the initial dose was still in the source. Aqueous solutions of ClO 2 can be used as a source of ClO 2 gas in various laboratory experiments involving treatment of fruits or vegetables. Movement from source to sink is continuous, which precludes the development of large headspace concentrations and the need for a tight chamber seal. When the source solution has dissipated, the chamber can be opened safely as there is little free ClO 2 remaining in the headspace. In tests with whole, wound-inoculated tomato fruit, at both green and pink stages of ripeness, the control of Salmonella enterica serotype Typhimurium in wounds varied with the weight of gas used. The number of viable cells of Typhimurium recovered was reduced by >5 log units when ≥0.5 mg of ClO 2 was applied to three pieces of fruit during a 2-h treatment.

This study investigated flow-through immunocapture (FTI), using the Pathatrix device, followed by plating on xylose lysine desoxycholate (XLD) agar (FTI-XLD) or analysis by real-time PCR (FTI-PCR) for the detection of Salmonella on smooth tomato surfaces and in potato salad and ground beef within 8 h. Food samples were inoculated with an appropriate dilution of a five-serovar Salmonella cocktail and enriched for 5 h. Following enrichment, samples were analyzed by the FTIXLD and FTI-PCR methods. Food samples were also analyzed by a modified U.S. Food and Drug Administration Bacteriological Analytical Manual (BAM) Salmonella culture method for comparison. Salmonella inoculated at 10 0 CFU per tomato or 10 0 CFU/25 g was detected by the FTI-XLD method in 6, 8, and 4 of 10 samples for tomatoes, potato salad, and ground beef, respectively. Salmonella inoculated at 10 0 CFU per tomato or 10 0 CFU/25 g was detected by the FTI-PCR method in 8, 9, and 9 of 10 samples for tomatoes, potato salad, and ground beef, respectively. The FTI-PCR method achieved significantly higher ( P < 0.05) detection of Salmonella on tomatoes, whereas the FTI-XLD method achieved significantly lower ( P < 0.05) detection of Salmonella in ground beef when compared with the modified BAM Salmonella culture method; however, all other comparisons to the modified BAM method were not significantly different. The FTI-XLD method demonstrated the ability to isolate presumptive Salmonella colonies up to 48 h faster than did the modified BAM Salmonella culture method.

This study reports a preliminary evaluation of flow-through immunocapture (FTI) followed by real-time PCR (FTI-PCR) for the detection of Salmonella serovars on tomato surfaces within 8 h. The FTI-PCR method was compared with real-time PCR, direct plating of FTI beads on xylose lysine desoxycholate (XLD), and the conventional culture method for Salmonella found in the U.S. Food and Drug Administration's (FDA) Bacteriological Analytical Manual (BAM). Unwaxed green tomatoes were spot inoculated with a five-serovar Salmonella cocktail on smooth surfaces at levels of 10 0 to 10 4 CFU per tomato and washed in lactose broth (LB) using a shake-rub method. The resulting LB rinse was incubated at 37°C for 4 h prior to analysis by FTI-XLD, real-time PCR, or FTI-PCR and for 24 h as the first step in the BAM Salmonella culture method. For FTI-XLD, the observed lowest detection level (LDL) was 4.6 × 10 1 CFU per tomato. There was no significant difference in performance between the FTI-XLD method and the BAM Salmonella culture method ( P > 0.05); however, the FTI-XLD method reduced the overall assay time by 48 h. For real-time PCR and FTI-PCR, the observed LDLs were 4.6 × 10 1 and 9.2 × 10 0 CFU per tomato, respectively. The FTI-PCR method was superior to the BAM Salmonella culture method ( P < 0.05) for the detection of Salmonella serovars on tomato surfaces and was completed within 8 h.