Keith R. Schneider

Human enteric pathogens in produce: un-answered ecological questions with direct implications for food safety.

Recent outbreaks of gastroenteritis linked to the consumption of fresh produce raise questions about the mechanisms by which human pathogens colonize plants and persist within marketable produce. Neither Salmonella nor Escherichia coli appear to produce enzymes that degrade plant cell walls, therefore it is not yet certain how these bacteria enter plant tissues and spread within them. Similar to plant-associated bacteria, enterics use cellulose and aggregative fimbriae for their attachment to plant surfaces. Salmonella can be an effective plant endophyte, even though it is capable of triggering plant defenses. Plant-associated microbiota contributes to the fitness and translocation of these human pathogens within plant hosts, although interactions and mechanisms of communication between plant-associated microbiota and enteric pathogens are not yet characterized.

Shigella as a Foodborne Pathogen and Current Methods for Detection in Food

Shigella, the causative agent of shigellosis or “bacillary dysentery,” has been increasingly involved in foodborne outbreaks. According to the Centers for Disease Control and Preventions Emerging Infections Program, Foodborne Diseases Active Surveillance Network (FoodNet), Shigella was the third most reported foodborne bacterial pathogen in 2002. Foods are most commonly contaminated with Shigella by an infected food handler who practices poor personal hygiene. Shigella is acid resistant, salt tolerant, and can survive at infective levels in many types of foods such as fruits and vegetables, low pH foods, prepared foods, and foods held in modified atmosphere or vacuum packaging. Survival is often increased when food is held at refrigerated temperatures. Detection methods for Shigella include conventional culture methods, immunological methods, and molecular microbiological methods. Conventional culture of Shigella in foods is often problematic due to the lack of appropriate selective media. Immunological methods for Shigella have been researched, yet there is only one commercially available test kit. Molecular microbiological methods such as PCR, oligonucleotide microarrays, and rep-PCR have also been developed for the detection and identification of Shigella. This manuscript reviews the general characteristics, prevalence, growth and survival, and methods for detection of Shigella in food.

Untangling metabolic and communication networks: interactions of enterics with phytobacteria and their implications in produce safety

Recent outbreaks of vegetable-borne gastrointestinal illnesses across the globe demonstrate that human enteric pathogens can contaminate produce at any stage of production. Interactions of enterics with native plant-associated microbiota influence the microbiological safety of produce by affecting the attachment, persistence and proliferation of human pathogens on plants. Supermarket surveys have revealed that bacteria, but not fungi or mechanical damage, promote the growth of Salmonella enterica on produce. Field and laboratory studies have indicated that some plant pathogenic bacteria and fungi facilitate the entry and internalization of human pathogens in plants. Conversely, some phytobacteria, including those involved in biocontrol of plant diseases, significantly inhibit attachment and plant colonization by non-typhoidal Salmonella and enterovirulent Escherichia coli by producing antibiotics or competing for nutrients in the phyllosphere. In this review, we attempt to elucidate the mechanisms of interactions between human enteric pathogens and plant-associated microbiota, and describe how these interactions affect produce safety.

Salmonella SdiA Recognizes N-acyl Homoserine Lactone Signals from Pectobacterium carotovorum in Vitro, but Not in a Bacterial Soft Rot

Genomes of Salmonella enterica isolates, including those linked to outbreaks of produce-associated gastroenteritis, contain sdiA, which encodes a receptor of N-acyl homoserine lactones (AHL). AHL are the quorum-sensing signals used by bacteria to coordinately regulate gene expression within -their populations. Because S. enterica does not produce its own AHL, SdiA is hypothesized to function in the interspecies cross-talk with AHL-producing bacteria. Under laboratory conditions, S. enterica responded to AHL from phytobacteria by upregulating expression of srgE. AHL-dependent expression of srgE required a functional sdiA. Essentially, no sdiA-dependent resolution of the srgE recombinase-based (RIVET) reporter was observed inside a soft rot formed on a tomato by an AHL-producing strain of Pectobacterium carotovorum. The results of the control experiments suggest that sdiA is not expressed inside tomato, pepper, green onion, or carrot affected by the soft rot, and the lack of sdiA expression in planta prevents Salmonella spp. from responding to AHL. Despite its inability to detect and respond to AHL during colonization of soft rots, S. enterica reached higher final cell numbers inside a tomato soft rot compared with its growth in intact tomato fruit. The synergistic effect was the strongest under the conditions that are typical for the Florida fall/winter production season.

Microbial Safety of Tropical Fruits

There are approximately 140 million tons of over 3,000 types of tropical fruits produced annually worldwide. Tropical fruits, once unfamiliar and rare to the temperate market, are now gaining widespread acceptance. Tropical fruits are found in a variety of forms, including whole, fresh cut, dried, juice blends, frozen, pulp, and nectars in markets around the world. Documented outbreaks of foodborne disease associated with tropical fruits have occurred. Norovirus and Salmonella are the leading viral and bacterial pathogens, respectively, documented to have caused outbreaks of infections associated with consumption of tropical fruits. Sources of contamination of tropical fruit have been identified in the production environment and postharvest handling, primarily related to sanitation issues. Limited data exist on the specific route of transmission from these sources. Research on the microbial safety of tropical fruits is minimal; with the growing market for tropical fruit expected to increase by 33% in 2010 this research area needs to be addressed. The aim of this review is to discuss the foodborne pathogen outbreaks associated tropical fruit consumption, research previously completed on pathogen behavior on tropical fruits, preventive strategies for pathogen contamination, and research needs.

Detection of salmonella by flow-through immunocapture real-time PCR in selected foods within 8 hours.

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 FTI-XLD 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 hfaster than did the modified BAM Salmonella culture method.

Evaluation of overhead spray-applied sanitizers for the reduction of Salmonella on tomato surfaces.

UNLABELLED Efficacy of sanitizers in an overhead spray and brush roller system was examined for reducing Salmonella on unwaxed, mature green tomatoes. Surface inoculated tomatoes were treated in the overhead spray system for 5, 15, 30, and 60 s. A sodium hypochlorite (NaOCl) study tested NaOCl (25, 50, and 100 mg/L) against a water control. A sanitizer study examined NaOCl (100 mg/L), chlorine dioxide (ClO₂; 5 mg/L), peroxyacetic acid (PAA; 80 mg/L), and water. The overhead spray system was also compared to a scale-model flume. All NaOCl concentrations were significantly more effective at removing Salmonella than water and achieved at least a 3-log₁₀ CFU/mL reduction at different treatment times (P < 0.05). NaOCl (100 mg/L) achieved a 4 ± 1.8 log₁₀ CFU/mL reduction at 15 s. In the sanitizer study, NaOCl, ClO₂, and PAA achieved at least a 3-log₁₀ CFU/mL reduction at 15 s and between 3.9 and 5.5 log₁₀ CFU/mL reductions at 30 to 60 s. NaOCl (100 mg/L) in the overhead spray system significantly reduced more Salmonella than in the flume at 15 to 60 s. NaOCl flume treatment only reached a 1.3 ± 1.1 log₁₀ CFU/mL reduction at 15 s. Results of this study demonstrate the ability of sanitizers in the laboratory model overhead spray system to reduce Salmonella on tomato surfaces. An overhead spray system could be implemented instead of flumes to achieve higher pathogen reduction with less water and sanitizer use, thereby lowering packing costs. PRACTICAL APPLICATION The use of a non-recirculating, overhead spray brush roller system could offer a cost effective and efficacious way of washing tomatoes. The use large communal dump tanks in tomato processing has been suspected as a source of contamination in the tomato processing process. If effective, the brush roller system could augment or possible replace currently used dump tanks.

Effects of tomato variety, temperature differential, and post-stem removal time on internalization of Salmonella enterica serovar Thompson in tomatoes.

Tomatoes have been implicated in salmonellosis outbreaks due to possible contamination through bacterial internalization during postharvest handling. This study was conducted to determine the effects of tomato variety, temperature differential between tomato pulp and bacterial suspension, and the time delay between stem removal and immersion in bacterial suspension on internalization of Salmonella enterica serovar Thompson in tomato fruit. Mature green tomatoes at 32.2°C were immersed in water containing approximately 10(6) CFU/ml S. enterica bacteria. Different tomato varieties (Mountain Spring, Applause, and BHN961), temperature differentials (-10, 0, and 10°F, or -5.6, 0, and 5.6°C, respectively), and post-stem removal times (0, 2, and 16 h) were evaluated for their effects on S. enterica internalization. The incidence and density of internalized cells were determined by culture enrichment and most-probable-number methods, respectively. Overall, variety and post-stem removal time by variety interaction significantly affected the incidence of S. enterica internalization (P < 0.0001), while temperature differential had no significant effect (P = 0.36). Mountain Spring tomatoes were less susceptible to S. enterica internalization than were Applause and BHN961. Increasing the time interval between stem removal and immersion greatly reduced pathogen internalization in BHN961 and Applause, while it had no effect in Mountain Spring tomatoes. The variety and interactions between varieties and post-stem removal times (P = 0.0363) and between temperature differentials and post-stem removal times (P = 0.0257) had significant effects on the populations of internalized S. enterica. Furthermore, all internalized S. enterica cells were found within the core tissue segments immediately underneath the stem scars.

A Framework for Developing Research Protocols for Evaluation of Microbial Hazards and Controls during Production That Pertain to the Application of Untreated Soil Amendments of Animal Origin on Land Used To Grow Produce That May Be Consumed Raw

Application of manure or soil amendments of animal origin (untreated soil amendments; UTSAs) to agricultural land has been a long-standing practice to maintain or improve soil quality through addition of organic matter, nitrogen, and phosphorus. Much smaller quantities of these types of UTSAs are applied to land used for food crops than to land used for animal grain and forage. UTSAs can harbor zoonotic enteric pathogens that may survive for extended periods after application. Additional studies are needed to enhance our understanding of preharvest microbial food safety hazards and control measures pertaining to the application of UTSAs especially for land used to grow produce that may be consumed raw. This document is intended to provide an approach to study design and a framework for defining the scope and type of data required. This document also provides a tool for evaluating the strength of existing data and thus can aid the produce industry and regulatory authorities in identifying additional research needs. Ultimately, this framework provides a means by which researchers can increase consistency among and between studies and facilitates direct comparison of hazards and efficacy of controls applied to different regions, conditions, and practices.

Ozone depuration of Vibrio vulnificus from the southern quahog clam, Mercenaria campechiensis.

Southern quahog clams, Mercenaria campechiensis, were dosed with Vibrio vulnificus and placed in a pilot-scale depuration system using ozonated recirculated artificial seawater. Twenty-four hours of treatment with ozone-treated recirculating artificial seawater reduced the numbers of V. vulnificus in the shellfish meats by an average of 2 log units when compared to natural die-off in control clams. The oxidant levels (up to 3 mg/liter) did not adversely affect shellfish pumping during the depuration process.