S294: Quality and Safety of Fresh-cut Vegetables and Fruits

(Multistate Research Project)

Status: Active

S294: Quality and Safety of Fresh-cut Vegetables and Fruits

Duration: 10/01/2022 to 09/30/2027

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

Statement of Issues and Justification

According to the Produce Marketing Association (PMA, 2014), the U.S. market for fresh-cut vegetables and fruits is estimated at $27 billion annually ($16 billion foodservice and $11 billion retail) with bagged salads representing 61% of the market, other fresh-cut vegetables 27%, and fresh-cut fruit 11%. Fresh-cuts thus account for 16% of total retail produce sales. Postharvest losses of fresh-cut produce are difficult to estimate but given the highly perishable nature of fresh-cuts compared to intact produce, the retail value of fresh-cut produce losses and wastage at all levels may exceed $9-10 billion annually. All previous iterations of this project have worked closely with industry, particularly the United Fresh Produce Association (UFPA; formed in 2006 by the merger of the United Fresh Fruit & Vegetable Association and the International Fresh-cut Produce Association), meeting annually at their convention and helping to plan and contributing to the convention educational program. Participants from previous iterations of S294 have joined with the UFPA’s Food Safety & Technology Council (FSTC) for an all-day meeting before each of our annual project meetings. The approach and objectives for this project were developed with input from the FSTC and approved by that body. With the merger of UFPA and PMA in January 2022, we expect S294 to closely cooperate with the new association, the International Fresh Produce Association (IFPA).

The appearance, convenience, and generally high nutritive value of fresh-cut vegetables and fruits drive sales of fresh produce, but repeat sales of fresh-cuts is dependent upon assurance of its safety and quality. The industry primarily relies on established technologies derived mainly from practical experience to maintain visual quality and shelf-life with less consideration of the quality characteristics that most strongly drive repeat sales such as good flavor retention, maintenance of an appealing texture (crispness, crunchiness), and increased microbial quality leading to extended shelf stability and food safety. Through interaction with the industry we know that current technologies, especially for fresh-cut fruits, do not provide the shelf stability needed to supply domestic markets with optimum shelf life and flavor quality.

As a result of physiological and microbial deterioration occurring during storage and marketing of fresh produce, and especially fresh-cut produce, there is a continuing need to develop effective, less-damaging treatments for maintaining the sensory quality (appearance, flavor, texture), nutritional value, and food safety of fresh harvested produce (How, 1990). Most of the sales of fresh-cut produce have been in the vegetable (salad, carrot slice) area (Garrett, 2002) and commercial handling practices for fresh-cut vegetables have been described (Barth et al., 2016). Beginning over a decade ago, research and commercial interest has focused more on fresh-cut fruits and melons (Beaulieu et al., 2004; Beaulieu and Gorny, 2016; Candir, 2017; Kader, 2008; Rojas-Grau and Martin-Belloso, 2008; Soliva-Fortuny and Martin-Belloso, 2003). With over 200 different vegetable and fruit crops with potential for development as fresh-cut products, each with unique physiology and handling requirements, an integrated, scientific approach to research and development including microbiological interactions with these products is critically needed. An area of focus for this iteration of the project will be creation of new fresh-cut products that include modifications, genetic or otherwise, targeting features that either limit or improve fresh-cut quality.

The conditions on the cut surface of fresh-cut products, with the presence of water and compounds that can be used for nutrition, are ideal for growth of microbes. Unfortunately, as produce consumption has increased in the U.S. in recent years, so has the number of produce-related outbreaks of foodborne illness. Produce-related outbreaks accounted for 12.3% of all reported foodborne outbreaks from 1990 to 2007, compared to only 0.7% in the 1970s (AFF, 2010, Sivapalasingam et al., 2004). More recently, about 24% of all foodborne illnesses from 2004 to 2013 were due to fresh produce (more than any other category; CSPI, 2015). Between 1996 and 2008, lettuce/leafy greens (32.9%), tomatoes (17.1%), and melons (15.9%) comprised two-thirds of produce-related outbreaks (Gravani, 2009). Pathogens of primary concern are Escherichia coli O157:H7, Salmonella spp., Listeria monocytogenes, and Norwalk-like viruses. From 1996 to 2006, 72 foodborne illness outbreaks were associated with fresh produce consumption with 18 of these connected to fresh-cut produce (FDA 2008). The economic yearly losses due to acute foodborne illness are estimated to be $152 billion, with $39 billion of this loss associated with fresh, processed, and canned produce (Scharff, 2010). The continuing nature of such produce-related outbreaks represents a threat to further increases in per capita consumption due to lowered confidence in the microbial safety of the product by the consuming public. Such outbreaks can also be very costly to growers, processors, shippers and restaurants.

It is very difficult to ascertain the efficacy of control measures for food safety as there are no direct measures of the effectiveness of intervention strategies on the rate of occurrence of foodborne illness in the general population. Instead, model systems are used to test the effectiveness of intervention strategies at selected stages of the processing chain. The hope is that by identifying and implementing numerous control strategies along the processing chain that were found to be effective in model systems, that the resulting net risk reduction will effectively reduce the real risk of foodborne illness. There are a number of opportunities to address food safety concerns as part of this project. Quality and safety concerns often overlap. For example, efforts to reduce spoilage organisms should also impact pathogenic organisms, since conditions that support spoilage organism growth also support human pathogen survival/proliferation. Removing damaged produce prior to production will reduce the risks associated with pathogen colonization of wounds.

The Food & Drug Administration (FDA) in collaboration with the USDA and the Centers for Disease Control (CDC) issued a series of guidelines in 1998 (since updated) referred as Good Agricultural Practices (GAPs) to reduce the risk of foodborne diseases from fresh fruits and vegetables (U.S. FDA, 2008). Since that time, indicative of the importance of fresh fruit and vegetable food safety and security research, USDA has emphasized the enhancement of safety and security in its strategic planning and created the Food Safety and Quality National Education Initiative (FSQ), Special Research Grants for Food Safety, the Food Safety Institute of the Americas, and the National Food Integrated Safety Initiative. Members of S294 have been actively involved in all of these programs and many also are involved in extension food safety programs. The Food Safety Modernization Act of 2011, currently being implemented, “…aims to ensure the U.S. food supply is safe by shifting the focus from responding to contamination to preventing it.” (https://www.fda.gov/food/guidanceregulation/fsma/).

In order to ascertain food safety risks involving fresh and fresh-cut produce, it is critical to be able to determine the survival and persistence of viable or infectious human pathogens under environmental conditions occurring in produce handling and processing facilities, on harvested crops, and on intact or fresh-cut products. Therefore, methods for detection and enumeration of target microbes, including bacteria and viruses, are of core relevancy to this project. Approaches for detection and enumeration of microbes on fresh produce can play an important role in mitigation of fresh produce-associated spoilage or foodborne disease, as well, providing decision makers with timely and actionable data, especially on the presence of human pathogens in these products (Brehm-Stecher et al., 2009). These data could help guide interventions such as: refusal of contaminated product from the field, cessation of processing for line or equipment sanitation, destruction of contaminated product held in inventory pending testing results, or product recall. Due to the relatively short shelf lives of most types of fresh and particularly fresh-cut produce, rapid methods for detection and enumeration are of special relevance to the goals of this work.

Integration of physiological, pathological, food safety, and instrumental and sensory quality measurement concepts is essential for developing the most effective handling procedures and innovative, new technologies for maintaining quality and shelf stability, and safety of fresh-cut products. This multistate project is structured to foster the cooperation and collaboration among AES, ARS and other scientists in multiple disciplines that is necessary to accomplish such outcomes.   Much experimental work is needed to optimize and integrate new and emerging treatments in diverse fresh-cut products. This fact supports the proposed integrated approach of having parallel projects in different states and of focusing the research into specific areas of importance. Alternative and emerging technologies for maintaining the quality and shelf stability of fresh-cut produce are being introduced at a rate that often precludes thorough evaluation of instrumental and sensory quality attributes, and their impact on product nutritional value, microbial quality and food safety. To do so, a multidisciplinary approach as proposed herein is needed to optimize the new and emerging treatments.

Related, Current and Previous Work

A search of the CRIS database revealed an absence of multistate project(s) or coordinating committees  concerning fresh-cut vegetables and fruits, and  collaborative participation involving plant physiologists, food scientists, and microbiologists working together. This multistate project is needed in order to provide coordination and collaboration among scientists working in this field if duplication of effort is to be avoided and the available time and resources are to be effectively applied. In this way, more effective approaches that are globally applicable to different fresh-cut products may be efficiently developed and utilized.

A. Critical Review of Previous Project Accomplishments

This proposal is for a replacement project to S-294, Postharvest Quality and Safety in Fresh-cut Vegetables and Fruits (2017-22), which has resulted in numerous collaborative activities including multiple federally funded grants with multistate collaboration. The project members developed information on postharvest treatment and storage effects on the nutritional value of fresh-cut products; standardized methods for subjectively evaluating postharvest quality-related changes; developed or evaluated new tools, treatments and cultivars to improve the quality and safety of fresh-cut vegetables and fruits; developed new information on the contamination and attachment of microbes to fresh-cut product as well as developing novel approaches to microbial control; developed standard practices for recovery, inoculation, detection, and enumeration of human microbial pathogens on produce; and elucidated the physiological processes underlying both positive and negative quality changes associated with fresh-cut processing.

The new project will emphasize standardization of microbiological procedures and instrumental and subjective methods for sensory quality analysis and flavor-based shelf life measurement and emerging treatments and techniques for assuring fresh-cut quality; similarly, we will consider physiological processes that control quality changes in fresh-cut products. We will develop standard protocols for evaluation of the efficacy of sanitizers and the appropriateness of experimental protocols for microbiological challenge studies with fresh-cut produce. New sanitizers, natural product antimicrobials, and physical treatments to control microbes will be tested. Because of the potential for treatment interactions between vegetable and fruit tissues and microbes, we plan on close coordination between microbiologists and plant/food scientists in all of the above activities.

B. Fresh-cut Product Quality

Pre-harvest conditions that stress the plant will affect the quality and shelf-life of the postharvest crop (Monselise and Goren, 1987; Nigh, 1990). Knowledge of these conditions is important for assessing postharvest potential of fresh-cut products (Blacharski et al., 2001; Borve and Sekse, 2000; Gorny et al., 1998; Kim et al., 1993). The maturity of fruits and vegetables intended for fresh-cut processing is a critical factor determining potential quality and shelf life (Bai et al., 2009; Beaulieu, 2005, 2007; Beaulieu et al., 2004; Soliva-Fortuny et al., 2002, 2004). Integration of cultivar selection, pre-harvest and postharvest conditions and treatments is needed to obtain the best possible quality of the marketed fresh-cut product (Beaulieu and Lea, 2003).

Understanding the components of eating quality involves comparison of instrumental with sensory analysis (Beaulieu and Baldwin, 2002; Jordán et al., 2001a,b; Plotto et al., 2000; Schieberle and Hofmann, 1997).  Detailed sensory analysis is required on fresh-cut produce treated in  various applications that  potentially affect flavor and textural attributes and impact overall eating quality. Many attempts at measuring texture have used sensory analysis coupled with instrumental measurements (Abbott et al., 1984; Beaulieu et al., 2004; DeBelie et al., 2002; Drake, 1962; Harker et al., 2002; Mohamed et al., 1982; Szczesniak, 1963; Vickers, 1981; Vickers and Bourne, 1976; Vincent, 1998). There are no generally accepted definitions of textural attributes applicable to fresh-cut products, sensory scale anchors, or methods for their measurement (Fillion and Kilcast, 2002; Harker et al., 1997). The proposed project will include study of practical methods for measuring flavor and texture of fresh-cut vegetables and fruits, and relating to sensory measurements using modern multivariate statistical techniques.

C. Technologies for Maintaining Quality and Shelf Stability

  1. Modified atmosphere packaging. Altering the gas mixture surrounding fresh-cut produce to a composition different from that of normal air is an effective tool to prolong quality retention, and this can be achieved by using packaging technologies (Kader et al., 1989). Essentially all fresh-cut products are handled in packages designed to maintain a modified atmosphere surrounding the enclosed product. Importantly however, good temperature control is essential for effective use of this modified atmosphere packaging (MAP; Beaudry et al., 2006).

MAP can reduce or delay the physical, chemical and microbiological changes caused in the produce during preparation (Gorny, 1997; Sandhya, 2010) and is largely used to suppress global metabolic activity through respiratory inhibition (Kaji et al., 1993; Soliva-Fortuny et al., 2005), inhibition of ethylene synthesis and action (Gil et al., 1998; Gorny et al., 1999; Rosen et al., 1989; Soliva-Fortuny et al., 2005), inhibition of oxidative processes (Escalona et al., 2010; Smyth et al., 1998), retention of ascorbic acid (Agar et al., 1999), and suppression of microbial growth (Bai et al., 2001; Gonzalez-Fandos et al., 2006). MAP also provides an enclosed system that minimizes water loss and thereby reduces shriveling, wilting, and the loss of fresh and glossy appearance. Dangers associated with MAP include potential induction of fermentation and resultant off-flavors (Zhang and Watkins, 2005).

  1. Active packaging. Active packages enhance quality and safety through either the scavenging of compounds that are involved in produce deterioration processes or the release of compounds that can mitigate the effect of factors involved in produce deterioration (Almenar, 2020; Almenar, 2021). Types of active packaging include modified moisture packaging (Chopra et al., 2016; Yue Bi et al., 2014), ethylene-removing packaging (Awalgaonkar et al., 2020; Chopra et al., 2017; Cao et al., 2015), oxygen-removing packaging (Kartal et al., 2012), carbon-dioxide-removing packaging (Wang et al., 2015), and antimicrobial packaging (Almenar, 2021; Almenar et al., 2007c; Almenar et al., 2009; Lopez et al., 2017). Bio-based plastics and biodegradable plastics are the main lines of current research in active packaging (Kuorwel et al., 2011).

The potential exists for the inclusion of additional bioactive compounds in the package or applied to the product that target human and plant pathogens. Such compounds include natural plant volatiles (Utama et al., 2002) such as hexanal and trans-2-hexenal (Kubo et al., 2004; Lanciotti et al.2003, Patrignani et al., 2008; Song et al., 1996; Utama et al, 2002). These volatiles can also be effectively encapsulated in cyclodextrins and their release manipulated (Almenar et al., 2007a; b; da Rocha Neto, 2018).

  1. 1-Methylcyclopropene (1-MCP). 1-MCP blocks ethylene receptors, preventing ethylene effects (Blankenship and Dole, 2003; Sisler and Blankenship, 1996; Sisler and Serek, 1997; Watkins, 2002). Thus, 1-MCP can potentially be used to control ripening and senescence of fresh-cut products (Jeong et al., 2004; Toivonen, 2008; Vilas-Boas et al., 2007), microbiological growth (Zhou, 2006), browning and hydrolysis of ascorbic acid (Buda et al., 2003). Further, the cyclodextrin can be incorporated into polymers to further modify release kinetics (Lee et al., 2006).

  2. Texture enhancers. Calcium salts have been used to decrease softening of fresh-cut fruits (Soliva-Fortuny et al., 2005). Calcium ions form cross-links between free carboxyl groups of pectin chains, resulting in strengthening of cell walls by formation of calcium pectates (Aguayo et al., 2008). Potassium has been reported to have both positive (Lester and Jifon, 2006) and negative (Pacheco et al., 2008) effects on fruit texture. Texture enhancers can be incorporated into the formulation of edible coatings (Oms-Oliu et al., 2008b; Rojas-Grau et al., 2009).

  3. Antibrowning agents. Browning caused by polyphenoloxidase (PPO) can be controlled by reducing the O2 content of the surrounding atmosphere or by using antibrowning agents, which can be grouped into different categories: acidulants, reducing agents, chelating agents, complexing agents, enzyme inhibitors, and synergistic combinations (Fan et al., 2005). Son et al. (2001) reported that the antibrowning compounds oxalic acid, oxalacetic acid, ascorbic acid-2-phosphate, cysteine, glutathione, N-acetylcysteine, kojic acid and 4-hexyl resorcinol showed the highest browning inhibitory activity on fresh-cut apples among 36 known antibrowning compounds.

  4. Naturally occurring compounds. Naturally occurring compounds have been also explored as flavor enhancers, and antibrowning agents. Some honeys and juices have shown antibrowning effects on fresh-cut apple slices (Jeon and Zhao, 2005; Song et al., 2007). Other natural antibrowning agents such as hexylresorcinol, N-acetylcysteine, and glutathione have been reported to reduce the browning of Fuji apple pieces with results significantly better than for ascorbic acid (Rojas-Grau et al., 2008a,b).

  5. Edible coatings. Edible coatings act via their barrier properties to water and gases (Baldwin et al., 1995a). Benefits of edible coatings on fresh-cut produce are reduced a) browning (Beaulieu and Gorny, 2004, Baldwin et al., 1996), b) respiration (Banks, 1984), c) ethylene production (Wong et al., 1994), d) moisture loss (Avena-Bustillos et al., 1997), and e) surface discoloration (Banks, 1984), as well as enhanced flavor volatiles and texture retention (Baldwin et al., 1995b; Guilbert et al., 1996).

Edible coatings can also extend fresh-cut shelf life by carrying active ingredients such as antibrowning agents, antimicrobial compounds, texture enhancers, and nutraceuticals (Han et al., 2004b; Hernandez-Munoz et al., 2008; Lee et al., 2003; Oms-Oliu et al., 2008a; Perez-Gago et al., 2006; Raybaudi-Massilia et al., 2008a; Rojas-Grau et al.; 2009; Waimaleongora-Ek et al., 2008; Zhao, 2010). New generation edible coatings focus on incorporating and controlling the release of active compounds using nanotechnology (Lopez-Rubio et al., 2006) and multi-layer nanolaminates (Weiss et al., 2006).

  1. Mild heat stress. Wounding of plant tissues results in degradative oxidative processes including browning, lipid peroxidation, and flavor changes (Hodges et al., 2004; Toivonen, 2003). However, plants also respond to abiotic and biotic stresses by up-regulation of their antioxidant system (Baker and Orlandi, 1995; Bray et al., 2000; Wang et al., 2003), which serves to protect against those processes by acting as reducing agents, free radical terminators, metal chelators, and singlet oxygen quenchers and by mediating the activity of various oxidizing enzymes (Ho, 1992; Okuda, 1993; Toivonen, 2004). Nonlethal heat stress has been shown to stimulate production of bioactive compounds (Lemoine et al., 2009), delay senescence, and improve quality of fresh-cut products (Sgroppo et al., 2009).

  2. Molecular approaches. Quality factors in fresh-cut produce, both positive and negative, are the result of physiological and biochemical processes. Enzymes have been identified that either control or limit the rate of most of these processes and it has been demonstrated that those enzyme proteins are, at their most basic level, controlled by the genes that are responsible for their production. An example is identification of the genes responsible for browning of fresh-cut (i.e., wounded) tissue as in lettuce (Daddiego et al., 2018), eggplant (Liu et al., 2021), and apple (Di Guardo et al., 2014; Zuo et al., 2021) along with molecular approaches for silencing those like antisense RNA, RNA interference, and CRISPR/Cas9 technology (El-Sappah et al., 2021).

D. Physiology of Fresh-cut Products

The physiology of fresh-cut vegetables and fruits is typical of that observed in plant tissues that have been wounded or exposed to stress conditions (Brecht et al., 2004). This includes increased respiration and ethylene production. Other consequences of wounding include oxidative browning, lipid oxidation, and enhanced water loss. Minimizing negative consequences of wounding in fresh-cut products can result in increased shelf-life and greater maintenance of nutritional, appearance, and taste quality.

  1. Appearance versus flavor-based shelf life. Shelf life is generally based on appearance, but internal quality characteristics may deteriorate faster than external characteristics. There are three main reasons for the general decline in flavor of fresh produce: genetics, harvest maturity, and postharvest handling (Baldwin and Plotto, 2007). Both harvest maturity and postharvest handling techniques are often directed toward extending the shelf life of fresh produce after harvest, sometimes with negative impacts on flavor quality. This is especially problematic for fresh-cut products, for which flavor loss may be due to metabolic changes related to wounding, off-gassing of volatile compounds due to removal of diffusion barriers, and altering of atmosphere through necessary packaging (Forney, 2008).

Quality and quantity of flavor components have been shown to vary among varieties of fruits and vegetables. Preharvest factors, including environmental influences, during the production season can affect flavor quality (Baldwin et al. 1995c; Beaulieu, 2005; Romani et al. 1983; Thybo et al., 2006; Wright and Harris 1985). Harvest maturity, especially for climacteric fruits that ripen after harvest, strongly affects flavor life (Bai et al., 2009; Beaulieu, 2005, 2007; Beaulieu et al., 2004; Soliva-Fortuny et al., 2002, 2004). Even nonclimacteric fruits like citrus have an optimal harvest maturity window for flavor (Obenland et al., 2009).

Quality of fresh-cut products is highly dependent on minimizing injury to the product. Wounding of plant tissues may cause elevated ethylene production and membrane lipid degradation (Deschene et al., 1991; Picchioni et al., 1994; Rolle and Chism, 1987; Zhuang et al., 1997). Extensive enzymatic degradation occurs in damaged membrane systems (Marangoni et al., 1996). Discoloration occurs when the products of phenylpropanoid metabolism and possibly other substrates (e.g., anthocyanins) are oxidized by phenolases such as polyphenoloxidase (PPO) or peroxidases. Wounding also induces synthesis of enzymes involved in browning reactions (Martinez and Whitaker, 1995) or substrate biosynthesis (Ke and Saltveit, 1989). It has been clearly shown that the degree of injury incurred during the cutting process has a tremendous influence on slice quality and shelf-life (Portela and Cantwell, 2001; Toivonen et al., 2005).

  1. Nutritional components in fresh-cut produce. Intact and fresh-cut vegetables and fruits are important dietary sources of vitamin A, C and E, minerals, carotenoids, polyphenols (flavonoids), and other antioxidant phytochemicals. However, little is known of the effects of fresh-cut processing technologies on the nutritional quality of fresh-cut products. Cutting increased the phenolic content and antioxidant capacity of fresh-cut lettuce and other vegetables (Heredia and Cisneros-Zevallos, 2009). This suggests that stressful treatments such as fresh cutting can increase nutrient levels in some commodities under certain circumstances. More information is needed relating to phytonutrient levels in fresh-cut produce and how emerging fresh cutting technologies are impacting phytonutrients. Measurement of antioxidant capacity will also be important in order to better understand the potential health implications of these changes. This can most expediently be accomplished by measuring changes in the two most important antioxidant functionalities: 1) reducing capacity, and 2) peroxyl radical scavenging capacity (Toivonen and Hampson, 2010).

E. Microbiology and Food Safety of Fresh-cut Products

  1. Methods for recovery, inoculation, detection, and enumeration/quantification of fecal contamination, biofilms, and human pathogens. Pre-analytical sample preparation: Although numerous rapid detection methods are available commercially or are being developed, detection methods alone are inadequate. As detection is a final downstream event, the value of the information that it provides is dependent on critical upstream inputs such as effective separation of target cells from the food matrix and their subsequent concentration to analytically suitable volumes (Brehm-Stecher et al., 2009; DSouza et al., 2009). Therefore, development of methods for effective pre-analytical sample preparation will play an integral role in the work proposed here.

Enumeration: Although presence/absence testing for pathogens serves a vital role in food safety surveillance, quantitative information on pathogen loads is required to understand pathogen behavior in foods or in processing environments, to explore kinetic phenomena such as pathogen growth or inactivation and to provide critical input for microbial risk analyses (Brehm-Stecher et al., 2009). Therefore, methods capable of providing enumerative data on target organisms, especially human pathogens, are expected to be of special importance to the proposed work.

Biofilms and inoculation studies: The dominant microbial phenotype in nature is the biofilm, not individual, free-living cells (Bisha and Brehm-Stecher, 2009a; Fett et al., 2006). Understanding the complex microbial interactions on produce surfaces will be an important focus of the proposed project. Approaches for artificial inoculation of microbes onto produce surfaces, with the goal of more accurately representing processes occurring in the field or in processing environments, will also be a focus of this work.

Microbial interactions on produce or processing surfaces: Tape-based sampling methods have recently been combined with whole-cell molecular identification via fluorescence in situ hybridization (FISH), to enable rapid, visual identification of specific pathogens on produce surfaces (Bisha and Brehm-Stecher, 2009b). Because tape-based sampling approaches preserve the spatial relationships of microflora present on plant surfaces, they could enable characterization of competitive or metabiotic interactions between plant saprophytes or spoilage organisms on these surfaces or on inanimate processing surfaces (Bisha and Brehm-Stecher, 2009b).

Viruses: Human noroviruses remain the leading cause of viral gastroenteritis outbreaks worldwide (Siebenga et al., 2009). Newly emergent strains are highly virulent and can be life threatening, especially to the elderly and immuno-compromised (Siebenga et al., 2009). Other viruses, such as hepatitis A, are significant because of the severity of disease they cause. Outbreaks of foodborne viruses have been linked to fresh produce such as green onions, lettuce, raspberries or frozen strawberries and raspberries. Methods for rapid and sensitive detection of viruses are needed. Promising approaches include real-time reverse-transcriptase-PCR, nucleic acid sequence based amplification (NASBA), and loop-mediated isothermal amplification (LAMP) assays.

  1. Sanitizers; alternatives/natural products; antimicrobial efficacy and sensory effects. Washing fresh-cut produce after cutting and prior to packaging is an important process in reducing microbial populations. Different washing chemical agents have been studied to determine their efficacy in the inactivation of pathogenic bacteria (Fan et al., 2009; Narciso and Plotto, 2005; O’Conner-Shaw et al., 1996; Ukuku and Fett, 2004; Ukuku et al., 2004; Yuk et al., 2005). Combining levulinic acid with sodium dodecyl sulfate (SDS) dramatically increased the bactericidal activity against both Salmonella and E. coli O157:H7 populations (Zhao et al., 2009). However, lettuce treated with levulinic acid and SDS developed sogginess and loss of texture (Guan et al., 2010). Organic acids have shown to be effective in reducing bacteria populations on some fresh-cut fruits and vegetables.

Formation of chlorine by-products is a concern due to their potential adverse health effects.  Scientists at ARS-PA studied the formation of trichloromethane (a chlorine by-product) in water, fresh-cut produce and as affected by the presence of citric acid. Results showed that trichloromethane was formed in chlorinated water, but not in ClO2 solution. Higher amount of trichloromethane (up to 280 ng/mL) was produced in the chlorine solution used for washing cut-lettuce than for diced onions while levels of trichloromethane in the final products (cut vegetables) were much lower (14-22 ng/g) than in the water. Citric acid reacted with chlorine, producing more than 1,000 ng/mL of trichloromethane in chlorine solution, suggesting that replacements should be explored for citric acid for pH adjustment.

Gaseous antimicrobials such as chlorine dioxide (ClO2) and ozone (O3) can penetrate to sites that liquid sanitizer can’t. This makes gaseous antimicrobials attractive for fresh-cut applications. Chlorine dioxide has been successfully applied to a number of crops (Du et al. 2003, Han et al. 2001; 2004a; Lee et al., 2004; Sapers et al. 2003), but ClO2 gas can have a negative impact on visual quality of leafy vegetables (Mahmoud and Linton 2008). Therefore, processing conditions would likely need to be altered for consumer acceptance and commercial applications. Gaseous O3 has been found to be effective in reducing S. enteritidis on tomatoes (Da_ et al., 2006) and Salmonella on cantaloupe melons (Selma et al., 2008).

Ionizing irradiation is effective in inactivating pathogens either on the surface or inside of fresh produce (Fan et al., 2008). Other new food processing technologies such as cold plasma, in-packaging sanitization, ultraviolet (UV) irradiation, and high hydrostatic pressure need to be investigated for their feasibility on fresh and fresh-cut produce. Tree-ripe fruit such as apricots and peaches cannot be washed with chemical sanitizers due to their softness.  Non-aqueous technologies are needed to minimize the risk of human pathogens on this type of fruit.  Scientists at ARS-PA evaluated the efficacy of UV-C light for inactivation of Salmonella spp. and E. coli O157:H7 on apricot. A short (10 sec) UV-C treatment could result in immediate reduction of 99% on apricot surface.  During post-UV storage, pathogen populations on UV-C exposed fruit continue to decrease: compared with the non-treated fruit, up to 99.999% of pathogens could be reduced after post-UV-C storage.

Unlike chemical sanitizers that only affect the surface of produce, hot water can inactivate bacteria below the produce surface (Breidt et al. 2000), and thus, is potentially more effective than chemical washes (Annous et al. 2004, Breidt et al. 2000, Lichter et al. 2000). Treatment of whole cantaloupes with 85°C water for 60 sec reduced Salmonella spp. by ca. 4.6 log CFU per cm2 of rind (Solomon et al., 2006). Hot water pasteurization of cantaloupes reduced native bacteria, yeast and mold populations, which also frequently resulted in lower microbial loads on fresh-cut fruit (Fan et al., 2008). No negative effect by the hot water treatment on sensory quality or vitamin C content of fresh-cut cantaloupes was observed.

Natural products and generally regarded as safe (GRAS) substances that often have antimicrobial/antioxidative activities or otherwise maintain the freshness and quality of the fresh-cut produce are of interest (Tiwari et al., 2009). In many cases, the concentration of the antimicrobial compound in its natural form is too low to be successfully used without damaging the sensory characteristics of the final product (Bagamboula et al., 2004). However, recent research (ARS-PA) showed that Salmonella typhimurium population on tomato fruit could be reduced by more than 99.99% without negatively impacting fruit quality after treatment with the certain naturally occurring plant extracts and their major constitutes. Also, cyclic sopholipids extracted from yeast were shown to inactivate Salmonella and Listeria spp.

Most chemical sanitizers are oxidants that can cause bleaching and other discoloration of fresh and fresh-cut produce. Because of their oxidative nature, these chemicals may not be able to be applied directly with antioxidants. However, treatment with oxidative sanitizers followed by treatment with an antioxidant may effectively maintain microbial quality of fresh-cut fruits. Several classes of surface active agents approved for food use can aid penetration of substances within apples (Saftner et al., 1997) and other produce, but their compatibility with fresh-cuts and their efficacy when combined with sanitizers have not been well studied.

  1. Antimicrobial edible coatings. Diverse coating materials have antimicrobial functions (Chien et al., 2007; Iverson and Ager, 2003; Park et al., 2005; Vargas et al., 2006; Zhang and Quantick, 1998) and antimicrobial agents may be incorporated into edible coatings (Du Plooya et al., 2009; Franssen and Krochta, 2003; Garcia et al., 2001; Lee et al., 2003; Park et al., 2004; 2005; Raybaudi-Massilia et al., 2008a; b; Rojas-Grau et al., 2007) to extend shelf-life and enhance microbial safety of fresh-cut produce (Lin and Zhao, 2007; Vargas et al., 2006). A patented edible film comprising organic acids, protein, and glycerol can inhibit human pathogen growth, including L. monocytogenes, S. gaminara, and E. coli 0157:H7 (Hettiarachchy and Satchithanandam, 2007). A novel class of phenolic fatty acids has shown potential for use as antimicrobials against Gram-positive bacteria (ARS-PA). The function and application of some antimicrobial coatings in fresh and fresh-cut produce have been discussed extensively in several review articles including Lin and Zhao (2007), McHugh et al., (2009), and Rojas-Grau et al. (2009).

  2. Antimicrobial packaging. Antimicrobial packaging involves incorporating an active compound with an antimicrobial capacity into the package in order to retard, reduce, or inhibit microbial growth during a desired period of time (Almenar, 2020; Almenar, 2021). Antimicrobial packaging has been shown to improve shelf life of many fruits and vegetables by lowering microbial load, retaining color, and maintaining firmness and weight (Almenar et al., 2007c; Almenar et al., 2009; Muriel-Galet et al., 2013; Rodriguez-Lafuente et al., 2010; da Rocha Neto et al., 2019).

Note that while we will not be conducting research on HACCP, the results from our project may help strength HACCP programs, such as development of rapid and sensitive detection methods for the identification of biological hazards (foodborne pathogens), and also development of new sanitizers, treatments and technologies for controlling pathogens.


  1. Evaluate methods of sampling and measuring flavor and nutrition of fresh-cut products to facilitate comparison to traditional shelf life factors.
    Comments: Activities on these objectives will be coordinated by the chairs of the 'Quality and Physiology Subcommittee' (Obj. 1, 2 & 3) and the 'Microbiology Subcommittee' (Obj. 4 & 5), who will be elected at the first annual meeting of the project participants (See: Organization and Governance).
  2. Develop new strategies to improve and maintain inherent fresh-cut product quality and nutrition.
    Comments: Activities on these objectives will be coordinated by the chairs of the 'Quality and Physiology Subcommittee' (Obj. 1, 2 & 3) and the 'Microbiology Subcommittee' (Obj. 4 & 5), who will be elected at the first annual meeting of the project participants (See: Organization and Governance).
  3. Improve understanding of physiological mechanisms that affect fresh-cut product quality.
    Comments: Activities on these objectives will be coordinated by the chairs of the 'Quality and Physiology Subcommittee' (Obj. 1, 2 & 3) and the 'Microbiology Subcommittee' (Obj. 4 & 5), who will be elected at the first annual meeting of the project participants (See: Organization and Governance).
  4. Determine critical factors in controlled inoculation studies with human pathogens and surrogates that influence the outcome of quantitative microbial risk assessments.
    Comments: Activities on these objectives will be coordinated by the chairs of the 'Quality and Physiology Subcommittee' (Obj. 1, 2 & 3) and the 'Microbiology Subcommittee' (Obj. 4 & 5), who will be elected at the first annual meeting of the project participants (See: Organization and Governance).
  5. Development and validation of novel diagnostic methods to determine presence of human pathogens and chemical hazards associated with fresh and fresh-cut products.
    Comments: Activities on these objectives will be coordinated by the chairs of the 'Quality and Physiology Subcommittee' (Obj. 1, 2 & 3) and the 'Microbiology Subcommittee' (Obj. 4 & 5), who will be elected at the first annual meeting of the project participants (See: Organization and Governance).


In this proposal section, Station abbreviations are indicated to show where collaboration between project participants will occur. Additional collaborations will develop over the course of the project as, at the annual meeting, like research objectives will be compared and new collaborations will be established as new research results occur.

There will be no economic analysis of new technologies or interventions as an objective of this project. Researchers may choose to include an economic analysis of their treatment but we do not have any economic faculty within this project. We feel that it would generally be premature to evaluate economics before efficacy is established, putting that activity outside the scope of the project.

All produce will be appropriately prepared and cut under highly sanitary conditions at refrigerated temperatures where the processing area, tools, and gloved hands are appropriately sanitized and personnel wear proper clothing to protect cut produce from contamination. Standard sanitation procedures to be used in conducting experiments will be determined by consensus of the participants. Any post-cutting treatments and packaging will also be performed using good manufacturing practices. After treatment, fresh-cut products, with intact control samples, will be stored using appropriate refrigerated temperatures and durations depending on commodity, stage of maturity at harvest or upon treatment, and storage temperature of the intact produce prior to processing.

Standard sensory and instrumental measures of flavor quality to be used in work encompassing the first three objectives will be developed by project members in CA, FL, and ARS-FL. Visual quality of fresh-cut products will be evaluated by applying standard hedonic scoring systems, reflectance color measurements, and spectrophotometric analysis of chlorophyll, anthocyanin, carotenoid, and phenolic pigments. Textural alterations will be analyzed using mechanical measurements of tissue firmness.

Since apparent responses to temperature, ethylene, etc. can be strongly affected by different fresh-cut preparation procedures, certain basic preparation procedures such as slicing procedures, slice or chunk sizes, and sanitation methods will need to be agreed upon, especially by those participants working with the same or similar types of products. The participants will agree on standard anti-browning and texture stabilizing treatments for products being studied at different stations. Similarly, standard hedonic scoring systems and physical measurement methods for color and texture for each common product will be used as much as possible. Flavor-related factors will be measured upon sampling or removal from packages or storage containers after a standard, specified period of time at a specified temperature.

Objective 1 - Evaluate methods of sampling and measuring flavor and nutrition of fresh-cut products to facilitate comparison to traditional shelf life factors.

Procedures: 1) We propose to compare methods currently used by project members for consumer and sensory panel testing of fresh-cut products and develop standard procedures. 2) We will conduct coordinated shelf life tests and develop standard procedures for measuring the shelf life of fresh-cut products in terms of retention of acceptable flavor versus physicochemical properties, including nutritionally important constituents. 3) We will relate instrumental with sensory measurements of fresh-cut product quality using newly developed instruments and statistical techniques.

For comparison of techniques between laboratories, fresh-cut products will be shared among participants and quantitative descriptive analysis, flash profiling, ranking, or sensory discrimination analysis in conjunction with signal detection theory will be used, depending on respective resources. Standardized preparation protocols for each product being tested will be used at all participating locations, and comparison between laboratories made for descriptive analyses.

Quality evaluations comparing sensory data with physicochemical analyses will be conducted with standard and novel instrumentation. Shelf life in terms of appearance, texture, nutritional value, and flavor will be compared and, specifically, interactions between volatile and non-volatile components of flavor will be determined. The chemical data for flavor compounds will be combined with sensory data to determine what type of aroma profile and sugar/acid ratios give the highest flavor quality (preference) or off-flavor (low preference) ratings for fresh-cut products.

Standardized sensory evaluation methods and objective methods of measuring color, texture, and composition of fresh-cut products will be proposed. Compositional measurements will include constituents related to flavor (sugars, organic acids, aroma volatiles) and nutritional value [ascorbic acid (vitamin C), carotenoids, polyphenols (flavonoids), and other antioxidant phytochemicals]. Potential methods will be discussed, agreed upon, and revisions made, and the procedures finalized and distributed within and outside the multistate project. The updated sensory analysis guide will be a collaborative effort between, but not exclusive to, participants from FL, ARS-FL, ARS-MD, and MI.

Objective 2 - Develop new strategies to improve and maintain inherent fresh-cut product quality and nutrition.

Procedures: 1) We will work with public and private entities involved in cultivar development to identify both conventional and genetically modified germplasm with outstanding sensory quality. 2) We will identify optimum initial (whole product) quality factors relating to improved flavor-based shelf life. 3) We will investigate improved processing and packaging strategies to better maintain fresh-cut product quality as compared to standard commercial practices.

Genotype selections will be based on reduced or delayed ripening characteristics, better appearance and flavor, enhanced texture retention, and higher nutritional value. This will form the basis for identification of the targets for genetic manipulation in Obj. 3. Sensory evaluations will be conducted by untrained panelists and by trained judges depending on the nature of the evaluation. Both intensity and overall acceptability characteristics of the fresh-cut products will be evaluated and later standardized. Work in this area will be conducted in FL, ARS-FL, MI, and WSU.

Selection of optimal quality produce for fresh-cut processing is another important consideration. Since fresh-cut products are intended for immediate consumption, fresh-cut fruit should be ripe or nearly ripe and vegetables should be fresh and showing no signs of senescence. Studies on initial product quality including optimal fruit maturity, preconditioning, and trait targeting will be emphasized; non-destructive instrumental measurements of vegetable and fruit quality will also be evaluated. Work in this area will be conducted in FL, ARS-FL, ARS-MD, and MI.

Pre-cutting treatments of intact produce destined for fresh-cut products will target ethylene synthesis or action, undesirable enzyme activities, and/or microbial loads of the fresh-cut products. We will evaluate 1-MCP and heat treatments and coordinate with the microbiologists in testing emerging sanitizers. Post-cutting treatments to be tested will include MAP; 1-MCP, natural products, and edible coatings (with or without food additives, antimicrobials and preservatives). Pectinesterase application with and without calcium in order to firm the tissue by creating pectin crosslinking will be evaluated for its effect on fresh-cut fruit tissue softening and watersoaking development during storage. Rinsing the cut fruit with buffered alkaline solution will also be tested to determine if hyper-acidification of the cut surface due to vacuole rupture might play a role in the development of watersoaking and softening due to activation of hydrolases in the cell wall and membrane.

New alternatives for fresh-cut produce packaging will be developed based on evaluating the effect of emerging packaging materials and technologies on fresh-cut product quality and safety. The emerging packaging materials will include bio-based plastics, modified paper, biodegradable multilayer structures, and biocomposites. Naturally occurring polymers and by-products from the food industry will be used as raw materials and processed with the same equipment as the one currently used by industry (blown film extruders, thermoforming machines, coating machines). Different processing variables will be explored for optimization. These materials will be evaluated for barrier, mechanical, optical, and thermal properties before and after use for optimization after determining produce/packaging interactions. Emerging technologies like active and intelligent packaging will be developed. Active compound selection for development of active packages will be based on the performance of highly porous compounds or not that have the capability to encapsulate and/or release substances that affect produce quality and safety (e.g., antimicrobials, water, carbon dioxide, ethylene). Furthermore, new control release technologies for active compounds will be developed and assessed on produce quality and safety. The effects of temperature, RH, and others on the adsorption/absorption and release of gases and vapors will be assessed. Novel techniques to add the active compounds to the packaging will be developed. New and current packaging systems will be compared to evaluate improvements in quality and safety of the fresh-cut products. The behavior of different packaging systems in the cold chain will be evaluated for improvement of safety and quality of fresh-cut fruits and vegetables. Consumers’ acceptance for some of these novel packaging technologies for fresh-cut produce will be evaluated using surveys and sensory evaluations. The contribution of produce to household food waste in the US and how this contribution relates to packaging will be evaluated using surveys as well.

Quality evaluations to assess the effects of product selection, pre- and post-cutting treatments, packaging technologies, and changes during storage include physicochemical analyses and sensory evaluations. Our strategy will be to first evaluate the effects of chemical and physical treatments on visual quality to determine whether the treatments are worth further study in terms of more complex physicochemical analyses and sensory evaluations during post-treatment storage periods at 1 to 10°C depending on commodity and commercial practices. Physicochemical techniques will include mostly instrumental measurements of surface pH, phytonutrient levels, surface color, firmness, sugar and acid levels, aromatic volatile abundance, microbial loads, and dissolved solid/electrolyte contents. Standardized methods developed in our previous project or in Objective 1 will be used.

The effects of various pre- and post-cutting treatments on fresh-cut vegetable and fruit quality attributes including microbe levels is an area that will involve interaction between the physiologists and microbiologists in the project. Mesophilic bacteria, yeasts, and molds will be quantified using standard plate count. A specific growing media, and storage temperature and time will be selected depending on the type of microorganism. Work in those areas described above will be conducted in ARS-FL, ARS-MD, FL, and MI.

Objective 3 - Improve understanding of physiological mechanisms that affect fresh-cut product quality

Procedures: In this objective, we will concentrate on investigating the role of ethylene and stress physiology in fresh-cut product quality and shelf life and using transcriptome analysis to identify genes involved in both positive and negative quality traits. Newer germplasm with traits that are desirable for fresh-cut products, including genetically modified organisms, will be used to evaluate the potential for manipulating physiology to improve marketability. An example is the Arctic series of non-browning apples lacking PPO.

We propose to determine the effects of wounding and heat stress on the tissue antioxidative capacity and concentrations of bioactive components during storage of fresh-cut vegetables and fruits. We will measure the levels of active oxygen species and other free radicals in response to fresh-cut processing and investigate how postharvest treatments such as MAP and heat can be used to enhance the plant antioxidant system in order to prevent the accumulation of those damaging compounds (FL and ARS-MD). Ethylene-dependent and ethylene-independent wound responses in fresh-cut vegetables and fruits will be investigated in FL by utilizing inhibitors of ethylene binding such as 1-MCP.

Standard spectrophotometric and HPLC analyses of chlorophyll, anthocyanins, carotenoids, flavonols, tocopherols, ascorbates, and phenolic pigments (Tomás-Barberán et al., 1997) will be used. Antioxidant capacity will be measured using the oxygen radical absorbance capacity (ORAC) assay. Contents of cysteine, reduced and oxidized glutathione and activities of dehydroascorbate reductase, ascorbate peroxidase, peroxidase, catalase, superoxide dismutase, glutathione reductase, and glutathione S-transferase, will be measured as previously described (Soto-Zamora et al., 2005). Lipid class analysis will be conducted as previously described by Picchioni et al. (1996), and lipid degrading enzyme activity as described by Todd et al. (1992).

We will use genomic tools to understand the changes occurring in fresh-cut tissues that affect sensory quality and to identify targets for genetic manipulation in order to improve fresh-cut quality. RNA will be extracted, RNA-seq libraries will be constructed, and the RNA data analyzed to determine differential expression between treatments or with and without cutting; transcript level studies will then be performed to validate the differentially expressed genes (DEGs) observed during the RNA-sequencing analysis (Liu et al., 2022). The identified genes will serve as targets for marker-assisted breeding and for genetic modification via CRISPR/Cas9 to create varieties with superior quality as fresh-cut products. Work in this area will be conducted in FL and WA.

Objective 4 - Determine critical factors in controlled inoculation studies with human pathogens and surrogates that influence the outcome of quantitative microbial risk assessments.

Procedures: 1) We propose to develop research best-practice guidance and standardized methods for food safety risk assessments of fresh-cut product quality enhancing treatments or efficacy of interventions and disinfection measures. 2) We will evaluate the influence of inoculum production, method of application, rate of drying, potential for sub-lethal injury on quantitative and qualitative recovery of pathogens, duration and condition of storage, biofilm development, and method of viable recovery. 3) We will validate culture-based and non-cultural methods for improved sensitivity and specificity of detecting fecal contaminants on fresh-cut products and processing environments. 4) We will determine the potential for fresh-cut product/modified atmosphere packaging combinations and conditions to influence pathogen gene expression related to virulence following consumption.

Whole and fresh-cut produce will be inoculated with the appropriate pathogen of interest or its surrogate to give an initial microbial load of circa 106 CFU/g. The potential for reducing the transfer of microorganisms of interest from the peel to the flesh of produce during fresh-cut preparation will be evaluated by testing resulting fresh-cut pieces for pathogen contamination. The potential for injured cells of interest to grow following storage will be also evaluated after 1 to 21 days in storage by plating to selective and nonselective media. Subtracting populations enumerated on selective media from those on nonselective media gives an estimate of the number of injured cells present.

Quality and safety concerns often overlap within the food industry. Although spoilage is a natural event in food shelf life, rapid spoilage may be an indicator of changes in the storage conditions (i.e. higher temperature, increased humidity/moisture, change in pH, etc.). These conditions also promote pathogenic microorganism growth or rapid growth. Additionally, spoilage organisms are typically found in a higher abundance in food types, which makes them an easy indicator of food safety concerns. Aerobic plate counts for example are utilized by the food industry to determine both quality and food safety risk within quality assurance plans. Therefore, research under this objective will simultaneously involve testing of various intervention strategies compared to standard commercial practices for effects on produce quality and naturally occurring and inoculated microorganisms. Factors such as concentrations, treatment time, intensity, power strength, storage condition (temperature, atmosphere, duration, etc.), and synergistic interactions of sanitizers and/or intervention technologies will be evaluated. Interventions may include hot water immersion (FL), ClO2and O3 gas (FL, MI), chemical sanitizers (acids, chlorine, peroxiacetic acid, and hydrogen peroxide; ARS-MD) combined with surfactants and other treatments (hurdle technology), such as ultrasonic and UV, and other novel chemical approaches, including potentially synergistic systems containing mixtures of functional food ingredients and antimicrobial packaging and coatings (FL, MI). Natural antimicrobial compounds (phyto and bacto antimicrobials) to inactivate food borne pathogens in fresh and fresh-cut produce will also be evaluated (ARS-PA and MI). The impact of novel methods such as cold plasma and plasma-infused water will be compared to more conventional methods (O3 and electrolyzed water) on the reduction of pathogens in fresh and fresh-cut produce and in water for quarantine treatment of specific commodities (MI).

Objective 5 - Development and validation of novel diagnostic methods to determine presence of human pathogens and chemical hazards associated with fresh and fresh-cut products

Procedures: Studies will evaluate the sensitivity and specificity of new or currently used tools to determine the presence of biological and chemical hazards that are associated with fresh and fresh-cut products. Research under this objective will be put into two areas: 1) Real-time PCR and 2) biosensor platform in MS. Studies will include development and evaluation of selective detection and quantification of target pathogen cells (Salmonella, E. coli O157:H7, Listeria monocytogenes and other target foodborne pathogens) in processing waters of fresh-cut produce.

Real-time RT-PCR methods for the detection of food safety organisms from intact and fresh-cut produce will be developed and (WSU). Real-time RT-PCR methods and rapid alternatives to direct nucleic acid sequencing for the detection or fingerprinting of Salmonella spp., Escherichia coli O157:H7, Listeria monocytogenes and other human pathogens like noroviruses from intact or fresh-cut produce, including leafy greens, tomatoes, and peppers, will be developed (MS).

Biosensor platforms will be tested for selective detection and quantification of target pathogen cells in processing waters of fresh-cut produce (MS). Biosensor surface will be functionalized using specifies-specific and genus-specific antibodies raised against Salmonella, E. coli O157:H7, Listeria monocytogenes and other target foodborne pathogens. The selective biosensor surfaces will be exposed to water samples from the processing lines of fresh-cut produce to capture target pathogen cells. For the real time label-free detection, the optical spectrum of the biosensor surfaces will be measured for changes in signal due to the binding of target cells to specific antibodies. These novel biosensor surfaces will be evaluated for their sensitivity by directly exposing to processing water samples from fresh-cut produce with and without pre-enrichment for a reliable fast real-time detection of target pathogen cells. These novel biosensor assays will be standardized using spiked pathogen cells and will be confirmed by real-time PCR and culture methods. Strong adherent foodborne bacterial cells to fresh produce and to food-contact processing surfaces and the resulting biofilm formation will be determined in the presence of traces of residue from fresh-cut produce. Samples collected before and after appropriate cleaning and sanitization chemicals will be tested for efficient removal of strongly adherent biofilms of target pathogen cells from produce and from different food-contact processing surfaces by culture and novel biosensor methods (MS).

Data will be collected and analyzed for commercialization of technology for produce industry use.


Measurement of Progress and Results


  • • The S-294 Technical Committee will issue annual project reports highlighting the results for the previous year, which will be made generally available on the project website.
  • • Participants will submit research findings for publication in peer reviewed and non-refereed journals and trade publications.
  • • Fruit and vegetable germplasm with outstanding sensory quality for use as fresh-cut products will be identified.
  • • Improved processing and packaging strategies to better maintain fresh-cut product quality will be identified or developed.
  • • Effects of wounding and heat stress on the tissue antioxidative capacity and concentrations of bioactive components during storage of fresh-cut vegetables and fruits will be determined.
  • • Ethylene-dependent and ethylene-independent wound responses in fresh-cut vegetables and fruits will be identified.
  • • Research best-practice guidance and standardized methods for food safety risk assessments of fresh-cut product treatments will be developed.
  • • We will validate culture-based and non-cultural methods for improved sensitivity and specificity of detecting fecal contaminants on fresh-cut products and processing environments.
  • • We will determine the potential for fresh-cut product/modified atmosphere packaging combinations and conditions to influence pathogen gene expression related to virulence following consumption.
  • • Novel produce packaging made from renewable resources and by-products will be developed.
  • • We will investigate consumers’ acceptance for novel packaging technologies for whole and fresh-cut produce.
  • • The tolerance at different whole and fresh-cut products to hot water immersion and effects on pathogen populations will be determined.
  • • The efficacy of ClO2 and O3 gas in reducing microbial populations on whole as well as fresh-cut fruits and vegetables will be determined.
  • • The changes occurring in fresh-cut tissues that affect sensory quality will be understood by the use of genomic tools.
  • • We will identify targets for genetic manipulation to improve fresh-cut quality using genomic tools.
  • • Potentially synergistic systems containing mixtures of functional food ingredients will be evaluated for the control of human pathogens on produce.
  • • Natural antimicrobial compounds to inactivate food borne pathogens in fresh and fresh-cut produce will be evaluated.
  • • Novel antimicrobial packaging and coating for fresh-cut fruits and vegetables will be developed.
  • • Novel diagnostic methods will be developed and tested to determine presence of human pathogens and chemical hazards associated with fresh and fresh-cut products.
  • • A final report will be issued at the conclusion of the project.

Outcomes or Projected Impacts

  • • Relevant information will be available to fresh-cut processors to assist in selection of optimal harvest maturity, processing procedures, handling and packaging conditions to best maintain fresh-cut product quality and safety.
  • • The fresh-cut industry will achieve considerable savings (potentially millions of dollars a year) from reductions of product losses and recalls.
  • • Incidence of fresh-cut products at retail with insufficient shelf life for consumer satisfaction will decrease.
  • • Human health will be improved as a result of increased consumption of vegetables and fruits.
  • • Availability of best-practice guidance and standardized methods for food safety risk assessments of fresh-cut product treatments will reduce the likelihood of food safety events by replacing ineffective food safety practices with science-based procedures.
  • • Food safety risk will be reduced through availability of new, more efficacious, strategies for controlling human pathogens.
  • • Researchers will have standard protocols for quantifying shelf life and standard microbiological methods.
  • • Fastest and most accurate diagnostic methods available for detecting presence of pathogens and chemical hazards will be identified and made available.
  • • Longer-term scientific benefits will be derived from obtaining a better understanding of ethylene and stress physiology of wounded plant tissues.


(2023):Recruit new members. Redesign, expand and update the content of the project website. Meet with IFPA; explore the possibility of conducting a research-industry symposium at ARS-MD Compile methods used by project members for consumer and sensory panel testing of fresh-cut products. Organize a task force to develop the procedures for assessing food safety risks in fresh-cut processing. Identify the most promising intervention strategies to reduce microbial populations on fresh-cut products. Identify the most promising diagnostic methods to determine presence of human pathogens and chemical hazards.

(2024):Identify fruit and vegetable cultivars that possess the highest levels of flavor-based quality factors and nutritionally important components for use in further project research efforts. Identify the critical factors in controlled inoculation studies with human pathogens and surrogates. Develop novel sustainable packaging. Initiate collaborations between participating institutions for multi-station coordinated testing of sanitizers for fresh-cut products and for measuring shelf life of fresh-cut products. Obtain stakeholder feedback on quality and safety procedures and activities planned so far.

(2025):Establish collaborations between participating institutions for research on new pre-cutting and post-cutting treatments including packaging to better maintain fresh-cut product quality. Establish collaborations for developing and testing diagnostic methods to determine presence of human pathogens and chemical hazards. Establish collaborations for developing novel sustainable packaging.

(2026):Identify additional or alternative intervention strategies to investigate for controlling human pathogens on fresh-cut produce; prioritize the approaches and conduct collaborative studies. Identify other pre-cutting and post-cutting treatments to better maintain fresh-cut product quality; prioritize the approaches and conduct collaborative studies.

(2027):Develop research best-practice guidance and standardized methods for food safety risk assessments of fresh-cut product, quality enhancing treatments, and efficacy of disinfection measures; present the recommendations to the industry. Issue final project report.

Projected Participation

View Appendix E: Participation

Outreach Plan

We will work closely with produce industry professionals, specifically the IFPA (Internatonal Food Produce Associaiton), to identify industry issues to be addressed in our research and outreach activities. Every year the S294 annual meeting is held in conjunction with the IFPA convention.  Members of S294 are appointed members of the IFPA Food Safety & Technology Council  and attend that group’s meeting the day before the S294 project meeting. During the meeting, current research being conducted by researchers in the S294 group is presented.  During the S294 meeting, we have members from the produce industry attend our meeting to provide guidance and insight into the current research.

Educational materials will be created related to fresh-cut processing best practices in terms of both product quality and safety. These materials will be incorporated into existing training programs being conducted by S294 participants. IFPA has insitituted a new emaphasis on trainings developed in collaboration with S294 – two joint Fresh-cut Produce trainings have been conducted so far, in November 2021 and March 2022.  We will also conduct directed studies to evaluate the food safety behavioral changes, attitudes, and knowledge changes of produce industry members as a result of educational materials and trainings developed by project members for fresh and fresh cut products.

Results from the proposed research activities will be disseminated via presentations at scientific and industry meetings and conventions (especially Institute of Food Technologists Annual Meeting and Food Expo, and American Society for Horticultural Science Annual Meeting), and will be published as project reports posted to the project web site, as well as in peer-reviewed and non-refereed (popular) publications. Participation by project members involved in undergraduate teaching, graduate student advisement, and extension activities associated with Land Grant Universities will promote the general dissemination of knowledge developed in the proposed project.


The offices of the Technical Committee will be the chair, vice-chair, secretary, and past-chair and will serve as the Executive Committee. The first three officers will be elected for two-year terms at the organizational meeting for the Technical Committee. Thereafter, a secretary will be elected biennially at the Technical Committee meeting. All voting members of the Technical Committee will be eligible for office. The officers will be promoted biennially in the following sequence: secretary to vice-chair, vice-chair to chair, chair to past-chair. These four officers will then constitute the Executive Committee to provide leadership and continuity and/or immediate action. The duties of the chair, vice-chair, and secretary will be as prescribed in the Guidelines for Multistate Research Activities; that of the past-chair will be to serve as a resource for the other committee members in carrying out their duties.

The project will have two subcommittees with chairs appointed by the Executive Committee chair. These will be the 'Quality and Physiology Subcommittee' and the 'Microbiology Subcommittee'. Members of each subcommittee will be those Technical Committee members whose research falls under the subcommittee area. The purpose of the subcommittees is to coordinate research activities within each associated research objective, foster grant-writing activities, and create reports as directed by the Executive Committee.

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Centro de Investigación y Tecnología Agroalimentaria de Aragón, Pennsylvania (ERRC), Tennessee Tech University, USDA ARS, USDA-ARS Beltsville Agricultural Resarch Center, USDA-ARS/Florida, USDA-ARS/WRRC
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