Project : USDA ARS

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Research Project: Integration of Multiple Interventions to Enhance Microbial Safety, Quality, and Shelf-life of Foods

Location: Microbial and Chemical Food Safety

2020 Annual Report

Objectives
This project will focus on the integration of effective intervention technologies and treatments to enhance microbial safety of fresh fruits and vegetables with a holistic approach addressing major elements (safety, quality, and shelf-life), necessary for the implementation of technologies. The ultimate goal is to reduce the risk of foodborne illnesses associated with consumption of fresh produce, while maintaining acceptable food quality and shelf-life. Specific objectives of the research program are: Objective 1 - Develop and optimize single intervention technologies to reduce pathogen populations, maintain quality, and extend shelf-life of foods. Sub-objective 1.1. Develop and optimize aerosolizing technology and pulsed light to reduce pathogen populations, maintain sensorial and nutritional quality, and extend shelf-life of fresh produce. Sub-objective 1.2. Develop new antimicrobial packaging/coating-based technologies by incorporating natural, modified and novel materials to inactivate foodborne pathogens on fresh produce. Objective 2 - Determine the synergistic/additive effectiveness of combining non-thermal processing, antimicrobial packaging and effective chemical interventions utilizing information generated from the first objective.

Approach
An integrated approach to enhance microbial safety while maintaining product quality and extending shelf life of fresh produce will be adopted by combining aerosolizing antimicrobials, pulsed light antimicrobial packaging and other interventions. The types of fresh and fresh-cut produce evaluated in the project will be those frequently involved in outbreaks of foodborne illnesses, those that are hard to sanitize due to surface characteristics, and those that cannot be washed. During the first part of the proposed project plan, we will develop and optimize new aerosolization systems, pulsed light technology, novel antimicrobial coating with incorporation of nature and bio-based substances, and antimicrobial packaging materials with controlled-release mechanisms triggered with either acids or pulsed light. The optimized/developed interventions and antimicrobial packaging will then be combined with each other, and with other effective antimicrobial treatments to study the synergistic or additive effects on pathogen inactivation while maintaining quality and shelf-life of fresh produce. When selecting combinations, technologies with different pathogen-inactivation mechanisms or synergistic interactions will be desirable. We will utilize the advanced oxidation, photochemical, and photothermal, and triggered-release mechanisms and other hurdle technologies to increase the efficacy of combined antimicrobial treatments. By combining effective intervention technologies and treatments, synergistic effects with a targeted 5-log reduction of common pathogens may be achieved. Pathogens to be included in the proposed project plan are Salmonella spp., E. coli O157:H7, L. monocytogenes and other emerging pathogens (such as non-O157 STECs). We will use a representative cocktail of 3-5 strains from each genera of bacteria that are associated with outbreaks of relevant fresh produce. Scientifically well-established inoculation, recovery, and enumeration procedures will be used. Appropriate controls will be included in each experiment, and experiments will be replicated independently at least three times. Inoculation of fresh produce will be achieved either by surface ‘spot inoculation’ where specific locations on the produce surface will be inoculated or by a ‘dip inoculation’ technique where the whole produce item will be submerged in the experimental inocula. The inoculated fresh produce will be drained and air dried in a laminar flow hood before being subjected to various treatments. After treatment with various chemicals and physical interventions, the total number of viable and injured bacteria will be determined using amended media. The effects of the individual and combined treatments on the physiochemical and sensorial quality and shelf-life will be evaluated during storage. Shelf-life will be determined based on the deterioration in product quality and increasing populations of microorganisms that render the product unacceptable to consumers.

Progress Report
Progress was made on Objective 2, which falls under National Program 108, Component I, Foodborne Contaminants, and National Action Plan Problem Statement 5, Intervention and Control Strategies. Experiments were conducted to develop formulations for the inactivation of Listeria monocytogenes while maintaining freshness of cut apples, and employ advanced oxidation processes by combining titanium dioxide (TiO2)-based antimicrobial packaging systems with pulsed light, and integrating pulsed light technology with aqueous sanitizers to achieve synergistic or additive reductions of human pathogens and spoilage microorganisms while maintaining quality of fresh produce. Two patent applications have been filed relating to the development of novel bio-based antimicrobials. Due to the COVID-19 pandemic, not all milestone projects are fully completed. Detailed progress to achieve the overall objectives is listed below. A number of natural and bio-based antimicrobials were screened either individually or in combinations for their efficacy in inactivating a cocktail of Listeria monocytogenes. Furthermore, those showing anti-listerial activity were applied onto cut apples to evaluate their ability in inhibiting surface browning of the fruits, while maintaining firmness. Results showed some combinations were able to achieve more than 99.999% reductions of the pathogen while preserving the freshness of cut-fruit. Research continues to optimize the formulations and assess shelf-life of treated cut fruits. Experiments were conducted to develop antimicrobial packaging films containing nano scale TiO2. Polylactic acid (PLA)-TiO2 antimicrobial films were produced using a solvent cast method by mixing PLA powder with TiO2 nano particles. The antimicrobial activity of the film was activated by normal fluorescent light and enhanced by pulsed light. The populations of Listeria and E. coli on PLA-TiO2 antimicrobial films were reduced by 96% and 98.5% under normal light conditions, respectively, while pulsed light for 5 seconds achieved over 99.9% reduction of the bacteria. Light-activated PLA-TiO2 antimicrobial films maintained antibacterial activity and had lower bacterial populations than that without light-activated films. The results demonstrate that the developed antimicrobial packaging material can be used to fabricate food containers or packaging films with pathogen-free surface. Further studies will be conducted with real food. Experiments were also conducted to develop and validate a method using low dose/short duration pulsed light (PL) in presence of hydrogen peroxide (H2O2) (1-3%) and in presence or absence of TiO2 to investigate the efficacy of advanced oxidation process (AOP). In the AOP treatment, PL interacts with H2O2 and TiO2, creating highly reactive hydroxyl radicals possessing strong oxidizing capability to inactivate pathogens and other microorganisms on the surfaces of produce. In a laboratory scale study, a brief 30 s pulsed light treatment with 3% H2O2 provided a 99.9% reduction of a three serotype Salmonella composite inoculated on blueberries. Effectiveness of inactivation was increased to 99.99% when TiO2 (10 ppm) was added to the system. No difference in decontamination efficacy was observed between 30 s and 60 s PL/H2O2 treatments. The AOP treatment also reduced native microbial populations on blueberries and slowed their growth during storage. Overall, results demonstrate that PL/H2O2 integrated AOP technology can be used to enhance microbial safety of produce.

Accomplishments
1. Combining cold plasma and hydrogen peroxide enhances the microbial safety of fruits. ARS scientists at Wyndmoor, Pennsylvania, combined cold plasma and hydrogen peroxide aerosols to produce highly reactive radicals that reduced the populations of bacteria on fresh fruits. Applying cold plasma to hydrogen peroxide increased the effectiveness of hydrogen peroxide aerosols, and killed almost 100 percent of Salmonella and Listeria on surfaces of apples, cantaloupe, and tomatoes. This new process did not affect appearance, color, texture or nutritional quality of the produce. These results would be of interest to produce industry seeking to enhance the microbial safety while keeping the fruits fresh longer. ARS scientists are collaborating with industry partners to explore the commercial applicability of the technology.

2. Pulsed light and Nisin-based sanitizer combination treatment can assure microbial safety of produce. Currently, produce industry employs chlorine-based sanitizers to reduce cross contamination despite issues related to their safety and effectiveness. Hence, there is a need to develop chlorine-free decontamination technologies. Researchers at Wyndmoor, Pennsylvania, developed a safe and effective method combining pulsed light with a Nisin-based sanitizer. This combined treatment method of pulsed light and the Nisin-based sanitizer, exhibited a synergistic activity killing almost one hundred percent of pathogens like Salmonella in tomatoes. The combination treatment is also proven effective in protecting sensory qualities like visual appearance and firmness of tomatoes. This new combined treatment can be used as a replacement for current chlorine-based sanitizers wash in the tomato industry.

3. Application of essential oil washing and vapor treatments for food safety and shelf life. Fresh blackberries are a nutritious fruit, but they have a short shelf life and are contaminated easily by pathogenic spoilage microorganisms. ARS scientists at Wyndmoor, Pennsylvania, investigated methods to reduce microbial contaminants on blackberries using essential oil wash, vapor, and their combination. The combination of essential oil wash and vapor treatments completely inhibited the growth of bacteria and fungi and extended the shelf life of fresh blackberries from 5 days to over 12 days at cool temperatures (10 degrees centigrade). The study shows that the essential oil treatment is good for maintaining the microbiological and nutritional qualities and extending the shelf life of blackberries.

U.S. DEPARTMENT OF AGRICULTURE

Microbial and Chemical Food Safety: Wyndmoor, PA