Project Number: 6022-32420-001-38-R
Project Type: Reimbursable Cooperative Agreement
Start Date: Oct 1, 2020
End Date: Jun 28, 2024
Objective 1. To reduce Salmonella (S.) on eggs using ozone microbubbles applied as a post-harvest wash treatment and study effect on egg quality parameters. Objective 2. To reduce Salmonella and Campylobacter (C.) jejuni on poultry carcass using ozone microbubbles wash treatments and study effect on carcass quality. Objective 3. To reduce Salmonella and Enteritidis (E.) coli O157:H7 on fresh produce using ozone microbubble wash treatments. Objective 4. Conduct a cost/benefit analysis on the research, extension and education efforts of the project. The extension/education objectives of this proposal are to: • Disseminate results of the project to the veteran farming community, through numerous delivery methods including conferences, electronic and printed materials, and training activities in a research and teaching oriented facility. • Conduct train the trainer programs for extension specialists and other interested stakeholders regarding the objectives and results of this project.
Our approach combines the antimicrobial efficacy of Generally Recognized As Safe (GRAS) status gas ozone with novel microbubble technology to develop natural, safe and rapid acting sanitizers with significant efficacy against foodborne pathogens such as Campylobacter (C.), Salmonella (S>) and Enteritidis (E.) coli O157:H7 in fresh produce and poultry products. Objective 1. The ozone bubbles will be generated in deionized water maintained at 4°C by using a micro/nanobubble generator. The size of ozone microbubbles will be characterized using a Nano ZS Zetasizer. The eggs will be spot-inoculated with 200 µl of a 5-serotype cocktail of S. Enteritidis on the egg surface, and air-dried for 1 h at 23°C in a biosafety cabinet to facilitate bacterial attachment onto egg surface. The efficacy of ozone microbubbles as an antimicrobial wash for killing S. Enteritidis on shell eggs will be investigated at 32°C. Five eggs will be used per treatment/time point (n=15) and the study will be repeated three times. Batches of 15 eggs each will be placed in a 2L sterile plastic bucket containing 1,000 ml of sterile deionized water (control) or water with ozone microbubbles and washed in a shaker water bath at 32°C for 1, 5, or 10 min. Water containing 200 ppm of chlorine and peracetic acid will be included as industry controls. S. Enteritidis will be enumerated by plating dilutions on XLD+NA and TSA+NA plates. The plates will be incubated at 37°C for 48 h for bacterial enumeration. Also, a batch of treated and control eggs will be refrigerated (to mimic post-harvest storage) in egg cartons for 3 weeks, and Salmonella populations on eggs will be determined on days 3, 7, 14 and 21 of storage. Objective 2. Two hundred microliters of a 5-strain mixture of S. Enteritidis and C. jejuni in Phosphate-buffered saline (PBS) will be used to spot inoculate the carcass and air-dried for 30 min at 23°C to facilitate bacterial attachment. Batches of 5 carcasses will be submerged in 20L of sterile deionized water (with or without ozone microbubbles) maintained at 4°C in a plastic tub for a period of 30 min and then refrigerated. Carcasses will be sampled at 0, 1, 3, 5 and 7 days of refrigerated storage to determine surviving Campylobacter and Salmonella counts. Objective 3. Two hundred microliters of a 5-strain mixture of S. Enteritidis and E. coli O157:H7 in PBS will be used to spot inoculate the selected fresh produce . The efficacy of ozone microbubbles in reducing Salmonella and E. coli O157:H7 on fresh produce without adversely affecting the produce quality will be assessed when used as antimicrobial wash at room temperature. Objective 4: We will perform a cost/benefit analysis that will allow for the comparison of the benefits of the ozone microbubble wash treatments, versus the cost of conducting such wash treatments. The costs will be derived in two ways. First, we will calculate the cost of the pilot program. Second, we will calculate the costs associated with a scaled-up version of the program, including extension and outreach components.