Plasma-activated liquids for mitigating biofilms on food and food contact surfaces.

Plasma-activated liquids (PALs) are emerging and promising alternatives to traditional decontamination technologies and have evolved as a new technology for applications in food, agriculture, and medicine. Contamination caused by foodborne pathogens and their biofilms has posed challenges and concerns to the food industry in terms of safety and quality. The nature of the food and the food processing environment are major factors that contribute to the growth of various microorganisms, followed by the biofilm characteristics that ensure their survival in severe environmental conditions and against traditional chemical disinfectants. PALs show an efficient impact against microorganisms and their biofilms, with various reactive species (short- and long-lived ones), physiochemical properties, and plasma processing factors playing a crucial role in mitigating biofilms. Moreover, there is potential to improve and optimize disinfection strategies using a combination of PALs with other technologies for the inactivation of biofilms. The overarching aim of this study is to build a better understanding of the parameters that govern the liquid chemistry generated in a liquid exposed to plasma and how these translate into biological effects on biofilms. This review provides a current understanding of PALs-mediated mechanisms of action on biofilms; however, the precise inactivation mechanism is still not clear and is an important part of the research. Implementation of PALs in the food industry could help overcome the disinfection hurdles and can enhance biofilm inactivation efficacy. Future perspectives in this field to expand existing state of the art to seek breakthroughs for scale-up and implementation of PALs technology in the food industry are also discussed.

Inactivation efficacy of atmospheric air plasma and airborne acoustic ultrasound against bacterial biofilms.

Biofilms are complex microbial communities that present serious contamination risks to our environment and health. In this study, atmospheric air plasma and airborne acoustic ultrasound technology were applied to inactivate Escherichia coli and Listeria innocua biofilms. Both technologies were efficient in controlling, or completely inactivating, the target bacterial biofilms. Viability and metabolic assays, along with microscopy analysis, revealed that atmospheric air plasma and airborne acoustic ultrasound damaged both the bacterial biofilm cells and its structural integrity. Scanning electron microscopy images highlighted the disruption of the biofilms and pore formation in bacterial cells exposed to both the plasma and acoustic treatments. Elevated reactive oxygen and nitrogen species in bacterial cells treated with atmospheric air plasma, demonstrated their primary role in the observed bacterial inactivation process. Our findings provide potential antimicrobial strategies to combat bacterial biofilms in the food and healthcare sectors.

Plasma-activated water: Physicochemical properties, microbial inactivation mechanisms, factors influencing antimicrobial effectiveness, and applications in the food industry.

Novel nonthermal inactivation technologies have been increasingly popular over the traditional thermal food processing methods due to their capacity in maintaining microbial safety and other quality parameters. Plasma-activated water (PAW) is a cutting-edge technology developed around a decade ago, and it has attracted considerable attention as a potential washing disinfectant. This review aims to offer an overview of the fundamentals and potential applications of PAW in the agri-food sector. A detailed description of the interactions between plasma and water can help to have a better understanding of PAW, hence the physicochemical properties of PAW are discussed. Further, this review elucidates the complex inactivation mechanisms of PAW, including oxidative stress and physical effect. In particular, the influencing factors on inactivation efficacy of PAW, including processing factors, characteristics of microorganisms, and background environment of water are extensively described. Finally, the potential applications of PAW in the food industry, such as surface decontamination for various food products, including fruits and vegetables, meat and seafood, and also the treatment on quality parameters are presented. Apart from decontamination, the applications of PAW for seed germination and plant growth, as well as meat curing are also summarized. In the end, the challenges and limitations of PAW for scale-up implementation, and future research efforts are also discussed. This review demonstrates that PAW has the potential to be successfully used in the food industry.

Antimicrobial effects of airborne acoustic ultrasound and plasma activated water from cold and thermal plasma systems on biofilms.

Bacterial biofilms are difficult to inactivate due to their high antimicrobial resistance. Therefore, new approaches are required for more effective bacterial biofilm inactivation. Airborne acoustic ultrasound improves bactericidal or bacteriostatic activity which is safe and environmentally friendly. While, plasma activated water (PAW) is attracting increasing attention due to its strong antimicrobial properties. This study determined efficacy of combined airborne acoustic ultrasound and plasma activated water from both cold and thermal plasma systems in inactivating Escherichia coli K12 biofilms. The application of airborne acoustic ultrasound (15 min) alone was significantly more effective in reducing E. coli counts in 48 and 72 h biofilms compared to 30 min treatment with PAW. The effect of airborne acoustic ultrasound was more pronounced when used in combination with PAW. Airborne acoustic ultrasound treatment for 15 min of the E. coli biofilm followed by treatment with PAW significantly reduced the bacterial count by 2.2-2.62 Log10 CFU/mL when compared to control biofilm treated with distilled water. This study demonstrates that the synergistic effects of airborne acoustic ultrasound and PAW for enhanced antimicrobial effects. These technologies have the potential to prevent and control biofilm formation in food and bio-medical applications.

Efficacy of cold plasma functionalised water for improving microbiological safety of fresh produce and wash water recycling.

Atmospheric cold plasma (ACP) is an effective method for microbiological decontamination. This study evaluated an alternative water-based decontamination approach for inactivation of bacterial population from fresh produce and in the wash water generated from fresh produce washing. The study characterised ACP inactivation of attached Listeria innocua and Pseudomonas fluorescens inoculated on lettuce in comparison to chlorine treatment. P. fluorescens was sensitive to ACP treatment and was reduced below detection limit within 3 min of treatment. L. innocua population was reduced by ∼2.4 Log10 CFU/g after 5 min of treatment; showing similar inactivation efficacy to chlorine treatment. The microbial load in wash water was continuously decreased and was below detection limits after 10 min of ACP treatment. Micro-bubbling along with agitation assisted the bacterial detachment and distribution of reactive species, thus increasing bacterial inactivation efficacy from fresh produce and wash water. A shift in pH of plasma functionalised water was observed along with high concentration of nitrate and ozone with a relative amount of nitrites which increased with plasma exposure time. Further, L. innocua treated at different independent pH conditions showed minimal or no effect of pH on ACP bacterial inactivation efficacy. Aqueous ACP treatment poses a promising alternative for decontamination of fresh produce and the associated wash-waters which could be applied in the food industry to replace continuous chlorine dosing of process waters.

High voltage atmospheric cold air plasma control of bacterial biofilms on fresh produce.

Atmospheric cold plasma (ACP) offers great potential for decontamination of food borne pathogens. This study examined the antimicrobial efficacy of ACP against a range of pathogens of concern to fresh produce comparing planktonic cultures, monoculture biofilms (Escherichia coli, Salmonella enterica, Listeria monocytogenes, Pseudomonas fluorescens) and mixed culture biofilms (Listeria monocytogenes and Pseudomonas fluorescens). Biotic and abiotic surfaces commonly occurring in the fresh food industry were investigated. Microorganisms showed varying susceptibility to ACP treatment depending on target and process factors. Bacterial biofilm populations treated with high voltage (80 kV) ACP were reduced significantly (p < 0.05) in both mono- and mixed species biofilms after 60 s of treatment and yielded non-detectable levels after extending treatment time to 120 s. However, an extended time was required to reduce the challenge mixed culture biofilm of L. monocytogenes and P. fluorescens inoculated on lettuce, which was dependent on biofilm formation conditions and substrate. Contained treatment for 120 s reduced L. monocytogenes and P. fluorescens inoculated as mixed cultures on lettuce (p < 0.05) by 2.2 and 4.2 Log10 CFU/ml respectively. When biofilms were grown at 4 °C on lettuce, there was an increased resistance to ACP treatment by comparison with biofilm grown at temperature abuse conditions of 15 °C. Similarly, L. monocytogenes and P. fluorescens exposed to cold stress (4 °C) for 1 h demonstrated increased tolerance to ACP treatment compared to non-stressed cells. These finding demonstrates that bacterial form, mono versus mixed challenges as well as environmental stress conditions play an important role in ACP inactivation efficacy.

The Effect of Atmospheric Cold Plasma on Bacterial Stress Responses and Virulence Using Listeria monocytogenes Knockout Mutants.

Listeria monocytogenes is an opportunistic intracellular pathogen commonly associated with serious infections and multiple food-borne outbreaks. In this study, we investigated the influence of atmospheric cold plasma (80 kV, 50 Hz) on L. monocytogenes (EGD-e) and its knockout mutants of sigB, rsbR, prfA, gadD, and lmo0799 genes at different treatment time intervals. Further, to ascertain if sub-lethal environmental stress conditions could influence L. monocytogenes survival and growth responses, atmospheric cold plasma (ACP) resistance was evaluated for the cultures exposed to cold (4°C) or acid (pH 4) stress for 1 h. The results demonstrate that both wild-type and knockout mutants were similarly affected after 1 min exposure to ACP (p > 0.05), with a difference in response noted only after 3 min of treatment. While all L. monocytogenes strains exposed to acid/cold stress were hypersensitive to ACP treatment and were significantly reduced or inactivated within 1 min of treatment (p < 0.05). The results indicate sigB and prfA are important for general stress resistance and biofilm, respectively, loss of these two genes significantly reduced bacterial resistance to ACP treatment. In addition, exposure to sub-lethal 1min ACP increased the gene expression of stress associated genes. SigB showed the highest gene expression, increasing by 15.60 fold, followed by gadD2 (7.19) and lmo0799 (8.6) after 1 min exposure. Overall, an increase in gene expression was seen in all stress associated genes analyzed both at 1 min treatment; while long treatment time reduced the gene expression and some cases down-regulated prfA and gadD3 gene expression. By comparing the response of mutants under ACP exposure to key processing parameters, the experimental results presented here provide a baseline for understanding the bacterial genetic response and resistance to cold plasma stress and offers promising insights for optimizing ACP applications.

Assessment of the disinfection capacity and eco-toxicological impact of atmospheric cold plasma for treatment of food industry effluents.

Generation of wastewater is one of the main environmental sustainability issues across food sector industries. The constituents of food process effluents are often complex and require high energy and processing for regulatory compliance. Wastewater streams are the subject of microbiological and chemical criteria, and can have a significant eco-toxicological impact on the aquatic life. Thus, innovative treatment approaches are required to mitigate environmental impact in an energy efficient manner. Here, dielectric barrier discharge atmospheric cold plasma (ACP) was evaluated for control of key microbial indicators encountered in food industry effluent. This study also investigated the eco-toxicological impact of cold plasma treatment of the effluents using a range of aquatic bioassays. Continuous ACP treatment was applied to synthetic dairy and meat effluents. Microbial inactivation showed treatment time dependence with significant reduction in microbial populations within 120 s, and to undetectable levels after 300 s. Post treatment retention time emerged as critical control parameter which promoted ACP bacterial inactivation efficiency. Moreover, ACP treatment for 20 min achieved significant reduction (≥2 Log10) in Bacillus megaterium endospores in wastewater effluent. Acute aquatic toxicity was assessed using two fish cell lines (PLHC-1 and RTG-2) and a crustacean model (Daphnia magna). Untreated effluents were toxic to the aquatic models, however, plasma treatment limited the toxic effects. Differing sensitivities were observed to ACP treated effluents across the different test bio-assays in the following order: PLHC-1 > RTG-2 ≥ D. magna; with greater sensitivity retained to plasma treated meat effluent than dairy effluent. The toxic effects were dependent on concentration and treatment time of the ACP treated effluent; with 30% cytotoxicity in D. magna and fish cells observed after 24 h of exposure to ACP treated effluent for concentrations up to 5%. The findings suggest the need to employ wider variety of aquatic organisms for better understanding and complete toxicity evaluation of long-term effects. The study demonstrates the potential to tailor ACP system parameters to control pertinent microbial targets (mono/poly-microbial, vegetative or spore form) found in complex and nutritious wastewater effluents whilst maintaining a safe eco-toxicity profile for aquatic species.

Recent Advances in the Application of Cold Plasma Technology in Foods.

The past decade has seen a surge in the scientific literature investigating the potential food-related applications of plasma. A multidisciplinary scientific effort has started to demonstrate process efficacy for a range of plasma applications, including antimicrobial, pesticidal, food functionalization, and waste treatment. Insights into the interactions of plasma species with food and the mechanisms of action are also emerging. This review examines the current status of cold plasma technology within the food sector with a particular emphasis on emerging applications. Opportunities and current challenges that need to be addressed for successful adoption of the approach by industry are detailed.

Controlling Brochothrix thermosphacta as a spoilage risk using in-package atmospheric cold plasma.

Brochothrix thermosphacta is the predominant spoilage microorganism in meat and its control in processing environments is important to maintain meat product quality. Atmospheric cold plasma is of interest for control of pathogenic and spoilage microorganisms in foods. This study ascertained the potential of dielectric barrier discharge atmospheric cold plasma (DBD-ACP) for control of B. thermosphacta, taking microbial and food environment factors into consideration, and investigated the shelf-life of lamb chop after in-package plasma treatment in modified atmosphere. Community profiling was used to assess the treatment effects on the lamb microflora. ACP treatment (80 kV) for 30s inactivated B. thermosphacta populations below detection levels in PBS, while 5 min treatment achieved a 2 Log cycle reduction using a complex meat model medium and attached cells. The antimicrobial efficacy of plasma was reduced but still apparent on lamb chop surface-inoculated with high concentrations of B. thermosphacta. Lamb chop treated under modified atmosphere exhibited reduced microbial growth over the product shelf-life and community profiling showed no evident changes to the microbial populations after the treatment. The overall results indicated potential of ACP to enhance microbial control leading to meat storage life extension through adjusting the modality of treatment.

Controlling Microbial Safety Challenges of Meat Using High Voltage Atmospheric Cold Plasma.

Atmospheric cold plasma (ACP) is a non-thermal technology, effective against a wide range of pathogenic microorganisms. Inactivation efficacy results from plasma generated reactive species. These may interact with any organic components in a test matrix including the target microorganism, thus food components may exert a protective effect against the antimicrobial mode of action. The effect of an in-package high voltage ACP process applied in conjunction with common meat processing MAP gas compositions as well as bacteria type and meat model media composition have been investigated to determine the applicability of this technology for decontamination of safety challenges associated with meat products. E. coli, L. monocytogenes, and S. aureus in PBS were undetectable after 60 s of treatment at 80 kVRMS in air, while ACP treatment of the contaminated meat model required post-treatment refrigeration to retain antimicrobial effect. The nutritive components in the meat model exerted a protective effect during treatment, where 300 s ACP exposure yielded a maximum reduction of 1.5 log using a high oxygen atmosphere, whilst using air and high nitrogen atmospheres yielded lower antimicrobial efficacy. Furthermore, an ROS assay was performed to understand the protective effects observed using the meat model. This revealed that nutritive components inhibited penetration of ROS into bacterial cells. This knowledge can assist the optimization of meat decontamination using ACP technology where interactions with all components of the food matrix require evaluation.