Atmospheric Cold Plasma as a Tool for Microbiological Control

Outbreaks of foodborne human illnesses resulting from contaminated raw or minimally processed fruits and vegetables have been widely reported globally. The microbiological challenges associated with fresh produce are diverse and respond differently to minimal processing technologies. Atmospheric cold plasma is a relatively new technology and represents a potential alternative to traditional methods for decontamination of foods. The objective of this work was to determine the influence of extrinsic atmospheric cold plasma (ACP) treatment control parameters and to optimize treatment parameters for decontamination with respect to different forms of key safety challenges pertinent to fresh produce. The optimisation studies demonstrated that inactivation efficacy of treatment, when tested against high populations of E. coli suspended in liquid media, was governed by the processing parameter of mode of exposure, treatment time, post treatment storage time, voltage levels, working gas and media composition. Post treatment storage time emerged as a critical treatment parameter for consistency and efficiency of bacterial inactivation with the system. The effect of media complexity was evident with higher inactivation rates achieved in media with simpler composition. Antimicrobial efficacy of ACP increased when voltage level and gas mixture with higher oxygen content was utilised, nullifying the effect of mode of ACP exposure and media composition. High voltage in-package indirect ACP treatment with 24 h of post treatment storage time, selected as the more favourable treatment approach in terms of produce quality retention, was highly effective for decontamination of cherry tomatoes and strawberries inoculated with Salmonella, E. coli and L. monocytogenes monocultures and against background microflora of produce. However, the produce type and the contaminating pathogen influenced decontaminating effect of ACP with higher inactivation rates achieved for Gram-negative bacteria and bacteria associated with smooth surface of produce. The antimicrobial potential of high voltage either direct or indirect in-package atmospheric air ACP treatment with subsequent 24 h of storage was proven to be effective for inactivation of pathogens in the form of monoculture biofilms commonly implicated in foodborne and healthcare associated human infections, E. coli, L. monocytogenes, S. aureus, P. aeruginosa established during 48 h on abiotic surface. However, the efficiency of ACP treatment was again bacterial type dependant. Although complete inactivation of metabolic activity of Gram-negative bacteria could not be achieved, electron microscopy analyses confirmed the destructive action of ACP treatment. In-package high voltage indirect ACP treatment was effective against Salmonella, L. monocytogenes and E. coli biofilms developed on lettuce. This study also demonstrated that produce storage conditions, such as temperature, light and storage time had interactive effects on bacterial proliferation, internalisation, stress response and susceptibility to the ACP treatment, highlighting the importance of preventive measures as key factors for the assurance of microbiological safety of fresh produce. Significant reductions of P. aeruginosa quorum sensing (QS)-regulated virulence factors, such as pyocyanin and elastase production, were achieved, suggesting that ACP technology could be a potential QS inhibitor and may play an important role in attenuation of virulence of pathogenic bacteria. Despite the varying parameters that influenced plasma bactericidal activity, high voltage in-package atmospheric air ACP decontamination approach showed an efficient reduction of high concentrations of bacteria in liquids, associated with produce and bacteria in their most resistant, biofilm form. These results represent significant technological advances in non-thermal bactericidal treatment with a key advantage of elimination of post-processing contamination of the product, thereby increasing microbiological safety and extension of produce shelf life

Atmospheric Cold Plasma as a Tool for Microbiological Control

Outbreaks of foodborne human illnesses resulting from contaminated raw or minimally processed fruits and vegetables have been widely reported globally. The microbiological challenges associated with fresh produce are diverse and respond differently to minimal processing technologies. Atmospheric cold plasma is a relatively new technology and represents a potential alternative to traditional methods for decontamination of foods. The objective of this work was to determine the influence of extrinsic atmospheric cold plasma (ACP) treatment control parameters and to optimize treatment parameters for decontamination with respect to different forms of key safety challenges pertinent to fresh produce. The optimisation studies demonstrated that inactivation efficacy of treatment, when tested against high populations of E. coli suspended in liquid media, was governed by the processing parameter of mode of exposure, treatment time, post treatment storage time, voltage levels, working gas and media composition. Post treatment storage time emerged as a critical treatment parameter for consistency and efficiency of bacterial inactivation with the system. The effect of media complexity was evident with higher inactivation rates achieved in media with simpler composition. Antimicrobial efficacy of ACP increased when voltage level and gas mixture with higher oxygen content was utilised, nullifying the effect of mode of ACP exposure and media composition. High voltage in-package indirect ACP treatment with 24 h of post treatment storage time, selected as the more favourable treatment approach in terms of produce quality retention, was highly effective for decontamination of cherry tomatoes and strawberries inoculated with Salmonella, E. coli and L. monocytogenes monocultures and against background microflora of produce. However, the produce type and the contaminating pathogen influenced decontaminating effect of ACP with higher inactivation rates achieved for Gramnegative bacteria and bacteria associated with smooth surface of produce. The antimicrobial potential of high voltage either direct or indirect in-package atmospheric air ACP treatment with subsequent 24 h of storage was proven to be effective for inactivation of pathogens in the form of monoculture biofilms commonly implicated in foodborne and healthcare associated human infections, E. coli, L. monocytogenes, S. aureus, P. aeruginosa established during 48 h on abiotic surface. However, the efficiency of ACP treatment was again bacterial type dependant. Although complete inactivation of metabolic activity of Gram-negative bacteria could not be achieved, electron microscopy analyses confirmed the destructive action of ACP treatment. In-package high voltage indirect ACP treatment was effective against Salmonella, L. monocytogenes and E. coli biofilms developed on lettuce. This study also demonstrated that produce storage conditions, such as temperature, light and storage time had interactive effects on bacterial proliferation, internalisation, stress response and susceptibility to the ACP treatment, highlighting the importance of preventive measures as key factors for the assurance of microbiological safety of fresh produce. Significant reductions of P. aeruginosa quorum sensing (QS)-regulated virulence factors, such as pyocyanin and elastase production, were achieved, suggesting that ACP technology could be a potential QS inhibitor and may play an important role in attenuation of virulence of pathogenic bacteria. Despite the varying parameters that influenced plasma bactericidal activity, high voltage in-package atmospheric air ACP decontamination approach showed an efficient reduction of high concentrations of bacteria in liquids, associated with produce and bacteria in their most resistant, biofilm form. These results represent significant technological advances in non-thermal bactericidal treatment with a key advantage of elimination of post-processing contamination of the product, thereby increasing microbiological safety and extension of produce shelf life

Cold Plasma For Insect Pest Control: Tribolium Castaneum Mortality and Defense Mechanisms in Response to Treatment

The insecticidal properties and mechanisms of high-voltage air-based atmospheric cold plasma using a contained dielectric barrier discharge reactor were investigated against Tribolium castaneum as an important bio-contaminant in stored grains spoilage. The mortality of 95.0%–100% for preadult stages can be achieved within seconds of treatment, but longer plasma exposure (5 min) is required to kill adult insects. Cold plasma treatment reduces both the respiration rate and the weight of insects and affects the levels of oxidative stress markers in adult populations. Sufficient toxicity is achievable through plasma process control in air to address the range of insect lifecycle stages that are disease vectors and pose risks for grain stability in storage. Balancing insecticidal activity with grains\u27 quality retention can provide a route to sustainable integrated pest management

Current and Future Technologies for Microbiological Decontamination of Cereal Grains

Cereal grains are the most important staple foods for mankind worldwide. The constantly increasing annual production and yield is matched by demand for cereals, which is expected to increase drastically along with the global population growth. A critical food safety and quality issue is to minimize the microbiological contamination of grains as it affects cereals both quantitatively and qualitatively. Microorganisms present in cereals can affect the safety, quality, and functional properties of grains. Some molds have the potential to produce harmful mycotoxins and pose a serious health risk for consumers. Therefore, it is essential to reduce cereal grain contamination to the minimum to ensure safety both for human and animal consumption. Current production of cereals relies heavily on pesticides input, however, numerous harmful effects on human health and on the environment highlight the need for more sustainable pest management and agricultural methods. This review evaluates microbiological risks, as well as currently used and potential technologies for microbiological decontamination of cereal grain

Hydra as a Model for Screening Ecotoxicological Effects of Plasma-Treated Water

Atmospheric cold plasma (ACP) has been widely researched to generate functionalized solutions for decontamination of liquids and wastewater treatment. The present investigation demonstrates that Hydra can be used as an additional in vivo tool to monitor the impact of plasma-processed solutions on the aquatic environment

Atmospheric Cold Plasma Inactivation of Escherichia Coli, Salmonella Enterica Serovar Typhimurium and Listeria Monocytogenes Inoculated on Fresh Produce

Atmospheric cold plasma (ACP) represents a potential alternative to traditional methods for non-thermal decontamination of foods. In this study, the antimicrobial efficacy of a novel dielectric barrier discharge ACP device against Escherichia coli, Salmonella enterica Typhimurium and Listeria monocytogenes inoculated on cherry tomatoes and strawberries, was examined. Bacteria were spot inoculated on the produce surface, air dried and sealed inside a rigid polypropylene container. Samples were indirectly exposed (i.e. placed outside plasma discharge) to a high voltage (70kVRMS) air ACP and subsequently stored at room temperature for 24 h. ACP treatment for 10, 60 and 120 s resulted in reduction of Salmonella, E. coli and L. monocytogenes populations on tomato to undetectable levels from initial populations of 3.1, 6.3, and 6.7 log10 CFU/sample, respectively. However, an extended ACP treatment time was necessary to reduce bacterial populations attached on the more complex surface of strawberries. Treatment time for 300 s resulted in reduction of E. coli, Salmonella and L. monocytogenes populations by 3.5, 3.8 and 4.2 log10 CFU/sample, respectively, and also effectively reduced the background microflora of tomatoes

Cold Plasma Inactivation of Internalised Bacteria and Biofilms for Salmonella Enterica Serovar Typhimurium, Listeria Monocytogenes and Escherichia Coli

Microbial biofilms and bacteria internalised in produce tissue may reduce the effectiveness of decontamination methods. In this study, the inactivation efficacy of in-package atmospheric cold plasma (ACP) afterglow was investigated against Salmonella Typhimurium, Listeria monocytogenes and Escherichia coli in the forms of planktonic cultures, biofilms formed on lettuce and associated bacteria internalised in lettuce tissue. Prepared lettuce broth (3%) was inoculated with bacteria resulting in a final concentration of ~ 7.0 log10 CFU/ml. For biofilm formation and internalisation, lettuce pieces (5 × 5 cm) were dip-inoculated in bacterial suspension of ~ 7.0 log10 CFU/ml for 2 h and further incubated for 0, 24 and 48 h at either 4 °C or room temperature (~ 22 °C) in combination with light/dark photoperiod or at 4 °C under dark conditions. Inoculated samples were sealed inside a rigid polypropylene container and indirectly exposed (i.e. placed outside plasma discharge) to a high voltage (80 kVRMS) air ACP with subsequent storage for 24 h at 4 °C. ACP treatment for 30 s reduced planktonic populations of Salmonella, L. monocytogenes and E. coli suspended in lettuce broth to undetectable levels. Depending on storage conditions, bacterial type and age of biofilm, 300 s of treatment resulted in reduction of biofilm populations on lettuce by a maximum of 5 log10 CFU/sample. Scanning electron and confocal laser microscopy pointed to the incidence of bacterial internalisation and biofilm formation, which influenced the inactivation efficacy of ACP. Measured intracellular reactive oxygen species (ROS) revealed that the presence of organic matter in the bacterial suspension might present a protective effect against the action of ROS on bacterial cells. This study demonstrated that high voltage in-package ACP could be a potential technology to overcome bacterial challenges associated with food produce. However, the existence of biofilms and internalised bacteria should be considered for further optimisation of ACP treatment parameters in order to achieve an effective control of the realistic challenges posed by foodborne pathogens

The Potential of Cold Plasma for Safe and Sustainable Food Production

In-package decontamination of foods using cold plasma has advanced this technology as a unit process for fresh foods decontamination and shelf-life extension. Chemical residues of agricultural pesticides of varying structure can be degraded to safe or less-toxic structures using cold plasma. Cold-plasma-mediated control of contaminants, along with the promotion of seed germination and plant growth, offers alternatives to current pesticides and fertilizers for agriculture. Controlling plasma reactive species formulations in dry and liquid delivery formats advances the potential for understanding and successful translation to multiple points along the agriculture and food sectors. Employing predictive microbiology, process optimization tools and a systems approach with controlled reactive species formulations may achieve risk- or problem-tailored solutions for whole food systems. Cold plasma science and technology is increasingly investigated for translation to a plethora of issues in the agriculture and food sectors. The diversity of the mechanisms of action of cold plasma, and the flexibility as a standalone technology or one that can integrate with other technologies, provide a rich resource for driving innovative solutions. The emerging understanding of the longer-term role of cold plasma reactive species and follow-on effects across a range of systems will suggest how cold plasma may be optimally applied to biological systems in the agricultural and food sectors. Here we present the current status, emerging issues, regulatory context, and opportunities of cold plasma with respect to the broad stages of primary and secondary food production

Investigation of mechanisms involved in germination enhancement of wheat (Triticum aestivum) by cold plasma: Effects on seed surface chemistry and characteristics

Recent reports indicate that atmospheric cold plasma (ACP) treatment of seeds can enhance their germination, however, the mechanisms of action are not yet entirely clear. In the present work, we report on the effects of plasma treatment on wheat seed germination and seedling growth. Additionally, changes in the surface chemistry and characteristics of the wheat seeds exposed to plasma were investigated. Treatments of 30–60 s significantly enhanced the germination rate and showed positive effects on seedling growth. ACP resulted in changes of seed surface and chemical characteristics including water uptake and contact angle values. Changes in seed pH and total titratable acidity, as well as nitrites, nitrates, and malondialdehyde concentrations were also recorded

The potential of atmospheric air cold plasma for control of bacterial contaminants relevant to cereal grain production

The aim of this work was to investigate the efficacy of dielectric barrier discharge atmospheric cold plasma (DBD ACP) against bacteria associated with grains quality and safety. ACP inactivation efficacy was tested against biofilms formed by different strains of E. coli, Bacillus and Lactobacillus in grain model media and against B. atrophaeus endospores either in grain media or attached on abiotic surfaces. Effects were dependent on bacterial strain, media composition and mode of ACP exposure. ACP treatment for 5min reduced E. coli spp., B. subtilis and Lactobacillus spp. biofilms by \u3e3 log10, whereas insignificant reductions were achieved for B. atrophaeus. ACP treatment of 5–20min reduced B. atrophaeus spores in liquids by \u3e5 log10. Treatment for 30min reduced spores on hydrophobic surface by \u3e6 log10, whereas maximum of 4.4 log reductions were achieved with spores attached to hydrophilic surface. Microscopy demonstrated that ACP caused significant damage to spores. In package ACP treatment has potential to inactivate grain contaminants in the form of biofilms, as well as spores and vegetative cells. Industrial relevance This study demonstrates that ACP technology is a promising tool for effective bio-decontamination which offers a wide range of possible applications including inactivation of microorganisms on cereal grains. However, due to the nature of the microbial contamination of grains and complex grain structures it may be necessary to optimise the potential for surface inactivation at several stages of grain processing and storage to enhance ACP efficacy against bacterial endospores