Efficacy of disinfectant and bacteriophage mixture against planktonic and biofilm state of Listeria monocytogenes to control in the food industry.

Fresh produce and animal-based products contaminated with Listeria monocytogenes have been the main cause of listeriosis outbreaks for many years. The present investigation explored the potential of combination treatment of disinfectants with a bacteriophage cocktail to control L. monocytogenes contamination in the food industry. A mixture of 1 minimal inhibitory concentration (MIC) of disinfectants (sodium hypochlorite [NaOCl], hydrogen peroxide [H2O2], and lactic acid [LA]) and multiplicity of infection (MOI) 100 of phage cocktail was applied to both planktonic cells in vitro and already-formed biofilm cells on food contact materials (FCMs; polyethylene, polypropylene, and stainless steel) and foods (celery and chicken meat). All the combinations significantly lowered the population, biofilm-forming ability, and the expression of flaA, motB, hlyA, prfA, actA, and sigB genes of L. monocytogenes. Additionally, in the antibiofilm test, approximately 4 log CFU/cm2 was eradicated by 6 h treatment on FCMs, and 3 log CFU/g was eradicated within 3 days on celery. However, <2 log CFU/g was eradicated in chicken meat, and regrowth of L. monocytogenes was observed on foods after 5 days. The biofilm eradication efficacy of the combination treatment was proven through visualization using scanning electron microscopy (SEM) and confocal microscopy. In the SEM images, the unusual behavior of L. monocytogenes invading from the surface to the inside was observed after treating celery with NaOCl+P or H2O2 + P. These results suggested that combination of disinfectants (NaOCl, H2O2, and LA) with Listeria-specific phage cocktail can be employed in the food industry as a novel antimicrobial and antibiofilm approach, and further research of L. monocytogenes behavior after disinfection is needed.

Control of Listeria monocytogenes in food industry by a combination treatment of natural aromatic compound with Listeria-specific bacteriophage cocktail.

Most Listeria monocytogenes found in the food industry are listeriosis-causing pathogens and possess the ability to form biofilms on food and food contact materials (FCMs). This study aims to evaluate the efficacy of the combination treatment of natural aromatic compounds (thymol, eugenol, carvacrol, and citral) with a Listeria-specific phage cocktail in mitigating the threat posed by L. monocytogenes in the food industry. In vitro combination treatment of 1 minimal inhibitory concentration (MIC) of natural aromatic compound with phage cocktail at multiplicity of infection (MOI) 100 reduced more than 4 log CFU/mL of L. monocytogenes planktonic cells and inhibited biofilm formation. In addition, the expression of virulence-related genes (flaA, motB, hlyA, prfA, and actA) and the stress response (sigB) gene were significantly downregulated. The combination of natural aromatic compound with phage cocktail reduced the biofilm cell population on contaminated celery by more than 2 log CFU/g and by more than 2 log CFU/cm2 on already-formed biofilm on FCMs, but it was less effective on chicken meat, with an approximate reduction of only 1 log CFU/g. The antibiofilm activity toward preformed L. monocytogenes biofilms was also observed using field-emission scanning electron microscopy (FESEM) and confocal laser scanning microscopy (CLSM). COMSTAT analysis of the structural change of biofilms revealed that major biofilm structure parameters (biovolume, thickness, diffusion distance, and microcolonies at substratum) were reduced after treatment. Our findings suggest that the combination of natural aromatic compounds with a phage cocktail has enormous potential as an antimicrobial and antibiofilm agent for controlling L. monocytogenes in the food industry.

Survival strategies of Listeria monocytogenes to environmental hostile stress: biofilm formation and stress responses.

Listeria monocytogenes is a critical foodborne pathogen that causes listeriosis and threatens public health. This pathogenic microorganism forms a transmission cycle in nature, food industry, and humans, expanding the areas of contamination among them and influencing food safety. L. monocytogenes forms biofilms to protect itself and promotes survival through stress responses to the various stresses (e.g., temperature, pH, and antimicrobial agents) that may be inflicted during food processing. Biofilms and mechanisms of resistance to hostile external or general stresses allow L. monocytogenes to survive despite a variety of efforts to ensure food safety. The current review article focuses on biofilm formation, resistance mechanisms through biofilms, and external specific or general stress responses of L. monocytogenes to help understand the unexpected survival rates of this bacterium; it also proposes the use of obstacle technology to effectively cope with it in the food industry.

Biofilm eradication ability of phage cocktail against Listeria monocytogenes biofilms formed on food contact materials and effect on virulence-related genes and biofilm structure.

Listeria monocytogenes is a foodborne pathogen that can form biofilms in food processing facilities even under unfavorable growth environment. This study aimed to evaluate the biofilm eradication ability of Listeria-specific bacteriophage (phage) cocktail (LMPC01+02+03) against L. monocytogenes young (1 day) and mature (3 days) biofilms formed on food contact materials (FCMs: polyethylene, polypropylene, and stainless steel) at 4, 15, and 30 °C. In addition, virulence-related genes and biofilm structure parameters of the phage-treated biofilms were investigated. The biofilm eradication ability of the phage cocktail was evaluated on 96 well and MBEC plate, and the results revealed that a multiplicity-of-infection (MOI) 100 of the phage cocktail exhibited the ability of eradicate biofilms. Using MOI 100, the phage cocktail treatment on the biofilms formed on FCMs for 8 h reduced over 2 log CFU/cm2 of the young biofilms, and approximately 1 log CFU/cm2 of the mature biofilms. In addition, the phage treatment against the biofilms resulted in a significant up-regulation of two genes (flaA and motB), and up/down-regulation or no changes in three genes (hlyA, prfA, and actA). Confocal and scanning electron microscopy images revealed the loss of the biofilm matrix after the phage treatment, and quantitative analysis revealed a reduction in the structural parameters of the biofilm, except the microcolonies at the substratum level, which increased. These results suggested that MOI 100 of the phage cocktail (LMPC01+02+03) was an effective tool for eradicating L. monocytogenes biofilms formed on FCMs, and it is essential to develop a countermeasure to eradicate the biofilm remaining after phage treatment.

Effect of sublethal concentrations of bactericidal antibiotics on mutation frequency and stress response of Listeria monocytogenes.

The purpose of this study was to investigate sublethal concentrations (SLC) of bactericidal antibiotics (ampicillin, gentamicin, kanamycin, and vancomycin) on the mutation frequency and stress response of antibiotic-induced-mutated (AIM) Listeria monocytogenes. Three L. monocytogenes strains (reference, clinical, and food isolate strains) were used in this study. SLC of bactericidal antibiotics significantly increased the mutation frequency in L. monocytogenes. It was found that AIM L. monocytogenes had a superior biofilm-forming ability than nontreated L. monocytogenes. This result correlated with the amounts of EPS produced (polysaccharide and protein) in the early stage of biofilm formation. AIM L. monocytogenes showed strong viability under food-associated stress (thermal, osmotic, and acidic) compared to nontreated L. monocytogenes. In addition, expression levels of motility (flaA) and virulence genes (hlyA, actA, and prfA) of AIM L. monocytogenes were significantly downregulated in the reference strain but significantly upregulated or similar to the expression levels in the clinical and food isolate strains compared to nontreated L. monocytogenes. Based on our results, SLC of bactericidal antibiotics increased the mutation frequency in L. monocytogenes, facilitated the adaptation of the bacterium to food-associated stress, and led to an increase in its pathogenicity.

Combination treatment of peroxyacetic acid or lactic acid with UV-C to control Salmonella Enteritidis biofilms on food contact surface and chicken skin.

The risk of salmonellosis is expected to increase with the rise in the consumption of poultry meat. The aim of this study was to investigate the combination treatment of peroxyacetic acid (PAA) or lactic acid (LA) with UV-C against Salmonella Enteritidis biofilms formed on food contact surface (stainless steel [SS], silicone rubber [SR], and ultra-high molecular weight polyethylene [UHMWPE]) and chicken skin. The biofilm on food contact surface and chicken skin was significantly decreased (P < 0.05) by combination treatment of PAA or LA with UV-C. Combination treatment of PAA (50-500 µg/mL) with UV-C (5 and 10 min) reduced 3.10-6.41 log CFU/cm2 and LA (0.5-2.0%) with UV-C (5 and 10 min) reduced 3.35-6.41 log CFU/cm2 of S. Enteritidis biofilms on food contact surface. Salmonella Enteritidis biofilms on chicken skin was reduced around 2 log CFU/g with minor quality changes in color and texture by combination treatment of PAA (500 µg/mL) or LA (2.0%) with UV-C (10 min). Additional reduction occurred on SS and UHMWPE by PAA or LA with UV-C, while only LA with UV-C caused additional reduction on chicken skin. Also, it was visualized that the biofilm on food contact surface and chicken skin was removed through field emission scanning electron microscopy (FESEM) and death of cells constituting the biofilm was confirmed through confocal laser scanning microscopy (CLSM). These results indicating that the combination treatment of PAA or LA with UV-C could be used for S. Enteritidis biofilm control strategy in poultry industry.

Isolation and characterization of Salmonella spp. from food and food contact surfaces in a chicken processing factory.

The presence of Salmonella serotypes is a major safety concern of the food industry and poultry farmers. This study aimed to isolate and identify Salmonella spp. from a chicken processing facility by PCR and pulsed-field gel electrophoresis (PFGE). In addition, the biofilm-forming abilities of the isolated bacteria on stainless steel, silicone rubber, plastic, and chicken skin were also investigated. PCR was used for the confirmation of Salmonella serotypes, and then gene similarity within the same serotype was analyzed by PFGE. As a result, 26 S. Enteritidis isolates were detected at a high rate from both food contact surfaces and chicken products during processing. All of them were 100% genetically identical to the same bacteria. The results indicated that the virulence factors and effective biofilm-forming ability of S. Enteritidis isolates could affect human health and economic revenue. It was also suggested that the visual observation of food and food contact surfaces could be a great concern in the future. The continuous monitoring of S. Enteritidis molecular and biofilm characteristics is needed to increase food safety.

COVID-19 pandemic crisis and food safety: Implications and inactivation strategies.

BACKGROUND: The COVID-19 pandemic that emerged in 2019 has imposed huge consequences, including economic losses and threats to human health, which are still affecting many aspects throughout the world. SCOPE AND APPROACH: This review provides an overview of SARS-CoV-2 infection, the cause of COVID-19, and explores its impact on the food supply system and food safety. This review examines the potential risk of transmission through food and environmental surfaces before discussing an effective inactivation strategy to control the COVID-19 pandemic in the aspect of food safety. This article also suggests effective food safety management post-COVID-19. KEY FINDINGS AND CONCLUSIONS: Respiratory viruses including SARS-CoV-2 are responsible for huge impacts on the global economy and human health. Although food and water are not currently considered priority transmission routes of SARS-CoV-2, infection through contaminated food and environmental surfaces where the virus can persist for several days cannot be ignored, particularly when the surrounding environment is unhygienic. This approach could help determine the exact transmission route of SARS-CoV-2 and prepare for the post-COVID-19 era in the food safety sector.

Efficacy of flavourzyme against Salmonella Typhimurium, Escherichia coli, and Pseudomonas aeruginosa biofilms on food-contact surfaces.

Food contamination is a major public health concern, with Salmonella Typhimurium, Escherichia coli, and Pseudomonas aeruginosa being the prominent causal agents. They often produce resistant shields in food through biofilm formation and are difficult to remove from food-contact surfaces using conventional cleaning agents. In the current study, we investigated the efficacy of flavourzyme, an industrial peptidase, in biofilm removal from ultra-high molecular weight polyethylene (UHMWPE) and rubber surfaces and compared the corresponding efficacies with those of the commonly used DNase I. We noticed a significant reduction of young (24-h-old) and mature (72-h-old) biofilms on both surfaces after treatment with flavourzyme. The overall reduction potentiality of flavourzyme was higher than that of DNase I. The flavourzyme-mediated removal of biofilms appears to be caused by the gradual disruption of amide (NH) and polysaccharide (C-O-C) stretching bands of the extracellular polymeric substances (EPS) released by the microbes. EPS elimination and the cell-friendly behavior of flavourzyme were further confirmed by field emission scanning electron microscopy. Based on these findings, we suggest that flavourzyme can reduce microbial EPS formation, thus possibly controlling microbial food contamination. This finding reveals a new opportunity for the development of a novel method for controlling foodborne illness as well as food spoilage.

Quantitative microbial risk assessment for Clostridium perfringens foodborne illness following consumption of kimchi in South Korea.

The purpose of this study was to conduct a risk assessment for Clostridium perfringens foodborne illness via kimchi consumption in South Korea. Prevalence of C. perfringens in kimchi, kimchi consumption amount and frequency, and distribution conditions (time and temperature) from manufacture to the home were determined. C. perfringens initial contamination level was estimated using Beta distribution [Beta (6, 79)]. Potential C. perfringens cell counts during distribution were predicted using the Weibull model (primary models, R 2 = 0.923-0.953) and a polynomial model [(δ = 1/(0.2385 + (- 0.0307 × Temp) + (0.0011 × Temp2)), R 2 = 0.719]. Average daily consumption data was assessed using Gamma distribution [1.0444, 91.767, RiskShift (0.16895), RiskTruncate (0, 1078)]. The mean risk of C. perfringens-associated foodborne illness following kimchi consumption was found to be 1.21 × 10-17. These results suggest that the risk of C. perfringens foodborne illness from kimchi consumption, under current conditions, can be considered to be very low in S. Korea.

Effect of UV-C irradiation on inactivation of Aspergillus flavus and Aspergillus parasiticus and quality parameters of roasted coffee bean (Coffea arabica L.).

This study investigated the antifungal effect of ultraviolet-C (UV-C) against Aspergillus flavus and Aspergillus parasiticus on roasted coffee beans. Also, any changes in the quality of the roasted coffee beans were measured after UV-C irradiation. As UV-C irradiation time increased (0-2 h), the number of surviving A. flavus and A. parasiticus spores significantly (P < .05) decreased. The reduction values of A. flavus in round part, crack part, and whole roasted coffee beans were 2.16, 0.71, and 1.58 log10 CFU g-1, respectively, and the reduction values of A. parasiticus in round part, crack part, and whole roasted coffee beans were 1.03, 0.37, and 0.72 log10 CFU g-1, respectively, after 2 h of UV-C irradiation. Field emission scanning electron microscopy showed that the morphology of A. flavus and A. parasiticus spores included expanded wrinkles that were deformed by UV-C irradiation. The Hunter colours were significantly reduced (P < .05). There was no significant change (P > .05) in moisture content, but the pH was significantly decreased (P < .05). Most of the sensory parameters did not change, but there was a significant difference (P < .05) in flavour. Based on this study, 2 h of UV-C irradiation was effective in reducing 90% of A. flavus, but it was not effective against A. parasiticus present on roasted coffee beans. Also, Hunter colour, pH, and sensory parameters (flavour) were changed by UV-C irradiation.

Effects of gamma ray, electron beam, and X-ray on the reduction of Aspergillus flavus on red pepper powder (Capsicum annuum L.) and gochujang (red pepper paste).

Aspergillus flavus is the potential pathogenic mold in red pepper powder (Capsicum annuum L.) and gochujang (red pepper paste), which can produce mycotoxins. This study investigated the effects of gamma ray, e-beam, and X-ray irradiation on the reduction of A. flavus on red pepper powder and gochujang and physicochemical and sensory quality changes. Gamma ray and e-beam at 3.5 kGy reduced A. flavus effectively (>4 log), without deteriorating the physicochemical quality. Same dose of X-ray did not cause any deterioration of the physicochemical quality. However, reduction effect of A. flavus in red pepper powder and gochujang by 3.5 kGy X-ray was under 2 log. Further, sensory quality analysis showed no significant difference in color, appearance, texture, and overall acceptability after three irradiations. However, flavor changes of red pepper powder and gochujang after three irradiations were mentioned by panelists. In this study, gamma ray and e-beam irradiation were effective in eliminating A. flavus present in red pepper powder and gochujang, but X-ray irradiation was not effective. The results indicate gamma ray and e-beam are effective in controlling microorganisms present in powdery or paste foods, but the X-ray was not effective.

Genetic Relationship, Virulence Factors, Drug Resistance Profile and Biofilm Formation Ability of Vibrio parahaemolyticus Isolated From Mussel.

The objective of this study was to investigate the virulence factors, genetic relationship, antibiotic resistance profile and the biofilm formation ability of Vibrio parahaemolyticus isolates on shrimp and mussel surfaces at 30°C. In this study, eight (n = 8) V. parahaemolyticus isolated from mussel were examined. We used the polymerase chain reaction (PCR) to examine the distribution of different genes, and Repetitive Extragenic Palindromic-PCR (REP-PCR) to compare the genetic relationship. Disk diffusion technique was used to assess antibiotic and multiple-antibiotic resistance. The biofilm formation assay, and field emission scanning electron microscopy (FE-SEM) were used to evaluate biofilm formation ability. Transmission Electron Microscope (TEM) was used to observe the morphological structure of bacterial cell. Our results indicated that the biofilm-associated genes, 16S rRNA, toxR, and tdh, were present in all the tested V. parahaemolyticus isolates (n = 8). Approximately, 62.5% (5 isolates among 8 isolates) isolates showed strong multiple-antibiotic resistance index with an average value of 0.56. All isolates (n = 8) showed strong genetic relationship and significant biofilm formation ability on shrimp and mussel surfaces. This study demonstrated that the presence of virulence factors, high multiple antibiotic resistance index (MARI) values, and effective biofilm formation ability of V. parahaemolyticus isolates could be a great threat to human health and economic values in future. It was also suggested that a high resistance rate to antibiotic could be ineffective for treating V. parahaemolyticus infections. The continuous monitoring of V. parahaemolyticus antibiotic, molecular and biofilm characteristics is needed to increase seafood safety.

Effects of microwaves on the reduction of Aspergillus flavus and Aspergillus parasiticus on brown rice (Oryza sativa L.) and barley (Hordeum vulgare L.).

Aspergillus flavus and Aspergillus parasiticus are primary pathogen moulds on brown rice and barley. This study investigated the effects of microwave irradiation (MWI) (2450 MHz, 700 W, 10-50 s) on inactivation of A. flavus and A. parasiticus on brown rice and barley and the quality of these samples. The counts of both strains were significantly (p < 0.05) reduced by the stepwise increase in MWI treatment time. The log reductions of A. flavus on brown rice and barley were 0.05 and 0.04 after 10 s; 1.06 and 1.05 after 20 s; 1.59 and 1.52 after 30 s; and 3.04 and 2.78 after 40 s. The log reductions of A. parasiticus on brown rice and barley were 0.06 and 0.10 after 10 s; 1.20 and 1.00 after 20 s; 2.04 and 1.61 after 30 s; and 2.89 and 2.90 after 40 s. Moreover, neither strain survived after 50 s of MWI. The Hunter colour 'L' gradually increased with increasing MWI treatment time. However, there were no significant differences in the 'L' of brown rice after 10-40 s of MWI treatment and of barley after 10-30 s of MWI treatment. The Hunter colour 'a' and 'b' gradually increased with increasing microwave time. No significant change was observed in the moisture content of either cereal treated with 10-20 s of MWI. The differences in the sensory quality (colour, appearance, flavour, texture and overall acceptability) after 0-30 s of MWI were not significant. However, values for colour, appearance, texture and overall acceptability were significantly reduced when treated with 40-50 s of MWI. Therefore, with 20 s of MWI at 2450 MHz, 700 W could be effective for > 90% reduction of mould without causing deleterious changes to the colour, moisture content and sensory qualities of these cereals.