Utilization of Piper betle L. Extract for Inactivating Foodborne Bacterial Biofilms on Pitted and Smooth Stainless Steel Surfaces.

Biofilms are a significant concern in the food industry. The utilization of plant-derived compounds to inactivate biofilms on food contact surfaces has not been widely reported. Also, the increasing negative perception of consumers against synthetic sanitizers has encouraged the hunt for natural compounds as alternatives. Therefore, in this study we evaluated the antimicrobial activities of ethanol extracts, acetone extracts, and essential oils (EOs) of seven culinary herbs against Salmonella enterica serotype Typhimurium and Listeria innocua using the broth microdilution assay. Among all tested extracts and EOs, the ethanol extract of Piper betle L. exhibited the most efficient antimicrobial activities. To evaluate the biofilm inactivation effect, S. Typhimurium and L. innocua biofilms on pitted and smooth stainless steel (SS) coupons were exposed to P. betle ethanol extract (12.5 mg/ml), sodium hypochlorite (NaClO; 200 ppm), hydrogen peroxide (HP; 1100 ppm), and benzalkonium chloride (BKC; 400 ppm) for 15 min. Results showed that, for the untreated controls, higher sessile cell counts were observed on pitted SS versus smooth SS coupons. Overall, biofilm inactivation efficacies of the tested sanitizers followed the trend of P. betle extract ≥ BKC > NaClO > HP. The surface condition of SS did not affect the biofilm inactivation effect of each tested sanitizer. The contact angle results revealed P. betle ethanol extract could increase the surface wettability of SS coupons. This research suggests P. betle extract might be utilized as an alternative sanitizer in food processing facilities.

Removal of Salmonella Typhimurium Biofilm from Food Contact Surfaces Using Quercus infectoria Gall Extract in Combination with a Surfactant.

Quercus infectoria (nutgall) has been reported to possess antimicrobial activities against a wide range of pathogens. Nevertheless, the biofilm removal effect of nutgall extract has not been widely investigated. In this study, we therefore evaluated the effect of nutgall extract in combination with cetrimonium bromide (CTAB) against preformed biofilm of Salmonella Typhimurium on polypropylene (PP) and stainless steel (SS) coupons in comparison with other sanitizers. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of nutgall extract and surfactants (CTAB and sodium dodecyl sulfate; SDS) were assessed. CTAB showed a more efficient antimicrobial activity than SDS and was selected to use in combination with nutgall extract for removing biofilm. To determine the biofilm removal efficacy, the PP and SS coupons were individually submerged in 2x MBC of nutgall extract (256 mg/ml) + 2x MBC of CTAB (2.5 mg/ml), nutgall extract alone (256 mg/ml), CTAB alone (2.5 mg/ml), distilled water, and 100 ppm sodium hypochlorite for 5, 15, and 30 min. The remaining sessile cells in biofilm were determined. Overall, the greatest biofilm removal efficacy was observed with nutgall extract + CTAB; the biofilm removal efficacy of sanitizers tended to increase with the exposure time. The SEM analysis demonstrated that S. Typhimurium biofilm on PP and SS coupons after exposure to nutgall extract + CTAB for 30 min displayed morphological alterations with wrinkles. This study suggests nutgall extract + CTAB may be an alternative to commonly used sanitizers to remove biofilm from food contact surfaces in the food industry and household.

Inhibition of Escherichia coli O157:H7 and Salmonella enterica Isolates on Spinach Leaf Surfaces Using Eugenol-Loaded Surfactant Micelles.

Spinach and other leafy green vegetables have been linked to foodborne disease outbreaks of Escherichia coli O157:H7 and Salmonella enterica around the globe. In this study, the antimicrobial activities of surfactant micelles formed from the anionic surfactant sodium dodecyl sulfate (SDS), SDS micelle-loaded eugenol (1.0% eugenol), 1.0% free eugenol, 200 ppm free chlorine, and sterile water were tested against the human pathogens E. coli O157:H7 and Salmonella Saintpaul, and naturally occurring microorganisms, on spinach leaf surfaces during storage at 5 °C over 10 days. Spinach samples were immersed in antimicrobial treatment solution for 2.0 min at 25 °C, after which treatment solutions were drained off and samples were either subjected to analysis or prepared for refrigerated storage. Whereas empty SDS micelles produced moderate reductions in counts of both pathogens (2.1-3.2 log10 CFU/cm2), free and micelle-entrapped eugenol treatments reduced pathogens by >5.0 log10 CFU/cm2 to below the limit of detection (<0.5 log10 CFU/cm2). Micelle-loaded eugenol produced the greatest numerical reductions in naturally contaminating aerobic bacteria, Enterobacteriaceae, and fungi, though these reductions did not differ statistically from reductions achieved by un-encapsulated eugenol and 200 ppm chlorine. Micelles-loaded eugenol could be used as a novel antimicrobial technology to decontaminate fresh spinach from microbial pathogens.

Application of Surfactant Micelle-Entrapped Eugenol for Prevention of Growth of the Shiga Toxin-Producing Escherichia coli in Ground Beef.

Beef safety may be compromised by O157 and non-O157 Shiga toxin-producing Escherichia coli (STEC) contamination. The capacity of surfactant micelles loaded with the plant-derived antimicrobial eugenol to reduce STEC on beef trimmings that were later ground and refrigerated for five days at 5 ± 1 °C was tested to determine their utility for beef safety protection. STEC-inoculated trimmings were treated with free eugenol, micelle-encapsulated eugenol, 2% lactic acid (55 °C), sterile distilled water (25 °C), or left untreated (control). Following treatment, trimmings were coarse-ground and stored aerobically at 5 ± 1 °C. Ground beef was then sampled for STEC immediately post-grinding, and again at three and five days of storage. STEC minimum inhibitory concentrations (MICs) in liquid medium for free eugenol and 1% sodium dodecyl sulfate (SDS)-loaded micelles were 0.5% and 0.125%, respectively. STEC numbers on beef trimmings treated by sterile water (6.5 log10 CFU/g), free eugenol (6.5 log10 CFU/g), micelle-loaded eugenol (6.4 log10 CFU/g), and lactic acid (6.4 log10 CFU/g) did not differ compared to untreated controls (6.6 log10 CFU/g) (p = 0.982). Conversely, STEC were significantly reduced by refrigerated storage (0.2 and 0.3 log10 CFU/g at three and five days of storage, respectively) (p = 0.014). Antimicrobial treatments did not significantly decontaminate ground beef, indicating their low utility for beef safety protection.

Inhibition of Bacterial Pathogens in Medium and on Spinach Leaf Surfaces using Plant-Derived Antimicrobials Loaded in Surfactant Micelles.

UNLABELLED: Encapsulation of hydrophobic plant essential oil components (EOC) into surfactant micelles can assist the decontamination of fresh produce surfaces from bacterial pathogens during postharvest washing. Loading of eugenol and carvacrol into surfactant micelles of polysorbate 20 (Tween 20), Surfynol® 485W, sodium dodecyl sulfate (SDS), and CytoGuard® LA 20 (CG20) was determined by identification of the EOC/surfactant-specific maximum additive concentration (MAC). Rheological behavior of dilute EOC-containing micelles was then tested to determine micelle tolerance to shearing. Antimicrobial efficacy of EOC micelles against Escherichia coli O157:H7 and Salmonella enterica serotype Saintpaul was first evaluated by the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Pathogen-inoculated spinach was treated with eugenol-containing micelles applied via spraying or immersion methods. SDS micelles produced the highest MACs for EOCs, while Tween 20 loaded the lowest amount of EOCs. Micelles demonstrated Newtonian behavior in response to shearing. SDS and CG20-derived micelles containing EOCs produced the lowest MICs and MBCs for pathogens. E. coli O157:H7 and S. Saintpaul were reduced on spinach surfaces by application of eugenol micelles, though no differences in numbers of surviving pathogens were observed when methods of antimicrobial micelle application (spraying, immersion) was compared (P ≥ 0.05). Data suggest eugenol in SDS and CG20 micelles may be useful for produce surface decontamination from bacterial pathogens during postharvest washing. PRACTICAL APPLICATION: Antimicrobial essential oil component (EOC)-containing micelles assist the delivery of natural food antimicrobials to food surfaces, including fresh produce, for decontamination of microbial foodborne pathogens. Antimicrobial EOC-loaded micelles were able to inhibit the enteric pathogens Escherichia coli O157:H7 and Salmonella Saintpaul in liquid medium and on spinach surfaces. However, pathogen reduction generally was not impacted by the method of micelle application (spraying, immersion washing) on spinach surfaces.