In a recent article published in Applied and Environmental Microbiology, researchers evaluated how effectively antimicrobial blue light (aBL) inactivated dried cells and biofilms of Listeria monocytogenes (Lm), a lethal foodborne pathogen.

Study: Inactivation of dried cells and biofilms of Listeria monocytogenes by exposure to blue light at different wavelengths and the influence of surface materials. Image Credit: Golubovy/Shutterstock.com

Background

Lm thrives in the environment and surfaces in the food industry, including floors, drains, and some equipment, even when cleaned and disinfected regularly. Also, Lm adheres to surfaces to form biofilms resistant to common disinfectants, another significant concern for the food industry.

Earlier ultraviolet light, particularly UV-C, was widely used as a disinfectant in the food industry. However, given its detrimental effects on the eyes and skin, there is an urgent need to develop new alternative antimicrobial disinfectants for processing and packing food production environments.

Most previous studies have reported the effectiveness of aBL against Lm in aqueous systems. However, data are scarce on its effects against Lm-dried cells and biofilms deposited on inert surfaces, primarily stainless steel (SS).

About the study

In the current investigation, the research team examined the antimicrobial properties of aBL (antimicrobial blue light) emitted by three distinct types of commercially available LED array lamps. These lamps emitted light at varying wavelengths, specifically 405, 420, and 460 nm. The target of this assessment was Listeria monocytogenes (Lm) existing in the form of dried cells on inert surfaces, such as stainless steel (SS), and colonizing these surfaces in the form of biofilms.

The researchers also explored how the application of an exogenous photosensitizer (Ps), for instance, gallic acid, interacted with the effects of aBL on Lm present on these surfaces. Another aspect under investigation was the specific emission doses (ED) and exposure durations required for different aBL wavelengths to combat Lm biofilms. In pursuit of this, the researchers exposed the surfaces to aBL at selected time intervals of four, eight, and 16 hours, and quantified the EDs in terms of Joules (J) per square centimeter (cm²).

To comprehensively examine the impact of material composition on Lm viability and the potential for aBL in surface decontamination, the researchers employed a mixture of five L. monocytogenes strains. They evaluated various inert surface materials, including stainless steel (SS), high-density polyethylene (HDPE), polyvinyl chloride (PVC), polystyrene (PS), and silicone rubber (SR).

The research team introduced 100 µL of the mixed strain cocktail onto the SS coupons and subsequently dried them in a biosafety cabinet for up to two hours to obtain dry cells. This process occurred in two series: one included the application of a photosensitizer, while the other did not. Additionally, biofilms were subjected to photosensitization tests.

Following each exposure to aBL, the team documented the average bacterial counts in terms of log colony-forming units (CFU) per square centimeter (cm²). Moreover, they utilized confocal laser scanning microscopy (CLSM) to analyze changes in the biofilm structure within the initial four hours of aBL exposure, across all three wavelengths.

Each experiment was replicated at least twice in triplicate, and statistical analysis, such as one-way analysis of variance (ANOVA), was employed to identify variations in bacterial populations and biofilm density. In addition to this, hierarchical cluster analysis was conducted to assess the similarity of Lm cells, and principal component analysis (PCA) was used to discern the impact of different factors on Lm cells.

Results

All aBL wavelengths consistently reduced the viability of Lm-dried cells and biofilms on SS in a dose-dependent fashion, with maximum effect attained at a constant dose. 

Irradiation at higher wavelengths of 420 and 460 nm did not result in >2 log reductions in Lm cells exposed to EDs of ≤1,000 J/cm² within the same time. On the contrary, irradiation at 405 nm aBL achieved the highest reductions in viability of dried Lm cells, including in biofilms.

It attained Lm cell viability reductions of 3log CFU/cm² on SS at an ED of 2,672 J/cm² within 16 hours. The application of Ps enhanced the efficacy of aBL by an additional 1 log reduction at 668 J/cm² ED against dried Lm cells, especially on hard-to-reach surfaces. 

Exposure to 405 nm aBL also caused the highest, i.e., 4 log CFU/cm² reduction in Lm cells viability with the lowest dose (=668 J/cm²) on PS, followed by HDPE and SR, suggesting the role of material composition on Lm inactivation. In all materials, PVC caused the minimum Lm cell viability reduction. 

Previous studies have suggested that aBL triggered the generation of reactive oxygen species (ROS) from porphyrins and flavins (photosensitizing molecules), which damaged the cell integrity of biofilms, causing Lm cell death.

Consistent with previous findings, in this study, exposing Lm in biofilms for four hours reduced biofilm biomass regardless of the aBL wavelength applied.

It damaged biofilm cell membrane integrity, which resulted in the loss of Lm cells' physiological functions, including membrane potential, permeability barrier, and efflux pump activity. 

Conclusion

This study provides considerable evidence of the ability of aBL technology to control surface contamination by foodborne pathogens, including Lm, with and without the use of Ps.

On SS, aBL irradiation at all three wavelengths, 405, 420, and 460 nm, reduced the counts of Lm dried cells gradually at all three EDs (668, 1336, 2,672 J/cm²) and exposure time durations (4, 8, and 16h) tested.

The application of Ps resulted in additional reductions of 1 log CFU/cm² at 668 J/cm², but the observed effects were inconsistent. 

The observed reductions in viable Lm cell counts on the biofilms at 420nm and 460nm were 1.9 and 1.6 log CFU/cm², respectively, at the highest EDs of 960 and 800J/cm².

Surface material composition affected reductions in Lm viability; however, increasing the ED to 4,008 J/cm² at 405 nm for 24 h only improved its efficacy on SS and PVC. Together, these results suggest that aBL is a potent intervention to mitigate surface contamination by Lm in food industries.

Written by

Neha Mathur

Neha is a digital marketing professional based in Gurugram, India. She has a Master’s degree from the University of Rajasthan with a specialization in Biotechnology in 2008. She has experience in pre-clinical research as part of her research project in The Department of Toxicology at the prestigious Central Drug Research Institute (CDRI), Lucknow, India. She also holds a certification in C++ programming.