Enhanced antimicrobial efficacy and energy efficiency of low irradiance 405-nm light for bacterial decontamination.

Due to its increased safety over ultraviolet light, there is interest in the development of antimicrobial violet-blue light technologies for infection control applications. To ensure compatibility with exposed materials and tissue, the light irradiances and dose regimes used must be suitable for the target application. This study investigates the antimicrobial dose responses and germicidal efficiency of 405 nm violet-blue light when applied at a range of irradiance levels, for inactivation of surface-seeded and suspended bacteria. Bacteria were seeded onto agar surfaces (101-108 CFUplate-1) or suspended in PBS (103-109 CFUmL-1) and exposed to increasing doses of 405-nm light (≤ 288 Jcm-2) using various irradiances (0.5-150 mWcm-2), with susceptibility at equivalent light doses compared. Bacterial reductions ≥ 96% were demonstrated in all cases for lower irradiance (≤ 5 mWcm-2) exposures. Comparisons indicated, on a per unit dose basis, that significantly lower doses were required for significant reductions of all species when exposed at lower irradiances: 3-30 Jcm-2/0.5 mWcm-2 compared to 9-75 Jcm-2/50 mWcm-2 for low cell density (102 CFUplate-1) surface exposures and 22.5 Jcm-2/5 mWcm-2 compared to 67.5 Jcm-2/150 mWcm-2 for low density (103 CFUmL-1) liquid exposures (P ≤ 0.05). Similar patterns were observed at higher densities, excluding S. aureus exposed at 109 CFUmL-1, suggesting bacterial density at predictable levels has minimal influence on decontamination efficacy. This study provides fundamental evidence of the greater energy efficacy of 405-nm light for inactivation of clinically-significant pathogens when lower irradiances are employed, further supporting its relevance for practical decontamination applications.

Evaluation of visible light and natural photosensitizers against Staphylococcus epidermidis and Staphylococcus saprophyticus planktonic cells and biofilm.

Antimicrobial photoinactivation (API) has shown some promise in potentially treating different nosocomial bacterial infections, however, its application on staphylococci, especially other than Staphylococcus aureus or methicillin-resistant S. aureus (MRSA) species is still limited. Although S. aureus is a well-known and important nosocomial pathogen, several other species of the genus, particularly coagulase-negative Staphylococcus (CNS) species such as Staphylococcus epidermidis and Staphylococcus saprophyticus, can also cause healthcare-associated infections and foodborne intoxications. CNS are often involved in resilient biofilm formation on medical devices and can cause infections in patients with compromised immune systems or those undergoing invasive procedures. In this study, the effects of chlorophyllin and riboflavin-mediated API on S. epidermidis and S. saprophyticus planktonic cells and biofilm are demonstrated for the first time. Based on the residual growth determination and metabolic reduction ability changes, higher inactivating efficiency of chlorophyllin-mediated API was determined against the planktonic cells of both tested species of bacteria and against S. saprophyticus biofilm. Some insights on whether aqueous solutions of riboflavin and chlorophyllin, when illuminated with optimal exciting wavelength (440 nm and 402 nm, respectively) generate O2-•, are also provided in this work.

Riboflavin- and chlorophyllin-based antimicrobial photoinactivation of Brevundimonas sp. ESA1 biofilms.

Some Brevundimonas spp. are globally emerging opportunistic pathogens that can be dangerous to individuals with underlying medical conditions and for those who are immunocompromised. Gram-negative Brevundimonas spp. can form resilient sessile biofilms and are found not only in different confined terrestrial settings (e.g., hospitals) but are also frequently detected in spacecraft which is inhabited by astronauts that can have altered immunity. Therefore, Brevundimonas spp. pose a serious health hazard in different environments, especially in its biofilm form. Conventional antimicrobials applied to disrupt, inactivate, or prevent biofilm formation have limited efficiency and applicability in different closed-loop systems. Therefore, new, effective, and safe biofilm control technologies are in high demand. The present work aimed to investigate antimicrobial photoinactivation (API) of Brevundimonas sp. ESA1 monocultural biofilms mediated by non-toxic, natural photosensitizers such as riboflavin (RF) and chlorophyllin (Chl) with an emphasis of this technology as an example to be safely used in closed-loop systems such as spacecraft. The present study showed that Chl-based API had a bactericidal effect on Brevundimonas sp. ESA1 biofilms at twice the lower irradiation doses than was needed when applying RF-based API. Long-term API based on RF and Chl using 450 nm low irradiance plate has also been studied in this work as a more practically applicable API method. The ability of Brevundimonas sp. ESA1 biofilms to reduce alamarBlue™ and regrowth analysis have revealed that after the applied photoinactivation, bacteria can enter a viable but non-culturable state with no ability to resuscitate in some cases.

Charge effect of water-soluble porphyrin derivatives as a prototype to fight infections caused by Acinetobacter baumannii by aPDT approaches.

In the last decade, Acinetobacter baumannii has emerged as a pathogen associated with infections in intensive care units worldwide, especially due to its ability to resist an extensive list of antibiotics. In this context, porphyrins have emerged as an important strategy in photodynamic therapy, since they are a group of tetrapyrrolic compounds with important photochemical and photobiological activities. In this study, the antimicrobial photodynamic activity of meso-tetra(4-N-methyl-pyridyl)porphyrin (H2TMePyP+) and meso-tetra(4-sulfonatophenyl)porphyrin (H2TPPS‒) was evaluated against A. baumannii by minimum inhibitory concentration (MIC), anti-biofilm activity, and the interaction with antibiotics after exposure to white-light LED irradiation. The cationic derivative H2TMePyP+ was more potent (MIC = 0.61 µM) than H2TPPS‒, with anti-biofilm activity and increased the antimicrobial activity of ciprofloxacin and amikacin. Given these findings, the tetra-cationic porphyrins can be assumed as prototypes to optimize and develop new agents by promoting oxidative stress and inducing free radical production.

Aerosol-based antimicrobial photoinactivation in the lungs: an action spectrum study.

Chronic lung infections are among the most diffused human infections, being often associated with multidrug-resistant bacteria. In this framework, the European project "Light4Lungs" aims at synthesizing and testing an inhalable light source to control lung infections by antimicrobial photoinactivation (aPDI), addressing endogenous photosensitizers only (porphyrins) in the representative case of S. aureus and P. aeruginosa. In the search for the best emission characteristics for the aerosolized light source, this work defines and calculates the photo-killing action spectrum for lung aPDI in the exemplary case of cystic fibrosis. This was obtained by applying a semi-theoretical modelling with Monte Carlo simulations, according to previously published methodology related to stomach infections and applied to the infected trachea, bronchi, bronchioles and alveoli. In each of these regions, the two low and high oxygen concentration cases were considered to account for the variability of in vivo conditions, together with the presence of endogenous porphyrins and other relevant absorbers/diffusers inside the illuminated biofilm/mucous layer. Furthermore, an a priori method to obtain the "best illumination wavelengths" was defined, starting from maximizing porphyrin and light absorption at any depth. The obtained action spectrum is peaked at 394 nm and mostly follows porphyrin extinction coefficient behavior. This is confirmed by the results from the best illumination wavelengths, which reinforces the robustness of our approach. These results can offer important indications for the synthesis of the aerosolized light source and definition of its most effective emission spectrum, suggesting a flexible platform to be considered in further applications.

Unsymmetrical cationic porphyrin-cyclodextrin bioconjugates for photoinactivation of Escherichia coli.

Photodynamic inactivation (PDI) of microorganisms has been used for the treatment of bacterial infection. PDI is based on the combination of three non-toxic elements: a photosensitizer (PS), light and molecular oxygen, which lead to the formation of reactive oxygen species (ROS) that cause lethal oxidative damage into the target pathogenic bacteria. For that, clinical approved tetrapyrrolic macrocycles, with particular emphasis on photoactive porphyrin (Por) dyes, have been used as PS in PDI for different biomedical applications. Two novel unsymmetrical free-base thiopyridyl Pors conjugated with α- or γ-CD units (Pors 2 and 3) were prepared and the corresponding cationic ones (Pors 2a and 3a) were assessed as water-soluble photosensitizer (PS) agents by photophysical, photochemical and E. coli photobiological studies. The presence of the CD unit and the positive charges on the Por periphery (2a and 3a) enhance their solubility in aqueous media. The photoactivity of the two cationic Pors 2a and 3a ensures their potential as PDI drugs against Gram-negative bacteria model, a bioluminescent E. coli, which the best PDI efficiency was determined for Por 3a that achieved the highest bacterial reduction of 4.0 log10 (ANOVA, p < 0.0001), reaching the detection limit of the method after 15 min.

A Comparative Study on Two Cationic Porphycenes: Photophysical and Antimicrobial Photoinactivation Evaluation.

Over the last decades, the number of pathogenic multi-resistant microorganisms has grown dramatically, which has stimulated the search for novel strategies to combat antimicrobial resistance. Antimicrobial photodynamic therapy (aPDT) is one of the promising alternatives to conventional treatments based on antibiotics. Here, we present a comparative study of two aryl tricationic porphycenes where photoinactivation efficiency against model pathogenic microorganisms is correlated to the photophysical behavior of the porphycene derivatives. Moreover, the extent of photosensitizer cell binding to bacteria has been assessed by flow cytometry in experiments with, or without, removing the unbound porphycene from the incubation medium. Results show that the peripheral substituent change do not significantly affect the overall behavior for both tricationic compounds neither in terms of photokilling efficiency, nor in terms of binding.