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Dual-light photodynamic therapy administered daily provides a sustained antibacterial effect on biofilm and prevents Streptococcus mutans adaptation

Sakari Nikinmaa, Heikki Alapulli, Petri Auvinen, Martti Vaara, Juha Rantala, Esko Kankuri, Timo Sorsa, Jukka Meurman, Tommi Pätilä

Published: May 6, 2020 https://doi.org/10.1371/journal.pone.0232775


Antibacterial photodynamic therapy (aPDT) and antibacterial blue light (aBL) are emerging treatment methods auxiliary to mechanical debridement for periodontitis. APDT provided with near-infrared (NIR) light in conjunction with an indocyanine green (ICG) photosensitizer has shown efficacy in several dental in-office-treatment protocols. In this study, we tested Streptococcus mutans biofilm sensitivity to either aPDT, aBL or their combination dual-light aPDT (simultaneous aPDT and aBL) exposure. Biofilm was cultured by pipetting diluted Streptococcus mutans suspension with growth medium on the bottom of well plates. Either aPDT (810 nm) or aBL (405 nm) or a dual-light aPDT (simultaneous 810 nm aPDT and 405 nm aBL) was applied with an ICG photosensitizer in cases of aPDT or dual-light, while keeping the total given radiant exposure constant at 100 J/cm2. Single-dose light exposures were given after one-day or four-day biofilm incubations. Also, a model of daily treatment was provided by repeating the same light dose daily on four-day and fourteen-day biofilm incubations. Finally, the antibacterial action of the dual-light aPDT with different energy ratios of 810 nm and 405 nm of light were examined on the single-day and four-day biofilm protocols. At the end of each experiment the bacterial viability was assessed by colony-forming unit method. Separate samples were prepared for confocal 3D biofilm imaging. On a one-day biofilm, the dual-light aPDT was significantly more efficient than aBL or aPDT, although all modalities were bactericidal. On a four-day biofilm, a single exposure of aPDT or dual-light aPDT was more efficient than aBL, resulting in a four logarithmic scale reduction in bacterial counts. Surprisingly, when the same amount of aPDT was repeated daily on a four-day or a fourteen-day biofilm, bacterial viability improved significantly. A similar improvement in bacterial viability was observed after repetitive aBL application. This viability improvement was eliminated when dual-light aPDT was applied. By changing the 405 nm to 810 nm radiant exposure ratio in dual-light aPDT, the increase in aBL improved the antibacterial action when the biofilm was older. In conclusion, when aPDT is administered repeatedly to Smutans biofilm, a single wavelength-based aBL or aPDT leads to a significant biofilm adaptation and increased Smutans viability. The combined use of aBL light in synchrony with aPDT arrests the adaptation and provides significantly improved and sustained antibacterial efficacy.



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PDT in oral hygiene- a split mouth randomized trial

Nikinmaa S, Meurman J, Moilanen N, Sorsa T, Rantala J, Alapulli H, Kankuri E, Kotiranta A, Auvinen P, Pätilä T. 

The effects of indocyanine green-mediated photodynamic therapy on the development of plaque, plaque bacteriological ecology, and early periodontitis markers - A randomised study 


Antimicrobial photodynamic therapy (aPDT) has been introduced as an adjunct method for dental hygiene. Although antibacterial and antiplaque effects resulting from aPDT have already been demonstrated in the literature, effects on bacterial flora diversity or early gingivitis biomarkers have not previously been established. 


Fifteen healthy adults were assigned to the study. Upper premolars (4. and 5.) were examined on both sides of the maxilla. After meticulous scaling and root planing, the maxillary dental arch was left without any mechanical cleaning for four days. Randomisation of the treatment side of the upper dental arch was performed, and following the initial sample collection, the mouth was rinsed with indocyanine green (ICG), and 100J/cm2 of 810 nm light was subsequently applied for eight minutes. The treatment was repeated daily for four days. ICG localisation after the mouth rinse procedure was measured after each treatment by near-infrared imaging. Plaque area, 16S rRNA bacteriological identification, and gingival crevicular fluid (GCF) matrix metalloproteinase 8 (MMP-8) samples were measured. Fluorescent imaging showed ICG adherence to dental plaque, enabling localised treatment at the target site.


Antimicrobial photodynamic therapy resulted in a significant reduction of plaque formation. An analysis of the 16S rRNA sequencing found reduction in the Streptococcus, AcinetobacterialCapnocytophagal, and Rothia bacteria species and a gain in Neisseria and Hemophilus bacteria on the aPDT-treated side. The gain in the latter group of bacteria superseded the relative loss of the former in the plaque, while alpha diversity remained stable. A reduction of the total amount of MMP-8 in the GCF was seen on the treated side, suggesting inhibition of early gingivitis.


In conclusion, ICG-based aPDT is effective and reduces the amount of known oral pathogens, with compensated bacterial growth in species associated with good oral health, but without a change in overall bacterial diversity. The treatment can be applied specifically to dental plaque, and the anti-inflammatory effect may prevent the development of early gingivitis.