## 4 Discussion  
Here, we show that BIC-40 system performs better than Brush in reducing _P. aeruginosa_ biofilm and microbial re-growth onto titanium disks. Dental implants are widely used in different clinical situations; osseointegration ability and long-lasting integrity bring about successful results [29]. Vice versa, microbial localization on the implant affects the surrounding area, eventually leading to osseointegration loss [3; 4]. As described in [30], microbial biofilm may happen, despite titanium disks' surface sharpness. This outline needs a targeted clinical action for patient benefit. Besides antibiotics or other therapies [31], decontamination techniques of dental implant surfaces deserve special attention as local cleaning systems. Such treatments seem to be less effective when applied to rough surfaces rather than to smooth surfaces. Furthermore, depending upon cleaning procedure used, undesirable effects may occur on implant surface morphology [32; 33; 34]. By a recently established model assessing in real time microbial biofilm formation onto medical devices [27], here we show that BLI-_Pseudomonas_ has the ability to adhere onto the titanium disks, thus rapidly forming a consistent biofilm on their surfaces within 24 h. Moreover, we provide the first evidence on the different efficacy of the two decontamination treatments, since Brush and BIC-40 are capable of impairing biofilm up to 76% and 99.9%, respectively. In particular, as evaluated by RLU analysis and subsequent conversion in CFU/disk, we show that microbial load is reduced of about 1 log  
Figure 2: Kinetic evaluation of _P. aeruginosa_ biofilm re-growth onto titanium disks, following decontamination by Ni-Ti Brush and AirPol BIC-40 systems. Microbial load was kinetically assessed on treated and untreated disks, by bioluminescence assay. The results are expressed as microbial load ratio between treated and untreated disks. The ratio = 1 (dotted line) indicates superimposable microbial load in treated and untreated groups. following Brush treatment, while more than 3 logs decrease occurs upon BIC-40 treatment. Furthermore, kinetic assessment of BLI signal demonstrates that microbial re-growth onto BIC-40-treated disks is delayed and remains mostly below the control values (about 1 log). In the Brush-treated disks, microbial re-growth occurs, rapidly reaching levels similar to those observed in the untreated control disks. It should be noted that _Pseudomonas_ produces one of the most complex and difficult to treat biofilms [35] even on dental implants [36]. Thus, we assume that the successful decontamination obtained by BIC-40, in our model, would likely be as effective when applied against other clinically relevant biofilms, including those produced during peri-mucositis/peri-implantitis. As recently published [25], a mechanical treatment for decontamination of titanium disks, such as the use of Brush, deeply affects the device surface producing deep grooves on it. In contrast, lack of damage occurs in BIC-40 treated titanium surface(s). Taken together, our previous [25] and present data favor the conclusion that the use of BIC-40, avoiding formation of niches onto device surface, would in turn prevent/minimize microbial adhesion, survival, and persistence. Moreover, we favor the hypothesis that the good performance of BIC-40 can also be related to the fact that sodium bicarbonate particles, remaining onto BIC-40-treated titanium disks [25], likely render the device surface hostile to subsequent microbial re-growth. In any case, using a BLI-based system that allows kinetic monitoring of microbial load onto medical devices [27], we have been able to show a different efficacy of the two decontamination procedures not only in impairing preformed biofilm but also in limiting subsequent re-growth onto titanium disks. Finally, it is worth noting that the present data have been obtained using titanium disks with rough surfaces, as representative of the worst condition clinically possible.