## 4 Discussion
Previous microbiological findings in an experimental peri-implantitis model mimicking natural disease progression supported the notion that large variation in microbial profiles makes interpretation of a correlation between disease progression and specific microbes difficult [35]. In the current study, Gram-negative _Escherichia coli_ was used to investigate bacterial adhesion and effects of laser decontamination on titanium implant surfaces. Although the biofilm compositions on contaminated implant surfaces in peri-implant diseases are complex and dominated by Gram-negativeanaerobes [6; 35; 36], several in vitro studies utilized single species bacterial biofilms for the investigation of implant surface contamination [18; 19; 22]. The adhesion rate of Gram-negative _Escherichia coli_ on the investigated implant surfaces prior to Er:YAG irradiation ranged from 1% to 3% (Table 2). Although these percentages seem low, the number of adhering bacteria reached 10\({}^{6}\) CFU/mL, which was equivalent to that described in the literature [27; 28]. Studying the effect of surface roughness of titanium implants on the adhesion and removal of bacterial contamination, [PERSON] et al. found no significant difference in _Escherichia coli_ bacterial adhesion among surfaces of varying roughness, except in 50 \(\upmu\)m and 250 \(\upmu\)m Al\({}_{2}\)O\({}_{3}\) sandblasting groups at 12 h of culture [27]. The cleaning modalities investigated did not induce significant surface alterations. However, _Escherichia coli_ adhesion was significantly reduced by air-powder abrasion and Er:YAG laser debridement in comparison to plastic curettage. The authors concluded that laser debridement could be a useful cleaning method for peri-implantitis therapy. Another in vitro study investigated bacterial adhesion on smooth and rough titanium disc surfaces after treatment with four different instruments using single species _Streptococcus sanguinis_ (American Type Culture Collection 10556) bacterial suspensions [28]. The authors found that smooth titanium surfaces were altered significantly by metal curettage, yet there were no significant rough surface alterations before and after treatment with Er:YAG laser irradiation, plastic curettage, metal curettage, or air-powder abrasion. The findings of the present study were in line with the literature, showing that significant effectiveness of sterilization by Er:YAG laser irradiation could be achieved on _Escherichia coli_-contaminated rough titanium implant surfaces. In the present study, clinical situations were simulated by real implant fixture placement in a 3D-printed standard human adult dento-alveolar model attached to a simulation manikin. However, effects of saliva and blood could not be simulated under the present experimental conditions. Considering the limitations of this model, it was difficult to determine whether implant diameter or access limitation caused a non-significant lower sterilization rate in wider implants at the mandibular posterior region. We concluded that effective sterilization rates could be achieved with Er:YAG laser irradiation with a proper contact tip and parameter settings irrespective of the depth and configuration of peri-implant intra-bony defects. Further studies are necessary to investigate similar effects on biofilms or multiple-species bacterial cultures. The use of integrated CAD and 3D-printing techniques to construct an in vitro laser standard treatment platform with different intra-bony defect configurations is demonstrated herein. Furthermore, these models were reverse-engineered from real peri-implantitis cases to visualize and simulate treatment procedures prior to surgery. All authors have made substantial contributions to conception and design of the study. [PERSON] and [PERSON] have been involved in data collection and data analysis. [PERSON] and [PERSON] have been involved in data interpretation, drafting the manuscript and revising it critically and have given final approval of the version to be published.