## 4 Discussion  
The presented results do not support the hypothesis that heated and not burned tobacco increases the levels of subgingival periodontitis in IQOS users without periodontal disease, when compared to non-smokers. Furthermore, the second hypothesis was that classic cigarette smokers have a higher prevalence and higher abundance of periodontitis when compared to IQOS users. This hypothesis can be partially accepted. _P. intermedia_ and _P. gingivalis_ appear to be specific to smokers with periodontal disease, but the prevalence of these two pathogens was higher in smokers regardless of whether tobacco is burned or heated. This research demonstrated that exposure to tobacco, regardless of whether it is burned (as in conventional cigarettes) or heated (as in THS products), has a modest effect on the composition of subgingival periodontitisen biofilms. Periodontal disease has a more pronounced effect on the abundance and diversity of subgingival dental microbiome than tobacco exposure. Our research study focused on nine main periodontal pathogen bacteria, and the analyzed results are mostly in line with previously conducted studies, with some new and surprising findings [43]. The present research demonstrated that subgingival dental plaque in classic cigarette smokers had the highest prevalence of _T. forystthia_ and _T. denticola_, while _P. micra_ is the least frequent in this group. Plaque in IQOS users abounds with _P. intermedia_ and _E. nodatum_, followed by _C. rectus_ and _P. gingivalis_, while _A. actinomycetemmmitans_ and _T. denticola_ were the least-present periodontopathogens. The most frequent periodontitis in non-smokers were _P. micra_ and _A. actinomycetemmmitans_, while _P. intermedia, T. forystthia_, and _E. nodatum_ were rarely found in this group. _E. nodatum_ is an orange complex bacterium that, according to the literature, is found in severe and advanced stages of periodontitis and is more frequent in smokers than in non-smokers [10]. Our investigation confirmed previous studies in this regard. We found that the numbers of _E. nodatum_ were much higher in subjects with periodontitis and in IQOS consumers with periodontitis. According to the literature, _E. nodatum_ is also more frequent in smokers [10]. Our study results indicated that _E. nodatum_ was the most numerous in IQOS users, when compared to classic smokers. \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Bacteria** & **Periodontitis** & **No Periodontitis** & _p *_ & **V** \\ \hline _Prevoltella intermediia_ & 27/30 (90\%) & 16/29 (55\%) & 0.003 & 0.392 \\ _Porplyomonas gingvalis_ & 29/30 (97\%) & 20/29 (69\%) & 0.005 & 0.369 \\ _Treponema deticola_ & 27/30 (90\%) & 22/29 (76\%) & 0.148 & 0.188 \\ _Campylobacter rectus_ & 20/30 (67\%) & 18/29 (62\%) & 0.712 & 0.048 \\ _Tannerella forystthia_ & 30/30 (100\%) & 23/29 (79\%) & 0.001 & 0.342 \\ _Eubacterium nodatum_ & 28/30 (93\%) & 16/29 (55\%) & 0.011 & 0.438 \\ _Parcinomas micra_ & 1/30 (3\%) & 2/29 (7\%) & 0.533 & 0.081 \\ _Fusobacterium nucleatum_ & 30/30 (100\%) & 29/29 (100\%) & - & - \\ _Aggregathacter_ & 5/30 (17\%) & 5/29 (17\%) & 0.958 & 0.008 \\ _actinomycetemmmitans_ & & & & \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Prevalence of perio-pathogenic bacteria in subjects with and without periodontitis. Therefore, it appears that in the case of _E. nodatum_, IQOS aerosol affects its abundance more than cigarette smoke. This unexpected result suggests that IQOS users are at higher risk of developing severe forms of periodontal disease. The keystone perio-pathogens _P. gingivalis_ and _T. forysthia_ were found in higher amounts in the subgingival plaque of patients suffering from periodontitis when compared to those without periodontitis. _T. denticola_ was present in similar numbers in all groups (i.e., smokers and non-smokers, with or without periodontal disease) and, so, we cannot strictly connect it to any group. However, its prevalence was highest in smokers with periodontitis.