br However since a diversity observed
However, since a-diversity (observed index) was increased as a whole in the OSCC patients compared to the healthy controls, changes in the oxygen exposure state, the influence of exudates from necrotic tissue, and changes in the immune state can occur as the tumor increases. It is also possible that the microbiota changes.
For the N factor indicating lymph node metastasis, progressing a-diversity (observed index) was not observed (Fig. 5B). Regarding the presence or absence of lymph node metastasis, a relationship with tumor size could not be clarified. The participants of this study showed lymph node metastasis at T4 and also at T1, due to which the relevance could not be confirmed. The nature of the tumor surface tissue influences the changes in microbiota. Thus, if caused by microbiota changes, the result is not contradictory in that, the T factor was involved in the microbiota change, but not the presence or absence of N factor.
In this study, Rothia revealed a negative correlation showing a decrease with the progression of the T factor (Fig. 5C), and from a previous study, we know that it is remarkably low in OSCC patients [9,10,23]. Moreover, although this bacterium is consid-ered to be phylogenetically related to Actinomyces, its pathoge-nicity itself is considered low. Thus, it is possible that Rothia is common among healthy people.
In this study, we examined factors influencing cancer incidence using logistic regression analysis (Table 3). So far, sex, smoking, and drinking habits have been reported as cancer risk factors [3,33,34], but in this study we obtained similar results only with regard to sex. An odds ratio of 10.85 also indicated a high risk.
The genera Rothia, Alloprevotella, Capnocytophaga, Fusobacte-rium, Haemophilus, and family Peptostreptococcus, which showed a significant difference in abundance among OSCC patients compared to the healthy controls, had no influence on cancer incidence, however the Chao 1 index, indicating a-diversity, showed a significant difference regarding cancer incidence. Although the odds ratio was 1.006, it is possible that the Trizma maleate index indicates an influencing factor for cancer incidence.
4.3. Relationship between oral microbiota and disease
It is suggested that there is also an age difference in the occur-rence of oral cancer .
Recent studies have reported that gut microbiota and age are related and that the microbiota changes with age [36,37]. In addi-tion, a relationship between intestinal microbiota and the general condition of the immune status has also been reported .
There have been several reports showing differences between the oral microbiota in healthy people and periodontitis patients [39,40]. Changes in the oral microbial flora have also been re-ported using saliva samples for comparison between aged care facility residents and the healthy group among the elderly in Japan. A significant difference was observed in a-diversity be-tween these two groups . From the results obtained in this study, differences in diversity and the existence of specific minor bacterial species might influence the incidence of OSCC and microbiota composition.
Although their influence is not strong, some bacteria related to dental caries and periodontal disease also suggest that the cancer risk can be reduced by maintaining good oral hygiene.
Despite the microbiota distribution evaluation at this time, it has been reported that the ratio as well as the number of bacteria influence disease onset . There is a possibility of identifying new causative bacteria for the onset by comparing the number of bacteria.
In addition, bacteria as well as fungi are involved in the for-mation of oral microbiota. It is known that Candida and other fungi are detected at high proportions in the oral cavity if the immune status is reduced as in the elderly and HIV patients [43,44]. It is suggested that these fungal species are also associated with oral cancer and other cancers, but the specifics of how they are involved are not clear . In recent years, analysis of microbiota using NGS has been carried out widely, but research on the characteristics of the fungal flora and their relation to the microbiota in OSCC pa-tients will be necessary in the future.