Effects of the Oral Health Promotion Program on oral health and oral microbiota changes in diabetic elderly individuals: a quasi-experimental study
As population aging continues to become a major demographic trend worldwide, diabetes has become one of the most crucial public health problems. Globally, there are 422 million diabetic adults and the prevalence of diabetes is still increasing at an alarming rate . Especially in China, the number of diabetic patients is expected to be 174 million by 2045 , which is considered to lead to a significant socioeconomic burden . Therefore, as one of the key aspects of treatment, it is emergent to improve oral health for diabetic individuals efficiently. Recent remarkable studies have shown unequivocal evidence of a bidirectional link between type 2 diabetes and periodontitis, where diabetes can worsen oral infections and vice versa . In particular, older diabetic individuals with poor glycemic control have a greater tendency to suffer severe periodontitis, dental caries, salivary dysfunction, delayed wound healing, halitosis  and provides a hostile microenvironment for altering the oral microbiota , Conversely, the aggravation of oral diseases in turn leads to the deteriorating of diabetes , all of which bring greater distress physically and mentally. In general, the present popular oral health management is sufficient for reversing mild oral conditions, improving the oral microbiota, and controlling glycated haemoglobin (HbA1c) in diabetic patients in the short term ; however, more common and comprehensive management methods for those patients are still neglected and lacking . Crucially, the present care strategy conducted by the medical staff mainly focused on the effect of dental visit, the oral health attitudes, and the knowledge on oral health of diabetic patients , but the oral health knowledge and self-care cognition of the patients themselves are generally insufficient and rarely to be considered. As a result, correct practices can always hardly be executed precisely, which is also an important factor that gives rise to the bidirectional whammy of age-related oral changes and diabetes . This study aims to propose a scientific evidence-based Oral Health Promotion Program (OHPP) for strengthening knowledge and ability of self-motivation for proactive oral health management among elderly diabetic patients at home or in communities . In addition to the regular indicators including oral hygiene and oral health-related quality of life that represent the oral condition and general health of diabetic elderly individuals, the changes of oral microbiota and the guiding role of nurses were also considered as crucial in enhancing awareness and initiative of the patients toward better oral health management . Therefore, a comprehensive intervention consisting of oral cleaning guidance and evaluation, family education and patient self-directed learning guidance, and oral health assessment were demonstrated in this study. 

Design and setting One of the groups with 26 elderly diabetic individuals was assigned to be the intervention group using the OHPP, and the other group consisting of 26 elderly diabetic individuals was as the control group receiving routine oral care. The two groups were assigned and treated in two regionally representative community health centers located in the east‒west direction of Chengdu, China, from August 2022 to January 2023. Participation The inclusion criteria for elderly individuals were as follows: (i) ≥ 60 years old; (ii) diagnosed with type 2 diabetes; (iii) having more than 20 natural teeth; (iv) being able to use smartphone; (v) can complete brushing independently; and (vi) can provide written informed consent for participation. The exclusion criteria were as follows: (i) diagnosed with hypoglycemia or cardiovascular disease within the last 3 months; (ii) suffering from periodontitis stage III and IV or oral tumors; (iii) with physical dysfunction as inability to clean oral independently; (iv) with cognitive impairment or mental disorders; (v) have systemically used antibiotics or immunosuppressants within the last 6 months; (vi) have high-dosely used probiotics for the last 6 months; (vii) have used topical antibiotics within 7 days; and (viii) with restricted mouth opening. Individuals who met the inclusion criteria were recruited through poster exhibitions or family doctor recommendations, and eligibilities were verified by reviewing electronic health records and patient interviews, where the online preliminary screening was conducted by telephone, and the second screening was conducted via offline interviews. Sample size The sample size was evaluated by the abundance of species in the two groups from previous studies . A medium Cohen's effect size (d) of 0.8 was adopted for the sample size calculation by using the commercial software G*Power Version 3.1.9.7 , where the required sample size was 42 participants (21 in each group) to attain 80% power at a type 1 error rate (two-tailed) of 5%. After considering an attrition rate of 20%, the required sample size was finally adjusted to 56. Training of nurses For the maintenance of a high level of homogeneity and fidelity in the community, all the involved nurses received standardized training through lectures, videos and operation demonstration before the experiment, where the training content includes five parts as oral health assessment, oral cleaning behavior guidance, denture cleaning behavior guidance, oral cleaning effect evaluation and oral health guidance. Moreover, all the nurses were also required to pass the assessment by two teaching experts. Correspondingly, one theoretical teaching researcher worked closely with the operator and two data collectors on-site in the intervention group community during the study, whereas the control group had no operator with the others being the same. Implementation of the intervention group As depicted in Fig. , the quasi-experiment was conducted over a period of three months for the implementation of three main phases as cognization, intensification, and consolidation. Phase I. Cognition The stage of cognization was carried out with the main purpose of popularizing theoretical knowledge and mastering basic skills, and the intervention was carried out in the forms of slide presentations, videos and live operations, aiming to populate the basic knowledge of oral health and teach the patients the correct method of brushing teeth and flossing. Trained clinical nurse specialists majoring in oral health were arranged to deliver all detailed OHPP content for learning and practicing. In addition to be instructed to brush teeth with soft toothbrush and fluoride toothpaste, participants were also guided and required to use dental bow floss in the daily oral clean, especially after meals. Bass method of technique was taught to the participants, and the cleaning duration was required to be three to five minutes. Dental cleaning was required at least twice a day, especially after meals and before bedtime. For those participants with removable dentures, the cleaning method was recommended as using denture brush and mild soap, and leave cleaned denture soaked in cold water overnight. Phase II. Intensification The aim of this stage was to continuously enhance the awareness of oral cleaning of the participants. The participants were regularly reminded of the details of oral cleaning twice a week, and the managers of the intervention group pushed interesting short videos and popular science tweets on oral health to the participants through social media platforms to emphasize the importance of oral health behaviors. Phase III. Consolidation In this stage, participants were expected to build up the ability of proactive self-management of keeping oral hygiene. The research team delivered the OHPP via three modalities (lecture, practice, and online teaching), where the theoretical frameworks were precisely based on health promotion theory, and rigorously incorporated behavioral self-management approaches designed to help participants set goals, solve problems, make behavioral changes, and increase self-efficacy. Implementation of the control group In contrast, the control group received routine nursing based on science education on oral health knowledge and routine oral hygiene guidance only at the beginning of the study, such as the frequency of brushing teeth should be more than 2 times a day and last more than 3 min each time. Meanwhile, in contrast to the intervention group, the participants of the control group were only regularly reminded of brushing teeth once a week. Measurements The primary outcome was oral health-related indicators, including changes in oral health-related quality of life measured through a modified version of the Oral Health Impact Profile (OHIP-14) questionnaire that covers 7 social dimensions (functional limitation, physical pain, psychological discomfort, physical disability, psychological disability, social disability, and handicap) out of 14 items of oral health impact , the dental plaque index (PLI) assessed by Turesky’s modified method , the glycated haemoglobin (HbA1c) level by analyzing the venous blood, and the mastery of oral health knowledge using the rigorously self-made questionnaires containing 15 true or false questions that cover the correct brushing method, the choice of toothpaste, the usage of dental floss, and the cleaning of dentures. The secondary outcomes were the species richness (Ace index and Sobs index), diversity (Shannon index) and composition of oral microbiota. The presence of the microbiota in the saliva was detected by the 16S ribosome RNA (16SrRNA) assay. and the collection of saliva was carried out according to the previous literature . The main reagents, instruments and procedures employed in the experiments are presented in detail in the Supplementary Material. Statistical methods The database was established by two researchers who recorded the original data independently, and all the further statistical analyses were conducted by two-sided tests with statistical significance ( P < 0.05). For non-microbiota data, Stata 14.0 software was used. The measurements of the corresponding baseline data followed the independent samples t test or paired samples Wilcoxon test, where the data confirming a normal distribution were further expressed as the mean ± standard deviation (SD), and those unconfirmed were represented by median and interquartile intervals. The count of the baseline data was described by the frequency and proportion of occurrence. Target outcomes of oral health were compared between groups via Paired samples t tests or Wilcoxon tests at two different time points (T0: before the study; T1: 3 months after intervention onset) and were compared via general linear regression analysis between groups (see details of methods for handling confounders in the Supplemental Material). All statistical analyses of the microbiome were carried out by R 3.1.1 software. The Ace, Sobs and Shannon index were calculated via mothur  software ( http://www.mothur.org/wiki/Calculators ), and then paired samples Wilcoxon test was used for the analysis of within-group differences in alpha diversity(boot 1.3.18 and stats 3.3.1 packages). Beta-diversity was assessed by PCoA analysis based on Bray–Curtis dissimilarity for testing the similarity of microbial community structure between samples (PCoA statistical analysis and graphing) and the Adonis nonparametric test was used to analyse whether the differences in microbial community structure between sample groups were significant (vegan packages). Linear discriminant analysis effect size (LEfSe) ( http://huttenhower.sph.harvard.edu/LEfSe ) was used to identify bacterial taxa with significant differences in abundance from phylum to genus level among different groups (LDA > 2, p < 0.05) . 

One of the groups with 26 elderly diabetic individuals was assigned to be the intervention group using the OHPP, and the other group consisting of 26 elderly diabetic individuals was as the control group receiving routine oral care. The two groups were assigned and treated in two regionally representative community health centers located in the east‒west direction of Chengdu, China, from August 2022 to January 2023. 

The inclusion criteria for elderly individuals were as follows: (i) ≥ 60 years old; (ii) diagnosed with type 2 diabetes; (iii) having more than 20 natural teeth; (iv) being able to use smartphone; (v) can complete brushing independently; and (vi) can provide written informed consent for participation. The exclusion criteria were as follows: (i) diagnosed with hypoglycemia or cardiovascular disease within the last 3 months; (ii) suffering from periodontitis stage III and IV or oral tumors; (iii) with physical dysfunction as inability to clean oral independently; (iv) with cognitive impairment or mental disorders; (v) have systemically used antibiotics or immunosuppressants within the last 6 months; (vi) have high-dosely used probiotics for the last 6 months; (vii) have used topical antibiotics within 7 days; and (viii) with restricted mouth opening. Individuals who met the inclusion criteria were recruited through poster exhibitions or family doctor recommendations, and eligibilities were verified by reviewing electronic health records and patient interviews, where the online preliminary screening was conducted by telephone, and the second screening was conducted via offline interviews. 

The sample size was evaluated by the abundance of species in the two groups from previous studies . A medium Cohen's effect size (d) of 0.8 was adopted for the sample size calculation by using the commercial software G*Power Version 3.1.9.7 , where the required sample size was 42 participants (21 in each group) to attain 80% power at a type 1 error rate (two-tailed) of 5%. After considering an attrition rate of 20%, the required sample size was finally adjusted to 56. 

For the maintenance of a high level of homogeneity and fidelity in the community, all the involved nurses received standardized training through lectures, videos and operation demonstration before the experiment, where the training content includes five parts as oral health assessment, oral cleaning behavior guidance, denture cleaning behavior guidance, oral cleaning effect evaluation and oral health guidance. Moreover, all the nurses were also required to pass the assessment by two teaching experts. Correspondingly, one theoretical teaching researcher worked closely with the operator and two data collectors on-site in the intervention group community during the study, whereas the control group had no operator with the others being the same. 

As depicted in Fig. , the quasi-experiment was conducted over a period of three months for the implementation of three main phases as cognization, intensification, and consolidation. Phase I. Cognition The stage of cognization was carried out with the main purpose of popularizing theoretical knowledge and mastering basic skills, and the intervention was carried out in the forms of slide presentations, videos and live operations, aiming to populate the basic knowledge of oral health and teach the patients the correct method of brushing teeth and flossing. Trained clinical nurse specialists majoring in oral health were arranged to deliver all detailed OHPP content for learning and practicing. In addition to be instructed to brush teeth with soft toothbrush and fluoride toothpaste, participants were also guided and required to use dental bow floss in the daily oral clean, especially after meals. Bass method of technique was taught to the participants, and the cleaning duration was required to be three to five minutes. Dental cleaning was required at least twice a day, especially after meals and before bedtime. For those participants with removable dentures, the cleaning method was recommended as using denture brush and mild soap, and leave cleaned denture soaked in cold water overnight. Phase II. Intensification The aim of this stage was to continuously enhance the awareness of oral cleaning of the participants. The participants were regularly reminded of the details of oral cleaning twice a week, and the managers of the intervention group pushed interesting short videos and popular science tweets on oral health to the participants through social media platforms to emphasize the importance of oral health behaviors. Phase III. Consolidation In this stage, participants were expected to build up the ability of proactive self-management of keeping oral hygiene. The research team delivered the OHPP via three modalities (lecture, practice, and online teaching), where the theoretical frameworks were precisely based on health promotion theory, and rigorously incorporated behavioral self-management approaches designed to help participants set goals, solve problems, make behavioral changes, and increase self-efficacy. 

The stage of cognization was carried out with the main purpose of popularizing theoretical knowledge and mastering basic skills, and the intervention was carried out in the forms of slide presentations, videos and live operations, aiming to populate the basic knowledge of oral health and teach the patients the correct method of brushing teeth and flossing. Trained clinical nurse specialists majoring in oral health were arranged to deliver all detailed OHPP content for learning and practicing. In addition to be instructed to brush teeth with soft toothbrush and fluoride toothpaste, participants were also guided and required to use dental bow floss in the daily oral clean, especially after meals. Bass method of technique was taught to the participants, and the cleaning duration was required to be three to five minutes. Dental cleaning was required at least twice a day, especially after meals and before bedtime. For those participants with removable dentures, the cleaning method was recommended as using denture brush and mild soap, and leave cleaned denture soaked in cold water overnight. 

The aim of this stage was to continuously enhance the awareness of oral cleaning of the participants. The participants were regularly reminded of the details of oral cleaning twice a week, and the managers of the intervention group pushed interesting short videos and popular science tweets on oral health to the participants through social media platforms to emphasize the importance of oral health behaviors. 

In this stage, participants were expected to build up the ability of proactive self-management of keeping oral hygiene. The research team delivered the OHPP via three modalities (lecture, practice, and online teaching), where the theoretical frameworks were precisely based on health promotion theory, and rigorously incorporated behavioral self-management approaches designed to help participants set goals, solve problems, make behavioral changes, and increase self-efficacy. 

In contrast, the control group received routine nursing based on science education on oral health knowledge and routine oral hygiene guidance only at the beginning of the study, such as the frequency of brushing teeth should be more than 2 times a day and last more than 3 min each time. Meanwhile, in contrast to the intervention group, the participants of the control group were only regularly reminded of brushing teeth once a week. 

The primary outcome was oral health-related indicators, including changes in oral health-related quality of life measured through a modified version of the Oral Health Impact Profile (OHIP-14) questionnaire that covers 7 social dimensions (functional limitation, physical pain, psychological discomfort, physical disability, psychological disability, social disability, and handicap) out of 14 items of oral health impact , the dental plaque index (PLI) assessed by Turesky’s modified method , the glycated haemoglobin (HbA1c) level by analyzing the venous blood, and the mastery of oral health knowledge using the rigorously self-made questionnaires containing 15 true or false questions that cover the correct brushing method, the choice of toothpaste, the usage of dental floss, and the cleaning of dentures. The secondary outcomes were the species richness (Ace index and Sobs index), diversity (Shannon index) and composition of oral microbiota. The presence of the microbiota in the saliva was detected by the 16S ribosome RNA (16SrRNA) assay. and the collection of saliva was carried out according to the previous literature . The main reagents, instruments and procedures employed in the experiments are presented in detail in the Supplementary Material. 

The database was established by two researchers who recorded the original data independently, and all the further statistical analyses were conducted by two-sided tests with statistical significance ( P < 0.05). For non-microbiota data, Stata 14.0 software was used. The measurements of the corresponding baseline data followed the independent samples t test or paired samples Wilcoxon test, where the data confirming a normal distribution were further expressed as the mean ± standard deviation (SD), and those unconfirmed were represented by median and interquartile intervals. The count of the baseline data was described by the frequency and proportion of occurrence. Target outcomes of oral health were compared between groups via Paired samples t tests or Wilcoxon tests at two different time points (T0: before the study; T1: 3 months after intervention onset) and were compared via general linear regression analysis between groups (see details of methods for handling confounders in the Supplemental Material). All statistical analyses of the microbiome were carried out by R 3.1.1 software. The Ace, Sobs and Shannon index were calculated via mothur  software ( http://www.mothur.org/wiki/Calculators ), and then paired samples Wilcoxon test was used for the analysis of within-group differences in alpha diversity(boot 1.3.18 and stats 3.3.1 packages). Beta-diversity was assessed by PCoA analysis based on Bray–Curtis dissimilarity for testing the similarity of microbial community structure between samples (PCoA statistical analysis and graphing) and the Adonis nonparametric test was used to analyse whether the differences in microbial community structure between sample groups were significant (vegan packages). Linear discriminant analysis effect size (LEfSe) ( http://huttenhower.sph.harvard.edu/LEfSe ) was used to identify bacterial taxa with significant differences in abundance from phylum to genus level among different groups (LDA > 2, p < 0.05) . 

Participant characteristics Twenty-six elderly participants were enrolled in the intervention group and the control group. The mean age of the participants was 71.77 (SD 5.75) years, with 28 (53.8%) male, 48 (92.3%) married, and an average of 25.87 (SD 3.99) remaining with natural teeth. Importantly, all the participants in the two groups were well matched for demographics and factors, including age, number of remaining teeth, duration of diabetes, smoking status and alcohol consumption (Table ). Importantly, the median duration of diabetes among the participants was 11.4 (3.9,15.3) years. Changes in oral health-related indicators in the intervention group and control group As expected, the changes in the OHIP-14 scores and PLI in both the intervention and control groups were statistically significant (all p < 0.05). In the intervention group, the OHIP-14 score at T0 and T1 was 6.21 ± 5.7 and 0.41 ± 0.88, and the PLI score changed from 1.73 ± 0.13 to 1.33 ± 0.14, respectively. As a comparison, in the control group the OHIP-14 score merely changed from 4.8 ± 1.17 to 1.84 ± 0.62, and the PLI score even increased from 1.84 ± 0.14 to 2.75 ± 0.19, which indicated the increase of dental plaque. Meanwhile, the mastery of oral health knowledge promoted from 0.63 ± 0.34 to 0.94 ± 0.01 (with all p < 0.05). The detailed outcome comparison of the oral health status within the two groups are illustrated in Fig. . Ulteriorly, to illustrate the advantages of the OHPP in improving the oral health of diabetic patients, the comparison between the intervention group and the control group is further presented. Notably, due to that education level was moderately correlated with the household registration ( r = 0.5373) and gender ( r = 0.4094), only one of the variables was included in the model. Meanwhile, there was no statistically significant difference in the HbA1c level between two groups, therefore the HbA1c was not included in the comparison. Accordingly, after respectively considering the oral health outcomes on oral-related quality of life, presence of dental plaque and mastering oral health knowledge as the dependent variable of the linear regression model where the other influencing factors (age, gender, registered permanent residence, annual family income, duration of diabetes, remaining natural teeth, and educational attainment) were as independent variables, the results showed that the OHIP-14, PLI and mastery of oral health knowledge of the intervention were all superior to those of the control group (all p < 0.05) (see details in Supplementary tables S2-S5). Changes in the oral microbiota in the intervention group and control group In terms of α -diversity, the Ace index increased with statistical significance after the implementation of OHPP in the intervention group and after the implementation of routine oral care in the control group (Fig. A and B, respectively), whereas the Sobs and Shannon indices increased significantly only in the intervention group. In terms of β -diversity, the PCoA plots (Fig. C and D, respectively) showed significant differences in oral microbial community structure both before and after the experiment in both two groups (all p < 0.05). Moreover, as depicted in Fig. E, the results of LEfSe analyses of the intervention group showed a significant increase in g_Fusobacterium , g_Campylobacter and a significant decrease in g_Streptococcus , g_Rothia after 12 weeks compared to baseline, which is distinct from the control group where a significant increase in g_Neisseria , g_Veillonella and a significant decrease in g_Streptococcusa , g_Porphyromonas , g_Gemella after 12 weeks compared to baseline (Fig. F). It should be noted that the results of the presented LEfSe analyses show only the top 10 phyla and genera in terms of LDA values. 

Twenty-six elderly participants were enrolled in the intervention group and the control group. The mean age of the participants was 71.77 (SD 5.75) years, with 28 (53.8%) male, 48 (92.3%) married, and an average of 25.87 (SD 3.99) remaining with natural teeth. Importantly, all the participants in the two groups were well matched for demographics and factors, including age, number of remaining teeth, duration of diabetes, smoking status and alcohol consumption (Table ). Importantly, the median duration of diabetes among the participants was 11.4 (3.9,15.3) years. 

As expected, the changes in the OHIP-14 scores and PLI in both the intervention and control groups were statistically significant (all p < 0.05). In the intervention group, the OHIP-14 score at T0 and T1 was 6.21 ± 5.7 and 0.41 ± 0.88, and the PLI score changed from 1.73 ± 0.13 to 1.33 ± 0.14, respectively. As a comparison, in the control group the OHIP-14 score merely changed from 4.8 ± 1.17 to 1.84 ± 0.62, and the PLI score even increased from 1.84 ± 0.14 to 2.75 ± 0.19, which indicated the increase of dental plaque. Meanwhile, the mastery of oral health knowledge promoted from 0.63 ± 0.34 to 0.94 ± 0.01 (with all p < 0.05). The detailed outcome comparison of the oral health status within the two groups are illustrated in Fig. . Ulteriorly, to illustrate the advantages of the OHPP in improving the oral health of diabetic patients, the comparison between the intervention group and the control group is further presented. Notably, due to that education level was moderately correlated with the household registration ( r = 0.5373) and gender ( r = 0.4094), only one of the variables was included in the model. Meanwhile, there was no statistically significant difference in the HbA1c level between two groups, therefore the HbA1c was not included in the comparison. Accordingly, after respectively considering the oral health outcomes on oral-related quality of life, presence of dental plaque and mastering oral health knowledge as the dependent variable of the linear regression model where the other influencing factors (age, gender, registered permanent residence, annual family income, duration of diabetes, remaining natural teeth, and educational attainment) were as independent variables, the results showed that the OHIP-14, PLI and mastery of oral health knowledge of the intervention were all superior to those of the control group (all p < 0.05) (see details in Supplementary tables S2-S5). 

In terms of α -diversity, the Ace index increased with statistical significance after the implementation of OHPP in the intervention group and after the implementation of routine oral care in the control group (Fig. A and B, respectively), whereas the Sobs and Shannon indices increased significantly only in the intervention group. In terms of β -diversity, the PCoA plots (Fig. C and D, respectively) showed significant differences in oral microbial community structure both before and after the experiment in both two groups (all p < 0.05). Moreover, as depicted in Fig. E, the results of LEfSe analyses of the intervention group showed a significant increase in g_Fusobacterium , g_Campylobacter and a significant decrease in g_Streptococcus , g_Rothia after 12 weeks compared to baseline, which is distinct from the control group where a significant increase in g_Neisseria , g_Veillonella and a significant decrease in g_Streptococcusa , g_Porphyromonas , g_Gemella after 12 weeks compared to baseline (Fig. F). It should be noted that the results of the presented LEfSe analyses show only the top 10 phyla and genera in terms of LDA values. 

The OHPP showed significant advantages on the oral health and oral microbiota of elderly patients with diabetes in communities, and five parts, namely, oral health assessment, oral cleaning behavior guidance, denture cleaning behavior guidance, oral cleaning effect evaluation and oral health guidance, included in the OHPP can directly guide daily oral cleaning comprehensively. In contrast to routine oral care methods, OHPP interventions delivered by generalist nurses in primary care conspicuously improved the oral health and oral microbiota of elderly diabetic patients, which indicates that proactively guiding patients to strengthen the subjective initiative in keeping the oral hygiene can promote oral health, and is beneficial for the holistic health in a more long-term manner, which is always underestimated in practice. Meanwhile, it is worth emphasizing that toothbrushing is always one of the key behaviors of self-maintaining oral cleaning. By conducting OHPP interventions, including brushing teeth with fluoride toothpaste, using auxiliary cleaning tools such as dental floss, and ensuring the daily brushing times and duration, the periodontal state can be maintained and improved significantly, which is similar to the outcome of a cohort study followed for six years . Accordingly, for diabetic elderly individuals who always suffer earlier and stronger gum inflammatory responses, in addition to focusing on cleaning measures, mastering and utilizing auxiliary cleaning tools also play a key role in improving oral conditions. Another important point is that the PLI score was significantly improved. As the OHPP being applied, the amount of dental plaque covering teeth decreased nearly half on average, which was consistent with the findings of a Finnish study of home oral care interventions  It is also worth noting that the PLI score of the control group actually increased after the experiment, which is possibly related to the absence of using dental flossing and lack of self-awareness of oral hygiene among the elderly participants in communities. On the other hand, the fact that the plaque was not removed completely also indicated that the current oral cleaning method may still have blind spots , and patients need further professional oral health care to prevent gingivitis and periodontitis in both of the groups. Moreover, though both routine care and the OHPP can improve the stability of the oral microbial community, the OHPP still showed more significant advantages in improving oral microecology, which strongly affect the macro-outcome of oral health. Evidence with statistical significance showed that the abundance of Rothia in which Rothia_mucilaginosa is commonly considered being associated with intraoral leukoplakia and chronic lung disease  and Streptococcus where Streptococcus_sanguinis is regarded being related to caries endocarditis  declined apparently in the intervention group, which implied that the OHPP may be more helpful to improve the general health level of elderly patients. On the other hand, Porphyromonas_gingivalis , Porphyromonas_endodontalis , and Prevotella_intermedia that can cause alveolar bone resorption also declined significantly in the control group as expected, which indicated the improvement of routine oral care for periodontal pathogens and that the regular guidance of oral management for elderly diabetic patients should always be guaranteed . Moreover, because the oral microecology is always in a dynamic balance , factors such as brushing, gargling, the use of dentures and the presence of oral ulcers could affect the oral microbiota, and further research design should be done for exploring the effect of OHPP on oral microbiome systematically. An extended result can also be proposed that the OHPP can alleviate the impacts of oral problems on the quality of life of diabetic elderly individuals. Commonly, poor oral status in elderly people is considered to negatively impact quality of life , such as daily eating, speaking, sleeping and socializing, and the concomitant malnutrition caused by poor oral function can even result in asthenia and dementia . As a result, the OHPP maintenance of mastery of oral health knowledge for a longer time, regular staining of dental plaque and self-feedback of oral hygiene status, and dynamic strengthening of oral health behaviors are expected to positively effects on the mastery of oral health knowledge and continuous oral health behavior of the patients, which significantly improve their oral health-related quality of life. Finally, and unexpectedly, the experiment results showed that the OHPP and routine care do not benefit the control of diabetes as the measured HbA1c have no significant difference, where some previous research have considered that home oral hygiene maneuvers are valuable in controlling the levels of glycated hemoglobin and preventing the onset of complications typical of diabetes . The possible reason is that diabetes being a complicated health outcome may be affected by other conditions or comorbidities, such as medications and dietary habits, which were not controlled in this work. Meanwhile, the control of diabetes was only monitored by assessing HbA1c, which may not reveal the overall results on diabetes control. Consequently, the direct benefits of OHPP-induced improvements on the glycemic control of diabetic elderly individuals in the short term need further research. There are also several limitations in this study. First, the relatively small sample size and short-term follow-up assessment may make it inapparent on some essential outcomes, and more methodologically rigorous studies, such as randomized controlled trials, are needed in the future. Meanwhile, elderly individuals with severe oral diseases were not included, therefore findings presented are lacking generalizability to high-risk groups. Moreover, due to the particularity of diabetes, the management of the patients was inevitable to be intervened by the communities in parallel, and the effects of dietary habits and medication on outcomes could hardly be considered. In addition, possible selection bias may also affect intervention adherence; for example, the in-cooperation of the participants may have led to the distortion of the questionnaire feedback. Therefore, continuous and long-term follow-up studies with larger sample sizes in different populations are further required. 

Superior results on changing oral microbiota and improving oral health of the OHPP demonstrated that correct guidance and advice management can better enhance the subjective initiative and self-regulation of elderly patients in contrast to the regular process. Once stable oral salutary behavior is established, the holistic health of elderly patients is expected to improve under auxiliary nursing management. By considering objective oral microecology and subjective oral health cognition and function, the participants were multidimensionally, scientifically and clinically evaluated from a comprehensive perspective. Above all, this study with comprehensive results on the oral health of representative diabetic elderly individuals may provide an innovative perspective for the new nursing methods of the elderly in different living backgrounds, and the OHPP is expected to be integrated into routine care for elderly diabetic patients. 

Supplementary Material 1. 

