Original Article Int J Clin Prev Dent 2015;11(3):143-152ㆍhttp://dx.doi.org/10.15236/ijcpd.2015.11.3.143 ISSN (Print) 1738-8546ㆍISSN (Online) 2287-6197 Changes in Oral Microflora with Oral Gargling and Interdental Brushing Su-Hyang Kim1, Chung-Jae Lee2, Kyu-Hwan Lee3, Su-Kyung Jwa4 1Department of Oral Health, Graduate School, Dankook University, Cheonan, 2Department of Dental Laboratory Technology, Shin Han University, Uijeongbu, 3Department of Health Promotion Center, Seoul National University Bundang Hospital, Seongnam, 4Department of Dental Hygiene, Daegu Health College, Daegu, Korea Objective: The study used phase contrast microscopy to examine the influence on the proximal oral bacterial microflora af- ter interdental brushing using cethyl pyridium chloride (CPC) solution in 50 adult volunteers. Methods: The study was done from August to September 2014 at the dental public health center in Deagu-si with Dankook University Hospital Institutional Review Board approval. Results: Numerous cocci present at the interproximal site prior to use of CPC were markedly reduced at the interdental brush tip following CPC use (p＜0.01). Coccai motility as an indication of motility was no different prior to and after CPC applica- tion(p＞0.05). The total number of Bacillus and number of motile Bacillus were significantly after CPC brushing (both p＜0.01). Total number and number of motile filamentous and spiral microorganisms were decreased after CPC applica- tion(all p＜0.05). The results indicate the effcacy of mouth rinsing with CPC. While there may be a relationship between the total and motile numbers of oral microorganisms, no relationship was routinely evident between age and microbe number. Conclusions: Use of CPC to sterilize the tip of the interdental brush after use the proximal area is recommended to achieve the best antiseptic effect. Keywords: cethyl pyridium chloride, interdental brush Introduction ing in Korea, with a 7.3% increase recorded from 2006 to 2011 [1]. Dental plaque is produced on the outer surface of the pelicle. The prevalence of periodontal disease, which is mainly due Primary plaque colonizers in the early stage of plaque formation to microorganisms resident in dental plaque, has been increas- include aerobic. Gram positive bacteria. As dental plaue thick- ens and oxygen becomes more limited in the interior of the pla- que, anaerobic bacteria including Gram negative bacteria in- Corresponding author Su-Kyung Jwa habit the plaque [2]. The latter include Prophyromonas gingivalis, Department of Dental Hygiene, Daegu Health College, 15 Prevolella intermidia, Aggregatibacter actinomycetemcomitans, Youngsong-ro, Buk-gu, Daegu 41453, Korea. Tel: +82-53- Tannerella forsythia, and Fusobacterium nucleatum [3]. 320-4516, Fax: +82-53-320-1345, E-mail: [email protected]. ac.kr Microorganisms that develop at the interproximal area and in the gingival pocket proximally are hard to remove. Received March 24, 2015, Revised June 16, 2015, Specialized oral hygiene devices like silk dental floss and inter- Accepted June 17, 2015 dental brush are needed. Indications for use of the interdental Copyright ⓒ 2015. Korean Academy of Preventive Dentistry. All rights reserved. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. 143 International Journal of Clinical Preventive Dentistry brush include patients with a wide interdental area. Periodontal Materials and Methods surgery, use of fixed prosthodontic appliance, implant and or- thodontic appliances [2]. 1. Subjects Interdental brushes and toothburshes should be anti- septically cleansed for optimal benefit and to avoid transfer of Fifty adult volunteers recruited at a public health center from microorganisms to tooth surfaces during brushing. Toothbrushes August to September 2014 participated in this study. The study can harbor millions to billions of bacteria, which can be deliv- was approved by Dankook University Hospital Institutional ered to the oral cavity [4]. Monthly replacement of a toothbrush Review Board. The age and the gender distribution of the sub- is recommended [5]. Differences in the numbers of micro- jects were shown in Table 1. organisms in the toothbrush bristle according to the cleaning 2. Materials method and cleansing time have been reported [6]. The tip of an interdental brush should be cleaned using an an- An interdental brush (Dr. Han’s Brush; Wooribiogen, Seoul, tiseptic, typically cethyl pyridium chloride (CPC). CPC is a Korea) and 0.05% CPC solution were provided to all subjects quatermary ammonium compound that exerts its bacteriostatic for cleaning of the proximal site and tip of interdental brush or bactericidal antiseptic action by disrupting the cell wall of (Figure 1). Gram postive bacterial and cell envelope of Gram negative bac- teria [7-10]. Xantolisol is an especially potent ingredient [11,12]. It’s merits include lack of staining of teeth and a palat- able taste compared to chlorhexidine [13]. Phase contrast microscopy has proven useful to measure the total number of oral microorganisms on the surface of interest. As well, the approach can detemine the number of motile mi- crobes as an indication of viability [14]. However, the method cannot speciate microorganisms and can be subject to examiner bias [15]. Oral microorganisms commonly identified using phase contrast microscopy are cocci, bacillus, filamentous and spiral shaped types. A caries-related environment tends to be enriched in motile cocci and bacilli with motile, with perio- dontal disease associated with filamentous and spiral micro- organisms [14-17]. Patient motivation for proper oral health could be boosted by Figure 1. Photograph of an interdental brush and cethyl pyridium the use of phase contrast microscopy, with images of the mi- chloride solution. crobes present in a patients’ mouth stimulating resolve fo proper dental hygiene [18,19]. The present study explored the use of phase contrast microscopy to chart the effect CPC sterilization of and interdental brush after use of the brush at the proximal region of the mouth. Table 1. Age and gender of the subjects Age (yr) Gender 30s 40s 50s 60s Total Male (n) 8 2 8 0 18 Female (n) 10 15 6 1 32 Figure 2. The interdental brush was moved back-and-forth times Total (n) 18 17 14 1 50 across the proximal area. 144 Vol. 11, No. 3, September 2015 Su-Hyang Kim, et al：Changes in Oral Microflora with Oral Gargling and Interdental Brushing Figure 3. Sterilzation with cethyl pyridium chloride. Figure 4. Applying of cethyl pyridium chloride at proximal area. 3. Method The interdental brush was used five times at the proximal area. Residue on the brush was smeared on a glass slide and cov- ered with cover glass. Motile bacteria were recorded for 10 sec- onds during phase contrast microscopy examination (Figure 2). The brush was shaken in CPC solution for 1 minute and the resi- due examined as just described (Figure 3). CPC was applied 5 times to at the proximal area of the interdental brush tip (Figure 4). Figure 5, 6 display representative images of oral micro- organisms recovered from the proximal area before and after 1-minute gargling with CPC solution, respectively. Dental pla- que recovered using a micro-brush was smeared on a slide glass, a drop of saline was added and the suspension was covered with Figure 5. Recovery of microorganisms from the proximal area before a cover glass (Figure 7). gargling with cethyl pyridium chloride. www.ijcpd.org 145 International Journal of Clinical Preventive Dentistry Figure 6. Microorganism recovery form the proximal area after cethyl pyridium chloride gargling. Figure 7. Preparation of a smear for phase contrast microscopy examination. and numbers of motile microorganisms were ranked on a scale of 0 to 3 (Table 2) [20]. 5. Statistical analysis SPSS ver. 12.0 software (SPSS Inc., Chicago, IL, USA) was used for Pearsons’ correlation coefficient statistical analysis af- ter calculating the mean and standard deviation. Results Total number and motile number of cocci (Table 3-5), bacilli (Table 6-8) and filamentous (Table 9-11) microorganisms are Figure 8. Various types of oral microorganism shown in a representative summarized. Correlatons between the total number and motile phase contrast microscopy image. numbers of microorganisms before and after CPC sterilization are presented in shown in Table 12-16. CPC sterilization for a minute in CPC solution marked re- 4. Microscopic findings duced the number of cocci, with the reduction in cell number Oral microorganisms were observed using a model DCS being more modest when CPC was applied at the proximal area 6002 phase contrast microscope (Dr. Prevent, Seoul, Korea) at for a minute (Table 3). The reduction in the number of motile a magnification of ×400 on the image monitor after recording cocci in the proximal region was not as evident (p>0.05). But the image for 10 seconds. The morphological types of the micro- sterilization the tip of the interdental brush by a 1-minute shakin organisms was classified as cocci, bacillus, filamentous and in CPC solution was effective (p<0.05) (Table 4). The number comma/spiral type (Figure 8). The number of micoorganisms of cocci prior to tip sterilization and following gargling with 146 Vol. 11, No. 3, September 2015 Su-Hyang Kim, et al：Changes in Oral Microflora with Oral Gargling and Interdental Brushing Table 2. Standardization of total number and motile number of oral microorganisms classified on the basis of morphology [20] Cocci type Bacilli type Filament type Comma/spiral type Score Amount Motile Amount Motile Amount Motile Amount Motile 0 None None None None None None None None 1 1-7 Trembling 1-7 Trembling 1-7 Trembling 1-3 Trembling 2 8-70 Slight movement 8-30 Slight movement 8-30 Slight movement 4-8 Slight movement 3 ＞70 Active movement ＞30 Active movement ＞30 Active movement ＞8 Active movement Table 3.