219

The Influence of Gingival Inflammation, Tooth Type, and Timing on the Rate of Plaque Formation* Marc

Quirynen,

Christel

Dekeyser,

and Daniel

van

Steenberghe

The undisturbed supragingival plaque formation was recorded twice over a 100 hours period in 14 healthy dental students; first after induction of gingival inflammation and a second time starting from a healthy gingiva. During each experimental segment plaque formation was recorded on 42 randomly selected and clean teeth at the start and after 6, 12, 24, 30, 36, 48, 54, 60, 72, 84, and 96 hours. Four hours later all teeth up to the first molars were examined. The presence of disclosable plaque was calculated planimetrically as percent of the total labial surface area of the tooth. The rate of plaque formation was found to be significantly greater (P 0.001) in the presence of gingival inflammation (+ 29.Wo). Moreover, this study indicates an important variation in the plaque growth rate within the dentition; the highest scores were found for the upper premolare, the upper molars, and the lower front teeth. Finally, the rate of plaque formation was found to decrease during the night. J Periodontol 1991; 62:219-222. =

Key Words: Dental plaque, occurrence; gingivitis/etiology; dental plaque/prevention and control.

Epidemiological surveys have repeatedly shown a definite relationship between bacterial plaque on teeth and periodontal breakdown.1 Clinical experiments in dogs and in man prove that plaque accumulation can lead to the inflammation of the periodontal tissues.2,3 The importance of plaque as the etiological factor of gingivitis and eventual Periodontitis has thus been sufficiently demonstrated. An efficient daily oral hygiene combined with a regular professional cleaning seems efficient and reliable to prevent this plaque accumulation,

even for many years.2 However, the bacterial recolonization of cleaned tooth surfaces occurs very rapidly,3-6 which is a major problem. Gingival inflammation seems to enhance this bacterial recolonization as indicated by clinical observations7'8 and by electron microscopy.9'10 The purpose of the present study was to quantify, in a practical way, the importance of the influence of gingival inflammation on the rate of plaque formation over a period of 96 hours. In addition we attempted to determine whether undisturbed plaque formation on the buccal tooth surface is different within different locations of the dentition and whether it is subjected to a diurnal variation.

MATERIALS AND METHODS

Study Population

Fourteen dental students, (5 females and 9 males; age range 20 to 23) participated in this experiment. They fulfilled the *

Catholic University of Leuven, Faculty of Medicine, School of Dentistry, Oral Pathology and Maxillo-Facial Surgery, Department of Periodontology, Leuven, Belgium.

following

criteria: 1) in good health without systemic dishad not used mouthrinses or medication that might eases; 2) have influenced their periodontal condition; 3) not using any orthodontic or removable prosthetic device; 4) no signs of Periodontitis nor mouth breathing.

Study Design

At their first visit the subjects received a thorough professional cleaning. They were then asked to refrain from any oral hygiene for 2 to 3 weeks in order to induce experimental gingivitis.11 At the end of this period the degree of gingival inflammation was scored, teeth were cleaned, and the first 100 hour period of undisturbed plaque formation (Phase I) started. The gingival inflammation of all teeth was measured by means of the Sulcus Bleeding Index (SBI).12 The mean SBI for all teeth per subject reached 1.31 (s.d. 0.38) and ranged from 0.8 to 1.9. The professional cleaning was performed with the help of a toothbrush, a toothpick, and curets if necessary. The teeth were also carefully polished with rubber cups and tap water. Using an 0.5% aqueous neutral red solution (3-Amino-7-dimethylamino-2-methylphenazine hydrochloride + ), a plaque disclosing solution with no antibacterial effect,13 the thorough plaque removal was checked and if necessary corrected. The experimental period always started before 12 a.m. The subjects were asked to refrain from all oral hygiene activity for the next 100 hours. A total of 42 teeth were selected at random: 7 central incisors, 7 canine teeth and 7 premolars in the left upper and right lower quadrant. One incisor, 1 canine, and

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INFLUENCES ON RATE OF PLAQUE FORMATION

1 premolar were chosen per subject. The selected teeth fulfilled the following criteria: no periodontal destruction, no carious lesions or restorations, no erosions nor hypo- or hyperplasia, a normal alignment in the dental arch, and no absence of antagonistic teeth or contact point. The presence of plaque on the labial surfaces of those teeth was recorded at the start and after 6, 12, 24, 30, 36, 48, 54, 60, 72, 84, and 96 hours from reproducible color slides, made at a magnification 2x, and after plaque disclosure with neutral red 0.5%. At the end of the experimental period (100 hours) the labial surfaces of all the teeth, including the first molars, were disclosed using an aqueous 4% erythrosine solution and once more color slides were taken, this time without magnification. At the end of Phase I, the dentition was professionally cleaned and the subjects were instructed to follow a thorough oral hygiene program until a gingiva free from any clinical sign of inflammation was obtained. Four to 6 weeks later, by which time a very healthy gingiva was established, Phase II of the experiment started. During Phase II the procedures carried out in Phase I of the experiment were repeated. At the start of Phase II the mean SBI for all teeth per subject reached 0.25 (S.D. 0.12) ranging from 0.0 to 0.5.

% PLAQUE

40r

I 30

20

10

J_l_

0

6 12

24 30 36

48 54 60

72

84

96

HOURS The influence of gingivitis on the plaque growth. Mean absolute increase in percentage of the labial tooth surface covered with plaque for the 42 selected teeth in Phase I, with gingivitis (·-·-·) and in Phase II, without gingivitis (O-O-O). The periods marked with a continuous line are day periods; the night periods are marked with a dotted line.

Figure 1:

Photographic Procedure To obtain reproducible slides the following measures were taken:14 the patient's head was immobilized in a head holder with a chin rest and a constant mouth opening was maintained by means of acrylic bite blocks placed on the second and third molars, standardizing the vertical and horizontal inclination between the camera and the tooth surface. Planimetrie

The

Plaque Analysis

extension on each tooth at each time interval evaluated was planimetrically. The color slides were en12.5 times and projected on white paper using a larged A projector. mirror, set at 45°, permitted us to leave the drawing paper in the horizontal plane. A rodium mirror was used to avoid parallax effects. The outline of the photographed tooth, the gingival margin, the papillae, and the area occupied by plaque were drawn. The total labial crown area and the plaque area were measured with a planimeter. The amount of plaque was expressed as a percentage of the total labial tooth surface. Occlusal contact regions, which for the lower teeth could interfere with undisturbed plaque growth, did not have to be taken into consideration since the plaque extension rarely reached these locations during the 100-hour observation period. More details on the methodology and the precautions of this scoring system can be found in a previous publication.14 All drawings and calculations were carried out by the same examiners in a blind randomized order. The reproducibility of duplicate registrations was examined in a previous study14 in which an intra-examiner correlation coefficient of 0.99 was found.

plaque

Data Analysis To examine the influence of gingivitis on the plaque formation rate, the results for each subject have been averaged across the three teeth for each time point of each phase. The average for a specific time point and phase served as the dependent variable for each subject. These averages have been compared across phases for each time point using paired i-tests. For the 100-hour data, the average plaque formation for each subject was calculated and a comparison between both phases was made by means of a paired i-test.

RESULTS The mean plaque growth on the 42 selected teeth during the first and second experimental phase is illustrated in Figure 1. The rate of plaque formation was clearly greater in the presence of gingival inflammation (mean SBI on the selected teeth: Phase I 1.33 vs. Phase II 0.23). This difference was already detectable after 36 hours. The statistical significance of the differences between both phases can be found in Table 1. Only after 36 hours a significant difference in plaque formation between both phases appeared. After 48 hours, this difference reached an important level of significance (P 0.002), which did not increase further on. After 96 hours of undisturbed plaque growth in the presence of gingivitis, a relative increase of 29% of the plaque extension was observed. =

=

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Volume 62 Number 3

QUIRYNEN, DEKEYSER, VAN STEENBERGHE

Table 1: Difference in Plaque Growth at Each Observation Between Phase I (with gingivitis) and Phase II (without gingivitis)

Time

value for 2-sided t Test

Absolute Difference in

Plaque (Gingivitis Healthy)

(hours)

-

0 6

-0.20 -0.09 0.07 0.88 0.89 1.84 3.44 3.89 5.04 5.80 7.67 7.79

12 24 30 36

48 54

60 72 84

96

Figure

2: The intra-oral

plaque

0.240 0.665 0.812 0.236 0.249 0.025 0.002 0.001 0.001 0.000 0.000 0.001

distribution. Distribution

of the

mean

percentage (14 subjects) of the labial tooth surface covered with plaque

after 100

hours

of undisturbed plaque formation; for gingivitis phase

(·-·-·) and for healthy phase (O-O-O).

Figure 1 also demonstrates that in both experimental phases, the plaque growth started very slowly during the first day. During day 2, the growth rate increased and remained relatively constant during day 3 and 4. Moreover, at least for the three types of teeth considered, the plaque growth rate seems to be reduced at night (period between 12 and 24, 36 and 48, 60 and 72, and 84 and 96 hours). The influence of the type of teeth on the supragingival plaque growth rate on the buccal surface is shown graphically in Figure 2. The mean 100 hours plaque score of all subjects is indicated for both phases and for each type of tooth. More disclosable plaque was found on the upper molars, the upper premolars, and the lower front teeth. The influence of the gingival inflammation on the plaque growth rate was also clear from this part of the experiment (P 0.001). =

DISCUSSION The results of the present study confirm the relationship between gingivitis and a higher plaque growth rate (P = 0.001). This is in agreement with previous reports.7-10,15 The presence of gingivitis explains a relative increase of 29% in plaque extension after 96 hours of undisturbed growth.

This is

221

an important difference, taking into account the small time interval in which it appeared. For some subjects, an increase of 50% in plaque growth rate was nearly reached due to the presence of gingival inflammation. Some methodological aspects can be put forward to explain the absence of a difference in growth rate during the first 24 hours, which could have been expected from previous scanning electron microscopical investigations910 which also compared plaque formation with and without the presence of gingivitis. It should be noted that in both these experiments, the examined teeth were surrounded by a plaque covered dentition, which implies a higher concentration of bacteria in the environment. In the present study, the lack of short-term differences may be due to the thorough cleaning of all teeth at the start of the experiment. A possible explanation for the faster plaque growth after the induction of gingivitis might be the increase in crevicular fluid due to gingival inflammation.1618 Indeed, Loesche19 proved the presence of important plaque nutrients in the crevicular fluid. Moreover, the gingival fluid might facilitate the bacterial adhesion in a number of ways, for example Silverman and Kleinberg20 established that divalent cations of calcium induced the aggregation of negatively charged bacteria and Hillam21 hypothesized that the high concentration of calcium present in gingival exúdate compared with that present in saliva might influence the rate of plaque formation. Proteins in saliva are known to bind macromolecules; a similar activity might be applied to those proteins found in the gingival fluid where protein precipitation might contribute to the aggregation of plaque bacteria.21 22 On the other hand, it has been postulated that the crevicular fluid might have a protective action.23 If this premise holds true, it certainly does not show in the evaluation of plaque growth. The protective role could be due to shifts among bacterial species composing the plaque, which was not evaluated here. Another hypothesis explaining the rapid plaque formation in the presence of gingivitis can be the increased bacterial concentration in the saliva. Van Houte et al.24 illustrated in vivo that the concentration of microorganisms has to exceed a certain threshold value before tooth colonization starts. Since brushing the tongue, which reduces the bacterial counts, does not influence plaque growth,25 this hypothesis remains open for further discussion. One could argue that the slight swelling of the gingiva due to inflammation favors an increased mechanical plaque retention at the cervical crown area and thus explains the higher plaque growth rate. It might also be argued that the more rapid plaque accumulation in the presence of gingivitis was due to the superficial enamel décalcification and roughening caused by the long-standing bacterial colonization during the induction of experimental gingivitis. These hypotheses cannot be assessed in this experiment. The plaque growth rate seems to decrease during the 12hour period in which the participants had their night rest, at least from the second day on when a minimal rate of

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INFLUENCES ON RATE OF PLAQUE FORMATION

plaque formation is established. This is in agreement with previous observations in which a 50% reduction in growth rate during the night has been recorded.26 Since it is known that the supragingival colonies can find their nutrients mainly in the saliva,27 the reduced salivary output at night28 seems an acceptable explanation for this decrease. The difference between day and night might even have been greater when

the students were instructed to retire 12 hours. The results from this study also indicate important variations in labial plaque growth within the dentition (Fig. 2). After lOO hours of undisturbed plaque growth, starting from a healthy gingiva, the upper molars areas scored the highest surface percentages (71.3%) followed by the upper premolare (56.5 %), the lower and upper canine teeth and the lower incisors (all around 40%). The upper lateral incisors, the lower molars and premolare scored 37.4%, 36.5%, and 34.1% respectively, while the upper central incisors accumulated the least amount of plaque (23.8%). The reproducibility of the plaque growth measured in both phases of the experiment, and the symmetrical distribution in a sagittal plane indicate the significance of this variation in plaque accumulation throughout the dentition. Moreover, it illustrates that the observed differences cannot be solely due to height variations in tooth crown. Although not dealing with undisturbed plaque growth two other studies, one dealing with variations in tooth brushing frequencies29 and one concerning habitual oral hygiene of not instructed dental students,30 found a similar intra-oral distribution of plaque. However, in the presence of mechanical plaque control, the lower front showed relatively less plaque. The large interindividual variations which were reported in the quoted studies and which were not obvious from our results can be explained by differences in oral hygiene.

Acknowledgments

The authors express their appreciation to Mrs. M. Hillen who corrected the English language and to Mrs. Vuylsteke (Computing Centre, K.U. Leuven) for the statistical analyses. REFERENCES 1. Theilade J. Dental plaque and dental calculus. 1975 In: Lindhe J, ed. Textbook of Clinical Periodontology. Copenhagen: Munksgaard, 1983:3. 2. Axelsson P, Lindhe J. Effect of controlled oral hygiene procedures on caries and periodontal disease in adults. Results after 6 years. / Clin Periodontol 1981;8:239-248. 3. Rönström A, Attström R, Egelberg J. Early formation of dental plaque on plastic films. 1. Light microscopic observations. J Periodont Res

1975;10:28-35. 4. Lie T. Ultrastructural study of early dental plaque formation. J Periodont Res 1978;13:391^109. 5. Berthold P. Formation of salivary coating and dental plaque on two different supporting materials. An electron microscopic study. J Periodontol 1979;50:397-405. 6. Quirynen M. Anatomical and inflammatory factors influence bacterial plaque growth and retention in man. [Thesis]. Leuven: Catholic Uni-

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1985;20:321-328. 15. Goldman RS, Abelson DC, Mandel ID, Chilton NW. The effect of various disclosants on plaque accumulation in human subjects. J Periodont Res 1974;9:381-385. 16. Rudin HJ, Overdiek HF, Rateitschak KH. Correlation between sulcus fluid rate and clinical and histological inflammation of the marginal gingiva. Helv Odontol Acta 1970;14:21-26. 17. Wilson AG, McHugh WD. Gingival exúdate an index of gingivitis? DentPractit 1971;21:261-266. 18. Attström R, Egelberg J. Presence of leukocytes within the gingival crevices during developing gingivitis in dogs. / Periodont Res 1971;6:110-114. 19. Loesche WJ. Importance of nutrition in gingival crevice microbial ecology. Periodontics 1968;6:245-249. 20. Silverman G, Kleinberg J. Studies on factors affecting the aggregation of the micro-organisms in human dental plaque. Arch Oral Biol -

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21. Hillam DG. Dental plaque and associated deposits on the teeth. In: Lavelle CLB, ed. Applied Physiology of the Mouth. Bristol: John Wright & Sons Limited, 1975:124. 22. Hay DI, Gibbons RJ, Spinell DM. Characteristics of some high molecular weight constituents with bacterial aggregating activity from whole saliva and dental plaque. Caries Res 1971;5:111-115. 23. Cimasoni G. The crevicular fluid. Monographs in Oral Science. Basel: Karger; 1974. 24. Van Houte J, Gibbons RJ, Banghart SB. Adherence as a determinant of the presence of Streptococcus salivarius and Streptococcus sanguis on the human tooth surface. Arch Oral Biol 1970 15:1025-1034. 25. Badersten A, Egelberg J, Jonsson G, Kroneng M. Effect of tongue brushing on formation of dental plaque. J Periodontol 1975;46:625627. 26. Quirynen M, van Steenberghe D. Is early plaque growth rate constant with time?/a/n Periodontol 1990;16:278-283. 27. Carlsson J. Microbiology of plaque associated periodontal disease. In: Lindhe J, ed. Textbook of Clinical Periodontology. Copenhagen:

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Send reprint requests to: Dr. M. Quirynen, Department of Periodontology, Catholic University of Leuven, Capucijnenvoer 7, B-3000 Leuven, Belgium. Accepted for publication September 27, 1990.