Vol. 46, No. 1INFECTION AND IMMUNITY, Oct. 1984, p. 1-60019-9567/84/100001-06$02.00/0Copyright © 1984, American Society for Microbiology

Bacteriology of Experimental Gingivitis in ChildrenW. E. C. MOORE,'* L. V. HOLDEMAN,' R. M. SMIBERT,' E. P. CATO,1 J. A. B3URMEISTER,2 K. G. PALCANIS,2

AND R. R. RANNEY2Department of Anaerobic Microbiology, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061,and Department of Periodontics, School of Dentistry, Virginia Commonwealth University, Richmond, Virginia 232982

Received 16 April 1984/Accepted 6 July 1984

Children are more resistant to gingivitis than are adults. To determine possible differences in theirperiodontal floras, an experimental gingivitis study, identical in design to one reported earlier with youngadults, was conducted with four 4- to 6-year-old children. The incidence of sites that developed gingival indexscores of 2 in children was less than one-third of the incidence observed in adults. The composition of the floraof each child was statistically significantly different from that of any other child or adult. The floras of thechildren as a group were statistically significantly different from those of the adults. Children had 3-fold greaterproportions of Leptotrichia species, 2.5-fold greater proportions of Capnocytophaga species, 2.3-fold greaterproportions of Selenomonas species, 2-fold greater proportions of bacterial species that require formate andfumarate, and 1.5-fold greater proportions of Bacteroides species. Adults had greater proportions ofFusobacterium, Eubacterium, and Lactobacillus species. Fusobacterium nucleatum, Actinomyces WVa 963,Selenomonas D04, and Treponema socranskii were predominant species that correlated with increasing gingivalindex scores in both children and adults.

Experimental gingivitis in humans (4) provides a model forassessing variables that may be related to the developmentof gingivitis. The bacteriology of this model with young adultsubjects, including the identification of all randomly selectedisolates and the most likely causative bacterial species, waspreviously described (12). Others (7, 9) have shown thatprepubertal children do not develop gingivitis in this modelto the same extent or severity as do adults. Together withcross-sectional data indicating a lesser severity of naturallyoccurring gingivitis in children than in adults (18), thisindicates a greater resistance of children to the developmentof gingivitis. There are suggestions in the literature that thehost responses expressed in the gingiva of children maydiffer from those of adults. Histologically, gingivitis inchildren is dominated by lymphocytes and includes fewerplasma cells than does gingivitis in adults (5, 6). The childgingivitis lesion has been characterized as T-cell dominated(15), whereas the predominant cells in the infiltrate of adultsare of the B-cell series (14). If the floras of developinggingivitis in children are identical to those in adults, thegreater resistance of children may be attributable to hostresponse differences, quite possibly the immunological re-sponse differences suggested by the distinct characteristicsof the inflammatory infiltrates. To determine whether thefloras are the same, the present study duplicated in childrenthe previous design and procedures in adults (12).

MATERIALS AND METHODSSubjects. Four Caucasian males, ages 4 to 6 years and

having primary dentition, participated as subjects. Theywere medically healthy by history and received no medica-tions during the course of the study and no antibiotics for 2months preceding the study. There were no instances ofprobeable gingival sulcus depth greater than 3 mm and noclinically detectable carious lesions of the teeth.

Experimental gingivitis. The experimental gingivitis modelas developed by Loe et al. (4), and as utilized in the previousstudy of young adults (12), was followed. This included thecleaning of all teeth and establishing base-line health, which

* Corresponding author.

was verified by examination of the preselected sites on day 0of the experimental phase (no brushing or other toothcleaning). The experimental phase continued for 25 days. Asecond, identical experiment was conducted with each childafter reestablishing gingival health and allowing a minimumof 3 months between the two trials. The gingival index (GI)(3) and plaque index (16) scores were determined for foursurfaces of all teeth present on each sample day. All exami-nations were performed by the same examiner, who also haddone all clinical scoring in the previous experiment withadults.Sample sites. As in the adult experiment, samples were

taken from two sites (one facial and one proximal) on each oftwo teeth (one mandibular and one maxillary) on each ofdays 4, 11, and 25 of the experimental phase (Table 1). Ingeneral, the sample sites occupied the same relative locationin the dental arches as the sites sampled previously in adults(12). However, samples were taken only from primary teeth.Additionally, sites were chosen to be nonadjacent to anysites of lost primary incisors. As in adults, a different surfacewas sampled each time within each experimental replicatetrial, and no proximal sample site was immediately adjacentto any other sample site. Left canines were used to avoiddisturbing mesial sample sites on right primary molars.Thus, each sample came f'rom a site in which the plaque wascompletely undisturbed for the designated time. There were24 facial and 24 proximal (mesial or distal) sites sampled ineach trial.

Sampling. The teeth were isolated with cotton rolls, andthe sample sites were gently dried with sterile cotton swabs.The accumulated supragingival plaque at the sites to besampled was removed as completely as possible with steriletoothpicks. The samples were then taken with sterile nickel-plated Morse 00 replaceable scaler tips by making a singlepass from the depth of the sulcus to just coronal to thegingival margin as was done previously for the adult experi-ment.

Microbiology. All sample dispersion, dilution, media, cul-tures on plates and roll tubes, spirochete, and other bacterialidentification were as described previously for the adultexperiment (12). In brief, randomly selected isolates from D-

1

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from

2 MOORE ET AL.

TABLE 1. Sample site schedule for both replicates ofexperimental gingivitis in children

Sample site schedule for childa:Day

1 2 3 4

4 H (F,D) A (F,M) H (F,D) B (F,M)M (F,D) T (F,M) S (F,M) M (F,D)

11 A (F,M) H (F,D) B (F,M) A (F,M)T (F,M) S (F,M) T (F,M) T (F,M)

25 B (F,M) B (F,M) A (F,M,Lb) H (F,D)S (F,M) M (F,D) M (F,D) S (F,M)J (F bMb) T (Lb)L (F bDb)

"Universal identification system for deciduous teeth (A to T), followed bysampled surfaces within parentheses: F, facial; D, distal; M, mesial; L,lingual.bExtra sites sampled in trial I in an attempt to increase the number of sites

with GI scores of 2 for statistical analyses.

4 medium (brain heart infusion base) were subcultured in D-5broth and identified by comparison with type and referencestrains after characterization by electrophoresis of solublecellular proteins (10), Gram stain, chromatographic analysisof fermentation acids and hydrogen, growth and pH in 30media, and tests for indole, esculin hydrolysis, catalase,protein digestion, gas production, motility, aerotolerance,ammonia production, and nitrate reduction (2), supplement-ed by other tests as required for differentiation of species inseveral genera. Strains of the genus Actinomyces wereserotyped with monovalent fluorescent antisera. Spirocheteswere isolated from OTI medium or RIP broth on peptone-fresh yeast extract-serum agar and identified by morpholo-gy, serology, fermentation patterns, and biochemical reac-tions (12).

RESULTS AND DISCUSSION

Of the 96 predesignated sample sites in both trials, 46 werefound to have GI scores of 0, 39 had GI scores of 1, and 11had GI scores of 2 (Table 2); this compares with GI scores of18, 42, and 36, respectively, in adults (12) under the sameexperimental conditions (Table 3). The mean GI scores ofsample sites were significantly lower for the children than forthe previously studied adults on corresponding sample days(P < 0.001 for days 4 and 11 and P < 0.005 for day 25 by ttests). At the same time the plaque index scores for samplesites in children were higher than for sample sites in adults atday 4 (1.75 ± 0.08 versus 1.28 ± 0.08) and essentially the

TABLE 3. Child versus adult experimental gingivitis

Subjects (yr) GI No. of No. of No. of %score samples isolates taxa Coverage

Children (4-6) 0 46 1,449 122 971 43a 1,363 122 972 13a 405 65 95

Adults (22-31)b 0 18 562 65 961 42 1,332 96 982 36 1,141 110 97

a Includes four extra sites with GI scores of 1 and two extra sites with GIscores of 2.

b Data are from reference 12.

same at days 11 and 25 (1.56 ± 0.09 versus 1.56 ± 0.12 and1.94 ± 0.04 versus 2.09 ± 0.07, respectively). These findingsare consistent with previous reports that children do notdevelop gingivitis to the same extent or severity as do adultsin the experimental gingivitis model, even though estimatesof plaque mass are comparable in the two populations (7, 9).

Trials. Neither the GI scores (P > 0.3) nor the plaqueindex scores (P > 0.6) at sample sites differed significantlybetween trials in the children (two-way analysis of variance).However, L analysis (12) showed that the composition of allsamples in trial I was different from that of all samples in trialII (P = 0.01). In the experiment with adults, there also was astatistically significant difference between trials I and II (P =0.004). In the adults it appeared that there may have beensome carry-over between trials (12), but there was noevidence for that in the present experiment. In the children,the floras of day 4 in trials I and II were not significantlydifferent (P = 0.29), nor were they significantly different onday 11 (P = 0.44). However, the composition of the floraswas significantly different on day 26 (P = 0.002). There weresix predesignated sites with GI scores of 2 on day 26 of thefirst trial and only two such sites on day 26 of the secondtrial. The proportions of most bacterial species in the floraswere reasonably similar in both trials on day 26. However,Actinomyces israelii II, Actinomyces meyeri (-) (phenotypi-cally like A. meyeri but not reactive with monovalent A.meyeri antiserum), Fusobacterium nucleatum, Streptococ-cus Dll, Streptococcus mitis I, Streptococcus sanguis I,Selenomonas D04, and Wolinella curva (19) were each threetimes or more as numerous in trial I as in trial II on day 26.Factors that may have caused the different responses in thetwo trials are not obvious.

Facial sites versus proximal sites. The floras of facial siteswere not significantly different from those of proximal (distalor mesial) sites in children (P = 0.19); neither were GI scores

TABLE 2. Distribution of GI scoresa among sites' sampled in childrenDistribution of GI scores in:

Trial Day Child 1 site: Child 2 site: Child 3 site: Child 4 site: Mean GIscore ± SE

1 2 3 4 1 2 3 4 1 2 3 4 1 2 3 4

I 4 0 0 0 0 0 0 0 0 0 0 1 1 0 0 1 0 0.19 ± 0.1011 0 1 0 0 0 0 2 0 1 1 1 1 0 1 1 0 0.56 0.1625 1 2 1 1 2 2 1 1 2 2 1 1 1 1 2 1 1.38 0.12

II 4 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0.00 0.0011 1 1 1 0 0 0 2 2 0 1 1 1 0 0 0 0 0.62 0.1125 1 1 2 1 1 1 1 1 2 1 1 1 1 1 1 0 1.06±0.11

aThe criteria of Loe and Silness (3) were used to determine GI scores; essentially, 0 is healthy, 1 is mild inflammation, and 2 is moderate inflammation andbleeding when gently probed.

b Sites sampled are given in Table 1 and in the text. Sites 1 and 2 were facial and proximal, respectively, of the first tooth listed; sites 3 and 4 were facial andproximal, respectively, of the second tooth listed.

INFECT. IMMUN.

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from

EXPERIMENTAL GINGIVITIS IN CHILDREN 3

TABLE 4. Percent similarity of floras of people' (720+ isolateseach)

% Similarity of floras of people:Person Cl C2 C3 C4 Al A2 A3 A4

Cl 100 58 62 64 58 58 50 46C2 100 53 56 51 56 49 53C3 100 65 52 52 44 36C4 100 61 62 45 46Al 100 63 52 56A2 100 55 49A3 100 52A4 100a C, Child; A, adult.

significantly different (0.69 + 0.10 for facial and 0.58 ± 0.10for proximal sample sites). These findings are in contrast tothose in adults, in which both the flora composition (P =

0.004) and GI scores differed between facial and proximalsites (12). These differences between children and adultsmay relate to differences in tooth shape or proximal toothcontact areas which could produce a different local environ-ment, flora, and consequent pattern of gingivitis develop-ment. The floras of facial sites in children were significantlydifferent from those of facial sites in adults (P < 0.001), andthe floras of proximal sites also differed between childrenand adults (P < 0.001). Comparisons of child proximal toadult facial sites, and child facial to adult proximal sites,indicated equally large differences (P < 0.001). Thus thedifferences in flora compositions apparently reflect a differ-ence in the total flora of children versus that of adults.Flora composition. There were 160 taxa among 3,217

isolates from all 102 samples from the children. The floras ofthe children were slightly more complex than those of theadults. For example, in children 122 taxa among 1,363isolates from sites with GI scores of 1 accounted for 97% ofthe cultivable floras, whereas in adults only 96 taxa among1,332 isolates accounted for 98% of the floras, as estimatedby formula Hg for coverage of Good (1) (Table 3).The floras of children and adults were compared by the L

analysis of Good (12). To compare the floras of children as a

group with the floras of adults, all 24 or more samples fromeach person were merged into a single sample of ca. 720isolates to represent the flora of that person. L analysisdemonstrated that the four children and the four adultsformed the two most distinct groups possible with a proba-bility of 0.028 (Table 4). Because L analysis is a Monte Carlotest, 0.028 is the minimum theoretical probability that can be

TABLE 5. Mean % similarity of 24 (30-colony) samples withinand between peopleaMean % similarity of floras of people

PersonCl C2 C3 C4 Al A2 A3 A4

Cl 25 18 25 24b 23 25 17 16C2 20 19 17 18 20 13 15C3 32 26 23 25 17 14C4 28 24 26 18 18Al 29 28 19 19A2 34 21 18A3 21 16A4 20a C, Child; A, adult.b By L analysis, P between Cl and C4 = 0.003, and P between all others

<0.001.

attained with two groups of only four items. (Randomassignment of samples will produce the identical two groupsof four with 2.8% frequency.) The mean similarity of thefloras of different children was 60%, of different adults was54%, and of children compared with adults was 51%.The compositions of all 24 to 28 samples from each child

or adult were significantly different from those of the sam-ples from each other child or adult when one pair of people ata time was tested (Table 5). In both children and adults themean similarity of 24 to 28 samples was 26%. The averagesimilarity of samples was 22% among different children and20% among different adults. The average similarity of sam-ples from children as compared with samples from adultswas 20%.

Differences in composition of the child and adult floras areshown in Tables 6 and 7. These tables list all taxa thatcomprised at least 0.5% of the floras of either adults orchildren. The children, who had greater resistance to gingivi-tis, had 2.5-fold higher proportions of all Capnocytophagaspecies combined, and 2.2-fold higher proportions of allanaerobic vibrios (species of the combined genera Campy-lobacter, Selenomonas, and Wolinella). These data suggestthat this flora composition is normal for children and thatfindings of significant numbers of Capnocytophaga speciesin young persons, as reported recently in juvenile diabetics(8), should be compared with findings in healthy controlsubjects of the same age to assess etiological significance.

Relationship of floras to GI scores. The composition of thefloras of sites with GI scores of 0 was significantly differentfrom those with GI scores of 1 (P < 0.001) or GI scores of 2(P = 0.002), but the floras of sites with GI scores of 1 werenot statistically significantly different from those of siteswith GI scores of 2 (P = 0.11). Flora compositions of siteswith GI scores of 0 in children were significantly differentfrom those in adults (P < 0.001), as were sites with GIscores of 1 (P < 0.001) and sites with GI scores of 2 (P =0.012). The lesser degree of significance between child andadult sites with GI scores of 2 may reflect the fact that therewere only 13 sites with GI scores of 2 in children and 36 siteswith GI scores of 2 in adults, whereas for GI scores of 0there were 46 in children and 18 in adults, and for GI scoresof 1 there were 43 in children and 42 in adults.

Bacterial species associated with gingivitis. The floras ofadults, who were more susceptible to gingivitis, containedgreater numbers of predominating gram-positive species(Table 7). The notable exception is F. nucleatum, which isgram negative.Among the predominant species in adult experimental

gingivitis, only five increase further in sulci of sites affectedwith moderate or severe periodontitis (Table 8). Of these fivespecies, both A. israelii I and Propionibacterium acnes aremost frequent in the gingival crevices of people with healthygingiva, whereas the incidence of Eubacterium timidum, F.nucleatum and Peptostreptococcus micros suggests thatthey might make the greatest contributions to the increasedsensitivity of adults to experimental gingivitis and to perio-dontitis.Mycoplasma. The incidence of mycoplasma among sam-

ples from different populations is shown in Table 9. Theirdistribution indicates that they do not contribute to gingivi-tis; rather, it appears that they occur as a result of gingivitisand might be important in periodontitis tissue destruction.Treponemes. Treponema socranskii was frequent in the

gingival crevice of people with healthy gingiva and in chil-dren and adults with experimental gingivitis. In both childrenand adults its frequency correlated with increasing GI

VOL. 46, 1984

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from

4 MOORE ET AL.

TABLE 6. Incidence of predominant species more common in children, as percent of total flora and as percent of (30-isolate) samplesthat are positive

% of flora % of samplesSpecies

Childa AdUltb Child/adult Childa Adultb Child/adult

Gram positiveActinomyces WVa 963 6.1 0.8 7.6 46 12 3.8Actinomyces israelii (-) 2.7 1.4 1.9 39 14 2.8Actinomyces israelii II 2.9 0.6 4.8 31 4 7.8Actinomyces naeslundii I 10.0 6.9 1.4 70 49 1.4Fac. Pos. Coccus D2SC 0.5 0.1 5.0 10 1 10.0Streptococcus anginosus 6.6 5.9 1.1 59 46 1.3Streptococcus D02 1.0 0.1 10.0 11 1 11.0Streptococcus D07 2.2 1.4 1.6 30 20 1.5

Gram negativeBacteroides D1C20 0.8 0.0 11 0Bacteroides gracilis 0.8 0.6 1.3 18 11 1.6Bacteroides intermedius 8944 1.4 0.6 2.3 20 7 2.9Capnocytophaga gingivalis 2.9 1.8 1.6 46 25 1.8Capnocytophaga ochracea 3.4 0.6 5.7 52 14 3.7Capnocytophaga sputigena 0.9 0.6 1.5 21 11 1.9Selenomonas sputigena 1.3 0.6 2.2 20 10 2.0Selenomonas DOI 0.8 0.0 10 0Selenomonas D02 0.7 0.3 2.3 8 3 2.7Selenomonas D04 2.4 0.8 3.0 28 10 2.8Veillonella dispar 0.7 0.2 3.5 17 4 4.2Wolinella curva 0.5 0.1 5.0 11 2 5.5Wolinella recta 1.2 0.7 1.7 23 15 1.5a Children: 3,217 isolates from 102 samples.b Adults: 3,035 isolates from 96 samples (12).c Unnamed species are designated by letters and numbers.

TABLE 7. Incidence of predominant species more common in adults, as percent of total flora and as percent of (30-isolate) samples thatare positive

% of flora % of samplesSpecies

Childa Adultb Child/adult Childa Adultb Child/adult

Gram positiveActinomyces israelii I 0.2 0.6 3.0 7 10 1.4Actinomyces naeslundii (-) 2.8 5.1 1.8 31 46 1.5Actinomyces naeslundii III 0.7 2.7 3.9 5 23 4.6Actinomyces NVC 3.1 4.0 1.3 37 41 1.1Actinomyces odontolyticus (-) 0.3 1.7 5.7 7 27 3.9Actinomyces odontolyticus I 2.1 3.7 1.8 31 51 1.6Actinomyces viscosus II 0.5 1.6 3.2 11 29 2.6Coccus D30d 0.0 0.6 0 9Coccus SM1 0.6 1.4 2.3 18 26 1.4Eubacterium saburreum 0.6 0.9 1.5 16 19 1.2Eubacterium timidum 0.1 0.5 5.0 3 9 3.0Lactobacillus D02 0.8 2.1 2.6 13 17 1.3Propionibacterium acnes 0.5 1.0 2.0 12 16 1.3Peptostreptococcus micros 1.0 1.4 1.4 16 21 1.3Streptococcus D16 1.6 2.6 1.6 22 32 1.5Streptococcus mitis 0.8 1.0 1.2 18 18 1.0Streptococcus SA 0.1 0.8 8.0 4 17 4.2Streptococcus sanguis I 1.0 6.7 6.7 20 56 2.8Streptococcus sanguis II 1.5 3.0 2.0 29 36 1.2Streptococcus SM 0.4 1.6 4.0 12 29 2.4

Gram negativeBacteroides oris 0.1 1.2 12.0 4 16 4.0Fusobacterium nucleatum 3.4 4.4 1.3 50 51 1.0Veillonella parvula 11.4 11.7 1.0 81 85 1.0a Children: 3,217 isolates from 102 samples.b Adults: 3,035 isolates from 96 samples (12).c Serologically related to both A. naeslundii and A. viscosus II.d Unnamed species are designated by letters and numbers.

INFECT. IMMUN.

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from

EXPERIMENTAL GINGIVITIS IN CHILDREN 5

TABLE 8. Concentrations of some suspect etiological agents in subgingival periodontal floras as % of floraConcn as % of flora in:

Species Adult Child Adult Adult Adult (severe(healthy)' (experimental (experimental (moderate Aeidultti(severgingivitis) gingivitis)b periodontitis) periodontitis)d

Actinomyces israelii I 1.3 0.2 0.6 0.8 0.8Eubacterium timidum 0.9 0.1 0.5 2.0 6.2Fusobacterium nucleatum 4.2 3.4 4.4 9.4 7.5Propionibacterium acnes 1.6 0.5 1.0 1.3 1.3Peptostreptococcus micros 2.8 1.0 1.4 5.2 4.4

a Data from periodontally healthy adults, studied as control subjects (11-13).bData from reference 12.c Data from reference 11.d Data from reference 13.

scores, and its higher incidence in moderate and severe detect the incidence of treponemes, each observation resultsperiodontitis suggests that this species may be important in from treponemes among 104 or more cells, whereas in theboth gingivitis and periodontitis. Several other treponemal other bacteriological data (Tables 6 and 7), the frequency ofspecies appear to occur as a result of gingivitis and are detection is based upon a sample size of only 30 colonies.associated primarily with moderate or severe periodontitis. The different floras among and between children, adults,The data (Table 9) indicate that dark-field microscopic and disease states may help to pinpoint the few bacterialobservations of samples dispersed in 1.5 ml of broth tend to species in common that could be causative agents. Of 160underestimate the incidence of treponemes that can be taxa detected in this study, only T. socranskii and 11cultured from healthy or gingivitis sites. Treponemal cul- nontreponemal species (Table 10) increased in both childrentures often were obtained from low dilutions of samples even and adults as the GI scores increased. In children, thethough no spirochetes were seen by dark-field examination. frequently detected nonspirochetal species (>0.05% of theConversely, cultural results may underestimate the number flora) which increased in proportion in the flora as GIof treponemes in moderate or severe periodontitis because score increased were Actinomyces WVA 963, A. israelii II,we have not cultured the "large treponeme" that is frequent Actinomyces naeslundii (-), Capnocytophaga gingivalis,in these sites. Capnocytophaga ochracea, Capnocytophaga sputigena,The treponeme data are not directly comparable to the Eubacterium saburreum, F. nucleatum, Selenomonas sputi-

nontreponeme data. Since selective media were used to gena, Selenomonas D01, Selenomonas D04, and Seleno-

TABLE 9. Incidence of mycoplasma and treponemes as percent of subgingival samples that were positiveIncidences as % of subgingival samples that were positive in:

Taxon Children Adults

GI = 0 GI = 1 GI = 2 GI=oa GI = la GI = 2a Hlb ModC Sevd

CulturalMycoplasma 0 0 0 0 0 3 0 16 55Treponema denticola 0 0 0 0 0 0 0 19 21Treponema pectinovorume 0 0 0 0 5 3 0 9 12Treponema socranskiif 20 28 41 17 24 50 27 62 60Treponema vincentii 0 0 0 0 0 0 0 4 0Treponeme C 0 0 0 0 2 6 0 0 0Treponeme D 0 0 0 0 0 3 0 11 17Treponeme E 0 0 0 0 2 0 0 2 0Treponeme F 0 0 0 0 0 3 0 0 2Treponeme G 0 0 0 0 0 3 0 3 0Treponeme L 0 0 0 0 0 3 0 1 2Treponeme M 0 0 0 0 2 0 0 0 0Treponeme N 0 0 0 0 2 0 0 0 2Treponeme P 0 0 0 0 0 3 0 0 0Treponeme R 0 0 0 0 2 0 0 0 0Treponeme S 0 0 0 0 0 3 0 0 0

MicroscopicLarge treponeme 0 0 0 0 0 0 0 11 64Treponemes observed 6 5 12 6 5 17 0 67 88

No. of samples 46 39 17 18 42 36 22 90 42a Data from reference 12.bData from periodontally healthy (Hl) adults studied as control subjects in references 11 to 13.C Data from reference 11. Mod, Moderate periodontitis.d Data from reference 13. Sev, Severe periodontitis.' Data from reference 17.f R. M. Smibert, J. L. Johnson, and R. R. Ranney, submitted for publication.

VOL. 46, 1984

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from

6 MOORE ET AL.

TABLE 10. Taxa that increased with increasing GI scores in bothchildren and adultsa

GITaxon

0 1 2Fusobacterium nucleatum 3.1 3.5 5.5Actinomyces WVa 963 3.2 3.2 4.5Selenomonas D04 0.5 2.1 2.2Bacteroides oris 0.1 0.6 1.4Eubacterium timidum 0.2 0.2 0.6Leptotrichia D22 0.1 0.2 0.4Selenomonas D06 0.2 0.3 0.4Bacteroides D-19 0.0 0.1 0.3FMNRD11 0.0 0.04 0.3FPCD40 0.0 0.04 0.2Bacteroides pneumosintes 0.05 0.11 0.13

a As percent of 2,011, 2,695, and 1,546 isolates from sites with GI scores of0, 1, 2, respectively, from the present study and reference 12.

monas D06. Of the frequently detected nonspirochetal spe-cies most closely correlated with developing gingivitis inadults (12), only Actinomyces WVA 963, F. nucleatum, andSelenomonas D04 appear in the above list.The three major species of the genus Capnocytophaga and

several of the most numerous anaerobic vibrios increased infrequency with increasing GI scores in children, which mayindicate that floras with larger proportions of these gram-negative species are somewhat less potent agents of gingivi-tis production than are the more gram-positive floras associ-ated with the more rapidly developing gingivitis in adults.Capnocytophaga species and anaerobic vibrios do not in-crease further in frequency in proportion in periodontitis(Table 8), which tends to diminish their potential significanceas periodontal pathogens and is consistent with the very lowpropensity of normal children to develop periodontitis.

If the floras in children had not been significantly differentfrom those in adults, we could have determined that thegreater resistance of children to experimental gingivitisreflects only host resistance. Since the incidence of somepredominant species was found to correlate with increasingGI scores in both children and adults, host resistance stillcould play the greater role, or the causative agents could bedifferent in the two populations.

ACKNOWLEDGMENTSWe thank Pauletta C. Atkins, Sandra S. Brown, Polly H. Cooper,

Luba S. Fabrycky, Leesa R. Miller, and Linda D. Stover forbacteriological analyses; Linda L. Long for chromatographic analy-ses; Karen S. Loferski for electrophoretic analyses; Ruth Z. Beyerfor data management; Linda B. Cook for glassware preparation; andKimberly Hardison and Felicia Caine for dental prophylaxis andsubject management.

This work was supported by Public Health Service grants DE-05054 from the National Institute of Dental Research and AI-15244from the National Institute of Allergy and Infectious Diseases andby project 2025790 from the Commonwealth of Virginia.

LITERATURE CITED1. Good, I. J. 1953. The population frequencies of species and the

estimation of population parameters. Biometrika 40:237-264.2. Holdeman, L. V., E. P. Cato, and W. E. C. Moore (ed.). 1977.

Anaerobe laboratory manual, 4th ed. Virginia Polytechnic Insti-tute and State University, Blacksburg.

3. Loe, H., and J. Silness. 1963. Periodontal disease in pregnancy.I. Prevalence and severity. Acta Odontol. Scand. 21:533-551.

4. Loe, H., E. Theilade, and S. B. Jensen. 1965. Experimentalgingivitis in man. J. Periodontol. 36:177-187.

5. Longhurst, P., R. Gillett, and N. W. Johnson. 1980. Electronmicroscope quantitation of inflammatory infiltrates in childhoodgingivitis. J. Periodontal. Res. 15:255-266.

6. Longhurst, P., N. W. Johnson, and R. M. Hopps. 1977. Differ-ences in lymphocyte and plasma cell densities in inflammedgingiva from adults and young children. J. Periodontol. 48:705-710.

7. Mackler, S. B., and J. J. Crawford. 1973. Plaque developmentand gingivitis in the primary dentition. J. Periodontol. 44:18-24.

8. Mashimo, P. A., Y. Yamamoto, J. Slots, B. H. Park, and R. J.Genco. 1983. The periodontal microflora of juvenile diabetics.Culture, immunofluorescence, and serum antibody studies. J.Periodontol. 54:420-430.

9. Mattson, L. 1978. Development of gingivitis in pre-schoolchildren and young adults. J. Clin. Periodontol. 5:24-38.

10. Moore, W. E. C., D. E. Hash, L. V. Holdeman, and E. P. Cato.1980. Polyacrylamide slab gel electrophoresis of soluble pro-teins for studies of bacterial floras. Appl. Environ. Microbiol.39:900-907.

11. Moore, W. E. C., L. V. Holdeman, E. P. Cato, R. M. Smibert,J. A. Burmeister, and R. R. Ranney. 1983. Bacteriology ofmoderate (chronic) periodontitis in mature adult humans. In-fect. Immun. 42:510-515.

12. Moore, W. E. C., L. V. Holdeman, R. M. Smibert, I. J. Good,J. A. Burmeister, K. G. Palcanis, and R. R. Ranney. 1982.Bacteriology of experimental gingivitis in young adult humans.Infect. Immun. 38:651-667.

13. Moore, W. E. C., L. V. Holdeman, R. M. Smibert, D. E. Hash,J. A. Burmeister, and R. R. Ranney. 1982. Bacteriology ofsevere periodontitis in young adult humans. Infect. Immun.38:1137-1148.

14. Page, R. C., and H. E. Schroeder. 1976. Pathogenesis ofinflammatory periodontal disease. Lab. Invest. 33:235-255.

15. Seymour, G. J., M. S. Crouch, R. N. Powell, D. Brooks, I.Beckman, H. Zola, J. Bradley, and G. F. Burns. 1982. Theidentification of lymphoid cell subpopulations in sections ofhuman lymphoid tissue and gingivitis in children using monoclo-nal antibodies. J. Periodontal Res. 17:247-256.

16. Silness, J., and H. Loe. 1964. Periodontal disease in pregnancy.II. Correlation between oral hygiene and periodontal condition.Acta Odontol. Scand. 22:112-135.

17. Smibert, R. M., and J. A. Burmeister. 1983. Treponema pectino-vorum sp. nov. isolated from humans with periodontitis. Int. J.Syst. Bacteriol. 33:852-856.

18. Smith, L. V., L. B. Golub, and D. F. Duperon. 1974. Anevaluation of crevicular fluid and gingival tissue in children. J.Dent. Child. 41:128-132.

19. Tanner, A. C. R., M. A. Listgarten, and J. L. Ebersole. 1984.Wolinella curva sp. nov.: "Vibrio succinogenes" of humanorigin. Int. J. Syst. Bacteriol. 34:275-282.

INFECT. IMMUN.

on January 9, 2020 by guesthttp://iai.asm

.org/D

ownloaded from