Oral Microbiol Immunot 1992: 7: 244-248
ActinobaeiUus actinomycetemcomitans, Capnoeytophaga and Porphyromonas gingivaiis in subgingival plaque of adolescents with Down's syndrome
M. Barr-Agholme\ G. Dahliof, L. Linder', T. Modeer' Departments of 'Pedodontics and ^Oral Microbiology, Faculty of Odontology, Karolinska institute. Huddinge, Sweden
Bair-Agtwtme M, Dahttof G. Linder L. Modeer T. Actinobacillus actinotnycetenicotnitans, Capnoeytophaga rt/;t/Porphyromonas gingivalis in subgingival plaque of adolescents with Down's syndrome. Oral Microbiol Immunot 1992: 7: 244-248. Levels of Aetinobaeittus actinomycetemeomitans, Capnocytoptiaga and Porphyromonas gingivalis were determitied in subgingival plaque satnples frotn 37 adolescents with Down's syndrome and 37 healthy controls tnatched with respect to age and sex. Gingival infiatntnation, supra- and subgingival calculus, periodontai pockets ( > 4 tntn) and alveolar bone loss were registered. Alveolar bone loss was tnore frequent in Down's syndrotne subjects (32"/.) than in the controls (3%). A. actinomycetemcomitans was detected in the subgingival plaque in 35'/o of the Down's syndrotne adolescents and in 5"/i of the controls. On site level, A. actinomycetemcomitans and Capnoeytophaga were tnore frequent in the subgingival plaque satnples of Down's syndrotne children than in those of controls. Cotnparing Down's syndrotne subjects positive or negative for A. aetinomyeetemeomitans and Capnoeytophaga, no significant differenees were foutid in tertns of gingival infiatntnation, periodontai pockets ( > 4 mm) or number of sites with alveolar bone loss. The results indicate an altered tnicrobial composition of the subgingival plaque of Down's syndrotne subjects cotnpared with healthy controls, with higher frequency of A. actinomycetemcomitans.
Down's syndrotne is an autosotnal chrotnosotnal anotnaly resulting frotn trisotny of the 2Ist chrotnosotne (15). Down's syndrotne children are more suseeptible than normal children to respiratory tract infections (35, 37) and have a high prevalence of periodontai disease (6, 10, 20, 29). Signs of alveolar bone loss, tnainly localized to the tnandibular anterior regioti, are seen as early as 11 years of age (20). Abnormalities in host response sueh as reduced chetnotaxis (7, 14), impaired phagocytosis (8) as well as an altered itntnune response may be important eontributing factors to the high prevalence of periodontai disease in Down's syndrotne (26, 27). In adolescents with juvenile periodotititis, Aetinobaeiltus aetinomyeetem-
eomitans is present in the subgingival microfiora in 41-90% (3, 30, 36), compared with 4-20% (2, 4, 30) in nondiseased adolescents. Capnoeytophaga has also been itnplieated in the pathogenesis of juvenile periodontitis (25, 28), but there are confiicting reports (13, 22). Of the black-pigmented anaerobic rod species, Porptiyromonas gingivatis (formerly Bacteroides gingivati.s) is associated with loss of attachtnent in adult periodonfitis (9, 16, 32). According to Meskin et al. (19), there is a significantly higher number of black-pigtnented anaerobic rods in supragingival plaque frotn Down's syndrotne ehildren than in non-gingivitis controls. However, Cutress et al. (II) found no differenee between tnentally
Key words: Down's syndrome: ActinobaeiUus actinomycetemcomitans) Capnocytoptiaga) Porptiyromonas gingivaiis] periodontai disease M. Barr-Agholme. Department of Pedodontics. Faculty of Odontology, Karolinska Institutet, Box 4064, S-141 04 Huddinge, Sweden Accepted for publication February 7. 1992
retarded patients, including Down's syndrotne and nortnal subjects, with respeet to the occurrenee of blaek-pigmented anaerobic rods in the supragingival plaque. The composition of the subgingival microfiora in Down's sytidrotne subjects has not been detertnined. The purpose of this study was therefore to determine the occurrence of A. aetinomyeetemeomitcms, Capnoeytophaga and P. gingivalis in the subgingival plaque of adolescents with Down's syndrotne. iVIateriai and methods
The subjects were selected, on the basis of cooperation, from a larger Down's syndrotne material described earlier (20)
A. actinotnycetetncomitans /// Down's .syndrome and comprised 20 boys and 17 girls aged 9-21 years living in the Stockhohn area. The mean age was 16.3 (SD + 3.4) years. The subjeets in the control group (« = 37), randotnly chosen to tnatch Down's syndrotne children with respect to age and sex, received their regular dental treattnent at the Department of Pedodontics.
the cetnentoenatnel junction to the alveolar bone crest on the radiographs exceeded 2.0 mtn on the tnesial and distal surfaces of fully erupted eentral incisors and first tnolars. Because of the discrepant craniofacial developmetit in Down's syndrotne patients (20), the distal surfaces of tnaxillary central incisors were excluded.
Interview
Microbiologicai anaiysis
The parents answered a questionnaire on the children's tnedical condition, disposition to infections and whether the child had undergone any antibiotic treattnent 3 tnonths before the clinical and tnicrobiological examination. One Down's syndrotne subject had cotnpleted a course of phenoxytnethylpenicillin one week before satnpling and one was being treated with oxytetracycline at the titne of exatnination.
Supragingival plaque was retnoved frotn eaeh satnpling site using eurettes and cotton pellets. Subgingival plaque was then collected by inserting a fine paperpoint to the bottotn of the sulcus. After 20 s, the paperpoint was retnoved atid itntnediately tratisferred to a screwcapped vial containing VMGA III transport tnedium (24). The satnple was processed in the laboratory within 2 h. Subgingival satnples were obtained frotn the tnesial sites on all first tnolars and central incisors (8 sites per individual) in all 592 satnples. Satnples diluted 10"- or undiluted wete cultured on TSBV mediutn (Trypticase soy agar with 10% serum, 75 /;g/tnl baeitracin and 5 /;g/tnl vancomycin) (31) and incubated for 3-5 days in 5% CO,. Isolates identified as A. aetinomyeetemeomitans were gratn-negative, eatalase-positive coccobacilli that exhibited stnall, convex, adherent colonies on TSBV-agar plates. Samples diluted 10"--10 •* were cultured on Brueella agar (BBL Microbiology Systetns, Cockeysville, MD) with 5% horse blood and tnenadinone (2.5 /;g/tnl) (Becton Dickinson, Cockeysville, MD) added. The agar tnedia were incubated at 37' C for 10 days in anaerobic jars (Gas Park, BBL Microbiology Systetns). Colonies of P. gingivalis atnong the black-pigmented anaerobic rods were differentiated as black or greenish-black colonies not
Gingivai condition
Gingival infiammation was assessed by bleeding on probing at 6 points around each tooth, according to the gingival bleeding index (5). The probing was performed by sweeping the probe aroutid the tooth. The percentage of surfaces with gingivitis was estitnated for each individual. Supragingival calculus was recorded when clearly visible. Probing deptii
The pocket depth was tneasured to the nearest millitneter with a graduated periodotital probe on tnesial and distal sites on all first tnolars and central ineisors. Radiographic examination
Two bitewings and two periapical radiographs were taken using a standardized, long cotie paralleling technique to examine the tnaxillary and tnandibular first tnolars and central itieisors. Subgitigival calculus was recorded when elearly discernible proximally on the radiographs. Aiveolar bone loss
The radiographs were exatnined usitig a peak scale loupe at 7-fold tnagnifieation, which permits measuretnents to the nearest 0.1 tntn. Alveolar bone loss was recorded when the distance frotn
245
demonstrating lotig-wave ultraviolet light tluoreseence (23). For isolation of Capnoeytophaga, samples diluted 10 ' or undiluted were cultured on BKV tnediutn (blood, kanatnycin and vancotnyciti) (34) for 3-4 days in 5% CO,. Capiiocvtoptiaga was identified as gratn-negative rods with "gliding tnotility" oti the agar surface. In each satnple, the total viable counts (numbers of eolonyfortning utiits/tnl transport tnediutn) were detertnined on Brucella blood agar. The nutnber of /i. actinomycetemcomitans, Capnoeytophaga and P. gingivatis were expressed as percentages of the total colony counts. All tnicrobiological detertninations were perfortned at the departtnent of Oral Microbiology, Karolinska Institute, Huddinge, Sweden.
Statistics
The chi-square test was used for statistical analyses of the cotnparative frequeneies between groups and Student's /-test (two-tailed) was applied to analysis of the variable Capnoevtophuga in relation to respiratory infeetion. Results Interview
Approximately half of the Down's syndrotne subjects reported a highly inereased frequency of infections; 38% had 10 or tnore infections per year, tnostly of the upper respiratory tract, and were therefore classified as disposed to infeetions. Five percent had congenital cardiac detcets (a septutn defect or a tnurtnur). Other tnedical conditions noted atnong the Down's syndrotne subjeets were; one with thyroid dysfunction, I diabetic and 2 with dertnatological conditions (such as skin boils). Of the control subjects, one showed a high
Tatite 1. Clinical data of the patients studied Control (/; = 37)
Down 's syndrome (;; = 37)
%
Variables
/;
"A:
Clinical variables Gingival bleeding index 0-25% Gingival bleeding index 25-5t)'/'n Gingival bleeding index > 50% Periodontai pockets ( > 4 tntn) Supragingival calculus
9 21 7 16 9
24 57 19 43 24
29 8 0 3 4
78*** 22 NS 0* 8** 11 NS
Radiographic variables Alveolar bone loss Subgingival calculus
12 ?
32 14
1 0
3** 0 NS
Chi-square: * P 4 tntn), calculus and alveolar bone loss is presented in Table 1. Periodontai pockets (> 4 mm) were tnore frequent in Down's syndrotne subjects than in the controls (P
ActinobaeiUus actinomycetemcomitans, Capnoeytophaga and Porphyromonas gingivaiis in subgingival plaque of adolescents with Down's syndrome
M. Barr-Agholme\ G. Dahliof, L. Linder', T. Modeer' Departments of 'Pedodontics and ^Oral Microbiology, Faculty of Odontology, Karolinska institute. Huddinge, Sweden
Bair-Agtwtme M, Dahttof G. Linder L. Modeer T. Actinobacillus actinotnycetenicotnitans, Capnoeytophaga rt/;t/Porphyromonas gingivalis in subgingival plaque of adolescents with Down's syndrome. Oral Microbiol Immunot 1992: 7: 244-248. Levels of Aetinobaeittus actinomycetemeomitans, Capnocytoptiaga and Porphyromonas gingivalis were determitied in subgingival plaque satnples frotn 37 adolescents with Down's syndrome and 37 healthy controls tnatched with respect to age and sex. Gingival infiatntnation, supra- and subgingival calculus, periodontai pockets ( > 4 tntn) and alveolar bone loss were registered. Alveolar bone loss was tnore frequent in Down's syndrotne subjects (32"/.) than in the controls (3%). A. actinomycetemcomitans was detected in the subgingival plaque in 35'/o of the Down's syndrotne adolescents and in 5"/i of the controls. On site level, A. actinomycetemcomitans and Capnoeytophaga were tnore frequent in the subgingival plaque satnples of Down's syndrotne children than in those of controls. Cotnparing Down's syndrotne subjects positive or negative for A. aetinomyeetemeomitans and Capnoeytophaga, no significant differenees were foutid in tertns of gingival infiatntnation, periodontai pockets ( > 4 mm) or number of sites with alveolar bone loss. The results indicate an altered tnicrobial composition of the subgingival plaque of Down's syndrotne subjects cotnpared with healthy controls, with higher frequency of A. actinomycetemcomitans.
Down's syndrotne is an autosotnal chrotnosotnal anotnaly resulting frotn trisotny of the 2Ist chrotnosotne (15). Down's syndrotne children are more suseeptible than normal children to respiratory tract infections (35, 37) and have a high prevalence of periodontai disease (6, 10, 20, 29). Signs of alveolar bone loss, tnainly localized to the tnandibular anterior regioti, are seen as early as 11 years of age (20). Abnormalities in host response sueh as reduced chetnotaxis (7, 14), impaired phagocytosis (8) as well as an altered itntnune response may be important eontributing factors to the high prevalence of periodontai disease in Down's syndrotne (26, 27). In adolescents with juvenile periodotititis, Aetinobaeiltus aetinomyeetem-
eomitans is present in the subgingival microfiora in 41-90% (3, 30, 36), compared with 4-20% (2, 4, 30) in nondiseased adolescents. Capnoeytophaga has also been itnplieated in the pathogenesis of juvenile periodontitis (25, 28), but there are confiicting reports (13, 22). Of the black-pigmented anaerobic rod species, Porptiyromonas gingivatis (formerly Bacteroides gingivati.s) is associated with loss of attachtnent in adult periodonfitis (9, 16, 32). According to Meskin et al. (19), there is a significantly higher number of black-pigtnented anaerobic rods in supragingival plaque frotn Down's syndrotne ehildren than in non-gingivitis controls. However, Cutress et al. (II) found no differenee between tnentally
Key words: Down's syndrome: ActinobaeiUus actinomycetemcomitans) Capnocytoptiaga) Porptiyromonas gingivaiis] periodontai disease M. Barr-Agholme. Department of Pedodontics. Faculty of Odontology, Karolinska Institutet, Box 4064, S-141 04 Huddinge, Sweden Accepted for publication February 7. 1992
retarded patients, including Down's syndrotne and nortnal subjects, with respeet to the occurrenee of blaek-pigmented anaerobic rods in the supragingival plaque. The composition of the subgingival microfiora in Down's sytidrotne subjects has not been detertnined. The purpose of this study was therefore to determine the occurrence of A. aetinomyeetemeomitcms, Capnoeytophaga and P. gingivalis in the subgingival plaque of adolescents with Down's syndrotne. iVIateriai and methods
The subjects were selected, on the basis of cooperation, from a larger Down's syndrotne material described earlier (20)
A. actinotnycetetncomitans /// Down's .syndrome and comprised 20 boys and 17 girls aged 9-21 years living in the Stockhohn area. The mean age was 16.3 (SD + 3.4) years. The subjeets in the control group (« = 37), randotnly chosen to tnatch Down's syndrotne children with respect to age and sex, received their regular dental treattnent at the Department of Pedodontics.
the cetnentoenatnel junction to the alveolar bone crest on the radiographs exceeded 2.0 mtn on the tnesial and distal surfaces of fully erupted eentral incisors and first tnolars. Because of the discrepant craniofacial developmetit in Down's syndrotne patients (20), the distal surfaces of tnaxillary central incisors were excluded.
Interview
Microbiologicai anaiysis
The parents answered a questionnaire on the children's tnedical condition, disposition to infections and whether the child had undergone any antibiotic treattnent 3 tnonths before the clinical and tnicrobiological examination. One Down's syndrotne subject had cotnpleted a course of phenoxytnethylpenicillin one week before satnpling and one was being treated with oxytetracycline at the titne of exatnination.
Supragingival plaque was retnoved frotn eaeh satnpling site using eurettes and cotton pellets. Subgingival plaque was then collected by inserting a fine paperpoint to the bottotn of the sulcus. After 20 s, the paperpoint was retnoved atid itntnediately tratisferred to a screwcapped vial containing VMGA III transport tnedium (24). The satnple was processed in the laboratory within 2 h. Subgingival satnples were obtained frotn the tnesial sites on all first tnolars and central incisors (8 sites per individual) in all 592 satnples. Satnples diluted 10"- or undiluted wete cultured on TSBV mediutn (Trypticase soy agar with 10% serum, 75 /;g/tnl baeitracin and 5 /;g/tnl vancomycin) (31) and incubated for 3-5 days in 5% CO,. Isolates identified as A. aetinomyeetemeomitans were gratn-negative, eatalase-positive coccobacilli that exhibited stnall, convex, adherent colonies on TSBV-agar plates. Samples diluted 10"--10 •* were cultured on Brueella agar (BBL Microbiology Systetns, Cockeysville, MD) with 5% horse blood and tnenadinone (2.5 /;g/tnl) (Becton Dickinson, Cockeysville, MD) added. The agar tnedia were incubated at 37' C for 10 days in anaerobic jars (Gas Park, BBL Microbiology Systetns). Colonies of P. gingivalis atnong the black-pigmented anaerobic rods were differentiated as black or greenish-black colonies not
Gingivai condition
Gingival infiammation was assessed by bleeding on probing at 6 points around each tooth, according to the gingival bleeding index (5). The probing was performed by sweeping the probe aroutid the tooth. The percentage of surfaces with gingivitis was estitnated for each individual. Supragingival calculus was recorded when clearly visible. Probing deptii
The pocket depth was tneasured to the nearest millitneter with a graduated periodotital probe on tnesial and distal sites on all first tnolars and central ineisors. Radiographic examination
Two bitewings and two periapical radiographs were taken using a standardized, long cotie paralleling technique to examine the tnaxillary and tnandibular first tnolars and central itieisors. Subgitigival calculus was recorded when elearly discernible proximally on the radiographs. Aiveolar bone loss
The radiographs were exatnined usitig a peak scale loupe at 7-fold tnagnifieation, which permits measuretnents to the nearest 0.1 tntn. Alveolar bone loss was recorded when the distance frotn
245
demonstrating lotig-wave ultraviolet light tluoreseence (23). For isolation of Capnoeytophaga, samples diluted 10 ' or undiluted were cultured on BKV tnediutn (blood, kanatnycin and vancotnyciti) (34) for 3-4 days in 5% CO,. Capiiocvtoptiaga was identified as gratn-negative rods with "gliding tnotility" oti the agar surface. In each satnple, the total viable counts (numbers of eolonyfortning utiits/tnl transport tnediutn) were detertnined on Brucella blood agar. The nutnber of /i. actinomycetemcomitans, Capnoeytophaga and P. gingivatis were expressed as percentages of the total colony counts. All tnicrobiological detertninations were perfortned at the departtnent of Oral Microbiology, Karolinska Institute, Huddinge, Sweden.
Statistics
The chi-square test was used for statistical analyses of the cotnparative frequeneies between groups and Student's /-test (two-tailed) was applied to analysis of the variable Capnoevtophuga in relation to respiratory infeetion. Results Interview
Approximately half of the Down's syndrotne subjects reported a highly inereased frequency of infections; 38% had 10 or tnore infections per year, tnostly of the upper respiratory tract, and were therefore classified as disposed to infeetions. Five percent had congenital cardiac detcets (a septutn defect or a tnurtnur). Other tnedical conditions noted atnong the Down's syndrotne subjeets were; one with thyroid dysfunction, I diabetic and 2 with dertnatological conditions (such as skin boils). Of the control subjects, one showed a high
Tatite 1. Clinical data of the patients studied Control (/; = 37)
Down 's syndrome (;; = 37)
%
Variables
/;
"A:
Clinical variables Gingival bleeding index 0-25% Gingival bleeding index 25-5t)'/'n Gingival bleeding index > 50% Periodontai pockets ( > 4 tntn) Supragingival calculus
9 21 7 16 9
24 57 19 43 24
29 8 0 3 4
78*** 22 NS 0* 8** 11 NS
Radiographic variables Alveolar bone loss Subgingival calculus
12 ?
32 14
1 0
3** 0 NS
Chi-square: * P 4 tntn), calculus and alveolar bone loss is presented in Table 1. Periodontai pockets (> 4 mm) were tnore frequent in Down's syndrotne subjects than in the controls (P
