Jpn. J. Infect. Dis., 54, 2001

Laboratory and Epidemiology Communications

Isolation of Opportunistic Pathogens in Dental Plaque, Saliva andTonsil Samples from Elderly
Mohamad Abdus Salam, Hidenobu Senpuku* , Ybshiaki Nomura,
Kbaiml Matin, Hideo Miyazakil and Nobuhiro Hanada
Department of Oral Science, National Institute ofInfectious Diseases,Toyama I‑23‑1, Shinjuku‑ku, Tobo 162‑8640 andJDivision of Preventive Dentistyy, Department of Oral Health Science.

Graduate School ofMedical and Dental Science, Niigata UniversiO!,
Gakkocho‑dori 2‑52 74, Niigata 951‑8514
Communicated by Nobuhiro Hanada(Accepted November 1 6, 200 1)

The oral cavlty lS a potential reservoir or pathogenic

forth with a cotton swab (SEEDSWAB No. 1 : Eiken Chemical

microorganisms (I‑3). In the elderly, oral hygiene is poor (4)Col, Ltd･ lEiken], Tokyo)･ In the case of subjectsusi.ng fulland dental biofllm contalnlng OPPOrtunistic pathogens forms

dentures, Samples were collected from the same regions Of

on tooth surfaces (4‑6)･ Oral pathogeでs are risk factors forthe dentures aRer removalfrom the oral cavity. From subjects

bacterial pneumonia as they may be asplrated into the resplra‑

not having any of the above一mentioned teeth, samples weretory tract (6,7). Pathogenic bacteria in the biofilm in the dental

obtained from the opposite teeth. The subjects were asked toplaque might also be released to colonize new surfaces, e.g.,chew paraffingum for 3 min to stimulate secretion of saliva,

the tonsils, pharynx, and resplratOry tract. To test thesewhich was collected into ice‑chilled sterile bottles uslng apossibilities, we studiedthe isolation frequencies of opportun‑

cotton swab. Samples from the tonsils were also collected

istic bacteria on tooth and tonsil surfaces, and in saliva in theOne hundred twenty‑five elderly people (mean age: 73.0

uslng a cotton swab. All specimens were transferred into I
ml of reduced transport nuid medium (0.4% agar, 0.15%
thioglycollate/phosphate buffered saline) in sterile bottles.on± 0･3 years old, 66 males and 59 females) from Niigata City

ice and processed within one night of collection. The entlre

in Japan partlClpated in this study, which was conducted insamples in tubes were inoculated onto chocolate agar, blood

July, 2000. None of the subjects was hospitalized or institu‑agar, OPA staphylococcus, and drigalski agar plates (NipponBecton Dickins?n Co･, Ltd･, Tokyo) with an aid ofa stick･
The plates were lnCubated in an atmosphere of 5% CO2 in H2
at 37oC for 24‑48 h. Representative microbial colomies fromelderly.

tionalized, and all came to the examination center either bythemselves or accompanied by their family. They were in good
general health and did not requlre SPeCial assistance for their

daily activities. Informed consent was obtained from each

each plate were gram stained and identi丘ed in tens of theirsubject pr10r tO the study. Ethical clearance for the study's

characteristic appearance, hemolytic, catalytic reaction, andmethods used was obtained from the Ethics Committee ofoxidase reaction (8). Colonies of microbes suspected to be

the Faculty of Dentistry, Niigata University.

those responsible for pneumonia and which were found in a

majorityofthe subjects were suspended in 1 ml ofO.5% saline
and tested by using diagnosis kits indicated below (9). We

SupraglnglVal plaque was collected from the upper rightsecond premolar and first molar teeth by swabbing back andidentified Staphylococcus aureus, both methicillin sensitive*Corresponding author: Tel: +8 1‑315285‑1 I 1 1 , Fax: +81 ‑3‑52851(MSSA) and resistant (MRSA) strains, by using PS latex,
rabbit plasma, and MRSA screening plates (Nippon Becton

1 172, E‑mail: hsenpuku@nih.gojp

193

Jpn. J. Infect. Dis., 54, 2001Dickinson); Pseudomonas sp. by using VITEK (BioMerieux

albicans, Enterobacteriaceae, Pseudomonas sp., S. aureus,Vitek Japan lBVJ], Tokyo); 8‑hemolytic streptococcus byXanthomonas maltophilia, Klebsiella pneumoniae, S･using a Seroidenstrepto kit (Eiken), Apュ strepto (BVJ), andmarcescens, coagulase‑negative staphylococci (CNS) and K.

VITEK; StTlePtOCOCCuS PneumOniae byuslng a Streptococcus
identification disk (Nippon Becton Dickinson); Haemophilus

Owtoca were most Bequently isolatedfrom these specimens.
The higher prevalence of these microorganisms in elderlyinjluenzae byusing a Haemophilus ID4 plate (Nippon BectonDickinson); Serratia mayICeSCenS by using VITEK; andsubjects indicates that this age population is at greaterrisk ofdeveloplng Systemic diseases such as pneumonia and heartdisease (6). Near 80% of the opportumistic pathogens isolated
from dental plaque were found in saliva or tonsils, while 63‑
64% of the microbes present in saliva were present in dental

Candida sp. by using Candida check (Iatron Laboratories lnc･,Tbkyo).

The isolationfrequencies of microorgamisms kom retrieved
from dental plaque, saliva, and tonsils, as determined uslng
the manufacturers'instruction, are shown in Table 1. C.

plaques or tonsils (Table 2)･ This may indicate that theseorganisms might be releasedfrom tooth surfaces into saliva

Table I. Isolation ficqucncy ofmicrobial pathogcns in dental plaque, saliva, and tonsil samples&om older adults
DcntalMicroorganisms

n=125

plaquenNo.

(%)≡Sa一iva

125

Tonsi
n

No.

(%)

ls=125No.

(%)

20 (16)

EnEerobacter cloacae13(ll)En terobacter aey10geneS2 (2)

3 (3)

6(5)

En terobacter sazakit'

0 (0)

2 (2)

0 (0)

EnteylDbacter sp.

5 (4)

7 (6)

7 (6)

En Eey10bacter agglom eyms

4 (3)

7 (6)

4(3)

En terococcus faecalis

4 (3)

5 (4)

9(8)

Enterococcus sp.

0 (0)

2 (2)

1(1)

Escherichia cotz'

I(I)

4 (3)

2 (2)

23 (19)El.kenelLa corndens

I(I)

0 (0)

0 (0)

Klebsiella pneumoniae

9 (7)29 (23)13(ll)

KJebsiella oqEoca

4 (3)

6 (5)

6 (5)

KIebsl'ella ozaenae

0 (0)0 (0)1(1)

Kluyvera sp.

1(1)

3 (2)

1(1)

Candida albicans

18(14)

21 (17)1(1)Candida pa rapsilosis

0 (0)

1(1)

I(I)

Candida EylDPica lis

0 (0)

I(I)

0 (0)

CorynebacEerium sp.

7 (6)

1(1)

2 (2)

CiEy10bacter.Peundii

3 (3)

3 (3)

I(1)

Comamonas acidovoran

3 (3)

I(1)

1(1)

Pseudomonas sp.

5 (4)

3 (3)

5 (4)

5 (4)

Pseudomonas.Puwescens

3 (3)

6 (5)

Pseudomon as putida

1(1)

3 (3)

5 (4)

Pseudomonas aenLglnOSa

I(1)

I(I)

0 (0)

Pseudomonas cepacl'a

1(I)

0 (0)

0 (0)

Ftavobacten'um sp.

0 (0)

1(1)

0 (0)

Flavobacterium indolgens

10(8)

6(5)

5 (4)

Flavobacterium m eningosepLicum

0 (0)

0 (0)

I(1)

Staphylococcus aureziS (MSSA)

3 (3)

5 (4)

13(ll)

Staphylococcus aweus (MRSA)

5 (4)

7 (6)

12 (10)

Staphylococcus aureus (CN S)

6 (5)

3 (3)

2 (2)

StylePtOCOCCuS ago lactiae

0 (0)

4 (3)

4 (3)

α‑hemolytic streptococcus

106 (85)111 (88)

lot (81)

B ‑hcmolytic streptococcus

0 (0)

1 (2)

0 (0)

β‑hemolytic streptococcus non group A

0 (0)

I (2)

0 (0)

γ‑hemolytic s什eptococcus

66 (53)74 (58)77 (61)Neisseria sp.

67 (54)85 (68)78 (63)

AcinetobacEer SP.

3 (3)

5 (4)

1(1)

A cin eEobacEer ca lcoacetl'cus

3 (3)

10(8)

5 (4)

A clln etobacter lwoj7iL'

1(1)

2 (2)

2 (2)

A Ica ligenes xylosoxydans

I(1)

2 (2)

1(I)

A Ica Ligenes faeca lis

0 (0)

1(1)

0 (0)

Serratia m ayICeSCenS

3 (3)4(3)4(3)

Serra tia liquefaciens

4(3)

6 (5)

6 (5)

Leclericia adeca rboxylata

1(1)

2 (2)

0 (0)

flaem oph iLus papa l'Iがuenzae

2 (2)

4 (3)

2 (2)

Edwardsiella sp.

0 (0)

1(1)

0 (0)

Morxella sp.

0 (0)

1(1)

0 (0)

Branhamella caEarrhalis

0 (0)

1(1)

Xanthomonas maLEophilia

5194(4)

5(4)

1(1)
4(3)

Jpn. J. Infect. Dis., 54, 2001
Table 2. Agreement rates of opportunistic pathogens isolated

C. D., Depaola, L G. and Suzuki, ∫. B. (1986): Oralfrom dental plaque, saliva, and tonsil samples
DentalplaqueDental plaque
SalivaTonsi1

Sativasuccesion gram‑negative bacilli in myelosuppressed
Tわnsils

79.8 ± 26.7*
63.7 ± 27,4

71.8 ±27.2

cancer patients. ∫. Clin. Microbio1., 24, 210‑213.

78.0 ± 27.3

4. Simons, D., Kidd, E. A. and Beighton, D. (1999): Oral

64.5 ± 26.7

health of elderly occupants in residential homes. Lancet,354, 515.

75.9 ±27.5

5. Scammpieco, F. A. and Mylotte, J. M. (1996): Relationship

Agreement rate: Number of opportunistlC Pathogens which was identicalbetween periodontal disease and bacterial pneumonia. J.

in bothsaliva and dentalplaque, divided by the number ofopportunistic

Periodonto1., 67, ll 14‑1 122.

pathogcns isolated from dental plaque (saliva/dental plaque).

*: P <0.05, saliva/dental plaque vs dental plaque/saliva.

6. Ybneyama, T., Yoshida, M., Matsui, T. and Sasaki, H.
and the Oral Care Working Group (1 999): Oral careandpneumonia. Lancet, 354, 5 15.

and then colonize on oral cavitysurfaces such as the tonsils.

7. Sasaki, H., Sekizawa, K., Yanai, M., Arai, H., Yamaya,

Professional oral hygiene to remove pathogens from dental

M. and Ohrui, T. (1997): New strategies fわr aspiration

biofilm may slgnificantly reduce the risk of systemic diseases

pneumonia. Int. Med., 36, 85 1‑855.
8. Murray, R. G. E., Brenner, D. J., Bryant, M. R., Holt, J.
G., Kiieg, N. R., Moulder, J. W., Pfennig, N., Sneath, P.

in elderly people.

H. A., Steley, ∫. T. and Williams, S. T. (1989): Bergey'sREFERENCES

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