An Investigation of the Oral
Mucosal and Periodontal diseases
with Exfoliative Cytology in Tonga
Sisilia Fusi FIFITA, Kayo KUYAMA, Takashi MATSUMOTO*,
Sililo Tomiki**, and Hirotsugu YAMAMOTO
Department of Pathology, Nihon University School of Dentistry at Matsudo*Clinical Pathology, Nihon University School of Dentistry at Matsudo** Department of Dentistry, Vaiola Hospital, Tonga

Objective
The diagnosis of a mucosal lesion cannot be based solelyon clinical findings. So far, there is no reliable method
applicable to the oral cavity that can replace a biopsy, butsome may be used as a supplement. Cultures are considered
the golden standard in microbiological evaluation of dental
diseases but they are expensive and time consuming toperform especially in third world countries. Cytologicalinvestigations of the mucosal problems in Tonga have notbeen previously reported. Therefore, the purpose of this study
was to assess the oral mucosal condition in Tonga, and to
estimate the method “exfoliative cytology”. We will use thisstudy’s finding to update the sparse information on oral
mucosal lesion in Tonga and as an additional aid toformulating additional strategies to enhance better
preventive oral health plan.

Introduction
Tonga, a South Pacific Polynesian island, a member of theCommonwealth of Nations, and with a population of about
104,000, consists of 171 islands (Fig.1). For the past 40 years ormore, Tonga had had a low caries rate but a high prevalenceof periodontal diseases, and other mucosal conditions wererelatively low. It was reported that the prevalence of
microbial plaque, calculus and gingival inflammation wasvery high in all subjects at all ages particularly over 30 years inTonga. A recent survey has shown that periodontal diseaseremains a concern in the adult population, and caries isgradually increasing among children. In Tonga, periodontal
abscess related to periodontitis is also a common clinicalfinding, but all reports were done by macroscopic
observation and no objective method including assessmentof distribution of microflora had been used.

Where isTonga?

† Tonga is*in the South
Pacific
Ocean, a
Polynesianisland nearFiji, Samoaand NZ †.
Fig.1

Materials and methods
1.Subjects
The study was conducted at Vaiola Hospital (main island), where 70 % of thepopulation lives, during 2001 to 2003, with clinical evidence of mucosal (minor) andperiodontal (major) lesions identified during clinical examination. The clinical oral
health status was briefly assessed prior to the delivery of cytological investigation,
and was expressed as satisfactory (good) or unsatisfactory (poor), depending on theclinical findings. A mouth with a lot of debris, calculus and gingival inflammation wasconsidered unsatisfactory.

2. The characteristics of periodontitis, gingivitis and mucosal lesionsThe periodontal check points; 1) periodontal pocket over 5 mm, 2) plaquedeposition, 3) existence of subgingival calculus, and 4) drainage from the pocket.
The site chosen for smearing was severely involved and was performed on the basisof the following check points. One ~ four were confirmed as for periodontitis. Amongthe six points of pocket surroundings (WHO probe), the deepest one was selected forsmearing.
The characteristics of gingivitis were the present or absent of gingival swellingwithout attachment loss and bleeding after probing. Oral mucosal lesions wereassessed as any visible lesions that included; pain, swelling, redness or whiteness,
nodular and rough surface were observed by an ocular inspection and palpation.

3. Exfoliative cytology
In all cases, the cells were exfoliated using Cytobrush (Teikokuzokiseiyaku K.K.,Japan) as shown in Fig. 2 from the pocket epithelium (77.5 %) or the supragingival
alveolar mucosa (22.5 %). The diameter of the cytobrush head was 6mm and its
length without the handle was 2 cm (marked as 0.5, 1.0, 1.5 cm) from the tip. Inorder to collect subgingival smears, the head of the brush was introduced
through the sulcus or pocket orifice as far apically as possible, then the size wasapproximated according to the WHO probe measurement. If it was impossible,supragingival mucosa was selected. The collected cells were smeared onlabeled glass slides.
4. Papanicolaou’s staining
Two glass slides were made for each case. All cytological smears wereimmediately fixed in 95 % ethyl alcohol, and stained by the Papanicolaou
method (Fig. 3). On the basis of the cytomorphological characteristics, thepresence of microorganisms, debris, inflammatory cells and changes ofsquamous cells were microscopically analyzed by a cytologist and 2 dentists. The
microorganisms were identified according to their Papanicolaou’s characteristicmorphology.

Fig. 2: The demonstration of using thecytobrush in the oral cavity of a patientwith a deep periodontal pocket(pocket depth=greater than 5 mm)

Fig. 3: The demonstration of sets of alcoholand xylene use in Papanicolau staining.

Results
1. Clinical findings
Hundred and twenty patients (62 males and 58 females), ranging from 3 to 76years old with a mean age of 32.7±20.9 years (Fig. 4). Regarding the patient’s
chief complaints; sore teeth and swollen gingiva were frequent and clinically foundin all cases. The locations of cytological areas in the oral cavity are outlined inTable 1. Clinical diagnosis was summarized in Table 2. Overall, 73 cases (60.8 %)were diagnosed with acute periodontal abscesses arising from periodontitis, caries
and pericoronitis, and 48.0 % of these (35 cases) had mild extra oral swelling (Fig. 5).
2. Children between the age of 3 and 14 years
In this study, 38 children (23 females and 15 males) were participated and theirage ranged from 3 to 14 years. The mean age was 8.5 ±3.5 years. Sixteen of the 38
children were identified as having one periodontal abscess related to the carious
tooth involved while 18 were identified as having one or more periodontal
abscesses related to many carious teeth, but no detection of any bony destruction.
Among children, 13 presented with mild facial swelling. The percentage of children(n=35) showing oral debris was 92.1 % and plaque was observed in almost all thesechildren. In contrast, the percentages of children showing calculus with caries (n=6),
and gingivitis (n=4) were low at 15.8 % and 6.0 % respectively.

3. Patients with gingivitis
Twenty adults (16.7 %) had mild to moderate gingivitis, bleeding on probing andhad fairly plaque accumulation. Regarding the distribution in the jaws, 8 (40 %) were
found in the lower jaw, and 12 (60 %) in the maxillary region. No gingival abscesses
were found (Fig. 6).
4. Patients with periodontitis
Abscess formation was readily identified clinically in these patients due toconcurrent signs such as redness, edema and swelling. Concerning about pocket
depth, there were more cases with greater than 5 mm (65.0 %) than less than 5 mm
(12.5 %). The mean smear pocket depth was 7 mm, ranging from 2 mm to 11 mm.
When the abscess condition was linked to the patients periodontal condition, 22(18.3 %) were diagnosed suffering from moderate to severe periodontits and werealso untreated, and 12 of them showed mild facial swelling of the affected side (Fig.
7).
5. Patients with pericoronitis
Other affected teeth were the lower third molars of adult patients (8.3 %). They had
localized gingival swelling, and pericoronal abscess developed around the crown ofpartially overlied (n=5) and the malposed (n=5) wisdom tooth. All the patients withmild to moderate facial swelling had surgical intervention and amoxicillin was theprincipal antibiotic.

6. Cytological findings
Light microscopical findings showed the abundance of microorganisms that
resembled Actinomyces and Entamoeba surrounded by varying amounts ofdebris based on the cytological characteristic morphology (60.8 %). Of this60.8 %, 28.3 % were children. In addition, the smears were infiltrated mainly withneutrophils and lymphocytes in all cases, and to a lesser extent withmacrophages (Fig. 8a).
The smears taken from one of deepest periodontal pockets identified the large
number of Actinomyces associated with Entamoeba like microorganisms. At
higher magnification, the Actinomyces like microorganisms exhibited finefilamentous structures (Fig. 8b), and the majority of the Entamoeba likemicroorganisms contained leukocytes within their digestive vacuoles and otherdebris could be observed within the cytoplasm, and nuclei were eccentricallylocated (Fig. 8c). Smears taken from inflamed alveolar mucosa of gingivitis cases
and from superficial pockets also revealed large bacterial mass of Actinomycesand debris but unable to find Entamoeba like microorganisms.

7. Changes in squamous epithelial cells
Cytological characteristics of epithelial cells were summarized in Table 3. Thecells showed reactive changes due to inflammation. Nuclear swelling ofsquamous epithelial cells was frequently visible. Other cytomorphologicalchanges were observed in the squamous
cells such as: perinuclear halo,and pyknotic chromatin.
Nuclear swelling
HaloPyknosis

8. Clinical and cytological findings
Concerning patients associated with microorganisms resemble toActinomyces and Entamoeba (45.3 %), they were mainly observed inperiodontitis and carious patients with 30.0 % and 23.1 %, respectively.
Characteristics of Actinomyces and Entamoeba like microorganisms wereidentified in 60.8 % cases. Clinically, these patients had unsatisfactory and poor
dental hygiene.

Table 1 Location of cytological smears
Locations
Pocket depth (mucosa) > 5 mm on theNo. of cases (%)
78 (65.0)

brush marking*
Mean pocket depth~7mm
Pocket depth (mucosa) < 5 mm on the15 (12.5)

brush marking**
Mean pocket depth~3mm
Inflamed alveolar gingival surface

24 (20.0)

Leukoplakia (buccal mucosa)

3 (2.5)*: periodontitis with no pocket abscess: n=20
: periodontal abscess associated to periodontitis: n=22
multiple caries: n=14
single caries: n=17
pericoronitis: n=5**: periodontal abscess associated to multiple decay: n=4, single decay: n=6,pericoronitis: n=5

Table 2: Clinical diagnosis and cytological findings
Clinical diagnosis
Periodontitis with no pocket abscessTotal Mean Age
No.
(yrs)
20

50.9

Swellingcase
%*
No.
16.7
0

22

59.8

18.3

12

54.5

19

15.8

18
23
10
24
3
120

8.0
10.0
21.7
27.9
42.0

15.0
19.2
8.3
20.0
2.5
100.0

67
1000
35

33.3
30.4
100.0
0.0
0.0

12
18
50
0
73

10.0
15.0
4.2
0.0
0.0
60.8

%**

Act+Enta .

0.0

No***
19%*
15.8

Periodontal abscess associated toPeriodontitis
Dental caries
Multiple decay aSingle decay b
Pericoronitis
Marginal gingivitis c
Leukoplakia
Total%*: Percentage of number/all cases (120)%**: Percentage of number/total cases
Act.+ Enta***.No.:Number of cases with Actinomyces + Entamoeba likemicroorganisms
a.: all childrenb.: 16 children, 7 adultsc.: 4 children , 20 adults

Table 3 : Summary of cytological characteristics
Characteristics
Nuclear Swelling (NS)Pyknosis (P)Halo (H)
Karyorrhexis (K)N/C ratio
Multinucleation (M)Vacuolation (V)Have all*(NS,P,H,K,N/C,M,V)
Cell size and shape

Severe-No. of cases(%)
Moderate
MildNegative
112 (93.3)
8 (6.67)
100 (83.3) 20 (16.7)
90 (75.0)30 (25.0)
75 (62.5)
45 (37.5)
110 (91.7)
10 (8.3)
Presence in small amounts
Presence in small amounts

120 (100%)
Variable within physiological limits*: all cases showed at least all characteristics: leukoplakia cases mainly showed P with lesser degree of other characteristics

N o.of patients

P atient population

127613951013116

M aleF em ale
61144

31~10 11~20 21~30 31~40 41~50 51~60 61~70 71~80
A ge range (yrs)Mean age=32.7, SD=20.8

Fig. 4: Characteristics of total patient population

Fig. 5: Extra- and intra- oral swellings associated with dental caries (arrowheads).

Tongan dentists at work

Fig. 6: Gingivitis and calculus

Fig. 7: Periodontitis

Fig. 8 (a): Papanicolaou stained oral smear with thebackground consisting of many inflammatory cells anddebris (arrowheads) (×100).

Fig. 8 (b): Well-formed branching of Actinomyceslike microorganism (arrows) with someinflammatory cells at the periphery (×1000).

Fig. 8 (c): Entamoeba like microorganism (arrows)
with digestive vacuoles containing leukocytes(×1000).

Discussion and conclusion
Our study showed that:* The clinicocytological results will provide knowledge of the periodontal diseases
and oral mucosal lesions in Tonga.
* The clinical findings showed that poor oral hygiene was a main feature.
* That periodontal abscesses have a high prevalence in periodontitis patientsand stems from pre-existing periodontal pocket (60.8 %).
* In younger population, periodontal abscesses developed in the absence ofperiodontitis due to impaction of foreign bodies as observed in dental caries
and acute pericoronitis.
* In adult population, abscesses occurred as an acute exacerbation of an
untreated periodontitis.
* The method highlighted the presence of large numbers of oral microorganisms
in patients with poor oral conditions ( Actinomyces and Entamoeba likemicroorganisms) and debris in the smears, giving the background the dirtyappearance.
* Only inflammatory atypia was found in the gingival mucosa.
Polymorpholeukocytes was observed in all clinically inflamed cases, and wasclearly suggestive of the hosts immediate response to infection.

* Exfoliative oral cytology might be effective assessment method in developing
countries as a simple and less expensive examination for the screening of oral
mucosal and periodontal diseases.
* It is worth integrating into the oral health programme planning, not only forfurther descriptive purposes, but also to enable the dental professional to tailor
practical oral health instruction.
* Ascertaining the bacterial population using this simple technique may signal animportant opportunity for dentists to improve their understanding of the nature oforal mucosal and periodontal diseases whereupon further preventive plans,reassurances and advice could be formulated.
* Since, the findings appear to be promoted by poor oral health, primaryprevention should be emphasized through hygiene educational programmes
ideally introduced in childhood.
* Because this is a pioneering study, further oral mucosal and periodontal studies
and investigations in Tonga based on it, are needed, especially to assess andreview the oral microflora and the health of the periodontal tissues using thissimple method. Hence, worsening of the mucosal problems may be prevented.
* The study showed the clinical diagnoses were correctly matched with thecytological diagnoses. Only the presence of oral microorganisms, inflammatory
cells and debris in varying quantities were revealed.