- 대한치과보철학회지 Vol. 24 No. 1, 1986 - 캔디다알비칸스의구강내빈도및분포도에관한연구 서울대학교치과대학보철학교실 이철규 김창회 Ⅰ. 서론 Ⅱ. 자료및방법 Ⅲ. 연구성적 Ⅳ. 총괄및고안 Ⅴ. 결론참고문헌영문초록 < 목차 > I. 서론 의치구내염은의치장착으로인해의치지지점막에나타나는단순한염증및유두상염증으로정의된다 (1). Cahn (2) 은 Denture sore mouth라는용어로이질환을언급했고최근에는만성위축성캔디다증 (chronic atrophic candidosis. 또는캔디다증 (candidosis) (3, 4) 이라는용어도자주사용되고있다. 이의치구내염의명확한병인론은규명되지않았으나외상 (5~8), 레진에대한과민반응 (9~11), 의치청결도 (12, 14), 완압실 ( 13), 취침시의치장착 (14) 이외에적합도및의치교합의부조화 (15, 16) 등이의치구내염의발병과깊은관계가있다고보고된바있다. 또한 Cahn (2) 이 Candida albicans가원인균일것이라고제안한후 Cawson (17), Turrell (18) 등이의치구내염환자에서 Candida albicans의양적인증가가일어났다고보고하였고 Lehner (4) 는면역형광법으로의치구내염환자의타액및혈액에서항체의증가가현저하므로 Candida가원인균으로간주된다고보고하였다. 이후로많은연구에서 Candida가원인요소중하나일것이라고보고되었다 (1, 19~32). Candida감염은엄밀한의미에서구강내상주균인 Candida albicans가편리공생관계에서기생관계로변화하는것인데이는병원체의변화로인한것이아닌숙주의조건이변화한때문이라고인정되며 (26) 이러한조건의변화란의치장착, 구강내포도당농도의증가및타액 ph의저하등으로 yeast가증식할수있는호조건이마련되거나방사선치료, 항암제복용및항생제의장기복용등에의한숙주의방어기전이억제되는등에기인한다고설명되고있다 (3). 이와같이의치구내염의발현은구강내 Candida albicans의양적증가와밀접한관계가있으나구강상주균인 Candida가정상성인에서그채취방법에따라 3~48% (28, 30) 의다양한수치로양성반응을보이는까닭에정상인지감염되었는지를알수있는양적증가를밝혀내는정량평가법이요구되어왔다 (27, 28). Davenport (20) 가 foam pad를이용한 imprint배양법을처음소개하여의치구내염환자에서 Candida의분포를밝혀냈으며 Arendorf와 Walker (27, 28) 는이를응용하여자연치군과의치군에서 Candida albicans의분포및단위면적당군락 (colony) 의밀도를조사하여구강내 Candida의정량평가를시도하였다. 91

Imprint 배양법을이용하여 Candida albicans 의분포와밀도및보균율의차이를조사하여분석하였는바이에의미있는결과를얻어보고하는바이다. 1. 연구대상 Ⅱ. 자료및방법 (1) 자연치군서울대학교치과대학재학생중정상치열을소유하고단순수복물이상의치과치료를받지않은 20세이상의남자 14명, 여자 14 명을연구대상으로하였으며이중남자의경우는흡연습관이있는자 7명, 흡연습관이없는자 7명으로구성하였다 (28). (2) 의치군서울대학교병원치과진료부보철과에내원하여상 하악에총의치를시술받은환자로저작등의기능수행에별다른문제점이없는환자중 20 명을선정하여대상으로삼았다. 다음의경우는연구대상에서제외하였다. 1 당뇨병 2 항생제및 steroid 장기복용자 3 항암제를복용중이거나방사선치료를받고있는환자 4 기타심각한전신적질환 Fig. 1 : Questionnaire form used in preliminary examination 2. 연구방법 (1) 설문조사 1 자연치군 : 연령 성별 흡연습관 간식정도 건강상태및칫솔질횟수 2 의치군 : 연령 성별 흡연습관 간식정도 건강상태 의치세척횟수및방법 현의치의장착연한무치악이된기간 추침시장착여부를기록하였다 (Fig.1). (2) 구강검사및의치검사 1 자연치군 i) 구강점막 : 점막의질환유무및질환양상 ii) 경험영구치 (DMFT) iii) 간이구강위생지수 : Green과 Vermillion씨방법 (1964) (35) 으로측정 Fig. 2 : Facial and basal surface quadrants for scoring plaque and stain deposits. iv) 타액 ph 측정 : ph 검사지 (Toyo Roshi 사제 ) 를설배면에 60 초간 (32) 접촉한뒤표준색과비교하여기록하였다. v) 보철물의존재여부및형태 2 의치군 i) 구강점막 : 점막의질환유무및양상 92

ii) 타액 ph의측정 : ph검사지를설배면에 60초간접 촉한상태로의치를장착하여시행한다. iii) 의치의위생검사 : 의치에부착된음식물잔사 치석 색소침착등을육안으로관찰하여기록하고보조 적수단으로상악의치를 plaque착색용액인 erythrosin(schein Port Reveal T.M. 사제 ) 에 30초 간담근다음흐르는물에가볍게씻어낸다. 이의치의 내 외면을 4등분하고각각의면에다음과같은기준으 로점수를준다 (36) (Fig.2). 0 : No plaque 착색 1 : Light plaque 1~25% 착색 2 : Moderate plaque 26~50% 착색 3 : Heavy plaque 51~75% 착색 4 : Very heavy plaque 76~100% (3) Imprint 배양법멸균된여과지를 Sabouraud s broth에담근다음각부위의구강점막에접촉시킨뒤다시멸균된 sponge로덮고균일한압력을 1분간가하여다음과같은부위에서각각채취하였다. 1 전설면 : 설배면의전방에사각형여과지의전방절단선양모서리를혀의측방변연부와일치시킨다. 2 후설면 : 설배면의후방중앙부에여과지의후방절단선을좌우측하악제2대구치의원심면을이은가상선과일치시킨다. 3 전방구개면 : 여과지의전방절단선을좌우상악견치의근심면을이은가상선과일치시킨다. 4 후방구개면 : 여과지의후방절단선을좌우상악제2대구치의원심면을이은가상선과일치시킨다. 5 좌우측협점막 : 여과지의전방절단선을각각하악제1소구치원심면의연장선과일치시킨다 (Fig.3). 의치환자의경우는인공치를기준치아로삼았으며전 후방구개면과동일한위치에대응이되는의치내면에서도채취하였다. 그외에보조적인수단으로설배면과구개면전체크기와모양에맞게여과지를절단한뒤이를각각설배면과구개면에압접하여 Candida의분포를전체적으로확인하기위해시행하였다. 위와같이 60초간점막에접촉한뒤여과지를제거하여 Sabouraud s dextrose agar plate에가압하여 8시간동안 37 부란기에호기배양하고, 8시간이후에는여과지를제거한다음 16시간동안다시동일한조건으로계속배양하여이를 Qubeec colony counter (American optical사제 ) 로측정한뒤단위면적당군락의수로환산하여기록했다. 예비실험에서인접부위나 Fig. 3 : Diagram showing how the sites sampled were determined. The sites were sampled with No. 2 filter paper. They are anterior tongue, posterior tongue and buccal cheek(a), anterior palate, posterior palate(b). 치아등에접촉하지않는가장큰사각형여과지는 2 2 cm2임이밝혀졌고, 또사용한선택배지는효모의적절한성장을위해 Streptomycin sulfate 40 μg / ml ( 한독 1g vial) 과 Benzathine penicillin G 40unit/ ml ( 한올 120 만 unit vial) 을첨가하고총 24 시간배양하는것이가장적당함이밝혀졌다. 이렇게배양이완료되얻어진군락을임의추출한후 Serum germ tube test 를시행하여 Candida albicans 임을추정하여최종확인하였다 (37). 93

(4) 도말표본제작 Imprint배양을시행한동일부위에서멸균된 1회용설압자를이용하여도말표본을얻어이를 Gram염색을한뒤 yeast상 organism의유무를관찰하였다 (19). 위의모든표본채취과정은식사후칫솔질을시행한뒤최소 2시간이경과된오전중으로 11시경을택하였고, 위과정에서얻어진모든결과는 Student s-test를이용하여 95% 유의수준으로검증하였다. Ⅲ. 연구성적 Imprint 배양법의결과 보균율의차이 : 자연치군의남성에서흡연자와비흡연자의보균율은흡연자에서 42.9%, 비흡연자에서 14.3% 로유의한차이를보였다 (P<0.05) 의치군의경우는흡연자의수가적어 (3 명 ) 그차이를비교할수없었다. 자연치군에서여성의보균율은 50% 로 36% 인남성보다높게나타났으나유의한차이는아니었다. 의치군에서도여성이 81.8% 로 55.6% 인남성보다높았으나유의한차이는아니었다. 자연치군과의치군을비교하면의치군은 70%, 자연치군은 39% 로유의한차이를보였다 (P<0.05) 의치군에서취침시장착습관이있는환자군이 80% 였고, 취침시의치를장착하지않는환자군은 60% 로나타났으나유의한차이는아니었다 (Fig.4). 채취부위에따른변화 : 자연치군에서채취부위에따른 Candida 의발현빈도와밀도모두가설배면에서높았으며특히후설면이가장높게나타났다 (Fig.5, 6). 의치군에서는의치내면, 특히구개융선부위가가장높게나타났고후설면에서도역시높게나타났다 (Fig.7, 8). 자연치군과의치군을비교해보면의치군이모든채취부위에서발현빈도및밀도모두가높게나타나며, 특히발현빈도에서는전 후방구개면의조직면이유의한차이로의치군에서높게나타났다 (P<0.05)(Fig.9, 10),(Tab.1, 2). 의치군에서취침시장착습관이있는사람은그렇지않은사람에비해발현빈도와밀도모두가높게나타났으나유의한차이는아니었다 (Fig.11, 12). 위의모든채취과정에서얻어진군락을임의추출하여 germ tube test 를시행한결과 85% 가 germ tube 양성반응을보여 Candida albicans 임을추정확인할수있었다 (37)(Fig.13, 14). 상피도말표본 : 상피도말표본을제작한시편으로자연치군에서 yeast 상 organism 을발견하기어려웠고의치군의경우는발현빈도가있다 (Fig.15). 사전조사 : 설문조사, 구강및의치검사의결과는보균여부에따라유의한차이를보이지않았다. 다만타액 ph, 자연치군의 plaque 지수, 의치의 plaque 점수항목에서보균자와비보균자사이에약간의차이를보였으나유의한차이는아니었다 (Tab.3~5). Fig. 4 : Comparison of candidal carrier rates in eachgroup of all subjects. The differences between A and B, I and J were significant, and others were not not significant. 94

Fig. 5 : Mean candidal density at various oral sites in dentate carriers(dentate group) Fig. 6 : Frequency of candidal detection in male and female(dentate group) Fig. 7 : Mean candidal density at various oral sites in dentate carriers of denture wearers(denture group) Fig. 8 : Frequency of candidal detection in male and female(denture group) Fig. 9 : Comparison of mean candidal density at various oral sites between dentate and denture group. Fig. 10 : Frequency of candidal detection in dentate and denture group. 95

Tab 1-2 : The mean candidal density and prevalence at various sites of dentate and denture subjects. Tab 3-4 : The mean of several factors examined with questionnaire, oral and denture examination. Fig. 11 : Mean candidal density at various oral sites in the carriers of denture wearers by night wearing(denture group) Fig. 12 : Frequency of candidal detection by night wearing of denture(denture group) 96

Fig. 13 : For the germ tube test the colonies were selected randomly and plated on the agar plates. Fig. 14 Typical germ tube is seen. This wad done with bovine calf serum(x 1000) Tab 5 Tab 5 : Several factors showed some differences between carriers and non carriers. Fig. 15 : Large numbers of yeast and hyphal form of Candida are seen. (Gram staining. x1000) Fig. 16-18 : Typical candidal colonies on the Sabouraud s dextrose agar plates are seen(dentate group). 97

Fig. 19-21 : Sabouraud s dextrose agar plates in denture wearers showing confluent growth(denture group). Ⅳ. 총괄및고안 많은문헌에서밝혀졌듯이 (27, 28, 29) imprint 배양법이타액표본법 인상배양법및도말표본법등에비해 Candida albicans의감지가훨씬예민하며정확한방법이라고할수있겠다. Arendorf와 Walker의연구 (28) 에서흡연자와비흡연자사이의보균율은유의한차이를보였으나그정확한기전은밝혀지지않았으며흡연으로인해구강점막이 Candida의군락형성에용이한조건을형성하는것이라고추정하고있다. 본연구에서도흡연자에서보균율이높게나타났다. 남녀간의보균율차이는자연치군과의치군공히여성이남성보다유의한차이는아니지만높게나타났으며이는 hormone의영향으로추정되고있다 (28). 타액 ph와 Candida 보균율과의관계는통계적유의성을보이지않으나보균자의평균 ph가비보균자에비해낮았는데이는 Young등의연구 (40) 결과와일치한다. Budtz-J rgensen (3, 41) 은의치내면에부착된 plaque 가의치구내염의원인으로작용한다고보고한바있으며 Miner (42) 에의한광학현미경연구와 Theilade 등 (43) 에의한전자현미경연구에서의치의 plaque 와자연치의 plaque 의기본구조가동일하다고밝혀졌다. 명확히의치의 plaque 내미생물과 Candida 가어떤작용을하는지는밝혀지지않았으나 Hodson 과 Craig (34) 는우식치가많을수록 yeast 의발현빈도가높았다고보고하였고, Samaranayake 등 (44, 45) 의시험관연구에서 Streptococcus salivarius 및기타 Streptococci 와 Lactobacilli 등이레진에 Candida 가부착되는정도에영향을미친다는연구가있었다. 본연구의결과의치 plaque 의점수와 Candida 보균율사이에통계적유의성은없었으나보균자에서 plaque 점수가높게나타났는데이는 Budtz-J rgensen (3, 41) 의연구결과와일치한다. Arendorf 와 Addy (32) 는교정장치의장착으로타액 ph 가감소하고 plaque 가증가한다고보고하면서구강내어떠한의치나장치도이러한변화를야기할것이라고발표했다. 이들연구결과 (28, 32) 와마찬가지로본연구에서도의치군에서이러한조건들의변화가인정되고있 98

다. 이와같이구강내환경변화에의치가차지하는비중이인지된이후의치내 plaque 를제거하는방법과효과적인세정제의개발에많은연구가발표된바있다 (46~56). 자연치군에서채취부위중빈도가가장높은곳은후설면이었다. 이는 Arendorf 등 (27, 28, 32) 의연구결과와일치하는데이는이부위의 filiform papilla( 사상유두 ) 로인해 Candida 부착이용이하게된것으로추정되고있다 (28). Cooke (33) 는이부위가 Candida 의주된서식처임은이부위에정중능형설염 (Median rhomboid glossitis) 의빈발과깊은관계가있다고보고한바있다. 그다음으로높은곳은전설면과전방구개면인데전방구개면은구개융선으로인해높은것으로사료된다. 의치군의경우의치내면에서빈도와밀도가가장높게나타난것은의치내면이거칠어음식물과미생물의부착을용이하게하고, 또의치세척이곤란한것도그요인으로간주되며 (46~56), 특히의치의전방구개면은칫솔이나기타도구로도세척하기어려워의치세정제의보급이시급하다고사료된다. 의치군의장착습관에따라 Candida 의발현빈도및밀도를살펴보면취침시의치를장착하는군은빈도와밀도모두가취침시의치를장착하지않는군에비해높게나타났으되유의한차이는아니었으나, Love (14) 의연구에서취침시의치장착이의치구내염발현과관련이깊다고보고된바있다. 본연구에서는타액 ph 및구강점막은수회의세척을경험한까닭에유의한차이를보이지않았을것으로사료된다. 측정시간이기상시간과가깝다면유의한차이를보이지않았을까라고추측할수있다. 도말표본의결과는자연치군에서균사의존재가발견되지않았고의치군에서는후설면, 구개면및의치내면에서간혹보였다. 상피도말법을이용한많은연구가구강내대부분의 yeast 는 Candida albicans 라고주장했으나 (20, 21, 40) Arendorf 와 Walker(27, 28) 의연구에서밝혀졌듯이상피도말법을이용해관찰된 yeast 는 Candida 인지의여부를알기위하여는부가적인실험이요구되며정량화하여양적비교가곤란하다. 이에비해 imprint 배양법은양적비교가가능하고 Candida 여부를확인하는과정이복잡하지않다. 이와같이도말표본법에비해 imprint 배양법이우수하여본연구에서도 imprint 배양법을위주로하고도말표본법은보조적인수단으로활용하였다. 본연구의결과로볼때도도말표본법은 Candida 의보균상태여부, 분포도및빈도관찰에적절한방법이아니라고사료된다. 본연구에서시행한여러가지사전조사결과, 그중타액 ph, 자연치군의 plaque지수, 의치의 plaque 점수등이보균자와비보균자사이에유의한차이는아니었으나보균자에서타액 ph는낮게 plaque지수및점수는높게나타나는경향을보였는데이와같은사실은 Arendorf 등 (27, 28) 과 Hodson 등 (34) 의연구결과와일치한다. 예비실험과정에서다음과같은몇가지사실을밝혀냈다. 그것은먼저 imprint 배양법을수행하기에는 sponge보다여과지가더우수한결과를보였다는점인데이는여과지가훨씬더균일하고긴밀하게조직에접촉할수있을뿐아니라흡습성도뛰어나기때문이라고사료된다. 상악모형을수집하여그를근거로인접조직에접촉하지않고채취할수있는최대크기는 2 2cm2임을알수있었는데, Arendorf등은 2.5 2.5cm2의크기를사용하였는바이러한결과로한국인과서양인과의치궁의크기및형태의차이가있었음을확인할수있었다. Arendorf와 Walker가 Sabouraud s dext rose agar 배지에다른미생물의성장을억제하기위해서첨가한항생제는그농도가 Streptomycin sulfate 5mg / ml, Benzathine penicillin G 1.5mg / ml인데이농도로시행한결과 Candida의성장이활발하지못하여본실험에서는 Streptomycin sulfate 40/ ml, Benzathine penicillin G 40unit/ ml로줄여서첨가하였고배양시간도48시간 (27, 28) 을 24 시간으로줄여서지나친성장을방지하였다. 이러한항생제농도및배양시간의결정은문헌 (38, 39) 에서제시된농도및시간을수차례시행한후가장적절한것으로택하였다. 따라서본연구에서얻어진단위면적당배양된군락의수를조건이다른 Arendorf등의연구결과와비교하는것은의미가없으며오히려양성반응빈도와보균율이더중요한의미를갖는다고사료된다. 본연구에서많은사실들이밝혀졌는데특히의치가구강내 Candida의증식과깊은관계가있고의치내면이구강내 Candida의주된증식처라는사실이나타났으나이러한결과만으로 Candida가의치구내염의유일한원인이라고단정할수없기때문에의치구내염의병인을밝히기위해서는보다더깊은연구가필요하다고생각된다. 이에의치를장착하는환자개개인의의치장착전 후에따른 Candida의빈도및분포도의변화를관찰하고의치구내염환자에서증상의심도에따른 Candida의빈도및밀도를연구하고나아가자연치군을대상으로장치물을이용해구개점막을피개하여그에따른 Candida의변화및의치구내염의발병기전을 99

좀더깊이연관지을수있으므로향후의연구는이러한점을감안하여이루어지는것이바람직하다고사료된다. Ⅴ. 결론 자연치군 28명, 의치군 20명을대상으로 imprint 배양법및상피도말표본법으로 Candida albicans의구강내빈도및분포도에대하여연구한결과다음과같은결론을얻었다. 1. Candida 보균율은의치군 (70.0%) 이자연치군 (39.0%) 에비해유의한차이로높았다 (P<0.05) 2. 남녀간보균율에서는여성이남성에비해높았으나유의한차이는아니었으며흡연여부에따른자연치군남성에서흡연자 (42.5%) 가비흡연자 (14.3%) 에비해유의한차이로높았다 (P<0.05). 3. Candida의빈도와분포도는자연치군에서는후설면이, 의치군에서는전방구개면이각각높았다. 4. 빈도에서의치군과자연치군사이에유의한차이를보이는조직면은전방및후방구개면이었다 (P<0.05) 5. 빈도와밀도에서취침시의치장착여부에따른차이는취침시의치를장착하는군이장착하지않는군에비해높았다. ( 끝으로본논문을완성하기까지시종지도편달하여주신김창회교수님께감사드리며, 조언과격려를아끼지않으신보철학교실여러교수님들과보철과의국원여러분께감사를드립니다. 그리고본연구를위해많은협조를하여주신서울대학교치과대학미생물학교실원여러분께도사의를표하는바입니다.) REFERENCES 1. E. Budtz-J rgensen, U. Bertram : Denture stomatitis I. The Etiology in relation to trauma and infection. Acta. Odont. Scand. 28 : 71, 1970. 2. Cahn, L. R. : The denture sore mouth. Ann. Dent. 3 : 36, 1936. 3. E. Budtz-J rgensen : Clinical aspects of Candida infection in denture wearers. JADA 96 : 474, 1978. 4. Lehner, T. : Immunofluorescent investigation of Candida. Dent. Practit. 16 : 142, 1965. 5. Hecht, S. S. : Chronic irritation of the epithelial tissue of the mouth associated with dentures. Amer. J. Orthodont. and Oral Surg. 25 : 574, 1939. 6. Nyquist, G. : Denture sore mouth. Acta. Odont. Scand. 10 : suppl 9, 1952. 7. Nyquist, G. : The influence of denture hygiene and the bacterial flora on the condition of the oral mucosa in full denture cases. Acta. Odont. Scand. 11 : 24, 1953. 8. Newton, A. V. : Denture sore mouth. Brit Dent. J. 112 : 357, 1962. 9. Rattner, H. : Stomatitis due to sensitization to dental plates. JADA 106 : 2230, 1936. 10. Cole, H.N. : Stomatitis and cheilitis due to denture plates. Arch. Derm. Syph. 37 : 338, 1938. 11. Vickers, H. R. : Sensitization to denture material as a cause of angular stomatitis. Brit. Med. J. 2 : 1091, 1949. 12. Fisher, A. a. : Allergic sensitization of the skin and oral mucosa to acrylic resin denture materials. J. Prosth. Dent. 6 : 593, 1956. 13. Campbell, R. L. : Relief chambers in complete dentures. J. Prosth. Dent. 11 : 234, 1961. 14. William, D. Love, F. A. Goska, R. J. Mixson : The etiology of mucosal inflammation associated with dentures. J. Prosth. Dent. 18 : 515, 1967. 15. Robinson, H.B.G. : Diagnosis of Lesions Associated with Dentures. J. Prosth. Dent. 7 : 338, 1957. 16. Hickey, J. C., Boucher, C.O. and Woelful, J.B. : Responsibility of the Dentist in complete dentures. J. Prosth. Dent. 12 : 638, 1962. 17. Cawson, R. A. : Denture sore mouth and angular cheilitis. Brit. Dent. J. 115 : 441, 1963 18. Turrell, A.S.W. : Etiology of inflamed upper denture-bearing tissues. Brit. Dent. J. 118 : 542, 1966. 19. R. P. Masella., C.T. Dalan and W.R. Laney : The prevention of the growth of candida on silastic 390 soft liner for denture. J. Prosth. Dent. 33 : 250, 1975. 100

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= Abstract = A STUDY ON THE PREVALENCE AND INTRA-ORAL DISTRIBUTION OF CANDIDA ALBICANS Cheol Gyu Lee, Chang Whe Kim Dept. of Prosthodontics, College of Dentistry, Seoul National University Using imprint cultures and epithelial smears, the density and prevalence of colonization of oral mucosal sites and denture surfaces by Candida albicans has been determined in 28 healthy dentate subjects and 20 denture wearers. With questionnaire, oral and denture examination, the relationship between the carrier rates and several factors; DMFT, oral hygiene index, salivary ph & denture plaque score were studied. But these factors have not significant relationship to the carrier rates. Imprint culture appears to be sensitive technique for detecting candidal carriers and be useful for distinguishing between the healthy carrier state and candidal infection. Cigarette smokers had a significantly increased carrier state(p<0.05) compared with nonsmoker in made dentate subjects. Female were more frequent carriers than male in dentate and denture group, but these differences were not significant. In denture wearers, there was a higher density and frequency of candidal colonization of all mucosal sites sampled, compared with that of healthy dentate group especially anterior palate and posterior palate showed highly significant differences in frequency of candidal colonization(p<0.05). The distribution of Candida albicans is not uniform throughout the mouth. The tongue in the healthy dentate subjects and the impression surfaces of upper dentures are the primary oral reservoirs for the fungus. Overnight wearing of dentures was associated with increased density and frequency of candidal colonization and density. 103