pissn: 2288-0402 eissn: 2288-0410 6(5):248-254, September 2018 https://doi.org/10.4168/aard.2018.6.5.248 ORIGINAL ARTICLE Polymerase chain reaction 양성소아마이코플라스마폐렴에서혈청 IgM enzyme-linked immunosorbent assays 의진단적가치 이혜진, 이윤태, 김경훈, 양은애, 김환수, 전윤홍, 윤종서, 김현희, 김진택 가톨릭대학교의과대학소아과학교실 Diagnostic value of serum IgM enzyme-linked immunosorbent assays in polymerase chain reaction-positive Mycoplasma pneumonia in children Hye Jin Lee, Yoon Tae Lee, Kyung Hoon Kim, Eun Ae Yang, Hwan Soo Kim, Yoon Hong Chun, Jong-Seo Yoon, Hyun Hee Kim, Jin Tack Kim Department of Pediatrics, College of Medicine, The Catholic University of Korea, Seoul, Korea Purpose: Mycoplasma pneumoniae (MP) is a common cause of community-acquired pneumonia (CAP) in children. MP serum IgM and polymerase chain reaction (PCR) are the methods that enable early diagnosis in patients with MP pneumonia. The objective of this study was to investigate the clinical value of serum MP-specific IgM antibodies in PCR-positive MP pneumonia for the early diagnosis of MP pneumonia in children with CAP. Methods: Out of 129 patients with lower respiratory tract infection aged over 3 years, 90 CAP children were enrolled in the study. Throat swab MP real-time PCR and serum enzyme-linked immunosorbent assays (ELISA) IgM antibodies were performed. A positive rate of MP PCR and serum IgM, the level of IgM index, clinical features, and laboratory findings were analyzed. Results: MP PCR was positive in 57 cases. Longer fever duration before admission (P< 0.001), higher rates of lobar or segmental pneumonia (P= 0.048), unilateral infiltration (P= 0.038), and extrapulmonary symptoms (P= 0.049) were associated with MP PCRpositive pneumonia. Serum IgM index was significantly higher in MP PCR-positive pneumonia them in MP PCR-negative pneumonia (3.9± 3.0 vs. 0.8± 1.3, P< 0.001). Using MP PCR as a gold standard, the sensitivity, specificity, positive predictive value and negative predictive value of serum IgM were 85.5%, 82.1%, 91.4%, and 71.9%, respectively. The area under the curves for serum IgM index was 0.892, and the ROC analysis indicated that an optimal cutoff value of 1.05 for serum IgM provided the highest sensitivity and specificity interestingly (83.9% vs. 85.7%, P< 0.001). Conclusion: Serum IgM ELISA has useful diagnostic value in PCR-positive MP pneumonia. Applying an IgM index cutoff of 1.05 improves diagnostic accuracy. ( 2018;6:248-254) Keywords: Mycoplasma pneumoniae, Diagnosis, Polymerase chain reaction, Immunoglobulin M, Pediatrics 서론마이코플라스마는전세계적으로지역사회획득폐렴의 10% 40% 를차지하며, 1,2 특히 3 15세사이소아의지역사회획득폐렴의중요한원인균으로최근에는발병주기가 4 7년에서 3년정도로단 축되고있고, 3 단체생활등으로인하여발병연령도학동기에서 4 6세로점차낮아지고있는추세이다. 3-5 대부분양성경과를취하며자연치유되는경향을보이지만일부에서는생명을위협하는전격성혹은괴사성폐렴으로진행하거나폐출혈을동반한급성호흡곤란증후군의원인이되기도한다. 6-9 Correspondence to: Jong-Seo Yoon https://orcid.org/0000-0002-5782-6175 Department of Pediatrics, Seoul St. Mary s Hospital, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul 06591, Korea Tel: +82-2-2258-7612, Fax: +82-2-537-4544, E-mail: pedjsyoon@catholic.ac.kr Received: March 14, 2018 Revised: August 28, 2018 Accepted: September 12, 2018 2018 The Korean Academy of Pediatric Allergy and Respiratory Disease The Korean Academy of Asthma, Allergy and Clinical Immunology This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/). 248 http://www.aard.or.kr
이혜진외 중합효소연쇄반응및 IgM ELISA 를통한소아마이코플라스마폐렴의조기진단 또한광범위한면역반응에의하여피부, 신경학적, 혈액학적, 근골격계질환등심각한폐외증상을동반할수있으며, 10,11 최근연구결과에따르면집중치료를요하는중등도이상의마이코플라스마폐렴환자가증가하고있다. 12,13 전세계적으로마크로라이드항생제내성폐렴마이코플라스마가증가하고있으며, 14 우리나라의경우에도 2011년국내다기관연구에서 62.9% 로발표된이후에도꾸준히증가하여 2017년한단일기관연구에서는 87.2% 까지보고되었다. 15 중증도이상의마이코플라스마폐렴에서조기에스테로이드치료를하는경우치료효과를보이는경우도있지만, 16,17 숙주의과도한세포면역반응과기도섬모기능장애를유발하여발열기간이길고, 병의경과가심한스테로이드저항성마이코플라스마폐렴도보고되고있다. 18 위와같은이유로마이코플라스마폐렴은지역사회획득폐렴에서바이러스혹은일반세균성폐렴과는차별적으로조기에의심하고진단하는것이중요하지만, 현실적으로마이코플라스마를조기에확진할수있는단일검사법은존재하지않기때문에임상에서어려움을겪는경우가많다. 전통적으로마이코플라스마폐렴의진단은인후면봉또는객담검체에서균의배양또는혈청특이 IgG 항체의 4배이상증가로확진할수있다. 그러나배양법은마이코플라스마균의배양조건이까다롭기때문에민감도가 23% 64% 정도로낮고, 19 시간이오래걸리며, 혈청 IgG 항체검사법은역가의상승을확인하기위한반복적채혈이필요하고수주이상시간이소요된다는단점이있어두진단방법모두임상에서활용하기에제한점이있다. 혈청 IgM 진단 kit, 리보솜단백질 (ribosomal protein) 신속항원검사 (rapid antigen kit), 고해상전산화단층촬영 (high resolution computerized tomography) 등마이코플라스마조기진단을위한다양한연구들이보고되었다. 20,21 따라서이연구에서는, 소아의마이코플라스마폐렴환자에서조기에결과를얻을수있는검사방법인중합효소연쇄반응 (polymerase chain reaction, PCR) 과혈청 IgM 검사를비교분석하고, PCR 양성마이코플라스마폐렴환자를대상으로혈청 IgM 검사의진단적유용성을알아보고자하였다. 대상및방법 1. 대상 2014년 8월 1일부터 2017년 7월 31일까지서울성모병원소아청소년과호흡기분과에입원한만 36개월이상의폐렴환자들을대상으로의무기록과방사선검사소견을후향적으로검토하였다. 폐렴의임상적진단기준은 38 C 이상발열, 급성호흡기증상, 기타원인으로설명되지않는단순흉부사진에서음영증가로하였다. 22 기저질환으로만성폐질환, 혹은악성혈액종양등의전신질환이있는 경우는대상에서제외하였다. 총 129명의 3세이상폐렴입원환자중혈청마이코플라스마 IgM 검사와구인후면봉 (throat swab) 실시간중합효소연쇄반응 (real-time PCR) 검사를모두시행한환자, 총 90명을연구대상으로하였다. 대상환자들의임상양상, 방사선검사결과, 혈액검사결과를비교하고혈청마이코플라스마 IgM과인후면봉 PCR 검사결과를조사하였다. 2. 방법대상환자들의의무기록을검토하여효소면역측정법 (enzymelinked immunosorbent assay, ELISA) 으로측정한혈청마이코플라스마 IgM index 수치와인후면봉 PCR 검사결과를분석하였다. 바이러스폐렴의감별진단을위하여비인두면봉검체로시행한다중역전사중합효소연쇄반응 (multiplex reverse transcriptionpolymerase chain reaction, RT-PCR) 결과를조사하였다. 대상환자를마이코플라스마 IgM과 PCR 양성여부에따라두그룹으로분류하여임상양상, 검사실검사결과, 방사선검사결과, 치료반응등을비교하였다. PCR 양성마이코플라스마폐렴에서혈청 IgM 검사의민감도, 특이도와양성예측도, 음성예측도, cutoff 값을분석하였다. 1) 혈청마이코플라스마 IgM 검사마이코플라스마 IgM 측정은 Chorus Trio 장비 (DIESSE diagnostic, Monteriggioni, Italy) 의 Chorus M. pneumonia IgM 시약을이용하여실시하였다. 폐렴마이코플라스마의항원 P1 cytoadhesin, 결합체 (conjugate), 희석액, tetramethylbenzidine (TMB) substrate 등의기질이담겨있는카트리지형식의시약에혈청 50 μl를분주하고장비에장착하여항체가를측정하였다. IgM 항체가는검체의 450 nm에서흡광도를측정하여보정물질의흡광도로나눈 index 수치로표기하였으며, index 0.9 미만은음성, 0.9 이상은양성으로판정하였다. 23 2) 중합효소연쇄반응구인후면봉법으로검체를채취하여 2 ml 생리식염수배지가담긴 falcon tube에넣고흔든후실시간중합효소연쇄반응 (M. pneumoniae Real-Time PCR Kit, Slan; Biocore, Seoul, Korea) 을이용하여 ATPase operon 을정성검출하였고, 검사방법은다음과같다. (1) Real-time PCR Reaction mixture 10 μl 당 Primer/Probe mixture 5 μl를혼합하여필요한만큼의 real-time PCR Master mixture를제조한다. (2) Real-time PCR Master mixture 15 μl를각각의 tube에분주하고시료의추출한 DNA 5 μl를 pipetting하여잘섞는다. (3) Real-time PCR machine 에넣어반응시킨다. (4) CT (threshold cycle) 값을 40으로설정후양성또는음성결 https://doi.org/10.4168/aard.2018.6.5.248 249
Lee HJ, et al. PCR and serum IgM ELISA for early diagnosis of Mycoplasma pneumonia 과를확인한다. 3. 마이코플라스마폐렴의진단및정의모든대상환자를대상으로입원 1병일에시행한마이코플라스마중합효소연쇄반응과혈청 IgM 검사결과를분석하였다. 진단적편의를위하여중합효소연쇄반응양성군과음성군으로분류하였으며, 이중중합효소연쇄반응양성인경우를마이코플라스마폐렴으로정의하였다. 24 환자들의발열및기침기간, 폐외동반증상및산소요구도등의임상양상, 일반혈액검사, 염증반응지표, 간기능검사, 방사선검사결과를분석하였다. 4. 통계연령은평균연령 ± 표준편차로측정값을표시하였다. 통계분석 은 IBM SPSS Statistics ver. 24.0 (IBM Co., Armonk, NY, USA) 을이용하였으며, 연속변수의비교에는 Student t-test를, 빈도변수의비교에는 chi-square test를이용하였고, 마이코플라스마폐렴과관련된독립적연관인자들의분석에는단변량분석에서유의한차이를보인변수들에대하여다변량로지스틱회귀분석 (multiple logistic regression analysis) 의후진형축차변수선택법 (backward stepwise regression method) 과그룹간성향점수매칭 (propensity score matching) 분석을시행하였다. Receiver operating characteristics (ROC) curves를이용하여 PCR 양성마이코플라스마폐렴진단에있어혈청 IgM index의유용성을평가하였으며, 상관분석은 Pearson 상관계수 (Pearson correlation coefficient) 를이용하여분석하였다. 모든분석에서 P 값이 0.05 미만인경우를유의한것으로판단하였다. Table 1. Demographic and clinical features and laboratory findings of study subjects Variable All patients (n= 90) M. pneumoniae PCR M. pneumoniae IgM P-value Positive (n= 62) Negative (n= 28) Positive (n= 58) Negative (n= 32) Age (yr) 5.7± 3.7 6.4± 3.6 4.2± 3.5 0.080 6.0± 3.5 5.1± 4.0 0.261 Male sex 37 (41.1) 25 (67.6) 12 (32.4) 0.172 22 (37.9) 15 (46.9) 0.503 Hospital days 5.5± 4.0 6.0± 4.7 4.5± 1.3 0.137 6.2± 4.9 4.2± 1.0 0.003 Symptoms Fever before admission (day) 4.9± 2.3 5.6± 2.1 3.4± 2.1 < 0.001 5.5± 2.2 3.8± 2.1 0.001 Fever after admission (day) 1.9± 2.7 2.2± 3.1 1.4± 1.3 0.068 2.3± 3.2 1.3± 1.1 0.082 Total fever duration (day) 6.9± 3.7 7.7± 3.7 4.7± 2.5 < 0.001 7.8± 3.8 5.1± 2.6 0.001 Cough before admission (day) 5.4± 3.0 5.9± 2.7 4.3± 3.3 0.001 5.9± 2.8 4.6± 3.2 0.046 Peak temperature ( C) 38.6± 0.8 38.7± 0.7 38.5± 1.0 0.413 38.5± 1.5 38.6± 0.9 0.413 Lobar/segmental pneumonia 46 (51.1) 36 (58.1) 10 (35.7) 0.048 33 (56.9) 13 (40.6) 0.139 Unilateral infiltration 47 (52.2) 37 (59.7) 10 (35.7) 0.038 33 (56.9) 14 (43.8) 0.232 Extra-pulmonary symptoms 13 (14.4) 12 (19.4) 1 (3.6) 0.049 2 (6.3) 11 (19) 0.100 Pleural effusion 11 (12.2) 9 (14.5) 2 (7.1) 0.492 9 (15.5) 2 (6.3) 0.199 Steroid use 61 (67.8) 47 (75.8) 14 (50.0) 0.015 46 (79.3) 15 (46.9) 0.002 O2 need 12 (13.3) 5 (8.1) 7 (25.0) 0.029 6 (10.3) 6 (18.8) 0.262 Positive RV PCR 36 (40.0) 19 (30.6) 17 (60.7) 0.013 21 (36.2) 15 (46.9) 0.248 Laboratory findings WBC (10/μL) 9.5± 4.9 9.8± 5.3 9.0± 4.0 0.788 10.1± 5.3 8.6± 4.1 0.179 Neutrophils 64.1± 14.4 66.6± 12.5 58.5± 17.0 0.056 66.3± 12.8 60.0± 16.5 0.047 Lymphocyte 25.4± 12.1 23.0± 9.6 30.8± 15.1 0.027 23.4± 10.2 29.0± 14.4 0.058 Monocyte 8.4± 3.2 7.9± 3.0 9.6± 3.4 0.052 8.0± 2.9 9.3± 3.6 0.068 Eosinophil 2.0± 3.7 2.4± 4.2 0.9± 1.5 0.003 2.6± 4.4 0.7± 1.1 0.002 ESR (mm/hr) 37.9± 19.2 39.1± 20.3 35.3± 16.5 0.599 39.4± 20.7 35.3± 16.3 0.333 CRP (mg/dl) 4.5± 4.9 4.8± 5.2 3.9± 4.1 0.705 4.7± 5.2 4.2± 4.2 0.664 AST (IU/L) 36.2± 17.7 35.8± 19.4 37.1± 13.6 0.621 36.8± 19.7 35.0± 13.5 0.644 ALT (IU/L) 26.9± 34.7 25.0± 25.7 31.1± 49.5 0.960 26.3± 26.6 28.0± 46.5 0.826 Serum MP IgM (index) 2.9± 3.0 3.9± 3.0 0.8± 1.3 < 0.001 4.3± 2.8 0.4± 0.2 < 0.001 Values are presented as mean± standard deviation or number. MP, Mycoplasma pneumoniae; PCR, polymerase chain reaction; RV, respiratory virus; WBC, white blood cells; ESR, erthyrocyte sedimentation rate; CRP, C-reactive protein; AST, aspartate aminotransferase; ALT, alanine aminotransferase. P-value 250 https://doi.org/10.4168/aard.2018.6.5.248
이혜진외 중합효소연쇄반응및 IgM ELISA 를통한소아마이코플라스마폐렴의조기진단 결과 1. 환자의특성총대상환자 90명의연령분포는 3세부터 15세까지로, 평균연령은 5.7세였고, 남아가 37명으로 41.1% 를차지하였다 (Table 1). 평균입원기간은 5.5일, 평균입원전발열기간은 4.9일이었다. 방사선검사결과, 대엽성 (lobar) 및분절성 (segmental) 폐렴환자가 46명 (51.1%), 한쪽폐만을침범한폐렴환자가 47명 (52.2%) 이었다. 입원기간중시행한마이코플라스마중합효소연쇄반응양성군은 62명 (68.9%) 으로, 호흡기바이러스중합효소연쇄반응양성군은 36명 (40%) 에비하여높은비율을보였다. 마이코플라스마중합효소연쇄반응음성군에서호흡기바이러스양성률은 60.7% 로전체에비하여높은비율을보였고, 검출된바이러스는 human rhinovirus, respiratory syncytial virus, parainfluenzavirus, adenovirus 순이었다. 2. 마이코플라스마 PCR, 혈청 IgM 양성군의임상양상 1) 중합효소연쇄반응양성군총 90명의대상환자중 62명의마이코플라스마중합효소연쇄반응양성폐렴환자에서입원전발열기간 (5.6±2.1 days vs. 3.4± 2.1 days, P< 0.001) 및총발열기간 (7.7±3.7 days vs. 4.7±2.5 days, P< 0.001), 대엽성및분절성폐렴 (58.1% vs. 35.7%, P = 0.048), 한쪽폐를침범한경우 (59.7% vs. 35.7%, P = 0.038), 그리고발진, 복통, 두통등의폐외동반증상 (19.4% vs. 3.6%, P = 0.049) 비율이음성군에비하여높았다. 중합효소연쇄반응양성군에서음성군에비하여혈청마이코플라스마 IgM의평균값이통계적으로유의하게높은것으로나타났다 (3.9± 3.0 vs. 0.8±1.3, P< 0.001). 2) 혈청마이코플라스마 IgM 양성군총 58명의혈청 IgM 양성환자에서항체음성환자군에비하여평균입원기간 (6.2±4.9 days vs. 4.2±1.0 days, P = 0.003), 입원전 (5.5±2.2 vs. 3.8± 2.1, P = 0.001) 및총발열기간 (7.8± 3.8 days vs. 5.1± 2.6 days, P = 0.001) 이길었으며, 입원당일말초혈액검사상호산구분율이 IgM 양성군환자에서유의하게높았다 (2.6% ± 4.4% vs. 0.7%±1.1%, P = 0.002). 입원전발열기간과혈청 IgM index 수치와의연관성분석을위하여 Pearson correlation coefficients로상관관계를분석하였으며, 입원전발열기간이길수록 IgM index 값이증가하였다 (r = 0.438, P< 0.001) (Fig. 1). 3) 혈청마이코플라스마 IgM index의 ROC 분석 ROC 분석을이용하여 PCR 양성마이코플라스마폐렴에대한혈청 IgM index 분석결과, AUC 0.892로혈청 IgM은통계적으로유의한진단적예측인자였으며, 혈청 IgM index cutoff 값은 1.05일때가장높은민감도 (83.9%) 와특이도 (85.7%) 를보였다 (P< 0.001) (Fig. 2). 4) 혈청마이코플라스마 IgM의민감도, 특이도와양성예측도, 음성예측도마이코플라스마 PCR 양성환자군분석결과, 총 62명중 53명에서혈청 IgM 검사양성이었으며, PCR 음성환자 28명중 23명에서혈청 IgM 검사가음성으로나타났다 (Table 2). 마이코플라스마 PCR 양성을기준으로한혈청 IgM 검사의민감도, 특이도, 양성예측도, 음성예측도는각각 85.5%, 82.1%, 91.4%, 71.9% 였으며, 양성검정우도비 (positive likelihood ratio) 는 4.79였다 (Table 3). 10 1.0 8 0.8 IgM (index) 6 4 Sensitivity 0.6 0.4 2 0.2 0 0 2 4 6 8 10 12 0 0.2 0.4 0.6 0.8 1.0 Fever before admission (day) 1-specificity Fig. 1. Correlation of the fever duration before admission with serum Mycoplasma pneumoniae IgM index in all patients (Pearson correlation coefficient r= 0.438, P< 0.001). Fig. 2. Receiver operating characteristic curve for serum M. pneumoniae (MP) IgM index in the diagnosis of MP pneumonia. The optimal cutoff values for serum MP IgM index were 1.05 (area under the curve, 0.892; sensitivity 83.9%, specificity 85.7%; P< 0.001). https://doi.org/10.4168/aard.2018.6.5.248 251
Lee HJ, et al. PCR and serum IgM ELISA for early diagnosis of Mycoplasma pneumonia Table 2. Relationship between serum Mycoplasma pneumoniae (MP) PCR and IgM antibodies of all study subjects (n= 90, P< 0.01) MP IgM antibodies Positive MP PCR 5) 마이코플라스마 PCR, 혈청 IgM 양성폐렴의임상적예측인자 마이코플라스마 PCR 및혈청 IgM 검사의단변량분석에서확인 된유의한개별임상양상들을동시에변량으로적용하여, 단계별 로마이코플라스마검사양성폐렴예측에기여정도가낮은임상 적인자를제거하는후진식다변량회귀분석을실시하였다. 분석 결과, 마이코플라스마 PCR 과혈청 IgM 양성군에서공통적으로 입원전발열기간이통계적으로유의한예측인자로추정되었다 (odds ratio [OR], 1.60; 95% confidence interval [CI], 1.11 2.31; P = 0.013; OR, 1.42; 95% CI, 1.11 1.82; P = 0.005). 마이코플라스마 PCR 음성과양성두그룹간유의한차이를보이는입원전발열기 간등을성향점수매칭한후 IgM 양성 OR 값을비교하였으며, 그 결과매칭전과후 IgM 양성확률이모두유의한것으로나타났다 (Table 4). 이외에 PCR 양성군에서폐외동반증상 (OR, 11.40; 95% CI, 1.07 121.10; P = 0.044) 과 IgM index 수치 (OR, 2.61; 95% CI, 1.44 4.73; P = 0.002) 가유의한예측인자로나타났다. 고찰 Negative Total Positive 53 (58.9) 5 (5.6) 58 (64.4) Negative 9 (10.0) 23 (25.6) 32 (35.6) Total (n= 90) 62 (68.9) 28 (31.1) 90 (100) Values are presented as number. PCR, polymerase chain reaction. Table 3. Diagnostic values of serum Mycoplasma pneumoniae (MP) IgM antibodies with a MP PCR as the gold standard Sensitivity Specificity PPV NPV MP IgM antibodies 85.5 82.1 91.4 71.9 4.79 PCR, polymerase chain reaction; PPV, positive predictive value; NPV, negative predictive value; PLR, positive likely hood ratio. 이번연구의목적은소아의마이코플라스마폐렴환자에서조기 에결과를얻을수있는검사방법인중합효소연쇄반응과혈청 IgM 검사를비교분석하고, PCR 양성마이코플라스마폐렴환자를대 상으로혈청 IgM 검사의진단적유용성을평가하고자하였다. 혈청 IgM 검사에관한연구들에서초회검사의민감도는 46% 78%, 특이도는 80% 98% 로보고되며, 25,26 이연구에서혈청 IgM 검사의민감도와특이도는각각 85.5%, 82.1% 로기존연구들에비 하여높은것으로나타났으나, 혈청 IgM 검사기법과 gold standard 에연구들마다차이가있어명확한비교에는제한점이있다. 혈청 PLR Table 4. Propensity score matching and multivariable Analysis of clinical features and laboratory findings of Mycoplasma pneumoniae PCR positive pneumonia Characteristic Adjusted odds ratio 95% CI P-value Crude analysis IgM positivity 27.09 8.80 99.34 < 0.001 Matched analysis* IgM positivity 13.50 3.69 60.68 < 0.001 Multivariable analysis Fever duration before admission 1.60 1.11 2.31 0.013 Unilateral infiltration 3.50 0.90 13.57 0.070 Extrapulmonary symptoms 11.40 1.07 121.10 0.044 O2 need 0.12 0.01 1.12 0.062 Eosinophil 1.25 0.93 1.67 0.140 IgM titer (index) 2.61 1.44 4.73 0.002 PCR, polymerase chain reaction; CI, confidence interval. *Matched on age, fever before admission, cough before admission, lobar/segmental pneumonia, unilateral infiltration, O2 need, extrapulmonary symptoms, Lymphocyte, eosinophil. Adjusted for fever before admission, cough before admission, lobar/segmental pneumonia, unilateral infiltration, O2 need, extrapulmonary symptoms, Lymphocyte, eosinophil. 마이코플라스마 IgM 은발병기간에따라혈청역가가증가하며감 염일주일이후부터의미있게증가하는것으로알려져있는데, 26 이연구에서혈청 IgM 양성군의평균발열기간은 5.5 일, 평균기침 기간수는 5.9 일로증상발생후수일이경과한이후에혈청 IgM 의 측정이이루어졌기때문으로추측할수있으며, 전체환자군을대 상으로한혈청 IgM 과발열기간간의 Pearson 상관관계분석에서 도의미있는양의상관관계를나타냈다 (Fig. 2). 급성감염의지표로이용되는혈청 IgM 은감염초기에는나타나 지않을수있고, 재감염의경우 20% 까지위음성이보고되며, 다른 종과의교차반응에의한위양성이있을수있고현성감염이후 1 년까지도혈청내에존재할수있어특이도가낮다고알려져있 다. 27,28 수년에걸쳐마이코플라스마감염이반복적으로이루어진 성인환자의경우, 최근현성감염에도혈청 IgM 의상승이없을수 있으나, 소아의경우재감염의빈도가낮아성인에비하여 IgM 증 가가특징적이라고알려져있고, 29 이연구에서도중합효소연쇄반 응과혈청 IgM 검사의일치백분율이 84.4% 로높았으며 80% 이상 의높은민감도와특이도를보였다. 또한마이코플라스마중합효소 연쇄반응을 gold standard 로한 ROC 분석에서혈청 IgM index 값 은 1.05 에서가장높은민감도 (83.9%) 와특이도 (85.7%) 를보여현 재 ELISA 의양성판정기준인 index 0.9 보다다소높은수치를적 용하는것이혈청학적진단에효과적이라고할수있다. 마이코플라스마중합효소연쇄반응은특정유전자를검출하여 균의존재를확인할수있고, ATPase, P1 adhesin, 16S rrna region 등이 primer 로사용되며 19,29 이연구에서는 ATPase 유전자를 이용하였다. 급성감염후수년까지위양성을보일수있으며감염 252 https://doi.org/10.4168/aard.2018.6.5.248
이혜진외 중합효소연쇄반응및 IgM ELISA 를통한소아마이코플라스마폐렴의조기진단 과보균상태를감별할수없다는단점이있으나, 29 한번의검체채취를통하여비교적높은민감도와특이도를가지며, 항체형성이전시기부터검출가능하므로혈청학적검사보다조기진단이가능하다는장점이있다. 중합효소연쇄반응검사는구인후면봉검체, 비인후흡인액, 객담등검체의종류에따라민감도가 78% 92% 까지다양하게보고되었다. 25,30 이연구에서이용한구인후면봉검체의경우, 소아환자에서는기타검사방법에비하여비교적쉽게검체를얻을수있다는장점이있고, 객담검체에비하여더높은민감도를보이기도하였다. 31 또한비인후검체와의비교에서는, 구인후면봉검체가하기도검체에가깝고균의집락율이높으며, PCR 저해 (inhibition) 를일으킬수있는기타물질의함량이적어비인후검체에비하여민감도가높다고보고되었다. 32,33 이번연구에서도 58명의 IgM 양성환자중 PCR 음성결과를보인환자는단 5명 (8.6%) 이었고, 이중한명의환자를제외하면 IgM index 값이 1.1 2.2 사이로낮았으며 3명의환자에서바이러스가검출되었으므로, 혈청 IgM의위양성가능성을배제할수없다. 비전형폐렴의임상양상을토대로마이코플라스마폐렴을조기진단하기위한연구들이이루어졌다. Japanese Respiratory Society scoring system에관한연구에는환자의연령, 동반질환, 기침양상, 청진소견, 세균감염의증거, 일반혈액검사등과같은 6개의임상지표를종합하여마이코플라스마폐렴을세균성또는기타바이러스폐렴과조기에감별할수있다고보고하였다. 21 이번연구에서살펴본바에의하면, 마이코플라스마중합효소연쇄반응양성폐렴환자에서입원전및총발열기간, 대엽성및분절성폐렴, 한쪽폐엽의침범, 폐외동반증상, 말초혈액호산구분율등이통계적으로의미있는임상적특징이었으며, 다변량회귀분석결과입원전발열기간은중합효소연쇄반응과혈청 IgM 양성을기준으로하였을때두경우모두마이코플라스마폐렴에서유의하게긴것으로나타났다. 이를토대로 60.7% 에서바이러스가검출된마이코플라스마음성폐렴군에비하여, 마이코플라스마양성폐렴군에서특징적임상양상이존재함을알수있었다. 입원당시마이코플라스마폐렴이의심되고발열기간이긴환자들을대상으로선택적으로스테로이드를투여하였으며, 중합효소연쇄반응양성군과 IgM 양성군의경우스테로이드를사용한환자가각각 75.8%, 79.3% 로음성군 50.0%, 46.9% 에비하여높았다. 전통적으로는기관지폐렴또는간질성, 망상소견이마이코플라스마에서특징적이라고알려져있으나 29,34 소아의세균성폐렴의빈도가감소함에따라대엽성폐렴에서마이코플라스마가차지하는비율이증가하고있으며, 국내연구에따르면 2015년대유행에서는마이코플라스마폐렴환자중대엽성폐렴의비율이 73.4% 까지보고된바있다. 35 따라서이연구결과는마이코플라스마폐렴에서대엽성, 분절성폐렴의경우긴발열기간, 폐외증상등합병증을 동반한심한임상경과를보인다는기존연구결과들과일치한다 고볼수있다. 3,36 이번연구는단일기관에서시행한연구로, 마이코플라스마중 합효소연쇄반응을 gold standard 로하였으며배양법과같은확진 검사를시행하지않았으므로중합효소연쇄반응의위음성가능성 을배제할수없다는한계점이있다. 현재까지존재하는마이코플 라스마의다양한진단방법중어느한가지로모든위양성과위음 성을배제하기는어렵기때문에, 많은연구들에서두가지이상검 사의조합을통하여진단하기를권고하고있으며, 25,37,38 한연구에 서는비인후흡인액 PCR 과혈청 IgM capture immunoassay 검사 의조합을통하여진단적민감도가 95% 까지증가한다고보고하였 다. 37 이연구에서는구인후면봉 PCR 양성마이코플라스마폐렴 환자를대상으로혈청 IgM ELISA 검사의진단적유용성을확인하 였고, index titer (>1.05) 를적용하는것이효과적임을확인하였다. 중합효소연쇄반응은혈청 IgM 이나타나기전시기의 PCR 양성환 자들을조기에진단가능하다는장점이있고, IgM index titer 를적 용하면 PCR 음성이면서위양성인환자를배제할수있으므로, 혈 청 IgM 과 PCR 두가지검사결과를모두고려하여진단할경우, 단 일결과를기준으로하는것에비하여보다많은수의환자를, 효과 적으로진단가능하다고할수있다. 기존연구에비하여인후면봉 검체 PCR 을시행하여 IgM 검사의진단적가치를알아보았다는 점, 혈청 ELISA IgM index cutoff 를제시하였다는점등은이연구 의차별점이라고할수있다. 따라서이번연구를통하여, 혈청 IgM ELISA index 검사는 PCR 양성마이코플라스마폐렴의진단에매우민감하고신속한검사 방법이며, 임상적으로유용하게사용될수있을것으로생각한다. REFERENCES 1. Liu WK, Liu Q, Chen DH, Liang HX, Chen XK, Chen MX, et al. Epidemiology of acute respiratory infections in children in Guangzhou: a threeyear study. PLoS One 2014;9:e96674. 2. He XY, Wang XB, Zhang R, Yuan ZJ, Tan JJ, Peng B, et al. Investigation of Mycoplasma pneumoniae infection in pediatric population from 12,025 cases with respiratory infection. Diagn Microbiol Infect Dis 2013;75:22-7. 3. Yang EA, Gang MH, You SY, Kim JH, Lee JH. Clinical characteristics of children with lobar pneumonia caused by Mycoplasma pneumoniae. Pediatr Allergy Respir Dis 2012;22:256-64. 4. Park GY, Lee YI, Shin M, Park JO, Kim CH. Clinical differences according to radiological patterns in childhood Mycoplasma pneumoniae pneumonia. 2013;1:362-9. 5. Kim KW, Kim KE. Mycoplasma and chlamydia infection in Korea. Korean J Pediatr 2009;52:277-82. 6. Izumikawa K, Izumikawa K, Takazono T, Kosai K, Morinaga Y, Nakamura S, et al. Clinical features, risk factors and treatment of fulminant Mycoplasma pneumoniae pneumonia: a review of the Japanese literature. J Infect Chemother 2014;20:181-5. 7. Pellan M, Bastian C, Gaudelus J, Delacourt C, de Pontual L. Pulmonary https://doi.org/10.4168/aard.2018.6.5.248 253
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