pissn 2508-4887 eissn 2508-4895 Original article Vol. 28, No. 4, December, 2017 Associated Factors with Respiratory Virus Detection in Newborn with Sus pected Infection Jin-Hyeok Lee, MD, Sun-Young Cho, MD, Myo-Jing Kim, MD Department of Pediatrics, Dong-A University College of Medicine, Busan, Korea Received: 28 April 2017 Revised: 28 June 2017 Accepted: 27 September 2017 Correspondence to Myo-Jing Kim, MD Department of Pediatrics, Dong-A University College of Medicine, 26 Daesingongwon-ro, Seo-gu, Busan 49201, Korea Tel: +82-51-240-2589 Fax: +82-51-242-2765 E-mail: myojing@dau.ac.kr Copyright 2017 by The Korean Society of This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/ license/by-nc/4.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided that the original work is properly cited. Objective: The objective of this study was to determine the detection rate of respiratory viruses and investigate the associated factors with respiratory virus detection in newborn infants with suspected infection. Methods: From January 2013 to December 2015, respiratory virus real-time polymerase chain reaction (RT-PCR) results were obtained from 136 newborn infants aged 28 days who admitted to the neonatal intensive care unit (NICU) of Dong-A University Hospital with suspected infectious diseases. We performed a retrospective analysis of the detection rate of respiratory virus, classes of respiratory viruses, clinical characteristics, and social environment characteristics associated with respiratory virus detection. Results: Of the 136 infants, 36 infants (26.5%) had the 37 following respiratory viruses: Respiratory syncytial virus (n=23), Rhinovirus (n=10), Parainfluenza virus (n=2), Influenza virus (n=1), and Corona virus (n=1). The detection of respiratory viruses was significantly associated with the old age at admission (17.3±5.8 vs. 10.6±6.8 days), the presence of respiratory symptoms: cough (72.2% vs. 7%), rhinorrhea (63.9% vs. 10%), rale (16.7% vs. 1%), a family history of respiratory illness (38.9% vs. 13%), especially siblings respiratory illness (33.3% vs. 8%), and a seasonal preference (October-March) (80.6% vs. 50%) (P<0.01). Conclusion: Respiratory virus is an important pathogen in newborn infants admitted to the NICU, who are suspected with infectious diseases. Respiratory virus detection was associated with admission age, presence of respiratory symptoms, a family history of respiratory illness, and seasonality. Key Words: Newborn, Respiratory tract infection, Viruses 서론 신생아패혈증으로대표되는신생아감염은신생아기사망과장기이환율의주요원인이 다. 하지만임상증상, 검사실검사로는세균감염과바이러스감염을감별하는데에한계가 있다. 1-4 다른연령군에비해신생아의세균성감염은심각한합병증으로이어지기쉬워감 염이의심되는경우패혈증이배제될때까지경험적항생제치료를받는경우가많다. 5-8 신 생아기호흡기바이러스감염은대부분출생후가족이나다른감염자들과의밀접접촉에의 해발생하고경한임상경과를보이지만, 드물게는사망에이르는중증의질병경과를보일 수도있다. 9-12 또한영아기호흡기세포융합바이러스 (respiratory syncytial virus, RSV) 를 비롯한호흡기바이러스감염이이후반복적인천명, 천식등비정상적인폐기능과관련이 있다고보고되고있다. 13-17 따라서환자의임상경과이해와신생아집중치료실의감염관리 및환자격리, 부적절한항생제남용예방을위해감염이의심되는신생아에서조기에호흡 기바이러스감염을감별하는것이필요하다. 국내외에서호흡기바이러스실시간중합효소 연쇄반응 (real-time polymerase chain reaction, RT-PCR) 검사를통해호흡기바이러스의
2017 December;28(4):134-139 유행양상및감염의임상증상을확인하는연구들이있으나, 18-24 28일이전의신생아를대상으로한연구는드물다. 이에저자들은감염이의심되는생후 28일이내의신생아에서 RT-PCR 검사를통해호흡기바이러스검출빈도를알고, 신생아기호흡기바이러스감염과관련된연관인자들을알아보고자하였다. 대상및방법 유형태, 입원전거주환경, 입원시점등을조사하였다. 통계치는평균 ± 표준편차로표기하였으며, IBM SPSS Statistics 22 프로그램 (IBM Corp., Armonk, NY, USA) 을이용하여분석하였다. 모든통계적검정은양측검정을하였으며, P값이 0.05 미만인경우에만통계적유의성이있는것으로판단하였다. 독립집단인두군사이의연속형변수의평균분석은 Mann- Whitney U test를이용하였으며, 범주형변수의빈도와분율의비교는 Fisher s exact test를이용하였다. 2013년 1월부터 2015년 12월까지신생아감염이의심되어동아대학교병원신생아집중치료실에입원한 154명가운데호흡기바이러스 RT-PCR 검사를실시하였던 136명을대상으로하였으며, 전체대상군중컨디션저하로인한수유곤란이아닌, 구토, 구역, 설사등의뚜렷한위장관증상을보이면서입원과동시에시행한로타바이러스신속항원검사에서양성을확인할수있었던 18명은호흡기 RT-PCR 검사시행대상에서제외되었다. 발열, 호흡기감염증상 ( 기침, 콧물등 ), 호흡곤란증상 ( 무호흡, 빈호흡 >60회 / 분, 흉골함몰, 산소포화도감소, 신음소리, 청색증등 ) 이있거나, 호흡기질환의가족력이있는경우, 또는증상이모호하거나, 발열의원인이명확하지않은경우검사를시행하였다. 호흡기바이러스검사는입원 24시간이내에멸균된솜면봉 (flocked swab) 을이용하였으며, 비강을통해외비공부터귀까지해당하는길이를삽입하여비인두검체를채취하였다. 검사는신생아집중치료실에근무하는숙련된담당간호사가시행하였으며, 검체는채취즉시검사실로이송하였다. 핵산추출을위해 Viral Gene-Spin Viral DNA/RNA Extraction Kit (IntronBio, Seongnam, Korea) 를사용하였으며, Advansure RV real-time RT-PCR Kit (LG Life Sciences, Seoul, Korea) 와 SLAN Realtime PCR Detection System (LG Life Sciences ) 을통하여 RSV, Rhinovirus, Influenza virus, Parainfluenza virus, Human metapneumovirus, Adenovirus, Coronavirus, Bocavirus의감염여부를확인하였다. 입원에서결과확인까지평균 40시간이소요되었으며, RT-PCR 결과를바탕으로호흡기바이러스검출양성군과음성군으로분류하여호흡기바이러스검출과관련된특징들을비교분석하였다. 의무기록은표준화된양식으로기입된전자기록지를통해후향적으로분석하였다. 임상적인특징에대해알기위해재태주수, 성별, 출생체중, 분만방법, 입원시나이와체중, 임상증상및활력징후, 입원중산소치료여부, 항생제치료여부, 입원기간, 검사실결과등을조사하였다. 사회환경적인특징을알기위해호흡기질환의가족력, 형제자매의호흡기질환여부, 수 결과 1. 호흡기바이러스검출빈도 호흡기바이러스 RT-PCR 검사를실시하였던 136 명중 RSV, Parainfluenza virus 가동시에검출된 1 명을포함하여 36 명 (26.5%) 에서 37 건의바이러스가검출되었다. 검출된바이러스 는 RSV 23 건, Rhinovirus 10 건, Parainfluenza virus 2 건, Influenza virus 1 건, Corona virus 1 건으로 RSV 가가장많이검출되 었다 (Fig. 1). 이들중혈액, 소변, 대변등의다른검체에서세균, 바이러스등이검출된경우는 2 건으로 Escherichia coli 에의한 요로감염, 로타바이러스감염이각각 1 건씩이었으며, 2 건모두 발열이외의증상은없었다. RT-PCR 음성군 100 명중에서는혈액, 소변, 대변등의다른 검체에서세균, 바이러스등이확인된경우는 57 건이었고, 로타 바이러스감염이 40 건으로가장흔하였으며, 이중 21 건에서위 장관증상이있었다. 그외장바이러스감염 (6 건 ), 대장균요로 감염 (5 건 ), B 군연쇄상구균뇌수막염 (2 건 ), 균배양검사로확진 된메티실린내성포도상구균패혈증 (2 건 ), 연쇄상구균패혈증 (1 건 ), 장내구균패혈증 (1 건 ) 이었다. 이외구체적인세균, 바이 러스등이검출이되지않은경우는 43 건이었고, 흡인성폐렴, 폐렴, 위장관염, 감염성관절염, 말초정맥염, 배꼽염, 바이러스 Fig. 1. Respiratory virus detection classes. RT-PCR, real-time polymerase chain reaction; RSV, respiratory syncytial virus. www.e-kjp.org 135
Lee JH, et al. Respiratory virus detection in newborn 성뇌수막염의심등이포함되었다. 2. 호흡기바이러스검출과관련된인자 1) 임상적인특징 호흡기바이러스양성군과음성군으로나눠비교분석하였 다. 호흡기바이러스양성군은음성군에비해입원당시나이가 더많았다 (17.3±5.8 일 vs. 10.6±6.8 일, P<0.01). 입원당시임상 증상및신체진찰에서발열의빈도는차이가없었고, 호흡기바 이러스양성군에서기침 (72.2% vs. 7%), 콧물 (63.9% vs. 10%), 수포음 (16.7% vs. 1%) 의호흡기증상의빈도가의미있게많았 다 (P<0.01). 임상경과의비교에서산소치료빈도는차이가없 Table 1. Clinical Characteristics Related with Respiratory Virus Detection RT-PCR (+) (n=36) RT-PCR (-) (n=100) P-value Gestational age (weeks) 38.6±1.8 38.4±1.8 0.57 Prematurity 3 (8.3) 12 (12) 0.76 Male 18 (50) 62 (62) 0.24 Birth weight (g) 3,216.1±437.8 3,246.5±570.4 0.77 NSVD 25 (69.4) 63 (63) 0.55 Age at admission (days) 17.3±5.8 10.6±6.8 <0.01 Weight at admission (g) 3,611.7±556.5 3,394±672.8 0.09 Fever 16 (44.4) 55 (55) 0.33 Cough 26 (72.2) 7 (7) <0.01 Rhinorrhea 23 (63.9) 10 (10) <0.01 Irritability 2 (5.6) 8 (8) 1.00 Moaning 2 (5.6) 11 (11) 0.51 Chest retraction 6 (16.7) 33 (33) 0.09 Tachypnea 9 (25) 30 (30) 0.67 Desaturation 7 (19.4) 25 (25) 0.65 Apnea 1 (2.8) 6 (6) 0.68 Wheezing 1 (2.8) 1 (1) 0.46 Rale 6 (16.7) 1 (1) <0.01 Rash 0 7 (7) 0.19 Oxygen treatment 6 (16.7) 21 (21) 0.64 WBC (/mm 3 ) 10,963.1±4,848.4 12,797.8±5,635.0 0.09 ANC (/mm 3 ) 3,998.6±3,195.3 7,028.7±4,512.1 <0.01 Positive CRP 11 (30.6) 39 (39) 0.42 Antibiotics treatment 12 (33.3) 81 (81) <0.01 Hospital stay (days) 5.3±2.1 8.9±5.4 <0.01 Values are presented as mean±standard deviation or number (%). Abbreviations: RT-PCR, real-time polymerase chain reaction; NSVD, normal spontaneous vaginal delivery; WBC, whole blood count; ANC, absolute neutrophil count; CRP, C-reactive protein. 었고, 호흡기바이러스음성군에서입원기간이길고 (8.9±5.4 일 vs. 5.3±2.1 일 ), 경험적항생제치료 (81% vs. 33.3%) 를더많이 받았으며, 항생제투여기간 (4.0 일 vs. 1.6 일 ) 도길었다 (P< 0.01). 검사실결과에서혈액 C- 반응단백, 총백혈구수수치는 차이가없었고, 호흡기바이러스양성군의경우절대호중구수 가의미있게낮았다 (3,998.6±3,195.3/mm 3 vs. 7,028.7± 4,512.1/mm 3, P<0.01) (Table 1). 2) 사회환경적인특징 호흡기바이러스양성군의경우호흡기질환의가족력 (38.9% vs. 13%, P<0.01) 이의미있게많았고, 특히형제자매의호흡기 질환력 (33.3% vs. 8%, P<0.01) 이통계적으로의미있게많았다. 수유형태 ( 모유수유 / 혼합수유 / 분유수유 ) 나입원전거주형태 ( 집 / 신생아실및조리원 / 모름 ) 에따른차이는없었다 (Table 2). 발 병시기에따른비교에서전통적으로가을 / 겨울로여겨지는시기 (9-2 월 ) 에따른차이는없었고, 3 월에 10 건으로호흡기바이러스 가가장많이검출되었고, 11 월 6 건, 12 월 5 건, 2 월 4 건, 1 월 2 건 으로 10 월에서 3 월사이에집중적으로발생하였다 (Fig. 2). 고찰 본연구는감염증이의심되어입원하였던생후 28 일이내의 신생아에서호흡기바이러스검출빈도와바이러스의종류에 대해조사하였고, 당시임상양상과 RT-PCR 결과를비교분석 Table 2. Socio-Environment Characteristics Related to Respiratory Virus Detection RT-PCR (+) (n=36) RT-PCR (-) (n=100) P-value Family history of respiratory illness 14 (38.9) 13 (13) <0.01 Sibling history of respiratory illness 12 (33.3) 8 (8) <0.01 Having sibling 18 (50) 40 (40) 0.33 Feeding type 0.24 Breast milk 5 (13.9) 28 (28) Mixed feeding 23 (63.9) 54 (54) Formula feeding 8 (22.2) 18 (18) Stay before admission 0.12 Home 22 (61.1) 42 (42) Medium center 14 (38.9) 52 (52) None 0 6 (6) Season (October-March) 29 (80.6) 50 (50) <0.01 Values are presented as number (%). 136 www.e-kjp.org
2017 December;28(4):134-139 Fig. 2. Monthly distribution of respiratory virus detection cases. PCR, poly merase chain reaction; RSV, respiratory syncytial virus. 하여바이러스검출과관련된연관인자들을확인하였다. 신생 아감염증이의심되었던환자중호흡기바이러스가검출된환 아는 26.5% 였고, 검출된바이러스는 RSV, Rhinovirus, Parainfluenza virus 순이었다. 한가지이상의바이러스가공존한경 우는 1 예로이전연구결과에비해낮았는데, 23,24 이는대상환자 들이생후 28 일이내의신생아로국한되었기때문으로생각된 다. 신생아에서호흡기바이러스감염은발열, 빈호흡, 호흡곤란, 무기력, 수유부진등으로나타날수있어질병의초기에는신생 아패혈증과의감별이어렵다. 9 본연구에서호흡기바이러스양 성군의경우입원당시임상증상및신체진찰에서발열보다는 기침, 콧물, 수포음이더많았다. 검사실검사에서혈액 C- 반응 단백의양성률과절대값에는차이가없었으나, 호흡기바이러 스양성군에서절대호중구수치가유의하게낮았다. 모유수유여부는호흡기바이러스검출과관련이없었는데 이는모유수유가급성호흡기감염의발생빈도와이환기간을 줄인다는기존의연구와는상반된결과로, 25,26 대상환자들의평 균입원시점이생후 2 주전후로비교적짧은기간동안모유를 섭취하였기때문에모유의감염예방효과를정확히평가할수 없었을것으로여겨진다. 또한최근산후조리원문화가확산되 고, 조리원내바이러스집단발병들이보고되고있어 27 입원전 거주형태에따라호흡기바이러스검출을비교분석하였으나, 신생아실이나조리원등집단시설내거주에따른호흡기바이 러스검출률은차이가없었다. 반면, 가족중에호흡기질환이 있는경우, 특히형제자매가호흡기질환이있었던경우뚜렷하 게호흡기바이러스검출이많아신생아호흡기바이러스감염 의경우밀접하게접촉하는가족구성원에의해발생하는것을 알수있었다. 호흡기바이러스양성군에서입원당시나이가더 많았다는점역시바이러스접촉기회가더증가하는것으로설 명가능하다. 바이러스검출시기가전통적으로호흡기바이러스유행시 기로여겨지는가을 / 겨울 (9-2월) 보다오히려 3월에가장많은바이러스가검출되었고, 발병시기가 10-3월사이인경우의미있게바이러스검출이더많았다. 이는최근기후변화로인해 9월까지의기온이높아지고, 3월까지낮아진결과로호흡기바이러스감염위험시기의변화를잘반영한다고할수있으며, 호흡기바이러스의유행을예측하고, 계절에따른호흡기감염의예방및치료전략을세우는정보가될수있다. 또한호흡기바이러스양성군에서경험적항생제치료빈도가적었고, 치료기간과입원기간이짧았다. 입원당시호흡기바이러스감염의주증상인기침, 콧물, 수포음등의임상증상및가족의호흡기질환력등의병력그리고신속하게확인가능한 RT-PCR 검사결과를바탕으로바이러스감염을판단하였다. 경험적항생제치료에대한일반적인지침은병력에서호흡기감염증이의심되는경우, 호흡기감염증상이뚜렷한경우는경험적항생제치료를시작하지않거나, 다른감염증을배제할수없어경험적항생제치료를시작한경우는 RT-PCR 결과가양성이확인된경우항생제사용을중단하였다. 반면감염증의임상증상이있으면서 RT-PCR 검사에서바이러스가검출되지않는경우혈액배양검사를통해세균감염이배제될때까지경험적항생제투여를유지하였다. 그결과호흡기바이러스양성군에서경험적항생제치료빈도와치료기간과그에따른입원기간을줄일수있었던것으로생각된다. 하지만근무당시진료의사의판단에따른차이가있을수있고, 호흡기바이러스음성군에서일정기간이상항생제치료를필요로하였던요로감염, 패혈증등의감염증이있었던경우가포함되어있어이에따른항생제치료기간의영향도고려할수있다. RT-PCR 양성결과가직접적으로급성호흡기바이러스감염을의미하는지에대한의문이있다. Non-viable organism이나소량의바이러스량도검출가능한 RT-PCR 검사의특성상무증상기나회복기에바이러스량이감소하여도결과가양성일수있기때문이다. 28,29 이런위양성의가능성을보완하기위해본연구에서는감염증의임상증상이동반된경우로대상환자를한정하였다. 또한채취자간채취방법이다를수있어바이러스검출의오차가있을수있는데본연구에서는숙련된신생아집중치료실담당간호사가표준화된방법으로직접검체를채취하여오차를줄이고자하였다. 호흡기바이러스는감염에서임상증상의발현까지 24시간에서수일이소요될수있어언제어디서감염이되었는지에대해정확하게평가할수없다는점역시한계이다. 이러한한계들은항생제투여및중단을결정하는과정에도영향을미치게된다. 실제로세균감염과바이러스감염의동시감염등의가능성을고려하여혈액배양검사결과까지확인한후항생제투여를중지하였던경우도있었다. www.e-kjp.org 137
Lee JH, et al. Respiratory virus detection in newborn 또한임상증상이명확하지않을수있는신생아감염의특성상입원시점에 RT-PCR 검사의시행여부를결정하기쉽지않다. 호흡기증상이명확하지않거나, 재원기간중뒤늦게타장기증상이발현된경우, 병력청취과정중호흡기질환의가족력이정확하게확인되지못한경우등호흡기질환여부를가늠하기힘든상황에서는원인감별을위해 RT-PCR 검사가필요한상황이생기게된다. 본연구에서도임상증상으로판단하기힘든다수의환아에서 RT-PCR 검사를시행하는경향을보였다. 입원시점에서의자세한병력청취와정확한의료진의판단이불필요한검사를줄일수있다. 신생아감염이의심되는경우확인즉시독립된공간으로격리하여외부로의병원체전파가능성을차단하고, 이후검사결과및임상양상에따라격리해제를포함한격리조치를변경하는수순이일반적인격리원칙이다. 실제로신생아집중치료실격리실에는다양한원인의감염환아가입원하여있으며, 각각원인병원체에따라구분되어있다. 대상환아들은호흡기바이러스 RT-PCR 결과가양성으로확인되면다른감염증환아와구분되어격리되었으며, 비말감염등의전파형태에따른세부원칙에따라관리하였다. RT-PCR 음성군에서많은수가분변- 경구감염형태의로타바이러스감염등인것을감안하였을때호흡기바이러스 RT-PCR 결과에따른격리조치의변화는감염전파예방을위한구체적인치료전략이될수있다. 다만각집중치료실의시설, 인력, 비용등의한계로병원체의종류나환아컨디션을고려하는수준까지의세밀한격리가어려울수있다. RT-PCR 검사양성이확인된대상환아중인공호흡기치료까지필요한경우도 1건있었으며, 그외높은수준의집중치료가필요한환아가다수있어시설및인력이필요함을감안할때신생아감염관리에대한구체적인논의와지원이수반되어야한다. 본연구를통해감염이의심되는신생아에서많은경우호흡기바이러스가원인이되며, RT-PCR을통한호흡기바이러스검출이신생아감염증의유용한진단방법임을알수있었다. 입원당시호흡기증상이나호흡기질환의가족력이있는경우호흡기바이러스검출이유의하게높았다는점을유념하여초기병력청취나신체진찰을시행하고, 호흡기바이러스 RT-PCR 검사를시행하는것이필요하다. 이를통해치료기간동안의불필요한항생제사용을줄일수있으며, 환자의임상경과및예후를예측하고, 효과적인환자격리방안을마련할수있다. References 1) Gerdes JS. Diagnosis and management of bacterial infections in the neonate. Pediatr Clin North Am 2004;51:939-59. 2) de Assis Meireles L, Vieira AA, Costa CR. Evaluation of the neonatal sepsis diagnosis: use of clinical and laboratory parameters as diagnosis factors. Rev Esc Enferm USP 2011;45:33-9. 3) Polin RA; the Committee on Fetus and Newborn. Management of neonates with suspected or proven early-onset bacterial sepsis. Pediatrics 2012;129:1006-15. 4) Simonsen KA, Anderson-Berry AL, Delair SF, Davies HD. Early-onset neonatal sepsis. Clin Microbiol Rev 2014;27:21-47. 5) van den Anker JN. How to optimize the evaluation and use of antibiotics in neonates. Early Hum Dev 2014;90 Suppl 1:S10-2. 6) Muller-Pebody B, Johnson AP, Heath PT, Gilbert RE, Henderson KL, Sharland M, et al. Empirical treatment of neonatal sepsis: are the current guidelines adequate? Arch Dis Child Fetal Neonatal Ed 2011;96:F4-8. 7) Jacqz-Aigrain E, Zhao W, Sharland M, van den Anker JN. Use of antibacterial agents in the neonate: 50 years of experience with vancomycin administration. Semin Fetal Neonatal Med 2013;18:28-34. 8) Edmond K, Zaidi A. New approaches to preventing, diagnosing, and treating neonatal sepsis. PLoS Med 2010;7:e1000213. 9) Tregoning JS, Schwarze J. Respiratory viral infections in infants: causes, clinical symptoms, virology, and immunology. Clin Microbiol Rev 2010; 23:74-98. 10) Diniz EM, Vieira RA, Ceccon ME, Ishida MA, Vaz FA. Incidence of respiratory viruses in preterm infants submitted to mechanical ventilation. Rev Inst Med Trop Sao Paulo 2005;47:37-44. 11) Bennett NJ, Tabarani CM, Bartholoma NM, Wang D, Huang D, Riddell SW, et al. Unrecognized viral respiratory tract infections in premature infants during their birth hospitalization: a prospective surveillance study in two neonatal intensive care units. J Pediatr 2012;161:814 8. 12) Piippo-Savolainen E, Korppi M. Wheezy babies-wheezy adults? Review on long-term outcome until adulthood after early childhood wheezing. Acta Paediatr 2008;97:5-11. 13) García-García ML, Calvo C, Casas I, Bracamonte T, Rellán A, Gozalo F, et al. Human metapneumovirus bronchiolitis in infancy is an important risk factor for asthma at age 5. Pediatr Pulmonol 2007;42:458-64. 14) Jackson DJ, Gangnon RE, Evans MD, Roberg KA, Anderson EL, Pappas TE, et al. Wheezing rhinovirus illnesses in early life predict asthma development in high-risk children. Am J Respir Crit Care Med 2008;178: 667-72. 15) Lemanske RF Jr, Jackson DJ, Gangnon RE, Evans MD, Li Z, Shult PA, et al. Rhinovirus illnesses during infancy predict subsequent childhood wheezing. J Allergy Clin Immunol 2005;116:571-7. 16) Fjaerli HO, Farstad T, Rød G, Ufert GK, Gulbrandsen P, Nakstad B. Acute bronchiolitis in infancy as risk factor for wheezing and reduced pulmonary function by seven years in Akershus County, Norway. BMC Pediatr 2005;5:31. 138 www.e-kjp.org
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