儿童EBV感染及相关疾病诊断(讲)

单击此处编辑母版标题样式,单击此处编辑母版文本样式,第二级,第三级,第四级,第五级,*,单击此处编辑母版标题样式,*,*,单击此处编辑母版文本样式,第二级,第三级,第四级,第五级,单击此处编辑母版标题样式,单击此处编辑母版文本样式,第二级,第三级,第四级,第五级,*,*,儿童EBV感染及相关疾病诊断(讲),常用名,正式命名,所属亚科,生物学特性,单纯疱疹V-1型,人疱疹V1型,在感染N节中潜伏,单纯疱疹V-2型,人疱疹V2型,在感染N节中潜伏,水痘-带状疱疹,人疱疹V3型,在感染N节中潜伏,EBV,人疱疹V4型,在淋巴细胞中潜伏,CMV,人疱疹V5型,在淋巴细胞及分泌腺体中潜伏,人疱疹V6型,人疱疹V6型,同上,人疱疹V7型,人疱疹V7型,同上,人疱疹V8型,人疱疹V8型,同EBV,人类疱疹病毒,Burkitts lymphoma,in Kenya,Epstein-Barr virus (EBV),1964, discovered from Burkitts lymphoma tissue by Epstein, Achong, and Barr,1968, the etiologic agent for infectious mononucleosis,1970, nasopharyngeal carcinoma,1980, non-Hodgkins lymphoma,EBV,双链DNA病毒，疱疹病毒科，,亚科，基因组,Genome: 172282 bp，有环状和线性两种形式,人群感染率高，我国35岁儿童95%已血清转化,EBV原发感染在婴幼儿及学龄前儿童主要为亚临床感染，在青少年和成人致IM（国外）,EBV与许多疾病相关,Circular form of the EBV genome (latent infection),Linear form of the EBV genome (lytic infection),EBV 相关疾病,传染性单核细胞增多症（Infectious mononucleosis，IM）,慢性活动性EB病毒感染（Chronic active EBV infection，CAEBV）,EB病毒相关性嗜血细胞综合征（EBV-associated hemophagocytic syndrome, EBVAHS),EBV相关疾病,伴性淋巴增殖综合征（X-linked lymphoproliferative syndrom）,鼻咽癌（Nasopharyngeal carcinoma）,Burkitt,s,淋巴瘤（Burkitt,s lymphoma）,何奇金淋巴瘤（,Hodgkins lymphoma,）,B,CD21,gp350,MHC-II,EBV,Asymptomatic infection,Symptomatic infection（IM),Primary EBV Infection,Latent infection,EBV-related other diseases,Saliva,Oropharynx,Lymphoid tissue and peripheral blood,Resting B cell,Epithelium,Lytic,EBV-infected,B cell,EBV-infected,B-cell blast,Reactivated EBV-infected,B-cell,Lytic,EBV-infected,B cell,EBV,EBV,Latently infected, resting memory B cells,Cytotoxic T cell,Cytotoxic T cell,Natural killer cell,LMP-1,LMP-1,LMP-2,LMP-2,LMP-2,LMP-2,Primary infection,Persistent infection,EBNAs,EBNAs,EBNA-1,Cohen, JI N Engl J Med 343:481-492, 2004,Model of EBV infection in humans,传染性单核细胞增多症（Infectious mononucleosis，IM）,IM,IM,嗜异白细胞阳性：EBV,嗜异白细胞阴性：EBV CMV Rub HHV6 Adv,EBV-IM的临床表现,发热： 约1周，严重者2周或更久，幼儿可不明显,淋巴结肿大：任何淋巴结，颈部最易受累,咽炎：,50有渗出物,，25上腭有瘀点,脾肿大：病程23周，50出现,肝炎：肿大1015，而GPT升高80,皮疹：红斑、斑丘疹或麻疹样疹，50可有眼睑浮肿,EBV-IM的临床表现,其它：,1.间质性肺炎,2.CNS: 脑炎、格林巴利综合征等,3.心肌炎,4.血液系统：溶血性贫血、再障、粒细胞减少,5.肾炎,6.关节炎,7.胰腺炎,IM的诊断,IM的拟诊：临床表现（发热,+,渗出性咽峡炎,+,淋巴结肿大、脾肿大）,+,异型淋巴细胞升高（10）,EBV-IM的诊断,嗜异白细胞凝集抗体,特异性EBV抗原的抗体检测:衣壳抗原（CA）IgM,荧光定量PCR检测外周血中EBV-DNA,嗜异白细胞抗体,IgM抗体,IM病人的血清在经吸收几内亚猪肾抗原后引起山羊红细胞的凝集,第12周出现，持续约6个月；小于5岁者，很可能阴性,外周血中EBV-DNA检测,普通PCR,荧光定量PCR：荧光定量PCR检测 EBV-DNA（血清、血浆、全血、外周血单核细胞），急性期（病程10天内）敏感性和特异性100,衣壳抗原（CA）IgM,一般情况下：一周左右升高，持续存在48周，类风湿因子和IgG抗体可致结果假阳性,临床要注意以下情况：,1、有的病人抗,EB,病毒,CA-IgM,产生延迟,2、少部分病人感染,EBV,后，,CA-IgM,持续阴性,3、也有的病人,CA-IgM,持续几个月阳性,抗体亲合力检测,机体在受到病原体入侵时首先产生低亲合力抗体，随感染的继续和进展，抗体亲合力升高。因此，低亲合力抗体的检出提示原发性急性感染。,原发EBV感染，100的病人在第一个月内可检测到抗EB病毒CA-IgG低亲合力抗体,原发性EBV感染后的免疫抗体反应,IM,EBV抗体四项,VCA-IgG,VCA-IgM,EA-IgG,NA-IgG,VCA-IgG亲合力,EBNA-IgG,阴性,VCA-IgG阳性,VCA-IgG阴性,未感染,VCA-IgM阳性,原发感染,VCA-IgM阴性,EAIgG强阳性,原发感染（ VCA-IgG 低亲合力）、感染后复发、或感染晚期抗EBNA抗体降低（ VCA-IgG 高亲合力）,EAIgG阴性,感染后复发或感染晚期抗EBNA抗体降低（ VCA-IgG 高亲合力）,EBNA-IgG阳性,仅VCA-IgG阳性,晚期感染,VCA-IgG阳性,和,EA-IgG强阳性,或,VCA-IgA强阳性,或,VCA-IgM阳性,感染后复发,Cervical lymphadenopathy,Cervical lymphadenopathy,Hepatosplenomegaly,Eyelid edema,Palatal petechiae,Atypical lymphocytes,慢性活动性EBV感染,CAEBV is characterized by severe, chronic or recurrent infectious mononucleosis-like symptoms after a primary EBV infection, and has a high morbidity and mortality from hepatic failure, lymphoma, sepsis, or hemophagocytic syndrome.,1. Unusual pattern of anti-EBV antibodies,(high levels of IgG anti-VCA and EA, absence of anti-EBNA),High EBV viral load in peripheral blood,Clonal,expansion of EBV-infected T cells and NK cells,Historical milestones of CAEBV,1948, Issacs: prolonged fever, malaise, lymphadenopathy, hepatosplenomegaly,1975, Horwitz et al:such clinical manifestations with mildly or moderately high or positive IgG against VCA and EA,1982, Tobi et al: similar atypical illness associated with serological evidence of persistent EBV infection,Historical milestones of CAEBV,1984, Dubois et al: criteria for such cases termed chronic mononucleosis syndrome:(1) disabling fatigue and malaise;(2) low-grade afternoon fever;(3) variable other nonspecific symptoms: myalgias, sore throat, depression,lasting 6 months or longer, with EBV serologies of (1)VCA-IgG,160, (2)EA-IgG 5, (3) postive anti-EBNA, (4)absent VCA-IgM, (5) absent Paul-Bunnell heterophil antibody.,发病机制,EBV感染的T细胞或NK细胞克隆性增殖,存在的问题：,1.如何感染T细胞或NK细胞,2.如何引起临床症状,EBV-infected cells in Japanese patients with CAEBV,临床表现,发热：间断性发热,淋巴结肿大,肝脾肿大,间质性肺炎,贫血,肝炎,眼葡萄膜炎,Diagnostic criteria of CAEBV,I. Severe illness of greater than 6 months duration that,:,1. Began as primary EBV infection OR,2. Is associated with grossly abnormal EBV antibody titers,(IgG to VCA1:5,120; antibody to EA1:640; or antibody,to EBNA1:2), AND,(Straus S.E.),II. Histological evidence of major organ involvement, such as:,1. Interstitial pneumonia,2. Hypoplasia of some bone marrow elements,3. Uveitis,4. Lymphadenitis,5. Persistent hepatitis,6. Splenomegaly, AND,III. Detection of increased quantities of EBV in affected tissues by,1. Anticomplementary immunofluorescence for EBNA, or,2. Nucleic acid hybridization,Reproduced from Straus S.E. (1988, J. Infect. Dis. 157:405_,/,412),CAEBV,CEBV: persistent IM-like illness with relatively good prognosis,SCEBV: rather severe manifestation with generally poor prognosis,Diagnostic criteria of a case definition for SCAEBV,Category,Criteria,Clinical,Hematologic,Virological,Other,Intermittent fever, lymphadenopathy, and hepatosplenomegaly,Anemia, thrombocytopenia, lymphocytopenia or lymphocytosis, neutropenia, and polyclonal gammopathy,Elevated antibody titers and positivity for antibodies to EBV-related antigens (VCA IgG5,120; VCA IgA, positive; EA D IgG640; EA D IgA, positive; and EA D and EA R IgG640) and/or detection of EBV genomes in affected tissues,Chronic illness which cannot be explained by other known disease processes,b,Reproduced from Okano M., et al. (1991, Clin. Microbiol. Rev. 4:129_,/,135),(Okano M),Revise of Criteria of CAEBV,2001,Kimura et al. extremely high antibody titers against EBV-replicative antigens are not absolutely necessary, but demonstrated significantly increased circulating EBV-DNA,Revised virological criteria: either or both extrmely against EBV-repicative antigens and/or increased genome copies in tissues.,伴性淋巴增殖综合征（X-linked lymphoproliferative syndrom,）,历史,1975年，Purtilo等发现一个家系中，18个男性有6个人出现良性或恶性淋巴细胞增生和组织细胞增加等征候，取名Duncan病,1998年，缺陷基因被确定：,SH2D1A/DHSP,or,SAP,(SLAM-associated protein,）,Called “Duncans disease”,after the family name,SH2D1A,编码含128个氨基酸的蛋白质- SAP (signaling lymphocytic activation molecule SLAM-associated protein),表达于活化的T和NK细胞表面，通过与SLAM及其他免疫球蛋白超家族如2B4等结合，参与信号传递，调节CTL的功能，如产生IFN-gamma 的能力,临床表现,家族史，仅见男性,发病年龄从6个月22岁（原发性EBV感染后）,IM样症状：发热、咽峡炎、淋巴结和肝脾肿大、异型淋巴细胞增加,免疫球蛋白异常：无,球蛋白血症、多克隆性高球蛋白血症,高IgM的免疫不全症,患者血清中EBV抗体阴性,临床分型,A型：属于急性致死性IM,多发病4周后死亡，占55,B型：同时有急性致死性IM和恶性淋巴瘤，占15,C型：EBV感染后免疫机能不全、低,球蛋白血症、骨髓增生低下、EBV抗体能力产生低下，占15,D型：无明显EBV感染表现而发生的恶性淋巴瘤，占15,诊断标准（Hamilton）,6个月至22岁男性有2个以上下述表现型,1. 增殖性改变,（1）有致死性或慢性IM,（2）有B免疫母细胞性淋巴肉瘤,（3）有非何杰金氏淋巴瘤,（4）IM继发高IgM免疫不全症,2. 非增生性改变,（1）粒细胞缺乏症或再生障碍性贫血,（2）,球蛋白异常：获得性无或低球蛋白血症,3. 先天异常,（1）心血管系,（2）中枢神经系,诊断标准,在母系直系亲属中有2人以上具备上述表现型者，可诊断本征,本征男性的B淋巴细胞体外感染EBV后，能自发的发育增殖；患者的唾液可使脐带血中的淋巴细胞发生形态改变；患者血清中缺乏EBV抗体,鉴别诊断,CAEBV: chronic active EBV infection,GLPD: granular lymphoproliferative disorder,ALPS: autoimmune lymphoproliferative syndrome,CAEBV,GLPD,XLP,ALPS,Heredity,?,?,X-linked,AD(AR),Sex,M,&,F,M,&,F,M,M,&,F,Gene localization,?,?,Xq25,10q23,Responsible gene,?,?,SH2D1A/SAP,Fas/FasL/caspase 10,Clinical manifestation,LPD,LPD,FIM/LPD/AGL,LPD,VAHS,+,+,+,+,Lymphadenopathy,+/malig benign,+/malig benign,+/malig,+/ benign malig,Immunoglobulin,High,High,Low,High,Autoimmune disease,+,-,-,+,EBV-antibodies,High,High,Low,Normal-high,EBV-genome,High,High,High,Normal-high,基因诊断,a monoclonal antibody, termed KST-3, against the XLP gene product, SAP.,Using a flow cytometric assay using KST-3,Shinozaki, K. et al. Int. Immunol.,14(10):1215-23, 2002,.,The patient exhibited markedly,deficient SAP expression,normal,porband,mother,father,小结,1、EBV感染与许多临床疾病相关，应引起临床医师的高度重视和警惕,2、临床诊断EBV感染要注意几个问题：,（1）是否感染EBV?,（2）感染的时期如何？,（3）是否活动感染？与本次临床表现是否有关？,临床检测结果的分析,1.,IgM阳性只能是近期感染的一个指标，而并不一定是急性期感染，更不能说某病原IgM阳性就是病原。通常IgM会持续46周或更长，如风疹、CMV等早期妊娠感染或先天性感染的儿童，特异性IgM可能持续达1年或更长时间。,2.,人类疱疹病毒如,CMV,、,EBV,等在儿童有一个血清阳性转化的问题，即隐性感染，同样会有,IgM,产生，但无临床表现。,3.,肠道病毒是夏季儿童呼吸道感染的主要病原，,PCR,结果提示阳性率在以上。（参考,2000,年秋冬至,2002,年夏北京地区急性呼吸道感染病毒病原学研究，临床儿科杂志，,2003,，,21,（,1,）：,25,28,）。,4.血清IgM检测结果阴性可能是：非该病原所致；标本采集太早；再感染或激活；免疫抑制病人。,5.任何实验均,存在一定的假阳性和假阴性，以及交叉反应的问题。,6.病毒室目前已基本建立室内质量控制体系，每年参加卫生部临床检验中心的室间质量评价，成绩合格。,以上解释供临床医师参考，有问题请联系病毒室，电话2892、2893。,Thanks,谢谢！,