Molecular epidemiology of enterovirus 71 infection in the Western Pacific Region

BACKGROUND: Recently, there have been large outbreaks of hand, foot and mouth disease (HFMD) mainly caused by enterovirus 71 (EV71) associated with severe neurological diseases in the Western Pacific Region (WPR). To monitor the realtime trend of EV71 transmission throughout the WPR, the authors conducted a molecular epidemiological analysis of EV71 infection. METHODS: Viruses were isolated from clinical samples from patients with HFMD or those with neurological complications. The EV71 isolates were identified by microneutralization assay. The VP4 and/or VP1 regions of recent EV71 isolates were sequenced and subjected to phylogenetic analysis using reference EV71 strains. RESULTS: The phylogenetic analysis of EV71 isolates from the WPR revealed two major genogroups, B and C, based on the nucleotide sequence alignment of the VP1 or VP4 region. These two major genogroups were further divided into subgenogroups, B1, B2, B3, and B4 and C1, C2, C3 and C4, respectively. CONCLUSIONS: The molecular epidemiological analyses of recent and previous EV71 isolates in the WPR indicated that two major genogroups of EV71 are co-circulating in Australia, Malaysia, Singapore, Taiwan and Japan. Recent EV71 isolates in Mainland China constitute a new distinct genetic cluster, subgenogroup C4. Two major lineages of EV71 are the major causative agents of the present HFMD epidemics in the WPR and both are considered to be neurovirulent.     

Critical management in patients with severe enterovirus 71 infection

OBJECTIVE: The aim of this study was to analyze clinical details occurring in children with severe enterovirus 71 (EV71) infection and synthesize the critical care experience for patients with severe EV71 infection. METHODS: A retrospective clinical, laboratory, and hemodynamic study was performed in a pediatric intensive care unit in a university hospital. From March 1998 to April 2000, seven consecutive pediatric patients with severe EV71 infection were retrospectively analyzed as the comparison group. From May 2000 to March 2003, eight consecutive patients with severe EV71 infection who had received the protocol therapy were enrolled as the study group. Detailed information about clinical treatment and pharmacological therapy was collected for comparison. RESULTS: The clinical presentations and laboratory findings between the comparison and the study groups were not significantly different. The amount of intravenous fluid in the first 24 h was significantly higher in the comparison group (9.2+/-5.0 vs 4.9+/-1.3 mL/kg per h). More patients in the study group received low doses of dopamine infusion, patients in the comparison group received more epinephrine, and none of them received milrinone. The acute-stage and long-term survival rates were higher in the study group (100% vs 43%, 87% vs 29%). CONCLUSION: Early cardiopulmonary support may prevent the vicious cycle of cardiopulmonary failure and improve the clinical outcome of severe EV71 infection. Milrinone may be the ideal inotropic agent for these patients. Echocardiography, a central line, and an arterial line could be an alternate method to replace direct intracardiac hemodynamic monitoring for guiding critical management.     

Different proinflammatory reactions in fatal and non-fatal enterovirus 71 infections: implications for early recognition and therapy

AIM: The mechanism of pulmonary oedema, a life-threatening manifestation of enterovirus 71 (EV71) encephalitis, is unclear. Our aim was to assess the relationship of proinflammatory cytokines to EV71-related pulmonary oedema. METHODS: Proinflammatory responses in 33 EV71 patients with various complications and 21 normal healthy children were measured using an enzyme-linked immunosorbent assay. RESULTS: EV71 patients with both encephalitis and pulmonary oedema were found to have much higher levels of blood interleukin-6 (IL-6) (947 +/- 1239 vs 4.9 +/- 3.1 pg/ml, p = 0.0003), tumour necrosis factor-alpha (TNF-alpha) (22.4 +/- 29.5 vs 5.3 +/- 1.0 pg/ml, p = 0.0035), interleukin Ibeta (IL-1beta) (48.4 +/- 85.2 vs 4.9 +/- 10.1 pg/ml, p = 0.01), white blood cell count (28.3 +/- 7.6 vs 15.5 +/- 6.8 10(9)/L, p > or = 0.0001) and blood glucose (501 +/- 186 vs 165 +/- 117 mg/dL, p = 0.0009) than patients with EV71 encephalitis alone. In fact, the cytokine levels in patients with encephalitis only or in those without complications were not significantly different from the levels found in normal children. The sensitivity, specificity, positive and negative predictive values of IL-6 > 70 pg/ml for EV71 encephalitis with pulmonary oedema were all 100%. CONCLUSION: Patients with EV71-related encephalitis combined with pulmonary oedema were found to have significantly elevated levels of proinflammatory cytokines and the best predictor for this complicated condition was found to be the level of serum IL-6.     

肠道病毒71型感染患儿免疫功能探讨

目的 探讨肠道病毒71型(EV71)感染患儿免疫功能变化与病情程度的关系.方法 患儿46例,健康同龄儿童12例,根据病情由轻到重将患儿分为4组:手足口病组11例、中枢神经系统病变组20例、中枢神经系统病变伴自主神经功能失调组10例、神经源性肺水肿组(pulmonaryedema,PE组)5例.进行下述检测:CD14~+单核细胞人类白细胞DR抗原(HLA-DR)表达率、淋巴细胞免疫分型、CD4~+CD25~+Foxp3~(high)调节性T细胞(regulatory T cells,Treg cells)及TH17细胞比例;白细胞介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)、白细胞介素10(IL-10)、转录生长因子β(TGF-β)、白细胞介素6(IL-6)、白细胞介素17A(IL-17A)血浓度;CD~4T细胞Foxp3、ROR-γt基因表达;血清免疫球蛋白及补体等.结果 (1)前炎症细胞因子TNF-α及IL-1β在轻症患儿中增高,随病情加重而下降,PE组明显降低(P<0.05);抗炎细胞因子IL-10及IL-10/TNF-α比值随病情加重增高,PE组增高明显(P<0.05).(2)HLA-DR、CD3~+T细胞、CD4~+T细胞、CD8~+T细胞、NK细胞随病情加重呈现逐步下降趋势,PE组下降最为明显(P<0.05).各组间B淋巴细胞及抗体差异无统计学意义.(3)Treg细胞比例、转录因子Foxp3 mRNA及诱导因子TGF-β血浓度随病情加重降低,而TH17细胞比例、IL-17A血浓度、转录因子ROR-γt mRNA及诱导因子IL-6血浓度随病情加重升高.结论 EV71感染患儿机体免疫功能随病情程度而变化,轻症患儿处于全身炎症反应状态,重症或危重症病例处于代偿性抗炎症反应或混合性拮抗反应状态,对EV71感染的免疫调控治疗应强调分阶段、个体化.     

肠道病毒71型感染致重症脑干脑炎的临床特征和治疗

目的：分析肠道病毒71型（EV71）感染所致重症脑干脑炎的临床特征及治疗方法。方法：回顾性分析2008年5月至2008年12月32例EV71感染所致重症脑干脑炎的住院患儿病例资料。结果：32例中3岁以下的婴幼儿28例（88%）,主要表现为持续发热（>38.5℃）,频繁呕吐,四肢抖动。8例在病程3~4 d病情迅速恶化,突发心动过速、呼吸喘促、四肢末端凉等交感神经亢进征象,直至口鼻腔涌出血性泡沫痰,发生神经源性肺水肿或肺出血。给予积极降颅压,静脉点滴丙种球蛋白和甲基强的松龙抑制炎症反应,使用血管活性药物,保护心功能,限制液体入量,早期气管插管正压通气等治疗,痊愈23例,好转4例,放弃治疗2例,死亡3例。结论：EV71感染所致脑干脑炎多有突出的植物神经功能损害的临床特征,早期识别并正确处理神经源性肺水肿是救治成功的关键。［中国当代儿科杂志,2009,11（12）：967-969］     

肠道病毒71型疫苗研究进展

手足口病(HFMD)是一种婴幼儿常见的传染性疾病。肠道病毒71型(EV-A71)是HFMD的主要病原之一,可导致严重并发症甚至造成死亡。因此,预防EV-A71感染尤为必要。开发EV-A71疫苗是预防、控制EV-A71感染最有效的方法。正在研发的EV-A71疫苗主要包括:灭活全病毒疫苗、减毒活疫苗、病毒样颗粒疫苗、重组VP1蛋白疫苗和合成多肽疫苗。现就EV-A71候选疫苗的开发和进展作一综述。     

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目的研究广州地区2011年手足口病病原流行特点及EV71的基因特征。方法收集2011年1～11月广州市妇女儿童研究中心63份不同患者来源的肛拭子、咽拭子以及血液样本,选择12份EV71阳性标本进行病毒分离,扩增EV71VP1区,测序并与EV71各血清型代表株序列比对,进行进化分析。结果 2011年广州地区12个临床分离株同A、B亚型的亲缘性较远,核苷酸同源性都不足85%,氨基酸同源性不足96.5%。其同C亚型的亲缘型较近,核苷酸、氨基酸同源性超过88.5%和97.5%,尤其与C4亚型的亲缘性最近,核苷酸和氨基酸的同源性分别为92.9%～94.6%和97.9%～98.9%。结论 2011年广州地区的12株EV71分离株都属于C4a亚型进化分支,并处于选择进化过程中。     

肠道病毒71型感染手足口病合并急性弛缓性麻痹临床分析

目的探讨肠道病毒71型感染手足口病合并急性弛缓性麻痹的临床特点。方法对10例肠道病毒71型感染手足口病合并急性弛缓性麻痹的患儿进行临床观察,并作病原学、头颅和脊髓磁共振成像、神经电生理及脑脊液检查。结果 10例中8例为2岁以下儿童,瘫痪前期均伴发热和皮疹,单侧肢体瘫痪占70%,1周左右患肢运动功能开始恢复,轻症多于1～3个月完全恢复。磁共振成像及神经电生理检查结果与临床症状具有高度一致性,提供了神经受累的定位证据。所有病例随访4～12周,7例(70%)肌力恢复至Ⅴ级。结论急性弛缓性麻痹是肠道病毒71型感染手足口病的严重并发症,磁共振成像及神经电生理检查对评估病情及预后有重要价值。     

肠道病毒71型感染9例临床及尸体解剖特点

目的了解肠道病毒71型(EV71)感染的危重病例的临床特点及尸体解剖病理特点。方法收集2010年3-6月桂林市7家医院收治的9例EV71感染死亡病例的临床资料,并对患儿尸体系统解剖,对脑、肺、肠、心、肝、胃、脾、肾等脏器行组织常规HE染色,光镜下观察。结果患儿年龄7个月～3岁。临床特点为病程短、进展快。6例病程中手足口或肛周有手足口样皮疹,3例无皮疹。9例患儿均在发病第1-2天出现神经系统症状,表现为精神差,呕吐,肌张力下降,抬头不稳、行走不稳,肢体抖动;3例在病程中有抽搐。病程第3-4天,9例患儿均出现肺水肿、肺出血而死亡。尸解结果显示其脑、肺部病变明显。脑部主要累及蛛网膜、脑干、软脑膜、脊髓、大脑、脊髓膜等处,表现为水肿、炎症、坏死,有明显的软化灶、出血灶形成;肺部主要为肺淤血、肺水肿及肺出血;心脏无炎症及心肌炎的表现;肝、脾、胃肠道、肾等器官病理改变不明显。结论 EV71感染危重病例的病变主要累及中枢神经系统,呼吸系统为继发病变;严重的脑干脑炎和神经源性肺水肿、肺出血是死亡的主要病因。     

肠道病毒71型脑干脑炎二例尸检报道及文献复习

目的 探讨手足口病的临床与病理特征.方法 分析2例肠道病毒71型(EV71)脑干脑炎死亡病例尸检结果及临床资料,并复习相关文献.结果 死者年龄≤3岁,病程短,进展快,1例有典型皮疹,1例皮疹不明显,2例均有不同程度的神经系统症状,最终出现肺水肿和肺出血死亡.尸检显示:病变主要在大脑和脑干,可见明显水肿、炎症、坏死等;肺脏主要为水肿和出血;心肌无炎细胞浸润及心肌炎的改变,肝、脾、肾、胰腺及胃肠道病理改变不明显.结论 本组2例尸解结果显示神经系统、呼吸系统病变明显.支持EV71感染的手足口病,常并发中枢神经系统损害;严重的脑干脑炎和神经源性肺水肿、肺出血是死亡的主要原因;也可以解释患儿以咳嗽、气促、抽搐、昏迷为主的临床表现.     

实时荧光定量RT-PCR法检测手足口病患儿大便标本中肠道病毒71型

目的 了解2008年手足口病流行期间住院手足口病患儿是否存在EV71病毒感染,并探讨实时荧光定量RT-PCR法对手足口病患儿大便标本中EV71病毒载最进行定量检测的可行性.方法 采用RT-PCR荧光探针体外扩增法对47例住院的手足口病患儿大便标本中抽提的RNA进行抽样检测.结果 22例(46.81%)手足口病患儿大便标本中EV71的病毒载量大于103 copies/ml,最高者达1.03 × 107copies/ml.实时荧光定量RT-PCR法其标准曲线显示,Ct值与病毒拷贝数的对数(log10)之间的相关系数为1.000,相关性较好.结论 2008年手足口病流行期间入住儿科医院的手足口病患儿近半数为EV71病毒感染.实时荧光定量RT-PCR法对大便标本中的EV71 RNA定量检测较方便快捷,结果直观,为进一步研究病毒载量与临床表现之间的关系打下了基础.     

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??Abstract??Objective??To study the clinical features of hand-foot-mouth disease ??HFMD?? caused by enterovirus 71 ??EV71?? infection. Methods Clinical data of hospitalized children with hand-foot-mouth disease caused by EV71 infection from May 2010 to August 2010 were analyzed retrospectively. The difference of clinical manifestation and results of auxiliary examination between intensive HFMD group and serious HFMD group were compared. Results??High fever and nontypical skin rash showed significant difference between intensive group and serious group??P = 0.002??P = 0.000??respectively???? 120 EV71-positived HFMD cases ??99.17%?? showed neurological impairments. The major neurological features included fatigue ??84.30%????frequent vomiting ??65.30%????limb tremble ??60.33%?? and sleep disorders ??53.72%??. The rate of abnormal knee reflex was 52.07% in physical examination. The incidence of vomiting??P = 0.001????unconsciousness ??P = 0.000????abnormal muscular tension??P = 0.000????abnormal heart rate ??P = 0.000????dysarteriotony ??P = 0.000????capillary refill time being more than 3 seconds ??CRT > 3 s?? ??P = 0.000????tachypnea or dyspnea ??P = 0.000?? and pulmonary exudative lesion in chest X-ray ??P = 0.000?? was morefrequent in serious group compared with intensive group??There were 51 cases ??42.15% ?? with a peripheral blood WBC count of more than 12 × 109/L or less than 4×109/L??52 cases ??42.98% ?? with blood glucose level of more than 6 mmol/L and cardiac troponin I elevated in 22 cases ??18.18%??. The above three indexes were significantly different between two groups ??P < 0.000??respectively??. Conclusion??HFMD caused by EV71 infection often shows neurological impairments. High fever??nontypical skin rash??frequent vomiting??unconsciousness??abnormal muscular tension??abnormal heart rate??dysarteriotony??CRT > 3s??tachypnea or dyspnea??and pulmonary exudative lesion are risk factors of serious HFMD. Early identification and correct treatment are the key to the rescue of serious HFMD.     

肠道病毒71型感染患儿胸X线片表现

目的：研究肠道病毒71型感染患儿的胸部X线影像特点。方法：选取2010年4月至2011年7月肠道病毒71型感染患儿120例,按病情分成轻型组（31例）、重型组（43例）、危重型组（46例）,对患儿发病至首次拍片时间以及首次胸部X线片影像进行比较。结果：各组发病至首次胸部X线片检查时间分别为：轻型组26~48 h（中位时间37 h）;重型组10~36 h（中位时间23 h）;危重型组2~36 h（中位时间19 h）。最早发现胸部X线片异常的时间轻型组约为发病后30 h;重型组约为发病后23 h;危重型组约为发病后2 h;3组胸部X线片最早出现异常的时间差异有统计学意义（P<0.01）。首次影像异常率：轻型组5.8%,重型组81.3%,危重型组100%,危重型组首次X线片的异常比例明显高于其他两组（P<0.01）。在胸部X线片的表现上,轻型组最常见的征象为肺纹理增粗、模糊;重型组常见表现为渗出、实变影;危重型则表现为肺水肿征像。危重型组病灶分布广,累及多个肺叶。结论：肠道病毒71型感染患儿首次胸片检查时间、胸片出现异常的时间、异常率、胸部X线片表现严重度与临床病情严重程度相关。     

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??Objective To investigate the relationship between CCL2-2518A/G gene polymorphism and severity of enterovirus 71??EV71?? infection. Methods CCL2 gene polymorphism in 188 EV71-infected patients and 235 healthy controls were detected by the improved multiplex ligation detection reaction technique??iMLDR??. The level of CCL2 in two groups was determined by enzyme-linked immunosorbent assay??ELISA??. Results No significant differences were found in the distribution of genotype CCL2-2518A/G between EV71-infected patients and the healthy control group??P??0.05??. The G allele??genotypes AG or GG?? in the CCL2-2518A/G??P??0.001?? was more frequent in patients with severe EV71 infection. The level of CCL2 in infected patients was higher than that of the heathy controls ??P??0.05???? the severe cases had higher level of CCL2 than that of the slight cases and healthy controls??P??0.05??. The level of CCL2 in GG gene group was significantly higher than that in AG gene group and the level of CCL2 in AG gene group was significantly higher than that in AA gene group.The people with CCL2-2518G allele??GG+AG?? had higher level of CCL2 than those only with CCL2-2518A allele??AA????P??0.05??. Conclusion The G carrier of the CCL2-2518A/G is found to be associated with severity of EV71 infection??and could be susceptibility factors in the development of EV71 infection.     

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??The respiratory and circulatory support treatments are essential to critically ill viral encephalitis and postinfectious encephalitis patients in PICU. This support treatments ensure other therapy measures to give full play and can reduce braindamage and win time for recovery. Respiratory and circulatory dysfunction is common in severe encephalitis patients. Early and proper respiratory and circulatory support treatments can help to bring down the mortality and improve its prognosis.     

Basal ganglia involvement in a child with herpes simplex encephalitis

Herpes simplex encephalitis (HSE) is a leading cause of sporadic, nonepidemic viral encephalitis in children and adults. We report a very rare case of HSE with involvement of bilateral thalamus, putamen, upper pons and midbrain, with development of extrapyramidal symptoms which responded to corticosteroid therapy. A 15-mth-old female baby admitted with complaint of fever for 5 days and generalised tonic clonic seizure 10 hours before admission. On clinical examination patient was drowsy, temperature was 39.4 °C and vitals were stable with signs of increased intracranial tension. There were no signs of meningeal irritation. Patient gradually become unconscious in the next few hours and pupils were constricted bilaterally with development of atonia in all four limbs and neck muscles. Doll’s eye phenomenon was absent.     

Tick-borne encephalitis in a 3-month-old child

Tick-borne encephalitis has not been reported in infants younger than 12 months of age. We report a 3.5-month-old child with a serologically proven tick-borne encephalitis. The infant had a history of a tick bite 3.5 weeks before the first symptoms of encephalitis appeared. The family lives in an endemic area of the disease. There were no prodromal signs and the course of the disease was monophasic. In an endemic area, prophylactic treatment with hyperimmunoglobulin after a tick bite should be considered even in very young infants, but in most children active immunization is probably not necessary because of infrequent exposure. Acitve immunization is still recommended after the 1st year of life.     

TLR3-1377C/T基因多态性及表达水平与儿童肠道病毒71型脑炎易感性的探讨

目的 研究Toll样受体(TLRs)基因TLR3-1377C/T位点基因多态性及TLR3表达水平与儿童肠道病毒71型(EV71)脑炎易感性之间的关系。方法 收集EV71感染患儿187例(脑炎组59例和无脑炎组128例)与同期健康体检儿童232例进行病例对照研究。采用聚合酶链反应限制性片段长度多态性(PCR-RFLP)方法对TLR3-1377C/T基因多态性进行检测,采用ELISA检测血清TLR3水平。结果 与EV71感染无脑炎组相比,脑炎组TLR3-1377C/T位点基因型分布及等位基因频率的差异无统计学意义(P0.05)。与对照组相比,EV71感染脑炎组、无脑炎组的血清TLR3水平均显著增高(P0.05),以无脑炎组最高(P0.05)。脑炎组的EV71病毒载量高于无脑炎组(P0.01)。1岁或≥1岁的EV71感染脑炎组、无脑炎组患儿的血清TLR3水平均较相应对照组增高(P0.05),其中无脑炎组TLR3水平高于相应年龄的脑炎组(P0.05)。脑炎组≥1岁患儿的TLR3浓度高于1岁者(P0.05);无脑炎组及对照组的TLR3浓度在1岁或≥1岁患儿之间的差异无统计学意义(P0.05)。EV71感染脑炎组,1岁患儿所占比例高于≥1岁者(P0.05)。结论 TLR3-1377C/T位点的基因多态性与EV71脑炎的发生无明显相关性。TLR3的低表达可能导致对病毒复制的抑制作用减弱,促进了EV71脑炎的发生。婴儿EV71感染后血清TLR3的表达不足可能是其合并脑炎的重要因素。     

肠道病毒71型感染18例临床分析

目的 探讨儿童肠道病毒71型(EV71)感染的临床特征.方法 回顾性分析我院2008年4月至7月间18例EV71感染病例的流行病学特征及临床表现.结果 发病年龄以3岁以下为主(89%),外地移居儿童发病率明显高于本地儿童.所有患儿均出现发热及皮疹(100%),合并肺炎5例(28%),肺水肿并肺出血2例(11%),心肌炎5例(28%),病毒性脑炎4例(22%).治愈13例,好转2例,自动出院1例,死亡2例.结论 重症病例主要由EV71感染引起;肢体抖动、膝反射亢进是重症病例较具特征的表现;白细胞增高、血糖升高对危重病例的早期发现有提示意义.