Molecular epidemiology of enterovirus B77 isolated from non polio acute flaccid paralytic patients in Pakistan during 2013

Human enteroviruses are associated with various clinical syndromes and severe neurological disorders. The aim of this study was to determine the molecular epidemiology of non polio enteroviruses and their correlation with acute flaccid paralysis (AFP) patients living in Khyber Pakhtunkhwa (KP) and Federally Administered Tribal Areas (FATA) of Pakistan. The stool samples collected from these patients were used for isolation of non polio enteroviruses (NPEVs). Out of 38 samples, 29 (76.3%) were successfully typed by microneutralization assay into eleven serotypes including echovirus (E)-3 (5.3%), E-7 (2.6%), E-11 (13.2%), E-12 (7.9%), E-13 (10.5%), E-20 (7.9%), E-27 (5.3%), E-29 (10.5%), E-30 (7.9%), E-33 (2.6%), coxsackievirus (CV) B5 (2.6%) and nine isolates (23.7%) remained untyped which were confirmed as NPEVs by real time RT-PCR. Complete VP1 genetic sequencing data characterized untypeable isolates into enterovirus B77 (EV-B77). Moreover, molecular phylogenetic analysis classified these viruses into two new genotypes having high genetic diversity (at least 17.7%) with prototype.This study provides valuable information on extensive genetic diversity of EV-B77 genotypes. Although, its association with neurological disorder has not yet been known but isolation of nine EV-B77 viruses from AFP cases highlights the fact that they may have a contributing role in the etiology of AFP. In addition, it is needed to establish enterovirus surveillance system and laboratory diagnostic facilities for early detection of NPEVs that may cause poliomyelitis like paralysis especially in the situation when we are at the verge of polio eradication.     

浙江省急性驰缓性麻痹相关肠道病毒鉴定分析

目的 对浙江省2006-2008年急性弛缓性麻痹病例(AFP)中的非脊灰肠道病毒(NPEV)分离株进行分子生物学分型鉴定.方法 采用简并引物和逆转录-聚合酶链反应(RT-PCR),对31株非脊灰肠道病毒分离株进行目的基因片段扩增,然后测定序列,将测定的结果与GenBank进行BLAST比较分析,运用DNAMAN软件进行同源性分析,并构建基因树.结果 31株非脊灰肠道病毒分离株经分子生物学分型,其中人类肠道病毒(HEV)-A组5株,有2个血清型;HEV-B组24株,有13个血清型;HEV-C组2株,有1个血清型.结论 浙江省2006-2008年分离到的非脊灰肠道病毒以HEV-B组为主.     

云南省急性弛缓性麻痹病例中柯萨奇病毒B5VP1区基因特征分析

目的 分析云南省2015年从急性弛缓性麻痹（AFP）病例中分离到的3株柯萨奇病毒B5（CVB5） VP1区核苷酸基因特征。方法 采集RD细胞和L20B细胞对AFP标本进行病毒分离,应用反转录-聚合酶链反应（RT-PCR）扩增病毒VP1基因,目的基因送至基因公司进行序列测定;应用MEGA 6软件对分离到的病毒与基因库代表株进行进化树分析。结果 213例AFP标本中分离到23株非脊髓灰质炎肠道病毒（NPEV）,包括3株CVB5;3株CVB5与原型株VP1区之间核苷酸（nt）相似性为80.7%~81.9%,氨基酸（aa）相似性为95.1%~96.5%;系统进化树结果显示,本研究分离的CVB5与山东分离株（JN712675）的nt相似性为91.7%~92%,与安徽分离株（KF751191）的nt相似性为92%~93%。结论 云南省2015年从AFP病例中分离到的3株CVB5与山东代表株（JN712675）和安徽代表株（KF751191）相似性较高。     

广西急性弛缓性麻痹病例中非脊髓灰质炎肠道病毒感染分析

目的了解广西2003～2009年非脊髓灰质炎(脊灰)肠道病毒(NPEV)病例流行病学情况,为控制NPEV感染提供科学依据。方法按照WHO要求对AFP病例进行病毒分离与鉴定并对其流行病学特征进行分析。结果从1 851例AFP病例中检出NPEV 206株,分离率为11.13%。NPEV感染病例分布于全区各市县,无地区聚集性,有明显季节性,3岁以下儿童占78.8%,男性高于女性,临床诊断以短暂性肢体麻痹为主。结论 NPEV在广西AFP病例中具有较高的感染率,加强对其基因型别的研究,为AFP病例的诊断、治疗、预防措施提供依据。     

贵州省2012年急性弛缓性麻痹病例非脊髓灰质炎肠道病毒分型鉴定分析

目的 运用分子生物学检测方法对从贵州省2012年AFP (Acute Flaccid Paralysis)病例中分离到的30株非脊灰肠道病毒进行分型鉴定.方法 采用简并引物RT-PCR对分离株进行VP1区基因扩增和序列测定,测序结果进行BLAST比较和基因进化分析.结果 序列分析结果表明,30株NPEV中,有4株无法扩增VP1基因,另外26株共包括11个血清型.包括EV71、ECH06、COXA4、EV96、ECHO14、COXA10、ECH025、ECHO13、ECHO11、COXB5、COXB4.其中EV71有6株,所占比例最高.ECH06、COXA4各3株,EV96、ECH014、COXA10、ECH025、ECH013、ECHO11各2株,COXB5、COXB4各1株.结论 贵州省2012年AFP病例的病原谱可能包括多型别的NPEV.     

宁夏地区1997-2011年急性弛缓性麻痹病例非脊髓灰质炎肠道病毒分型鉴定分析

目的 对1997-2011年宁夏地区急性弛缓性麻痹(AFP)病例中分离的73株非脊髓灰质炎肠道病毒(NPEV)进行分型鉴定.方法 采用简并引物RT-PCR对分离株进行VP1区基因扩增和序列测定,测序结果进行BLAST比较和基因进化分析.结果 经过基因测序分析鉴定,73株NPEV中,有4株无法定型,另69株共包括27个血清型,分别属于A组肠道病毒(HEV-A)和HEV-B组,无HEV-C和HEV-D组毒株,其中HEV-A组毒株包括8个血清型23株,HEV-B组毒株包括19个血清型46株.69株病毒中,以EV71所占比例最大(9/69),以CVA4、CVA16、Echo24、Echo6和CoxA9所占比例较高.结论 1997-2011年宁夏地区AFP病例分离的NPEV以HEV-B组为主要型别66.7％(46/69).     

杭州市2001-2008年急性弛缓性麻痹监测病例中非脊髓炎质炎肠道病毒感染分析

目的为保持急性弛缓性麻痹（AFP）病例监测系统处于敏感状态,对非脊髓灰质炎肠道病毒（NPEV）引起AFP病例监测,为维持无脊髓灰质炎（简称脊灰）状态提供可靠依据。方法根据全市各个区、县（市）2001-2008年连续8年的AFP病例监测,进行系统评估。结果连续8年共有AFP病例181例,无脊灰确诊病例,AFP病例报告发病率在1.59/10万～3.09/10万之间。181例AFP病例共有11例为NPEV引起,发病全部为3岁以下儿童,2岁组发病6人。都没有出现残留麻痹。发病月份主要集中在7～9月,占54.55%。结论本研究连续8年的AFP监测表明,杭州市的AFP病例监测系统处于敏感状态。杭州市存在由NPEV感染引起的麻痹病例。     

浙江省2004～2007年急性驰缓性麻痹病例中非脊髓灰质炎肠道病毒分析

目的:对浙江省2004～2007年急性弛缓性麻痹病例(AFP)粪便标本中分离到的非脊髓灰质炎肠道病毒(NPEV)进行监测结果分析.方法:按照WHO规定的方法,用RDa和L2OB细胞对AFP患者的粪便标本进行病毒分离,对分离到的NPEV毒株进行型别鉴定.结果:4年共分离到44株NPEV,分别为柯萨奇A组病毒1株(COXA4),埃可病毒(ECHO)28株,不能定型15株.结论:浙江省2004年～2007年分离到的NPEV中主要为ECHO病毒,未分离到COXB组株病毒,且NPEV感染有明显的夏秋季高峰,主要集中在小年龄组儿童.     

Epidemiology and clinical characteristics of acute flaccid paralysis associated with non-polio enterovirus isolation: the experience in the Americas.

The Pan American Health Organization adopted as a goal the interruption of transmission of wild poliovirus from the americas by 1990. Collection and processing of stool specimens from patients with acute flaccid paralysis (AFP) to identify wild poliovirus is critical for monitoring the success of the eradication programme. In the study described, cases of AFP in children less than 15 years of age reported in the Americas from 1989 to 1991 were evaluated to investigate the epidemiology of AFP associated with the isolation of non-polio enterovirus (NPEV), to characterize their clinical presentation, and to compare their clinical characteristics with those of AFP cases associated with the isolation of wild poliovirus (confirmed as poliomyelitis). The results show that the notification pattern for AFP associated with NPEV isolates is similar to that for all AFP cases. While AFP associated with NPEV isolates generally differs clinically from confirmed poliomyelitis, 2-21% of cases met one of three case definitions for poliomyelitis. Following the eradication of poliomyelitis, countries can therefore anticipate the continued reporting of cases of AFP that clinically mimic poliomyelitis but which are associated with NPEV. The study also confirms that NPEV circulation is common and that most isolates were from cases that did not resemble poliomyelitis. It is therefore questionable whether characterization of NPEV isolates is essential for global eradication of poliomyelitis and consequently whether allocation of resources for that purpose can be justified.     

Screening of cases of acute flaccid paralysis for poliomyelitis eradication: ways to improve specificity.

The Pan American Health Organization in 1985 adopted an initiative to eradicate poliomyelitis from the Western Hemisphere. In 1990, over 2000 cases of acute flaccid paralysis (AFP) were reported in this region, of which < 1% were determined to be caused by wild poliovirus. At present, the eradication programme uses AFP as the criterion for surveillance of children aged < 15 years; this is 100% sensitive, but not specific. To minimize unnecessary diagnostic investigations, we studied all 4333 cases of AFP reported to the programme during 1989 and 1990 in order to develop more efficient operational screening criteria for cases of AFP. Among children with AFP, the use of criteria such as age < 6 years and either presence of fever at the onset of paralysis or a < 4-day period for complete development of paralysis resulted in a sensitivity of 96% (95% C.I. 90-103%) and specificity of 49% (C.I. 47-52%). With criteria of age < 6 years and fever present at the onset of paralysis the sensitivity was 75% (C.I. 61-89%) and specificity was 73% (C.I. 71-75%). These results suggest that by screening young children with AFP who either had fever at the onset or showed a rapid progression of paralysis, the number of cases of AFP requiring investigation can be reduced by one half, with minimal compromise in the sensitivity of confirmed poliomyelitis case detection.     

江苏省2012—2014年脊髓灰质炎疫苗病毒阳性急性弛缓性麻痹病例分析

目的为加强急性弛缓性麻痹(AFP),尤其是疫苗相关麻痹型脊灰病例(Vaccine-associated Paralytic Poliomyelitis,VAPP)监测和妥善处理提供依据。方法采用回顾性方法,对江苏省2012—2014年全省报告的AFP病例中检测脊灰疫苗株阳性的病例资料进行汇总分析。结果江苏省2012—2014年共报告AFP病例887例,下肢麻痹的占96.15%,粪便标本采便率96.96%。脊灰病毒检出率3.02%,全部为疫苗株,其中Ⅰ型占26.92%(7例),Ⅱ型占19.23%(5例),Ⅲ型占34.62%(9例),Ⅰ+Ⅱ型占11.54%(3例),Ⅰ+Ⅲ型占3.85%(1例),Ⅰ+Ⅱ+Ⅲ型占3.85%(1例)。南京、苏州、泰州3个市病例占57.69%,病例集中在4—5月,夏季Ⅰ型占54.55%,秋冬季Ⅲ型占87.50%(P=0.007)。男性多于女性,<5岁病例占73.08%。免疫史>3次占84.62%,采便距最后一次服苗时间<30d的混合型占75.00%,≥30d的单型占90.91%(P=0.031)。结论在保持高水平免疫接种率的同时,应继续保持高质量的AFP病例流行病学监测和病毒学监测,及时发现可能的输入病例,及时采取措施控制病毒的进一步传播及疫苗衍生脊髓灰质炎病毒(Vaccine-derived Polioviruses,VDPV)病例的发生。     

Poliomyelitis eradication programme: acute flaccid paralysis surveillance in mardin and five other provinces around Mardin, Turkey 1998

After the successful eradication of small pox, the eradication of poliomyelitis was planned. In the poliomyelitis eradication programme, conducted since 1989, routine vaccination, supplemental immunisation activities and acute flaccid paralysis (AFP) surveillance were the main strategies. In 1997, in Mardin provinces six poliomyelitis cases were reported. Therefore it was planned to strengthen the eradication programme in this province and those around it. In 1998 in Mardin and five neighboring provinces, a study was conducted that included monthly visits, educational activities, etc. At the result of this study, 64 AFP cases (22 of them poliomyelitis) were reported. Non-poliomyelitis AFP rate had increased from 0.9 in 1997 to 2.8 in 1998. An adequate two specimen ratio was 72%. We recognised that more poliomyelitis cases were reported in hot seasons when compared with non-poliomyelitis cases reported throughout the year. This difference was statistically significant (P<0.001). Most AFP cases were aged under 35 months. At the 60th day follow-up visits of the patients, we found 90.9% of the poliomyelitis cases but only 19.0% of the non-poliomyelitis case had residual paralysis. Presence of prodromal fever was another finding that distinguished poliomyelitis cases from non-poliomyelitis AFP cases. 90.9% of the poliomyelitis cases but only 64.3% of the non-poliomyelitis cases had fever at onset. This finding was also statistically significant (P<0.05).     

2012 - 2017年郑州市急性弛缓性麻痹病例监测结果分析

目的 对郑州市2012 - 2017年所监测的急性迟缓性麻痹（Acute Flaccid Paralysis, AFP）病例进行流行病学特征分析,为提高医生AFP的诊断水平和AFP防控策略的制定提供依据。方法 对AFP专病信息报告管理系统中2012 - 2017年郑州市16个市（区县）所报告的AFP病例进行分析。结果 2012 - 2017年这6年中郑州市共报告AFP病例226例,年平均发病率为2.91/10万,8月份报告病例最多,达32例;16个市区县中登封市报告病例数最多,达32例;有23个医院参与病例报告,其中报告最多是郑州市儿童医院（69 例）;病例主要集中于5岁以下人群,完成免疫者占总病例数的84.96%;实验室监测出PEV和NPEV阳性毒株;临床症状以发热和肌肉疼痛为主,治疗后仍有19.46%的患者有不同程度的残留麻痹。结论 郑州市2012 - 2017年AFP病例年平均发病率整体上呈现出下降趋势,提示该市脊灰防治工作效果良好,但基层医院的诊断水平有待提高,治疗后仍有患者存在麻痹症状,需进行长期监测。     

Surveillance of patients with acute flaccid paralysis in Finland: report of a pilot study

WHO recommends that surveillance of patients with acute flaccid paralysis (AFP) be used to demonstrate the eradication of wild poliovirus. In this article we report the results of a study to assess the frequency of AFP patients referred to Finnish hospitals and whether virological diagnostic coverage could be improved by repeated reminders and active feedback. For this purpose, we sent monthly questionnaires to all neurological and paediatric neurological units in Finland, requesting retrospective reporting on investigated paralytic patients with defined clinically relevant diagnoses, rather than AFP. Reminder letters included a pre-paid return envelope. Virological investigations were offered cost free. Of the 492 reporting forms sent, 415 (84%) were returned, evenly covering both the population and the study period (July 1997 to June 1998). Of the 90 patients reported, 83 were evaluable. The apparent incidences of the diagnoses covered were 1.6 per 100,000 at any age, and 1.0 per 100,000 for under--15-year-olds. Guillain-Barré syndrome was the most common diagnosis (0.80 per 100,000). The two faecal specimens required were virologically investigated in nine out of the 10 patients under 15 years of age, but in only 46% of all patients. Four adenovirus strains, but no polioviruses or other enteroviruses, were isolated. We conclude that a satisfactory monthly reporting system was readily established and that a sufficient number of patients with diagnoses resembling AFP are being referred to Finnish hospitals. Active feedback did not increase the proportion of virologically investigated patients to an acceptable level in all age groups. It is clear that other approaches must be used to quantify the circulation of poliovirus in Finland.     

Surveillance of acute flaccid paralysis in Italy: 1996–1997

The last case of poliomyelitis due to transmission of indigenous wild poliovirus occurred in Italy in 1982. To achieve the certification of the eradication of poliomyelitis in Italy, an active surveillance of acute flaccid paralysis (AFP) in the population aged less than 15 years was set up following the World Health Organization (WHO) guidelines. The survey started in 1996 with a pilot study involving 4 out of 21 regions, and was gradually extended to a national level in 1997. The two-year survey identified five patients with diagnosis of vaccine associated paralytic poliomyelitis (VAPP). Polioviruses type 2 and 3 Sabin-like were isolated and characterized in three of them. In the remaining two cases, samples were collected late after the onset of symptoms, and poliovirus could not be isolated. No wild polioviruses were detected during the survey. The rate of non-polio AFP found in Italy in 1997 was 0.61 cases per 100,000, which is lower than the level of 1.0 case per 100,000 considered as acceptable by the WHO. This was mainly due to the delay in organizing the hospital network and starting the active search of AFP cases in the largest and most densely populated regions. Therefore, the overall rate of AFP found in Italy underestimates the global effectiveness of the program, which however will be better evaluated in the next few years. This study is the first systematic attempt to determine the rate of AFP in Italy.     

2016年河南省急性弛缓性麻痹病例中脊髓灰质炎病毒监测结果分析

目的鉴定分析河南省15岁以下儿童疑似急性弛缓性麻痹(acute flaccid paralysis,AFP)病例中实验室确诊感染脊髓灰质炎(脊灰)病毒(Poliovirus,PV)的型别,为河南省制定更完善的脊灰防控策略及维持无脊灰野病毒状态提供科学依据。方法依据《WHO脊髓灰质炎实验室手册第4版》操作规程,使用RD和L20B两种细胞对河南省2016年AFP标本进行病毒分离,采用荧光定量PCR法及逆转录PCR法进行病毒序列鉴定和分析。结果共检测453例疑似AFP病例,其中有7例(1.55%)病例分离出10株脊灰病毒,其中Ⅰ型、Ⅲ型和Ⅰ+Ⅲ型分别为5株,3株和2株。分离到脊灰毒株与Sabin株相比,核苷酸VP1区发生0、1、2、4和6个变异的数量分别为3、1、3、2和1株。结论 2016年河南省分离出脊灰病毒均为脊灰疫苗株,毒株型别以Ⅰ型为主,VP1区核苷酸以低变异为主,未发现疫苗衍生脊灰病毒(Vaccine-derived poliovirus,VDPV)及脊灰野病毒(Wild poliovirus,WPV)。     

贵州省2012—2019年急性弛缓性麻痹病例监测分析

目的 分析贵州省2012—2019年急性弛缓性麻痹（acute flaccid paralysis,AFP）病例的流行病学特征,为开展AFP病例监测工作提供科学依据。方法 采用描述性流行病学方法对贵州省2012—2019年AFP病例监测数据进行分析。结果 贵州省2012—2019年共报告1 714例AFP病例,年均报告发病率为2.65例/10万,无明显季节性分布,男性发病率（3.14/10万）高于女性发病率（2.07/10万）,1～6岁儿童居多（55.43%）,以散居儿童（45.22%）和集体单位儿童（托幼机构、学校）（42.47%）为主;脊灰疫苗免疫史≥3剂次的病例占91.72%;AFP病例肠道病毒分离率为8.38%;27.48%的病例为短暂性肢体麻痹;16.45%的病例残留麻痹。结论 贵州省仍需保持AFP病例监测的高敏感性和及时性,保持常住儿童脊灰疫苗高覆盖率,提高流动儿童接种率,筑牢免疫屏障。随访时加强健康教育,促进病例肌力恢复,提高生存质量,保护成年后劳动生产力。