A Centralized Report on Pediatric Japanese Encephalitis Cases from Beijing Children's Hospital, 2013 / 生物医学与环境科学（英文）

Fifteen pediatric cases of suspected Japanese encephalitis (JE) were reported in Beijing Children's Hospital during the late summer of 2013. The clinical manifestations in most cases included high fever, seizures, and abnormal magnetic resonance imaging findings. Twelve of 15 cases were laboratory-confirmed as JE cases by pathogen identification. Epidemiological investigations showed that five of the 12 laboratory-confirmed patients had an incomplete JE vaccination history. Follow-up investigations after discharge indicated that seven laboratory-confirmed JE patients without JE vaccinations had relatively poor prognoses, with an average Modified Rankin Scale (MRS) score of 2.6 when compared with the other five laboratory-confirmed, JE-vaccinated patients with an average MRS score of 0.5. The observation of pediatric JE cases among those with a history of JE vaccination warrants further attention.

Immunoprotective effect of inactivated coxsackievirus A16 vaccine in mice / 病毒学报

This study aims to construct inactivated coxsackievirus A16 (CVA16) vaccine and to investigate its protective effect in ICR mice. A clinical isolate of CVA16, 521-01T, was cultured in VERO cells, inactivated by formaldehyde, and purified by ultracentrifugation for vaccine preparation. Purity and other characteristics of the vaccine were determined by SDS-PAGE and Western blot. Female ICR mice were subcutaneously inoculated with inactivated CVA16 or Al(OH)3-absorbed CVA16, followed by booster immunization at the end of 2 and 4 weeks. CVA16-specific IgG titers in serum were determined by ELISA, and titers of neutralizing antibodies were determined by viral neutralization assay. The immunity of T lymphocytes was evaluated by IFN-gamma ELISPOT assay. The protective effect was evaluated by challenging the neonatal offspring (< 48 hours) of vaccinated female mice with 1 000 LD50 of CVA16 521-01T. The mortality rates of different groups were compared. The results showed that Al(OH)3 +CVA16 could induce high titers of specific IgG antibodies in ICR mice. After being boosted two times, the serum IgG antibody titer could reach up to 1 : 1 x 10(5) (P = 0.000), and neutralizing antibody titer was higher than 1 : 256. Additionally, more spot forming cells were induced in the immunized groups than in the negative controls. The maternal antibodies showed protective effect in 100% of the neonatal mice challenged with 1 000 LD50 of CVA16 521-01T. The inactivated CVA16 vaccine has ideal immunogenicity and immunoprotective effect. This research lays a foundation for the development and evaluation of CVA16 vaccines.

Epidemiology of hand, foot, and mouth disease and genetic characterization of enterovirus A71: a survey from 2007 to 2012 in Linyi of Shandong Province, China / 病毒学报

To investigate the epidemiology of hand, foot, and mouth disease (HFMD) and the genetic characteristics of enterovirus A71 (EV-A71) in Linyi of Shandong Province, China during 2007-2012. The number of reported HFMD cases were obtained from the National Notifiable Disease Reporting System (NNDRS) were analyzed by descriptive epidemiology method; the VP1 region of EV-A71 isolated from HFMD patients in Linyi was amplified and sequenced. Finally, the genetic variability and phylogenecity of VP1 sequences of EV-A71 were analyzed by MEGA 5.0. The results showed that HFMD incidence was reported in each year from 2007 to 2012 in Linyi, and the highest incidence and mortality were reported in 2009, when there were total 14697 cases and 9 of death. The reported incidence was 140.28/100000, and the mortality was 0.086/100000. The peak incidence usually occurred between April and July, and the summit occurred in May. Scattered children accounted for 77.37%-92.00% of all cases. The peak age was 2.5 years during 2007-2009 and 1.5 years during 2010-2012. A total of 1365 laboratory-confirmed HFMD cases were reported in the 6 consecutive years, accounting for 2.98% of the gross number. Among these reports, the ratio of EV-A71 was 44.18%, and the ratio of coxsackievirus A16 (CVA16) was 46.59%. All EV-A71 strains isolated in Linyi during 2007-2012 belonged to the C4a evolutionary branch of C4 genotype. In conclusion, HFMD outbreaks occurred every year in Linyi during 2007-2012. Incidence varied significantly among different counties. The peak incidence in each year lasted from April to July. Most of the patients were children under 3 years of age, and scattered children took the highest proportion. Co-circulation of EV-A71 and CVA16 was the major cause of HFMD in each year. Since the first report of HFMD prevalence caused by EV-A71 (C4a) in 2007, the virus has been prevalent continuously in Linyi for 6 years.

Research advances in molecular epidemiology and vaccines of Coxsackievirus A16 / 病毒学报

Epidemics of hand, foot and mouth disease (HFMD) have mainly been caused by Coxsackievirus A16 (CVA16) and Enterovirus A 71 (EV-A71), which circulated alternatively or together in the affected area. CVA16 has caused numerous outbreaks and epidemics in multiple countries and geographical regions, and has become an important public health problem. Based on an analysis of the complete VP1 coding region, all CVA16 strains can be divided into genotypes A, B1, and B2. Furthermore, genotype B1 can be divided into subgenotypes B1a, B1b, and B1c. After 2000, no reports of genotype B2 virus strains have been reported. All of the CVA16 strains reported in mainland China have belonged to subgenotypes B1a and B1b. Most CVA16-associated infections cause only mild symptoms; however, some CVA16 infections can lead to severe complications and even death. Vaccination is considered to be the most effective method to control the transmission and infection rate of this virus. A number of research groups are studying various vaccine types, including inactivated vaccines, genetic engineering vaccines, and DNA vaccines, amongst others. In this review, an overview is provided of the research advances in molecular epidemiology and vaccines of CVA16.

Epidemiology and etiology of hand-foot-and-mouth disease in Hainan, 2010 / 病毒学报

To reveal the epidemiological and pathogenic characteristics of Hand-foot-and-mouth disease (HFMD) in Hainan province in 2010, epidemiology data of HFMD reporting cases were analyzed, clinical specimens from 1346 HFMD cases were collected for enterovirus (EV) detection. Viral isolation was performed for EV nucleic acid positive samples. Complete VP1 encoding region of EV71 were sequenced and analyzed with Sequencher (version 5.0) and MEGA software (version 5.0). The epidemiology data showed that all 18 prefectures in Hainan had reporting cases during 2010, with higher incidence in the northeast; and the children less than 4 years old accounted for the majority of the suffered; the epidemic reached peak during September to October, which was different from other Provinces in China. The laboratory results indicated that EV71 and CA16 were identified as the major causative pathogens in Hainan in 2010, however, EV71 infection was absolutely dominant among severe and fatal cases. In addition, some HFMD cases were identified associated with other serotypes of EV infections. Molecular epidemiological analysis showed that all the EV71 strains belonged to C4a evolutionary branch, which is the dominant evolutionary branch in China in recent years, and at least three transmission chains existed. This study has an important information in clarifying the characteristics of epidemics and transmission of HFMD in Hainan, and to provide the guidance for HFMD prevention and control in the future.

Etiology of hand-foot-and-mouth disease seen in Nanjing in 2010 and molecular characteristics analysis of EV71 / 中华儿科杂志

<p><b>OBJECTIVE</b>To study the etiological agent of hand-foot-and-mouth disease (HFMD), and to genetically characterize enterovirus 71 (EV71) isolated from clinical specimens of HFMD patients in Nanjing in 2010.</p><p><b>METHOD</b>Throat swab specimens were collected from 248 inpatients with HFMD in Nanjing Children's Hospital and then viral isolation was performed. Real-time PCR was used for detection of EV71, coxsackievirus A16 (Cox A16) and other enteroviruses from the positive isolates. Twenty EV71 strains from different clinical types of cases were selected for entire VP1 coding gene amplification and sequencing, finally a phylogenetic tree was constructed among the 20 EV71 strains and EV71 representative strains of known genotypes and subgenotypes.</p><p><b>RESULT</b>From the 248 throat swabs specimens, 110 EV71 strains, 28 Cox A16 strains, and 8 other enterovirus strains were isolated and the positive rate was 44.35%, 11.29%, 3.23%, respectively. Then nucleotide sequencing was performed on the 20 EV71 strains. There was little difference in the nucleotide and the amino acid sequences among the 20 EV71 strains, the homology was 95.51%-100% and 98.32%-100%, respectively. The phylogenetic tree showed that all the 20 EV71 strains belonged to C4 subgenotype.</p><p><b>CONCLUSION</b>EV71 was the main pathogen of HFMD in Nanjing in 2010 and all the analyzed isolates belonged to C4 serotype. No significant difference was found in sequence of VP1 region of EV71 isolated from different clinical types of cases.</p>

Multiple viral transmission chains of enterovirus 71 co-circulated in Jilin Province during 2009-2010 / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To analyze the genetic characterization of enterovirus type71 (EV71) associated with hand foot and mouth disease (HFMD) epidemics in Jilin province, during 2009-2010.</p><p><b>METHODS</b>Randomly selected 31 representative EV71 strains from the cases of 8 prefectures to amplify and sequences of VP1 genes of EV71 strains, and analyzed with Bioedit and Mega4.0 program.</p><p><b>RESULTS</b>All representative 31 EV71 strains belong to C4a subgenotype, the homology of nucleotide in VP1 region among the 31 EV71 strains were 94. 5%-100. 0%, and were clustered into 5 transmission chains respectively. 25 strains out of 31 strains were associated with a predominant transmission chain, and circulating in 8 prefectures, while other 6 strains clustered into 4 lineages.</p><p><b>CONCLUSION</b>Multiple transmission chains of EV71 C4a subgenotype were co-circulating in Jilin province during 2009-2010, and a predominant transmission chain was circulating in 8 prefectures, associated with HFMD outbreaks of Jilin province.</p>

Transmission and prevalence patterns of C4a evolutionary lineage of human enterovirus 71 circulating in mainland China, 2008-2010 / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To understand the evolutionary relationship between the C4a evolutionary lineage of human enterovirus 71 (HEV71) strains circulating in mainland of China during 2008-2010 and 2008 Fuyang strains and study the prevalence and transmission patterns of 2008 Fuyang strains.</p><p><b>METHODS</b>Download all the complete VP1 ( > or = 891 bp) or approximate complete VP1 (> or = 876 bp) gene nucleotide sequences from GenBank of HEV71 strains circulating in Mainland of China during 2008-2010. And analyze the phylogenetic relationship between Fuyang strains and other provinces' strains using the MEGA software, version 5.0.</p><p><b>RESULTS</b>All of the HEV71 isolates circulating in Mainland of China during 2008-2010 were clustered into evolutionary lineage C4a except for eight strains grouped in the genotype A and one isolate belongs to evolutionary lineage C4b; the homology analysis showed there were 96.5%-100% identity between C4a viruses circulating in mainland China during 2008-2010 and 2008 Fuyang strains, and they were evolved from C4b viruses of 1998. The transmission chains of Fuyang strains were mainly transmitted in Guangdong, Jiangsu, Shanghai, Hunan, Shandong provinces.</p><p><b>CONCLUSION</b>The predominant viruses circulating in Mainland of China during 2008-2010 were evolutionary lineage C4a of human Enterovirus 71; Fuyang transmission chains mainly distributed in southern of China and the Central China around Anhui provinces.</p>

Genetic characteristics of entervirus 71 isolated in Ningxia Hui Autonomous region in 2009 / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To analyze the genetic characteristics of EV71 strains isolated from HFMD cases in Ningxia Hui Autonomous Region in 2009.</p><p><b>METHODS</b>In 2009, totally 385 specimens from 344 HFMD cases were collected from Ningxia. Enterovirus isolation was performed in RD cell line from all the specimens. EV71 isolates were identified by specific RT-PCR from the positive cultures, and sequences of complete EV71 VP1 encoding region were determined for farther analyses.</p><p><b>RESULTS</b>Totally from 126 EV strains isolated in this study, 58 EV71 strains (46%) were identified. And complete VP1 sequences of 46 EV71 strains were determined, and genetic analyses were performed. It was showed that the nucleotide identity of 46 Ningxia strains with the representatives of A and B genotypes were 81.7%-82.8% and 83.1%-85.2%, and the amino acid identity were 93.9%-95.9% and 96. 2%-97.9% respectively. The nucleotide identity of NingXia EV71 isolates with representatives of subgenotype C1, C2, C3, C4a, C4b, and C5 were 88.3%-90.6% (97.9%-99.6%), 88.3%-90.1% (97.9%-99.3%), 87.8%-89.0% (97.6%-98.9%), 94.2%-98.9% (97.9%-100%), 91.8%-94.1% (98.6%-99.6%), and 86.7%-89.1% (97.9%-98.9%). Phylogenetic tree analysis revealed that 46 stains were clustered with reference stains of subgenotype C4 and the Ningxia EV71 isolates were belonged to subgenotype C4a.</p><p><b>CONCLUSION</b>EV71 of subgenotype C4a had spread widely in Ningxia in 2009, which was absolutely predominant type in Ningxia in 2009 and also as the predominant type in China mainland since 2005.</p>

Etiology of hand-foot-and-mouth disease and genetic characteristics of Coxsackievirus A16 in Beijing, 2009 / 病毒学报

This study aimed to reveal the pathogen spectrum of hand-foot-and-mouth disease (HFMD) and genetic characteristics of Coxsackievirus A16 (CoxA16) isolates in Beijing in 2009. From 1044 clinical specimens collected from 975 HFMD cases at Beijing Pediatrics Hospital, Beijing You'an Hospital and Beijing Ditan Hospital in 2009, viral nucleic acids of enterovirus were detected by reverse transcriptase polymerase chain reaction (RT-PCR). Enterovirus isolations were conducted with rhabdomyosarcoma (RD) cell line on 200 throat swabs having positive RT-PCR results. Sequencing and analyses of VP1 encoding gene were performed on 9 CoxA16 isolates in this study. The results showed that CoxA16 (49.4%) and EV71 (36.4%) were the major pathogens for the epidemics of HFMD in 2009 in Beijing, and CoxA16 was the predominant serotype, while there were also other enterovirus co-circulating, such as CoxA4, CoxA10, and CoxA9; the CoxA16 strains prevalent in Beijing in 2009 belonged to subgenotype B1a and B1b.

Epidemiology and etiology of hand-foot-and-mouth disease in Shanghai, 2009 / 病毒学报

To analyze the epidemiological and etiological characteristics of Hand-Foot-and-Mouth disease (HFMD) in Shanghai in 2009, epidemiological data was retrieved from the National Notifiable Disease Report System (NNDRS). Nucleic acid of enterovirus (EV) was detected by real-time RT-PCR from 799 HFMD cases from 15 districts/counties in Shanghai; the complete sequences of VP1 encoding region of several identified EV71 strains and sequences of VP4 encoding region of several untyped EV were determined and analyzed. Analysis and summary of the epidemiological data was conducted with Microsoft Excel, and sequence analyses were conducted with both BioEdit and MEGA software. Untyped EV was identified through comparing the VP4 sequence to sequence database using BLAST online service. It was showed that all the 18 districts/counties had reported HFMD cases; children less than 6 years old were the most susceptible population group; the peak of epidemics of HFMD was from April to July; EV71 and Coxsackievirus A16 (CA16) were the major pathogens for this epidemic, but the constituent ratio of EV71 and CA16 was different in different months and regions; CA16 infection was mainly responsible for the mild HFMD, but EV71 for most of the severe cases; EV71 strains of Shanghai were clustered with representatives of subgenotype C4a and showed the highest identity to them, based on the sequence analyses of VP1 encoding region; 2 of the untyped EV were identified as CA2 and CA10 respectively. All the results indicated that EV71 and CA16 were the major pathogens for the epidemic of HFMD in Shanghai, 2009; the circulating EV71 belonged to subgenotype C4a. Besides, other types of EV (for example: CA2 and CA10) were also responsible for a few of the HFMD cases.

The epidemiologic and virological analysis of an outbreak of hand, foot, and mouth disease in Inner Mongolia in 2007 / 病毒学报

In 2007, an outbreak of hand, foot, and mouth disease (HFMD) occurred in Jungar Banner, Erdos city, Inner Mongolia Autonomous Region, China Fever, vesicular exanthema on the hands, feet, mouth, and buttocks were presented in most of the patients. Most of the patients were infants less than 5 years old, and an obvious peak period appeared in the disease outbreak. From 28 hospitalized patients, 23 stool specimens and 6 throat swab specimens were collected for enterovirus isolation, and 15 enteroviruses were isolated, 9 were identified as Human Enterovirus 71 (HEV71, the isolation rate is 31.03%) and 1 was identified as Coxsackievirus A16 (CVA16). According to the comprehensive analysis of clinical manifestation, epidemiology data and laboratory results, this outbreak was probably mainly caused by HEV71. The variability at nucleotide acid level and amino acid level among 9 HEV71 was relatively low, and the homology was more than 99.4% and 99.0% respectively, showing that this outbreak was caused by only one viral transmission chain. Phylogenetic analysis of 9 HEV71 strains isolated during this outbreak revealed that they all belonged to subgenotype C4, which has been continuously circulating in mainland China since its first reported occurrence in Shenzhen City in 1998. It was also suggested that subgenotype C4 HEV71 had a widely distribution and transmission in mainland China.

Human bocavirus in children suffering from acute lower respiratory tract infection in Beijing Children's Hospital / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Human bocavirus (HBoV) is a parvovirus recently found to possibly cause respiratory tract disease in children and adults. This study investigated HBoV infection and its clinical characteristics in children younger than five years of age suffering from acute lower respiratory tract infection in Beijing Children's Hospital.</p><p><b>METHODS</b>Nasopharyngeal aspirates were collected from children suffering from acute lower respiratory tract infection during the winters of 2004 to 2006 (from November through the following February). HBoV was detected by polymerase chain reaction amplification and virus isolation and the amplification products were sequenced for identification.</p><p><b>RESULTS</b>HBoV infection was detected in 16 of 333 study subjects. Coinfections with respiratory syncytial virus were detected in 3 of 16 HBoV positive patients with acute lower respiratory tract infection. The median age for HBoV positive children was 8 months (mean age, 17 months; range, 3 to 57 months). Among the HBoV positive children, 14 were younger than 3 years old, 9 were younger than 1 year old and 7 were younger than 6 months. These 16 positive HBoV children exhibited coughing and abnormal chest radiography findings and more than 60% of these children had wheezing and fever. Ten children were clinically diagnosed with pneumonia, 2 bronchiolitis, 2 acute bronchitis and 2 asthma. One child died.</p><p><b>CONCLUSIONS</b>HBoV was detected in about 5% of children with acute lower respiratory infection seen in Beijing Children's Hospital. Further investigations regarding clinical and epidemiologic characteristics of HBoV infection are needed.</p>