Analysis of Coxsackie virus A6 and A10 detection in hand foot and mouth disease in Guiyang area during 2015 / 重庆医学

Objective To understand the enterovirus(EV) pathogenic composition of Coxsackie virus(CV)A6 and A10 causing hand foot and mouth disesse(HFMD) in Guiyang area during 2015 to provide a basis for the prevention and treatment of HFMD in local area.Methods The specific primers were respectively designed according to CVA6 and CVA10 sequence published in GenBank.The fluorescence quantitative RT-PCR method system was established.The gene sequencing method was used for conducting verification.Then this method was used to detect the clinical samples from 607 cases of HFMD.Results A total of 607 samples of suspected HFMD were detected,the overall positive rate was 59.47 ％ (361/607),in which EV71 accounted for 7.25 ％ (44/607),CVA16 for 11.37％(69/607),EV71+CVA16 double positive accounted for 0.16％ (1/607) and other EV for 40.69％ (247/607).The positive samples of CVA6,CVA10 and CVA6+CVA10 detected by the established real time fluorescence RT-PCR were 11 cases,71 cases and 1 case.Conclusion CVA6 and CVA10 are the main pathogens causing new onset HFMD in Guiyang area and the CVA10 monitoring should be strengthened.

Neurologic Manifestations of Enterovirus 71 Infection in Korea

Enterovirus 71 frequently involves the central nervous system and may present with a variety of neurologic manifestations. Here, we aimed to describe the clinical features, magnetic resonance imaging (MRI) findings, and cerebrospinal fluid (CSF) profiles of patients presenting with neurologic complications of enterovirus 71 infection. We retrospectively reviewed the records of 31 pediatric patients hospitalized with acute neurologic manifestations accompanied by confirmed enterovirus 71 infection at Ulsan University Hospital between 2010 and 2014. The patients' mean age was 2.9 ± 5.5 years (range, 18 days to 12 years), and 80.6% of patients were less than 4 years old. Based on their clinical features, the patients were classified into 4 clinical groups: brainstem encephalitis (n = 21), meningitis (n = 7), encephalitis (n = 2), and acute flaccid paralysis (n = 1). The common neurologic symptoms included myoclonus (58.1%), lethargy (54.8%), irritability (54.8%), vomiting (48.4%), ataxia (38.7%), and tremor (35.5%). Twenty-five patients underwent an MRI scan; of these, 14 (56.0%) revealed the characteristic increased T2 signal intensity in the posterior region of the brainstem and bilateral cerebellar dentate nuclei. Twenty-six of 30 patients (86.7%) showed CSF pleocytosis. Thirty patients (96.8%) recovered completely without any neurologic deficits; one patient (3.2%) died due to pulmonary hemorrhage and shock. In the present study, brainstem encephalitis was the most common neurologic manifestation of enterovirus 71 infection. The characteristic clinical symptoms such as myoclonus, ataxia, and tremor in conjunction with CSF pleocytosis and brainstem lesions on MR images are pathognomonic for diagnosis of neurologic involvement by enterovirus 71 infection.

The pulmonary edema caused by enterovirus71 / 中国医师杂志

Enterovirus 71 (EV71) infection can cause hand,foot and mouth disease,due to its neurotropic nature,severe cases can affect the central nervous system and emerge neurogenic pulmonary edema (NPE),respiratory and circulatory failure,and even lead to death.The advances in the pathogenesis of severe HFMD neurogenic pulmonary edema after EV71 infection in recent years are described.

Etiology of hand foot and mouth disease in Guiyang area during 2012 / 重庆医学

Objective To understand the pathogen distribution situation among the children inpatients with hand ,foot and mouth diseases(HFMD) in Guiyang area during 2012 to provide the basis for the diagnosis ,treatment and prevention .Methods The data in 3 179 cases of HFMD were collected .The fluorescence quantitative RT‐PCR was adopted to perform the genotyping on universal enterovirus ,enterovirus 71(EV71)and Coxsackie virus A16(CA16) .Results A total 3 179 samples of HFMD were col‐lected ,among them ,151 cases (4 .75% ) were CA16 positive ,331 cases (10 .41% ) were EV71 positive ,7 cases (0 .22% ) were CA16 and EV71 co‐infection ,and 897 cases(28 .22% )were the other enterovirus .The whole year had 2 peaks of onset ,which were April to July and October to November .The onset age focused on the children aged under 5 years old (96 .16% ) ,among them ,0-3 years old had the highest onset ,moreover male children were more than female .Conclusion The etiology distribution of children HFMD in Guiyang area during 201 was dominated by the other genotypes of enterovirus and EV71 .

Application of combined detection of enterovirus nucleic acid and antibody in early etiological diagnosis for hand-foot-and-mouth disease / 中华检验医学杂志

Objective To assess the value of combined detection of enterovirus nucleic acid and antibody in early etiological diagnosis for hand-foot-and-mouth disease ( HFMD).Methods A case-control study was conducted.A total of 1 066 cases of children clinically diagnosed with HFMD from Hangzhou Children′s Hospital were involved into the research group from January to June 2014, consisting of 401 common cases and 665 severe cases; Throat swabs and serum samples from these children underwent combined detection for EV71/CA16/EV of enterovirus nucleic acid by fluorescence quantitative RT-PCR and for EV71/CA16-IgM by ELISA.All data were analyzed with SPSS 16.0.Results The total positive rate of enterovirus nucleic acid EV71/CA16/EV by fluorescence quantitative RT-PCR in the 1 066 cases of children clinically diagnosed with HFMD was 75.52%( 805/1 066 ) ( 95%CI: 72.80%-78.05%).But the total positive rate of combined detection was 91.46%( 975/1 066 ) ( 95%CI:89%.58-93.04%).The total positive rate of combined detection is higher than that of RT-PCR test(χ2 =98.338,P=0.000).The positive rate of EV71 type of combined detection was 64.63%(689/1 066)(95%CI:61.67%-67.49%),which is 15.38%higher than that of RT-PCR test 49.25%(525/1 066)(95%CI:46.21%-52.29%)(χ2 =51.453, P=0.000).In 665 severe cases of HFMD, the total positive rate of combined detection was 96.69%(643/665)(95%CI:94.95%-97.87%), which is higher than that of RT-PCR test 79.25%(527/665)(95%CI:75.92%-82.22%)(χ2 =95.607, P =0.000).In the severe cases, the positive rate of EV71 type of combined detection was 87.52%( 582/665 ) ( 95%CI:84.71%-89.89%) , which is 18.95% higher than that of RT-PCR test 68.57%(456/665) (95%CI:64.87%-72.06%) (χ2 =69.665, P=0.000).In the fatal cases, the positive rate of EV71 type of combined detection was 95.92%(94/98) (95%CI:89.28%-98.68%).Conclusions The combined detection of enterovirus nucleic acid and specific IgM antibody can significantly increase the positive rate of HFMD, especially for severe cases.The combine detection increases both the total positive rate and EV71 positive rate.Thus it has a high potential for becoming a new guidelines for laboratory diagnosis of HFMD.

Sequence analysis of VP1 and VP4 genes of enterovirus 71 strains isolated from severe and mild hanD-foot-mouth disease cases in Shanghai, China, 2011 / 中华传染病杂志

Objective To analyze the VP1 and VP4 genetic region of enterovirus 71 (EV71)isolated from severe cases and mild cases with hanD-foot-mouth disease (HFMD) in Shanghai in 2011.Methods Five EV71 strains isolated from severe cases and five EV71 strains from mild cases in 2011 were included.Reverse transcription-polymerase chain reaction (RT-PCR) method was used to amplify and sequence the VP1 and VP4 genes of EV71,and then the sequencing results were compared with those of A,B,C genotype reference EV71 strains from GenBank by nucleotide alignment,amino acid alignment and phylogenetic tree analyses.Results The homogeneity between EV71 strains from severe cases and mild cases was 96.0％-98.1％ and 93.7％-99.5％ for VP1 and VP4 nucleotide sequences,respectively.The VP1 nucleotide sequences of 5 strains isolated from severe cases and 5 strains from mild cases in Shanghai shared 86.9％-98.2％ and 87.4％ 98.5％ identity with genotype C,respectively,while the homogeneity of VP4 nucleotide sequence was 85.5％-100.0％and 84.5％-99.5％,respectively.In addition,compared with the Fuyang EV71 strains (representative of C4 subtype),the strains from mild and severe cases shared homogeneity of 97.0％-98.2％ and 97.9％-98.5％ for VP1 gene,respectively,96.1％-100.0％ and 97.1％-99.5％ for VP4 gene,respectively.Among 3 strains isolated from severe cases,mutations at the residue 282 in the VP1 protein (N→S) and residue 7 in the VP4 protein (T→A) were discovered simultaneously.Conclusions The 10 EV71 strains isolated from severe and mild cases in Shanghai belong to subgenogroup C4.Among 3 strains isolated from severe cases,mutations at the residue 282 in the VP1 protein (N→S) and residue 7 in the VP4 protein(T→A) are discovered simultaneously.

Establishment and clinical application of triplex taqman probes real-time PCR for pathogenesis of hand foot mouth disease / 中华检验医学杂志

Objective To establish the triplex Taqman probes real-time RT-PCR method for simultaneously detecting of EV71,CA16 and EV.Methods Retrospective study.Specific primers and probes were designed based on conserved regions of EV71,CA16 and EV.The sensitivity,specificity and reproducibility were assessed by the optimized reaction system.A total of 176 throat swabs as the experimental group were collected from children with suspected hand foot mouth disease (HFMD),who admitted from April 2012 to July 2012 in Nanjing Children's Hospital affiliated to Nanjing Medical University.During this time,10 cases of healthy children,10 cases of outpatients with flu-like symptoms and 90 cases of inpatients in pneumology department of our hospital were recruited as control group,whose throat swabs were also collected.All of 286 samples were tested by the triplex Taqman probes real-time RT-PCR for simultaneously detecting EV71,CA16 and EV.SPSS13.0 was used to analyze the results.Results The sensitivities of the triplex Taqman probes real-time RT-PCR was 1.0 × 103 copies per milliliter for EV71,CA16 and EV.It showed 100％ specificity for 9 enterovirus and 3 non-enterovirus.Analysis with 1.0 × 103-1.0 × 105 copies per milliliter constructed plasmids demonstrated high reproducibility with coefficient of variation of 0.44％-1.04％ for EV71,0.38％-0.73％ for CA16,and 0.46％-0.90％ for EV.More over 176 samples collected from children with suspected HFMD were detected by triplex Taqman probes real-time RTPCR and real-time RT-PCR.The results showed 97.2％ (171/176)agreement and 0.94 Kappa value with high concordance.Conclusions The triplex Taqman probes real-time RT-PCR detecting EV71,CA16 and EV simultaneously has been established successfully.The assay,with high sensitivity and specificity,provide good basis for the rapid clinical diagnosis of EV71,CA16 and EV and open up broad prospects for clinical and relevant researches.

Risk Factors for Neurologic Complications of Hand, Foot and Mouth Disease in the Republic of Korea, 2009

In 2009, the first outbreak of hand, foot and mouth disease (HFMD) or herpangina (HP) caused by enterovirus 71 occurred in the Republic of Korea. This study inquired into risk factors associated with complications of HFMD or HP. A retrospective medical records review was conducted on HFMD or HP patients for whom etiologic viruses had been verified in 2009. One hundred sixty-eight patients were examined for this investigation. Eighty patients were without complications while 88 were accompanied by complications, and 2 had expired. Enterovirus 71 subgenotype C4a was the most prevalent in number with 67 cases (54.9%). In the univariate analysis, the disease patterns of HFMD rather than HP, fever longer than 4 days, peak body temperature over 39degrees C, vomiting, headache, neurologic signs, serum glucose over 100 mg/dL, and having an enterovirus 71 as a causative virus were significant risk factors of the complications. After multiple logistic analysis, headache (Odds ratio [OR], 10.75; P < 0.001) and neurologic signs (OR, 42.76; P < 0.001) were found to be the most significant factors. Early detection and proper management of patients with aforementioned risk factors would be necessary in order to attain a better clinical outcome.

Survey on the enterovirus 71 survival ability on different surfaces under different climate / 中华传染病杂志

Objective To evaluate the survival ability of enterovirus 71 (EV71) on different surface and under different climate.Methods Each 1 × 105 tissue culture infective dose 50 (TCID50)EV71 was added on different aseptic surface of plastic,rubber,cloth and wood,respectively.Then these materials were put into biotron (artificial climatic chamber) which could simulate different temperature and moisture.The viruses were recovered after a definite time and then inoculated into Vero cell.The cytopathic effect (CPE) was observed everyday to survey the survival ability of EV71 on different medium surface.Results The recovery rates of EV71 on medium surface ranged from 89 ％-93 ％.The survival time of EV71 on medium surface varied under different climatic conditions.The longest survival time of the virus was observed under the condition of 20 ℃ as the temperature and 80％ as the humidity.After 24 hours of incubation,the infectious titer on plastic surface reduced about 4 lg.After 72 hours of incubation,the infectious titer reduced at least 3.89 lg on cloth and wood surface.Conclusions Temperature and humidity can affect the survival time of EV71 on medium surface,which is longer in the condition of low temperature and high humidity.The survival ability of EV71 on natural cloth and wood surface is better than that on synthetic plastic surface.

Survivability of hand foot mouth disease virus in tap water / 中华传染病杂志

Objective To evaluate the survivability of hand foot mouth disease(HFMD)virus,in tap water for daily use.Methods HFMD viruses were isolated from cases of HFMD in Shanghai and Zhejiang from in 2008.Six isolated strains (five subtype of enterovirus 71 and one coxackie virus)were selected in this study.These viruses were mixed with chloride 1.0 mg/L tap-water and then inoculated into Vero cells.The cytopathic effect (CPE)was checked everyday in order to survey the survivability of each virus strain.The decline of virus survivability was analyzed by scatter diagram.Results These six strains of HMFD virus could survive longer than one month in tap water with initial chloride concentration of 1.0 mg/L and still had celluar infectivity.The survivabilities were varied between viruses isolated from different HFMD cases.Conclusions The survivabilities of enterovirus 71 and coxackie virus stains are quite strong in water.Therefore,the transmission route of water-borne pathogens should be monitored in regions using tap water during HFMD epidemic period.

Detection and clinical analysis of EV71,CA16 and respiratory virus con-infection in patients with hand,foot and mouse disease / 中华检验医学杂志

Objective To explore the situation of respiratory virus co-infection with EV71 and CA16 in patients with hand,foot and mouse disease(HFMD) ,and analyze the influence of co-infection on clinical aspects.Methods From June to October of 2010,there were 348 patients enrolled in the study,with 248 hospitalization cases and 100 mild outpatients.All the patients were diagnosed as HFMD in Beijing You-an Hospital.The viral RNA from the pharynx swab samples were extracted and reversely transcribed by RT-PCR.All the samples were detected with the EV71 and CA16 by real-time fluorescence quantitative PCR.Twelve kinds of respiratory viruses were detected by a commercial multiplex-PCR method.The PCR products were confirmed by electrophoresis.Chi square test was used in the data analysis.Results Of the 348 HFMD patients,36 subjects were detected as positive for respiratory virus co-infection.In the 248 hospitalization cases,111 cases were positive for EV71 or CA16,with eight cases identified with respiratory virus co-infection(7.2%); the other 137 cases were negative for EV71 and CA16,with eleven cases identified with respiratory virus co-infection(7.4%).There was not significant difference between respiratory virus co-infection and the identification of EV71 /CA16(x2 = 0.059,P ＞ 0.05).In the 100 mild outpatients positive for EV71 or CA16,seventeen cases were identified with respiratory virus co-infection(17%).The rate of respiratory virus co-infection in the mild outpatients was much higher than in the severe hospitalization patients(x2 = 4.830,P＜ 0.05).Among the 111 EV71(+) or CA16(+) inpatients,there were 101 cases diagnosed as severe cases(91.0%); similarly,there were 132 cases diagnosed as severe cases(96.4%) among the 137 EV71(-) CA16(-) cases.There was not difference between the identification of EV71/ CA 16 and illness of HFMD(x2 = 3.099,P ＞ 0.05).The leading respiratory virus being identified were HRV A/B,PIV3 and FLU A in the 348 HFMD patients.Conclusions Co-infection with respiratory virus exists in the HFMD patients. However,the respiratory virus infection has no significant influence to the state of HFMD illness.

Detection and subtype analysis of VP1 gene of enterovirus 71 strains isolated from children with hand,foot and mouth disease in Shanghai area during the first half year of 2009 / 中华传染病杂志

Objective To understand the characteristics of molecular epidemiology of enterovirus 71(EV71) in children with hand, foot and mouth disease (HFMD) in Shanghai area during the first half year of 2009. Methods Seventy-three throat swabs and 38 stool samples were collected from 95 hospitalized children with clinical diagnosis of HFMD in Children's Hospital of Fudan University during April to May 2009. TaqMan real-time reverse transcription-polymerase chain reaction (RT-PCR) and nest RT-PCR were used to detect EV71 VP1, followed by gene sequencing analysis. Results Six of the 73 throat swabs were EV71 positive with the detection rate of 8.2%. In the 38 stool samples, 24 were EV71 positive with the detection rate of 63.2%. Twenty-eight nested RT-PCR positive samples were sequenced and the genetic analysis showed that 27 were C4 subtype,which were absolute dominant strain and the other one was C2 subtype. The isolated strain from a fatal case was C4 subtype and there was no obvious mutation found in VP1 region. Conclusions EV71 is an important pathogen in HFMD children in Shanghai area during April to May 2009. C4 subtype strains are absolutely dominant, and accompanied by epidemic strains of subtype C2.