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Sample records for rapid enterovirus molecular

  1. Producing infectious enterovirus type 71 in a rapid strategy

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    Qin E-De

    2010-06-01

    Full Text Available Abstract Background Enterovirus 71 (EV71 is an etiologic agent of hand-foot-and-mouth disease (HFMD, and recent HFMD epidemics worldwide have been associated with a severe form of brainstem encephalitis associated with pulmonary edema and high case-fatality rates. EV71 contains a positive-sense single-stranded genome RNA of approximately 7400 bp in length which encodes a polyprotein with a single open reading frame (ORF, which is flanked by untranslated regions at both the 5' and 3' ends. Results A long distance RT-PCR assay was developed to amplify the full length genome cDNA of EV71 by using specific primes carrying a SP6 promoter. Then the in vitro synthesized RNA transcripts from the RT-PCR amplicons were then transfected into RD cells to produce the rescued virus. The rescued virus was further characterized by RT-PCR and indirect fluorescent-antibody (IFA assay in comparison with the wild type virus. The rescued viruses were infectious on RD cells and neurovirulent when intracerebrally injected into suckling mice. Conclusions Thus, we established a rapid method to produce the infectious full length cDNA of EV71 directly from RNA preparations and specific mutations can be easily engineered into the rescued enterovirus genome by this method.

  2. Molecular typing and characterization of a new serotype of human enterovirus (EV-B111) identified in China.

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    Zhang, Yong; Hong, Mei; Sun, Qiang; Zhu, Shuangli; Tsewang; Li, Xiaolei; Yan, Dongmei; Wang, Dongyan; Xu, Wenbo

    2014-04-01

    Molecular methods, based on sequencing the region encoding the complete VP1 or P1 protein, have enabled the rapid identification of new enterovirus serotypes. In the present study, the complete genome of a newly discovered enterovirus serotype, strain Q0011/XZ/CHN/2000 (hereafter referred to as Q0011), was sequenced and analyzed. The virus, isolated from a stool sample from a patient with acute flaccid paralysis in the Tibet region of China in 2000, was characterized by amplicon sequencing and comparison to a GenBank database of enterovirus nucleotide sequences. The nucleotide sequence encoding the complete VP1 capsid protein is most closely related to the sequences of viruses within the species enterovirus B (EV-B), but is less than 72.1% identical to the homologous sequences of the recognized human enterovirus serotypes, with the greatest homology to EV-B101 and echovirus 32. Moreover, the deduced amino acid sequence of the complete VP1 region is less than 84.7% identical to those of the recognized serotypes, suggesting that the strain is a new serotype of enterovirus within EV-B. The virus was characterized as a new enterovirus type, named EV-B111, by the Picornaviridae Study Group of the International Committee on Taxonomy of Viruses. Low positive rate and titer of neutralizing antibody against EV-B111 were found in the Tibet region of China. Nearly 50% of children ≤5 years had no neutralizing antibody against EV-B111. So the extent of transmission and the exposure of the population to this new EV are very limited. This is the first identification of a new serotype of human enterovirus in China, and strain Q0011 was designated the prototype strain of EV-B111. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. Detection of enterovirus RNA in cerebrospinal fluid : Comparison of two molecular assays

    NARCIS (Netherlands)

    de Crom, S. C. M.; Obihara, C. C.; van Loon, A. M.; Argilagos-Alvarez, A. A.; Peeters, M. F.; van Furth, A. M.; Rossen, J. W. A.

    Enterovirus (EV) and human parechovirus (HPeV) are a major cause of infection in childhood. A rapid diagnostic test may improve the management of patients with EV and HPeV infection. The aim of this study is to evaluate the performance of the GeneXpert enterovirus assay (GXEA) for detection of EV

  4. Enterovirus genotypes causing hand foot and mouth disease in Shanghai, China: a molecular epidemiological analysis

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    2013-01-01

    Background A rapid expansion of hand, foot, and mouth disease (HFMD) outbreaks has occurred and caused deaths in China in recent years, but little is known about the other etiologic agents except enterovirus 71 (EV71) and coxsackievirus A 16 (CA16). The objective of this study is to determine the genotype compositions of enterovirus causing HFMD in Shanghai and identify any associations between enterovirus types and clinical manifestations. Methods Stool specimens were collected from patients hospitalized for treatment of HFMD, from May 2010 to April 2011. Enterovirus was detected by reverse transcription PCR and directly genotyped by sequencing the PCR products. Phylogenetic analysis was based on the VP1 partial gene. Results Of 290 specimens, 277 (95.5%) tested positive for enterovirus. The major genotypes were EV71 (63.8%), CA10 (9.0%), CA6 (8.3%), CA16 (6.9%), CA12 (2.4%), and CA4 (1.4%). The EV71 strains belonged to the C4a subtype and CA16 belonged to the B subtype. CA6 was closely related to strains detected in Japan, Taiwan and China, and CA10, CA12 and CA4 were phylogenetically similar to other strains circulating in China. Mean hospital stays and the prevalence of complications in patients with EV71 infection were higher than those in patients in CA6, CA10 or CA16 infection (P enterovirus genotypes. It deserves our attention as early identification of enterovirus genotypes is important for diagnosis and treatment of HFMD patients. PMID:24148902

  5. Enterovirus genotypes causing hand foot and mouth disease in Shanghai, China: a molecular epidemiological analysis.

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    Xu, Menghua; Su, Liyun; Cao, Lingfeng; Zhong, Huaqing; Dong, Niuniu; Xu, Jin

    2013-10-22

    A rapid expansion of hand, foot, and mouth disease (HFMD) outbreaks has occurred and caused deaths in China in recent years, but little is known about the other etiologic agents except enterovirus 71 (EV71) and coxsackievirus A 16 (CA16). The objective of this study is to determine the genotype compositions of enterovirus causing HFMD in Shanghai and identify any associations between enterovirus types and clinical manifestations. Stool specimens were collected from patients hospitalized for treatment of HFMD, from May 2010 to April 2011. Enterovirus was detected by reverse transcription PCR and directly genotyped by sequencing the PCR products. Phylogenetic analysis was based on the VP1 partial gene. Of 290 specimens, 277 (95.5%) tested positive for enterovirus. The major genotypes were EV71 (63.8%), CA10 (9.0%), CA6 (8.3%), CA16 (6.9%), CA12 (2.4%), and CA4 (1.4%). The EV71 strains belonged to the C4a subtype and CA16 belonged to the B subtype. CA6 was closely related to strains detected in Japan, Taiwan and China, and CA10, CA12 and CA4 were phylogenetically similar to other strains circulating in China. Mean hospital stays and the prevalence of complications in patients with EV71 infection were higher than those in patients in CA6, CA10 or CA16 infection (P treatment of HFMD patients.

  6. Molecular and epidemiological study of enterovirus D68 in Taiwan.

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    Huang, Yuan-Pin; Lin, Tsuey-Li; Lin, Ting-Han; Wu, Ho-Sheng

    2017-08-01

    As an immunofluorescence assay for enterovirus D68 (EV-D68) is not available in the enteroviruses surveillance network in Taiwan, EV-D68 may be the actual pathogen of untypeable enterovirus-suspected isolates. The untypeable isolates collected from 2007 through 2014 were identified by nucleic acid amplification-based methods and sequencing of the VP1 region to analyze the phylogeny and epidemiology of EV-D68 in Taiwan. Twenty-nine EV-D68 isolates were sequenced, including 15 Cluster 3 and 14 Cluster 1 viruses. Approximately 41% of the patients were children under 5 years of age and their infections peaked in August. The ratio of male to female patients was 1.5 and 3.67 for Cluster 3 and Cluster 1, respectively. Fever and respiratory symptoms were commonly reported in EV-D68-infected patients. The results of phylogenetic analyses showed that EV-D68 isolates between 2007 and 2014 belonged to different clusters and existed for years, indicating that endemic circulation of EV-D68 existed in Taiwan. This study showed that EV-D68 has been endemic in Taiwan for some years despite a small number of positive cases. The continuous monitoring and efforts towards the improvement of diagnostic techniques are required to complete the surveillance system. This study provided the genetic and epidemiological information which could contribute to understanding the etiology and epidemiology of EV-D68. Copyright © 2015. Published by Elsevier B.V.

  7. Enterovirus D68 disease and molecular epidemiology in Australia.

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    Levy, Avram; Roberts, Jason; Lang, Jurissa; Tempone, Simone; Kesson, Alison; Dofai, Alfred; Daley, Andrew J; Thorley, Bruce; Speers, David J

    2015-08-01

    Enterovirus D68 (EV-D68) has received considerable recent attention as a cause of widespread respiratory illness. Neurological syndromes such as acute flaccid paralysis following EV-D68 infection have also been reported in a small number of cases. To summarize the clinical and epidemiological characteristics of laboratory confirmed EV-D68 cases in Australia. We combined EV-D68 data acquired through laboratory surveillance in Western Australia with cases from national enterovirus surveillance and regional acute flaccid paralysis (AFP) surveillance. Clinical data was obtained for EV-D68 cases and capsid protein sequences were used for phylogenetic analysis. Sporadic cases of EV-D68 were recorded in Australia since 2008, with peaks in activity during 2011 and 2013. EV-D68 was primarily associated with respiratory disease, but was also detected in cerebrospinal fluid of one patient and faeces of two patients presenting with AFP. EV-D68 has been circulating in Western Australia and is likely to have also been present in the wider region for a number of years, causing primarily respiratory disease. Detection of EV-D68 in cerebrospinal fluid of one patient and in faeces of two AFP cases reinforces the association between EV-D68 and neurological disease. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. The Current Status of the Disease Caused by Enterovirus 71 Infections: Epidemiology, Pathogenesis, Molecular Epidemiology, and Vaccine Development

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    Chang, Ping-Chin; Chen, Shou-Chien; Chen, Kow-Tong

    2016-01-01

    Enterovirus 71 (EV71) infections have a major public health impact in the Asia-Pacific region. We reviewed the epidemiology, pathogenesis, and molecular epidemiology of EV71 infection as well as EV71 vaccine development. Previous studies were found using the search terms “enterovirus 71” and “epidemiology” or “pathogenesis” or “molecular epidemiology” or “vaccine” in Medline and PubMed. Articles that were not published in the English language, manuscripts without an abstract, and opinion articles were excluded from the review. The reported epidemiology of cases caused by EV71 infection varied from country to country; seasonal variations in incidence were observed. Most cases of EV71 infection that resulted in hospitalization for complications occurred in children less than five years old. The brainstem was the most likely major target of EV71 infection. The emergence of the EV71 epidemic in the Asia-Pacific region has been associated with the circulation of different genetic lineages (genotypes B3, B4, C1, C2, and C4) that appear to be undergoing rapid evolutionary changes. The relationship between the gene structure of the EV71 virus and the factors that ensure its survival, circulation, and evasion of immunity is still unknown. EV71 infection has emerged as an important global public health problem. Vaccine development, including the development of inactivated whole-virus live attenuated, subviral particles, and DNA vaccines, has been progressing. PMID:27618078

  9. The Current Status of the Disease Caused by Enterovirus 71 Infections: Epidemiology, Pathogenesis, Molecular Epidemiology, and Vaccine Development.

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    Chang, Ping-Chin; Chen, Shou-Chien; Chen, Kow-Tong

    2016-09-09

    Enterovirus 71 (EV71) infections have a major public health impact in the Asia-Pacific region. We reviewed the epidemiology, pathogenesis, and molecular epidemiology of EV71 infection as well as EV71 vaccine development. Previous studies were found using the search terms "enterovirus 71" and "epidemiology" or "pathogenesis" or "molecular epidemiology" or "vaccine" in Medline and PubMed. Articles that were not published in the English language, manuscripts without an abstract, and opinion articles were excluded from the review. The reported epidemiology of cases caused by EV71 infection varied from country to country; seasonal variations in incidence were observed. Most cases of EV71 infection that resulted in hospitalization for complications occurred in children less than five years old. The brainstem was the most likely major target of EV71 infection. The emergence of the EV71 epidemic in the Asia-Pacific region has been associated with the circulation of different genetic lineages (genotypes B3, B4, C1, C2, and C4) that appear to be undergoing rapid evolutionary changes. The relationship between the gene structure of the EV71 virus and the factors that ensure its survival, circulation, and evasion of immunity is still unknown. EV71 infection has emerged as an important global public health problem. Vaccine development, including the development of inactivated whole-virus live attenuated, subviral particles, and DNA vaccines, has been progressing.

  10. Molecular evolution of enterovirus 68 detected in the Philippines.

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    Tadatsugu Imamura

    Full Text Available BACKGROUND: Detection of Enterovirus 68 (EV68 has recently been increased. However, underlying evolutionary mechanism of this increasing trend is not fully understood. METHODS: Nasopharyngeal swabs were collected from 5,240 patients with acute respiratory infections in the Philippines from June 2009 to December 2011. EV68 was detected by polymerase chain reaction (PCR targeting for 5' untranslated region (5'UTR, viral protein 1 (VP1, and VP4/VP2. Phylogenetic trees were generated using the obtained sequences. RESULTS: Of the 5,240 tested samples, 12 EV68 positive cases were detected between August and December in 2011 (detection rate, 0.23%. The detection rate was higher among inpatients than outpatients (p<0.0001. Among VP1 sequences detected from 7 patients in 2011, 5 in lineage 2 were diverged from those detected in the Philippines in 2008, however, 2 in lineage 3 were not diverged from strains detected in the Philippines in 2008 but closely associated with strains detected in the United States. Combined with our previous report, EV68 occurrences were observed twice in the Philippines within the last four years. CONCLUSIONS: EV68 detections might be occurring in cyclic patterns, and viruses might have been maintained in the community while some strains might have been newly introduced.

  11. Clinical characteristics and molecular epidemiology of Enterovirus infection in infants <3 months in a referral paediatric hospital of Barcelona.

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    Rodà, Diana; Pérez-Martínez, Esther; Cabrerizo, María; Trallero, Gloria; Martínez-Planas, Aina; Luaces, Carles; García-García, Juan-José; Muñoz-Almagro, Carmen; Launes, Cristian

    2015-11-01

    Enterovirus (EV) infection is common in infants, but the information with regard to the molecular epidemiology and the associations between types and clinical variables is very scarce. This study includes 195 children Enteroviruses cause a normally benign illness in young infants, except in some cases. • The molecular epidemiology of Enterovirus infection is not well known in European countries. • This study describes a large number of infants with Enterovirus infection and shows the seasonality of different types, and their associations with epidemiologic and clinical variables.

  12. Epidemic 2014 enterovirus D68 cross-reacts with human rhinovirus on a respiratory molecular diagnostic platform.

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    McAllister, Shane C; Schleiss, Mark R; Arbefeville, Sophie; Steiner, Marie E; Hanson, Ryan S; Pollock, Catherine; Ferrieri, Patricia

    2015-01-01

    Enterovirus D68 (EV-D68) is an emerging virus known to cause sporadic disease and occasional epidemics of severe lower respiratory tract infection. However, the true prevalence of infection with EV-D68 is unknown, due in part to the lack of a rapid and specific nucleic acid amplification test as well as the infrequency with which respiratory samples are analyzed by enterovirus surveillance programs. During the 2014 EV-D68 epidemic in the United States, we noted an increased frequency of "low-positive" results for human rhinovirus (HRV) detected in respiratory tract samples using the GenMark Diagnostics eSensor respiratory viral panel, a multiplex PCR assay able to detect 14 known respiratory viruses but not enteroviruses. We simultaneously noted markedly increased admissions to our Pediatric Intensive Care Unit for severe lower respiratory tract infections in patients both with and without a history of reactive airway disease. Accordingly, we hypothesized that these "low-positive" RVP results were due to EV-D68 rather than rhinovirus infection. Sequencing of the picornavirus 5' untranslated region (5'-UTR) of 49 samples positive for HRV by the GenMark RVP revealed that 33 (67.3%) were in fact EV-D68. Notably, the mean intensity of the HRV RVP result was significantly lower in the sequence-identified EV-D68 samples (20.3 nA) compared to HRV (129.7 nA). Using a cut-off of 40 nA for the differentiation of EV-D68 from HRV resulted in 94% sensitivity and 88% specificity. The robust diagnostic characteristics of our data suggest that the cross-reactivity of EV-D68 and HRV on the GenMark Diagnostics eSensor RVP platform may be an important factor to consider in making accurate molecular diagnosis of EV-D68 at institutions utilizing this system or other molecular respiratory platforms that may also cross-react.

  13. Rapid and highly sensitive detection of Enterovirus 71 by using nanogold-enhanced electrochemical impedance spectroscopy

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    Li, Hsing-Yuan; Tseng, Shing-Hua; Cheng, Tsai-Mu; Chu, Hsueh-Liang; Lu, Yu-Ning; Wang, Fang-Yu; Tu, Lung-Chen; Chang, Chia-Ching; Tsai, Li-Yun; Shieh, Juo-Yu; Yang, Jyh-Yuan; Juan, Chien-Chang

    2013-01-01

    Enterovirus 71 (EV71) infection is an emerging infectious disease causing neurological complications and/or death within two to three days after the development of fever and rash. A low viral titre in clinical specimens makes the detection of EV71 difficult. Conventional approaches for detecting EV71 are time consuming, poorly sensitive, or complicated, and cannot be used effectively for clinical diagnosis. Furthermore, EV71 and Coxsackie virus A16 (CA16) may cross react in conventional assays. Therefore, a rapid, highly sensitive, specific, and user-friendly test is needed. We developed an EV71-specific nanogold-modified working electrode for electrochemical impedance spectroscopy in the detection of EV71. Our results show that EV71 can be distinguished from CA16, Herpes simplex virus, and lysozyme, with the modified nanogold electrode being able to detect EV71 in concentrations as low as 1 copy number/50 μl reaction volume, and the duration between sample preparation and detection being 11 min. This detection platform may have the potential for use in point-of-care diagnostics. (paper)

  14. Rapid and highly sensitive detection of Enterovirus 71 by using nanogold-enhanced electrochemical impedance spectroscopy

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    Li, Hsing-Yuan; Tseng, Shing-Hua; Cheng, Tsai-Mu; Chu, Hsueh-Liang; Lu, Yu-Ning; Wang, Fang-Yu; Tsai, Li-Yun; Shieh, Juo-Yu; Yang, Jyh-Yuan; Juan, Chien-Chang; Tu, Lung-Chen; Chang, Chia-Ching

    2013-07-01

    Enterovirus 71 (EV71) infection is an emerging infectious disease causing neurological complications and/or death within two to three days after the development of fever and rash. A low viral titre in clinical specimens makes the detection of EV71 difficult. Conventional approaches for detecting EV71 are time consuming, poorly sensitive, or complicated, and cannot be used effectively for clinical diagnosis. Furthermore, EV71 and Coxsackie virus A16 (CA16) may cross react in conventional assays. Therefore, a rapid, highly sensitive, specific, and user-friendly test is needed. We developed an EV71-specific nanogold-modified working electrode for electrochemical impedance spectroscopy in the detection of EV71. Our results show that EV71 can be distinguished from CA16, Herpes simplex virus, and lysozyme, with the modified nanogold electrode being able to detect EV71 in concentrations as low as 1 copy number/50 μl reaction volume, and the duration between sample preparation and detection being 11 min. This detection platform may have the potential for use in point-of-care diagnostics.

  15. Rapid Resolution of Enterovirus 71-Associated Opsoclonus Myoclonus Syndrome on Intravenous Immunoglobulin

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    Ahmed Sahly MD

    2017-09-01

    Full Text Available Nonparaneoplastic opsoclonus–myoclonus ataxia syndrome is a rare neuroinflammatory condition featured by opsoclonus, myoclonus, ataxia, and cognitive behavioral disturbance. The authors report an observation of enterovirus 71-associated opsoclonus–myoclonus ataxia syndrome evolving toward full recovery on intravenous intravenous immunoglobulin (IG treatment. Based on this case report, enterovirus 71 should be added to the list of infectious agents likely involved in opsoclonus–myoclonus ataxia syndrome, including the emerging subgroup of opsoclonus–myoclonus ataxia syndrome recovering without aggressive or prolonged immunosuppressive intervention. Further studies are mandatory to define the precise role, incidence, treatment, and outcome of enterovirus 71 and other infectious agents in benign forms of opsoclonus–myoclonus ataxia syndrome.

  16. Molecular epidemiology of enterovirus 71, coxsackievirus A16 and A6 associated with hand, foot and mouth disease in Spain.

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    Cabrerizo, M; Tarragó, D; Muñoz-Almagro, C; Del Amo, E; Domínguez-Gil, M; Eiros, J M; López-Miragaya, I; Pérez, C; Reina, J; Otero, A; González, I; Echevarría, J E; Trallero, G

    2014-03-01

    Hand, foot and mouth disease (HFMD) is a childhood illness frequently caused by genotypes belonging to the enterovirus A species, including coxsackievirus (CV)-A16 and enterovirus (EV)-71. Between 2010 and 2012, several outbreaks and sporadic cases of HFMD occurred in different regions of Spain. The objective of the present study was to describe the enterovirus epidemiology associated with HFMD in the country. A total of 80 patients with HFMD or atypical rash were included. Detection and typing of the enteroviruses were performed directly in clinical samples using molecular methods. Enteroviruses were detected in 53 of the patients (66%). CV-A6 was the most frequent genotype, followed by CV-A16 and EV-71, but other minority types were also identified. Interestingly, during almost all of 2010, CV-A16 was the only causative agent of HFMD but by the end of the year and during 2011, CV-A6 became predominant, while CV-A16 was not detected. In 2012, however, both CV-A6 and CV-A16 circulated. EV-71 was associated with HFMD symptoms only in three cases during 2012. All Spanish CV-A6 sequences segregated into one major genetic cluster together with other European and Asian strains isolated between 2008 and 2011, most forming a particular clade. Spanish EV-71 strains belonged to subgenogroup C2, as did most of the European sequences circulated. In conclusion, the recent increase of HFMD cases in Spain and other European countries has been due to a larger incidence of circulating species A enteroviruses, mainly CV-A6 and CV-A16, and the emergence of new genetic variants of these viruses. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  17. Rapid and Accurate Sequencing of Enterovirus Genomes Using MinION Nanopore Sequencer.

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    Wang, Ji; Ke, Yue Hua; Zhang, Yong; Huang, Ke Qiang; Wang, Lei; Shen, Xin Xin; Dong, Xiao Ping; Xu, Wen Bo; Ma, Xue Jun

    2017-10-01

    Knowledge of an enterovirus genome sequence is very important in epidemiological investigation to identify transmission patterns and ascertain the extent of an outbreak. The MinION sequencer is increasingly used to sequence various viral pathogens in many clinical situations because of its long reads, portability, real-time accessibility of sequenced data, and very low initial costs. However, information is lacking on MinION sequencing of enterovirus genomes. In this proof-of-concept study using Enterovirus 71 (EV71) and Coxsackievirus A16 (CA16) strains as examples, we established an amplicon-based whole genome sequencing method using MinION. We explored the accuracy, minimum sequencing time, discrimination and high-throughput sequencing ability of MinION, and compared its performance with Sanger sequencing. Within the first minute (min) of sequencing, the accuracy of MinION was 98.5% for the single EV71 strain and 94.12%-97.33% for 10 genetically-related CA16 strains. In as little as 14 min, 99% identity was reached for the single EV71 strain, and in 17 min (on average), 99% identity was achieved for 10 CA16 strains in a single run. MinION is suitable for whole genome sequencing of enteroviruses with sufficient accuracy and fine discrimination and has the potential as a fast, reliable and convenient method for routine use. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

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

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    Angez, Mehar; Shaukat, Shahzad; Zahra, Rabaab; Khurshid, Adnan; Sharif, Salmaan; Alam, Muhammad Masroor; Zaidi, Syed Sohail Zahoor

    2015-01-01

    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. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. A novel enterovirus species identified from severe diarrheal goats.

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    Wang, Mingyue; He, Jia; Lu, Haibing; Liu, Yajing; Deng, Yingrui; Zhu, Lisai; Guo, Changming; Tu, Changchun; Wang, Xinping

    2017-01-01

    The Enterovirus genus of the family of Picornaviridae consists of 9 species of Enteroviruses and 3 species of Rhinoviruses based on the latest virus taxonomy. Those viruses contribute significantly to respiratory and digestive disorders in human and animals. Out of 9 Enterovirus species, Enterovirus E-G are closely related to diseases affecting on livestock industry. While enterovirus infection has been increasingly reported in cattle and swine, the enterovirus infections in small ruminants remain largely unknown. Virology, molecular and bioinformatics methods were employed to characterize a novel enterovirus CEV-JL14 from goats manifesting severe diarrhea with morbidity and mortality respectively up to 84% and 54% in China. CEV-JL14 was defined and proposed as a new Enterovirus species L within the genus of Enterovirus of the family Picornaviridae. CEV-JL14 had a complete genome sequence of 7461 nucleotides with an ORF encoding 2172 amino acids, and shared 77.1% of genomic sequence identity with TB4-OEV, an ovine enterovirus. Comparison of 5'-UTR and structural genes of CEV-JL14 with known Enterovirus species revealed highly genetic variations among CEV-JL14 with known Enterovirus species. VP1 nucleotide sequence identities of CEV-14 were 51.8%-53.5% with those of Enterovirus E and F, 30.9%-65.3% with Enterovirus G, and 43.8-51. 5% with Enterovirus A-D, respectively. CEV-JL14 was proposed as a novel species within the genus of Enterovirus according to the current ICTV demarcation criteria of enteroviruses. CEV-JL14 clustered phylogenetically to neither Enterovirus E and F, nor to Enterovirus G. It was defined and proposed as novel species L within the genus of Enterovirus. This is the first report of caprine enterovirus in China, the first complete genomic sequence of a caprine enterovirus revealed, and the unveiling of significant genetic variations between ovine enterovirus and caprine enterovirus, thus broadening the current understanding of enteroviruses.

  20. Rapid detection of Enterovirus and Coxsackievirus A10 by a TaqMan based duplex one-step real time RT-PCR assay.

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    Chen, Jingfang; Zhang, Rusheng; Ou, Xinhua; Yao, Dong; Huang, Zheng; Li, Linzhi; Sun, Biancheng

    2017-06-01

    A TaqMan based duplex one-step real time RT-PCR (rRT-PCR) assay was developed for the rapid detection of Coxsackievirus A10 (CV-A10) and other enterovirus (EVs) in clinical samples. The assay was fully evaluated and found to be specific and sensitive. When applied in 115 clinical samples, a 100% diagnostic sensitivity in CV-A10 detection and 97.4% diagnostic sensitivity in other EVs were found. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Development of single-step multiplex real-time RT-PCR assays for rapid diagnosis of enterovirus 71, coxsackievirus A6, and A16 in patients with hand, foot, and mouth disease.

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    Puenpa, Jiratchaya; Suwannakarn, Kamol; Chansaenroj, Jira; Vongpunsawad, Sompong; Poovorawan, Yong

    2017-10-01

    Real-time reverse-transcription polymerase chain reaction (rRT-PCR) to detect enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16) has facilitated the rapid and accurate identification of the two most common etiological agents underlying hand, foot, and mouth disease (HFMD). However, the worldwide emergence of CV-A6 infection in HFMD necessitates development of an improved multiplex rRT-PCR method. To rapidly determine the etiology of HFMD, two rRT-PCR assays using TaqMan probes were developed to differentiate among three selected common enteroviruses (EV-A71, CV-A16 and CV-A6) and to enable broad detection of enteroviruses (pan-enterovirus assay). No cross-reactions were observed with other RNA viruses examined. The detection limits of both assays were 10 copies per microliter for EV-A71, CV-A6 and CV-A16, and pan-enterovirus. The methods showed high accuracy (EV-A71, 90.6%; CV-A6, 92.0%; CV-A16, 100%), sensitivity (EV-A71, 96.5%; CV-A6, 95.8%; CV-A16, 99.0%), and specificity (EV-A71, 100%; CV-A6, 99.9%; CV-A16, 99.9%) in testing clinical specimens (n=1049) during 2014-2016, superior to those of conventional RT-PCR. Overall, the multiplex rRT-PCR assays enabled highly sensitive detection and rapid simultaneous typing of EV-A71, CV-A6 and CV-A16, and enteroviruses, rendering them feasible and attractive methods for large-scale surveillance of enteroviruses associated with HFMD outbreaks. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Improvement of the management of infants, children and adults with a molecular diagnosis of Enterovirus meningitis during two observational study periods.

    Directory of Open Access Journals (Sweden)

    Christine Archimbaud

    Full Text Available Enteroviruses (EVs are a major cause of aseptic meningitis, and RNA detection using molecular assay is the gold standard diagnostic test. The aim of this study was to assess the impact of an EV positive diagnosis on the clinical management of patients admitted for meningitis over the course of two observational study periods (2005 and 2008-09 in the same clinical departments. We further investigated in multivariate analysis various factors possibly associated with hospital length of stay (LOS in all age groups (infants, children, and adults. The results showed an overall improvement in the management of patients (n = 142 between the study periods, resulting in a significantly shorter hospital LOS for adults and children, and a shorter duration of antibiotic use for adults and infants. In multivariate analysis, we observed that the time from molecular test results to discharge of patients and the median duration of antibiotic treatment were associated with an increase in LOS in all age groups. In addition, among adults, the turnaround time of the molecular assay was significantly correlated with LOS. The use of CT scan in children and hospital admission outside the peak of EV prevalence in infants tended to increase LOS. In conclusion, the shorter length of stay of patients with meningitis in this study was due to various factors including the rapidity of the EV molecular test (particularly in adults, greater physician responsiveness after a positive result (in adults and children, and greater experience on the part of physicians in handling EV meningitis, as evidenced by the shorter duration of antibiotic use in adults and infants.

  3. Improvement of the Management of Infants, Children and Adults with a Molecular Diagnosis of Enterovirus Meningitis during Two Observational Study Periods

    Science.gov (United States)

    Archimbaud, Christine; Ouchchane, Lemlih; Mirand, Audrey; Chambon, Martine; Demeocq, François; Labbé, André; Laurichesse, Henri; Schmidt, Jeannot; Clavelou, Pierre; Aumaître, Olivier; Regagnon, Christel; Bailly, Jean-Luc; Henquell, Cécile; Peigue-Lafeuille, Hélène

    2013-01-01

    Enteroviruses (EVs) are a major cause of aseptic meningitis, and RNA detection using molecular assay is the gold standard diagnostic test. The aim of this study was to assess the impact of an EV positive diagnosis on the clinical management of patients admitted for meningitis over the course of two observational study periods (2005 and 2008–09) in the same clinical departments. We further investigated in multivariate analysis various factors possibly associated with hospital length of stay (LOS) in all age groups (infants, children, and adults). The results showed an overall improvement in the management of patients (n = 142) between the study periods, resulting in a significantly shorter hospital LOS for adults and children, and a shorter duration of antibiotic use for adults and infants. In multivariate analysis, we observed that the time from molecular test results to discharge of patients and the median duration of antibiotic treatment were associated with an increase in LOS in all age groups. In addition, among adults, the turnaround time of the molecular assay was significantly correlated with LOS. The use of CT scan in children and hospital admission outside the peak of EV prevalence in infants tended to increase LOS. In conclusion, the shorter length of stay of patients with meningitis in this study was due to various factors including the rapidity of the EV molecular test (particularly in adults), greater physician responsiveness after a positive result (in adults and children), and greater experience on the part of physicians in handling EV meningitis, as evidenced by the shorter duration of antibiotic use in adults and infants. PMID:23874676

  4. A novel enterovirus species identified from severe diarrheal goats.

    Directory of Open Access Journals (Sweden)

    Mingyue Wang

    Full Text Available The Enterovirus genus of the family of Picornaviridae consists of 9 species of Enteroviruses and 3 species of Rhinoviruses based on the latest virus taxonomy. Those viruses contribute significantly to respiratory and digestive disorders in human and animals. Out of 9 Enterovirus species, Enterovirus E-G are closely related to diseases affecting on livestock industry. While enterovirus infection has been increasingly reported in cattle and swine, the enterovirus infections in small ruminants remain largely unknown.Virology, molecular and bioinformatics methods were employed to characterize a novel enterovirus CEV-JL14 from goats manifesting severe diarrhea with morbidity and mortality respectively up to 84% and 54% in China.CEV-JL14 was defined and proposed as a new Enterovirus species L within the genus of Enterovirus of the family Picornaviridae. CEV-JL14 had a complete genome sequence of 7461 nucleotides with an ORF encoding 2172 amino acids, and shared 77.1% of genomic sequence identity with TB4-OEV, an ovine enterovirus. Comparison of 5'-UTR and structural genes of CEV-JL14 with known Enterovirus species revealed highly genetic variations among CEV-JL14 with known Enterovirus species. VP1 nucleotide sequence identities of CEV-14 were 51.8%-53.5% with those of Enterovirus E and F, 30.9%-65.3% with Enterovirus G, and 43.8-51. 5% with Enterovirus A-D, respectively. CEV-JL14 was proposed as a novel species within the genus of Enterovirus according to the current ICTV demarcation criteria of enteroviruses.CEV-JL14 clustered phylogenetically to neither Enterovirus E and F, nor to Enterovirus G. It was defined and proposed as novel species L within the genus of Enterovirus. This is the first report of caprine enterovirus in China, the first complete genomic sequence of a caprine enterovirus revealed, and the unveiling of significant genetic variations between ovine enterovirus and caprine enterovirus, thus broadening the current understanding of

  5. A novel enterovirus species identified from severe diarrheal goats

    Science.gov (United States)

    Liu, Yajing; Deng, Yingrui; Zhu, Lisai; Guo, Changming; Tu, Changchun; Wang, Xinping

    2017-01-01

    Backgrounds The Enterovirus genus of the family of Picornaviridae consists of 9 species of Enteroviruses and 3 species of Rhinoviruses based on the latest virus taxonomy. Those viruses contribute significantly to respiratory and digestive disorders in human and animals. Out of 9 Enterovirus species, Enterovirus E-G are closely related to diseases affecting on livestock industry. While enterovirus infection has been increasingly reported in cattle and swine, the enterovirus infections in small ruminants remain largely unknown. Methods Virology, molecular and bioinformatics methods were employed to characterize a novel enterovirus CEV-JL14 from goats manifesting severe diarrhea with morbidity and mortality respectively up to 84% and 54% in China. Results CEV-JL14 was defined and proposed as a new Enterovirus species L within the genus of Enterovirus of the family Picornaviridae. CEV-JL14 had a complete genome sequence of 7461 nucleotides with an ORF encoding 2172 amino acids, and shared 77.1% of genomic sequence identity with TB4-OEV, an ovine enterovirus. Comparison of 5’-UTR and structural genes of CEV-JL14 with known Enterovirus species revealed highly genetic variations among CEV-JL14 with known Enterovirus species. VP1 nucleotide sequence identities of CEV-14 were 51.8%-53.5% with those of Enterovirus E and F, 30.9%-65.3% with Enterovirus G, and 43.8–51. 5% with Enterovirus A-D, respectively. CEV-JL14 was proposed as a novel species within the genus of Enterovirus according to the current ICTV demarcation criteria of enteroviruses. Conclusions CEV-JL14 clustered phylogenetically to neither Enterovirus E and F, nor to Enterovirus G. It was defined and proposed as novel species L within the genus of Enterovirus. This is the first report of caprine enterovirus in China, the first complete genomic sequence of a caprine enterovirus revealed, and the unveiling of significant genetic variations between ovine enterovirus and caprine enterovirus, thus broadening the

  6. Enterovirus D68

    Science.gov (United States)

    Non-polio enterovirus ... Centers for Disease Control and Prevention website. Enterovirus D68. www.cdc.gov/non-polio-enterovirus/about/ev-d68.html#us . Updated October 20, 2017. Accessed October 26, 2017. Romero JR, Modlin ...

  7. Non-Polio Enterovirus

    Science.gov (United States)

    ... Controls Cancel Submit Search The CDC Non-Polio Enterovirus Note: Javascript is disabled or is not supported ... visit this page: About CDC.gov . Non-Polio Enterovirus Home About Non-Polio Enterovirus Symptoms Transmission Prevention & ...

  8. Structure determination of enterovirus 71

    Energy Technology Data Exchange (ETDEWEB)

    Plevka, Pavel; Perera, Rushika; Cardosa, Jane; Kuhn, Richard J.; Rossmann, Michael G. (Purdue); (Sentinext)

    2013-02-20

    Enterovirus 71 is a picornavirus that causes hand, foot and mouth disease but may induce fatal neurological illness in infants and young children. Enterovirus 71 crystallized in a body-centered orthorhombic space group with two particles in general orientations in the crystallographic asymmetric unit. Determination of the particle orientations required that the locked rotation function excluded the twofold symmetry axes from the set of icosahedral symmetry operators. This avoided the occurrence of misleading high rotation-function values produced by the alignment of icosahedral and crystallographic twofold axes. Once the orientations and positions of the particles had been established, the structure was solved by molecular replacement and phase extension.

  9. Microchip capillary electrophoresis with laser-induced fluorescence combined with one-step duplex reverse-transcription polymerase chain reaction for the rapid detection of Enterovirus 71 and Coxsackievirus A16 in throat swab specimens.

    Science.gov (United States)

    Jia, Ruan; Chengjun, Sun; Heng, Chen; Chen, Zhou; Yuanqian, Li; Yongxin, Li

    2015-07-01

    Enterovirus 71 and Coxsackievirus A16 are the main pathogens causing hand-foot-mouth disease. In this paper, microchip capillary electrophoresis with laser-induced fluorescence combined with one-step duplex reverse transcript-polymerase chain reaction has been developed for the detection of Enterovirus 71 and Coxsackievirus A16 in throat swab specimens. The specific reverse transcription-polymerase chain reaction amplicons labeled with SYBR Orange were separated by microchip capillary electrophoresis and detected by laser induced fluorescence detector within 7 min. The intraday and interday relative standard deviation of migration time for DNA Marker was in the range of 1.36-2.94 and 2.78-3.96%, respectively. The detection limits were as low as 2.06 × 10(3) copies/mL for Enterovirus 71 and 5 × 10(3) copies/mL for Coxsackievirus A16. No cross-reactivity was observed with rotavirus, astrovirus, norovirus, and adenovirus, which showed good specificity of the method. This assay was validated using 100 throat swab specimens that were detected by real-time reverse-transcript polymerase chain reaction in parallel and the two methods produced the same results. This study provided a rapid, sensitive and specific method for the detection of Enterovirus 71 and Coxsackievirus A16, which make a contribution to significant time and cost saving for the identification and treatment of patients. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Rapid detection of enterovirus in cerebrospinal fluid by a fully-automated PCR assay is associated with improved management of aseptic meningitis in adult patients.

    Science.gov (United States)

    Giulieri, Stefano G; Chapuis-Taillard, Caroline; Manuel, Oriol; Hugli, Olivier; Pinget, Christophe; Wasserfallen, Jean-Blaise; Sahli, Roland; Jaton, Katia; Marchetti, Oscar; Meylan, Pascal

    2015-01-01

    Enterovirus (EV) is the most frequent cause of aseptic meningitis (AM). Lack of microbiological documentation results in unnecessary antimicrobial therapy and hospitalization. To assess the impact of rapid EV detection in cerebrospinal fluid (CSF) by a fully-automated PCR (GeneXpert EV assay, GXEA) on the management of AM. Observational study in adult patients with AM. Three groups were analyzed according to EV documentation in CSF: group A = no PCR or negative PCR (n=17), group B = positive real-time PCR (n = 20), and group C = positive GXEA (n = 22). Clinical, laboratory and health-care costs data were compared. Clinical characteristics were similar in the 3 groups. Median turn-around time of EV PCR decreased from 60 h (IQR (interquartile range) 44-87) in group B to 5h (IQR 4-11) in group C (p<0.0001). Median duration of antibiotics was 1 (IQR 0-6), 1 (0-1.9), and 0.5 days (single dose) in groups A, B, and C, respectively (p < 0.001). Median length of hospitalization was 4 days (2.5-7.5), 2 (1-3.7), and 0.5 (0.3-0.7), respectively (p < 0.001). Median hospitalization costs were $5458 (2676-6274) in group A, $2796 (2062-5726) in group B, and $921 (765-1230) in group C (p < 0.0001). Rapid EV detection in CSF by a fully-automated PCR improves management of AM by significantly reducing antibiotic use, hospitalization length and costs. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Development of a multiplex polymerase chain reaction assay for simultaneous identification of human enterovirus 71 and coxsackievirus A16

    Science.gov (United States)

    Thao, Nguyen Thi Thanh; Ngoc, Nguyen Thi Kim; Tú, Phan Văn; Thúy, Trần Thi; Cardosa, Mary Jane; McMinn, Peter Charles; Phuektes, Patchara

    2010-01-01

    Human enterovirus 71 (HEV71) and coxsackievirus A16 (CVA16) are two major aetiological agents of hand, foot and mouth disease (HFMD) in children. Recently there have been several large outbreaks of HFMD in Vietnam and the Asia-Pacific region. In this study, a multiplex RT-PCR assay was developed in order to detect simultaneously HEV71, CVA16 and other human enteroviruses. Enterovirus detection was performed with a mixture of three pairs of oligonucleotide primers: one pair of published primers for amplifying all known enterovirus genomes and two new primer pairs specific for detection of the VP1 genes of HEV71 and CVA16. Enterovirus isolates, CVA16 and HEV71 strains identified previously from patients with HFMD were examined to evaluate the sensitivity and specificity of the multiplex RT-PCR assay. The assay was then applied to the direct detection of these viruses in clinical specimens obtained from HFMD cases identified at Children's Hospital Number 2, Ho Chi Minh City, Vietnam. The multiplex RT-PCR assay showed 100% specificity in screening for enteroviruses and in identifying HEV71 and CVA16. Similar results were obtained when using the multiplex RT-PCR assay to screen for enteroviruses and to identify HEV71 and CVA16 in clinical specimens obtained from HFMD cases identified at the hospital. This multiplex RT-PCR assay is a rapid, sensitive and specific assay for the diagnosis of HEV71 or CVA16 infection in cases of HFMD and is also potentially useful for molecular epidemiological investigations. PMID:20863857

  12. Improved detection limit in rapid detection of human enterovirus 71 and coxsackievirus A16 by a novel reverse transcription-isothermal multiple-self-matching-initiated amplification assay.

    Science.gov (United States)

    Ding, Xiong; Nie, Kai; Shi, Lei; Zhang, Yong; Guan, Li; Zhang, Dan; Qi, Shunxiang; Ma, Xuejun

    2014-06-01

    Rapid detection of human enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) is important in the early phase of hand-foot-and-mouth disease (HFMD). In this study, we developed and evaluated a novel reverse transcription-isothermal multiple-self-matching-initiated amplification (RT-IMSA) assay for the rapid detection of EV71 and CVA16 by use of reverse transcriptase, together with a strand displacement DNA polymerase. Real-time RT-IMSA assays using a turbidimeter and visual RT-IMSA assays to detect EV71 and CVA16 were established and completed in 1 h, and the reported corresponding real-time reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assays targeting the same regions of the VP1 gene were adopted as parallel tests. Through testing VP1 RNAs transcribed in vitro, the real-time RT-IMSA assays exhibited better linearity of quantification, with R(2) values of 0.952 (for EV71) and 0.967 (for CVA16), than the real-time RT-LAMP assays, which had R(2) values of 0.803 (for EV71) and 0.904 (for CVA16). Additionally, the detection limits of the real-time RT-IMSA assays (approximately 937 for EV71 and 67 for CVA16 copies/reaction) were higher than those of real-time RT-LAMP assays (approximately 3,266 for EV71 and 430 for CVA16 copies/reaction), and similar results were observed in the visual RT-IMSA assays. The new approaches also possess high specificities for the corresponding targets, with no cross-reactivity observed. In clinical assessment, compared to commercial reverse transcription-quantitative PCR (qRT-PCR) kits, the diagnostic sensitivities of the real-time RT-IMSA assays (96.4% for EV71 and 94.6% for CVA16) were higher than those of the real-time RT-LAMP assays (91.1% for EV71 and 90.8% for CVA16). The visual RT-IMSA assays also exhibited the same results. In conclusion, this proof-of-concept study suggests that the novel RT-IMSA assay is superior to the RT-LAMP assay in terms of detection limit and has the potential to rapidly detect EV71

  13. Development and implementation of real-time nucleic acid amplification for the detection of enterovirus infections in comparison to rapid culture of various clinical specimens

    NARCIS (Netherlands)

    van Doornum, G J J; Schutten, Martin; Voermans, J; Guldemeester, G J J; Niesters, H G M

    2007-01-01

    Several real-time PCR and nucleic acid sequence-based amplification (NASBA) primer pairs and a modified real-time PCR primer pair for the detection of enteroviruses were compared. The modified real-time PCR primer pair was evaluated on clinical samples in comparison with cell culture using the

  14. ENTEROVIRUS INFECTION: VARIETY OF ETIOLOGICAL FACTORS AND CLINICAL MANIFESTATIONS

    Directory of Open Access Journals (Sweden)

    O. I. Kanaeva

    2014-01-01

    Full Text Available Abstract. Enteroviruses are widely distributed human infectious pathogens. In spite of infection a disease does not manifest in majority number of cases. However, in some infected persons the different kind of symptoms can be observed; from common cold signs up to  aseptic (serous meningitis and myocarditis. Severe enteroviral cases with lethal outcomes are rarely reported. Ability of enteroviruses to cause large outbreaks and even epidemic distribution is very significant for health care systems. Taking in account a high genetic diversity of enteroviruses it is possible appearance of new highly pathogenic strains in the future. In some countries including the Russian Federation the permanent surveillance for enteroviral infections is provided besides of WHO polio elimination program. The laboratory diagnostics of enterovirus infections is complicated by numerous of pathogen serotypes. Thus, classical virological methods should be supported by molecular-biological tools to sequence pathogen genome and to define phylogenetic relations between different enterovirus strains.

  15. Molecular epidemiology of enterovirus and parechovirus infections according to patient age over a 4-year period in Spain.

    Science.gov (United States)

    Cabrerizo, María; Díaz-Cerio, María; Muñoz-Almagro, Carmen; Rabella, Núria; Tarragó, David; Romero, María Pilar; Pena, María José; Calvo, Cristina; Rey-Cao, Sonia; Moreno-Docón, Antonio; Martínez-Rienda, Inés; Otero, Almudena; Trallero, Gloria

    2017-03-01

    The epidemiology and clinical association of enterovirus (EV) and parechovirus (HPeV) infections, as well as the type-distribution-according-to-age, were determined during a 4-year study period in Spain. During 2010-2013, a total of 21,832 clinical samples were screened for EV and the detection frequency was 6.5% (1,430). Of the total EV-negative samples, only 1,873 samples from 2011 to 2013 were available for HPeV testing. HPeV was detected in 42 (2%) of them. Positive samples were genotyped using PCR and sequencing. EV infections occurred in all age groups of patients: neonates (17%), children 28 days to 2 years (29%), children 2-14 years (40%), and adults (14%). Thirty-four different EV types were identified. HPeV infections were detected exclusively in infants 2 years and adults (P < 0.05). Clinically, meningitis was associated with EV (P < 0.01) whereas, encephalitis was more frequent in HPeV-infected patients. CV-B types were associated with myocarditis (90%; P < 0.05) and EV species A with hand-foot-mouth-disease/atypical exanthema (88%; P < 0.05). J. Med. Virol. 89:435-442, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Immunology in the clinic review series; focus on type 1 diabetes and viruses: the enterovirus link to type 1 diabetes: critical review of human studies

    Science.gov (United States)

    Stene, L C; Rewers, M

    2012-01-01

    OTHER THEMES PUBLISHED IN THIS IMMUNOLOGY IN THE CLINIC REVIEW SERIES Metabolic diseases, host responses, cancer, autoinflammatory diseases, allergy. The hypothesis that under some circumstances enteroviral infections can lead to type 1 diabetes (T1D) was proposed several decades ago, based initially on evidence from animal studies and sero-epidemiology. Subsequently, enterovirus RNA has been detected more frequently in serum of patients than in control subjects, but such studies are susceptible to selection bias and reverse causality. Here, we review critically recent evidence from human studies, focusing on longitudinal studies with potential to demonstrate temporal association. Among seven longitudinal birth cohort studies, the evidence that enterovirus infections predict islet autoimmunity is quite inconsistent in our interpretation, due partially, perhaps, to heterogeneity in study design and a limited number of subjects studied. An association between enterovirus and rapid progression from autoimmunity to T1D was reported by one longitudinal study, but although consistent with evidence from animal models, this novel observation awaits replication. It is possible that a potential association with initiation and/or progression of islet autoimmunity can be ascribed to a subgroup of the many enterovirus serotypes, but this has still not been investigated properly. There is a need for larger studies with frequent sample intervals and collection of specimens of sufficient quality and quantity for detailed characterization of enterovirus. More research into the molecular epidemiology of enteroviruses and enterovirus immunity in human populations is also warranted. Ultimately, this knowledge may be used to devise strategies to reduce the risk of T1D in humans. PMID:22385232

  17. Rapid molecular technique to distinguish Fusarium species

    CSIR Research Space (South Africa)

    Lodolo, EJ

    1993-03-01

    Full Text Available The nuclear DNA (nDNA) of different isolates of three closely related, toxin-producing Fusarium species, F. moniliforme, F. nygamai and F. napiforme, was compared to ascertain the sensitivity of a molecular method to distinguish these three species...

  18. Rapid differentiation of rocky mountain spotted fever from chickenpox, measles, and enterovirus infections and bacterial meningitis by frequency-pulsed electron capture gas-liquid chromatographic analysis of sera.

    Science.gov (United States)

    Brooks, J B; McDade, J E; Alley, C C

    1981-01-01

    Normal sera and sera from patients with Rocky Mountain spotted fever, chickenpox, enterovirus infections, measles, and Neisseria meningitidis infections were extracted with organic solvents under acidic and basic conditions and then derivatized with trichloroethanol or heptafluorobutyric anhydride-ethanol to form electron-capturing derivatives of organic acids, alcohols, and amines. The derivatives were analyzed by frequency-pulsed electron capture gas-liquid chromatography (FPEC-GLC). There were unique differences in the FPEC-GLC profiles of sera obtained from patients with these respective diseases. With Rocky Mountain spotted fever patients, typical profiles were detected as early as 1 day after onset of disease and before antibody could be detected in the serum. Rapid diagnosis of Rocky Mountain spotted fever by FPEC-GLC could permit early and effective therapy, thus preventing many deaths from this disease. PMID:7276147

  19. Human enterovirus 71 epidemics: what's next?

    Science.gov (United States)

    Yip, Cyril C. Y.; Lau, Susanna K. P.; Woo, Patrick C. Y.; Yuen, Kwok-Yung

    2013-01-01

    Human enterovirus 71 (EV71) epidemics have affected various countries in the past 40 years. EV71 commonly causes hand, foot and mouth disease (HFMD) in children, but can result in neurological and cardiorespiratory complications in severe cases. Genotypic changes of EV71 have been observed in different places over time, with the emergence of novel genotypes or subgenotypes giving rise to serious outbreaks. Since the late 1990s, intra- and inter-typic recombination events in EV71 have been increasingly reported in the Asia-Pacific region. In particular, ‘double-recombinant’ EV71 strains belonging to a novel genotype D have been predominant in mainland China and Hong Kong over the last decade, though co-circulating with a minority of other EV71 subgenotypes and coxsackie A viruses. Continuous surveillance and genome studies are important to detect potential novel mutants or recombinants in the near future. Rapid and sensitive molecular detection of EV71 is of paramount importance in anticipating and combating EV71 outbreaks. PMID:24119538

  20. Tissue tropism, pathology and pathogenesis of enterovirus infection.

    Science.gov (United States)

    Muehlenbachs, Atis; Bhatnagar, Julu; Zaki, Sherif R

    2015-01-01

    Enteroviruses are very common and cause infections with a diverse array of clinical features. Enteroviruses are most frequently considered by practising pathologists in cases of aseptic meningitis, encephalitis, myocarditis and disseminated infections in neonates and infants. Congenital infections have been reported and transplacental transmission is thought to occur. Although skin biopsies during hand, foot and mouth disease are infrequently obtained, characteristic dermatopathological findings can be seen. Enteroviruses have been implicated in lower respiratory tract infections. This review highlights histopathological features of enterovirus infection and discusses diagnostic modalities for formalin-fixed paraffin-embedded tissues and their associated pitfalls. Immunohistochemistry can detect enterovirus antigen within cells of affected tissues; however, assays can be non-specific and detect other viruses. Molecular methods are increasingly relied upon but, due to the high frequency of asymptomatic enteroviral infections, clinical-pathological correlation is needed to determine significance. Of note, diagnostic assays on central nervous system or cardiac tissues from immunocompetent patients with prolonged disease courses are most often negative. Histopathological, immunohistochemical and molecular studies performed on clinical specimens also provide insight into enteroviral tissue tropism and pathogenesis. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

  1. Cellular receptors for human enterovirus species A

    Directory of Open Access Journals (Sweden)

    Yorihiro eNishimura

    2012-03-01

    Full Text Available Human enterovirus species A (HEV-A is one of the four species of HEV in the genus Enterovirus in the family Picornaviridae. Among HEV-A, coxsackievirus A16 (CVA16 and enterovirus 71 (EV71 are the major causative agents of hand, foot, and mouth disease (HFMD. Some other types of HEV-A are commonly associated with herpangina. Although HFMD and herpangina due to HEV-A are common febrile diseases among infants and children, EV71 can cause various neurological diseases, such as aseptic meningitis and fatal encephalitis.Recently, two human transmembrane proteins, P-selectin glycoprotein ligand-1 (PSGL-1 and scavenger receptor class B, member 2 (SCARB2, were identified as functional receptors for EV71 and CVA16. In in vitro infection experiments using the prototype HEV-A strains, PSGL-1 and SCARB2 could be responsible for the specific receptors for EV71 and CVA16. However, the involvement of both receptors in the in vitro and in vivo infections of clinical isolates of HEV-A has not been clarified yet. To elucidate a diverse array of the clinical outcome of HEV-A-associated diseases, the identification and characterization of HEV-A receptors may provide useful information in understanding the HEV-A pathogenesis at a molecular level.

  2. AN INTEGRATED CELL CULTURE/RT-PCR METHOD FOR DETECTING ENTEROVIRUS IN WATER

    Science.gov (United States)

    Echovirus and coxsackievirus can cause mild to severe disease following consumption of contaminated drinking water. However, comprehensive occurrence studies of enteroviruses in drinking water matrices are limited, in part because of the lack of available methods that are rapid, ...

  3. Pathological examinations of an enterovirus 71 infection: an autopsy case.

    Science.gov (United States)

    Gao, Lulu; Lin, Peixin; Liu, Shuguang; Lei, Bin; Chen, Qing; Yu, Shouyi; Shen, Hong

    2014-01-01

    We report an 8-month-old female infant with the fatal enterovirus 71 infection here. Clinically, she developed respiratory failure and severe pulmonary edema rapidly. Histologically, the lung specimen showed diffuse, severe pulmonary congestion and edema with focal intra-alveolar hemorrhage and typical features of acute encephalitis were easily identified under light microscope. Immunohistochemically, enterovirus 71 antigen was positive in the cerebella and brainstem. We measured the viral loads of different tissues and found that the brainstem and mesenteric lymph nodes showed the highest viral loads among all tissues. We hope that our case report may help to have a better understanding of the enterovirus 71 infection and provide clues to the prevention and treatment of this disease.

  4. Production of reference enteroviruses.

    Science.gov (United States)

    Kalter, S S; Rodriguez, A R; Armour, V

    1968-10-01

    Forty-five human enterovirus reagents of certified purity and quality were prepared for use as seed viruses and as immunizing antigens. One of the reagents was ampouled as "untreated" seed virus, whereas 14 were ampouled as "MgCl(2)-stabilized" reagents. The remaining 30 reagents were ampouled as "untreated" seed viruses and as "MgCl(2)-stabilized" reagents. Thirty of the reagents were propagated on primary African green monkey kidney cells, 3 on primary baboon kidney cells, 3 on primary rhesus monkey kidney cells, and the remaining 9 on human amnion cells. Forty-two of the viral antigens were concentrated for use in the production of high-titered specific antisera in large animals.

  5. The Autophagic Machinery in Enterovirus Infection.

    Science.gov (United States)

    Lai, Jeffrey K F; Sam, I-Ching; Chan, Yoke Fun

    2016-01-27

    The Enterovirus genus of the Picornaviridae family comprises many important human pathogens, including polioviruses, rhinovirus, enterovirus A71, and enterovirus D68. They cause a wide variety of diseases, ranging from mild to severe life-threatening diseases. Currently, no effective vaccine is available against enteroviruses except for poliovirus. Enteroviruses subvert the autophagic machinery to benefit their assembly, maturation, and exit from host. Some enteroviruses spread between cells via a process described as autophagosome-mediated exit without lysis (AWOL). The early and late phases of autophagy are regulated through various lipids and their metabolizing enzymes. Some of these lipids and enzymes are specifically regulated by enteroviruses. In the present review, we summarize the current understanding of the regulation of autophagic machinery by enteroviruses, and provide updates on recent developments in this field.

  6. Genetic Determinants of Enterovirus Infections

    DEFF Research Database (Denmark)

    Witsø, Elisabet; Cinek, Ondrej; Tapia, German

    2015-01-01

    Enteroviruses have been suggested as triggers of type 1 diabetes (T1D). We aimed to assess whether established T1D susceptibility single nucleotide polymorphisms (SNPs) and candidate SNPs in innate immune genes were associated with the frequency of enterovirus infection in otherwise healthy...... essentially all enterovirus serotypes, in 7,393 longitudinal stool samples collected monthly (age range 3-36 months). The most significant association was with two T1D SNPs, rs12150079 (ZPBP2/ORMDL3/GSDMB region) (enterovirus frequency: AA 7.3%, AG 8.7%, GG 9.7%, RR = 0.86, overall p = 1.87E-02) and rs229541...... (C1QTNF6/SSTR3/RAC2) (enterovirus frequency: CC 7.8%, CT 9.7%, TT 9.4%, RR = 1.13, overall p = 3.6E-02), followed by TLR8 (rs2407992) (p = 3.8E-02), TLR3 (1914926) (p = 4.9E-02), and two other T1D SNPs (IFIH1 rs3747517, p = 4.9E-02 and PTPN22, rs2476601, p = 5.3E-02). However, the quantile...

  7. Molecular Comparison and Evolutionary Analyses of VP1 Nucleotide Sequences of New African Human Enterovirus 71 Isolates Reveal a Wide Genetic Diversity

    Science.gov (United States)

    Nougairède, Antoine; Joffret, Marie-Line; Deshpande, Jagadish M.; Dubot-Pérès, Audrey; Héraud, Jean-Michel

    2014-01-01

    Most circulating strains of Human enterovirus 71 (EV-A71) have been classified primarily into three genogroups (A to C) on the basis of genetic divergence between the 1D gene, which encodes the VP1 capsid protein. The aim of the present study was to provide further insights into the diversity of the EV-A71 genogroups following the recent description of highly divergent isolates, in particular those from African countries, including Madagascar. We classified recent EV-A71 isolates by a large comparison of 3,346 VP1 nucleotidic sequences collected from GenBank. Analysis of genetic distances and phylogenetic investigations indicated that some recently-reported isolates did not fall into the genogroups A-C and clustered into three additional genogroups, including one Indian genogroup (genogroup D) and 2 African ones (E and F). Our Bayesian phylogenetic analysis provided consistent data showing that the genogroup D isolates share a recent common ancestor with the members of genogroup E, while the isolates of genogroup F evolved from a recent common ancestor shared with the members of the genogroup B. Our results reveal the wide diversity that exists among EV-A71 isolates and suggest that the number of circulating genogroups is probably underestimated, particularly in developing countries where EV-A71 epidemiology has been poorly studied. PMID:24598878

  8. Rapid Molecular Identification of Human Taeniid Cestodes by Pyrosequencing Approach

    Science.gov (United States)

    Thanchomnang, Tongjit; Tantrawatpan, Chairat; Intapan, Pewpan M.; Sanpool, Oranuch; Janwan, Penchom; Lulitanond, Viraphong; Tourtip, Somjintana; Yamasaki, Hiroshi; Maleewong, Wanchai

    2014-01-01

    Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse. PMID:24945530

  9. Rapid molecular identification of human taeniid cestodes by pyrosequencing approach.

    Directory of Open Access Journals (Sweden)

    Tongjit Thanchomnang

    Full Text Available Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1 gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse.

  10. [Enterovirus sequencing as a new approach to the laboratory diagnosis for clinical and epidemiological purposes].

    Science.gov (United States)

    Rainetová, P; Jiřincová, H; Musílek, M; Nováková, L; Vodičková, I; Štruncová, V; Švecová, M; Pazdiora, P; Piskunová, N; Trubač, P; Zajíc, T; Havlíčková, M

    2015-06-01

    Introducing enterovirus sequencing as an advanced approach to classify the viruses isolated according to the novel nomenclature and to characterize isolates in detail. Seventy-five specimens collected from 64 patients in two hospitals, Liberec Regional Hospital, and Plzeň University Hospital, were analyzed. The study patients' age ranged from four to 54 years, with a median of 15 years in males and 16 years in females. In most patients, the reasons for admission were intense headache, fever, vomiting, tiredness, meningeal symptoms, intestinal symptoms (in two patients), and skin symptoms (in one patient). The specimens collected were rectal and throat swabs, cerebrospinal fluid (CSF) and stool specimens. Molecular detection and typing were performed using the RT-PCR method. A segment of the 5´non-coding RNA was selected for typing. Specimens were amplified using single-step PCR with external primers and with the same primers extended to include M13 sequences (Generi-Biotech). The LASERGENE software (DIASTAR) was used in sequence editing, alignment, and quality check. The sequences obtained were checked against the central GenBank sequence database using the BLAST algorithm. The identification of the study isolates resulted in 61 ECHO viruses 30, three coxsackie viruses B1, one coxsackie virus B3, one coxsackie virus A9, one enterovirus 86, one enterovirus 71, Two ECHO viruses 13/coxsackie virus B5, one ECHO virus 7/30/coxsackie virus B4, one coxsackie virus B4/enterovirus B, one enterovirus 87/ECHO virus 30/enterovirus B, and one ECHO virus 3. All viruses isolated, except enterovirus 71 classified into group A, were of group B. The enteroviruses were identified unambigously, although the sequencing only targeted a short, conserved segment that showed considerable variability. The sequencing was an effective alternative to enterovirus identification by the neutralisation test and allowed for detailed characterization of the isolates. The predominance of ECHO 30 as

  11. Enterovirus 71 in Taiwan

    Directory of Open Access Journals (Sweden)

    Luan-Yin Chang

    2008-08-01

    Full Text Available The enterovirus 71 (EV71 outbreak in Taiwan in 1998 proved fatal in many children. A seroepidemiological study performed prior to the 1998 outbreak showed pre-epidemic (1997 EV71 seroprevalence rates to be about 60–70% in adults and children older than 6 years of age. A retrospective case review carried out from 1980–81 identified 16 cases of hand, foot and mouth disease associated with central nervous system involvement, two of whom died soon after hospitalization. There were 405 severe cases and 78 deaths reported in the 1998 epidemic, and dozens of fatal EV71 cases were still reported from 2000 to 2002. A stage-based management strategy was developed to reduce fatality, but most survivors of brainstem encephalitis with cardio pulmonary failure have neurologic sequelae and impaired cognition. Continuous clinical and laboratory surveillance of EV71 disease is required to enable earlier implementation of control and prevention measures. Development of EV71-specific antiviral therapy, a novel class of imidazolidinones, and development of a vaccine are ongoing.

  12. [Enteroviruses responsible for acute hemorrhagic conjunctivitis].

    Science.gov (United States)

    Lévêque, N; Huguet, P; Norder, H; Chomel, J-J

    2010-04-01

    Acute hemorrhagic conjunctivitis (AHC) is an epidemic form of highly contagious conjunctivitis, characterized by conjunctival hemorrhages. The first AHC outbreak was described in 1969 in Ghana, West Africa, and was called Apollo disease, from the Apollo landing on the moon. This outbreak was caused by Enterovirus 70 (EV70) together with a Coxsackievirus A24 (CVA24v) variant, which are the major etiological agents involved in AHC outbreaks worldwide. AHC is known to be directly transmitted by close person-to-person contact or indirectly through soiled ophthalmological materials or unsafe recreational water. Recently, a possible airborne virus spread was suggested which could explain the high transmission rate of the disease. In the absence of a specific antiviral therapy, a rapid diagnosis of the causative agent is required to distinguish AHC due to enteroviruses from other ocular infectious diseases, for there are active drugs, or to quickly implement proper public health measures to limit the extension of the outbreak. However, virus identification remains difficult and time-consuming. Moreover, virological diagnosis is difficult to implement in developing countries where AHC has recently become a major problem for public health. Copyright 2009 Elsevier Masson SAS. All rights reserved.

  13. Enterovirus and herpesviridae family as etiologic agents of lymphomonocytary meningitis, Southern Brazil

    Directory of Open Access Journals (Sweden)

    Luine Rosele Renaud Vidal

    2011-06-01

    Full Text Available Viral meningitis is a common infectious disease of the central nervous system (CNS that occurs worldwide. The aim of this study was to identify the etiologic agent of lymphomonocytary meningitis in Curitiba, PR, Brazil. During the period of July 2005 to December 2006, 460 cerebrospinal fluid (CSF samples with lymphomonocytary meningitis were analyzed by PCR methodologies. Fifty nine (12.8% samples were positive. Enteroviruses was present in 49 (83% samples and herpes virus family in 10 (17%, of these 6 (10% herpes simplex virus, 1 (2% Epstein Barr virus, 2 (3% human herpes virus type 6 and 1 (2% mixed infection of enterovirus and Epstein Barr virus. As conclusion enterovirus was the most frequent virus, with circulation during summer and was observed with higher frequency between 4 to 17 years of age. PCR methodology is an important method for rapid detection of RNA enterovirus and DNA herpesvirus in CSF.

  14. [Control of poliomyelitis and enterovirus infection in several areas of Russian Federation].

    Science.gov (United States)

    Romanenkova, N I; Bichurina, M A; Rosaeva, N R

    2011-01-01

    Control of poliovirus circulation by study of material from patients with acute flaccid paralysis and contact individuals, from children of risk groups; molecular characteristics of isolated polioviruses; monitoring of circulation of polioviruses and nonpoliomyelitis enteroviruses in population and the environment. Isolation and study of polioviruses and nonpoliomyelitis enteroviruses from various sources was performed in accordance with WHO recommendations. Prolonged persistence and circulation of vaccine related strains of polioviruses in children is demonstrated. Enterovirus serotypes that circulate in the population and the environment more frequently are determined. CONCLUSION. Long term control of poliomyelitis and acute flaccid paralysis in combination with additional control variants in children from risk groups and objects of the environment allowed to obtain valuable data on poliovirus and nonpoliomyelitis enteroviruses circulation for the Program of eradication of poliomyelitis.

  15. Rapid Isolation and Molecular Detection of Streptomycin-Producing Streptomycetes

    Directory of Open Access Journals (Sweden)

    M Motovali-bashi

    2006-07-01

    Full Text Available Introduction: Streptomyces species are mycelial, aerobic gram-positive bacteria that are isolated from soil and produce a diverse range of antibiotics. Streptomyces griseus produces the antibiotic, streptomycin and forms spores even in a liquid culture. The gene cluster for the production of Streptomycin antibiotic contains strR gene that encodes StrR, a pathway-specific regulator. Then, this pathway-specific regulator induces transcription of other streptomycin production genes in the gene cluster. The overall aim of this work was rapid isolation and molecular detection of streptomycin-producing Streptomycetes, especially S. griseus, from Iranian soils in order to manipulate them for increased production of streptomycin. Methods: This research used new initiative half-specific medium for isolation of Streptomycetes from natural environments, called FZmsn. The fifty colonies of Streptomyces strains grown on the surface of FZmsn medium isolated from environmental samples were defined on the basis of their morphological characteristics and light microscope studies. A set of primers was designed to detect strR by OLIGO software. Results: In colony-PCR reactions followed by gel electrophoresis, 6 colonies from Streptomyces strains colonies were detected as S. griseus colonies. Conclusion: These native Streptomyces strains will be used for genetic manipulation of S. griseus in order to increase production levels of streptomycin.

  16. A novel enterovirus species identified from severe diarrheal goats

    OpenAIRE

    Wang, Mingyue; He, Jia; Lu, Haibing; Liu, Yajing; Deng, Yingrui; Zhu, Lisai; Guo, Changming; Tu, Changchun; Wang, Xinping

    2017-01-01

    Backgrounds The Enterovirus genus of the family of Picornaviridae consists of 9 species of Enteroviruses and 3 species of Rhinoviruses based on the latest virus taxonomy. Those viruses contribute significantly to respiratory and digestive disorders in human and animals. Out of 9 Enterovirus species, Enterovirus E-G are closely related to diseases affecting on livestock industry. While enterovirus infection has been increasingly reported in cattle and swine, the enterovirus infections in small...

  17. 21 CFR 866.3225 - Enterovirus nucleic acid assay.

    Science.gov (United States)

    2010-04-01

    ... (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3225 Enterovirus... tests, aids in the clinical laboratory diagnosis of viral meningitis caused by enterovirus. (b...

  18. Rapid sampling of molecular motions with prior information constraints.

    Science.gov (United States)

    Raveh, Barak; Enosh, Angela; Schueler-Furman, Ora; Halperin, Dan

    2009-02-01

    Proteins are active, flexible machines that perform a range of different functions. Innovative experimental approaches may now provide limited partial information about conformational changes along motion pathways of proteins. There is therefore a need for computational approaches that can efficiently incorporate prior information into motion prediction schemes. In this paper, we present PathRover, a general setup designed for the integration of prior information into the motion planning algorithm of rapidly exploring random trees (RRT). Each suggested motion pathway comprises a sequence of low-energy clash-free conformations that satisfy an arbitrary number of prior information constraints. These constraints can be derived from experimental data or from expert intuition about the motion. The incorporation of prior information is very straightforward and significantly narrows down the vast search in the typically high-dimensional conformational space, leading to dramatic reduction in running time. To allow the use of state-of-the-art energy functions and conformational sampling, we have integrated this framework into Rosetta, an accurate protocol for diverse types of structural modeling. The suggested framework can serve as an effective complementary tool for molecular dynamics, Normal Mode Analysis, and other prevalent techniques for predicting motion in proteins. We applied our framework to three different model systems. We show that a limited set of experimentally motivated constraints may effectively bias the simulations toward diverse predicates in an outright fashion, from distance constraints to enforcement of loop closure. In particular, our analysis sheds light on mechanisms of protein domain swapping and on the role of different residues in the motion.

  19. Rapid sampling of molecular motions with prior information constraints.

    Directory of Open Access Journals (Sweden)

    Barak Raveh

    2009-02-01

    Full Text Available Proteins are active, flexible machines that perform a range of different functions. Innovative experimental approaches may now provide limited partial information about conformational changes along motion pathways of proteins. There is therefore a need for computational approaches that can efficiently incorporate prior information into motion prediction schemes. In this paper, we present PathRover, a general setup designed for the integration of prior information into the motion planning algorithm of rapidly exploring random trees (RRT. Each suggested motion pathway comprises a sequence of low-energy clash-free conformations that satisfy an arbitrary number of prior information constraints. These constraints can be derived from experimental data or from expert intuition about the motion. The incorporation of prior information is very straightforward and significantly narrows down the vast search in the typically high-dimensional conformational space, leading to dramatic reduction in running time. To allow the use of state-of-the-art energy functions and conformational sampling, we have integrated this framework into Rosetta, an accurate protocol for diverse types of structural modeling. The suggested framework can serve as an effective complementary tool for molecular dynamics, Normal Mode Analysis, and other prevalent techniques for predicting motion in proteins. We applied our framework to three different model systems. We show that a limited set of experimentally motivated constraints may effectively bias the simulations toward diverse predicates in an outright fashion, from distance constraints to enforcement of loop closure. In particular, our analysis sheds light on mechanisms of protein domain swapping and on the role of different residues in the motion.

  20. Crystal Structure of Human Enterovirus 71

    Energy Technology Data Exchange (ETDEWEB)

    Plevka, Pavel; Perera, Rushika; Cardosa, Jane; Kuhn, Richard J.; Rossmann, Michael G. (Purdue); (Sentinext)

    2013-04-08

    Enterovirus 71 is a picornavirus associated with fatal neurological illness in infants and young children. Here, we report the crystal structure of enterovirus 71 and show that, unlike in other enteroviruses, the 'pocket factor,' a small molecule that stabilizes the virus, is partly exposed on the floor of the 'canyon.' Thus, the structure of antiviral compounds may require a hydrophilic head group designed to interact with residues at the entrance of the pocket.

  1. Therapeutic Use of Native and Recombinant Enteroviruses

    Directory of Open Access Journals (Sweden)

    Jani Ylä-Pelto

    2016-02-01

    Full Text Available Research on human enteroviruses has resulted in the identification of more than 100 enterovirus types, which use more than 10 protein receptors and/or attachment factors required in cell binding and initiation of the replication cycle. Many of these “viral” receptors are overexpressed in cancer cells. Receptor binding and the ability to replicate in specific target cells define the tropism and pathogenesis of enterovirus types, because cellular infection often results in cytolytic response, i.e., disruption of the cells. Viral tropism and cytolytic properties thus make native enteroviruses prime candidates for oncolytic virotherapy. Copy DNA cloning and modification of enterovirus genomes have resulted in the generation of enterovirus vectors with properties that are useful in therapy or in vaccine trials where foreign antigenic epitopes are expressed from or on the surface of the vector virus. The small genome size and compact particle structure, however, set limits to enterovirus genome modifications. This review focuses on the therapeutic use of native and recombinant enteroviruses and the methods that have been applied to modify enterovirus genomes for therapy.

  2. A novel RT-multiplex PCR for enteroviruses, hepatitis A and E viruses and influenza A virus among infants and children with diarrhea in Vietnam.

    Science.gov (United States)

    Phan, T G; Nguyen, T A; Yan, H; Okitsu, S; Ushijima, H

    2005-06-01

    A novel reverse transcription-multiplex polymerase chain reaction (RT-multiplex PCR) assay that can detect enteroviruses, hepatitis A and E viruses and influenza A virus from various hosts (avian species, human, swine and horse) was developed. The identification of that group of viruses was performed with the mixture of four pairs of published specific primers (F1 and R1, P3 and P4, 2s and 2as, MMU42 and MMU43) for amplifying viral genomes and specifically generated four different amplicon sizes of 440, 267, 146 and 219 bp for enteroviruses, hepatitis A and E viruses and influenza A virus, respectively. A total of 276 fecal specimens (previously screened for rotavirus, adenovirus, norovirus, sapovirus and astrovirus-negative) from infants and children admitted into hospital with acute gastroenteritis in Ho Chi Minh city, Vietnam during October 2002 and September 2003 were collected and further tested for the presence of those viruses by RT-multiplex PCR. Enteroviruses were identified in 27 specimens and this represented 9.8%. No hepatitis A and E viruses and influenza A virus was found among these subjects. The sensitivity and specificity of RT-multiplex PCR were also assessed and demonstrated the strong validation against RT-monoplex PCR. Taken together, the findings clearly indicated that this novel RT-multiplex PCR is a simple and potential assay for rapid, sensitive, specific and cost-effective laboratory diagnosis to investigate molecular epidemiology of acute gastroenteritis caused by enteroviruses, hepatitis A and E viruses and influenza A virus. This report is the first, to our knowledge, detecting these kinds of viruses in diarrheal feces from infants and children in Vietnam.

  3. Enterovirus 74 Infection in Children

    Science.gov (United States)

    Peacey, Matthew; Hall, Richard J.; Wang, Jing; Todd, Angela K.; Yen, Seiha; Chan-Hyams, Jasmine; Rand, Christy J.; Stanton, Jo-Ann; Huang, Q. Sue

    2013-01-01

    Enterovirus 74 (EV74) is a rarely detected viral infection of children. In 2010, EV74 was identified in New Zealand in a 2 year old child with acute flaccid paralysis (AFP) through routine polio AFP surveillance. A further three cases of EV74 were identified in children within six months. These cases are the first report of EV74 in New Zealand. In this study we describe the near complete genome sequence of four EV74 isolates from New Zealand, which shows only limited sequence identity in the non-structural proteins when compared to the other two known EV74 sequences. As is typical of enteroviruses multiple recombination events were evident, particularly in the P2 region and P3 regions. This is the first complete EV74 genome sequenced from a patient with acute flaccid paralysis. PMID:24098514

  4. NONPOLIO ENTEROVIRUSES WHICH CAUSED THE RISE OF ENTEROVIRUS INFECTION ON SOME TERRITORIES OF RUSSIA IN 2016

    Directory of Open Access Journals (Sweden)

    N. I. Romanenkova

    2017-01-01

    Full Text Available Aim: Characteristics of the peculiarities and the etiological factor of enterovirus infection on some territories of Russia in 2016. Materials and methods: We investigated 2138 samples from the patients with enterovirus infection. The isolation and identification of enteroviruses were conducted by the virological method and by partial sequencing of the genome region VP1. Phylogenic trees were constructed according to the method of Bayesian Monte Carlo Markov Chain. Results: Epidemic peaks of enterovirus infection were fixed on some territories of Russia. In Saratov region the morbidity index of enterovirus infection in 2016 was twice as high as the median morbidity index over previous years. The morbidity level of enterovirus meningitis – 3, 21 for 100000 of the population (77% from all the cases of enterovirus infection was higher than on the other territories. In Kostroma region the morbidity index of enterovirus infection in 2016 was 11 times higher than the index of the previous year. On both territories the rise of morbidity depends on the active circulation of enterovirus ЕСНО30. Enteroviruses ECHO30 from Saratov region belonged to two phylogenic groups of genotype h. To one of them belonged viruses ECHO30 from Kostroma region. In Murmansk and Leningrad regions in 2016 most cases of enterovirus infection were represented by hand, foot and mouth disease (HFMD. The grouped foci of infection were registered in some preschool institutions. The etiological factor of this clinical form was Coxsackieviruses A6 belonging to different genetic variants. Conclusion: Epidemic peaks of enterovirus infection with the prevalence of different clinical forms of the disease were provoked by different etiological factors. On territories where enterovirus meningitis prevailed strains of enterovirus ECHO30 belonging to different variants of genotype h were detected. In patients with clinical picture of HFMD from territories where this form was leading

  5. Introducing molecular selectivity in rapid impedimetric sensing of phthalates

    KAUST Repository

    Zia, Asif I.; Mukhopadhyay, Subhas Chandra; Al-Bahadly, Ibrahim H.; Yu, Paklam; Gooneratne, Chinthaka Pasan; Kosel, Jü rgen

    2014-01-01

    This research article reports a real-time and non-invasive detection technique for phthalates in liquids by Electrochemical Impedance Spectroscopy (EIS), incorporating molecular imprinting technique to introduce selectivity for the phthalate

  6. A simple and rapid molecular method for Leptospira species identification

    NARCIS (Netherlands)

    Ahmed, Ahmed; Anthony, Richard M.; Hartskeerl, Rudy A.

    2010-01-01

    Serological and DNA-based classification systems only have little correlation. Currently serological and molecular methods for characterizing Leptospira are complex and costly restricting their world-wide distribution and use. Ligation mediated amplification combined with microarray analysis

  7. Using molecular techniques for rapid detection of Salmonella ...

    African Journals Online (AJOL)

    PRECIOUS

    2010-02-01

    Feb 1, 2010 ... A total of 152 samples of chicken and chicken products ... detection of Salmonella species in the collected field samples ... that 16 million new cases of typhoid fever occur each ... vative methods for the rapid identification of Salmonella ... saved for the PCR-Non Selective test (PCR-NS) and 1 ml of the.

  8. Using current molecular techniques for rapid differentiation of ...

    African Journals Online (AJOL)

    Typhoid fever is responsible for the deaths of many people annually. However, conventional and timeconsuming detection methods for Salmonella Typhi still dominate. By using a molecular based approach, it was possible to identify Salmonella Typhi by amplifying two specific genes (viaB and tyv) and by using RFLP ...

  9. Enterovirus D68 and other enterovirus serotypes identified in South African patients with severe acute respiratory illness, 2009-2011.

    Science.gov (United States)

    Hellferscee, Orienka; Treurnicht, Florette K; Tempia, Stefano; Variava, Ebrahim; Dawood, Halima; Kahn, Kathleen; Cohen, Adam L; Pretorius, Marthi; Cohen, Cheryl; Madhi, Shabir A; Venter, Marietjie

    2017-05-01

    Human enteroviruses (EV) have been associated with severe acute respiratory illness (SARI) in South Africa. We aimed to describe the molecular epidemiology of EV serotypes among patients hospitalized with SARI during 2009-2011. Study samples from patients were tested for the presence of enterovirus using a polymerase chain reaction assay. 8.2% (842/10 260) of SARI cases tested positive for enterovirus; 16% (7/45) were species EV-A, 44% (20/45) EV-B, 18% (8/45) EV-C and 22% (10/45) EV-D. Seventeen different EV serotypes were identified within EV-A to EV-D, of which EV-D68 (22%; 10/45) and Echovirus 3 (11%; 5/45) were the most prevalent. EV-D68 should be monitored in South Africa to assess the emergence of highly pathogenic strains. © 2017 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

  10. Introducing molecular selectivity in rapid impedimetric sensing of phthalates

    KAUST Repository

    Zia, Asif I.

    2014-05-01

    This research article reports a real-time and non-invasive detection technique for phthalates in liquids by Electrochemical Impedance Spectroscopy (EIS), incorporating molecular imprinting technique to introduce selectivity for the phthalate molecule in the detection system. A functional polymer with Bis (2-ethylhexyl) phthalate (DEHP) template was immobilized on the sensing surface of the inter-digital (ID) capacitive sensor with sputtered gold sensing electrodes fabricated over a native layer of silicon dioxide on a single crystal silicon substrate. Various concentrations (10 to 200 ppm) of DEHP in deionized MilliQ water were exposed to the sensor surface functionalized with molecular imprinted polymer (MIP) in order to capture the analyte molecule, hence introducing molecular selectivity to the testing system. Impedance spectra were obtained using EIS in order to determine sample conductance for evaluation of phthalate concentration in the solution. Electrochemical Spectrum Analyzer algorithm was used to deduce equivalent circuit and equivalent component parameters from the experimentally obtained impedance spectra employing Randle\\'s cell model curve fitting technique. Experimental results confirmed that the immobilization of the functional polymer on sensing surface introduces selectivity for phthalates in the sensing system. The results were validated by testing the samples using High Performance Liquid Chromatography (HPLC-DAD). © 2014 IEEE.

  11. Direct Identification of Enteroviruses in Cerebrospinal Fluid of Patients with Suspected Meningitis by Nested PCR Amplification

    Directory of Open Access Journals (Sweden)

    Alexandr Krasota

    2016-01-01

    Full Text Available Enteroviruses, the most common human viral pathogens worldwide, have been associated with serous meningitis, encephalitis, syndrome of acute flaccid paralysis, myocarditis and the onset of diabetes type 1. In the future, the rapid identification of the etiological agent would allow to adjust the therapy promptly and thereby improve the course of the disease and prognosis. We developed RT-nested PCR amplification of the genomic region coding viral structural protein VP1 for direct identification of enteroviruses in clinical specimens and compared it with the existing analogs. One-hundred-fifty-nine cerebrospinal fluids (CSF from patients with suspected meningitis were studied. The amplification of VP1 genomic region using the new method was achieved for 86 (54.1% patients compared with 75 (47.2%, 53 (33.3% and 31 (19.5% achieved with previously published methods. We identified 11 serotypes of the Enterovirus species B in 2012, including relatively rare echovirus 14 (E-14, E-15 and E-32, and eight serotypes of species B and 5 enteroviruses A71 (EV-A71 in 2013. The developed method can be useful for direct identification of enteroviruses in clinical material with the low virus loads such as CSF.

  12. Detection of hepatitis A virus and Enterovirus In the output water purification stations and charguia Jdeida

    International Nuclear Information System (INIS)

    Tallous Chaowki

    2010-01-01

    The objective of this study is looking for two enteric viruses (enteroviruses and hepatitis A) in the treated wastewater from two sewage treatment plants, Jdeida and charguia. The detection of these viruses is performed by RT-PCR. The detection limit of this technique is estimated at 10PFU/ml. The molecular study showed that HAV found in 10 pour cent of wastewater analyse samples.Enteroviruses were detected in 15 pour cent of tested samples. The presence of these viruses in treated water showed a lack of purification function of these stations on virology terms.

  13. MRI of Enterovirus 71 myelitis with monoplegia

    International Nuclear Information System (INIS)

    Shen, W.C.; Tsai, C.H.; Chiu, H.H.; Chow, K.C.

    2000-01-01

    We report two boys diagnosed as having herpangina and hand-foot-mouth disease complicated by monoplegia during the outbreak enterovirus infection in Taiwan in 1998. Enterovirus 71 was identified in the stool and throat swab; neither polio nor Coxsackie viruses was identified. MRI showed unilateral lesions in the anterior horns of the spinal cord at T11-12 and C2-5. Although the MRI findings and sites of these lesions were similar to those of poliovirus-associated poliomyelitis, the virological data indicated that these boys were infected with enterovirus type 71. (orig.)

  14. MRI of Enterovirus 71 myelitis with monoplegia

    Energy Technology Data Exchange (ETDEWEB)

    Shen, W.C. [Department of Radiology, and School of Medicine, China Medical College, China Medical College Hospital, Taiwan (Taiwan); Tsai, C.H. [Department of Paediatrics, and School of Medicine, China Medical College, China Medical College Hospital, Taiwan (Taiwan); Chiu, H.H. [Department of Paediatrics, China Medical College Hospital, No. 2 Yuh-Der Road, 407 Taichung, Taiwan (Taiwan); Chow, K.C. [Department of Medical Research, China Medical College Hospital, Taichung, Taiwan (Taiwan)

    2000-02-01

    We report two boys diagnosed as having herpangina and hand-foot-mouth disease complicated by monoplegia during the outbreak enterovirus infection in Taiwan in 1998. Enterovirus 71 was identified in the stool and throat swab; neither polio nor Coxsackie viruses was identified. MRI showed unilateral lesions in the anterior horns of the spinal cord at T11-12 and C2-5. Although the MRI findings and sites of these lesions were similar to those of poliovirus-associated poliomyelitis, the virological data indicated that these boys were infected with enterovirus type 71. (orig.)

  15. Rapid methods for the extraction and archiving of molecular grade fungal genomic DNA.

    Science.gov (United States)

    Borman, Andrew M; Palmer, Michael; Johnson, Elizabeth M

    2013-01-01

    The rapid and inexpensive extraction of fungal genomic DNA that is of sufficient quality for molecular approaches is central to the molecular identification, epidemiological analysis, taxonomy, and strain typing of pathogenic fungi. Although many commercially available and in-house extraction procedures do eliminate the majority of contaminants that commonly inhibit molecular approaches, the inherent difficulties in breaking fungal cell walls lead to protocols that are labor intensive and that routinely take several hours to complete. Here we describe several methods that we have developed in our laboratory that allow the extremely rapid and inexpensive preparation of fungal genomic DNA.

  16. Rapid molecular identification and characteristics of Lactobacillus strains.

    Science.gov (United States)

    Markiewicz, L H; Biedrzycka, E; Wasilewska, E; Bielecka, M

    2010-09-01

    Eleven type strains and 24 Lactobacillus isolates, preliminarily classified to the species due to phenotypic features, were investigated. Standard methods of identification with species-specific PCRs and typing with PFGE (with ApaI, NotI and SmaI restriction enzymes) allowed us to distinguish 16 unique strains belonging to 5 species (L. acidophilus, L. delbrueckii ssp. bulgaricus, L. plantarum, L. rhamnosus, L. salivarius). Alternative approach with 16S-23S rDNA ARDRA identification (with merely two restrictases, BsuRI and TaqI) and PCR-based typing (RAPD with two random- and rep-PCR with (GTG)(5) primers) showed to be more discriminative, i.e. 21 unique strains were classified in the same species as above. As a result, 7 out of 24 phenotypically species-assigned isolates were reclassified. The alternative procedure of rapid identification and typing of Lactobacillus isolates appeared to be equally effective and shortened from 1 week to 2-3 d (in comparison to the standard methods).

  17. Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

    Science.gov (United States)

    Cazanave, Charles; Greenwood-Quaintance, Kerryl E.; Hanssen, Arlen D.; Karau, Melissa J.; Schmidt, Suzannah M.; Gomez Urena, Eric O.; Mandrekar, Jayawant N.; Osmon, Douglas R.; Lough, Lindsay E.; Pritt, Bobbi S.; Steckelberg, James M.

    2013-01-01

    We previously showed that culture of samples obtained by prosthesis vortexing and sonication was more sensitive than tissue culture for prosthetic joint infection (PJI) diagnosis. Despite improved sensitivity, culture-negative cases remained; furthermore, culture has a long turnaround time. We designed a genus-/group-specific rapid PCR assay panel targeting PJI bacteria and applied it to samples obtained by vortexing and sonicating explanted hip and knee prostheses, and we compared the results to those with sonicate fluid and periprosthetic tissue culture obtained at revision or resection arthroplasty. We studied 434 subjects with knee (n = 272) or hip (n = 162) prostheses; using a standardized definition, 144 had PJI. Sensitivities of tissue culture, of sonicate fluid culture, and of PCR were 70.1, 72.9, and 77.1%, respectively. Specificities were 97.9, 98.3, and 97.9%, respectively. Sonicate fluid PCR was more sensitive than tissue culture (P = 0.04). PCR of prosthesis sonication samples is more sensitive than tissue culture for the microbiologic diagnosis of prosthetic hip and knee infection and provides same-day PJI diagnosis with definition of microbiology. The high assay specificity suggests that typical PJI bacteria may not cause aseptic implant failure. PMID:23658273

  18. The rapid evolution of molecular genetic diagnostics in neuromuscular diseases.

    Science.gov (United States)

    Volk, Alexander E; Kubisch, Christian

    2017-10-01

    The development of massively parallel sequencing (MPS) has revolutionized molecular genetic diagnostics in monogenic disorders. The present review gives a brief overview of different MPS-based approaches used in clinical diagnostics of neuromuscular disorders (NMDs) and highlights their advantages and limitations. MPS-based approaches like gene panel sequencing, (whole) exome sequencing, (whole) genome sequencing, and RNA sequencing have been used to identify the genetic cause in NMDs. Although gene panel sequencing has evolved as a standard test for heterogeneous diseases, it is still debated, mainly because of financial issues and unsolved problems of variant interpretation, whether genome sequencing (and to a lesser extent also exome sequencing) of single patients can already be regarded as routine diagnostics. However, it has been shown that the inclusion of parents and additional family members often leads to a substantial increase in the diagnostic yield in exome-wide/genome-wide MPS approaches. In addition, MPS-based RNA sequencing just enters the research and diagnostic scene. Next-generation sequencing increasingly enables the detection of the genetic cause in highly heterogeneous diseases like NMDs in an efficient and affordable way. Gene panel sequencing and family-based exome sequencing have been proven as potent and cost-efficient diagnostic tools. Although clinical validation and interpretation of genome sequencing is still challenging, diagnostic RNA sequencing represents a promising tool to bypass some hurdles of diagnostics using genomic DNA.

  19. Global emergence of enterovirus D68

    DEFF Research Database (Denmark)

    Holm-Hansen, Charlotte Carina; Midgley, Sofie Elisabeth; Fischer, Thea Kølsen

    2016-01-01

    be assessed in terms of capacity and ability to detect and report any upsurge of respiratory viruses such as enterovirus D68 in a timely manner, and focus should be paid to development of preventive measures against these emerging enteroviruses that have potential for severe disease.......Since its discovery in California in 1962, reports of enterovirus D68 have been infrequent. Before 2014, infections were confirmed in only 699 people worldwide. In August, 2014, two paediatric hospitals in the USA reported increases in the number of patients with severe respiratory illness......, with an over-representation in children with asthma. Shortly after, the authorities recognised a nationwide outbreak, which then spread to Canada, Europe, and Asia. In 2014, more than 2000 cases of enterovirus D68 were reported in 20 countries. Concurrently, clusters of children with acute flaccid paralysis...

  20. Enterovirus and Norovirus Monitoring under UCMR3

    Science.gov (United States)

    This presentation describes the Unregulated Contaminant Monitoring Rule round 3 (UCMR3) monitoring program for enterovirus and norovirus in groundwater. It provides the data on microbial indicators and virus occurrence during the monitoring period. Enteric virus occurrence was ab...

  1. Antiviral Combination Approach as a Perspective to Combat Enterovirus Infections.

    Science.gov (United States)

    Galabov, Angel S; Nikolova, Ivanka; Vassileva-Pencheva, Ralitsa; Stoyanova, Adelina

    2015-01-01

    Human enteroviruses distributed worldwide are causative agents of a broad spectrum of diseases with extremely high morbidity, including a series of severe illnesses of the central nervous system, heart, endocrine pancreas, skeleton muscles, etc., as well as the common cold contributing to the development of chronic respiratory diseases, including the chronic obstructive pulmonary disease. The above mentioned diseases along with the significantly high morbidity and mortality in children, as well as in the high-risk populations (immunodeficiencies, neonates) definitely formulate the chemotherapy as the main tool for the control of enterovirus infections. At present, clinically effective antivirals for use in the treatment of enteroviral infection do not exist, in spite of the large amount of work carried out in this field. The main reason for this is the development of drug resistance. We studied the process of development of resistance to the strongest inhibitors of enteroviruses, WIN compounds (VP1 protein hydrophobic pocket blockers), especially in the models in vivo, Coxsackievirus B (CV-B) infections in mice. We introduced the tracing of a panel of phenotypic markers (MIC50 value, plaque shape and size, stability at 50℃, pathogenicity in mice) for characterization of the drug-mutants (resistant and dependent) as a very important stage in the study of enterovirus inhibitors. Moreover, as a result of VP1 RNA sequence analysis performed on the model of disoxaril mutants of CVB1, we determined the molecular basis of the drug-resistance. The monotherapy courses were the only approach used till now. For the first time in the research for anti-enterovirus antivirals our team introduced the testing of combination effect of the selective inhibitors of enterovirus replication with different mode of action. This study resulted in the selection of a number of very effective in vitro double combinations with synergistic effect and a broad spectrum of sensitive

  2. Understanding Enterovirus 71 Neuropathogenesis and Its Impact on Other Neurotropic Enteroviruses.

    Science.gov (United States)

    Ong, Kien Chai; Wong, Kum Thong

    2015-09-01

    Enterovirus A71 (EV-A71) belongs to the species group A in the Enterovirus genus within the Picornaviridae family. EV-A71 usually causes self-limiting hand, foot and mouth disease or herpangina but rarely causes severe neurological complications such as acute flaccid paralysis and encephalomyelitis. The pathology and neuropathogenesis of these neurological syndromes is beginning to be understood. EV-A71 neurotropism for motor neurons in the spinal cord and brainstem, and other neurons, is mainly responsible for central nervous system damage. This review on the general aspects, recent developments and advances of EV-A71 infection will focus on neuropathogenesis and its implications on other neurotropic enteroviruses, such as poliovirus and the newly emergent Enterovirus D68. With the imminent eradication of poliovirus, EV-A71 is likely to replace it as an important neurotropic enterovirus of worldwide importance. © 2015 International Society of Neuropathology.

  3. Enterovirus related metabolic myopathy: a postviral fatigue syndrome

    OpenAIRE

    Lane, R; Soteriou, B; Zhang, H; Archard, L

    2003-01-01

    Objective: To detect and characterise enterovirus RNA in skeletal muscle from patients with chronic fatigue syndrome (CFS) and to compare efficiency of muscle energy metabolism in enterovirus positive and negative CFS patients.

  4. Clinical librarian support for rapid review of clinical utility of cancer molecular biomarkers.

    Science.gov (United States)

    Geng, Yimin; Fowler, Clara S; Fulton, Stephanie

    2015-01-01

    The clinical librarian used a restricted literature searching and quality-filtering approach to provide relevant clinical evidence for the use of cancer molecular biomarkers by institutional policy makers and clinicians in the rapid review process. The librarian-provided evidence was compared with the cited references in the institutional molecular biomarker algorithm. The overall incorporation rate of the librarian-provided references into the algorithm was above 80%. This study suggests the usefulness of clinical librarian expertise for clinical practice. The searching and filtering methods for high-level evidence can be adopted by information professionals who are involved in the rapid literature review.

  5. Implementation of Rapid Molecular Infectious Disease Diagnostics: the Role of Diagnostic and Antimicrobial Stewardship.

    Science.gov (United States)

    Messacar, Kevin; Parker, Sarah K; Todd, James K; Dominguez, Samuel R

    2017-03-01

    New rapid molecular diagnostic technologies for infectious diseases enable expedited accurate microbiological diagnoses. However, diagnostic stewardship and antimicrobial stewardship are necessary to ensure that these technologies conserve, rather than consume, additional health care resources and optimally affect patient care. Diagnostic stewardship is needed to implement appropriate tests for the clinical setting and to direct testing toward appropriate patients. Antimicrobial stewardship is needed to ensure prompt appropriate clinical action to translate faster diagnostic test results in the laboratory into improved outcomes at the bedside. This minireview outlines the roles of diagnostic stewardship and antimicrobial stewardship in the implementation of rapid molecular infectious disease diagnostics. Copyright © 2017 American Society for Microbiology.

  6. Therapeutic and prevention strategies against human enterovirus 71 infection

    Science.gov (United States)

    Kok, Chee Choy

    2015-01-01

    Human enterovirus 71 (HEV71) is the cause of hand, foot and mouth disease and associated neurological complications in children under five years of age. There has been an increase in HEV71 epidemic activity throughout the Asia-Pacific region in the past decade, and it is predicted to replace poliovirus as the extant neurotropic enterovirus of highest global public health significance. To date there is no effective antiviral treatment and no vaccine is available to prevent HEV71 infection. The increase in prevalence, virulence and geographic spread of HEV71 infection over the past decade provides increasing incentive for the development of new therapeutic and prevention strategies against this emerging viral infection. The current review focuses on the potential, advantages and disadvantages of these strategies. Since the explosion of outbreaks leading to large epidemics in China, research in natural therapeutic products has identified several groups of compounds with anti-HEV71 activities. Concurrently, the search for effective synthetic antivirals has produced promising results. Other therapeutic strategies including immunotherapy and the use of oligonucleotides have also been explored. A sound prevention strategy is crucial in order to control the spread of HEV71. To this end the ultimate goal is the rapid development, regulatory approval and widespread implementation of a safe and effective vaccine. The various forms of HEV71 vaccine designs are highlighted in this review. Given the rapid progress of research in this area, eradication of the virus is likely to be achieved. PMID:25964873

  7. Rapid molecular diagnostics for multi-drug resistant tuberculosis in India.

    Science.gov (United States)

    Ramachandran, Rajeswari; Muniyandi, M

    2018-03-01

    Rapid molecular diagnostic methods help in the detection of TB and Rifampicin resistance. These methods detect TB early, are accurate and play a crucial role in reducing the burden of drug resistant tuberculosis. Areas covered: This review analyses rapid molecular diagnostic tools used in the diagnosis of MDR-TB in India, such as the Line Probe Assay and GeneXpert. We have discussed the burden of MDR-TB and the impact of recent diagnostic tools on case detection and treatment outcomes. This review also discusses the costs involved in establishing these new techniques in India. Expert commentary: Molecular methods have considerable advantages for the programmatic management of drug resistant TB. These include speed, standardization of testing, potentially high throughput and reduced laboratory biosafety requirements. There is a desperate need for India to adopt modern, rapid, molecular tools with point-of-care tests being currently evaluated. New molecular diagnostic tests appear to be cost effective and also help in detecting missing cases. There is enough evidence to support the scaling up of these new tools in India.

  8. Enterovirus RNA in longitudinal blood samples and risk of islet autoimmunity in children with a high genetic risk of type 1 diabetes: the MIDIA study.

    Science.gov (United States)

    Cinek, Ondrej; Stene, Lars C; Kramna, Lenka; Tapia, German; Oikarinen, Sami; Witsø, Elisabet; Rasmussen, Trond; Torjesen, Peter A; Hyöty, Heikki; Rønningen, Kjersti S

    2014-10-01

    Only a few longitudinal molecular studies of enterovirus and islet autoimmunity have been reported, and positive results seem to be limited to Finland. We aimed to investigate an association between enterovirus RNA in blood and islet autoimmunity in the MIDIA study from Norway, a country which largely shares environmental and economic features with Finland. We analysed serial blood samples collected at ages 3, 6, and 9 months and then annually from 45 children who developed confirmed positivity for at least two autoantibodies (against insulin, GAD65 and IA-2) and 92 matched controls, all from a cohort of children with a single high-risk HLA-DQ-DR genotype. Enterovirus was tested in RNA extracted from frozen blood cell pellets, using real-time RT-PCR with stringent performance control. Out of 807 blood samples, 72 (8.9%) were positive for enterovirus. There was no association between enterovirus RNA and islet autoimmunity in samples obtained strictly before (7.6% cases, 10.0% controls, OR 0.75 [95% CI 0.36, 1.57]), or strictly after the first detection of islet autoantibodies (10.5% case, 5.8% controls, OR 2.00 [95% CI 0.64, 6.27]). However, there was a tendency towards a higher frequency of enterovirus detection in the first islet autoantibody-positive sample (15.8%) compared with the corresponding time point in matched controls (3.2%, OR 8.7 [95% CI 0.97, 77]). Neither of these results was changed by adjusting for potential confounders, restricting to various time intervals or employing various definitions of enterovirus positivity. Positivity for enterovirus RNA in blood did not predict the later induction of islet autoantibodies, but enterovirus tended to be detected more often at the islet autoantibody seroconversion stage.

  9. Enterovirus IgM detection

    International Nuclear Information System (INIS)

    Frisk, G.; Nilsson, E.; Diderholm, H.

    1989-01-01

    A predominantly type-specific L-capture radio-immunoassay (RIA) of IgM antibodies to Coxsackie B1-B5 (CB1-CB5) viruses was previously described. The present study is concerned with the specificity of this assay, using as antigen different strains of one serotype (CB5) and procapsids of two serotypes (CB3 and CB5). Eight strains of CB5 virions were tested against acute and/or convalescent sera from 10 patients from whom CB5 had been isolated. It is concluded that the choice of strain is important if a high frequency of homotypic titres with no or only a few heterotypic reactions is to be obtained. When procapsids were used as antigen, both homotypic and heterotypic titres were seen to a large extent. All patients with homotypic IgM against CB3 or CB5 virions showed IgM against the CB3 or the CB5 procapsids, respectively. It is concluded that the use of two or more procapsids in combination is of value for the diagnosis of a recent or current enterovirus infection (author). 18 refs.; 2 figs.; 4 tabs

  10. Highly sensitive and rapid bacteria detection using molecular beacon-Au nanoparticles hybrid nanoprobes.

    Science.gov (United States)

    Cao, Jing; Feng, Chao; Liu, Yan; Wang, Shouyu; Liu, Fei

    2014-07-15

    Since many diseases are caused by pathogenic bacterial infections, accurate and rapid detection of pathogenic bacteria is in urgent need to timely apply appropriate treatments and to reduce economic costs. To end this, we designed molecular beacon-Au nanoparticle hybrid nanoprobes to improve the bacterial detection efficiency and sensitivity. Here, we show that the designed molecular beacon modified Au nanoparticles could specifically recognize synthetic DNAs targets and can readily detect targets in clinical samples. Moreover, the hybrid nanoprobes can recognize Escherichia coli within an hour at a concentration of 10(2) cfu/ml, which is 1000-folds sensitive than using molecular beacon directly. Our results show that the molecular beacon-Au nanoparticle hybrid nanoprobes have great potential in medical and biological applications. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Inactivation of enteroviruses in sewage with ozone

    Energy Technology Data Exchange (ETDEWEB)

    Ivanova, O.E.; Bogdanov, M.V.; Kazantseva, V.A.; Gabrilevskaia, L.N.; Kodkind, G.K.H.

    The study of ozone inactivation of enteroviruses in sewage showed the presence in sewage of suspensions of organic origin and bacterial flora to influence the rate of inactivation. The inactivation rate of poliomyelitis virus in sewage free from organic suspension and bacterial flora was significantly higher than that in sewage containing such suspension and bacterial flora. The inactivation rate of enteroviruses was found not to depend upon the protein and salt composition and pH of sewage or strain appurtenance of viruses. The inactivation rate of enteroviruses directly depended upon the dose of ozone and time of contact with it. Differences in the resistance of different types of poliomyelitis virus, ECHO and Coxsackie viruses to the effect of ozone are likely exist. These differences are manifested within the range of relatively small doses of ozone. E. coli is more resistant to ozone than entero-viruses. The results of laboratory studies were used to choose the regimen of sanitation of urban sewage to be used in technological cycles of industrial enterprises.

  12. Genetic Diversity of Enterovirus A71, India

    Science.gov (United States)

    Saxena, Vinay K.; Sane, Sudhir; Nadkarni, Sushma S.; Sharma, Deepa K.

    2015-01-01

    We have identified circulation of 3 genogroups of enterovirus (EV) A71 in India. A new genogroup (proposed designation G) was discovered during this study. We isolated genogroups D and G in wide geographic areas but detected subgenogroup C1 only in 1 focus in western India. A systematic nationwide search for EV-A71 is warranted. PMID:25531549

  13. Quantum state specific reactant preparation in a molecular beam by rapid adiabatic passage

    Science.gov (United States)

    Chadwick, Helen; Hundt, P. Morten; van Reijzen, Maarten E.; Yoder, Bruce L.; Beck, Rainer D.

    2014-01-01

    Highly efficient preparation of molecules in a specific rovibrationally excited state for gas/surface reactivity measurements is achieved in a molecular beam using tunable infrared (IR) radiation from a single mode continuous wave optical parametric oscillator (cw-OPO). We demonstrate that with appropriate focusing of the IR radiation, molecules in the molecular beam crossing the fixed frequency IR field experience a Doppler tuning that can be adjusted to achieve complete population inversion of a two-level system by rapid adiabatic passage (RAP). A room temperature pyroelectric detector is used to monitor the excited fraction in the molecular beam and the population inversion is detected and quantified using IR bleaching by a second IR-OPO. The second OPO is also used for complete population transfer to an overtone or combination vibration via double resonance excitation using two spatially separated RAP processes.

  14. Inactivation of a bovine enterovirus and a bovine parvovirus in cattle manure by anaerobic digestion, heat treatment, gamma irradiation, ensilage and composting

    Energy Technology Data Exchange (ETDEWEB)

    Monteith, H.D.; Shannon, E.E.; Derbyshire, J.B.

    1986-08-01

    A bovine enterovirus and a bovine parvovirus seeded into liquid cattle manure were rapidly inactivated by anaerobic digestion under thermophilic conditions (55/sup 0/C), but the same viruses survived for up to 13 and 8 days respectively under mesophilic conditions (35/sup 0/C). The enterovirus was inactivated in digested liquid manure heated to 70/sup 0/C for 30 min, but the parvovirus was not inactivated by this treatment. The enterovirus, seeded into single cell protein (the solids recovered by centrifugation of digested liquid manure), was inactivated by a gamma irradiation dose of 1.0 Mrad, but the parvovirus survived this dose. When single cell protein seeded with bovine enterovirus or bovine parvovirus was ensiled with cracked corn, the enterovirus was inactivated after a period of 30 days, while the parvovirus survived for 30 days in one of two experiments. Neither the enterovirus nor the parvovirus survived composting for 28 days in a thermophilic aerobic environment when seeded into the solid fraction of cattle manure. It was concluded that, of the procedures tested, only anaerobic digestion under thermophilic conditions appeared to be reliable method of viral inactivation to ensure the safety of single cell protein for refeeding to livestock. Composting appeared to be a suitable method for the disinfection of manure for use as a soil conditioner.

  15. Neurotropic Enterovirus Infections in the Central Nervous System

    Directory of Open Access Journals (Sweden)

    Hsing-I Huang

    2015-11-01

    Full Text Available Enteroviruses are a group of positive-sense single stranded viruses that belong to the Picornaviridae family. Most enteroviruses infect humans from the gastrointestinal tract and cause mild symptoms. However, several enteroviruses can invade the central nervous system (CNS and result in various neurological symptoms that are correlated to mortality associated with enteroviral infections. In recent years, large outbreaks of enteroviruses occurred worldwide. Therefore, these neurotropic enteroviruses have been deemed as re-emerging pathogens. Although these viruses are becoming large threats to public health, our understanding of these viruses, especially for non-polio enteroviruses, is limited. In this article, we review recent advances in the trafficking of these pathogens from the peripheral to the central nervous system, compare their cell tropism, and discuss the effects of viral infections in their host neuronal cells.

  16. Neurotropic Enterovirus Infections in the Central Nervous System.

    Science.gov (United States)

    Huang, Hsing-I; Shih, Shin-Ru

    2015-11-24

    Enteroviruses are a group of positive-sense single stranded viruses that belong to the Picornaviridae family. Most enteroviruses infect humans from the gastrointestinal tract and cause mild symptoms. However, several enteroviruses can invade the central nervous system (CNS) and result in various neurological symptoms that are correlated to mortality associated with enteroviral infections. In recent years, large outbreaks of enteroviruses occurred worldwide. Therefore, these neurotropic enteroviruses have been deemed as re-emerging pathogens. Although these viruses are becoming large threats to public health, our understanding of these viruses, especially for non-polio enteroviruses, is limited. In this article, we review recent advances in the trafficking of these pathogens from the peripheral to the central nervous system, compare their cell tropism, and discuss the effects of viral infections in their host neuronal cells.

  17. [Enterovirus non-poliomyelitis infections in Krasnodar region].

    Science.gov (United States)

    Zhukova, L I; Rafeenko, G K; Larin, F I; Shcherbina, L I; Shut', I N; Davydova, M A; Vaniukov, A A

    2014-01-01

    Evaluation of epidemic situation by non-poliomyelitis enterovirus infections in Krasnodar region in multi-year dynamics and characterization of clinical course of enterovirus serous meningitis in hospitalize patients. Retrospective analysis of non-poliomyelitis enterovirus infection epidemi process manifestations during 2002-2012 in Krasnodar region territory based on data of Center of Hygien and Epidemiology in Krasnodar Region. Clinical-epidemiologic characteristics of enterovirus infections in Krasnodar region are presented. Landscape ofenteroviruses isolated from the environment of some territories of the region and from the biological material of patients with various diseases is demonstrated. Clinical features ofenterovirus meningitis course are characterized. Enterovirus transmission b contact route was established to be the most frequent. A lack of pathognomonic symptoms and awareness o physicians of various specialties regarding diagnostics of this infection are the clinical problems of non-po liomyelitis enterovirus diseases.

  18. First isolation of enterovirus 71 (EV-71 from Northern Brazil

    Directory of Open Access Journals (Sweden)

    Lamarão Leticia Martins

    2003-01-01

    Full Text Available Enterovirus 71 (EV-71 has been associated to cases of neurological disease in many countries including Brazil. This virus has now been reported from three of the five Brazilian regions. Our study relates the findings concerning to the first isolate of this virus in Northern region of Brazil. A 15-month old female patient, from the rural zone of the municipality of Santana do Araguaia in southern Pará state was admitted at the hospital with acute, flaccid, asymmetric and ascending motor deficiency, located in the right lower limb. Stools samples from this child were inoculated in RD cells and was isolated an EV-71. We plan to sequence our strain and compare it to other isolates in Brazil. Differences at the molecular level can explain why EV-71 strains circulating in other continents, such as Asia, appear to be more virulent.

  19. Enterovirus strain and type-specific differences in growth kinetics and virus-induced cell destruction in human pancreatic duct epithelial HPDE cells.

    Science.gov (United States)

    Smura, Teemu; Natri, Olli; Ylipaasto, Petri; Hellman, Marika; Al-Hello, Haider; Piemonti, Lorenzo; Roivainen, Merja

    2015-12-02

    diabetes. In addition, the capability for rapid adaptation to different cell types suggests that, on occasion, enterovirus strains with different pathogenetic properties may arise from less pathogenic ancestors. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. Rapid Molecular detection of citrus brown spot disease using ACT gene in Alternaria alternata

    Directory of Open Access Journals (Sweden)

    Hamid Moghimi

    2017-06-01

    Full Text Available Introduction:Using rapid detection methods is important for detection of plant pathogens and also prevention through spreading pests in agriculture. Citrus brown spot disease caused by pathogenic isolates of Alternaria alternata is a common disease in Iran. Materials and methods: In this study, for the first time a PCR based molecular method was used for rapid diagnosis of brown spot disease. Nine isolates of A. Alternata were isolated in PDA medium from different citrus gardens. The plant pathogenic activity was examined in tangerine leaves for isolates. Results showed that these isolates are the agents of brown spot disease. PCR amplification of specific ACT-toxin gene was performed for DNA extracted from A. alternata isolates, with 11 different fungal isolates as negative controls and 5 DNA samples extracted from soil. Results: Results showed that A. alternata, the causal agent of brown spot disease, can be carefully distinguished from other pathogenic agents by performing PCR amplification with specific primers for ACT toxin gene. Also, the results from Nested-PCR method confirmed the primary reaction and the specificity of A. alternata for brown spot disease. PCR results to control samples of the other standard fungal isolates, showed no amplification band. In addition, PCR with the DNA extracted from contaminated soils confirmed the presence of ACT toxin gene. Discussion and conclusion: Molecular procedure presented here can be used in rapid identification and prevention of brown spot infection in citrus gardens all over the country.

  1. Quantum Dot-Fullerene Based Molecular Beacon Nanosensors for Rapid, Highly Sensitive Nucleic Acid Detection.

    Science.gov (United States)

    Liu, Ye; Kannegulla, Akash; Wu, Bo; Cheng, Li-Jing

    2018-05-15

    Spherical fullerene (C 60 ) can quench the fluorescence of a quantum dot (QD) through energy transfer and charge transfer processes, with the quenching efficiency regulated by the number of proximate C 60 on each QD. With the quenching property and its small size compared with other nanoparticle-based quenchers, it is advantageous to group a QD reporter and multiple C 60 -labeled oligonucleotide probes to construct a molecular beacon (MB) probe for sensitive, robust nucleic acid detection. We demonstrated a rapid, high-sensitivity DNA detection method using the nanosensors composed of QD-C 60 based MBs carried by magnetic nanoparticles (MNPs). The assay was accelerated by first dispersing the nanosensors in analytes for highly efficient DNA capture resulting from short-distance 3-dimensional diffusion of targets to the sensor surface and then concentrating the nanosensors to a substrate by magnetic force to amplify the fluorescence signal for target quantification. The enhanced mass transport enabled a rapid detection (< 10 min) with a small sample volume (1-10 µl). The high signal-to-noise ratio produced by the QD-C 60 pairs and magnetic concentration yielded a detection limit of 100 fM (~106 target DNA copies for a 10 µl analyte). The rapid, sensitive, label-free detection method will benefit the applications in point-of-care molecular diagnostic technologies.

  2. Immune responses to baculovirus-displayed enterovirus 71 VP1 antigen.

    Science.gov (United States)

    Kiener, Tanja K; Premanand, Balraj; Kwang, Jimmy

    2013-04-01

    The increased distribution and neurovirulence of enterovirus 71 is an important health threat for young children in Asia Pacific. Vaccine design has concentrated on inactivated virus with the most advanced undergoing Phase III clinical trials. By using a subunit vaccine approach, production costs could be reduced by lowering the need for biocontainment. In addition, novel mutations could be rapidly incorporated to reflect the emergence of new enterovirus 71 subgenogroups. To circumvent the problems associated with conventional subunit vaccines, the antigen can be displayed on a viral vector that conveys stability and facilitates purification. Additional advantages of viral-vectored subunit vaccines are their ability to stimulate the innate immune system by transducing cells and the possibility of oral or nasal delivery, which dispenses with the need for syringes and medical personnel. Baculovirus-displayed VP1 combines all these benefits with protection that is as efficient as inactivated virus.

  3. Recombinase polymerase amplification: Emergence as a critical molecular technology for rapid, low-resource diagnostics.

    Science.gov (United States)

    James, Ameh; Macdonald, Joanne

    2015-01-01

    Isothermal molecular diagnostics are bridging the technology gap between traditional diagnostics and polymerase chain reaction-based methods. These new techniques enable timely and accurate testing, especially in settings where there is a lack of infrastructure to support polymerase chain reaction facilities. Despite this, there is a significant lack of uptake of these technologies in developing countries where they are highly needed. Among these novel isothermal technologies, recombinase polymerase amplification (RPA) holds particular potential for use in developing countries. This rapid nucleic acid amplification approach is fast, highly sensitive and specific, and amenable to countries with a high burden of infectious diseases. Implementation of RPA technology in developing countries is critically required to assess limitations and potentials of the diagnosis of infectious disease, and may help identify impediments that prevent adoption of new molecular technologies in low resource- and low skill settings. This review focuses on approaching diagnosis of infectious disease with RPA.

  4. Enterovirus D68 in Viet Nam (2009-2015).

    Science.gov (United States)

    Ny, Nguyen Thi Han; Anh, Nguyen To; Hang, Vu Thi Ty; Nguyet, Lam Anh; Thanh, Tran Tan; Ha, Do Quang; Minh, Ngo Ngoc Quang; Ha, Do Lien Anh; McBride, Angela; Tuan, Ha Manh; Baker, Stephen; Tam, Pham Thi Thanh; Phuc, Tran My; Huong, Dang Thao; Loi, Tran Quoc; Vu, Nguyen Tran Anh; Hung, Nguyen Van; Minh, Tran Thi Thuy; Xang, Nguyen Van; Dong, Nguyen; Nghia, Ho Dang Trung; Chau, Nguyen Van Vinh; Thwaites, Guy; van Doorn, H Rogier; Anscombe, Catherine; Le Van, Tan

    2017-01-01

    Since 1962, enterovirus D68 (EV-D68) has been implicated in multiple outbreaks and sporadic cases of respiratory infection worldwide, but especially in the USA and Europe with an increasing frequency between 2010 and 2014. We describe the detection, associated clinical features and molecular characterization of EV-D68 in central and southern Viet Nam between 2009 and 2015. Enterovirus/rhinovirus PCR positive respiratory or CSF samples taken from children and adults with respiratory/central nervous system infections in Viet Nam were tested by an EV-D68 specific PCR. The included samples were derived from 3 different observational studies conducted at referral hospitals across central and southern Viet Nam between 2009 and 2015. Whole-genome sequencing was carried out using a MiSeq based approach. Phylogenetic reconstruction and estimation of evolutionary rate and recombination were carried out in BEAST and Recombination Detection Program, respectively. EV-D68 was detected in 21/625 (3.4%) enterovirus/rhinovirus PCR positive respiratory samples but in none of the 15 CSF. All the EV-D68 patients were young children (age range: 11.8 - 24.5 months) and had moderate respiratory infections. Phylogenetic analysis suggested that the Vietnamese sequences clustered with those from Asian countries, of which 9 fell in the B1 clade, and the remaining sequence was identified within the A2 clade. One intra sub-clade recombination event was detected, representing the second reported recombination within EV-D68. The evolutionary rate of EV-D68 was estimated to be 5.12E -3 substitutions/site/year. Phylogenetic analysis indicated that the virus was imported into Viet Nam in 2008. We have demonstrated for the first time EV-D68 has been circulating at low levels in Viet Nam since 2008, associated with moderate acute respiratory infection in children. EV-D68 in Viet Nam is most closely related to Asian viruses, and clusters separately from recent US and European viruses that were

  5. Enterovirus 71, One Virus and Many Stories

    Directory of Open Access Journals (Sweden)

    Shih-Min Wang

    2008-08-01

    Full Text Available Enterovirus 71 (EV71 has emerged as a significant cause of brainstem encephalitis and acute flaccid paralysis in Taiwan. It may be complicated by autonomic nervous system dysregulation and pulmonary edema (PE. Cytokines in the central nervous system and systemic inflammatory responses play important roles in the pathogenesis of EV71-associated PE. Pathogenesis-based management with intravenous immunoglobulin and milrinone has been associated with reduced mortality in children with severe EV71 infections.

  6. Challenges to Licensure of Enterovirus 71 Vaccines

    Science.gov (United States)

    Wang, Jen-Ren; Chi, Chia-Yu; Chong, Pele; Su, Ih-Jen

    2012-01-01

    Human enteroviruses usually cause self-limited infections except polioviruses and enterovirus 71 (EV71), which frequently involve neurological complications. EV71 vaccines are being evaluated in humans. However, several challenges to licensure of EV71 vaccines need to be addressed. Firstly, EV71 and coxsackievirus A (CA) are frequently found to co-circulate and cause hand-foot-mouth disease (HFMD). A polyvalent vaccine that can provide protection against EV71 and prevalent CA are desirable. Secondly, infants are the target population of HFMD vaccines and it would need multi-national efficacy trials to prove clinical protection and speed up the licensure and usage of HFMD vaccines in children. An international network for enterovirus surveillance and clinical trials is urgently needed. Thirdly, EV71 is found to evolve quickly in the past 15 years. Prospective cohort studies are warranted to clarify clinical and epidemiological significances of the antigenic and genetic variations between different EV71 genogroups, which is critical for vaccine design. PMID:22953003

  7. Challenges to licensure of enterovirus 71 vaccines.

    Directory of Open Access Journals (Sweden)

    Min-Shi Lee

    Full Text Available Human enteroviruses usually cause self-limited infections except polioviruses and enterovirus 71 (EV71, which frequently involve neurological complications. EV71 vaccines are being evaluated in humans. However, several challenges to licensure of EV71 vaccines need to be addressed. Firstly, EV71 and coxsackievirus A (CA are frequently found to co-circulate and cause hand-foot-mouth disease (HFMD. A polyvalent vaccine that can provide protection against EV71 and prevalent CA are desirable. Secondly, infants are the target population of HFMD vaccines and it would need multi-national efficacy trials to prove clinical protection and speed up the licensure and usage of HFMD vaccines in children. An international network for enterovirus surveillance and clinical trials is urgently needed. Thirdly, EV71 is found to evolve quickly in the past 15 years. Prospective cohort studies are warranted to clarify clinical and epidemiological significances of the antigenic and genetic variations between different EV71 genogroups, which is critical for vaccine design.

  8. Enterovirus and parechovirus infection in children : a brief overview

    NARCIS (Netherlands)

    de Crom, S. C. M.; Rossen, J. W. A.; van Furth, A. M.; Obihara, C. C.

    Enterovirus and parechovirus are a frequent cause of infection in children. This review is an overview of what is known from enterovirus and parechovirus infection in children and contains information about the epidemiology, pathogenesis, clinical presentation, diagnosis, treatment, and prognosis of

  9. Detection of Enterovirus D68 in Canadian Laboratories

    Science.gov (United States)

    Hatchette, Todd F.; Drews, Steven J.; Grudeski, Elsie; Booth, Tim; Martineau, Christine; Dust, Kerry; Garceau, Richard; Gubbay, Jonathan; Karnauchow, Tim; Krajden, Mel; Levett, Paul N.; Mazzulli, Tony; McDonald, Ryan R.; McNabb, Alan; Mubareka, Samira; Needle, Robert; Petrich, Astrid; Richardson, Susan; Rutherford, Candy; Smieja, Marek; Tellier, Raymond; Tipples, Graham

    2015-01-01

    The recent emergence of a severe respiratory disease caused by enterovirus D68 prompted investigation into whether Canadian hospital and provincial laboratories can detect this virus using commercial and laboratory-developed assays. This study demonstrated analytical sensitivity differences between commercial and laboratory-developed assays for the detection of enterovirus D68. PMID:25740765

  10. Epidemiology of childhood enterovirus infections in Hangzhou, China.

    Science.gov (United States)

    Li, Wei; Zhang, Xiao; Chen, Xi; Cheng, Yu-Ping; Wu, Yi-Dong; Shu, Qiang; Chen, Xue-Jun; Shang, Shi-Qiang

    2015-04-14

    There are over 100 serotypes of enterovirus species A-D, which are the common cause of various symptoms in infants, such as meningitis, encephalitis and hand foot mouth disease (HFMD). This study aims to investigate the epidemiological characteristics of enteroviruses in Hangzhou, Zhejiang province, China, and to provide relevant information to guide public health responses and interventions. Systematic surveillance was conducted on enterovirus infections. Samples were collected from children admitted to the inpatient wards and outpatient departments between January 2010 and December 2012 in the Children's Hospital, Zhejiang University School of Medicine. Enteroviruses from all specimens were detected by RT-PCR using a commercialized detection kit. From 13026 samples collected and examined, 2673 (21.21%) were found positive for enteroviruses. The annual enterovirus-positive rate decreased from 32.78% in 2010 to 14.23% in 2012. Positivity rate for enteroviruses was highest among children aged less than 5 years. The monthly positivity rate for enterovirus infection ranged from 2.6% to 34.83%, with a peak in June and July. Serotypes causing severe symptoms such as HFMD including EV71 and CA16 were decreasing, while the proportion of unidentified EV serotypes causing herpangina and viral encephalitis were on the rise. EV infection is highly prevalent among young children in Hangzhou, as it is in the most other parts of the world. Further surveillance using methods that can subtype all EVs is warranted to better monitor these infections and their etiology.

  11. [Contamination of protozoa by enteroviruses in fresh water and sewages].

    Science.gov (United States)

    Skachkov, M V; Al'misheva, A Sh; Plotnikov, A O; Nemtseva, N V; Skvortsov, V O

    2009-01-01

    To determine rate of infection of protozoa by enteroviruses to assess the potential role of protozoa as a natural reservoir of enteroviruses. The samples were collected from flowing and stagnant water reservoirs in Orenburg region in summer and autumn. The samples of sewages were taken in all stages of their treatment. Cultures of protozoa were isolated with micromanipulator equipped with micropipette, incubated on Pratt's medium at 25 degrees C and fed with Pseudomonas fluorescens culture. RNA of enteroviruses was detected by reverse transcription polymerase chain reaction (RT-PCR). Seventy-two protozoan species were found in Ural river, whereas 15 and 38 species were found in lakes and sewages respectively. Enteroviruses were detected by RT-PCR in 61.8% cultures of protozoa belonging to 23 species of flagellates, amoebae and ciliates isolated from natural water bodies undergoing anthropogenic impact as well as from sewages in all stages of their treatment. Predominant localization of enteroviruses in dominant taxons of protozoa (Paraphysomonas sp., Spumella sp., Petalomonas poosilla, Amoeba sp.) was noted. Obtained data confirm presence of enteroviruses in protozoa living both in flowing and stagnant recreation natural water bodies as well as in sewages and confirm the hypothesis of persistence of enteroviruses in protozoa and the reservoir role of the latter. Contingency of life cycles of viruses and protozoa allows to explain the seasonality of aseptic meningitis incidence caused by enteroviruses, which peaks in summer and autumn when protozoa massively multiply in water bodies.

  12. Rapid and recent diversification patterns in Anseriformes birds: Inferred from molecular phylogeny and diversification analyses.

    Directory of Open Access Journals (Sweden)

    Zhonglou Sun

    Full Text Available The Anseriformes is a well-known and widely distributed bird order, with more than 150 species in the world. This paper aims to revise the classification, determine the phylogenetic relationships and diversification patterns in Anseriformes by exploring the Cyt b, ND2, COI genes and the complete mitochondrial genomes (mito-genomes. Molecular phylogeny and genetic distance analyses suggest that the Dendrocygna species should be considered as an independent family, Dendrocygnidae, rather than a member of Anatidae. Molecular timescale analyses suggests that the ancestral diversification occurred during the Early Eocene Climatic Optimum (58 ~ 50 Ma. Furthermore, diversification analyses showed that, after a long period of constant diversification, the median initial speciation rate was accelerated three times, and finally increased to approximately 0.3 sp/My. In the present study, both molecular phylogeny and diversification analyses results support that Anseriformes birds underwent rapid and recent diversification in their evolutionary history, especially in modern ducks, which show extreme diversification during the Plio-Pleistocene (~ 5.3 Ma. Therefore, our study support that the Plio-Pleistocene climate fluctuations are likely to have played a significant role in promoting the recent diversification for Anseriformes.

  13. Rapid and recent diversification patterns in Anseriformes birds: Inferred from molecular phylogeny and diversification analyses.

    Science.gov (United States)

    Sun, Zhonglou; Pan, Tao; Hu, Chaochao; Sun, Lu; Ding, Hengwu; Wang, Hui; Zhang, Chenling; Jin, Hong; Chang, Qing; Kan, Xianzhao; Zhang, Baowei

    2017-01-01

    The Anseriformes is a well-known and widely distributed bird order, with more than 150 species in the world. This paper aims to revise the classification, determine the phylogenetic relationships and diversification patterns in Anseriformes by exploring the Cyt b, ND2, COI genes and the complete mitochondrial genomes (mito-genomes). Molecular phylogeny and genetic distance analyses suggest that the Dendrocygna species should be considered as an independent family, Dendrocygnidae, rather than a member of Anatidae. Molecular timescale analyses suggests that the ancestral diversification occurred during the Early Eocene Climatic Optimum (58 ~ 50 Ma). Furthermore, diversification analyses showed that, after a long period of constant diversification, the median initial speciation rate was accelerated three times, and finally increased to approximately 0.3 sp/My. In the present study, both molecular phylogeny and diversification analyses results support that Anseriformes birds underwent rapid and recent diversification in their evolutionary history, especially in modern ducks, which show extreme diversification during the Plio-Pleistocene (~ 5.3 Ma). Therefore, our study support that the Plio-Pleistocene climate fluctuations are likely to have played a significant role in promoting the recent diversification for Anseriformes.

  14. Survey of enterovirus infections from hand, foot and mouth disease outbreak in china, 2009

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    Yang Fan

    2011-11-01

    Full Text Available Abstract Background In China, a rapid expansion of Hand, foot, and mouth disease (HFMD outbreaks has occurred since 2004 and HFMD has become an important issue for China. However, people are still only concerned with human enterovirus 71(HEV-71 and coxsackie virus A16 (CV-A16. Much of what is known about the other enterovirus infections relies on fractional evidence and old epidemic data, with little knowledge concerning their distribution. To alert potential threatens of the other enteroviruses, our study genetically characterized specimens from different regions of China and yielded novel information concerning the circulating and phylogenetic characteristics of enteroviral strains from HFMD cases. Findings A total of 301 clinical throat swabs were randomly obtained from patients suffering from HFMD from the southern, northern and central regions of China during outbreaks in 2009. 266 of 301 (88.4% HFMD cases were found positive for HEV and seven genotypes, HEV-71, CV-A16, -B5, -A4, -A6, -A10, and -A12, were detected. Conclusions The HFMD pathogen compositions in the different regions of China were significantly different. HFMD epidemics might persist for a long time in China due to the multiple pathogen compositions, the enteroviral characteristic of recombination and co-infection, the ever-increasing travel and migration and the deficiency of effective vaccine. Our study deserves the attention on HFMD control and vaccine development.

  15. Binding of glutathione to enterovirus capsids is essential for virion morphogenesis.

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    Hendrik Jan Thibaut

    2014-04-01

    Full Text Available Enteroviruses (family of the Picornaviridae cover a large group of medically important human pathogens for which no antiviral treatment is approved. Although these viruses have been extensively studied, some aspects of the viral life cycle, in particular morphogenesis, are yet poorly understood. We report the discovery of TP219 as a novel inhibitor of the replication of several enteroviruses, including coxsackievirus and poliovirus. We show that TP219 binds directly glutathione (GSH, thereby rapidly depleting intracellular GSH levels and that this interferes with virus morphogenesis without affecting viral RNA replication. The inhibitory effect on assembly was shown not to depend on an altered reducing environment. Using TP219, we show that GSH is an essential stabilizing cofactor during the transition of protomeric particles into pentameric particles. Sequential passaging of coxsackievirus B3 in the presence of low GSH-levels selected for GSH-independent mutants that harbored a surface-exposed methionine in VP1 at the interface between two protomers. In line with this observation, enteroviruses that already contained this surface-exposed methionine, such as EV71, did not rely on GSH for virus morphogenesis. Biochemical and microscopical analysis provided strong evidence for a direct interaction between GSH and wildtype VP1 and a role for this interaction in localizing assembly intermediates to replication sites. Consistently, the interaction between GSH and mutant VP1 was abolished resulting in a relocalization of the assembly intermediates to replication sites independent from GSH. This study thus reveals GSH as a novel stabilizing host factor essential for the production of infectious enterovirus progeny and provides new insights into the poorly understood process of morphogenesis.

  16. Binding of Glutathione to Enterovirus Capsids Is Essential for Virion Morphogenesis

    Science.gov (United States)

    Thibaut, Hendrik Jan; Thys, Bert; Canela, María-Dolores; Aguado, Leire; Wimmer, Eckard; Paul, Aniko; Pérez-Pérez, María-Jesús; van Kuppeveld, Frank J. M.; Neyts, Johan

    2014-01-01

    Enteroviruses (family of the Picornaviridae) cover a large group of medically important human pathogens for which no antiviral treatment is approved. Although these viruses have been extensively studied, some aspects of the viral life cycle, in particular morphogenesis, are yet poorly understood. We report the discovery of TP219 as a novel inhibitor of the replication of several enteroviruses, including coxsackievirus and poliovirus. We show that TP219 binds directly glutathione (GSH), thereby rapidly depleting intracellular GSH levels and that this interferes with virus morphogenesis without affecting viral RNA replication. The inhibitory effect on assembly was shown not to depend on an altered reducing environment. Using TP219, we show that GSH is an essential stabilizing cofactor during the transition of protomeric particles into pentameric particles. Sequential passaging of coxsackievirus B3 in the presence of low GSH-levels selected for GSH-independent mutants that harbored a surface-exposed methionine in VP1 at the interface between two protomers. In line with this observation, enteroviruses that already contained this surface-exposed methionine, such as EV71, did not rely on GSH for virus morphogenesis. Biochemical and microscopical analysis provided strong evidence for a direct interaction between GSH and wildtype VP1 and a role for this interaction in localizing assembly intermediates to replication sites. Consistently, the interaction between GSH and mutant VP1 was abolished resulting in a relocalization of the assembly intermediates to replication sites independent from GSH. This study thus reveals GSH as a novel stabilizing host factor essential for the production of infectious enterovirus progeny and provides new insights into the poorly understood process of morphogenesis. PMID:24722756

  17. Near instrument-free, simple molecular device for rapid detection of herpes simplex viruses.

    Science.gov (United States)

    Lemieux, Bertrand; Li, Ying; Kong, Huimin; Tang, Yi-Wei

    2012-06-01

    The first near instrument-free, inexpensive and simple molecular diagnostic device (IsoAmp HSV, BioHelix Corp., MA, USA) recently received US FDA clearance for use in the detection of herpes simplex viruses (HSV) in genital and oral lesion specimens. The IsoAmp HSV assay uses isothermal helicase-dependent amplification in combination with a disposable, hermetically-sealed, vertical-flow strip identification. The IsoAmp HSV assay has a total test-to-result time of less than 1.5 h by omitting the time-consuming nucleic acid extraction. The diagnostic sensitivity and specificity are comparable to PCR and are superior to culture-based methods. The near instrument-free, rapid and simple characteristics of the IsoAmp HSV assay make it potentially suitable for point-of-care testing.

  18. A rapid, one step molecular identification of Trichoderma citrinoviride and Trichoderma reesei.

    Science.gov (United States)

    Saroj, Dina B; Dengeti, Shrinivas N; Aher, Supriya; Gupta, Anil K

    2015-06-01

    Trichoderma species are widely used as production hosts for industrial enzymes. Identification of Trichoderma species requires a complex molecular biology based identification involving amplification and sequencing of multiple genes. Industrial laboratories are required to run identification tests repeatedly in cell banking procedures and also to prove absence of production host in the product. Such demands can be fulfilled by a brief method which enables confirmation of strain identity. This communication describes one step identification method for two common Trichoderma species; T. citrinoviride and T. reesei, based on identification of polymorphic region in the nucleotide sequence of translation elongation factor 1 alpha. A unique forward primer and common reverse primer resulted in 153 and 139 bp amplicon for T. citrinoviride and T. reesei, respectively. Simplification was further introduced by using mycelium as template for PCR amplification. Method described in this communication allows rapid, one step identification of two Trichoderma species.

  19. Enterovirus Species B Bias of RD Cell Line and Its Influence on Enterovirus Diversity Landscape.

    Science.gov (United States)

    Faleye, Temitope Oluwasegun Cephas; Adeniji, Johnson Adekunle

    2015-12-01

    Despite its widespread use in poliovirus isolation, studies show that most RD cell line isolates are species B enteroviruses (EB), it was therefore employed to further catalogue the EB diversity in two different regions of Nigeria. Concentrates of 18 environmental samples were inoculated into RD cell line. Isolates were subjected to PCR assays to detect enteroviruses, species C and B members and partial VP1 gene which was subsequently sequenced and used for identification and phylogenetic analysis. Isolates were further passaged in L20B cell line to detect polioviruses. Sixty-eight isolates were recovered from the 18 concentrates, all of which were positive for the enterovirus 5'-UTR screen. Thirteen of the 68 isolates were positive for the species C screen and replicated in L20B cell line, eleven of which also contained species B enteroviruses. Some of the mixed isolates were successfully typed, but as species B members. In all, isolates recovered in this study were identified as CVB5, E6, E7, E11, E13, E19, E20, E33, EVB75 and WPV3, while some could not be typed. Alongside the ten different enterovirus serotypes confirmed, results of this study document for the first time in Nigeria, EVB75. It showed the EB bias of RD cell line might indicate something much more fundamental in its biology. Finally, the finding of WPV3 in a region considered low risk for poliovirus emphasizes the need to expand poliovirus environmental surveillance to enable early detection of poliovirus silent circulation before occurrence of clinical manifestations.

  20. Fabrication of Polymerase Chain Reaction Plastic Lab-on-a-Chip Device for Rapid Molecular Diagnoses

    Directory of Open Access Journals (Sweden)

    Kieu The Loan Trinh

    2016-05-01

    Full Text Available Purpose: We aim to fabricate a thermoplastic poly(methylmethacrylate (PMMA Lab-on-a-Chip device to perform continuous- flow polymerase chain reactions (PCRs for rapid molecular detection of foodborne pathogen bacteria. Methods: A miniaturized plastic device was fabricated by utilizing PMMA substrates mediated by poly(dimethylsiloxane interfacial coating, enabling bonding under mild conditions, and thus avoiding the deformation or collapse of microchannels. Surface characterizations were carried out and bond strength was measured. The feasibility of the Lab-on-a-Chip device for performing on-chip PCR utilizing a lab-made, portable dual heater was evaluated. The results were compared with those obtained using a commercially available thermal cycler. Results: A PMMA Lab-on-a-Chip device was designed and fabricated for conducting PCR using foodborne pathogens as sample targets. A robust bond was established between the PMMA substrates, which is essential for performing miniaturized PCR on plastic. The feasibility of on-chip PCR was evaluated using Escherichia coli O157:H7 and Cronobacter condimenti, two worldwide foodborne pathogens, and the target amplicons were successfully amplified within 25 minutes. Conclusions: In this study, we present a novel design of a low-cost and high-throughput thermoplastic PMMA Lab-on-a-Chip device for conducting microscale PCR, and we enable rapid molecular diagnoses of two important foodborne pathogens in minute resolution using this device. In this regard, the introduced highly portable system design has the potential to enable PCR investigations of many diseases quickly and accurately.

  1. Ancient low-molecular-weight organic acids in permafrost fuel rapid carbon dioxide production upon thaw.

    Science.gov (United States)

    Drake, Travis W; Wickland, Kimberly P; Spencer, Robert G M; McKnight, Diane M; Striegl, Robert G

    2015-11-10

    Northern permafrost soils store a vast reservoir of carbon, nearly twice that of the present atmosphere. Current and projected climate warming threatens widespread thaw of these frozen, organic carbon (OC)-rich soils. Upon thaw, mobilized permafrost OC in dissolved and particulate forms can enter streams and rivers, which are important processors of OC and conduits for carbon dioxide (CO2) to the atmosphere. Here, we demonstrate that ancient dissolved organic carbon (DOC) leached from 35,800 y B.P. permafrost soils is rapidly mineralized to CO2. During 200-h experiments in a novel high-temporal-resolution bioreactor, DOC concentration decreased by an average of 53%, fueling a more than sevenfold increase in dissolved inorganic carbon (DIC) concentration. Eighty-seven percent of the DOC loss to microbial uptake was derived from the low-molecular-weight (LMW) organic acids acetate and butyrate. To our knowledge, our study is the first to directly quantify high CO2 production rates from permafrost-derived LMW DOC mineralization. The observed DOC loss rates are among the highest reported for permafrost carbon and demonstrate the potential importance of LMW DOC in driving the rapid metabolism of Pleistocene-age permafrost carbon upon thaw and the outgassing of CO2 to the atmosphere by soils and nearby inland waters.

  2. Ancient low–molecular-weight organic acids in permafrost fuel rapid carbon dioxide production upon thaw

    Science.gov (United States)

    Drake, Travis W.; Wickland, Kimberly P.; Spencer, Robert G. M.; McKnight, Diane M.; Striegl, Robert G.

    2015-01-01

    Northern permafrost soils store a vast reservoir of carbon, nearly twice that of the present atmosphere. Current and projected climate warming threatens widespread thaw of these frozen, organic carbon (OC)-rich soils. Upon thaw, mobilized permafrost OC in dissolved and particulate forms can enter streams and rivers, which are important processors of OC and conduits for carbon dioxide (CO2) to the atmosphere. Here, we demonstrate that ancient dissolved organic carbon (DOC) leached from 35,800 y B.P. permafrost soils is rapidly mineralized to CO2. During 200-h experiments in a novel high–temporal-resolution bioreactor, DOC concentration decreased by an average of 53%, fueling a more than sevenfold increase in dissolved inorganic carbon (DIC) concentration. Eighty-seven percent of the DOC loss to microbial uptake was derived from the low–molecular-weight (LMW) organic acids acetate and butyrate. To our knowledge, our study is the first to directly quantify high CO2 production rates from permafrost-derived LMW DOC mineralization. The observed DOC loss rates are among the highest reported for permafrost carbon and demonstrate the potential importance of LMW DOC in driving the rapid metabolism of Pleistocene-age permafrost carbon upon thaw and the outgassing of CO2 to the atmosphere by soils and nearby inland waters.

  3. Rapid Characterization of Molecular Chemistry, Nutrient Make-Up and Microlocation of Internal Seed Tissue

    International Nuclear Information System (INIS)

    Yu, P.; Block, H.; Niu, Z.; Doiron, K.

    2007-01-01

    Wheat differs from corn in biodegradation kinetics and fermentation characteristics. Wheat exhibits a relatively high rate (23% h 01 ) and extent (78% DM) of biodegradation, which can lead to metabolic problems such as acidosis and bloat in ruminants. The objective of this study was to rapidly characterize the molecular chemistry of the internal structure of wheat (cv. AC Barrie) and reveal both its structural chemical make-up and nutrient component matrix by analyzing the intensity and spatial distribution of molecular functional groups within the intact seed using advanced synchrotron-powered Fourier transform infrared (FTIR) microspectroscopy. The experiment was performed at the U2B station of the National Synchrotron Light Source at Brookhaven National Laboratory, New York, USA. The wheat tissue was imaged systematically from the pericarp, seed coat, aleurone layer and endosperm under the peaks at ∼1732 (carbonyl C(double b ond)O ester), 1515 (aromatic compound of lignin), 1650 (amide I), 1025 (non-structural CHO), 1550 (amide II), 1246 (cellulosic material), 1160, 1150, 1080, 930, 860 (all CHO), 3350 (OH and NH stretching), 2928 (CH 2 stretching band) and 2885 cm -1 (CH 3 stretching band). Hierarchical cluster analysis and principal component analysis were applied to analyze the molecular FTIR spectra obtained from the different inherent structures within the intact wheat tissues. The results showed that, with synchrotron-powered FTIR microspectroscopy, images of the molecular chemistry of wheat could be generated at an ultra-spatial resolution. The features of aromatic lignin, structural and non-structural carbohydrates, as well as nutrient make-up and interactions in the seeds, could be revealed. Both principal component analysis and hierarchical cluster analysis methods are conclusive in showing that they can discriminate and classify the different inherent structures within the seed tissue. The wheat exhibited distinguishable differences in the

  4. Impact of cell lines included in enterovirus isolation protocol on perception of nonpolio enterovirus species C diversity.

    Science.gov (United States)

    Adeniji, Johnson Adekunle; Faleye, Temitope Oluwasegun Cephas

    2014-10-01

    There has been under-reporting of nonpolio enterovirus species Cs (NPESCs) in Nigeria despite the fact that most isolates recovered from the Nigerian vaccine derived poliovirus serotype 2 (VDPV2) outbreak were recombinants with nonstructural region of NPESC origin. It has been suggested that cell lines included in enterovirus isolation protocols might account for this phenomenon and this study examined this suggestion. Fifteen environmental samples concentrated previously and analysed using L20B and RD cell lines as part of the poliovirus environmental surveillance (ES) program in Nigeria were randomly selected and inoculated into two cell lines (MCF-7 and LLC-MK2). Isolates were identified as enteroviruses and species C members using different RT-PCR assays, culture in L20B cell line and sequencing of partial VP1. Forty-eight (48) isolates were recovered from the 15 samples, 47 (97.9%) of which were enteroviruses. Of the enteroviruses, 32 (68.1%) belonged to enterovirus species C (EC) of which 19 (40.4%) were polioviruses and 13 (27.7%) were NPESC members. All 13 NPESC isolates were recovered on MCF-7. Results of the study show that NPESCs are circulating in Nigeria and their under-reporting was due to the combination of cell lines used for enterovirus isolation in previous reports. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. Improving the diagnosis of meningitis due to enterovirus and herpes simplex virus I and II in a tertiary care hospital

    Science.gov (United States)

    2013-01-01

    Background Enterovirus and herpes simplex viruses are common causes of lymphocytic meningitis. The purpose of this study was to analyse the impact of the use molecular testing for Enteroviruses and Herpes simplex viruses I and II in all suspected cases of viral meningitis. Methods From November 18, 2008 to November 17, 2009 (phase II, intervention), all patients admitted with suspected viral meningitis (with pleocytosis) had a CSF sample tested using a nucleic acid amplification test (NAAT). Data collected during this period were compared to those from the previous one-year period, i.e. November 18, 2007 to November 17, 2008 (phase I, observational), when such tests were available but not routinely used. Results In total, 2,536 CSF samples were assessed, of which 1,264 were from phase I, and 1,272 from phase II. Of this total, a NAAT for Enterovirus was ordered in 123 cases during phase I (9.7% of the total phase I sample) and in 221 cases in phase II (17.4% of the total phase II sample). From these, Enterovirus was confirmed in 35 (28.5%, 35/123) patients during phase I and 71 (32.1%, 71/221) patients during phase II (p = 0.107). The rate of diagnosis of meningitis by HSV I and II did not differ between the groups (13 patients, 6.5% in phase I and 13, 4.7% in phase II) (p = 1.0), from 200 cases in phase I and 274 cases in phase II. Conclusions The number of cases diagnosed with enteroviral meningitis increased during the course of this study, leading us to believe that the strategy of performing NAAT for Enterovirus on every CSF sample with pleocytosis is fully justified. PMID:24138798

  6. COPI is required for enterovirus 71 replication.

    Directory of Open Access Journals (Sweden)

    Jianmin Wang

    Full Text Available Enterovirus 71 (EV71, a member of the Picornaviridae family, is found in Asian countries where it causes a wide range of human diseases. No effective therapy is available for the treatment of these infections. Picornaviruses undergo RNA replication in association with membranes of infected cells. COPI and COPII have been shown to be involved in the formation of picornavirus-induced vesicles. Replication of several picornaviruses, including poliovirus and Echovirus 11 (EV11, is dependent on COPI or COPII. Here, we report that COPI, but not COPII, is required for EV71 replication. Replication of EV71 was inhibited by brefeldin A and golgicide A, inhibitors of COPI activity. Furthermore, we found EV71 2C protein interacted with COPI subunits by co-immunoprecipitation and GST pull-down assay, indicating that COPI coatomer might be directed to the viral replication complex through viral 2C protein. Additionally, because the pathway is conserved among different species of enteroviruses, it may represent a novel target for antiviral therapies.

  7. Precise genotyping and recombination detection of Enterovirus

    Science.gov (United States)

    2015-01-01

    Enteroviruses (EV) with different genotypes cause diverse infectious diseases in humans and mammals. A correct EV typing result is crucial for effective medical treatment and disease control; however, the emergence of novel viral strains has impaired the performance of available diagnostic tools. Here, we present a web-based tool, named EVIDENCE (EnteroVirus In DEep conception, http://symbiont.iis.sinica.edu.tw/evidence), for EV genotyping and recombination detection. We introduce the idea of using mixed-ranking scores to evaluate the fitness of prototypes based on relatedness and on the genome regions of interest. Using phylogenetic methods, the most possible genotype is determined based on the closest neighbor among the selected references. To detect possible recombination events, EVIDENCE calculates the sequence distance and phylogenetic relationship among sequences of all sliding windows scanning over the whole genome. Detected recombination events are plotted in an interactive figure for viewing of fine details. In addition, all EV sequences available in GenBank were collected and revised using the latest classification and nomenclature of EV in EVIDENCE. These sequences are built into the database and are retrieved in an indexed catalog, or can be searched for by keywords or by sequence similarity. EVIDENCE is the first web-based tool containing pipelines for genotyping and recombination detection, with updated, built-in, and complete reference sequences to improve sensitivity and specificity. The use of EVIDENCE can accelerate genotype identification, aiding clinical diagnosis and enhancing our understanding of EV evolution. PMID:26678286

  8. Infectious Entry Pathway of Enterovirus B Species

    Directory of Open Access Journals (Sweden)

    Varpu Marjomäki

    2015-12-01

    Full Text Available Enterovirus B species (EV-B are responsible for a vast number of mild and serious acute infections. They are also suspected of remaining in the body, where they cause persistent infections contributing to chronic diseases such as type I diabetes. Recent studies of the infectious entry pathway of these viruses revealed remarkable similarities, including non-clathrin entry of large endosomes originating from the plasma membrane invaginations. Many cellular factors regulating the efficient entry have recently been associated with macropinocytic uptake, such as Rac1, serine/threonine p21-activated kinase (Pak1, actin, Na/H exchanger, phospholipace C (PLC and protein kinase Cα (PKCα. Another characteristic feature is the entry of these viruses to neutral endosomes, independence of endosomal acidification and low association with acidic lysosomes. The biogenesis of neutral multivesicular bodies is crucial for their infection, at least for echovirus 1 (E1 and coxsackievirus A9 (CVA9. These pathways are triggered by the virus binding to their receptors on the plasma membrane, and they are not efficiently recycled like other cellular pathways used by circulating receptors. Therefore, the best “markers” of these pathways may be the viruses and often their receptors. A deeper understanding of this pathway and associated endosomes is crucial in elucidating the mechanisms of enterovirus uncoating and genome release from the endosomes to start efficient replication.

  9. Development of Novel Vaccines against Enterovirus-71

    Science.gov (United States)

    Yee, Pinn Tsin Isabel; Poh, Chit Laa

    2015-01-01

    The hand, foot and mouth disease is caused by a group of Enteroviruses such as Enterovirus 71 (EV-A71) and Coxsackievirus CV-A5, CV-A8, and CV-A16. Mild symptoms of EV-A71 infection in children range from high fever, vomiting, rashes and ulcers in mouth but can produce more severe symptoms such as brainstem and cerebellar encephalitis, leading up to cardiopulmonary failure and death. The lack of vaccines and antiviral drugs against EV-A71 highlights the urgency of developing preventive and treatment agents against EV-A71 to prevent further fatalities. Research groups have developed experimental inactivated vaccines, recombinant Viral Protein 1 (VP1) vaccine and virus-like particles (VLPs). The inactivated EV-A71 vaccine is considered the safest viral vaccine, as there will be no reversion to the infectious wild type strain. The recombinant VP1 vaccine is a cost-effective immunogen, while VLPs contain an arrangement of epitopes that can elicit neutralizing antibodies against the virus. As each type of vaccine has its advantages and disadvantages, increased studies are required in the development of such vaccines, whereby high efficacy, long-lasting immunity, minimal risk to those vaccinated, safe and easy production, low cost, dispensing the need for refrigeration and convenient delivery are the major goals in their design. PMID:26729152

  10. Rapid molecular evolution of human bocavirus revealed by Bayesian coalescent inference.

    Science.gov (United States)

    Zehender, Gianguglielmo; De Maddalena, Chiara; Canuti, Marta; Zappa, Alessandra; Amendola, Antonella; Lai, Alessia; Galli, Massimo; Tanzi, Elisabetta

    2010-03-01

    Human bocavirus (HBoV) is a linear single-stranded DNA virus belonging to the Parvoviridae family that has recently been isolated from the upper respiratory tract of children with acute respiratory infection. All of the strains observed so far segregate into two genotypes (1 and 2) with a low level of polymorphism. Given the recent description of the infection and the lack of epidemiological and molecular data, we estimated the virus's rates of molecular evolution and population dynamics. A dataset of forty-nine dated VP2 sequences, including also eight new isolates obtained from pharyngeal swabs of Italian patients with acute respiratory tract infections, was submitted to phylogenetic analysis. The model parameters, evolutionary rates and population dynamics were co-estimated using a Bayesian Markov Chain Monte Carlo approach, and site-specific positive and negative selection was also investigated. Recombination was investigated by seven different methods and one suspected recombinant strain was excluded from further analysis. The estimated mean evolutionary rate of HBoV was 8.6x10(-4)subs/site/year, and that of the 1st+2nd codon positions was more than 15 times less than that of the 3rd codon position. Viral population dynamics analysis revealed that the two known genotypes diverged recently (mean tMRCA: 24 years), and that the epidemic due to HBoV genotype 2 grew exponentially at a rate of 1.01year(-1). Selection analysis of the partial VP2 showed that 8.5% of sites were under significant negative pressure and the absence of positive selection. Our results show that, like other parvoviruses, HBoV is characterised by a rapid evolution. The low level of polymorphism is probably due to a relatively recent divergence between the circulating genotypes and strong purifying selection acting on viral antigens.

  11. Human Rhinovirus 87 and Enterovirus 68 Represent a Unique Serotype with Rhinovirus and Enterovirus Features

    Science.gov (United States)

    Blomqvist, Soile; Savolainen, Carita; Råman, Laura; Roivainen, Merja; Hovi, Tapani

    2002-01-01

    It has recently been reported that all but one of the 102 known serotypes of the genus Rhinovirus segregate into two genetic clusters (C. Savolainen, S. Blomqvist, M. N. Mulders, and T. Hovi, J. Gen. Virol. 83:333-340, 2002). The only exception is human rhinovirus 87 (HRV87). Here we demonstrate that HRV87 is genetically and antigenically highly similar to enterovirus 68 (EV68) and is related to EV70, the other member of human enterovirus group D. The partial nucleotide sequences of the 5′ untranslated region, capsid regions VP4/VP2 and VP1, and the 3D RNA polymerase gene of the HRV87 prototype strain F02-3607 Corn showed 97.3, 97.8, 95.2, and 95.9% identity to the corresponding regions of EV68 prototype strain Fermon. The amino acid identities were 100 and 98.1% for the products of the two capsid regions and 97.9% for 3D RNA polymerase. Antigenic cross-reaction between HRV87 and EV68 was indicated by microneutralization with monotypic antisera. Phylogenetic analysis showed definite clustering of HRV87 and EV68 with EV70 for all sequences examined. Both HRV87 and EV68 were shown to be acid sensitive by two different assays, while EV70 was acid resistant, which is typical of enteroviruses. The cytopathic effect induced by HRV87 or EV68 was inhibited by monoclonal antibodies to the decay-accelerating factor known to be the receptor of EV70. We conclude that HRV87 and EV68 are strains of the same picornavirus serotype presenting features of both rhinoviruses and enteroviruses. PMID:12409401

  12. Enterovirus 71 Brainstem Encephalitis and Cognitive and Motor Deficits

    Directory of Open Access Journals (Sweden)

    J Gordon Millichap

    2006-12-01

    Full Text Available Follow-up studies were conducted in 63 previously healthy children with enterovirus 71 brainstem encephalitis (49 stage II, 7 stage Ilia, and 7 stage Illb at National Cheng Kung University Hospital, Tainan, Taiwan.

  13. Enterovirus species and serotype distributions in monthly municipal wastewater samples

    Data.gov (United States)

    U.S. Environmental Protection Agency — This dataset contains the enterovirus genome copies measured per liter of sample and read counts obtained from amplicon sequencing of the VP1 and VP4 genes. This...

  14. Meteorite Impact-Induced Rapid NH3 Production on Early Earth: Ab Initio Molecular Dynamics Simulation

    Science.gov (United States)

    Shimamura, Kohei; Shimojo, Fuyuki; Nakano, Aiichiro; Tanaka, Shigenori

    2016-12-01

    NH3 is an essential molecule as a nitrogen source for prebiotic amino acid syntheses such as the Strecker reaction. Previous shock experiments demonstrated that meteorite impacts on ancient oceans would have provided a considerable amount of NH3 from atmospheric N2 and oceanic H2O through reduction by meteoritic iron. However, specific production mechanisms remain unclear, and impact velocities employed in the experiments were substantially lower than typical impact velocities of meteorites on the early Earth. Here, to investigate the issues from the atomistic viewpoint, we performed multi-scale shock technique-based ab initio molecular dynamics simulations. The results revealed a rapid production of NH3 within several picoseconds after the shock, indicating that shocks with greater impact velocities would provide further increase in the yield of NH3. Meanwhile, the picosecond-order production makes one expect that the important nitrogen source precursors of amino acids were obtained immediately after the impact. It was also observed that the reduction of N2 proceeded according to an associative mechanism, rather than a dissociative mechanism as in the Haber-Bosch process.

  15. Rapid molecular diagnostics of severe primary immunodeficiency determined by using targeted next-generation sequencing.

    Science.gov (United States)

    Yu, Hui; Zhang, Victor Wei; Stray-Pedersen, Asbjørg; Hanson, Imelda Celine; Forbes, Lisa R; de la Morena, M Teresa; Chinn, Ivan K; Gorman, Elizabeth; Mendelsohn, Nancy J; Pozos, Tamara; Wiszniewski, Wojciech; Nicholas, Sarah K; Yates, Anne B; Moore, Lindsey E; Berge, Knut Erik; Sorte, Hanne; Bayer, Diana K; ALZahrani, Daifulah; Geha, Raif S; Feng, Yanming; Wang, Guoli; Orange, Jordan S; Lupski, James R; Wang, Jing; Wong, Lee-Jun

    2016-10-01

    Primary immunodeficiency diseases (PIDDs) are inherited disorders of the immune system. The most severe form, severe combined immunodeficiency (SCID), presents with profound deficiencies of T cells, B cells, or both at birth. If not treated promptly, affected patients usually do not live beyond infancy because of infections. Genetic heterogeneity of SCID frequently delays the diagnosis; a specific diagnosis is crucial for life-saving treatment and optimal management. We developed a next-generation sequencing (NGS)-based multigene-targeted panel for SCID and other severe PIDDs requiring rapid therapeutic actions in a clinical laboratory setting. The target gene capture/NGS assay provides an average read depth of approximately 1000×. The deep coverage facilitates simultaneous detection of single nucleotide variants and exonic copy number variants in one comprehensive assessment. Exons with insufficient coverage (diagnostic yield of severe primary immunodeficiency. Establishing a molecular diagnosis enables early immune reconstitution through prompt therapeutic intervention and guides management for improved long-term quality of life. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  16. Rapid Evaporation of Water on Graphene/Graphene-Oxide: A Molecular Dynamics Study.

    Science.gov (United States)

    Li, Qibin; Xiao, Yitian; Shi, Xiaoyang; Song, Shufeng

    2017-09-07

    To reveal the mechanism of energy storage in the water/graphene system and water/grapheme-oxide system, the processes of rapid evaporation of water molecules on the sheets of graphene and graphene-oxide are investigated by molecular dynamics simulations. The results show that both the water/graphene and water/grapheme-oxide systems can store more energy than the pure water system during evaporation. The hydroxyl groups on the surface of graphene-oxide are able to reduce the attractive interactions between water molecules and the sheet of graphene-oxide. Also, the radial distribution function of the oxygen atom indicates that the hydroxyl groups affect the arrangement of water molecules at the water/graphene-oxide interface. Therefore, the capacity of thermal energy storage of the water/graphene-oxide system is lower than that of the water/graphene system, because of less desorption energy at the water/graphene-oxide interface. Also, the evaporation rate of water molecules on the graphene-oxide sheet is slower than that on the graphene sheet. The Leidenfrost phenomenon can be observed during the evaporation process in the water/grapheme-oxide system.

  17. Seroprevalence of enterovirus 71 and no evidence of crossprotection of enterovirus 71 antibody against the other enteroviruses in kindergarten children in Taipei city.

    Science.gov (United States)

    Huang, Wen-Chan; Huang, Li-Min; Kao, Chuan-Liang; Lu, Chun-Yi; Shao, Pei-Lan; Cheng, Ai-Ling; Fan, Tsui-Yien; Chi, Hsin; Chang, Luan-Yin

    2012-04-01

    Enterovirus 71 (EV71) infection may cause severe neurological and cardiopulmonary complications, especially in preschool children. This study is to investigate the seroprevalence and seroconversion of EV71, and the crossprotection of EV71 antibody against other enteroviruses among kindergarteners. Overall 228 children in a public kindergarten were enrolled during two academic years, 2006 and 2007, in Taipei, Taiwan and we measured their EV71 neutralizing antibody. When the participants had herpangina; hand, foot and mouth disease (HFMD); febrile illness or respiratory symptoms, throat swabs were sampled and processed for viral culture and enterovirus real-time reverse transcriptase polymerase chain reaction (RT-PCR). Questionnaires, completed by the participants' guardians, surveyed the history of allergy and annual incidence of symptoms related to enterovirus infection. Seropositive rates of EV71 were 20% (32/163) in 2006 and 6% (4/65) in 2007. The rate of EV71 seropositivity increased with age (p Taipei City from September 2006 to June 2008. Presence of EV71 neutralizing antibody was not associated with lower incidence of enterovirus infection caused by non-71 serotypes. Copyright © 2011. Published by Elsevier B.V.

  18. Clinical severity of pediatric respiratory illness with enterovirus D68 compared with rhinovirus or other enterovirus genotypes.

    Science.gov (United States)

    Mertz, Dominik; Alawfi, Abdulsalam; Pernica, Jeffrey M; Rutherford, Candy; Luinstra, Kathy; Smieja, Marek

    2015-11-17

    Enterovirus D68 (EV-D68) resulted in a reported increase in the number of children needing hospital or critical care admission because of respiratory insufficiency during 2014. It remains unclear, however, whether EV-D68 infections were more severe than rhinovirus or non-EV-D68 enterovirus infections. We evaluated consecutive children presenting to a pediatric hospital between Aug. 1 and Oct. 31, 2014, with positive nasopharyngeal swabs for rhinovirus or enterovirus that were sent automatically for EV-D68 testing. We compared characteristics and outcomes of patients with EV-D68 with those with rhinovirus or non-EV-D68 enterovirus in a matched cohort study. A total of 93/297 (31.3%) of rhinovirus or enterovirus samples tested positive for EV-D68, and it was possible to compare 87 matched pairs. Children with EV-D68 infection were more likely to have difficulty breathing (odds ratio [OR] 3.00, 95% confidence interval [CI] 1.47-6.14). There was no significant difference in admission to the critical care unit or death among children with EV-D68 infection compared with those with other rhinovirus or enterovirus infections (adjusted OR 1.47, 95% CI 0.61-3.52). Children with EV-D68 infection were more often admitted to hospital, but not significantly so (adjusted OR 2.29, 95% CI 0.96-5.46). Enterovirus D68 seems to be a more virulent pulmonary pathogen than rhinovirus or non-EV-D68 enterovirus, but we did not find a significant difference in death or need for critical care. © 2015 Canadian Medical Association or its licensors.

  19. Milrinone in Enterovirus 71 Brain Stem Encephalitis

    Directory of Open Access Journals (Sweden)

    SHIH-MIN eWANG

    2016-03-01

    Full Text Available Enterovirus 71 (EV71 was implicated in a widespread outbreak of hand-foot-and-mouth disease (HFMD across the Asia Pacific area since 1997 and has also been reported sporadically in patients with brain stem encephalitis. Neurogenic shock with pulmonary edema (PE is a fatal complication of EV71 infection. Among inotropic agents, milrinone is selected as a therapeutic agent for EV71- induced PE due to its immunopathogenesis. Milrinone is a type III phosphodiesterase inhibitor that has both inotropic and vasodilator effects. Its clinical efficacy has been shown by modulating inflammation, reducing sympathetic over-activity, and improving survival in patients with EV71-associated PE. Milrinone exhibits immunoregulatory and anti-inflammatory effects in the management of systemic inflammatory responses in severe EV71 infection.

  20. Multidrug-resistant tuberculosis: Rapid molecular detection with MTBDRplus® assay in clinical samples

    Directory of Open Access Journals (Sweden)

    Rita Macedo

    2009-05-01

    Full Text Available Nowadays, the greatest concern of tuberculosis control programmes is the appearance of multidrug-resistant tuberculosis and extensively drug-resistant tuberculosis. Rapid determination of drug resistance in clinical samples, with Mycobacterium tuberculosis complex (MTC, is the prerequisite for initiating effective chemotherapy, ensuring successful treatment of the patient and preventing further spread of drugresistant isolates.The aim of our study was to determine the sensitivity of the new MTBDRplus® assay in comparison to culture, identification and classic DST, directly from smear-positive clinical specimens.A total of 68 smear-positive sputum specimens were processed by both the classical mycobacteriological methods and the molecular assay, MTBDRplus®.MTBDRplus® assay allowed an accurate identification of MTC species by detection of the specific band in all samples, from which we also isolated and identified MTC strains by culture methods. In the samples from which we isolated susceptible strains (63.2%, wild type patterns were found using MTBDRplus® assay. The samples from which we isolated resistant strains (36.8% showed specific mutations associated with the correspondent resistant phenotype.Our study indicated that this assay allows rapid detection of resistance, always in agreement with classic methods. Resumo: Uma das principais problematicas no controlo da tuberculose e o aparecimento de casos de tuberculose multirresistente (TB-MR e tuberculose extensivamente resistente (TB-XDR. A deteccao precoce da resistencia a farmacos, directamente a partir de amostras respiratorias, e essencial para que se assegure o tratamento atempado, adequado e eficaz da tuberculose, bem como para prevenir a disseminacao destes casos de especial gravidade.O nosso objectivo foi avaliar a sensibilidade e comparar os resultados obtidos com um metodo de genetica molecular disponivel comercialmente – MTBDRplus® – e o isolamento

  1. Fourteen types of co-circulating recombinant enterovirus were associated with hand, foot, and mouth disease in children from Wenzhou, China.

    Science.gov (United States)

    Guo, Wen-Ping; Lin, Xian-Dan; Chen, Yi-Ping; Liu, Qi; Wang, Wen; Wang, Cai-Qiao; Li, Ming-Hui; Sun, Xiao-Yu; Shi, Mang; Holmes, Edward C; Zhang, Yong-Zhen

    2015-09-01

    Although hand, foot, and mouth disease (HFMD) is a major public concern in China, the prevalence and clinical symptoms associated with the different agents of HFMD in this country remain poorly understood. We investigated the clinical and molecular characteristics of enteroviruses in patients with HFMD from Wenzhou, China. Patients with laboratory-confirmed HFMD admitted to the Yuying Children's Hospital in Wenzhou, China during 2013 were included in this study. Viral RNA sequences were amplified using RT-PCR, determined by sequencing, and compared by phylogenetic analysis. A total of 955 clinically diagnosed HFMD cases were determined using PCR, with whole viral genomes obtained for each enterovirus type. 14 types of enterovirus belonging to two viral species were identified. Notably, Coxsackievirus A6 (CV-A6) was the most common species detected (77.8%), followed by EV-A71 (8.2%) and CV-A10 (8.1%). Phylogenetic analysis revealed multiple independent introductions of these viruses into Wenzhou. In addition, the enterovirus observed in Wenzhou had a recombinant history, with two or three recombination breakpoints. Although the illness associated with CV-A6 was milder than that of EV-A71, CV-A6 infection caused more widespread rash, larger blisters, and subsequent skin peeling and/or nail shedding. Our study revealed the co-circulation of 14 types of enteroviruses in a single location - Wenzhou, China - with CV-A6 virus the predominant agent of HFMD. This work highlights the need to perform larger-scale surveillance to fully understand the epidemiology of enteroviruses in China and the wider Asia-Pacific region. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Rapid molecular evolution across amniotes of the IIS/TOR network.

    Science.gov (United States)

    McGaugh, Suzanne E; Bronikowski, Anne M; Kuo, Chih-Horng; Reding, Dawn M; Addis, Elizabeth A; Flagel, Lex E; Janzen, Fredric J; Schwartz, Tonia S

    2015-06-02

    The insulin/insulin-like signaling and target of rapamycin (IIS/TOR) network regulates lifespan and reproduction, as well as metabolic diseases, cancer, and aging. Despite its vital role in health, comparative analyses of IIS/TOR have been limited to invertebrates and mammals. We conducted an extensive evolutionary analysis of the IIS/TOR network across 66 amniotes with 18 newly generated transcriptomes from nonavian reptiles and additional available genomes/transcriptomes. We uncovered rapid and extensive molecular evolution between reptiles (including birds) and mammals: (i) the IIS/TOR network, including the critical nodes insulin receptor substrate (IRS) and phosphatidylinositol 3-kinase (PI3K), exhibit divergent evolutionary rates between reptiles and mammals; (ii) compared with a proxy for the rest of the genome, genes of the IIS/TOR extracellular network exhibit exceptionally fast evolutionary rates; and (iii) signatures of positive selection and coevolution of the extracellular network suggest reptile- and mammal-specific interactions between members of the network. In reptiles, positively selected sites cluster on the binding surfaces of insulin-like growth factor 1 (IGF1), IGF1 receptor (IGF1R), and insulin receptor (INSR); whereas in mammals, positively selected sites clustered on the IGF2 binding surface, suggesting that these hormone-receptor binding affinities are targets of positive selection. Further, contrary to reports that IGF2R binds IGF2 only in marsupial and placental mammals, we found positively selected sites clustered on the hormone binding surface of reptile IGF2R that suggest that IGF2R binds to IGF hormones in diverse taxa and may have evolved in reptiles. These data suggest that key IIS/TOR paralogs have sub- or neofunctionalized between mammals and reptiles and that this network may underlie fundamental life history and physiological differences between these amniote sister clades.

  3. Characterization and specificity of the linear epitope of the enterovirus 71 VP2 protein

    Directory of Open Access Journals (Sweden)

    Kiener Tanja K

    2012-02-01

    Full Text Available Abstract Background Enterovirus 71 (EV71 has emerged as a major causative agent of hand, foot and mouth disease in the Asia-Pacific region over the last decade. Hand, foot and mouth disease can be caused by different etiological agents from the enterovirus family, mainly EV71 and coxsackieviruses, which are genetically closely related. Nevertheless, infection with EV71 may occasionally lead to high fever, neurologic complications and the emergence of a rapidly fatal syndrome of pulmonary edema associated with brainstem encephalitis. The rapid progression and high mortality of severe EV71 infection has highlighted the need for EV71-specific diagnostic and therapeutic tools. Monoclonal antibodies are urgently needed to specifically detect EV71 antigens from patient specimens early in the infection process. Furthermore, the elucidation of viral epitopes will contribute to the development of targeted therapeutics and vaccines. Results We have identified the monoclonal antibody 7C7 from a screen of hybridoma cells derived from mice immunized with the EV71-B5 strain. The linear epitope of 7C7 was mapped to amino acids 142-146 (EDSHP of the VP2 capsid protein and was characterized in detail. Mutational analysis of the epitope showed that the aspartic acid to asparagine mutation of the EV71 subgenogroup A (BrCr strain did not interfere with antibody recognition. In contrast, the serine to threonine mutation at position 144 of VP2, present in recently emerged EV71-C4 China strains, abolished antigenicity. Mice injected with this virus strain did not produce any antibodies against the VP2 protein. Immunofluorescence and Western blotting confirmed that 7C7 specifically recognized EV71 subgenogroups and did not cross-react to Coxsackieviruses 4, 6, 10, and 16. 7C7 was successfully used as a detection antibody in an antigen-capture ELISA assay. Conclusions Detailed mapping showed that the VP2 protein of Enterovirus 71 contains a single, linear, non

  4. The absence of exanthema is related with death and illness severity in acute enterovirus infection

    Directory of Open Access Journals (Sweden)

    Hong-Tao Zhou

    2014-11-01

    Conclusions: A considerable proportion of children with an acute enterovirus infection in Guangdong Province, China during 2009–2012 presented no exanthema, and the absence of exanthema was found to be related to death and illness severity for these acute enterovirus infections. Clinicians in China should consider enterovirus as the possible pathogen when treating children with an acute pathogen infection without exanthema.

  5. Enterovirus type D68 en acute slappe verlamming: een nieuw duo?

    NARCIS (Netherlands)

    van Dissel, J T; Meijer, A

    2017-01-01

    Following a case of acute flaccid paralysis after infection with enterovirus type D68, we highlight current understanding of the causal role of enterovirus infection in this neurological syndrome. Acute flaccid paralysis is a rare complication of enterovirus infections. Such viruses have become a

  6. Coxsackievirus A21, Enterovirus 68, and Acute Respiratory Tract Infection, China

    OpenAIRE

    Xiang, Zichun; Gonzalez, Richard; Wang, Zhong; Ren, Lili; Xiao, Yan; Li, Jianguo; Li, Yongjun; Vernet, Guy; Paranhos-Baccalà, Gláucia; Jin, Qi; Wang, Jianwei

    2012-01-01

    During August 2006–April 2010, in Beijing, China, 2 rare human enterovirus serotypes, coxsackievirus A21 and enterovirus 68, were detected most frequently in human enterovirus–positive adults with acute respiratory tract infections. Thus, during some years, these 2 viruses cause a substantial proportion of enterovirus-associated adult acute respiratory tract infections.

  7. Heat shock protein-90-beta facilitates enterovirus 71 viral particles assembly

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Robert Y.L., E-mail: yuwang@mail.cgu.edu.tw [Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333 Taiwan (China); Kuo, Rei-Lin [Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Department of Medical Biotechnology and Laboratory Science and Graduate Program of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Ma, Wei-Chieh [Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333 Taiwan (China); Huang, Hsing-I [Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Department of Medical Biotechnology and Laboratory Science and Graduate Program of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Yu, Jau-Song [Department of Cell and Molecular Biology, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Molecular Medicine Research Center, Chang Gung University, Tao-Yuan 333, Taiwan (China); Yen, Sih-Min [Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333 Taiwan (China); Huang, Chi-Ruei [Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333 Taiwan (China); Shih, Shin-Ru [Research Center for Emerging Viral Infections, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China); Department of Medical Biotechnology and Laboratory Science and Graduate Program of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan 333, Taiwan (China)

    2013-09-01

    Molecular chaperones are reported to be crucial for virus propagation, but are not yet addressed in Human Enterovirus 71 (EV71). Here we describe the specific association of heat shock protein-90-beta (Hsp90β), but not alpha form (Hsp90α), with EV71 viral particles by the co-purification with virions using sucrose density gradient ultracentrifugation, and by the colocalization with viral particles, as assessed by immunogold electron microscopy. The reduction of the Hsp90β protein using RNA interference decreased the correct assembly of viral particles, without affecting EV71 replication levels. Tracking ectopically expressed Hsp90β protein associated with EV71 virions revealed that Hsp90β protein was transmitted to new host cells through its direct association with infectious viral particles. Our findings suggest a new antiviral strategy in which extracellular Hsp90β protein is targeted to decrease the infectivity of EV71 and other enteroviruses, without affecting the broader functions of this constitutively expressed molecular chaperone. - Highlights: • Hsp90β is associated with EV71 virion and is secreted with the release virus. • Hsp90β effects on the correct assembly of viral particles. • Viral titer of cultured medium was reduced in the presence of geldanamycin. • Viral titer was also reduced when Hsp90β was suppressed by siRNA treatment. • The extracellular Hsp90β was also observed in other RNA viruses-infected cells.

  8. Heat shock protein-90-beta facilitates enterovirus 71 viral particles assembly

    International Nuclear Information System (INIS)

    Wang, Robert Y.L.; Kuo, Rei-Lin; Ma, Wei-Chieh; Huang, Hsing-I; Yu, Jau-Song; Yen, Sih-Min; Huang, Chi-Ruei; Shih, Shin-Ru

    2013-01-01

    Molecular chaperones are reported to be crucial for virus propagation, but are not yet addressed in Human Enterovirus 71 (EV71). Here we describe the specific association of heat shock protein-90-beta (Hsp90β), but not alpha form (Hsp90α), with EV71 viral particles by the co-purification with virions using sucrose density gradient ultracentrifugation, and by the colocalization with viral particles, as assessed by immunogold electron microscopy. The reduction of the Hsp90β protein using RNA interference decreased the correct assembly of viral particles, without affecting EV71 replication levels. Tracking ectopically expressed Hsp90β protein associated with EV71 virions revealed that Hsp90β protein was transmitted to new host cells through its direct association with infectious viral particles. Our findings suggest a new antiviral strategy in which extracellular Hsp90β protein is targeted to decrease the infectivity of EV71 and other enteroviruses, without affecting the broader functions of this constitutively expressed molecular chaperone. - Highlights: • Hsp90β is associated with EV71 virion and is secreted with the release virus. • Hsp90β effects on the correct assembly of viral particles. • Viral titer of cultured medium was reduced in the presence of geldanamycin. • Viral titer was also reduced when Hsp90β was suppressed by siRNA treatment. • The extracellular Hsp90β was also observed in other RNA viruses-infected cells

  9. Using rapid diagnostic tests as source of malaria parasite DNA for molecular analyses in the era of declining malaria prevalence

    DEFF Research Database (Denmark)

    Ishengoma, Deus S; Lwitiho, Sudi; Madebe, Rashid A

    2011-01-01

    was conducted to examine if sufficient DNA could be successfully extracted from malaria rapid diagnostic tests (RDTs), used and collected as part of routine case management services in health facilities, and thus forming the basis for molecular analyses, surveillance and quality control (QC) testing of RDTs....... continued molecular surveillance of malaria parasites is important to early identify emerging anti-malarial drug resistance, it is becoming increasingly difficult to obtain parasite samples from ongoing studies, such as routine drug efficacy trials. To explore other sources of parasite DNA, this study...

  10. In Vivo Imaging with Bioluminescent Enterovirus 71 Allows for Real-Time Visualization of Tissue Tropism and Viral Spread.

    Science.gov (United States)

    Caine, Elizabeth A; Osorio, Jorge E

    2017-03-01

    Hand, foot, and mouth disease (HFMD) is a reemerging illness caused by a variety of enteroviruses. The main causative agents are enterovirus 71 (EV71), coxsackievirus A16 (CVA16), and, most recently, coxsackievirus A6 (CVA6). Enterovirus infections can vary from asymptomatic infections to those with a mild fever and blisters on infected individuals' hands, feet, and throats to infections with severe neurological complications. Viral persistence for weeks postinfection (wpi) has also been documented by the demonstration of virus in children's stools. However, little is known about disease progression, viral spread, and tissue tropism of these viruses. These types of studies are limited because many recently developed mouse models mimic the severe neurological complications that occur in a small percentage of enterovirus infections. In the present study, we documented real-time EV71 infection in two different mouse strains by the use of in vivo imaging. Infection of BALB/c mice with a bioluminescent mouse-adapted EV71 construct (mEV71-NLuc) resulted in a lack of clinical signs of disease but in relatively high viral replication, as visualized by luminescence, for 2 wpi. In contrast, mEV71-NLuc infection of AG129 mice (alpha/beta and gamma interferon receptor deficient) showed rapid spread and long-term persistence of the virus in the brain. Interestingly, AG129 mice that survived infection maintained luminescence in the brain for up to 8 wpi. The results we present here will allow future studies on EV71 antiviral drug susceptibility, vaccine efficacy, transmissibility, and pathogenesis. IMPORTANCE We report here that a stable full-length enterovirus 71 (EV71) reporter construct was used to visualize real-time viral spread in AG129 and BALB/c mice. To our knowledge, this is the first report of in vivo imaging of infection with any member of the Picornaviridae family. The nanoluciferase (NLuc) gene, one of the smallest luciferase genes currently available, was shown to

  11. Re-Analysis of Metagenomic Sequences from Acute Flaccidmyelitis Patients Reveals Alternatives to Enterovirus D68 Infection

    Science.gov (United States)

    2015-07-13

    caused in some cases by infection with enterovirus D68. We found that among the patients whose symptoms were previously attributed to enterovirus D68...distribution is unlimited. Re-analysis of metagenomic sequences from acute flaccidmyelitis patients reveals alternatives to enterovirus D68...Street Baltimore, MD 21218 -2685 ABSTRACT Re-analysis of metagenomic sequences from acute flaccidmyelitis patients reveals alternatives to enterovirus

  12. In Vitro Assessment of Combinations of Enterovirus Inhibitors against Enterovirus 71.

    Science.gov (United States)

    Wang, Yizhuo; Li, Guiming; Yuan, Shilin; Gao, Qianqian; Lan, Ke; Altmeyer, Ralf; Zou, Gang

    2016-09-01

    Enterovirus 71 (EV-A71) is a major causative pathogen of hand, foot, and mouth disease (HFMD) epidemics. No antiviral therapies are currently available for treating EV-A71 infections. Here, we selected five reported enterovirus inhibitors (suramin, itraconazole [ITZ], GW5074, rupintrivir, and favipiravir) with different mechanisms of action to test their abilities to inhibit EV-A71 replication alone and in combination. All selected compounds have anti-EV-A71 activities in cell culture. The combination of rupintrivir and ITZ or favipiravir was synergistic, while the combination of rupintrivir and suramin was additive. The combination of suramin and favipiravir exerted a strong synergistic antiviral effect. The observed synergy was not due to cytotoxicity, as there was no significant increase in cytotoxicity when compounds were used in combinations at the tested doses. To investigate the potential inhibitory mechanism of favipiravir against enterovirus, two favipiravir-resistant EV-A71 variants were independently selected, and both of them carried an S121N mutation in the finger subdomain of the 3D polymerase. Reverse engineering of this 3D S121N mutation into an infectious clone of EV-A71 confirmed the resistant phenotype. Moreover, viruses resistant to ITZ or favipiravir remained susceptible to other inhibitors. Most notably, combined with ITZ, rupintrivir prevented the development of ITZ-resistant variants. Taken together, these results provide a rational basis for the design of combination regimens for use in the treatment of EV-A71 infections. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Susceptibilities of enterovirus D68, enterovirus 71, and rhinovirus 87 strains to various antiviral compounds.

    Science.gov (United States)

    Smee, Donald F; Evans, W Joseph; Nicolaou, K C; Tarbet, E Bart; Day, Craig W

    2016-07-01

    Compounds were evaluated for antiviral activity in rhabdomyosarcoma (RD) cells against a recent 2014 clinical isolate of enterovirus D68 (EV-D68), a 1962 strain of EV-68D, rhinovirus 87 (RV-87, serologically the same as EV-D68), and enterovirus 71 (EV-71). Test substances included known-active antipicornavirus agents (enviroxime, guanidine HCl, pirodavir, pleconaril, and rupintrivir), nucleobase/nucleoside analogs (3-deazaguanine and ribavirin), and three novel epidithiodiketopiperazines (KCN-2,2'-epi-19, KCN-19, and KCN-21). Of these, rupintrivir was the most potent, with 50% inhibition of viral cytopathic effect (EC50) and 90% inhibition (EC90) of virus yield at 0.0022-0.0053 μM against EV-D68. Enviroxime, pleconaril and the KCN compounds showed efficacy at 0.01-0.3 μM; 3-deazaguanine and pirodavir inhibited EV-D68 at 7-13 μM, and guanidine HCl and ribavirin were inhibitory at 80-135 μM. Pirodavir was active against EV-71 (EC50 of 0.78 μM) but not against RV-87 or EV-D68, and all other compounds were less effective against EV-71 than against RV-87 and EV-D68. The most promising compound inhibiting both virus infections at low concentrations was rupintrivir. Antiviral activity was confirmed for the ten compounds in virus yield reduction (VYR) assays in RD cells, and for enviroxime, guanidine HCl, and pirodavir by cytopathic effect (CPE) assays in A549, HeLa-Ohio-1, and RD cells. These studies may serve as a basis for further pre-clinical discovery of anti-enterovirus inhibitors. Furthermore, the antiviral profiles and growth characteristics observed herein support the assertion that EV-D68 should be classified together with RV-87. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Design of two molecular methodologies for the rapid identification of Colombian community-acquired methicillin-resistant Staphylococcus aureus isolates

    OpenAIRE

    Escobar, Javier Antonio; Gómez, Ingrid Tatiana; Murillo, Martha Johanna; Castro, Betsy Esperanza; Chavarro, Bibiana; Márquez, Ricaurte Alejandro; Vanegas, Natasha

    2012-01-01

    Introduction. Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infections are found with increasing the frequency, both in healthy individuals in the community and in hospitalized patients. In Colombia and the Andean region, CA-MRSA isolates have a genetic background that is related to the pandemic USA300 clone. Objective. Two molecular methods are designed and standardized for the rapid differentiation of Colombian community-acquired and hospital-acquired methicillin-...

  15. Efficacy of Inactivation of Human Enteroviruses by Multiple ...

    Science.gov (United States)

    Ultraviolet (UV) light has been successfully used for treating a broad suite of pathogens without the concomitant formation of carcinogenic disinfection by-products (DBPs). However, conventional mercury UV lamps have some practical limitations in water treatment applications, such as the inefficiency of energy consumption and more importantly potential mercury contamination upon disposal of the lamps. The recent invention of a novel light-emitting-diodes (LED) device generating germicidal UV wavelengths could eliminate the aforementioned limitations. In this study, we investigated the efficacy of multiple-wavelength UV LEDs for inactivating USEPA contaminant candidate list (CCL) RNA enteroviruses. Of 12 enterovirus species, serotype representatives of the four human enteric species (enterovirus A-D) such as coxsackievirus A10 (CVA10), echovirus 30 (Echo30), poliovirus 1 (PV1), and enterovirus 70 (EV70) respectively were selected as testing RNA viruses. Bench-scale performance evaluation was conducted using a collimated beam (CB) apparatus with LEDs emitting at 260 nm, 280 nm, and the combination of 260|280 nm together, as well as a monochromatic low-pressure (LP) UV lamp at 254 nm for comparison. The CB tests were performed with mixed stocks of four viruses. Infectious virus concentrations were determined using an integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR). The 260 nm LED was most effective at inactivating all enteroviruses teste

  16. Enterovirus and Parechovirus Surveillance - United States, 2014-2016.

    Science.gov (United States)

    Abedi, Glen R; Watson, John T; Nix, W Allan; Oberste, M Steven; Gerber, Susan I

    2018-05-11

    Infections caused by enteroviruses (EV) and parechoviruses (PeV), members of the Picornaviridae family, are associated with various clinical manifestations, including hand, foot, and mouth disease; respiratory illness; myocarditis; meningitis; and sepsis; and can result in death. The genus Enterovirus includes four species of enterovirus (A-D) known to infect humans, and the genus Parechovirus includes one species (A) that infects humans. These species are further divided into types, some of which are associated with specific clinical manifestations. During 2014-2016, a total of 2,967 U.S. cases of EV and PeV infections were reported to the National Enterovirus Surveillance System (NESS). The largest number of reports during that time (2,051) occurred in 2014, when a large nationwide outbreak of enterovirus D68 (EV-D68) occurred, accounting for 68% of cases reported to NESS that year (1). Reports to the National Respiratory and Enteric Virus Surveillance System (NREVSS) during 2014-2016 indicated that circulation of EV peaks annually in the summer and early fall. Because the predominant types of EV and PeV circulating from year to year tend to vary, tracking these trends requires consistent and complete reports from laboratories with the capacity to perform typing.

  17. MicroRNA and Pathogenesis of Enterovirus Infection.

    Science.gov (United States)

    Ho, Bing-Ching; Yang, Pan-Chyr; Yu, Sung-Liang

    2016-01-06

    There are no currently available specific antiviral therapies for non-polio Enterovirus infections. Although several vaccines have entered clinical trials, the efficacy requires further evaluation, particularly for cross-strain protective activity. Curing patients with viral infections is a public health problem due to antigen alterations and drug resistance caused by the high genomic mutation rate. To conquer these limits in the development of anti-Enterovirus treatments, a comprehensive understanding of the interactions between Enterovirus and host cells is urgently needed. MicroRNA (miRNA) constitutes the biggest family of gene regulators in mammalian cells and regulates almost a half of all human genes. The roles of miRNAs in Enterovirus pathogenesis have recently begun to be noted. In this review, we shed light on recent advances in the understanding of Enterovirus infection-modulated miRNAs. The impacts of altered host miRNAs on cellular processes, including immune escape, apoptosis, signal transduction, shutdown of host protein synthesis and viral replication, are discussed. Finally, miRNA-based medication provides a promising strategy for the development of antiviral therapy.

  18. Deferoxamine compensates for decreases in B cell counts and reduces mortality in enterovirus 71-infected mice.

    Science.gov (United States)

    Yang, Yajun; Ma, Jing; Xiu, Jinghui; Bai, Lin; Guan, Feifei; Zhang, Li; Liu, Jiangning; Zhang, Lianfeng

    2014-07-07

    Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore, deferoxamine relieved symptoms and reduced mortality and muscle damage caused by enterovirus 71 infection. This work suggested that deferoxamine has the potential for further development as a B cell-immunomodulator against enterovirus 71.

  19. Clinical Characteristics and Functional Motor Outcomes of Enterovirus 71 Neurological Disease in Children.

    Science.gov (United States)

    Teoh, Hooi-Ling; Mohammad, Shekeeb S; Britton, Philip N; Kandula, Tejaswi; Lorentzos, Michelle S; Booy, Robert; Jones, Cheryl A; Rawlinson, William; Ramachandran, Vidiya; Rodriguez, Michael L; Andrews, P Ian; Dale, Russell C; Farrar, Michelle A; Sampaio, Hugo

    2016-03-01

    . Focal paresis was evident in 23 of 57 (40%) at presentation and was the most common persisting clinical and functional problem at 12 months (observed in 5 of 6 patients), with 1 patient also requiring invasive ventilation. Patients initially seen with acute flaccid paralysis or pulmonary edema had significantly greater frequencies of motor dysfunction at follow-up compared with patients initially seen with other syndromes (odds ratio, 15; 95% CI, 3-79; P Enterovirus 71 may cause serious neurological disease in young patients. The distinct clinicoradiological syndromes, predominantly within the spinal cord and brainstem, enable rapid recognition within evolving outbreaks. Long-term functional neurological morbidity is associated with paresis linked to involvement of gray matter in the brainstem or spinal cord.

  20. Milrinone therapy for enterovirus 71-induced pulmonary edema and/or neurogenic shock in children: a randomized controlled trial.

    Science.gov (United States)

    Chi, Chia-Yu; Khanh, Truong Huu; Thoa, Le Phan Kim; Tseng, Fan-Chen; Wang, Shih-Min; Thinh, Le Quoc; Lin, Chia-Chun; Wu, Han-Chieh; Wang, Jen-Ren; Hung, Nguyen Thanh; Thuong, Tang Chi; Chang, Chung-Ming; Su, Ih-Jen; Liu, Ching-Chuan

    2013-07-01

    Enterovirus 71-induced brainstem encephalitis with pulmonary edema and/or neurogenic shock (stage 3B) is associated with rapid mortality in children. In a small pilot study, we found that milrinone reduced early mortality compared with historical controls. This prospective, randomized control trial was designed to provide more definitive evidence of the ability of milrinone to reduce the 1-week mortality of stage 3B enterovirus 71 infections. Prospective, unicenter, open-label, randomized, controlled study. Inpatient ward of a large tertiary teaching hospital in Ho Chi Minh City, Vietnam. Children (≤ 18 yr old) admitted with proven enterovirus 71-induced pulmonary edema and/or neurogenic shock. Patients were randomly assigned to receive intravenous milrinone (0.5 μg/kg/min) (n = 22) or conventional management (n = 19). Both groups received dopamine or dobutamine and intravenous immunoglobulin. The primary endpoint was 1-week mortality. The secondary endpoints included length of ventilator dependence and hospital stay and adverse events. The median age was 2 years with a predominance of boys in both groups. The 1-week mortality was significantly lower, 18.2% (4/22) in the milrinone compared with 57.9% (11/19) in the conventional management group (relative risk = 0.314 [95% CI, 0.12-0.83], p = 0.01). The median duration of ventilator-free days was longer in the milrinone treatment group (p = 0.01). There was no apparent neurologic sequela in the survivors in either group, and no drug-related adverse events were documented. Milrinone significantly reduced the 1-week mortality of enterovirus 71-induced pulmonary edema and/or neurogenic shock without adverse effects. Further studies are needed to determine whether milrinone might be useful to prevent progression of earlier stages of brainstem encephalitis.

  1. Detection and identification of enteroviruses from various drinking water sources in Taiwan

    Science.gov (United States)

    Hsu, Bing-Mu; Chen, Chien-Hsien; Wan, Min-Tao; Chang, Po-Jen; Fan, Cheng-Wei

    2009-02-01

    SummaryTwenty-three water samples, including seventeen from surface water reservoirs, three from the raw water of groundwater treatment plants, and three from small water systems, were collected in Taiwan and investigated for the presence of, as well as the species of enteroviruses. RT-PCR was used for the detection of enteroviruses. Results revealed that 23.5% of raw water samples from reservoirs were positive for enteroviruses. In addition, one of the three groundwater samples and two of the three small system water samples were positive for enteroviruses. Water samples that were positive for enteroviruses subsequently were evaluated by real-time PCR. The results indicated that enterovirus concentration in groundwater was lower than that in samples obtained from surface water sources. Enteroviruses were identified by nucleic acid sequencing in the 5'-untranslated regions. Three clusters of enteroviruses were identified as coxsackievirus A2, coxsackievirus A6, and enterovirus 71. The presence of enteroviruses indicates the possibility of waterborne transmission of enteroviruses in Taiwan, if water is not adequately treated.

  2. Enterovirus Control of Translation and RNA Granule Stress Responses.

    Science.gov (United States)

    Lloyd, Richard E

    2016-03-30

    Enteroviruses such as poliovirus (PV) and coxsackievirus B3 (CVB3) have evolved several parallel strategies to regulate cellular gene expression and stress responses to ensure efficient expression of the viral genome. Enteroviruses utilize their encoded proteinases to take over the cellular translation apparatus and direct ribosomes to viral mRNAs. In addition, viral proteinases are used to control and repress the two main types of cytoplasmic RNA granules, stress granules (SGs) and processing bodies (P-bodies, PBs), which are stress-responsive dynamic structures involved in repression of gene expression. This review discusses these processes and the current understanding of the underlying mechanisms with respect to enterovirus infections. In addition, the review discusses accumulating data suggesting linkage exists between RNA granule formation and innate immune sensing and activation.

  3. Direct Detection and Identification of Enteroviruses from Faeces of Healthy Nigerian Children Using a Cell-Culture Independent RT-Seminested PCR Assay

    Directory of Open Access Journals (Sweden)

    Temitope Oluwasegun Cephas Faleye

    2016-01-01

    Full Text Available Recently, a cell-culture independent protocol for detection of enteroviruses from clinical specimen was recommended by the WHO for surveillance alongside the previously established protocols. Here, we investigated whether this new protocol will show the same enterovirus diversity landscape as the established cell-culture dependent protocols. Faecal samples were collected from sixty apparently healthy children in Ibadan, Nigeria. Samples were resuspended in phosphate buffered saline, RNA was extracted, and the VP1 gene was amplified using WHO recommended RT-snPCR protocol. Amplicons were sequenced and sequences subjected to phylogenetic analysis. Fifteen (25% of the 60 samples yielded the expected band size. Of the 15 amplicons sequenced, 12 were exploitable. The remaining 3 had electropherograms with multiple peaks and were unexploitable. Eleven of the 12 exploitable sequences were identified as Coxsackievirus A1 (CVA1, CVA3, CVA4, CVA8, CVA20, echovirus 32 (E32, enterovirus 71 (EV71, EVB80, and EVC99. Subsequently, the last exploitable sequence was identified as enterobacteriophage baseplate gene by nucleotide BLAST. The results of this study document the first description of molecular sequence data on CVA1, CVA8, and E32 strains present in Nigeria. The result further showed that species A enteroviruses were more commonly detected in the region when cell-culture bias is bypassed.

  4. Rapid and molecular selective electrochemical sensing of phthalates in aqueous solution

    KAUST Repository

    Zia, Asif I.; Mukhopadhyay, Subhas Chandra; Yu, Paklam; Al-Bahadly, Ibrahim H.; Gooneratne, Chinthaka Pasan; Kosel, Jü rgen

    2015-01-01

    molecular imprinted polymer (MIP) to introduce selectivity for the di(2-ethylhexyl) phthalate (DEHP) molecule. Various concentrations (1-100. ppm) of DEHP in deionized MilliQ water were tested using the functionalized sensing surface to capture the analyte

  5. Bedside immunochromatographic test for enterovirus 71 infection in children.

    Science.gov (United States)

    Huang, Kuan-Ying Arthur; Yang, Shuan; Tsao, Kuo-Chien; Chen, Chih-Jung; Hsieh, Yu-Chia; Chiu, Cheng-Hsun; Hsieh, Juo-Yu; Yang, Jyh-Yuan; Huang, Yhu-Chering

    2013-11-01

    Enterovirus 71 (EV71) causes frequent outbreaks worldwide, particularly in the Asia-Pacific area. Its quick spread is a critical challenge for public health and timely preventive measures and clinical management therefore rely on early detection. There is a need for a rapid, easy-to-use, and reliable method for detecting EV71 infections. The study aimed to evaluate a bedside immunochromatography (ICT) kit for diagnosing acute EV71 infection in children. Pediatric patients with herpangina or hand-foot-mouth disease were randomly and prospectively enrolled from hospitals across Taiwan. Throat or rectal swabs were collected for viral culture and reverse-transcriptase polymerase chain reaction (RTPCR). For the ICT kit, whole blood was obtained by ear piercing, finger-sticking, or venipuncture. The results of ICT, virus isolation and RTPCR in clinical samples were compared. Of the 156 patients enrolled, 91 (58%), 64 (41%) and 72 (46%) had positive results of the ICT kit, viral culture and RTPCR, respectively. Laboratory-confirmed infection with either positive EV71 culture or RTPCR was used as the diagnostic standard. The sensitivity and specificity of the ICT kit was 84% and 77%, respectively. The viral culture and RTPCR had relatively lower sensitivity but higher specificity. The patient's age did not affect the performance of the ICT, viral culture and RTPCR. However, a low sensitivity of ICT kit was noted before the second day of disease onset. The ICT kit may serve as a simple, quick and reliable method for the bedside diagnosis of acute EV71 infection in children. Copyright © 2013 Elsevier B.V. All rights reserved.

  6. Outbreak of Enterovirus - 71 Meningitis in Calicut

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    CK Sasidharan

    2016-04-01

    Full Text Available Objective: Enterovirus 71(EV 71 causes wide spectrum of infections ranging from asymptomatic conditions to clinical syndromes like diarrhea, rash, hand-foot-and mouth disease (HFMD, herpangina, aseptic meningitis, encephalitis, myocarditis, acute flaccid paralysis, bulbar and brainstem encephalitis Guillain Barre syndrome, pulmonary haemorrhage. This study deals with an outbreak of aseptic meningitis in children caused by EV 71 virus. Methods: The authors report an outbreak of aseptic meningitis in children in and around Calicut in June 2008. Clinical and laboratory study was done in collaboration with National Centre for Disease Control, New Delhi. 149 children with aseptic meningitis were studied and followed up from June 2008 to May 2009. Result: All children had clinical features suggestive of aseptic meningitis and serology showed the rising antibody titre against EV 71 virus infection. CSF analysis also showed four fold rise in antibodies in one and ≥ 1:2 neutralising antibodies titer against EV- 71 in four samples indicating meningitis due to EV-71. Conclusion: EV 71 was identified as the causative agent of the outbreak of aseptic meningitis in the study and the fact that the EV 71 infection has evolved from minor illness like HFMD to major illness like aseptic meningitis from the same locality is truly alarming.

  7. Replication and Inhibitors of Enteroviruses and Parechoviruses

    Directory of Open Access Journals (Sweden)

    Lonneke van der Linden

    2015-08-01

    Full Text Available The Enterovirus (EV and Parechovirus genera of the picornavirus family include many important human pathogens, including poliovirus, rhinovirus, EV-A71, EV-D68, and human parechoviruses (HPeV. They cause a wide variety of diseases, ranging from a simple common cold to life-threatening diseases such as encephalitis and myocarditis. At the moment, no antiviral therapy is available against these viruses and it is not feasible to develop vaccines against all EVs and HPeVs due to the great number of serotypes. Therefore, a lot of effort is being invested in the development of antiviral drugs. Both viral proteins and host proteins essential for virus replication can be used as targets for virus inhibitors. As such, a good understanding of the complex process of virus replication is pivotal in the design of antiviral strategies goes hand in hand with a good understanding of the complex process of virus replication. In this review, we will give an overview of the current state of knowledge of EV and HPeV replication and how this can be inhibited by small-molecule inhibitors.

  8. Enterovirus-71 genotype C isolated in Peru between 2006 and 2009.

    Science.gov (United States)

    Huaman, Jose L; Carrion, Gladys; Ampuero, Julia S; Ocaña, Victor; Laguna-Torres, V Alberto; Hontz, Robert D

    2016-12-01

    Enterovirus-71 (EV71) was first isolated in California, United States in 1969, belongs to the genus Enterovirus, family Picornaviridae. Although infection normally causes mild, often undiagnosed illness, it can cause central nervous system infections that could turn fatal. Based on VP1 gene analysis, EV71 has been classified into six separate genotypes. Although the molecular epidemiology of EV71 has been well described via studies originating from Asia and Europe, it is mostly unknown in South America. From our study, four EV71 isolates from Peru were characterized using phylogenetic methods to determine their relationship with known reference strains. These four Peruvian EV71 isolates from between 2006 and 2009 were analyzed by RT-PCR using primers capable of amplifying the entire VP1 gene. Reference strains representing all six known genotypes were used to determine any recognizable phylogenetic relationships. In fact, all of our isolates clustered together within the genotype C1 lineage- separate from Asian, European, North American, and Australian strains. We present evidence that EV71 genotype C1 exists in Peru, and this is the first such report documenting EV71 genotype C1 circulating in South America. Gathering additional isolates will help elucidate a more complete global epidemiological picture of EV71 infections. Published by Elsevier B.V.

  9. Divergent Requirement for a DNA Repair Enzyme during Enterovirus Infections.

    Science.gov (United States)

    Maciejewski, Sonia; Nguyen, Joseph H C; Gómez-Herreros, Fernando; Cortés-Ledesma, Felipe; Caldecott, Keith W; Semler, Bert L

    2015-12-29

    Viruses of the Enterovirus genus of picornaviruses, including poliovirus, coxsackievirus B3 (CVB3), and human rhinovirus, commandeer the functions of host cell proteins to aid in the replication of their small viral genomic RNAs during infection. One of these host proteins is a cellular DNA repair enzyme known as 5' tyrosyl-DNA phosphodiesterase 2 (TDP2). TDP2 was previously demonstrated to mediate the cleavage of a unique covalent linkage between a viral protein (VPg) and the 5' end of picornavirus RNAs. Although VPg is absent from actively translating poliovirus mRNAs, the removal of VPg is not required for the in vitro translation and replication of the RNA. However, TDP2 appears to be excluded from replication and encapsidation sites during peak times of poliovirus infection of HeLa cells, suggesting a role for TDP2 during the viral replication cycle. Using a mouse embryonic fibroblast cell line lacking TDP2, we found that TDP2 is differentially required among enteroviruses. Our single-cycle viral growth analysis shows that CVB3 replication has a greater dependency on TDP2 than does poliovirus or human rhinovirus replication. During infection, CVB3 protein accumulation is undetectable (by Western blot analysis) in the absence of TDP2, whereas poliovirus protein accumulation is reduced but still detectable. Using an infectious CVB3 RNA with a reporter, CVB3 RNA could still be replicated in the absence of TDP2 following transfection, albeit at reduced levels. Overall, these results indicate that TDP2 potentiates viral replication during enterovirus infections of cultured cells, making TDP2 a potential target for antiviral development for picornavirus infections. Picornaviruses are one of the most prevalent groups of viruses that infect humans and livestock worldwide. These viruses include the human pathogens belonging to the Enterovirus genus, such as poliovirus, coxsackievirus B3 (CVB3), and human rhinovirus. Diseases caused by enteroviruses pose a major problem

  10. Rapid molecular sexing of three-spined sticklebacks, Gasterosteus aculeatus L., based on large Y-chromosomal insertions.

    Science.gov (United States)

    Bakker, Theo C M; Giger, Thomas; Frommen, Joachim G; Largiadèr, Carlo R

    2017-08-01

    There is a need for rapid and reliable molecular sexing of three-spined sticklebacks, Gasterosteus aculeatus, the supermodel species for evolutionary biology. A DNA region at the 5' end of the sex-linked microsatellite Gac4202 was sequenced for the X chromosome of six females and the Y chromosome of five males from three populations. The Y chromosome contained two large insertions, which did not recombine with the phenotype of sex in a cross of 322 individuals. Genetic variation (SNPs and indels) within the insertions was smaller than on flanking DNA sequences. Three molecular PCR-based sex tests were developed, in which the first, the second or both insertions were covered. In five European populations (from DE, CH, NL, GB) of three-spined sticklebacks, tests with both insertions combined showed two clearly separated bands on agarose minigels in males and one band in females. The tests with the separate insertions gave similar results. Thus, the new molecular sexing method gave rapid and reliable results for sexing three-spined sticklebacks and is an improvement and/or alternative to existing methods.

  11. A rapid PCR-based approach for molecular identification of filamentous fungi.

    Science.gov (United States)

    Chen, Yuanyuan; Prior, Bernard A; Shi, Guiyang; Wang, Zhengxiang

    2011-08-01

    In this study, a novel rapid and efficient DNA extraction method based on alkaline lysis, which can deal with a large number of filamentous fungal isolates in the same batch, was established. The filamentous fungal genomic DNA required only 20 min to prepare and can be directly used as a template for PCR amplification. The amplified internal transcribed spacer regions were easy to identify by analysis. The extracted DNA also can be used to amplify other protein-coding genes for fungal identification. This method can be used for rapid systematic identification of filamentous fungal isolates.

  12. Enterovirus no polio de casos con parálisis residual. Colombia, 1992-1995

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    Dioselina Peláez

    1999-06-01

    Full Text Available Las parálisis fláccidas agudas (PFA tienen una amplia variedad de orígenes y de agentes causales: físicos, fisiopatológicos, tóxicos e infecciosos. Entre estos últimos, el virus salvaje (silvestre de la poliomielitis y el enterovirus 71 (EV71, parecen ser los agentes virales más frecuentes. Habiendo eliminado el poliovirus salvaje autóctono como agente causal de enfermedad paralítica en Colombia desde junio de 1991 y teniendo aislamientos de virus no polio en el 20,8% del total de casos de PFA notificados anualmente, quisimos conocer el papel que juegan los enterovirus en la incidencia de parálisis fláccida aguda y la dinámica de circulación y distribución de los mismos en Colombia. Se revisó la base de datos de los casos notificados al Programa de Erradicación de la Poliomielitis en Colombia entre el 1"" de enero de 1992 y el 31 de diciembre de 1995, al cual se notificaron 856 casos sospechosos de niños menores de 15 años, y se escogieron 69 casos para el estudio pero se recuperaron 57 aislamientos virales por reinoculación en células RD y Hep-2C. Estos se identificaron mediante neutralización con mezclas de antisueros de Lim & Benyesh-Melnick (LBM.Todos ellos fueron sometidos a caracterización molecular mediante reacción en cadena de la polimerasa -PCR-, utilizando iniciadores complementarios a la regiónVP1 del genoma viral, seguida del análisis de secuencia de nucleótidos de los fragmentos amplificados por PCR. La identificación final del serotipo se realizó por comparación de nucleótidos en auto assemblery el análisis descriptivo de los datos. No se estableció circulación mayor de ningún serotipo específico en región geográfica alguna del país. Tampoco hubo asociación causal a ninguna patología característica con ninguno de los enterovirus aislados. Se describe el hallazgo de EV71 en un caso de PFA con diagnóstico clínico de síndrome de Guillain-Barré. La descripción de 22 serotipos diferentes

  13. Enterovirus Migration Patterns between France and Tunisia.

    Science.gov (United States)

    Othman, Ines; Mirand, Audrey; Slama, Ichrak; Mastouri, Maha; Peigue-Lafeuille, Hélène; Aouni, Mahjoub; Bailly, Jean-Luc

    2015-01-01

    The enterovirus (EV) types echovirus (E-) 5, E-9, and E-18, and coxsackievirus (CV-) A9 are infrequently reported in human diseases and their epidemiologic features are poorly defined. Virus transmission patterns between countries have been estimated with phylogenetic data derived from the 1D/VP1 and 3CD gene sequences of a sample of 74 strains obtained in France (2000-2012) and Tunisia (2011-2013) and from the publicly available sequences. The EV types (E-5, E-9, and E-18) exhibited a lower worldwide genetic diversity (respective number of genogroups: 4, 5, and 3) in comparison to CV-A9 (n = 10). The phylogenetic trees estimated with both 1D/VP1 and 3CD sequence data showed variations in the number of co-circulating lineages over the last 20 years among the four EV types. Despite the low number of genogroups in E-18, the virus exhibited the highest number of recombinant 3CD lineages (n = 10) versus 4 (E-5) to 8 (E-9). The phylogenies provided evidence of multiple transportation events between France and Tunisia involving E-5, E-9, E-18, and CV-A9 strains. Virus spread events between France and 17 other countries in five continents had high probabilities of occurrence as those between Tunisia and two European countries other than France. All transportation events were supported by BF values > 10. Inferring the source of virus transmission from phylogenetic data may provide insights into the patterns of sporadic and epidemic diseases caused by EVs.

  14. Epidemiology of Enterovirus 71 Infections in Taiwan

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    Min-Yuan Chia

    2014-08-01

    Full Text Available Enterovirus 71 (EV71 was first described in USA in 1969 but retrospective studies in The Netherlands further detected EV71 in the clinical specimens collected in 1963. EV71 has one single serotype measured by using hyperimmune animal antisera but can be phylogenetically classified into three genogroups (A, B, and C including 11 genotypes (A, B1–B5, C1–C5. In Taiwan, EV71 caused a large-scale nationwide epidemic in 1998. Retrospective studies further detected EV71 in clinical specimens collected from hand–foot–mouth disease patients in 1980 and 1986. Therefore, EV71 may have circulated in Taiwan prior to 1980. Since 1998, EV71 has cyclically caused nationwide epidemics with different predominant genotypes in 1998 (genotype C2, 2000–2001 (B4, 2005 (C4, 2008 (B5, and 2012 (B5. Phylogenetic analysis revealed that C4 viruses isolated in 2005 were probably from China, B5 viruses isolated in 2008 were probably from South Eastern Asia, and B5 viruses isolated in 2012 were probably from Xiamen, China. Several studies have collected postinfection sera from children to measure cross-reactive neutralizing antibody titers against different EV71 genotypes and found that antigenic differences between genogroup B and C viruses did not have a clear pattern but that genotype A virus was antigenically different from genogroup B and C viruses. In conclusion, EV71 cyclically caused nationwide epidemics through international importations. EV71 surveillance in Taiwan should combine genetic and serological methods.

  15. Anti-Enterovirus 71 Agents of Natural Products

    Directory of Open Access Journals (Sweden)

    Liyan Wang

    2015-09-01

    Full Text Available This review, with 42 references, presents the fascinating area of anti-enterovirus 71 natural products over the last three decades for the first time. It covers literature published from 2005–2015 and refers to compounds isolated from biogenic sources. In total, 58 naturally-occurring anti-EV71 compounds are recorded.

  16. Detection of enteroviruses from urban sewage in Parma.

    Science.gov (United States)

    Cesari, C; Colucci, M E; Veronesi, L; Giordano, R; Paganuzzi, F; Affanni, P; Bracchi, M T; Capobianco, E; Ferrari, G; Tanzi, Maria Luisa

    2010-12-01

    The human enteroviruses (EV) are the most common and widespread human viruses in the world. They have bowel as their natural habitat and they can spread in the environment through the faecal excretion. In the continental climate Regions these viruses may cause epidemic outbreaks in summer and fall, while in the tropical Regions the EV infections present a high incidence during all year. The symptoms can be minor or subclinic, but they can be also associated to rare and serious diseases. The aim of this study was to evaluate the environmental circulation of polioviruses and non-polio enteroviruses (NPEV) using standard methods of urban wastewater surveillance recommended by the WHO. A total of 188 wastewater samples were collected between February 2005 and December 2008 from two sewage treatment plants in Parma. The sampling was carried out twice a month. Environmental variables were collected for each day of sampling. Out of the 188 examined wastewater samples, 78.7% were positive to the enterovirus research. One out of the 148 positive samples was identified as poliovirus Sabin-like type 3. The remaining 147 positive samples were enteroviruses non polio: Coxsakieviruses and Echoviruses. All Coxsakieviruses isolated were of type B. The proposed method has shown high sensibility, also in presence of very low expected prevalence of vaccine poliovirus. It allows to verify the kind and relative frequency of enteric viruses circulating in the country, whose characteristics (virulence and pathogenicity) may vary with reference to a different epidemiologic and demographic structure of the resident population.

  17. Severe neonatal parechovirus infection and similarity with enterovirus infection

    NARCIS (Netherlands)

    Verboon-Maciolek, Malgorzata A.; Krediet, Tannette G.; Gerards, Leo J.; de Vries, Linda S.; Groenendaal, Floris; van Loon, Anton M.

    Background: Enteroviruses (EV) are an important cause of neonatal disease including hepatitis, meningoencephalitis, and myocarditis that can lead to death or severe long-term sequelae. Less is known about severe neonatal infection caused by the parechoviruses (PeV) of which type 1 (PeV1) and type 2

  18. Seroepidemiology of Human Enterovirus 71 Infection among Children, Cambodia

    Science.gov (United States)

    Horwood, Paul F.; Andronico, Alessio; Tarantola, Arnaud; Salje, Henrik; Duong, Veasna; Mey, Channa; Ly, Sovann; Dussart, Philippe; Cauchemez, Simon

    2016-01-01

    Enterovirus 71 is reported to have emerged in Cambodia in 2012; at least 54 children with severe encephalitis died during that outbreak. We used serum samples collected during 2000–2011 to show that the virus had been widespread in the country for at least a decade before the 2012 outbreak. PMID:26690000

  19. Is enterovirus 71-vaccinatie ook nodig in Nederland?

    NARCIS (Netherlands)

    Wolthers, Katja C.; de Jong, Menno D.

    2014-01-01

    Enterovirus 71 (EV71) is an emerging infection, causing large outbreaks in Asia with high morbidity and mortality in children. New vaccines against EV71 have been developed and tested in China, leading to two recent publications on the efficacy of these vaccines in The New England Journal of

  20. Anti-Enterovirus 71 Agents of Natural Products.

    Science.gov (United States)

    Wang, Liyan; Wang, Junfeng; Wang, Lishu; Ma, Shurong; Liu, Yonghong

    2015-09-09

    This review, with 42 references, presents the fascinating area of anti-enterovirus 71 natural products over the last three decades for the first time. It covers literature published from 2005-2015 and refers to compounds isolated from biogenic sources. In total, 58 naturally-occurring anti-EV71 compounds are recorded.

  1. Synergistic antiviral activity of gemcitabine and ribavirin against enteroviruses

    NARCIS (Netherlands)

    Kang, Hyunju; Kim, Chonsaeng; Kim, Dong-Eun; Song, Jae-Hyoung; Choi, Miri; Choi, Kwangman; Kang, Mingu; Lee, Kyungjin; Kim, Hae Soo; Shin, Jin Soo; Kim, Janghwan; Han, Sang-Bae; Lee, Mi-Young; Lee, Su Ui; Lee, Chong-Kyo; Kim, Meehyein; Ko, Hyun-Jeong; van Kuppeveld, Frank J M; Cho, Sungchan

    2015-01-01

    Enteroviruses are major causative agents of various human diseases, and some of them are currently considered to be an enormous threat to public health. However, no effective therapy is currently available for the treatment of these infections. We identified gemcitabine, a nucleoside-analog drug

  2. A Naturally Occurring Recombinant Enterovirus Expresses a Torovirus Deubiquitinase.

    Science.gov (United States)

    Shang, Pengcheng; Misra, Saurav; Hause, Ben; Fang, Ying

    2017-07-15

    Enteroviruses (EVs) are implicated in a wide range of diseases in humans and animals. In this study, a novel enterovirus (enterovirus species G [EVG]) (EVG 08/NC_USA/2015) was isolated from a diagnostic sample from a neonatal pig diarrhea case and identified by using metagenomics and complete genome sequencing. The viral genome shares 75.4% nucleotide identity with a prototypic EVG strain (PEV9 UKG/410/73). Remarkably, a 582-nucleotide insertion, flanked by 3C pro cleavage sites at the 5' and 3' ends, was found in the 2C/3A junction region of the viral genome. This insertion encodes a predicted protease with 54 to 68% amino acid identity to torovirus (ToV) papain-like protease (PLP) (ToV-PLP). Structural homology modeling predicts that this protease adopts a fold and a catalytic site characteristic of minimal PLP catalytic domains. This structure is similar to those of core catalytic domains of the foot-and-mouth disease virus leader protease and coronavirus PLPs, which act as deubiquitinating and deISGylating (interferon [IFN]-stimulated gene 15 [ISG15]-removing) enzymes on host cell substrates. Importantly, the recombinant ToV-PLP protein derived from this novel enterovirus also showed strong deubiquitination and deISGylation activities and demonstrated the ability to suppress IFN-β expression. Using reverse genetics, we generated a ToV-PLP knockout recombinant virus. Compared to the wild-type virus, the ToV-PLP knockout mutant virus showed impaired growth and induced higher expression levels of innate immune genes in infected cells. These results suggest that ToV-PLP functions as an innate immune antagonist; enterovirus G may therefore gain fitness through the acquisition of ToV-PLP from a recombination event. IMPORTANCE Enteroviruses comprise a highly diversified group of viruses. Genetic recombination has been considered a driving force for viral evolution; however, recombination between viruses from two different orders is a rare event. In this study, we

  3. Neurologic Manifestations of Enterovirus 71 Infection in Korea.

    Science.gov (United States)

    Lee, Kyung Yeon; Lee, Myoung Sook; Kim, Dong Bin

    2016-04-01

    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.

  4. Rapid molecular assays for the detection of yellow fever virus in low-resource settings.

    Science.gov (United States)

    Escadafal, Camille; Faye, Oumar; Sall, Amadou Alpha; Faye, Ousmane; Weidmann, Manfred; Strohmeier, Oliver; von Stetten, Felix; Drexler, Josef; Eberhard, Michael; Niedrig, Matthias; Patel, Pranav

    2014-03-01

    Yellow fever (YF) is an acute viral hemorrhagic disease transmitted by Aedes mosquitoes. The causative agent, the yellow fever virus (YFV), is found in tropical and subtropical areas of South America and Africa. Although a vaccine is available since the 1930s, YF still causes thousands of deaths and several outbreaks have recently occurred in Africa. Therefore, rapid and reliable diagnostic methods easy to perform in low-resources settings could have a major impact on early detection of outbreaks and implementation of appropriate response strategies such as vaccination and/or vector control. The aim of this study was to develop a YFV nucleic acid detection method applicable in outbreak investigations and surveillance studies in low-resource and field settings. The method should be simple, robust, rapid and reliable. Therefore, we adopted an isothermal approach and developed a recombinase polymerase amplification (RPA) assay which can be performed with a small portable instrument and easy-to-use lyophilized reagents. The assay was developed in three different formats (real-time with or without microfluidic semi-automated system and lateral-flow assay) to evaluate their application for different purposes. Analytical specificity and sensitivity were evaluated with a wide panel of viruses and serial dilutions of YFV RNA. Mosquito pools and spiked human plasma samples were also tested for assay validation. Finally, real-time RPA in portable format was tested under field conditions in Senegal. The assay was able to detect 20 different YFV strains and demonstrated no cross-reactions with closely related viruses. The RPA assay proved to be a robust, portable method with a low detection limit (<21 genome equivalent copies per reaction) and rapid processing time (<20 min). Results from real-time RPA field testing were comparable to results obtained in the laboratory, thus confirming our method is suitable for YFV detection in low-resource settings.

  5. The SUCCESS model for laboratory performance and execution of rapid molecular diagnostics in patients with sepsis.

    Science.gov (United States)

    Dekmezian, Mhair; Beal, Stacy G; Damashek, Mary Jane; Benavides, Raul; Dhiman, Neelam

    2015-04-01

    Successful performance and execution of rapid diagnostics in a clinical laboratory hinges heavily on careful validation, accurate and timely communication of results, and real-time quality monitoring. Laboratories must develop strategies to integrate diagnostics with stewardship and evidence-based clinical practice guidelines. We present a collaborative SUCCESS model for execution and monitoring of rapid sepsis diagnostics to facilitate timely treatment. Six months after execution of the Verigene Gram-Positive Blood Culture (BC-GP) and the AdvanDx PNA-FISH assays, data were collected on 579 and 28 episodes of bacteremia and fungemia, respectively. Clinical testing was executed using a SUCCESS model comprising the following components: stewardship, utilization of resources, core strategies, concierge services, education, support, and surveillance. Stewardship needs were identified by evaluating the specialty services benefiting from new testing. Utilization of resources was optimized by reviewing current treatment strategies and antibiogram and formulary options. Core strategies consisted of input from infectious disease leadership, pharmacy, and laboratory staff. Concierge services included automated Micro-eUpdate and physician-friendly actionable reports. Education modules were user-specific, and support was provided through a dedicated 24/7 microbiology hotline. Surveillance was performed by daily audit by the director. Using the SUCCESS model, the turnaround time for the detailed report with actionable guidelines to the physician was ∼3 hours from the time of culture positivity. The overall correlation between rapid methods and culture was 94% (546/579). Discrepant results were predominantly contaminants such as a coagulase-negative staphylococci or viridans streptococci in mixed cultures. SUCCESS is a cost-effective and easily adaptable model for clinical laboratories with limited stewardship resources.

  6. Rapid molecular assays for the detection of yellow fever virus in low-resource settings.

    Directory of Open Access Journals (Sweden)

    Camille Escadafal

    2014-03-01

    Full Text Available BACKGROUND: Yellow fever (YF is an acute viral hemorrhagic disease transmitted by Aedes mosquitoes. The causative agent, the yellow fever virus (YFV, is found in tropical and subtropical areas of South America and Africa. Although a vaccine is available since the 1930s, YF still causes thousands of deaths and several outbreaks have recently occurred in Africa. Therefore, rapid and reliable diagnostic methods easy to perform in low-resources settings could have a major impact on early detection of outbreaks and implementation of appropriate response strategies such as vaccination and/or vector control. METHODOLOGY: The aim of this study was to develop a YFV nucleic acid detection method applicable in outbreak investigations and surveillance studies in low-resource and field settings. The method should be simple, robust, rapid and reliable. Therefore, we adopted an isothermal approach and developed a recombinase polymerase amplification (RPA assay which can be performed with a small portable instrument and easy-to-use lyophilized reagents. The assay was developed in three different formats (real-time with or without microfluidic semi-automated system and lateral-flow assay to evaluate their application for different purposes. Analytical specificity and sensitivity were evaluated with a wide panel of viruses and serial dilutions of YFV RNA. Mosquito pools and spiked human plasma samples were also tested for assay validation. Finally, real-time RPA in portable format was tested under field conditions in Senegal. CONCLUSION/SIGNIFICANCE: The assay was able to detect 20 different YFV strains and demonstrated no cross-reactions with closely related viruses. The RPA assay proved to be a robust, portable method with a low detection limit (<21 genome equivalent copies per reaction and rapid processing time (<20 min. Results from real-time RPA field testing were comparable to results obtained in the laboratory, thus confirming our method is suitable for

  7. Rapid and molecular selective electrochemical sensing of phthalates in aqueous solution

    KAUST Repository

    Zia, Asif I.

    2015-05-01

    Reported research work presents real time non-invasive detection of phthalates in spiked aqueous samples by employing electrochemical impedance spectroscopy (EIS) technique incorporating a novel interdigital capacitive sensor with multiple sensing thin film gold micro-electrodes fabricated on native silicon dioxide layer grown on semiconducting single crystal silicon wafer. The sensing surface was functionalized by a self-assembled monolayer of 3-aminopropyltrietoxysilane (APTES) with embedded molecular imprinted polymer (MIP) to introduce selectivity for the di(2-ethylhexyl) phthalate (DEHP) molecule. Various concentrations (1-100. ppm) of DEHP in deionized MilliQ water were tested using the functionalized sensing surface to capture the analyte. Frequency response analyzer (FRA) algorithm was used to obtain impedance spectra so as to determine sample conductance and capacitance for evaluation of phthalate concentration in the sample solution. Spectrum analysis algorithm interpreted the experimentally obtained impedance spectra by applying complex nonlinear least square (CNLS) curve fitting in order to obtain electrochemical equivalent circuit and corresponding circuit parameters describing the kinetics of the electrochemical cell. Principal component analysis was applied to deduce the effects of surface immobilized molecular imprinted polymer layer on the evaluated circuit parameters and its electrical response. The results obtained by the testing system were validated using commercially available high performance liquid chromatography diode array detector system.

  8. Association of Tic Disorders and Enterovirus Infection

    Science.gov (United States)

    Tsai, Ching-Shu; Yang, Yao-Hsu; Huang, Kuo-You; Lee, Yena; McIntyre, Roger S.; Chen, Vincent Chin-Hung

    2016-01-01

    Abstract There has been growing interest in the association between infectious disease and mental disorders, but an association between enterovirus (EV) infection and tic disorders has not been sufficiently explored. Herein, we aim to investigate the association between EV infection and incidence of tic disorders in a nationwide population-based sample using Taiwan's National Health Insurance Research Database. We identified individuals aged ≤18 years prior to 2005 with an inpatient diagnosis of EV infection and/or history of EV infection. Tic disorder was operationalized using International Classification of Disease, Revision 9, Clinical Modification (ICD-9-CM) codes 307.20–307.23. A total of 47,998 individuals with history of EV infection were compared to 47,998 sex-, age-, and urbanization-matched controls on incidence of tic disorders. The mean ± standard deviation follow-up period for all subjects was 9.7 ± 3.6 years; the mean latency period between initial EV infection and incident diagnosis of tic disorder diagnosis was 5.4 ± 2.8 years. EV infection was significantly associated with greater incidence of tic disorders (hazard ratio [HR] = 1.24, 95% CI: 1.07–1.45). When subgrouped on the basis of central nervous system (CNS) involvement, EV infection with CNS involvement was not significantly associated with greater incidence of tic disorders when compared to controls (HR = 1.25, 95% CI: 0.64–2.43); EV infection without CNS involvement was significantly associated greater incidence of tic disorders when compared to controls (HR = 1.24, 95% CI: 1.07–1.45). In addition, hospitalization for an EV infection did not increase the hazard for greater incidence of tic disorders (HR = 1.32, 95% CI: 1.04–1.67 with hospitalization and 1.22, 95% CI: 1.04–1.44 without hospitalization). EV infection is temporally associated with incidence of tic disorders. Our observations add to the growing body of literature implicating immune

  9. Detection of enteroviruses and hepatitis a virus in water by consensus primer multiplex RT-PCR

    Science.gov (United States)

    Li, Jun-Wen; Wang, Xin-Wei; Yuan, Chang-Qing; Zheng, Jin-Lai; Jin, Min; Song, Nong; Shi, Xiu-Quan; Chao, Fu-Huan

    2002-01-01

    AIM: To develop a rapid detection method of enteroviruses and Hepatitis A virus (HAV). METHODS: A one-step, single-tube consensus primers multiplex RT-PCR was developed to simultaneously detect Poliovirus, Coxsackie virus, Echovirus and HAV. A general upstream primer and a HAV primer and four different sets of primers (5 primers) specific for Poliovirus, Coxsacki evirus, Echovirus and HAV cDNA were mixed in the PCR mixture to reverse transcript and amplify the target DNA. Four distinct amplified DNA segments representing Poliovirus, Coxsackie virus, Echovirus and HAV were identified by gel electrophoresis as 589-, 671-, 1084-, and 1128 bp sequences, respectively. Semi-nested PCR was used to confirm the amplified products for each enterovirus and HAV. RESULTS: All four kinds of viral genome RNA were detected, and producing four bands which could be differentiated by the band size on the gel. To confirm the specificity of the multiplex PCR products, semi-nested PCR was performed. For all the four strains tested gave positive results. The detection sensitivity of multiplex PCR was similar to that of monoplex RT-PCR which was 24 PFU for Poliovrus, 21 PFU for Coxsackie virus, 60 PFU for Echovirus and 105 TCID50 for HAV. The minimum amount of enteric viral RNA detected by semi-nested PCR was equivalent to 2.4 PFU for Poliovrus, 2.1 PFU for Coxsackie virus, 6.0 PFU for Echovirus and 10.5 TCID50 for HAV. CONCLUSION: The consensus primers multiplex RT-PCR has more advantages over monoplex RT-PCR for enteric viruses detection, namely, the rapid turnaround time and cost effectiveness. PMID:12174381

  10. Establishment of a panel of in-house polyclonal antibodies for the diagnosis of enterovirus infections.

    Science.gov (United States)

    Kotani, Osamu; Iwata-Yoshikawa, Naoko; Suzuki, Tadaki; Sato, Yuko; Nakajima, Noriko; Koike, Satoshi; Iwasaki, Takuya; Sata, Tetsutaro; Yamashita, Teruo; Minagawa, Hiroko; Taguchi, Fumihiro; Hasegawa, Hideki; Shimizu, Hiroyuki; Nagata, Noriyo

    2015-04-01

    The aim of this study was to establish a reliable method of virus detection for the diagnosis of critical enterovirus infections such as acute infective encephalitis, encephalomyelitis and myocarditis. Because histopathological and immunohistochemical analyses of paraffin-embedded tissues play an important role in recognizing infectious agents in tissue samples, six in-house polyclonal antibodies raised against three representative enteroviruses using an indirect immunofluorescence assay and immunohistochemistry were examined. This panel of polyclonal antibodies recognized three serotypes of enterovirus. Two of the polyclonal antibodies were raised against denatured virus particles from enterovirus A71, one was raised against the recombinant VP1 protein of coxsackievirus B3, and the other for poliovirus type 1 were raised against denatured virus particles, the recombinant VP1 protein and peptide 2C. Western blot analysis revealed that each of these antibodies recognized the corresponding viral antigen and none cross-reacted with non-enteroviruses within the family Picornaviridae. However, all cross-reacted to some extent with the antigens derived from other serotypes of enterovirus. Indirect immunofluorescence assay and immunohistochemistry revealed that the virus capsid and non-structural proteins were localized in the cytoplasm of affected culture cells, and skeletal muscles and neurons in neonatal mice experimentally-infected with human enterovirus. The antibodies also recognized antigens derived from recent clinical isolates of enterovirus A71, coxsackievirus B3 and poliovirus. In addition, immunohistochemistry revealed that representative antibodies tested showed the same recognition pattern according to each serotype. Thus, the panel of in-house anti-enterovirus polyclonal antibodies described herein will be an important tool for the screening and pathological diagnosis for enterovirus infections, and may be useful for the classification of different

  11. Defining the Enterovirus Diversity Landscape of a Fecal Sample: A Methodological Challenge?

    Science.gov (United States)

    Faleye, Temitope Oluwasegun Cephas; Adewumi, Moses Olubusuyi; Adeniji, Johnson Adekunle

    2016-01-12

    Enteroviruses are a group of over 250 naked icosahedral virus serotypes that have been associated with clinical conditions that range from intrauterine enterovirus transmission withfataloutcome through encephalitis and meningitis, to paralysis. Classically, enterovirus detection was done by assaying for the development of the classic enterovirus-specific cytopathic effect in cell culture. Subsequently, the isolates were historically identified by a neutralization assay. More recently, identification has been done by reverse transcriptase-polymerase chain reaction (RT-PCR). However, in recent times, there is a move towards direct detection and identification of enteroviruses from clinical samples using the cell culture-independent RT semi-nested PCR (RT-snPCR) assay. This RT-snPCR procedure amplifies the VP1 gene, which is then sequenced and used for identification. However, while cell culture-based strategies tend to show a preponderance of certain enterovirus species depending on the cell lines included in the isolation protocol, the RT-snPCR strategies tilt in a different direction. Consequently, it is becoming apparent that the diversity observed in certain enterovirus species, e.g., enterovirus species B(EV-B), might not be because they are the most evolutionarily successful. Rather, it might stem from cell line-specific bias accumulated over several years of use of the cell culture-dependent isolation protocols. Furthermore, it might also be a reflection of the impact of the relative genome concentration on the result of pan-enterovirus VP1 RT-snPCR screens used during the identification of cell culture isolates. This review highlights the impact of these two processes on the current diversity landscape of enteroviruses and the need to re-assess enterovirus detection and identification algorithms in a bid to better balance our understanding of the enterovirus diversity landscape.

  12. Deferoxamine Compensates for Decreases in B Cell Counts and Reduces Mortality in Enterovirus 71-Infected Mice

    OpenAIRE

    Yang, Yajun; Ma, Jing; Xiu, Jinghui; Bai, Lin; Guan, Feifei; Zhang, Li; Liu, Jiangning; Zhang, Lianfeng

    2014-01-01

    Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore...

  13. Molecular phylogeny of Pompilinae (Hymenoptera: Pompilidae): Evidence for rapid diversification and host shifts in spider wasps.

    Science.gov (United States)

    Rodriguez, Juanita; Pitts, James P; Florez, Jaime A; Bond, Jason E; von Dohlen, Carol D

    2016-01-01

    Pompilinae is one of the largest subfamilies of spider wasps (Pompilidae). Most pompilines are generalist spider predators at the family level, but some taxa exhibit ecological specificity (i.e., to spider-host guild). Here we present the first molecular phylogenetic analysis of Pompilinae, toward the aim of evaluating the monophyly of tribes and genera. We further test whether changes in the rate of diversification are associated with host-guild shifts. Molecular data were collected from five nuclear loci (28S, EF1-F2, LWRh, Wg, Pol2) for 76 taxa in 39 genera. Data were analyzed using maximum likelihood (ML) and Bayesian inference (BI). The phylogenetic results were compared with previous hypotheses of subfamilial and tribal classification, as well as generic relationships in the subfamily. The classification of Pompilus and Agenioideus is also discussed. A Bayesian relaxed molecular clock analysis was used to examine divergence times. Diversification rate-shift tests accounted for taxon-sampling bias using ML and BI approaches. Ancestral host family and host guild were reconstructed using MP and ML methods. Ancestral host guild for all Pompilinae, for the ancestor at the node where a diversification rate-shift was detected, and two more nodes back in time was inferred using BI. In the resulting phylogenies, Aporini was the only previously proposed monophyletic tribe. Several genera (e.g., Pompilus, Microphadnus and Schistonyx) are also not monophyletic. Dating analyses produced a well-supported chronogram consistent with topologies from BI and ML results. The BI ancestral host-use reconstruction inferred the use of spiders belonging to the guild "other hunters" (frequenting the ground and vegetation) as the ancestral state for Pompilinae. This guild had the highest probability for the ML reconstruction and was equivocal for the MP reconstruction; various switching events to other guilds occurred throughout the evolution of the group. The diversification of

  14. Rapid and Accurate Molecular Identification of the Emerging Multidrug-Resistant Pathogen Candida auris.

    Science.gov (United States)

    Kordalewska, Milena; Zhao, Yanan; Lockhart, Shawn R; Chowdhary, Anuradha; Berrio, Indira; Perlin, David S

    2017-08-01

    Candida auris is an emerging multidrug-resistant fungal pathogen causing nosocomial and invasive infections associated with high mortality. C. auris is commonly misidentified as several different yeast species by commercially available phenotypic identification platforms. Thus, there is an urgent need for a reliable diagnostic method. In this paper, we present fast, robust, easy-to-perform and interpret PCR and real-time PCR assays to identify C. auris and related species: Candida duobushaemulonii , Candida haemulonii , and Candida lusitaniae Targeting rDNA region nucleotide sequences, primers specific for C. auris only or C. auris and related species were designed. A panel of 140 clinical fungal isolates was used in both PCR and real-time PCR assays followed by electrophoresis or melting temperature analysis, respectively. The identification results from the assays were 100% concordant with DNA sequencing results. These molecular assays overcome the deficiencies of existing phenotypic tests to identify C. auris and related species. Copyright © 2017 Kordalewska et al.

  15. OpenMM 7: Rapid development of high performance algorithms for molecular dynamics.

    Directory of Open Access Journals (Sweden)

    Peter Eastman

    2017-07-01

    Full Text Available OpenMM is a molecular dynamics simulation toolkit with a unique focus on extensibility. It allows users to easily add new features, including forces with novel functional forms, new integration algorithms, and new simulation protocols. Those features automatically work on all supported hardware types (including both CPUs and GPUs and perform well on all of them. In many cases they require minimal coding, just a mathematical description of the desired function. They also require no modification to OpenMM itself and can be distributed independently of OpenMM. This makes it an ideal tool for researchers developing new simulation methods, and also allows those new methods to be immediately available to the larger community.

  16. BPIFB6 Regulates Secretory Pathway Trafficking and Enterovirus Replication.

    Science.gov (United States)

    Morosky, Stefanie; Lennemann, Nicholas J; Coyne, Carolyn B

    2016-05-15

    Bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 3 (BPIFB3) is an endoplasmic reticulum (ER)-localized host factor that negatively regulates coxsackievirus B (CVB) replication through its control of the autophagic pathway. Here, we show that another member of the BPIFB family, BPIFB6, functions as a positive regulator of CVB, and other enterovirus, replication by controlling secretory pathway trafficking and Golgi complex morphology. We show that similar to BPIFB3, BPIFB6 localizes exclusively to the ER, where it associates with other members of the BPIFB family. However, in contrast to our findings that RNA interference (RNAi)-mediated silencing of BPIFB3 greatly enhances CVB replication, we show that silencing of BPIFB6 expression dramatically suppresses enterovirus replication in a pan-viral manner. Mechanistically, we show that loss of BPIFB6 expression induces pronounced alterations in retrograde and anterograde trafficking, which correlate with dramatic fragmentation of the Golgi complex. Taken together, these data implicate BPIFB6 as a key regulator of secretory pathway trafficking and viral replication and suggest that members of the BPIFB family participate in diverse host cell functions to regulate virus infections. Enterovirus infections are associated with a number of severe pathologies, such as aseptic meningitis, dilated cardiomyopathy, type I diabetes, paralysis, and even death. These viruses, which include coxsackievirus B (CVB), poliovirus (PV), and enterovirus 71 (EV71), co-opt the host cell secretory pathway, which controls the transport of proteins from the endoplasmic reticulum to the Golgi complex, to facilitate their replication. Here we report on the identification of a novel regulator of the secretory pathway, bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 6 (BPIFB6), whose expression is required for enterovirus replication. We show that loss of BPIFB6 expression

  17. Epidemiology and seroepidemiology of human enterovirus 71 among Thai populations

    Science.gov (United States)

    2014-01-01

    Background Human enterovirus 71 (EV71) is an important pathogen caused large outbreaks in Asian-Pacific region with severe neurological complications and may lead to death in young children. Understanding of the etiological spectrum and epidemic changes of enterovirus and population’s immunity against EV71 are crucial for the implementation of future therapeutic and prophylactic intervention. Results A total of 1,182 patients who presented with the symptoms of hand foot and mouth disease (67.3%) or herpangina (HA) (16.7%) and admitted to the hospitals during 2008-2013 were tested for enterovirus using pan-enterovirus PCR targeting 5′-untranslated region and specific PCR for viral capsid protein 1 gene. Overall, 59.7% were pan-enterovirus positive comprising 9.1% EV71 and 31.2% coxsackievirus species A (CV-A) including 70.5% CV-A6, 27.6% CV-A16, 1.1% CV-A10, and 0.8% CV-A5. HFMD and HA occurred endemically during 2008-2011. The number of cases increased dramatically in June 2012 with the percentage of the recently emerged CV-A6 significantly rose to 28.4%. Co-circulation between different EV71 genotypes was observed during the outbreak. Total of 161 sera obtained from healthy individuals were tested for neutralizing antibodies (NAb) against EV71 subgenotype B5 (EV71-B5) using microneutralization assay. The seropositive rate of EV71-B5 was 65.8%. The age-adjusted seroprevalence for individuals was found to be lowest in children aged >6 months to 2 years (42.5%). The seropositive rate remained relatively low in preschool children aged > 2 years to 6 years (48.3%) and thereafter increased sharply to more than 80% in individuals aged > 6 years. Conclusions This study describes longitudinal data reflecting changing patterns of enterovirus prevalence over 6 years and demonstrates high seroprevalences of EV71-B5 NAb among Thai individuals. The rate of EV71 seropositive increased with age but without gender-specific significant difference. We identified

  18. BPIFB6 Regulates Secretory Pathway Trafficking and Enterovirus Replication

    Science.gov (United States)

    Morosky, Stefanie; Lennemann, Nicholas J.

    2016-01-01

    ABSTRACT Bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 3 (BPIFB3) is an endoplasmic reticulum (ER)-localized host factor that negatively regulates coxsackievirus B (CVB) replication through its control of the autophagic pathway. Here, we show that another member of the BPIFB family, BPIFB6, functions as a positive regulator of CVB, and other enterovirus, replication by controlling secretory pathway trafficking and Golgi complex morphology. We show that similar to BPIFB3, BPIFB6 localizes exclusively to the ER, where it associates with other members of the BPIFB family. However, in contrast to our findings that RNA interference (RNAi)-mediated silencing of BPIFB3 greatly enhances CVB replication, we show that silencing of BPIFB6 expression dramatically suppresses enterovirus replication in a pan-viral manner. Mechanistically, we show that loss of BPIFB6 expression induces pronounced alterations in retrograde and anterograde trafficking, which correlate with dramatic fragmentation of the Golgi complex. Taken together, these data implicate BPIFB6 as a key regulator of secretory pathway trafficking and viral replication and suggest that members of the BPIFB family participate in diverse host cell functions to regulate virus infections. IMPORTANCE Enterovirus infections are associated with a number of severe pathologies, such as aseptic meningitis, dilated cardiomyopathy, type I diabetes, paralysis, and even death. These viruses, which include coxsackievirus B (CVB), poliovirus (PV), and enterovirus 71 (EV71), co-opt the host cell secretory pathway, which controls the transport of proteins from the endoplasmic reticulum to the Golgi complex, to facilitate their replication. Here we report on the identification of a novel regulator of the secretory pathway, bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 6 (BPIFB6), whose expression is required for enterovirus replication. We show that loss of

  19. A novel, broad-spectrum inhibitor of enterovirus replication that targets host cell factor phosphatidylinositol 4-kinase IIIβ

    NARCIS (Netherlands)

    van der Schaar, H.M.; Leyssen, Pieter; Thibaut, H.J.; de Palma, Armando; van der Linden, Lonneke; Lanke, Kjerstin H.W.; Lacroix, Céline; Verbeken, Erik; Conrath, Katja; Macleod, Angus M; Mitchell, Dale R; Palmer, Nicholas J; van de Poël, Hervé; Andrews, Martin; Neyts, Johan; van Kuppeveld, F.J.M.

    2013-01-01

    Despite their high clinical and socioeconomic impacts, there is currently no approved antiviral therapy for the prophylaxis or treatment of enterovirus infections. Here we report on a novel inhibitor of enterovirus replication, compound 1,

  20. Evaluation of PCR electrospray-ionization mass spectrometry for rapid molecular diagnosis of bovine mastitis.

    Science.gov (United States)

    Perreten, Vincent; Endimiani, Andrea; Thomann, Andreas; Wipf, Juliette R K; Rossano, Alexandra; Bodmer, Michèle; Raemy, Andreas; Sannes-Lowery, Kristin A; Ecker, David J; Sampath, Rangarajan; Bonomo, Robert A; Washington, Cicely

    2013-06-01

    Bovine mastitis, an inflammatory disease of the mammary gland, is one of the most costly diseases affecting the dairy industry. The treatment and prevention of this disease is linked heavily to the use of antibiotics in agriculture and early detection of the primary pathogen is essential to control the disease. Milk samples (n=67) from cows suffering from mastitis were analyzed for the presence of pathogens using PCR electrospray-ionization mass spectrometry (PCR/ESI-MS) and were compared with standard culture diagnostic methods. Concurrent identification of the primary mastitis pathogens was obtained for 64% of the tested milk samples, whereas divergent results were obtained for 27% of the samples. The PCR/ESI-MS failed to identify some of the primary pathogens in 18% of the samples, but identified other pathogens as well as microorganisms in samples that were negative by culture. The PCR/ESI-MS identified bacteria to the species level as well as yeasts and molds in samples that contained a mixed bacterial culture (9%). The sensitivity of the PCR/ESI-MS for the most common pathogens ranged from 57.1 to 100% and the specificity ranged from 69.8 to 100% using culture as gold standard. The PCR/ESI-MS also revealed the presence of the methicillin-resistant gene mecA in 16.2% of the milk samples, which correlated with the simultaneous detection of staphylococci including Staphylococcus aureus. We demonstrated that PCR/ESI-MS, a more rapid diagnostic platform compared with bacterial culture, has the significant potential to serve as an important screening method in the diagnosis of bovine clinical mastitis and has the capacity to be used in infection control programs for both subclinical and clinical disease. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  1. Rapid molecular detection of rifampicin resistance facilitates early diagnosis and treatment of multi-drug resistant tuberculosis: case control study.

    Directory of Open Access Journals (Sweden)

    Philly O'Riordan

    2008-09-01

    Full Text Available Multi-drug resistant tuberculosis (MDR-TB is a major public health concern since diagnosis is often delayed, increasing the risk of spread to the community and health care workers. Treatment is prolonged, and the total cost of treating a single case is high. Diagnosis has traditionally relied upon clinical suspicion, based on risk factors and culture with sensitivity testing, a process that can take weeks or months. Rapid diagnostic molecular techniques have the potential to shorten the time to commencing appropriate therapy, but have not been put to the test under field conditions.This retrospective case-control study aimed to identify risk factors for MDR-TB, and analyse the impact of testing for rifampicin resistance using RNA polymerase B (rpoB mutations as a surrogate for MDR-TB. Forty two MDR-TB cases and 84 fully sensitive TB controls were matched by date of diagnosis; and factors including demographics, clinical presentation, microbiology findings, management and outcome were analysed using their medical records. Conventionally recognised risk factors for MDR-TB were absent in almost half (43% of the cases, and 15% of cases were asymptomatic. A significant number of MDR-TB cases were identified in new entrants to the country. Using rpoB mutation testing, the time to diagnosis of MDR-TB was dramatically shortened by a median of 6 weeks, allowing patients to be commenced on appropriate therapy a median of 51days earlier than those diagnosed by conventional culture and sensitivity testing.MDR-TB is frequently an unexpected finding, may be asymptomatic, and is particularly prevalent among TB infected new entrants to the country. Molecular resistance testing of all acid fast bacilli positive specimens has the potential to rapidly identify MDR-TB patients and commence them on appropriate therapy significantly earlier than by conventional methods.

  2. Rapid, efficient and selective preconcentration of benzo[a]pyrene (BaP) by molecularly imprinted composite cartridge and HPLC

    Energy Technology Data Exchange (ETDEWEB)

    Çorman, Mehmet Emin, E-mail: mecorman@sinop.edu.tr [Hacettepe University, Department of Chemistry, Ankara (Turkey); Sinop University, Department of Bioengineering, Sinop (Turkey); Armutcu, Canan [Hacettepe University, Department of Chemistry, Ankara (Turkey); Uzun, Lokman, E-mail: lokman@hacettepe.edu.tr [Hacettepe University, Department of Chemistry, Ankara (Turkey); Denizli, Adil [Hacettepe University, Department of Chemistry, Ankara (Turkey)

    2017-01-01

    In this study, cryogel-based molecularly imprinted composite cartridges were designed for the rapid, efficient, and selective preconcentration of benzo[a]pyrene (BaP) from water samples. First, a BaP-imprinted poly(2-hydroxyethyl methacrylate-N-methacryloyl-(L)-phenylalanine) composite cartridge was synthesized under semi-frozen conditions and characterized by scanning electron microscopy, elemental analysis, Fourier transform infrared spectroscopy, and swelling tests. After the optimization of preconcentration parameters, i.e., pH and initial BaP concentration, the selectivity and preconcentration efficiency, and reusability of these cartridges were also evaluated. In selectivity experiments, BaP imprinted composite cartridge exhibited binding capacities 3.09, 9.52, 8.87, and 8.77-fold higher than that of the non-imprinted composite cartridge in the presence of competitors, such as benzo[b]fluoranthene (BbF), benzo[k]fluoranthene (BkF), indeno[1,2,3-cd]pyrene (IcdP), and 1-naphthol, respectively. The method detection limit (MDL), relative standard deviation (RSD) and preconcentration efficiency (PE) of the synthesized composite cartridge were calculated as 24.86 μg/L, 1.60%, and 349.6%, respectively. - Highlights: • Cryogel based molecularly imprinted composite cartridges as solid-phase extraction sorbents • Combination unique structural features of cryogels with MIP • An excellent ability to recognize the BaP molecule even if single-run contact • Rapid, efficient, selective and cost-friendly PAH preconcentration • Hydrophobic interactions via N-methacryloyl-(L)-phenylalanine.

  3. Reciprocal Regulation between Enterovirus 71 and the NLRP3 Inflammasome

    Directory of Open Access Journals (Sweden)

    Hongbin Wang

    2015-07-01

    Full Text Available Enterovirus 71 (EV71 is the major etiological agent of hand, foot, and mouth disease (HFMD. Early studies showed that EV71-infected patients with severe complications exhibited elevated plasma levels of IL-1β, indicating that EV71 may activate inflammasomes. Our current study demonstrates that the NLRP3 inflammasome plays a protective role against EV71 infection of mice in vivo. EV71 replication in myeloid cells results in the activation of the NLRP3 inflammasome and secretion of IL-1β. Conversely, EV71 counteracts inflammasome activation through cleavage of NLRP3 by viral proteases 2A and 3C, which cleave NLRP3 protein at the G493-L494 or Q225-G226 junction, respectively. Moreover, EV71 3C interacts with NLRP3 and inhibits IL-1β secretion when expressed in mammalian cells. These results thus reveal a set of reciprocal regulations between enterovirus 71 and the NLRP3 inflammasome.

  4. Infecciones del sistema nervioso central por enterovirus en niños atendidos en un hospital de Lima, Perú Enteroviral central nervous system infections in children treated at a hospital in Lima, Peru

    Directory of Open Access Journals (Sweden)

    Iván O Espinoza

    2011-12-01

    presence of enterovirus, herpes simplex virus 1 (HSV-1, herpes simplex virus 2 (HSV-2 and varicella-zoster virus (VZV by polymerase chain reaction (PCR. Results. 97 patients were included, out of which 69 % had acute encephalitis and 31 % acute meningitis. Enteroviruses were identified in 52,6% of all acute non-bacterial central nervous system infections; corresponding to 83,3 % of meningitis and 38,8 % of encephalitis. There were no cases of infection due to HSV-1, HSV-2 or VZV. Enterovirus infections reached 82,9 % in the warm months (November-January and 28,6 % in the colder months (May-July. Conclusions. Enteroviruses are the principal etiologic agents in acute aseptic meningitis and encephalitis in pediatric patients in Lima, Peru. Enteroviruses have a seasonal epidemiological pattern with a clear increase in the number of cases during the summer months. It is useful to have this rapid diagnostic method available as an aid in the management of acute central nervous system infections.

  5. Structure, Immunogenicity, and Protective Mechanism of an Engineered Enterovirus 71-Like Particle Vaccine Mimicking 80S Empty Capsid.

    Science.gov (United States)

    Wang, Xiaoli; Ku, Zhiqiang; Zhang, Xiang; Ye, Xiaohua; Chen, Jinhuan; Liu, Qingwei; Zhang, Wei; Zhang, Chao; Fu, Zhenglin; Jin, Xia; Cong, Yao; Huang, Zhong

    2018-01-01

    Enterovirus 71 (EV71) is the major causative agent of severe hand, foot, and mouth disease, which affects millions of young children in the Asia-Pacific region annually. In this study, we engineered a novel EV71 virus-like particle (VLP) that lacks VP4 (therefore designated VLP ΔVP4 ) and investigated its structure, antigenicity, and vaccine potential. The cryo-electron microscopy (cryo-EM) structure of VLP ΔVP4 was reconstructed to 3.71-Å resolution. Results from structural and biochemical analyses revealed that VLP ΔVP4 resembles the end product of the viral uncoating process, the 80S empty capsid. VLP ΔVP4 is able to elicit high-titer neutralizing antibodies and to fully protect mice against lethal viral challenge. Mechanistic studies showed that, at the cellular level, the anti-VLP ΔVP4 sera exert neutralization effects at both pre- and postattachment stages by inhibiting both virus attachment and internalization, and at the molecular level, the antisera can block multiple interactions between EV71 and its key receptors. Our study gives a better understanding of EV71 capsid assembly and provides important information for the design and development of new-generation vaccines for EV71, and perhaps for other enteroviruses, as well. IMPORTANCE Enterovirus 71 (EV71) infection may lead to severe hand, foot, and mouth disease, with significant morbidity and mortality. Knowledge regarding EV71 particle assembly remains limited. Here, we report the generation and characterization of a novel EV71 virus-like particle that lacks the VP4 capsid subunit protein. This particle, termed VLP ΔVP4 , structurally mimics the 80S empty capsid, which is the end stage of EV71 uncoating. We further show that VLP ΔVP4 exhibits desirable immunogenicity and protective efficacy in proof-of-concept studies. In addition, the inhibitory mechanisms of the VLP ΔVP4 -induced antibodies are unraveled at both the cellular and molecular levels. Our work provides the first evidence of

  6. Reemergence of enterovirus 71 epidemic in northern Taiwan, 2012.

    Science.gov (United States)

    Luo, Shu-Ting; Chiang, Pai-Shan; Chung, Wan-Yu; Chia, Min-Yuan; Tsao, Kuo-Chien; Wang, Ying-Hsiang; Lin, Tzou-Yien; Lee, Min-Shi

    2015-01-01

    Enterovirus 71 (EV71) belongs to picornavirus family and could be classified phylogenetically into three major genogroups (A, B and C) including 11 genotypes (A, B1-B5 and C1-C5). Since 1997, EV71 has caused large-scale of epidemics with neurological complications in Asian children. In Taiwan, nationwide EV71 epidemics with different predominant genotypes have occurred cyclically since 1998. A nationwide EV71 epidemic occurred again in 2012. We conducted genetic and antigenic characterizations of the 2012 epidemic. Chang Gung Memorial Hospital (CGMH) is a medical center in northern Taiwan. In CGMH, specimens were collected from pediatric inpatients with suspected enterovirus infections for virus isolation. Enterovirus isolates were serotyped and genotyped and sera from EV71 inpatients were collected for measuring neutralizing antibody titers. There were 10, 16 and 99 EV71 inpatients identified in 2010, 2011 and 2012, respectively. There were 82 EV71 isolates genotyped, which identified 17 genotype C4a viruses and 65 genotype B5 viruses. The genotype B5 viruses were not detected until November 2011 and caused epidemics in 2012. Interestingly, the B5-2011 viruses were genetically distinguishable from the B5 viruses causing the 2008 epidemic and are likely introduced from China or Southeastern Asia. Based on antigenic analysis, minor antigenic variations were detected among the B5-2008, B5-2011, C4a-2008 and C4a-2012 viruses but these viruses antigenically differed from genotype A. Genotype B5 and C4a viruses antigenically differ from genotype A viruses which have disappeared globally for 30 years but have been detected in China since 2008. Enterovirus surveillance should monitor genetic and antigenic variations of EV71.

  7. Enteroviruses, hygiene and type 1 diabetes: toward a preventive vaccine.

    Science.gov (United States)

    Drescher, Kristen M; von Herrath, Matthias; Tracy, Steven

    2015-01-01

    Enteroviruses and humans have long co-existed. Although recognized in ancient times, poliomyelitis and type 1 diabetes (T1D) were exceptionally rare and not epidemic, due in large part to poor sanitation and personal hygiene which resulted in repeated exposure to fecal-oral transmitted viruses and other infectious agents and viruses and the generation of a broad protective immunity. As a function of a growing acceptance of the benefits of hygienic practices and microbiologically clean(er) water supplies, the likelihood of exposure to diverse infectious agents and viruses declined. The effort to vaccinate against poliomyelitis demonstrated that enteroviral diseases are preventable by vaccination and led to understanding how to successfully attenuate enteroviruses. Type 1 diabetes onset has been convincingly linked to infection by numerous enteroviruses including the group B coxsackieviruses (CVB), while studies of CVB infections in NOD mice have demonstrated not only a clear link between disease onset but an ability to reduce the incidence of T1D as well: CVB infections can suppress naturally occurring autoimmune T1D. We propose here that if we can harness and develop the capacity to use attenuated enteroviral strains to induce regulatory T cell populations in the host through vaccination, then a vaccine could be considered that should function to protect against both autoimmune as well as virus-triggered T1D. Such a vaccine would not only specifically protect from certain enterovirus types but more importantly, also reset the organism's regulatory rheostat making the further development of pathogenic autoimmunity less likely. Copyright © 2014 John Wiley & Sons, Ltd.

  8. Enterovirus and parechovirus infection in children: a brief overview.

    Science.gov (United States)

    de Crom, S C M; Rossen, J W A; van Furth, A M; Obihara, C C

    2016-08-01

    Enterovirus and parechovirus are a frequent cause of infection in children. This review is an overview of what is known from enterovirus and parechovirus infection in children and contains information about the epidemiology, pathogenesis, clinical presentation, diagnosis, treatment, and prognosis of enterovirus and parechovirus infection in children. EV and HPeV infections are a frequent cause of infection in childhood. The clinical presentation is diverse. RT-qPCR is the best way to detect an EV or HPeV. Cerebrospinal fluid, blood and feces have the highest sensitivity for detecting an EV or HPeV. There is no treatment for EV and HPeV infections. Two vaccines against EV 71 are just licensed in China and will be available on the private market. Little is known about the prognosis of EV and HPeV infections. •EV and HPeV are a frequent cause of infection in children. What is new: •This review gives a brief overview over EV and HPeV infection in children.

  9. A highly conserved amino acid in VP1 regulates maturation of enterovirus 71.

    Directory of Open Access Journals (Sweden)

    Yong-Xin Zhang

    2017-09-01

    Full Text Available Enterovirus 71 (EV71 is the major causative agent of hand, foot and mouth disease (HFMD in children, causing severe clinical outcomes and even death. Here, we report an important role of the highly conserved alanine residue at position 107 in the capsid protein VP1 (VP1A107 in the efficient replication of EV71. Substitutional mutations of VP1A107 significantly diminish viral growth kinetics without significant effect on viral entry, expression of viral genes and viral production. The results of mechanistic studies reveal that VP1A107 regulates the efficient cleavage of the VP0 precursor during EV71 assembly, which is required, in the next round of infection, for the transformation of the mature virion (160S into an intermediate or A-particle (135S, a key step of virus uncoating. Furthermore, the results of molecular dynamic simulations and hydrogen-bond networks analysis of VP1A107 suggest that flexibility of the VP1 BC loop or the region surrounding the VP1107 residue directly correlates with viral infectivity. It is possible that sufficient flexibility of the region surrounding the VP1107 residue favors VP0 conformational change that is required for the efficient cleavage of VP0 as well as subsequent viral uncoating and viral replication. Taken together, our data reveal the structural role of the highly conserved VP1A107 in regulating EV71 maturation. Characterization of this novel determinant of EV71 virulence would promote the study on pathogenesis of Enteroviruses.

  10. A neonatal murine model for evaluation of enterovirus E HY12 virus infection and pathogenicity.

    Directory of Open Access Journals (Sweden)

    Xiaochun Gai

    Full Text Available HY12 viruses are enteroviruses recently isolated from cattle characterized by severe respiratory and digestive disease with high morbidity and mortality in China. While the viruses exhibit unique biological and molecular characters distinct from known enterovirus E, the pathogenicity and viral pathogenesis remains largely unknown.Neonatal mice of Balb/C, ICR, and Kunming strain are infected with HY12 to determine the susceptible mouse strain. The minimal infection dose, the virus infection routes, the pathogenicity and tissue tropism for HY12 were determined by infecting susceptible mice with HY12 viruses, and confirmed by different approaches including virus isolation and recovery, virus detection, histopathology, and immunohistochemistry.A murine model for HY12 infection was successfully established and employed to investigate the pathogenicity of HY12 viruses. ICR mouse strain is the most susceptible strain for HY12 infection with a minimal infective dose as 2×106TCID50/mouse. HY12 viruses have the capability of infecting ICR suckling mice via all infection routes including intranasal administration, oral administration, intraperitoneal injection, subcutaneous injection, and intramuscular injection, which are confirmed by the isolation and recovery of viruses from HY12-infected mice; detection of viruses by RT-PCR; observations of pathological lesions and inflammatory cell infiltrations in the intestine, lung, liver, and brain; uncovering of HY12 virus antigens in majority of tissues, especially in intestine, lung, and infected brain of mice by immunohistochemistry assay.A neonatal murine model for HY12 infection is successfully established for determining the susceptible mouse strain, the minimal infective dose, the infection route, the viral pathogenicity and the tropism of HY12, thus providing an invaluable model system for elucidating the pathogenesis of HY12 viruses and the elicited immunity.

  11. Polio and Measles Down the Drain: Environmental Enterovirus Surveillance in the Netherlands, 2005 to 2015.

    NARCIS (Netherlands)

    Benschop, Kimberley S M; van der Avoort, Harrie G; Jusic, Edin; Vennema, Harry; van Binnendijk, Rob; Duizer, Erwin

    2017-01-01

    Polioviruses (PVs) are members of the genus Enterovirus In the Netherlands, the exclusion of PV circulation is based on clinical enterovirus (EV) surveillance (CEVS) of EV-positive cases and routine environmental EV surveillance (EEVS) conducted on sewage samples collected in the region of the

  12. Continued seasonal circulation of enterovirus D68 in the Netherlands, 2011-2014.

    NARCIS (Netherlands)

    Meijer, A.; Benschop, K.S.; Donker, G.A.; Avoort, H.G. van der

    2014-01-01

    Enterovirus D68 (EV-D68) continued to circulate in a seasonal pattern in the Netherlands, after the outbreak in 2010. Outpatient EV-D68 cases, mainly in the under 20 and 50–59 years age groups, presented with relatively mild respiratory disease. Hospital-based enterovirus surveillance identified

  13. Retrospective Surveillance of Wastewater To Examine Seasonal Dynamics of Enterovirus Infections

    Science.gov (United States)

    Enteroviruses are RNA viruses that are responsible for both mild gastroenteritis and mild respiratory illnesses as well as debilitating diseases such as meningitis and myocarditis. The disease burden of enteroviruses in the United States is difficult to assess because most infect...

  14. Impact of Enterovirus Testing on Resource Use in Febrile Young Infants: A Systematic Review.

    Science.gov (United States)

    Wallace, Sowdhamini S; Lopez, Michelle A; Caviness, A Chantal

    2017-02-01

    Enterovirus infection commonly causes fever in infants aged 0 to 90 days and, without testing, is difficult to differentiate from serious bacterial infection. To determine the cost savings of routine enterovirus testing and identify subgroups of infants with greater potential impact from testing among infants 0 to 90 days old with fever. Studies were identified systematically from published and unpublished literature by using Embase, Medline, the Cochrane database, and conference proceedings. Inclusion criteria were original studies, in any language, of enterovirus infection including the outcomes of interest in infants aged 0 to 90 days. Standardized instruments were used to appraise each study. The evidence quality was evaluated using Grading of Recommendations Assessment, Development, and Evaluation criteria. Two investigators independently searched the literature, screened and critically appraised the studies, extracted the data, and applied the Grading of Recommendations Assessment, Development, and Evaluation criteria. Of the 257 unique studies identified and screened, 32 were completely reviewed and 8 were included. Routine enterovirus testing was associated with reduced hospital length of stay and cost savings during peak enterovirus season. Cerebrospinal fluid pleocytosis was a poor predictor of enterovirus meningitis. The studies were all observational and the evidence was of low quality. Enterovirus polymerase chain reaction testing, independent of cerebrospinal fluid pleocytosis, can reduce length of stay and achieve cost savings, especially during times of high enterovirus prevalence. Additional study is needed to identify subgroups that may achieve greater cost savings from testing to additionally enhance the efficiency of testing. Copyright © 2017 by the American Academy of Pediatrics.

  15. Deferoxamine Compensates for Decreases in B Cell Counts and Reduces Mortality in Enterovirus 71-Infected Mice

    Directory of Open Access Journals (Sweden)

    Yajun Yang

    2014-07-01

    Full Text Available Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore, deferoxamine relieved symptoms and reduced mortality and muscle damage caused by enterovirus 71 infection. This work suggested that deferoxamine has the potential for further development as a B cell-immunomodulator against enterovirus 71.

  16. Viral kinetics of Enterovirus 71 in human abdomyosarcoma cells

    Science.gov (United States)

    Lu, Jing; He, Ya-Qing; Yi, Li-Na; Zan, Hong; Kung, Hsiang-Fu; He, Ming-Liang

    2011-01-01

    AIM: To characterise the viral kinetics of enterovirus 71 (EV71). METHODS: In this study, human rhabdomyosarcoma (RD) cells were infected with EV71 at different multiplicity of infection (MOI). After infection, the cytopathic effect (CPE) was monitored and recorded using a phase contrast microscope associated with a CCD camera at different time points post viral infection (0, 6, 12, 24 h post infection). Cell growth and viability were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in both EV71 infected and mock infected cells at each time point. EV71 replication kinetics in RD cells was determined by measuring the total intracellular viral RNA with real-time reverse-transcription polymerase chain reaction (qRT-PCR). Also, the intracellular and extracellular virion RNA was isolated and quantified at different time points to analyze the viral package and secretion. The expression of viral protein was determined by analyze the levels of viral structure protein VP1 with Western blotting. RESULTS: EV71 infection induced a significant CPE as early as 6 h post infection (p.i.) in both RD cells infected with high ratio of virus (MOI 10) and low ratio of virus (MOI 1). In EV71 infected cells, the cell growth was inhibited and the number of viable cells was rapidly decreased in the later phase of infection. EV71 virions were uncoated immediately after entry. The intracellular viral RNA began to increase at as early as 3 h p.i. and the exponential increase was found between 3 h to 6 h p.i. in both infected groups. For viral structure protein synthesis, results from western-blot showed that intracellular viral protein VP1 could not be detected until 6 h p.i. in the cells infected at either MOI 1 or MOI 10; and reached the peak at 9 h p.i. in the cells infected with EV71 at both MOI 1 and MOI 10. Simultaneously, the viral package and secretion were also actively processed as the virus underwent rapid replication. The viral package kinetics

  17. Frequency of isolation of polioviruses and non polio enteroviruses from patients with acute flaccid paralysis, enterovirus infection and children from groups at risk

    Directory of Open Access Journals (Sweden)

    N. I. Romanenkova

    2012-01-01

    Full Text Available The article describes the frequency of isolation of polioviruses and non polio enteroviruses from different categories of the investigated children. The percentage of detection of polioviruses from the patients with acute flaccid paralysis was lower than that from the children from groups at risk. Among the patients with the enterovirus infection the polioviruses were rarely revealed. The frequency of isolation of non polio enteroviruses from these patients was significantly higher than that from the other categories of investigated persons. The improvement of poliomyelitis surveillance and the reinforcement of virological surveillance of children from groups at risk and those with enterovirus infection will provide the important data for Global Polio Eradication Initiative and the maintenance of polio free status of the Russian Federation.

  18. Exploration of the anti-enterovirus activity of a series of pleconaril/pirodavir-like compounds.

    Science.gov (United States)

    Bernard, Angela; Lacroix, Céline; Cabiddu, Maria G; Neyts, Johan; Leyssen, Pieter; Pompei, Raffaello

    2015-04-01

    The Enterovirus genus of the Picornaviridae is represented by several viral pathogens that are associated with human disease, namely Poliovirus 1, Enterovirus 71 and Rhinoviruses. Enterovirus 71 has been associated with encephalitis, while Rhinoviruses are a major cause of asthma exacerbations and chronic obstructive pulmonary disease. Based on the structure of both pleconaril and pirodavir, we previously synthesized some original compounds as potential inhibitors of Rhinovirus replication. These compounds were explored for in vitro antiviral potential on other human pathogenic Enteroviruses, namely Enterovirus 71 on rhabdo-myosarcoma cells, Coxsackievirus B3 on Vero cells, Poliovirus 1 and Echovirus 11 on BGM cells. Activity was confirmed for compound against Rhinovirus 14. Furthermore, few compounds showed a cell-protective effect on Enterovirus 71, presented a marked improvement as compared to the reference drug pleconaril for inhibitory activity on both Enterovirus 71 and Poliovirus 1. The most striking observation was the clear cell protective effect for the set of analogues in a virus-cell-based assay for Echovirus 11 with an effective concentration (EC50) as low as 0.3 µM (Selectivity index or SI = 483), and selectivity indexes greater than 857 (EC50 = 0.6 µM) and 1524 (EC50 = 0.33 µM). Some of the evaluated compounds showed potent and selective antiviral activity against several enterovirus species, such as Enterovirus 71 (EV-A), Echovirus 11 (EV-B), and Poliovirus 1 (EV-C). This could be used as a starting point for the development of other pleconaril/pirodavir-like enterovirus inhibitors with broad-spectrum activity and improved effects as compared to the reference drugs. © The Author(s) 2015.

  19. Impact of rapid molecular diagnostic tests on time to treatment initiation and outcomes in patients with multidrug-resistant tuberculosis, Tamil Nadu, India.

    Science.gov (United States)

    Nair, Dina; Navneethapandian, Pooranaganga D; Tripathy, Jaya Prasad; Harries, Anthony D; Klinton, Joel S; Watson, Basilea; Sivaramakrishnan, Gomathi N; Reddy, Devarajulu S; Murali, Lakshmi; Natrajan, Mohan; Swaminathan, Soumya

    2016-09-01

    India is replacing culture and drug sensitivity testing (CDST) with rapid molecular tests for diagnosing MDR-TB. We assessed the impact of rapid tests on time to initiation of treatment and outcomes in patients with MDR-TB compared with CDST. A retrospective cohort study involving MDR-TB patients from six districts in Tamil Nadu state, who underwent CDST (2010-2011) and rapid tests (2012-2013). There were 135 patients in the CDST group and 389 in the rapid diagnostic test group. Median time from sputum receipt at the laboratory to initiation of MDR-TB treatment was 130 days (IQR 75-213) in the CDST group and 22 days (IQR 14-38) in the rapid diagnostic test group (p30% in both groups and missing data were higher in CDST (13%) compared with rapid tests (3%). There were significantly higher risks of unfavourable treatment outcomes in males (aRR 1.3, 95% CI 1.1-1.5) and those with treatment initiation delays >30 days (aRR 1.3, 95% CI 1.0-1.6). Rapid molecular diagnostic tests shortened the time to initiate treatment which was associated with reduced unfavourable outcomes in MDR-TB patients. This supports the policy to scale up these tests in India. © The Author 2016. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Molecular dynamics study of dynamic and structural properties of supercooled liquid and glassy iron in the rapid-cooling processes

    Energy Technology Data Exchange (ETDEWEB)

    Cao, Qi-Long; Huang, Duo-Hui; Yang, Jun-Sheng; Wan, Min-Jie; Wang, Fan-Hou, E-mail: eatonch@gmail.com

    2014-10-01

    Molecular dynamics simulations were applied to study the dynamic and structural properties of supercooled liquid and glassy iron in the rapid-cooling processes. The mean-square displacement and the non-Gaussian parameter were used to describe the dynamic properties. The evolution of structural properties was investigated using the pair distribution functions and bond-angle distribution functions. Results for dynamic and structural relaxations indicate that the dynamic features are consistently correlated with the structure evolution, and there are three temperature regions as the temperature decreases: (1) at higher temperatures (1500 K, 1300 K, and 1100 K), the system remains in the liquid characteristics during the overall relaxation process. (2) At medial temperatures (1050 K, 900 K, and 700 K), a fast β-relaxation is followed by a much slower α-relaxation. There is a little change in the structural properties in the β-relaxation region, while major configuration rearrangements occurred in the α-relaxation range and the crystallization process was completed at the end of α-relaxation region. (3) At lower temperature (500 K), the system shows glassy characteristics during the overall relaxation process. In addition, the melting temperature, glass transition temperature and diffusion coefficients of supercooled liquid iron are also computed.

  1. Rapid molecular cytogenetic analysis of X-chromosomal microdeletions: Fluorescence in situ hybridization (FISH) for complex glycerol kinase deficiency

    Energy Technology Data Exchange (ETDEWEB)

    Worley, K.C.; Lindsay, E.A.; McCabe, E.R.B. [Baylor College of Medicine, Houston, TX (United States)] [and others

    1995-07-17

    Diagnosis of X-chromosomal microdeletions has relied upon the traditional methods of Southern blotting and DNA amplification, with carrier identification requiring time-consuming and unreliable dosage calculations. In this report, we describe rapid molecular cytogenetic identification of deleted DNA in affected males with the Xp21 contiguous gene syndrome (complex glycerol kinase deficiency, CGKD) and female carriers for this disorder. CGKD deletions involve the genes for glycerol kinase, Duchenne muscular dystrophy, and/or adrenal hypoplasia congenita. We report an improved method for diagnosis of deletions in individuals with CGKD and for identification of female carriers within their families using fluorescence in situ hybridization (FISH) with a cosmid marker (cosmid 35) within the glycerol kinase gene. When used in combination with an Xq control probe, affected males demonstrate a single signal from the control probe, while female carriers demonstrate a normal chromosome with two signals, as well as a deleted chromosome with a single signal from the control probe. FISH analysis for CGKD provides the advantages of speed and accuracy for evaluation of submicroscopic X-chromosome deletions, particularly in identification of female carriers. In addition to improving carrier evaluation, FISH will make prenatal diagnosis of CGKD more readily available. 17 refs., 2 figs.

  2. Divergent Requirement for a DNA Repair Enzyme during Enterovirus Infections

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    Sonia Maciejewski

    2015-12-01

    Full Text Available Viruses of the Enterovirus genus of picornaviruses, including poliovirus, coxsackievirus B3 (CVB3, and human rhinovirus, commandeer the functions of host cell proteins to aid in the replication of their small viral genomic RNAs during infection. One of these host proteins is a cellular DNA repair enzyme known as 5′ tyrosyl-DNA phosphodiesterase 2 (TDP2. TDP2 was previously demonstrated to mediate the cleavage of a unique covalent linkage between a viral protein (VPg and the 5′ end of picornavirus RNAs. Although VPg is absent from actively translating poliovirus mRNAs, the removal of VPg is not required for the in vitro translation and replication of the RNA. However, TDP2 appears to be excluded from replication and encapsidation sites during peak times of poliovirus infection of HeLa cells, suggesting a role for TDP2 during the viral replication cycle. Using a mouse embryonic fibroblast cell line lacking TDP2, we found that TDP2 is differentially required among enteroviruses. Our single-cycle viral growth analysis shows that CVB3 replication has a greater dependency on TDP2 than does poliovirus or human rhinovirus replication. During infection, CVB3 protein accumulation is undetectable (by Western blot analysis in the absence of TDP2, whereas poliovirus protein accumulation is reduced but still detectable. Using an infectious CVB3 RNA with a reporter, CVB3 RNA could still be replicated in the absence of TDP2 following transfection, albeit at reduced levels. Overall, these results indicate that TDP2 potentiates viral replication during enterovirus infections of cultured cells, making TDP2 a potential target for antiviral development for picornavirus infections.

  3. Inhibition of enterovirus 71 entry by transcription factor XBP1

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    Jheng, Jia-Rong; Lin, Chiou-Yan [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China); Horng, Jim-Tong, E-mail: jimtong@mail.cgu.edu.tw [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China); Lau, Kean Seng [Department of Biochemistry and Research Center for Emerging Viral Infections, Chang Gung University, 259 Wen-Hwa First Road, Kweishan, Taoyuan 333, Taiwan (China)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer IRE1 was activated but no XBP1 splicing was detected during enterovirus 71 infection. Black-Right-Pointing-Pointer XBP1 was subject to translational shutoff by enterovirus 71-induced eIF4G cleavage. Black-Right-Pointing-Pointer The uptake of UV-irradiated virus was decreased in XBP1-overexpressing cells. -- Abstract: Inositol-requiring enzyme 1 (IRE1) plays an important role in the endoplasmic reticulum (ER), or unfolded protein, stress response by activating its downstream transcription factor X-box-binding protein 1 (XBP1). We demonstrated previously that enterovirus 71 (EV71) upregulated XBP1 mRNA levels but did not activate spliced XBP1 (XBP1s) mRNA or its downstream target genes, EDEM and chaperones. In this study, we investigated further this regulatory mechanism and found that IRE1 was phosphorylated and activated after EV71 infection, whereas its downstream XBP1s protein level decreased. We also found that XBP1s was not cleaved directly by 2A{sup pro}, but that cleavage of eukaryotic translation initiation factor 4G by the EV71 2A{sup pro} protein may contribute to the decrease in XBP1s expression. Knockdown of XBP1 increased viral protein expression, and the synthesis of EV71 viral protein and the production of EV71 viral particles were inhibited in XBP1-overexpressing RD cells. When incubated with replication-deficient and UV-irradiated EV71, XBP1-overexpressing RD cells exhibited reduced viral RNA levels, suggesting that the inhibition of XBP1s by viral infection may underlie viral entry, which is required for viral replication. Our findings are the first indication of the ability of XBP1 to inhibit viral entry, possibly via its transcriptional activity in regulating molecules in the endocytic machinery.

  4. Inhibition of enterovirus 71 entry by transcription factor XBP1

    International Nuclear Information System (INIS)

    Jheng, Jia-Rong; Lin, Chiou-Yan; Horng, Jim-Tong; Lau, Kean Seng

    2012-01-01

    Highlights: ► IRE1 was activated but no XBP1 splicing was detected during enterovirus 71 infection. ► XBP1 was subject to translational shutoff by enterovirus 71-induced eIF4G cleavage. ► The uptake of UV-irradiated virus was decreased in XBP1-overexpressing cells. -- Abstract: Inositol-requiring enzyme 1 (IRE1) plays an important role in the endoplasmic reticulum (ER), or unfolded protein, stress response by activating its downstream transcription factor X-box-binding protein 1 (XBP1). We demonstrated previously that enterovirus 71 (EV71) upregulated XBP1 mRNA levels but did not activate spliced XBP1 (XBP1s) mRNA or its downstream target genes, EDEM and chaperones. In this study, we investigated further this regulatory mechanism and found that IRE1 was phosphorylated and activated after EV71 infection, whereas its downstream XBP1s protein level decreased. We also found that XBP1s was not cleaved directly by 2A pro , but that cleavage of eukaryotic translation initiation factor 4G by the EV71 2A pro protein may contribute to the decrease in XBP1s expression. Knockdown of XBP1 increased viral protein expression, and the synthesis of EV71 viral protein and the production of EV71 viral particles were inhibited in XBP1-overexpressing RD cells. When incubated with replication-deficient and UV-irradiated EV71, XBP1-overexpressing RD cells exhibited reduced viral RNA levels, suggesting that the inhibition of XBP1s by viral infection may underlie viral entry, which is required for viral replication. Our findings are the first indication of the ability of XBP1 to inhibit viral entry, possibly via its transcriptional activity in regulating molecules in the endocytic machinery.

  5. Class I ADP-ribosylation factors are involved in enterovirus 71 replication.

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    Jianmin Wang

    Full Text Available Enterovirus 71 is one of the major causative agents of hand, foot, and mouth disease in infants and children. Replication of enterovirus 71 depends on host cellular factors. The viral replication complex is formed in novel, cytoplasmic, vesicular compartments. It has not been elucidated which cellular pathways are hijacked by the virus to create these vesicles. Here, we investigated whether proteins associated with the cellular secretory pathway were involved in enterovirus 71 replication. We used a loss-of-function assay, based on small interfering RNA. We showed that enterovirus 71 RNA replication was dependent on the activity of Class I ADP-ribosylation factors. Simultaneous depletion of ADP-ribosylation factors 1 and 3, but not three others, inhibited viral replication in cells. We also demonstrated with various techniques that the brefeldin-A-sensitive guanidine nucleotide exchange factor, GBF1, was critically important for enterovirus 71 replication. Our results suggested that enterovirus 71 replication depended on GBF1-mediated activation of Class I ADP-ribosylation factors. These results revealed a connection between enterovirus 71 replication and the cellular secretory pathway; this pathway may represent a novel target for antiviral therapies.

  6. Accuracy of Diagnostic Methods and Surveillance Sensitivity for Human Enterovirus, South Korea, 1999–2011

    Science.gov (United States)

    Hyeon, Ji-Yeon; Hwang, Seoyeon; Kim, Hyejin; Song, Jaehyoung; Ahn, Jeongbae; Kang, Byunghak; Kim, Kisoon; Choi, Wooyoung; Chung, Jae Keun; Kim, Cheon-Hyun; Cho, Kyungsoon; Jee, Youngmee; Kim, Jonghyun; Kim, Kisang; Kim, Sun-Hee; Kim, Min-Ji

    2013-01-01

    The epidemiology of enteroviral infection in South Korea during 1999–2011 chronicles nationwide outbreaks and changing detection and subtyping methods used over the 13-year period. Of 14,657 patients whose samples were tested, 4,762 (32.5%) samples were positive for human enterovirus (human EV); as diagnostic methods improved, the rate of positive results increased. A seasonal trend of outbreaks was documented. Genotypes enterovirus 71, echovirus 30, coxsackievirus B5, enterovirus 6, and coxsackievirus B2 were the most common genotypes identified. Accurate test results correlated clinical syndromes to enterovirus genotypes: aseptic meningitis to echovirus 30, enterovirus 6, and coxsackievirus B5; hand, foot and mouth disease to coxsackievirus A16; and hand, foot and mouth disease with neurologic complications to enterovirus 71. There are currently no treatments specific to human EV infections; surveillance of enterovirus infections such as this study provides may assist with evaluating the need to research and develop treatments for infections caused by virulent human EV genotypes. PMID:23876671

  7. Class I ADP-ribosylation factors are involved in enterovirus 71 replication.

    Science.gov (United States)

    Wang, Jianmin; Du, Jiang; Jin, Qi

    2014-01-01

    Enterovirus 71 is one of the major causative agents of hand, foot, and mouth disease in infants and children. Replication of enterovirus 71 depends on host cellular factors. The viral replication complex is formed in novel, cytoplasmic, vesicular compartments. It has not been elucidated which cellular pathways are hijacked by the virus to create these vesicles. Here, we investigated whether proteins associated with the cellular secretory pathway were involved in enterovirus 71 replication. We used a loss-of-function assay, based on small interfering RNA. We showed that enterovirus 71 RNA replication was dependent on the activity of Class I ADP-ribosylation factors. Simultaneous depletion of ADP-ribosylation factors 1 and 3, but not three others, inhibited viral replication in cells. We also demonstrated with various techniques that the brefeldin-A-sensitive guanidine nucleotide exchange factor, GBF1, was critically important for enterovirus 71 replication. Our results suggested that enterovirus 71 replication depended on GBF1-mediated activation of Class I ADP-ribosylation factors. These results revealed a connection between enterovirus 71 replication and the cellular secretory pathway; this pathway may represent a novel target for antiviral therapies.

  8. MALDI-TOF MS enables the rapid identification of the major molecular types within the Cryptococcus neoformans/C. gattii species complex.

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    Carolina Firacative

    Full Text Available BACKGROUND: The Cryptococcus neoformans/C. gattii species complex comprises two sibling species that are divided into eight major molecular types, C. neoformans VNI to VNIV and C. gattii VGI to VGIV. These genotypes differ in host range, epidemiology, virulence, antifungal susceptibility and geographic distribution. The currently used phenotypic and molecular identification methods for the species/molecular types are time consuming and expensive. As Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS offers an effective alternative for the rapid identification of microorganisms, the objective of this study was to examine its potential for the identification of C. neoformans and C. gattii strains at the intra- and inter-species level. METHODOLOGY: Protein extracts obtained via the formic acid extraction method of 164 C. neoformans/C. gattii isolates, including four inter-species hybrids, were studied. RESULTS: The obtained mass spectra correctly identified 100% of all studied isolates, grouped each isolate according to the currently recognized species, C. neoformans and C. gattii, and detected potential hybrids. In addition, all isolates were clearly separated according to their major molecular type, generating greater spectral differences among the C. neoformans molecular types than the C. gattii molecular types, most likely reflecting a closer phylogenetic relationship between the latter. The number of colonies used and the incubation length did not affect the results. No spectra were obtained from intact yeast cells. An extended validated spectral library containing spectra of all eight major molecular types was established. CONCLUSIONS: MALDI-TOF MS is a rapid identification tool for the correct recognition of the two currently recognized human pathogenic Cryptococcus species and offers a simple method for the separation of the eight major molecular types and the detection of hybrid strains within this

  9. Rapid Colorimetric Detection of Cartap Residues by AgNP Sensor with Magnetic Molecularly Imprinted Microspheres as Recognition Elements

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    Mao Wu

    2018-06-01

    Full Text Available The overuse of cartap in tea tree leads to hazardous residues threatening human health. A colorimetric determination was established to detect cartap residues in tea beverages by silver nanoparticles (AgNP sensor with magnetic molecularly imprinted polymeric microspheres (Fe3O4@mSiO2@MIPs as recognition elements. Using Fe3O4 as supporting core, mesoporous SiO2 as intermediate shell, methylacrylic acid as functional monomer, and cartap as template, Fe3O4@mSiO2@MIPs were prepared to selectively and magnetically separate cartap from tea solution before colorimetric determination by AgNP sensors. The core-shell Fe3O4@mSiO2@MIPs were also characterized by FT-IR, TEM, VSM, and experimental adsorption. The Fe3O4@mSiO2@MIPs could be rapidly separated by an external magnet in 10 s with good reusability (maintained 95.2% through 10 cycles. The adsorption process of cartap on Fe3O4@mSiO2@MIPs conformed to Langmuir adsorption isotherm with maximum adsorption capacity at 0.257 mmol/g and short equilibrium time of 30 min at 298 K. The AgNP colorimetric method semi-quantified cartap ≥5 mg/L by naked eye and quantified cartap 0.1–5 mg/L with LOD 0.01 mg/L by UV-vis spectroscopy. The AgNP colorimetric detection after pretreatment with Fe3O4@mSiO2@MIPs could be successfully utilized to recognize and detect cartap residues in tea beverages.

  10. A novel Enterovirus 96 circulating in China causes hand, foot, and mouth disease.

    Science.gov (United States)

    Xu, Yi; Sun, Yisuo; Ma, Jinmin; Zhou, Shuru; Fang, Wei; Ye, Jiawei; Tan, Limei; Ji, Jingkai; Luo, Dan; Li, Liqiang; Li, Jiandong; Fang, Chunxiao; Pei, Na; Shi, Shuo; Liu, Xin; Jiang, Hui; Gong, Sitang; Xu, Xun

    2017-06-01

    Enterovirus 96 (EV-96) is a recently described member of the species Enterovirus C and is associated with paralysis and myelitis. In this study, using metagenomic sequencing, we identified a new enterovirus 96 strain (EV-96-SZ/GD/CHN/2014) as the sole pathogen causing hand, foot, and mouth disease (HFMD). A genomic comparison showed that EV-96-SZ/GD/CHN/2014 is most similar to the EV-96-05517 strain (85% identity), which has also been detected in Guangdong Province. This is the first time that metagenomic sequencing has been used to identify an EV-96 strain shown to be associated with HFMD.

  11. Management strategies of enterovirus D68 outbreaks: current perspectives

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    Milhano N

    2018-03-01

    Full Text Available Natacha Milhano, Kaja Sverdrup Borge, Karoline Bragstad, Susanne G Dudman Domain for Environmental Health and Infectious Disease Control, Norwegian Institute of Public Health, Oslo, Norway Abstract: Following its discovery in California in 1962, enterovirus D68 (EV-D68 was reported only sporadically around the world. In August 2014, a marked increase of EV-D68 cases in young children with severe respiratory infections was reported in the USA and Canada and later in Europe and Asia. Some of these cases were also found to be associated with acute flaccid paralysis, which exacerbated public health concern, and has since triggered international efforts to strengthen both EV-D68 and acute flaccid paralysis surveillance systems. This review summarizes the current knowledge on EV-D68, offering an overview of EV-D68 epidemiology, clinical presentations, diagnostic methodologies, and treatment strategies, as well as surveillance and outbreak management. Keywords: enterovirus D68, AFP, diagnostics, treatment, surveillance, outbreak 

  12. African Non-Human Primates Host Diverse Enteroviruses.

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    Illich Manfred Mombo

    Full Text Available Enteroviruses (EVs belong to the family Picornaviridae and are responsible for mild to severe diseases in mammals including humans and non-human primates (NHP. Simian EVs were first discovered in the 1950s in the Old World Monkeys and recently in wild chimpanzee, gorilla and mandrill in Cameroon. In the present study, we screened by PCR EVs in 600 fecal samples of wild apes and monkeys that were collected at four sites in Gabon. A total of 32 samples were positive for EVs (25 from mandrills, 7 from chimpanzees, none from gorillas. The phylogenetic analysis of VP1 and VP2 genes showed that EVs identified in chimpanzees were members of two human EV species, EV-A and EV-B, and those identified in mandrills were members of the human species EV-B and the simian species EV-J. The identification of two novel enterovirus types, EV-B112 in a chimpanzee and EV-B113 in a mandrill, suggests these NHPs could be potential sources of new EV types. The identification of EV-B107 and EV90 that were previously found in humans indicates cross-species transfers. Also the identification of chimpanzee-derived EV110 in a mandrill demonstrated a wide host range of this EV. Further research of EVs in NHPs would help understanding emergence of new types or variants, and evaluating the real risk of cross-species transmission for humans as well for NHPs populations.

  13. Characterization of a novel enterovirus serotype and an enterovirus EV-B93 isolated from acute flaccid paralysis patients.

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    Shahzad Shaukat

    Full Text Available Non-polio enteroviruses (NPEVs are among the most common viruses infecting humans worldwide. Most of these infections are asymptomatic but few can lead to systemic and neurological disorders like Acute Flaccid Paralysis (AFP. Acute Flaccid Paralysis is a clinical syndrome and NPEVs have been isolated frequently from the patients suffering from AFP but little is known about their causal relationship. The objective of this study was to identify and characterize the NPEV serotypes recovered from 184 stool samples collected from AFP patients in Federally Administered Tribal Areas (FATA in north-west of Pakistan. Overall, 44 (95.6 % isolates were successfully typed through microneutralization assay as a member of enterovirus B species including echovirus (E-2, E-3, E-4, E-6, E-7, E-11, E-13, E-14, E-21 and E-29 while two isolates (PAK NIH SP6545B and PAK NIH SP1202B remained untypeable. The VP1 and capsid regions analysis characterized these viruses as EV-B93 and EV-B106. Phylogenetic analysis confirmed that PAK NIH isolates had high genetic diversity and represent distinct genotypes circulating in the country. Our findings highlight the role of NPEVs in AFP cases to be thoroughly investigated especially in high disease risk areas, with limited surveillance activities and health resources.

  14. Direct-acting antivirals and host-targeting strategies to combat enterovirus infections.

    Science.gov (United States)

    Bauer, Lisa; Lyoo, Heyrhyoung; van der Schaar, Hilde M; Strating, Jeroen Rpm; van Kuppeveld, Frank Jm

    2017-06-01

    Enteroviruses (e.g., poliovirus, enterovirus-A71, coxsackievirus, enterovirus-D68, rhinovirus) include many human pathogens causative of various mild and more severe diseases, especially in young children. Unfortunately, antiviral drugs to treat enterovirus infections have not been approved yet. Over the past decades, several direct-acting inhibitors have been developed, including capsid binders, which block virus entry, and inhibitors of viral enzymes required for genome replication. Capsid binders and protease inhibitors have been clinically evaluated, but failed due to limited efficacy or toxicity issues. As an alternative approach, host-targeting inhibitors with potential broad-spectrum activity have been identified. Furthermore, drug repurposing screens have recently uncovered promising new inhibitors with disparate viral and host targets. Together, these findings raise hope for the development of (broad-range) anti-enteroviral drugs. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. [Genomic characteristics and recombination of enterovirus 71 strains isolated in Henan Province between 2008 and 2010].

    Science.gov (United States)

    Wei, Hai-Yan; Xu, Yu-Ling; Huang, Xue-Yong; Ma, Hong; Chen, Hao-Min; Xu, Bian-Li

    2011-09-01

    To reveal the genetic features and recombination of enterovirus 71 isolates between 2008 and 2010. A total of 5 enterovirus 71 isolates were sequenced completely and phylogenetic analysis and recombination were performed. Phylogenetic analysis based on VP1 regions revealed that the Henan enterovirus 71 between 2008 and 2010 belonged to C4a in subgenotype C4. Bootscan analyses and phylogenetic analysis based on the 5'UTR, P1, P2, P3 genomic regions revealed the recombinations between EV71 genotypes B and C at the 2A-2B junction, and between EV71 genotype B and CA16 strain G-10 at the 3B-3C junction. Henan enterovirus 71 isolates between 2008 and 2010 belonged to C4a in subgenotype C4 which was the predominant virus genotype circulating in mainland China since 2004, a combination of intratypic and intertypic recombination were found in EV71 subgenotype C4.

  16. Seroprevalence of Enterovirus A71 and Coxsackievirus A16 in Healthy People in Shandong Province, China.

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    Jian-Xing Wang

    Full Text Available Hand, foot, and mouth disease has become very common in mainland of China in recent years, and enterovirus A71 and coxsackievirus A16 are its major etiologic factors. Here we investigated the seroprevalence of enterovirus A71 and coxsackievirus A16 based on a large group of healthy individuals in Shandong province, China.A total of 1378 healthy individuals were tested for serum neutralizing antibodies against enterovirus A71 and coxsackievirus A16 using a micro neutralization test.The overall seroprevalence of enterovirus A71 neutralizing antibodies was 74.75%. It increased significantly from 48.84% in children aged 0-1 years old to 88.64% in those aged 20-29 years (p 40 years old with a significant gender-specific difference (p 40 years without a gender-specific difference. Nearly 50% of the children <1 year were susceptible to enterovirus A71 infection versus 40% to coxsackievirus A16 infection. Sample collection time and place also played a role in the enterovirus A71 and coxsackievirus A16 positive rates. The overall rates in January were significantly lower than those in April and August (p < 0.01; enterovirus A71 positive rates in Jinan city (capital city of Shandong province were lower than those in Jining city and Zibo city (p < 0.05; and oxsackievirus A16 positive rates in Jining city were significantly higher than those in Jinan city and Zibo city (p < 0.01.There were significant differences among age groups, locations, and time points in the seroprevalence rates of enterovirus A71 and coxsackievirus A16 neutralizing antibodies in healthy people in Shandong province.

  17. A novel dromedary camel enterovirus in the family Picornaviridae from dromedaries in the Middle East.

    Science.gov (United States)

    Woo, Patrick C Y; Lau, Susanna K P; Li, Tong; Jose, Shanty; Yip, Cyril C Y; Huang, Yi; Wong, Emily Y M; Fan, Rachel Y Y; Cai, Jian-Piao; Wernery, Ulrich; Yuen, Kwok-Yung

    2015-07-01

    The recent emergence of Middle East respiratory syndrome coronavirus from the Middle East and the discovery of the virus from dromedary camels have boosted interest in the search for novel viruses in dromedaries. Whilst picornaviruses are known to infect various animals, their existence in dromedaries was unknown. We describe the discovery of a novel picornavirus, dromedary camel enterovirus (DcEV), from dromedaries in Dubai. Among 215 dromedaries, DcEV was detected in faecal samples of four (1.9 %) dromedaries [one (0.5 %) adult dromedary and three (25 %) dromedary calves] by reverse transcription PCR. Analysis of two DcEV genomes showed that DcEV was clustered with other species of the genus Enterovirus and was most closely related to and possessed highest amino acid identities to the species Enterovirus E and Enterovirus F found in cattle. The G+C content of DcEV was 45 mol%, which differed from that of Enterovirus E and Enterovirus F (49-50 mol%) by 4-5 %. Similar to other members of the genus Enterovirus, the 5' UTR of DcEV possessed a putative type I internal ribosome entry site. The low ratios of the number of nonsynonymous substitutions per non-synonymous site to the number of synonymous substitutions per synonymous site (Ka/Ks) of various coding regions suggested that dromedaries are the natural reservoir in which DcEV has been stably evolving. These results suggest that DcEV is a novel species of the genus Enterovirus in the family Picornaviridae. Western blot analysis using recombinant DcEV VP1 polypeptide showed a high seroprevalence of 52 % among serum samples from 172 dromedaries for IgG, concurring with its much higher infection rates in dromedary calves than in adults. Further studies are important to understand the pathogenicity, epidemiology and genetic evolution of DcEV in this unique group of animals.

  18. Development and evaluation of a real-time method for testing human enteroviruses and coxsackievirus A16.

    Science.gov (United States)

    Chen, Qian; Hu, Zheng; Zhang, Qihua; Yu, Minghui

    2016-05-01

    Hand, foot, and mouth disease (HFMD) is a common infectious disease caused by a group of the human enteroviruses (HEV), including coxsackievirus A16 (CA16) and enterovirus 71 (EV71). In recent years, another HEV-A serotype, CA6 or CA10, has emerged to be one of the major etiologic agents that can induce HFMD worldwide. The objective of this study is to develop specific, sensitive, and rapid methods to help diagnose HEV and CA16 specifically by using simultaneous amplification testing (SAT) based on isothermal amplification of RNA and real-time detection of fluorescence technique, which were named as SAT-HEV and SAT-CA16, respectively (SAT-HEV/SAT-CA16). The specificity and sensitivity of SAT were tested here. SAT-HEV/SAT-CA16 could measure viral titers that were at least 10-fold lower than those measured by real-time PCR. Non-false cross-reactive amplification indicated that SAT-HEV/SAT-CA16 were highly specific with the addition of internal control (IC) RNA (5000 copies/reaction). A total of 198 clinical specimens were assayed by SAT comparing with real-time PCR. The statistically robust assessment of SAT-HEV and HEV-specific real-time PCR plus sequencing reached 99.0% (196/198), with a kappa value of 0.97, and 99.5% (197/198) and a kappa value of 0.99 for CA16, respectively. Additionally, IC prevented false-negative readings and assured the SAT-HEV/SAT-CA16 method's accuracy. Overall, SAT-HEV/SAT-CA16 method may serve as a platform for the simple and rapid detection of HEV/CA16 in time of HFMD outbreak. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. [MANIFESTATIONS OF EPIDEMIC PROCESS AND TRANSMISSION ROUTES OF CAUSATIVE AGENT OF ENTEROVIRUS SEROUS MENINGITIS].

    Science.gov (United States)

    Sergevnin, V I; Tryasolobova, M A; Kudrevatykh, E V; Kuzovnikova, E Zh

    2015-01-01

    Study the manifestations of epidemic process and leading transmission routes of causative agents of enterovirus serous meningitis (SM) by results of laboratory studies and epidemiologic examination of epidemic nidi. During 2010 - 2014 a study for enterovirus was carried out in cerebrospinal fluid in 743 patients, hospitalized into medical organizations of Perm with primary diagnosis "serous meningitis", feces of 426 individuals, that had communicated with patients with SM of enterovirus etiology; 827 water samples from the distribution network, 295 water samples from open water and 57 washes from surface of vegetables and fruits. All the samples were studied in polymerase chain reaction, part--by a virological method. Epidemiologic examination of 350 epidemic nidi of SM was carried out. Enterovirus and (or) its RNA were detected in 62.0% of patients and 61.9% of individuals that had communicated with patients with enteroviris SM. ECHO 6 serotype enterovirus dominated among the causative agents. Maximum intensity of epidemic process of enterovirus SM, based on data from laboratory examination of patients, was detected in a group of organized pre-school and school age children during summer-autumn period. . Examination of epidemic nidi and laboratory control of environmental objects have shown that CV causative agent transmission factors are, in particular, unboiled water from decentralized sources (boreholes, wells, springs), water from open waters during bathing, as well as fresh vegetables, fruits, berries and meals produced from them. .

  20. Australian National Enterovirus Reference Laboratory annual report, 2013.

    Science.gov (United States)

    Roberts, Jason A; Hobday, Linda K; Ibrahim, Aishah; Aitkin, Thomas; Thorley, Bruce R

    2015-06-30

    Australia conducts surveillance for cases of acute flaccid paralysis (AFP) in children less than 15 years of age as the main method to monitor its polio-free status in accordance with the World Health Organization (WHO) recommendations. Cases of AFP in children are notified to the Australian Paediatric Surveillance Unit or the Paediatric Active Enhanced Disease Surveillance System and faecal specimens are referred for virological investigation to the National Enterovirus Reference Laboratory. In 2013, no cases of poliomyelitis were reported from clinical surveillance and Australia reported 1.4 non-polio AFP cases per 100,000 children, meeting the WHO performance criterion for a sensitive surveillance system. Non-polio enteroviruses can also be associated with AFP and enterovirus A71 was identified from nine of the 61 cases classified as non-polio AFP in 2013, which was part of a larger outbreak associated with this virus. A Sabin poliovirus was detected in an infant recently returned from Pakistan and who had been vaccinated while abroad. Globally, 416 cases of polio were reported in 2013, with the 3 endemic countries: Afghanistan; Nigeria; and Pakistan, accounting for 38% of the cases. To safeguard the progress made towards polio eradication, in May 2014, WHO recommended travellers from the 10 countries that are currently reporting wild poliovirus transmission have documented evidence of recent polio vaccination before departure. This work is copyright. You may download, display, print and reproduce the whole or part of this work in unaltered form for your own personal use or, if you are part of an organisation, for internal use within your organisation, but only if you or your organisation do not use the reproduction for any commercial purpose and retain this copyright notice and all disclaimer notices as part of that reproduction. Apart from rights to use as permitted by the Copyright Act 1968 or allowed by this copyright notice, all other rights are reserved

  1. Structural characterization and antiviral activity of a novel heteropolysaccharide isolated from Grifola frondosa against enterovirus 71.

    Science.gov (United States)

    Zhao, Chao; Gao, Luying; Wang, Chunyang; Liu, Bin; Jin, Yu; Xing, Zheng

    2016-06-25

    A novel heteropolysaccharide from Grifola frondosa mycelia was extracted and purified using DEAE Sephadex A-50 and Sephadex G-200 chromatography. Fourier transform infrared (FT-IR) spectroscopy and nuclear magnetic resonance ((1)H NMR and (13)C NMR) spectroscopy were used to decipher the structure of the purified G. frondosa polysaccharide (GFP1). Chemical and spectral analysis revealed that GFP1, with an average molecular weight of 40.5kDa, possessed a 1,6-β-d-glucan backbone with a single 1,3-α-d-fucopyranosyl side-branching unit. Enterovirus 71 (EV71) is the causative pathogen of hand-foot-and-mouth disease. GFP1 was tested for its anti-EV71 activity in cultured cells, which showed that EV71 viral replication was blocked and viral VP1 protein expression and genomic RNA synthesis were suppressed. Moreover, GFP1 exhibited apoptotic and other activities by suppressing the EV71-induced caspase-3 cleavage and IκBα down regulation. Our results demonstrate that the novel G. frondosa polysaccharide has antiviral activity, which could be valuable as a potentially new anti-EV71 therapeutic compound. Copyright © 2016. Published by Elsevier Ltd.

  2. Analysis of miRNAs Involved in Mouse Brain Damage upon Enterovirus 71 Infection.

    Science.gov (United States)

    Yang, Xiaoxia; Xie, Jing; Jia, Leili; Liu, Nan; Liang, Yuan; Wu, Fuli; Liang, Beibei; Li, Yongrui; Wang, Jinyan; Sheng, Chunyu; Li, Hao; Liu, Hongbo; Ma, Qiuxia; Yang, Chaojie; Du, Xinying; Qiu, Shaofu; Song, Hongbin

    2017-01-01

    Enterovirus 71 (EV71) infects the central nervous system (CNS) and causes brainstem encephalitis in children. MiRNAs have been found to play various functions in EV71 infection in human cell lines. To identify potential miRNAs involved in the inflammatory injury in CNS, our study, for the first time, performed a miRNA microarray assay in vivo using EV71 infected mice brains. Twenty differentially expressed miRNAs were identified (four up- and 16 down-regulated) and confirmed by qRT-PCR. The target genes of these miRNAs were analyzed using KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis, revealing that the miRNAs were mainly involved in the regulation of inflammation and neural system function. MiR-150-5p, -3082-5p, -3473a, -468-3p, -669n, -721, -709, and -5107-5p that regulate MAPK and chemokine signaling were all down-regulated, which might result in increased cytokine production. In addition, miR-3473a could also regulate focal adhesion and leukocyte trans-endothelial migration, suggesting a role in virus-induced blood-brain barrier disruption. The miRNAs and pathways identified in this study could help to understand the intricate interactions between EV71 and the brain injury, offering new insight for the future research of the molecular mechanism of EV71 induced brainstem encephalitis.

  3. Rapid Determination of Six Low Molecular Carbonyl Compounds in Tobacco Smoke by the APCI-MS/MS Coupled to Data Mining

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    Wuduo Zhao

    2017-01-01

    Full Text Available A simple method was established for the rapid determination of low molecular carbonyl compounds by the combination of atmospheric pressure chemical ionization tandem mass spectrometry (APCI-MS/MS and data mining. The ionization was carried out in positive mode, and six low molecular carbonyl compounds of acrolein, acetone, propionaldehyde, crotonaldehyde, butanone, and butyraldehyde were analyzed by both full scan mode and daughter scan mode. To overcome the quantitative difficulties from isomer of acetone/propionaldehyde and butanone/butyraldehyde, the quantitation procedure was performed with the characteristic ion of [CH3O]+ under CID energy of 5 and 15 eV. Subsequently, the established method was successfully applied to analysis of six low molecular carbonyl compounds in tobacco smoke with analytical period less than four minutes. The contents of acrolein, acetone, propionaldehyde, crotonaldehyde, butanone, and butyraldehyde for a cigarette were about 63±5.8, 325±82, 55±9.7, 11±1.4, 67±5.9, and 12±1.8 μg/cig, respectively. The experimental results indicated that the established method had the potential application in rapid determination of low molecular carbonyl compounds.

  4. HIV-TRACE (Transmission Cluster Engine): a tool for large scale molecular epidemiology of HIV-1 and other rapidly evolving pathogens.

    Science.gov (United States)

    Kosakovsky Pond, Sergei L; Weaver, Steven; Leigh Brown, Andrew J; Wertheim, Joel O

    2018-01-31

    In modern applications of molecular epidemiology, genetic sequence data are routinely used to identify clusters of transmission in rapidly evolving pathogens, most notably HIV-1. Traditional 'shoeleather' epidemiology infers transmission clusters by tracing chains of partners sharing epidemiological connections (e.g., sexual contact). Here, we present a computational tool for identifying a molecular transmission analog of such clusters: HIV-TRACE (TRAnsmission Cluster Engine). HIV-TRACE implements an approach inspired by traditional epidemiology, by identifying chains of partners whose viral genetic relatedness imply direct or indirect epidemiological connections. Molecular transmission clusters are constructed using codon-aware pairwise alignment to a reference sequence followed by pairwise genetic distance estimation among all sequences. This approach is computationally tractable and is capable of identifying HIV-1 transmission clusters in large surveillance databases comprising tens or hundreds of thousands of sequences in near real time, i.e., on the order of minutes to hours. HIV-TRACE is available at www.hivtrace.org and from github.com/veg/hivtrace, along with the accompanying result visualization module from github.com/veg/hivtrace-viz. Importantly, the approach underlying HIV-TRACE is not limited to the study of HIV-1 and can be applied to study outbreaks and epidemics of other rapidly evolving pathogens. © The Author 2018. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Synthesis of molecularly imprinted dye-silica nanocomposites with high selectivity and sensitivity: Fluorescent imprinted sensor for rapid and efficient detection of τ-fluvalinate in vodka.

    Science.gov (United States)

    Wang, Yunyun; Wang, Jixiang; Cheng, Rujia; Sun, Lin; Dai, Xiaohui; Yan, Yongsheng

    2018-04-01

    An imprinted fluorescent sensor was fabricated based on SiO 2 nanoparticles encapsulated with a molecularly imprinted polymer containing allyl fluorescein. High fluorine cypermethirin as template molecules, methyl methacrylate as functional monomer, and allyl fluorescein as optical materials synthesized a core-shell fluorescent molecular imprinted sensor, which showed a high and rapid sensitivity and selectivity for the detection of τ-fluvalinate. The sensor presented appreciable sensitivity with a limit of 13.251 nM, rapid detection that reached to equilibrium within 3 min, great linear relationship in the relevant concentration range from 0 to 150 nM, and excellent selectivity over structural analogues. In addition, the fluorescent sensor demonstrated desirable regeneration ability (eight cycling operations). The molecularly imprinted polymers ensured specificity, while the fluorescent dyes provided the stabile sensitivity. Finally, an effective application of the sensor was implemented by the detection of τ-fluvalinate in real samples from vodka. The molecularly imprinted fluorescent sensor showed a promising potential in environmental monitoring and food safety. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Molecular tools for bathing water assessment in Europe: Balancing social science research with a rapidly developing environmental science evidence-base.

    Science.gov (United States)

    Oliver, David M; Hanley, Nick D; van Niekerk, Melanie; Kay, David; Heathwaite, A Louise; Rabinovici, Sharyl J M; Kinzelman, Julie L; Fleming, Lora E; Porter, Jonathan; Shaikh, Sabina; Fish, Rob; Chilton, Sue; Hewitt, Julie; Connolly, Elaine; Cummins, Andy; Glenk, Klaus; McPhail, Calum; McRory, Eric; McVittie, Alistair; Giles, Amanna; Roberts, Suzanne; Simpson, Katherine; Tinch, Dugald; Thairs, Ted; Avery, Lisa M; Vinten, Andy J A; Watts, Bill D; Quilliam, Richard S

    2016-02-01

    The use of molecular tools, principally qPCR, versus traditional culture-based methods for quantifying microbial parameters (e.g., Fecal Indicator Organisms) in bathing waters generates considerable ongoing debate at the science-policy interface. Advances in science have allowed the development and application of molecular biological methods for rapid (~2 h) quantification of microbial pollution in bathing and recreational waters. In contrast, culture-based methods can take between 18 and 96 h for sample processing. Thus, molecular tools offer an opportunity to provide a more meaningful statement of microbial risk to water-users by providing near-real-time information enabling potentially more informed decision-making with regard to water-based activities. However, complementary studies concerning the potential costs and benefits of adopting rapid methods as a regulatory tool are in short supply. We report on findings from an international Working Group that examined the breadth of social impacts, challenges, and research opportunities associated with the application of molecular tools to bathing water regulations.

  7. Enterovirus-associated changes in blood transcriptomic profiles of children with genetic susceptibility to type 1 diabetes.

    Science.gov (United States)

    Lietzen, Niina; An, Le T T; Jaakkola, Maria K; Kallionpää, Henna; Oikarinen, Sami; Mykkänen, Juha; Knip, Mikael; Veijola, Riitta; Ilonen, Jorma; Toppari, Jorma; Hyöty, Heikki; Lahesmaa, Riitta; Elo, Laura L

    2018-02-01

    Enterovirus infections have been associated with the development of type 1 diabetes in multiple studies, but little is known about enterovirus-induced responses in children at risk for developing type 1 diabetes. Our aim was to use genome-wide transcriptomics data to characterise enterovirus-associated changes in whole-blood samples from children with genetic susceptibility to type 1 diabetes. Longitudinal whole-blood samples (356 samples in total) collected from 28 pairs of children at increased risk for developing type 1 diabetes were screened for the presence of enterovirus RNA. Seven of these samples were detected as enterovirus-positive, each of them collected from a different child, and transcriptomics data from these children were analysed to understand the individual-level responses associated with enterovirus infections. Transcript clusters with peaking or dropping expression at the time of enterovirus positivity were selected as the enterovirus-associated signals. Strong signs of activation of an interferon response were detected in four children at enterovirus positivity, while transcriptomic changes in the other three children indicated activation of adaptive immune responses. Additionally, a large proportion of the enterovirus-associated changes were specific to individuals. An enterovirus-induced signature was built using 339 genes peaking at enterovirus positivity in four of the children, and 77 of these genes were also upregulated in human peripheral blood mononuclear cells infected in vitro with different enteroviruses. These genes separated the four enterovirus-positive samples clearly from the remaining 352 blood samples analysed. We have, for the first time, identified enterovirus-associated transcriptomic profiles in whole-blood samples from children with genetic susceptibility to type 1 diabetes. Our results provide a starting point for understanding the individual responses to enterovirus infections in blood and their potential connection to

  8. Development of duplex real-time RT-PCR based on Taqman technology for detecting simultaneously the genome of pan-enterovirus and enterovirus 71.

    Science.gov (United States)

    Hwang, Seoyeon; Kang, Byunghak; Hong, Jiyoung; Kim, Ahyoun; Kim, Hyejin; Kim, Kisang; Cheon, Doo-Sung

    2013-07-01

    Human enterovirus (EV) 71 is the main etiological agent of hand, foot, and mouth disease (HFMD). It is associated with neurological complications, and caused fatalities during recent outbreaks in the Asia-Pacific region. Infections caused by EV71 could lead to many complications, ranging from brainstem encephalitis to pulmonary oedema, resulting in high mortality. In this study, a duplex real-time RT-PCR assay was developed in order to simultaneously detect pan-EV and EV71. EV71-specific primers and probes were designed based on the highly conserved VP1 region of EV71. Five EV71 strains were detected as positive, and no positive fluorescence signal was observed in the duplex real-time RT-PCR for other viral RNA, which showed 100% specificity for the selected panel, and no cross-reactions were observed in this duplex real-time RT-PCR. The EV71-specific duplex real-time RT-PCR was more sensitive than conventional RT-PCR, and detected viral titers that were 10-fold lower than those measured by the latter. Of the 381 HFMD clinical specimens, 196 (51.4%) cases were pan-EV-positive, of which 170 (86.7%) were EV71-positive when tested by pan-EV and EV71-specific duplex real-time RT-PCR. EV71-specific duplex real-time RT-PCR offers a rapid and sensitive method to detect EV71 from clinical specimens, and will allow quarantine measures to be taken more effectively during outbreaks. Copyright © 2013 Wiley Periodicals, Inc.

  9. Proactive approach to containment of enterovirus infection in the nursery.

    Science.gov (United States)

    Fuchs, Inbal; Golan, Agneta; Borer, Abraham; Shemer-Avni, Yonat; Dagan, Ron; Greenberg, David

    2013-07-01

    Administration of prophylactic intravenous immunoglobulins to contacts of infants actively shedding enterovirus during a hospital nursery outbreak may attenuate severity of disease in those contacts and aid in containment of the outbreak. Four cases of neonatal enteroviral disease were treated in our hospital nursery in July and August 2011; 3 were presumed or proven vertical transmission cases and 1 was a presumed horizontal transmission. We aimed to prevent development of severe illness in contacts of affected neonates following a ministry of health advisory during the summer of 2011 warning of increased neonatal enteroviral morbidity and mortality in Israel. Strict infection control measures were implemented, including meticulous decontamination of the nursery environment and administration of intravenous immunoglobulin prophylaxis to contacts. No further horizontal transmission occurred after infection control interventions. Immunoglobulin prophylaxis to control enteroviral infection in the nursery should be considered as an auxiliary infection control intervention during a nursery outbreak.

  10. The Antiviral Effect of Baicalin on Enterovirus 71 In Vitro

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    Xiang Li

    2015-08-01

    Full Text Available Baicalin is a flavonoid compound extracted from Scutellaria roots that has been reported to possess antibacterial, anti-inflammatory, and antiviral activities. However, the antiviral effect of baicalin on enterovirus 71 (EV71 is still unknown. In this study, we found that baicalin showed inhibitory activity on EV71 infection and was independent of direct virucidal or prophylactic effect and inhibitory viral absorption. The expressions of EV71/3D mRNA and polymerase were significantly blocked by baicalin treatment at early stages of EV71 infection. In addition, baicalin could decrease the expressions of FasL and caspase-3, as well as inhibit the apoptosis of EV71-infected human embryonal rhabdomyosarcoma (RD cells. Altogether, these results indicate that baicalin exhibits potent antiviral effect on EV71 infection, probably through inhibiting EV71/3D polymerase expression and Fas/FasL signaling pathways.

  11. Interleukin-18 protects mice from Enterovirus 71 infection.

    Science.gov (United States)

    Li, Zheng; Wang, Hongbin; Chen, Yihui; Niu, Junling; Guo, Qiuhong; Leng, Qibin; Huang, Zhong; Deng, Zhirui; Meng, Guangxun

    2017-08-01

    Previous study has demonstrated that the NLRP3 inflammasome is essential for protecting murine host against Enterovirus 71 (EV71) infection. However, the underlying mechanism remained unknown. Here we discovered that the pleiotropic cytokine interleukin-18 (IL-18), an NLRP3 inflammasome-dependent effector protein, exhibits a protective capability against EV71 challenge. Deficiency of IL-18 in mice exacerbated EV71 infection, which was reflected by increased viral replication, elevated production of interferons (IFN-β, IFN-γ), proinflammatory cytokines (TNF-α, IL-6) and chemokine CCL2,as well as decreased survival of experimental animals. Conversely, administration of recombinant IL-18 considerably restrained EV71 infection in IL-18 deficient mice. Thus, our results revealed a protective role for IL-18 against EV71 challenge, and indicated a novel therapeutic application for IL-18 in EV71 associated hand, foot, and mouth disease (HFMD). Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Human enteroviruses in oysters and their overlying waters.

    Science.gov (United States)

    Goyal, S M; Gerba, C P; Melnick, J L

    1979-03-01

    The presence of enteroviruses in oysters and oyster-harvesting waters of the Texas Gulf coast was monitored over a period of 10 months. Viruses were detected in water and oyster samples obtained from areas both open and closed to shellfish harvesting. Viruses were detected periodically in waters that met current bacteriological standards for shellfish harvesting. No significant statistical relationship was demonstrated between virus concentration in oysters and the bacteriological and physiochemical quality of water and shellfish. Viruses in water were, however, moderately correlated with total coliforms in water and oysters and with fecal coliforms in oysters. Total coliforms in water were realted to total coliforms in sediment were related only to total coliforms in sediment. Among the physiochemical characteristics of water, turbidity was related statistically to the organic matter content of water and to fecal coliforms in water. There was a marked effect of rainfall on the bacteriological quality of water. Of a total of 44 water samples, 26 yielded virus in concentrations from 4 to 167 plaque-forming units per 100-gallon (ca. 378.5-liter) sample. Of a total of 40 pools of 10 to 12 oysters each, virus was found in 14 pools at a concentration of 6 to 224 plaque-forming units per 100 g of oyster meat. On five occasions, virus was found in water samples when no virus could be detected in oysters harvested from the same sites. This study indicates that current bacteriological standards for determining the safety of shellfish and shellfish-growing waters do no reflect the occurrence of enteroviruses.

  13. Enterovirus D68 in Viet Nam (2009-2015 [version 1; referees: 2 approved, 1 approved with reservations

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    Nguyen Thi Han Ny

    2017-06-01

    Full Text Available Background: Since 1962, enterovirus D68 (EV-D68 has been implicated in multiple outbreaks and sporadic cases of respiratory infection worldwide, but especially in the USA and Europe with an increasing frequency between 2010 and 2014. We describe the detection, associated clinical features and molecular characterization of EV-D68 in central and southern Viet Nam between 2009 and 2015. Methods: Enterovirus/rhinovirus PCR positive respiratory or CSF samples taken from children and adults with respiratory/central nervous system infections in Viet Nam were tested by an EV-D68 specific PCR. The included samples were derived from 3 different observational studies conducted at referral hospitals across central and southern Viet Nam between 2009 and 2015. Whole-genome sequencing was carried out using a MiSeq based approach. Phylogenetic reconstruction and estimation of evolutionary rate and recombination were carried out in BEAST and Recombination Detection Program, respectively. Results: EV-D68 was detected in 21/625 (3.4% enterovirus/rhinovirus PCR positive respiratory samples but in none of the 15 CSF. All the EV-D68 patients were young children (age range: 11.8 – 24.5 months and had moderate respiratory infections. Phylogenetic analysis suggested that the Vietnamese sequences clustered with those from Asian countries, of which 9 fell in the B1 clade, and the remaining sequence was identified within the A2 clade. One intra sub-clade recombination event was detected, representing the second reported recombination within EV-D68. The evolutionary rate of EV-D68 was estimated to be 5.12E-3 substitutions/site/year. Phylogenetic analysis indicated that the virus was imported into Viet Nam in 2008. Conclusions: We have demonstrated for the first time EV-D68 has been circulating at low levels in Viet Nam since 2008, associated with moderate acute respiratory infection in children. EV-D68 in Viet Nam is most closely related to Asian viruses, and clusters

  14. Visual detection of human enterovirus 71 subgenotype C4 and Coxsackievirus A16 by reverse transcription loop-mediated isothermal amplification with the hydroxynaphthol blue dye.

    Science.gov (United States)

    Nie, Kai; Zhang, Yong; Luo, Le; Yang, Meng-Jie; Hu, Xiu-Mei; Wang, Miao; Zhu, Shuang-Li; Han, Feng; Xu, Wen-Bo; Ma, Xue-Jun

    2011-08-01

    A sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for rapid visual detection of human enterovirus 71 subgenotype C4 (EV71-C4) and Coxsackievirus A16 (CVA16) infection, respectively. The reaction was performed in one step in a single tube at 65°C for 60 min with the addition of the hydroxynaphthol blue (HNB) dye prior to amplification. The detection limits of the RT-LAMP assay were 0.33 and 1.58 of a 50% tissue culture infective dose (TCID(50)) per reaction based on 10-fold dilutions of a titrated EV71 or CVA16 strain, respectively. No cross-reaction was observed with Coxsackievirus A (CVA) viruses (CVA2, 4, 5, 7, 9, 10, 14, and 24), Coxsackievirus B (CVB) viruses (CVB1,2,3,4, and 5) or ECHO viruses (ECHO3, 6, 11, and 19). The assay was further evaluated with 47 clinical stool specimens diagnosed previously with EV71, CVA16 or other human enterovirus infections. Virus isolates from stool samples were confirmed by virus neutralization testing and sequencing. RT-LAMP with HNB dye was demonstrated to be a sensitive and cost-effective assay for rapid visual detection of human EV71-C4 and CVA16. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. Peptidyl aldehyde NK-1.8k suppresses enterovirus 71 and enterovirus 68 infection by targeting protease 3C.

    Science.gov (United States)

    Wang, Yaxin; Yang, Ben; Zhai, Yangyang; Yin, Zheng; Sun, Yuna; Rao, Zihe

    2015-05-01

    Enterovirus (EV) is one of the major causative agents of hand, foot, and mouth disease in the Pacific-Asia region. In particular, EV71 causes severe central nervous system infections, and the fatality rates from EV71 infection are high. Moreover, an outbreak of respiratory illnesses caused by an emerging EV, EV68, recently occurred among over 1,000 young children in the United States and was also associated with neurological infections. Although enterovirus has emerged as a considerable global public health threat, no antiviral drug for clinical use is available. In the present work, we screened our compound library for agents targeting viral protease and identified a peptidyl aldehyde, NK-1.8k, that inhibits the proliferation of different EV71 strains and one EV68 strain and that had a 50% effective concentration of 90 nM. Low cytotoxicity (50% cytotoxic concentration, >200 μM) indicated a high selective index of over 2,000. We further characterized a single amino acid substitution inside protease 3C (3C(pro)), N69S, which conferred EV71 resistance to NK-1.8k, possibly by increasing the flexibility of the substrate binding pocket of 3C(pro). The combination of NK-1.8k and an EV71 RNA-dependent RNA polymerase inhibitor or entry inhibitor exhibited a strong synergistic anti-EV71 effect. Our findings suggest that NK-1.8k could potentially be developed for anti-EV therapy. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  16. Rapid degradation of Congo red by molecularly imprinted polypyrrole-coated magnetic TiO2 nanoparticles in dark at ambient conditions

    International Nuclear Information System (INIS)

    Wei, Shoutai; Hu, Xiaolei; Liu, Hualong; Wang, Qiang; He, Chiyang

    2015-01-01

    Highlights: • Molecularly imprinted polypyrrole-coated magnetic TiO 2 catalyst was prepared. • The catalyst degraded Congo red rapidly in dark at ambient conditions. • Degradation mechanism was proposed according to LC–MS analysis. • The catalyst can be easily recycled by a magnet. - Abstract: A novel molecularly imprinted polymer (MIP)-coated magnetic TiO 2 nanocomposite was prepared, using methyl orange (MO) as the dummy template and pyrrole as functional monomer, for degradation of Congo red (CR). The nanocomposite was characterized by Fourier transform infrared spectroscopy, thermo-gravimetric analysis, X-ray diffraction, transmission electron microscopy, and vibrating sample magnetometer. The imprinting efficiency of the imprinted nanoparticles was investigated by static binding test, and their degradation ability toward CR was also studied. Moreover, the effects of pH, temperature, dissolved oxygen and oscillation rate on degradation rate of CR were investigated. Results showed that the imprinted nanocomposite had higher adsorption ability for MO compared with the non-imprinted one. Moreover, it could degrade CR rapidly in dark at room temperature and atmospheric pressure and could be recycled easily by a magnet with a good reusability. A degradation mechanism was proposed according to LC–MS analysis of degradation products of CR. The new imprinted nanoparticles showed high catalytic activity at ambient conditions without light illumination and additional chemicals, and therefore, it can be potentially applied to the rapid, “green” and low-cost degradation of CR in industrial printing and dyeing wastewater

  17. Rapid degradation of Congo red by molecularly imprinted polypyrrole-coated magnetic TiO{sub 2} nanoparticles in dark at ambient conditions

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Shoutai; Hu, Xiaolei; Liu, Hualong; Wang, Qiang; He, Chiyang, E-mail: chiyanghe@hotmail.com

    2015-08-30

    Highlights: • Molecularly imprinted polypyrrole-coated magnetic TiO{sub 2} catalyst was prepared. • The catalyst degraded Congo red rapidly in dark at ambient conditions. • Degradation mechanism was proposed according to LC–MS analysis. • The catalyst can be easily recycled by a magnet. - Abstract: A novel molecularly imprinted polymer (MIP)-coated magnetic TiO{sub 2} nanocomposite was prepared, using methyl orange (MO) as the dummy template and pyrrole as functional monomer, for degradation of Congo red (CR). The nanocomposite was characterized by Fourier transform infrared spectroscopy, thermo-gravimetric analysis, X-ray diffraction, transmission electron microscopy, and vibrating sample magnetometer. The imprinting efficiency of the imprinted nanoparticles was investigated by static binding test, and their degradation ability toward CR was also studied. Moreover, the effects of pH, temperature, dissolved oxygen and oscillation rate on degradation rate of CR were investigated. Results showed that the imprinted nanocomposite had higher adsorption ability for MO compared with the non-imprinted one. Moreover, it could degrade CR rapidly in dark at room temperature and atmospheric pressure and could be recycled easily by a magnet with a good reusability. A degradation mechanism was proposed according to LC–MS analysis of degradation products of CR. The new imprinted nanoparticles showed high catalytic activity at ambient conditions without light illumination and additional chemicals, and therefore, it can be potentially applied to the rapid, “green” and low-cost degradation of CR in industrial printing and dyeing wastewater.

  18. Moving toward rapid and low-cost point-of-care molecular diagnostics with a repurposed 3D printer and RPA.

    Science.gov (United States)

    Chan, Kamfai; Wong, Pui-Yan; Parikh, Chaitanya; Wong, Season

    2018-03-15

    Traditionally, the majority of nucleic acid amplification-based molecular diagnostic tests are done in centralized settings. In recent years, point-of-care tests have been developed for use in low-resource settings away from central laboratories. While most experts agree that point-of-care molecular tests are greatly needed, their availability as cost-effective and easy-to-operate tests remains an unmet goal. In this article, we discuss our efforts to develop a recombinase polymerase amplification reaction-based test that will meet these criteria. First, we describe our efforts in repurposing a low-cost 3D printer as a platform that can carry out medium-throughput, rapid, and high-performing nucleic acid extraction. Next, we address how these purified templates can be rapidly amplified and analyzed using the 3D printer's heated bed or the deconstructed, low-cost thermal cycler we have developed. In both approaches, real-time isothermal amplification and detection of template DNA or RNA can be accomplished using a low-cost portable detector or smartphone camera. Last, we demonstrate the capability of our technologies using foodborne pathogens and the Zika virus. Our low-cost approach does not employ complicated and high-cost components, making it suitable for resource-limited settings. When integrated and commercialized, it will offer simple sample-to-answer molecular diagnostics. Copyright © 2018 Elsevier Inc. All rights reserved.

  19. Synthetic peptides for efficient discrimination of anti-enterovirus antibodies at the serotype level.

    Science.gov (United States)

    Routsias, John G; Mavrouli, Maria D; Antonaki, Georgia; Spanakis, Nikolaos; Tsakris, Athanassios

    2014-08-01

    Enteroviruses are important human pathogens, causing a broad spectrum of diseases from minor common colds to fatal myocarditis. However, certain disease syndromes are caused by one or few serotypes. Serotype identification is difficult due to the laborious neutralization tests that lack of sensitivity, while in commercial ELISAs homotypic antibodies' activities are largely masked by the recognition of genera-specific epitopes by heterotypic antibodies. In the present study homotypic assays were developed with the ability to discriminate different enterovirus serotypes. Seventy-three children sera, positive for IgM antibodies against enterovirus genus and 49 healthy children were examined for the presence of antibodies against 14 synthetic peptides derived from a non-conserved region of the VP1 protein of coxsackieviruses B2, B3, B4, B5, A9, A16, A24, echoviruses 6, 7, 9, 11, 30, enterovirus 71 and parechovirus 1. 50% of the anti-enterovirus IgM positive sera (>150 BU) reacted with the peptides with the majority of them to preferentially recognize one of them, supporting the homotypic nature of our assay. Inhibition studies yielded homologous inhibition rates 67-95% suggesting that specific peptide recognition actually occurred. The diagnostic value of our assay was tested in blood samples drawn over a 1.5-year period from a 5-year old patient. The anti-enterovirus reactivity was clearly attributed to echovirus serotype 11. The IgM/IgG antibody ratio was reversed 4 months later and subsequently IgM antibodies dropped below the cutoff point. In this paper we demonstrate that our assay can be used to discriminate between antibodies targeting different enterovirus serotypes. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Rapid Formation of Molecular Bromine from Deliquesced NaBr Aerosol in the Presence of Ozone and UV Light

    Science.gov (United States)

    The formation of gas-phase bromine from aqueous sodium bromide aerosols is investigated through a combination of chamber experiments and chemical kinetics modeling. Experiments show that Br2(g) is produced rapidly from deliquesced NaBr aerosols in the presence of OH radicals prod...

  1. The survey of porcine teschoviruses, sapeloviruses and enteroviruses B infecting domestic pigs and wild boars in the Czech Republic between 2005 and 2011.

    Science.gov (United States)

    Prodělalová, Jana

    2012-10-01

    This study presents results of epidemiological survey and genetic characterisation of porcine enteric picornaviruses belonging to the genera Teschovirus, Sapelovirus, and Porcine enterovirus B. Faecal or gut content samples from domestic pigs (Sus scrofa f. domestica) and the cecal content of wild boars (Sus scrofa) of different ages (collected between 2005 and 2011) were analysed by molecular methods. Porcine enterovirus B was the most prevalent virus detected in both domestic pigs and wild boars (50.2% and 69.4%, respectively), followed by Porcine teschovirus and Porcine sapelovirus. The majority of positive domestic pigs (69.4%) and wild boars (64.3%) were infected with two or three tested viruses. There was no significant difference in prevalences of teschoviruses, sapeloviruses, and enteroviruses among healthy and diarrhoeic pigs. Results of epidemiological survey demonstrated that all target viral genera are common in Czech farms producing pigs and wild boars. Amplified nucleotide fragments of VP2 region obtained from randomly selected both historical and recent Teschovirus isolates were sequenced. Based on sequence data, historical Porcine teschovirus isolate CAPM V-180, previously determined as serotype 1 was reclassified into serotype 11. Moreover, another recent Porcine teschovirus isolate OH264/2010 was described and classified into serotype 11. Four nontypeable PTV strains (historical isolate CAPM V-182/1976 and recent isolates JA247/2010, NI429/2010, and BR1576/2007) identified in this study might represent novel serotypes. To the best of our knowledge, our study represents the first description of this serotype in the Czech Republic. Copyright © 2012 Elsevier B.V. All rights reserved.

  2. Rapid Analysis of Apolar Low Molecular Weight Constituents in Wood Using High Pressure Liquid Chromatography with Evaporative Light Scattering Detection

    NARCIS (Netherlands)

    Claassen, F.W.; Haar, van de C.; Beek, van T.A.; Dorado, J.; Martinez-Inigo, M.; Sierra-Alvarez, R.

    2000-01-01

    A new high pressure liquid chromatographic method with evaporative light scattering detection was developed for the qualitative and quantitative analysis of apolar, low molecular weight constituents in wood. The wood extractives were obtained by means of a 6 h Soxhlet extraction with acetone. The

  3. Illuminating the Sites of Enterovirus Replication in Living Cells by Using a Split-GFP-Tagged Viral Protein

    NARCIS (Netherlands)

    van der Schaar, H M; Melia, C E; van Bruggen, J A C; Strating, J R P M; van Geenen, M E D; Koster, A J; Bárcena, M; van Kuppeveld, F J M

    2016-01-01

    Like all other positive-strand RNA viruses, enteroviruses generate new organelles (replication organelles [ROs]) with a unique protein and lipid composition on which they multiply their viral genome. Suitable tools for live-cell imaging of enterovirus ROs are currently unavailable, as recombinant

  4. Estimation of contamination sources of human enteroviruses in a wastewater treatment and reclamation system by PCR-DGGE.

    Science.gov (United States)

    Ji, Zheng; Wang, Xiaochang C; Xu, Limei; Zhang, Chongmiao; Funamizu, Naoyuki; Okabe, Satoshi; Sano, Daisuke

    2014-06-01

    A polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was employed to estimate the contamination sources of human enteroviruses and understand how their dominant strains vary in a wastewater treatment and reclamation system consisting of sewage collection, wastewater treatment with membrane bioreactor and open lakes for reclaimed water storage and reuse. After PCR-DGGE using a selected primer set targeting enteroviruses, phylogenetic analysis of acquired enterovirus gene sequences was performed. Enteroviruses identified from the septic tank were much more diverse than those from grey water and kitchen wastewater. Several unique types of enterovirus different from those in wastewater samples were dominant in a biological wastewater treatment unit. Membrane filtration followed by chlorination was proved effective for physically eliminating enteroviruses; however, secondary contamination likely occurred as the reclaimed water was stored in artificial lakes. Enterovirus 71 (EV71), a hand-foot-and-mouth disease (HFMD) viral pathogen, was detected mainly from the artificial lakes, implying that wastewater effluent was not the contamination source of EV71 and that there were unidentified non-point sources of the contamination with the HFMD viral pathogen in the reclaimed water stored in the artificial lakes. The PCR-DGGE targeting enteroviruses provided robust evidence about viral contamination sources in the wastewater treatment and reclamation system.

  5. Molecular evidence of malaria and zoonotic diseases among rapid diagnostic test-negative febrile patients in low-transmission season, Mali

    DEFF Research Database (Denmark)

    Touré, Mahamoudou; Petersen, Pelle T; Bathily, Sidy N'd

    2017-01-01

    From November to December 2012 in Sélingué-Mali, blood samples from 88 febrile patients who tested negative by malaria Paracheck (®) rapid diagnostic tests (RDTs) were used to assess the presence of sub-RDT Plasmodium falciparum as well as Borrelia, Coxiella burnetii, and Babesia applying molecular...... tools. Plasmodium sp. was present among 57 (60.2%) of the 88 malaria RDT-negative patients, whereas the prevalence of Borrelia, C. burnetii, and Babesia were 3.4% (N = 3), 1.1% (N = 1), and 0.0%, respectively. The additional diagnostic use of polymerase chain reaction (PCR) identified a high proportion...

  6. Country-wide surveillance of molecular markers of antimalarial drug resistance in Senegal by use of positive Malaria Rapid Diagnostic Tests

    DEFF Research Database (Denmark)

    Ndiaye, Magatte; Sow, Doudou; Nag, Sidsel

    2017-01-01

    of drug resistance. Therefore, surveillance of drug resistance in the malaria parasites is essential. The objective of this pilot study was to test the feasibility of routinely sampled malaria rapid diagnostic tests (RDTs) at a national scale to assess the temporal changes in the molecular profiles...... of antimalarial drug resistance markers of Plasmodium falciparum parasites. Overall, 9,549 positive malaria RDTs were collected from 14 health facilities across the country. A limited random set of RDTs were analyzed regarding Pfcrt gene polymorphisms at codon 72-76. Overall, a high but varied prevalence (> 50...

  7. Enterovirus infection in febrile neonates: A hospital-based prospective cohort study.

    Science.gov (United States)

    Lv, Xiao-Qing; Qian, Ling-He; Wu, Tai; Yuan, Tian-Ming

    2016-08-01

    This study aims to investigate clinical characteristics and microbiological results and to assess the predictors for enterovirus infection in febrile neonates. A prospective cohort study was conducted on 334 febrile patients (age: 0.33-28 days) in 2011-2012 years. Enterovirus RNA was detected by reverse transcription polymerase chain reaction on faeces or cerebrospinal fluid (CSF). Clinical characteristics were compared, and non-conditional logistic regression analysis was performed to determine independent predictors for enterovirus infection. There were 131 episodes of neonatal enterovirus infection (39.22%). Forty-eight (36.64%) developed respiratory symptoms, 69 (52.67%) had diarrhoea, 22 (16.79%) had poor feeding and 34 (25.95%) had rash. Eighteen (13.74%) had lower platelet counts, and CSF specimens were positive for enterovirus RNA in 44.27% (58/131) whose CSF revealed a mean white blood cell counts of 100.38 ± 147.97 cells/mm(3) (range: 2-668 cells/mm(3) ). The positivity of stool 38.92% (130/334) was significantly higher than that of CSF specimens 26.24% (58/221) for enterovirus RNA (P enterovirus infection: abnormal CSF test (odds ratio (OR): 12.426, 95% confidence interval (CI): 5.633-27.413), thrombocytopenia (OR: 3.647, 95% CI: 1.312-10.136), duration of fever >3.25 (d) (OR: 2.293, 95% CI: 1.279-4.113), highest temperature >38.35 (°C) (OR: 2.094, 95% CI: 1.342-4.123) and negative bacterial culture (OR: 5.073, 95% CI: 1.504-17.114). Our data indicated that enteroviruses should be routinely considered in the differential diagnosis of febrile neonates. The factors, which may predict the risk of neonatal enterovirus infection, were abnormal CSF test, thrombocytopenia, duration of fever >3.25 (d), highest temperature >38.35 (°C) and negative bacterial culture. © 2016 Paediatrics and Child Health Division (The Royal Australasian College of Physicians).

  8. Enterovirus genotypes among patients with severe acute respiratory illness, influenza-like illness, and asymptomatic individuals in South Africa, 2012–2014

    Science.gov (United States)

    Hellferscee, Orienka; Tempia, Stefano; Walaza, Sibongile; Variava, Ebrahim; Dawood, Halima; Wolter, Nicole; Madhi, Shabir A.; du Plessis, Mignon; Cohen, Cheryl; Treurnicht, Florette K.

    2017-01-01

    Enteroviruses can cause outbreaks of severe acute respiratory illness (SARI) and EV-A, -B, -C, and -D species have different pathogenic profiles and circulation patterns. We aimed to characterize and determine the prevalence of enterovirus genotypes among South African patients with respiratory illness and controls during June 2012 to July 2014. Syndromic SARI and influenza-like illness (ILI) surveillance was performed at two sentinel sites. At each site nasopharyngeal/oropharyngeal specimens were collected from SARI and ILI patients as well as controls. Specimens were tested for enterovirus by real-time PCR. Positive specimenswere further genotyped by sequencing a region of theVP1 gene. The prevalence of enterovirus was 5.8% (87/1494), 3.4% (103/3079), and 3.4% (46/1367) among SARI, ILI, and controls, respectively (SARI/controls, P=0.002 and ILI/control, P = 0.973). Among the 101/236 (42.8%) enterovirus-positive specimens that could be genotyped, we observed a high diversity of circulating enterovirus genotypes (a total of 33 genotypes) from all four human enterovirus species with high prevalence of Enterovirus-B (60.4%; 61/101) and Enterovirus-A (21.8%; 22/101) compared to Enterovirus-C (10.9%; 11/101) and Enterovirus-D (6.9%; 7/101) (P = 0.477). Of the enterovirus genotypes identified, Echovirus 30 (9.9%, 10/101), Coxsackie virus B5 (7.9%, 8/101) and Enterovirus-D68 (6.9%, 7/101) were most prevalent. There was no difference in disease severity (SARI or ILI compared to controls) between the different enterovirus species (P = 0.167).We observed a high number of enterovirus genotypes in patients with respiratory illness and in controls from South Africa with no disease association of EV species with disease severity. PMID:28574589

  9. Enterovirus spectrum from the active surveillance of hand foot and mouth disease patients under the clinical trial of inactivated Enterovirus A71 vaccine in Jiangsu, China, 2012-2013.

    Science.gov (United States)

    Yao, Xin; Bian, Lian-Lian; Lu, Wei-Wei; Li, Jing-Xin; Mao, Qun-Ying; Wang, Yi-Ping; Gao, Fan; Wu, Xing; Ye, Qiang; Xu, Miao; Li, Xiu-Ling; Zhu, Feng-Cai; Liang, Zheng-Lun

    2015-12-01

    Epidemiological data from active surveillance on human enterovirus, which could cause hand, foot, and mouth disease, were limited. An active surveillance system was used to investigate the enterovirus spectrum and the incidence of different enteroviruses in infants aged 6-35 months in Jiangsu Province from 2012 to 2013. Fifty-nine infants were randomly selected from 522 non-EV-A71/CV-A16 HFMD patients. We collected 173 throat swabs and 174 rectal swabs from these infants. RT-PCR was used to amplify 5'-UTR and VP1 regions of enteroviruses and the serotypes were determined by the sequence comparison using BLAST. Twenty-one non-EV-A71/CA16 enterovirus serotypes were detected in those infants. E16, E18 were firstly reported in HFMD patients. The four top common non-EV-A71/CV-A enteroviruses among infants were CV-B3, CV-A10, CV-A6, and E9 with the HFMD incidence rates at 1.4%, 0.84%, 0.56%, and 0.47%, respectively. Over 20.8% patients were co-infected with multiple enteroviruses. Neither the course of sickness nor clinical symptoms of the co-infected patients was more severe than those infected with single enterovirus. Two patients were infected different enterovirus successively within 2 months. Several new enterovirus serotypes and multiple models of infection associated with HFMD were discovered through the active surveillance system. These data provide a better understanding of the viral etiology of HFMD. © 2015 Wiley Periodicals, Inc.

  10. Risk of leukaemia in children infected with enterovirus: a nationwide, retrospective, population-based, Taiwanese-registry, cohort study.

    Science.gov (United States)

    Lin, Jiun-Nong; Lin, Cheng-Li; Lin, Ming-Chia; Lai, Chung-Hsu; Lin, Hsi-Hsun; Yang, Chih-Hui; Sung, Fung-Chang; Kao, Chia-Hung

    2015-10-01

    The association between enterovirus infections in children and risk of leukaemia is unclear. We aimed to assess the risk of leukaemia after enterovirus infection in children. We did a nationwide retrospective cohort study by analysing data from the National Health Insurance Research Database (NHIRD) in Taiwan. Children with enterovirus infections aged younger than 18 years were identified. With use of computer-generated random numbers, children not infected with enterovirus were randomly selected and frequency matched (1:1) with children infected with enterovirus by sex, age, urbanisation level, parental occupation, and index year of enterovirus infection. We only included children with complete baseline data for age and sex and who had at least three clinic visits with the diagnosis of enterovirus infection. The diagnosis date of the first clinic visit for the enterovirus infection was defined as the index date for initiation of follow-up person-year measurement and participants. All study patients were followed up until they developed leukaemia, were lost to follow-up, withdrew from the NHI programme, or until the end of the study without leukaemia (censored). Our primary endpoint was a diagnosis of leukaemia during follow-up. Insurance claims data for 3 054 336 children younger than 18 years were randomly selected from all insured children in the NHIRD. We identified 282 360 children infected with enterovirus and 282 355 children not infected with enterovirus between Jan 1, 2000, and Dec 31, 2007. The incidence density rates of leukaemia were 3·26 per 100 000 person-years for the enterovirus-infected and 5·84 per 100 000 person-years for the non-enterovirus-infected cohorts. The risk of leukaemia was significantly lower in the enterovirus-infected cohort than in the non-enterovirus-infected cohort (adjusted subhazard ratio [SHR] 0·44, 95% CI 0·31-0·60; penterovirus have a reduced risk of both lymphocytic leukaemia (adjusted SHR 0·44, 0·30-0

  11. Case report: lymphogranuloma venereum proctitis-from rapid screening to molecular confirmation of a masked sexually transmitted disease.

    Science.gov (United States)

    Markowicz, Mateusz; Grilnberger, Evelyn; Huber, Florian; Leibl, Gabriele; Abrahamian, Heidemarie; Gartner, Manfred; Huber, Monika; Chott, Andreas; Reiter, Michael; Stanek, Gerold

    2013-08-01

    Proctitis caused by Chlamydia trachomatis L2b can manifest with very mild, nonspecific symptoms, and appropriate diagnostic evaluation is crucial. The case report demonstrates that rapid screening test, detection of specific antibodies in serum, and direct pathogen identification by PCR performed on tissue sample or rectal swab allow successful diagnosis of the still emerging sexually transmitted disease among homosexual patients. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Potent antiviral agents fail to elicit genetically-stable resistance mutations in either enterovirus 71 or Coxsackievirus A16.

    Science.gov (United States)

    Kelly, James T; De Colibus, Luigi; Elliott, Lauren; Fry, Elizabeth E; Stuart, David I; Rowlands, David J; Stonehouse, Nicola J

    2015-12-01

    Enterovirus 71 (EV71) and Coxsackievirus A16 (CVA16) are the two major causative agents of hand, foot and mouth disease (HFMD), for which there are currently no licenced treatments. Here, the acquisition of resistance towards two novel capsid-binding compounds, NLD and ALD, was studied and compared to the analogous compound GPP3. During serial passage, EV71 rapidly became resistant to each compound and mutations at residues I113 and V123 in VP1 were identified. A mutation at residue 113 was also identified in CVA16 after passage with GPP3. The mutations were associated with reduced thermostability and were rapidly lost in the absence of inhibitors. In silico modelling suggested that the mutations prevented the compounds from binding the VP1 pocket in the capsid. Although both viruses developed resistance to these potent pocket-binding compounds, the acquired mutations were associated with large fitness costs and reverted to WT phenotype and sequence rapidly in the absence of inhibitors. The most effective inhibitor, NLD, had a very large selectivity index, showing interesting pharmacological properties as a novel anti-EV71 agent. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  13. Adaptation of enterovirus 71 to adult interferon deficient mice.

    Directory of Open Access Journals (Sweden)

    Elizabeth A Caine

    Full Text Available Non-polio enteroviruses, including enterovirus 71 (EV71, have caused severe and fatal cases of hand, foot and mouth disease (HFMD in the Asia-Pacific region. The development of a vaccine or antiviral against these pathogens has been hampered by the lack of a reliable small animal model. In this study, a mouse adapted EV71 strain was produced by conducting serial passages through A129 (α/β interferon (IFN receptor deficient and AG129 (α/β, γ IFN receptor deficient mice. A B2 sub genotype of EV71 was inoculated intraperitoneally (i.p. into neonatal AG129 mice and brain-harvested virus was subsequently passaged through 12 and 15 day-old A129 mice. When tested in 10 week-old AG129 mice, this adapted strain produced 100% lethality with clinical signs including limb paralysis, eye irritation, loss of balance, and death. This virus caused only 17% mortality in same age A129 mice, confirming that in the absence of a functional IFN response, adult AG129 mice are susceptible to infection by adapted EV71 isolates. Subsequent studies in adult AG129 and young A129 mice with the adapted EV71 virus examined the efficacy of an inactivated EV71 candidate vaccine and determined the role of humoral immunity in protection. Passive transfer of rabbit immune sera raised against the EV71 vaccine provided protection in a dose dependent manner in 15 day-old A129 mice. Intramuscular injections (i.m. in five week-old AG129 mice with the alum adjuvanted vaccine also provided protection against the mouse adapted homologous strain. No clinical signs of disease or mortality were observed in vaccinated animals, which received a prime-and-boost, whereas 71% of control animals were euthanized after exhibiting systemic clinical signs (P<0.05. The development of this animal model will facilitate studies on EV71 pathogenesis, antiviral testing, the evaluation of immunogenicity and efficacy of vaccine candidates, and has the potential to establish correlates of protection

  14. Enterovirus serotypes in patients with central nervous system and respiratory infections in Viet Nam 1997-2010.

    Science.gov (United States)

    B'Krong, Nguyen Thi Thuy Chinh; Minh, Ngo Ngoc Quang; Qui, Phan Tu; Chau, Tran Thi Hong; Nghia, Ho Dang Trung; Do, Lien Anh Ha; Nhung, Nguyen Ngoc; Van Vinh Chau, Nguyen; Thwaites, Guy; Van Tan, Le; van Doorn, H Rogier; Thanh, Tran Tan

    2018-04-12

    Enteroviruses are the most common causative agents of human illness. Enteroviruses have been associated with regional and global epidemics, recently, including with severe disease (Enterovirus A71 and D68), and are of interest as emerging viruses. Here, we typed Enterovirus A-D (EV) from central nervous system (CNS) and respiratory infections in Viet Nam. Data and specimens from prospective observational clinical studies conducted between 1997 and 2010 were used. Species and serotypes were determined using type-specific RT-PCR and viral protein 1 or 4 (VP1, VP4) sequencing. Samples from patients with CNS infection (51 children - 10 CSF and 41 respiratory/rectal swabs) and 28 adults (28 CSF) and respiratory infection (124 children - 124 respiratory swabs) were analysed. Twenty-six different serotypes of the four Enterovirus species (A-D) were identified, including EV-A71 and EV-D68. Enterovirus B was associated with viral meningitis in children and adults. Hand, foot and mouth disease associated Enteroviruses A (EV-A71 and Coxsackievirus [CV] A10) were detected in children with encephalitis. Diverse serotypes of all four Enterovirus species were found in respiratory samples, including 2 polio-vaccine viruses, but also 8 CV-A24 and 8 EV-D68. With the exception of EV-D68, the relevance of these viruses in respiratory infection remains unknown. We describe the diverse spectrum of enteroviruses from patients with CNS and respiratory infections in Viet Nam between 1997 and 2010. These data confirm the global circulation of Enterovirus genera and their associations and are important for clinical diagnostics, patient management, and outbreak response.

  15. Application of next-generation sequencing for rapid marker development in molecular plant breeding: a case study on anthracnose disease resistance in Lupinus angustifolius L.

    Directory of Open Access Journals (Sweden)

    Yang Huaan

    2012-07-01

    Full Text Available Abstract Background In the last 30 years, a number of DNA fingerprinting methods such as RFLP, RAPD, AFLP, SSR, DArT, have been extensively used in marker development for molecular plant breeding. However, it remains a daunting task to identify highly polymorphic and closely linked molecular markers for a target trait for molecular marker-assisted selection. The next-generation sequencing (NGS technology is far more powerful than any existing generic DNA fingerprinting methods in generating DNA markers. In this study, we employed a grain legume crop Lupinus angustifolius (lupin as a test case, and examined the utility of an NGS-based method of RAD (restriction-site associated DNA sequencing as DNA fingerprinting for rapid, cost-effective marker development tagging a disease resistance gene for molecular breeding. Results Twenty informative plants from a cross of RxS (disease resistant x susceptible in lupin were subjected to RAD single-end sequencing by multiplex identifiers. The entire RAD sequencing products were resolved in two lanes of the 16-lanes per run sequencing platform Solexa HiSeq2000. A total of 185 million raw reads, approximately 17 Gb of sequencing data, were collected. Sequence comparison among the 20 test plants discovered 8207 SNP markers. Filtration of DNA sequencing data with marker identification parameters resulted in the discovery of 38 molecular markers linked to the disease resistance gene Lanr1. Five randomly selected markers were converted into cost-effective, simple PCR-based markers. Linkage analysis using marker genotyping data and disease resistance phenotyping data on a F8 population consisting of 186 individual plants confirmed that all these five markers were linked to the R gene. Two of these newly developed sequence-specific PCR markers, AnSeq3 and AnSeq4, flanked the target R gene at a genetic distance of 0.9 centiMorgan (cM, and are now replacing the markers previously developed by a traditional DNA

  16. A novel molecularly imprinted material based on magnetic halloysite nanotubes for rapid enrichment of 2,4-dichlorophenoxyacetic acid in water

    International Nuclear Information System (INIS)

    Zhong, Shian; Zhou, Chengyun; Zhang, Xiaona; Zhou, Hui; Li, Hui; Zhu, Xiaohong; Wang, Yan

    2014-01-01

    Graphical abstract: - Highlights: • Successful preparation of a novel type of magnetic halloysite molecularly imprinted material. • Rapid enrichment for 2,4-dichlorophenoxyacetic acid in water. • This material possesses high adsorption capacity and specific recognition to 2,4-dichlorophenoxyacetic acid. • Magnetic halloysite were synthesized by co-precipitation method. - Abstract: A new type of magnetic halloysite nanotubes molecularly imprinted polymer (MHNTs@MIP) based on halloysite nanotubes (HNTs) with embedded magnetic nanoparticles was introduced in this study. MHNTs@MIP was prepared through surface imprinting technology, using 2,4-dichlorophenoxyacetic acid (2,4-D) as a template, 4-vinylpyridine as the monomer, divinylbenzene as cross-linking agents, and 2,2-azodiisobutyronitrile as initiator. MHNTs@MIP was characterized by Fourier Transform Infrared Spectrometer, transmission electron microscopy, X-ray diffraction, and vibrating sample magnetometer. MHNTs@MIP exhibited rapid and reliable analysis with supermagnetic properties, as well as repeated use and template-specific recognition. The adsorption capacity of magnetic halloysite nanotubes non-imprinted polymer (MHNTs@NIP) and MHNTs@MIP was 10.3 mg/g and 35.2 mg/g, respectively. In the detailed discussion on specific selectivity, MHNTs@MIP can be applied as an adsorbent for sample pretreatment extraction and obtain high recoveries of about 85–94%. After extraction, high-performance liquid chromatography was used to detect 2,4-D residue in water

  17. A novel molecularly imprinted material based on magnetic halloysite nanotubes for rapid enrichment of 2,4-dichlorophenoxyacetic acid in water

    Energy Technology Data Exchange (ETDEWEB)

    Zhong, Shian; Zhou, Chengyun; Zhang, Xiaona [School of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China); Zhou, Hui [Cancer Hospital of Xiangya Medical College, Central South University, Changsha 410013 (China); Li, Hui; Zhu, Xiaohong [School of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China); Wang, Yan, E-mail: yanwangcsu@163.com [School of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China)

    2014-07-15

    Graphical abstract: - Highlights: • Successful preparation of a novel type of magnetic halloysite molecularly imprinted material. • Rapid enrichment for 2,4-dichlorophenoxyacetic acid in water. • This material possesses high adsorption capacity and specific recognition to 2,4-dichlorophenoxyacetic acid. • Magnetic halloysite were synthesized by co-precipitation method. - Abstract: A new type of magnetic halloysite nanotubes molecularly imprinted polymer (MHNTs@MIP) based on halloysite nanotubes (HNTs) with embedded magnetic nanoparticles was introduced in this study. MHNTs@MIP was prepared through surface imprinting technology, using 2,4-dichlorophenoxyacetic acid (2,4-D) as a template, 4-vinylpyridine as the monomer, divinylbenzene as cross-linking agents, and 2,2-azodiisobutyronitrile as initiator. MHNTs@MIP was characterized by Fourier Transform Infrared Spectrometer, transmission electron microscopy, X-ray diffraction, and vibrating sample magnetometer. MHNTs@MIP exhibited rapid and reliable analysis with supermagnetic properties, as well as repeated use and template-specific recognition. The adsorption capacity of magnetic halloysite nanotubes non-imprinted polymer (MHNTs@NIP) and MHNTs@MIP was 10.3 mg/g and 35.2 mg/g, respectively. In the detailed discussion on specific selectivity, MHNTs@MIP can be applied as an adsorbent for sample pretreatment extraction and obtain high recoveries of about 85–94%. After extraction, high-performance liquid chromatography was used to detect 2,4-D residue in water.

  18. Effect of rapid thermal annealing on InP1−xBix grown by molecular beam epitaxy

    International Nuclear Information System (INIS)

    Wu, X Y; Wang, K; Pan, W W; Wang, P; Li, Y Y; Song, Y X; Gu, Y; Yue, L; Xu, H; Zhang, Z P; Cui, J; Gong, Q; Wang, S M

    2015-01-01

    The effect of post-growth rapid thermal annealing on structural and optical properties of InP 1−x Bi x thin films was investigated. InPBi shows good thermal stability up to 500 °C and a modest improvement in photoluminescence (PL) intensity with an unchanged PL spectral feature. Bismuth outdiffusion from InPBi and strain relaxation are observed at about 600 °C. The InPBi sample annealed at 800 °C shows an unexpected PL spectrum with different energy transitions. (paper)

  19. The presence of enterovirus, adenovirus, and parvovirus B19 in myocardial tissue samples from autopsies

    DEFF Research Database (Denmark)

    Nielsen, Trine Skov; Hansen, Jakob; Nielsen, Lars Peter

    2014-01-01

    of adenovirus, enterovirus, and parvovirus B19 (PVB) in myocardial autopsy samples from myocarditis related deaths and in non-inflamed control hearts in an effort to clarify their significance as the causes of myocarditis in a forensic material. METHODS: We collected all autopsy cases diagnosed with myocarditis...... from 1992 to 2010. Eighty-four suicidal deaths with morphologically normal hearts served as controls. Polymerase chain reaction was used for the detection of the viral genomes (adenovirus, enterovirus, and PVB) in myocardial tissue specimens. The distinction between acute and persistent PVB infection...... was made by the serological determination of PVB-specific immunoglobulins M and G. RESULTS: PVB was detected in 33 of 112 (29 %) myocarditis cases and 37 of 84 (44 %) control cases. All of the samples were negative for the presence of adenovirus and enterovirus. Serological evidence of an acute PVB...

  20. Onychomadesis outbreak in Valencia, Spain associated with hand, foot, and mouth disease caused by enteroviruses.

    Science.gov (United States)

    Davia, Javier López; Bel, Pablo Hernández; Ninet, Violeta Zaragoza; Bracho, María Alma; González-Candelas, Fernando; Salazar, Antonio; Gobernado, Miguel; Bosch, Isabel Febrer

    2011-01-01

    This report evaluates the June 2008 onychomadesis outbreak in Valencia, Spain. The study sample consisted of 221 onychomadesis cases and 77 nonaffected individuals who lived close to those affected. We collected data on dietary variables, hygiene products, and individual pathological histories. Feces and blood specimens were collected from 44 cases and 24 controls to evaluate exposure to infectious agents. Pathological background data revealed a high frequency (61%) of hand, foot, and mouth disease among the onychomadesis cases. Coxsackievirus A10 was the most commonly detected enterovirus in both case and control groups (49%). Other enteroviruses such as coxsackieviruses A5, A6, A16, B1, and B3; echoviruses 3, 4, and 9; and enterovirus 71 were present in low frequencies in the case and control groups (3-9%). The 2008 onychomadesis outbreak in the metropolitan area of Valencia was associated with an outbreak of hand, foot, and mouth disease primarily caused by coxsackievirus A10. © 2010 Wiley Periodicals, Inc.

  1. Increased Intracranial Pressure in the Setting of Enterovirus and Other Viral Meningitides

    Directory of Open Access Journals (Sweden)

    Jules C. Beal

    2017-01-01

    Full Text Available Increased intracranial pressure due to viral meningitis has not been widely discussed in the literature, although associations with Varicella and rarely Enterovirus have been described. Patients with increased intracranial pressure and cerebrospinal fluid analysis suggestive of a viral process are sometimes classified as having atypical idiopathic intracranial hypertension (IIH. However, a diagnosis of IIH requires normal cerebrospinal fluid, and therefore in these cases an infection with secondary intracranial hypertension may be a more likely diagnosis. Here seven patients are presented with elevated intracranial pressure and cerebrospinal fluid suggestive of viral or aseptic meningitis. Of these, 1 had Enterovirus and the remainder were diagnosed with nonspecific viral meningitis. These data suggest that viral meningitis may be associated with elevated intracranial pressure more often than is commonly recognized. Enterovirus has previously been associated with increased intracranial pressure only in rare case reports.

  2. [Molecular beacon based PNA-FISH method combined with fluorescence scanning for rapid detection of Listeria monocytogenes].

    Science.gov (United States)

    Wu, Shan; Zhang, Xiaofeng; Shuai, Jiangbing; Li, Ke; Yu, Huizhen; Jin, Chenchen

    2016-07-04

    To simplify the PNA-FISH (Peptide nucleic acid-fluorescence in situ hybridization) test, molecular beacon based PNA probe combined with fluorescence scanning detection technology was applied to replace the original microscope observation to detect Listeria monocytogenes The 5′ end and 3′ end of the L. monocytogenes specific PNA probes were labeled with the fluorescent group and the quenching group respectively, to form a molecular beacon based PNA probe. When PNA probe used for fluorescence scanning and N1 treatment as the control, the false positive rate was 11.4%, and the false negative rate was 0; when N2 treatment as the control, the false positive rate decreased to 4.3%, but the false negative rate rose to 18.6%. When beacon based PNA probe used for fluorescence scanning, taken N1 treatment as blank control, the false positive rate was 8.6%, and the false negative rate was 1.4%; taken N2 treatment as blank control, the false positive rate was 5.7%, and the false negative rate was 1.4%. Compared with PNA probe, molecular beacon based PNA probe can effectively reduce false positives and false negatives. The success rates of hybridization of the two PNA probes were 83.3% and 95.2% respectively; and the rates of the two beacon based PNA probes were 91.7% and 90.5% respectively, which indicated that labeling the both ends of the PNA probe dose not decrease the hybridization rate with the target bacteria. The combination of liquid phase PNA-FISH and fluorescence scanning method, can significantly improve the detection efficiency.

  3. Rapid determination of 239Pu in urine samples using molecular recognition technology product AnaLigRPu-02 gel

    International Nuclear Information System (INIS)

    Silvia Dulanska; Boris Remenec; Jan Bilohuscin; Miroslav Labaska; Bianka Horvathova; Andrej Matel

    2013-01-01

    This paper describes the use of IBC's AnaLig R Pu-02 molecular recognition technology product to effectively and selectively pre-concentrate, separate and recover plutonium from urine samples. This method uses two-stage column separations consisting of two different commercial products, Eichrom's Pre-filter Material and AnaLig R Pu-02 resin from IBC Advanced Technologies. By eliminating the co-precipitation techniques and the ashing steps to remove residual organics, the analysis time was reduced significantly. The method was successfully tested by adding known activities of reference solutions of 242 Pu and 239 Pu to urine samples. (author)

  4. Rapid molecular detection of invasive species in ballast and harbor water by integrating environmental DNA and light transmission spectroscopy.

    Science.gov (United States)

    Egan, Scott P; Grey, Erin; Olds, Brett; Feder, Jeffery L; Ruggiero, Steven T; Tanner, Carol E; Lodge, David M

    2015-04-07

    Invasive species introduced via the ballast water of commercial ships cause enormous environmental and economic damage worldwide. Accurate monitoring for these often microscopic and morphologically indistinguishable species is challenging but critical for mitigating damages. We apply eDNA sampling, which involves the filtering and subsequent DNA extraction of microscopic bits of tissue suspended in water, to ballast and harbor water sampled during a commercial ship's 1400 km voyage through the North American Great Lakes. Using a lab-based gel electrophoresis assay and a rapid, field-ready light transmission spectroscopy (LTS) assay, we test for the presence of two invasive species: quagga (Dreissena bugensis) and zebra (D. polymorpha) mussels. Furthermore, we spiked a set of uninfested ballast and harbor samples with zebra mussel tissue to further test each assay's detection capabilities. In unmanipulated samples, zebra mussel was not detected, while quagga mussel was detected in all samples at a rate of 85% for the gel assay and 100% for the LTS assay. In the spiked experimental samples, both assays detected zebra mussel in 94% of spiked samples and 0% of negative controls. Overall, these results demonstrate that eDNA sampling is effective for monitoring ballast-mediated invasions and that LTS has the potential for rapid, field-based detection.

  5. Clinical manifestations of severe enterovirus 71 infection and early assessment in a Southern China population.

    Science.gov (United States)

    Yang, Si-da; Li, Pei-Qing; Li, Yi-Min; Li, Wei; Lai, Wen-Ying; Zhu, Cui-Ping; Tao, Jian-Ping; Deng, Li; Liu, Hong-Sheng; Ma, Wen-Cheng; Lu, Jia-Ming; Hong, Yan; Liang, Yu-Ting; Shen, Jun; Hu, Dan-Dan; Gao, Yuan-Yuan; Zhou, Yi; Situ, Min-Xiong; Chen, Yan-Ling

    2017-02-17

    Enterovirus 71 (EV-A71) shows a potential of rapid death, but the natural history of the infection is poorly known. This study aimed to examine the natural history of EV-A71 infection. This was a prospective longitudinal observational study performed between January 1 st and October 31 st , 2012, at three hospitals in Guangdong, China. Subjects with positive EV-A71 RNA laboratory test results were included. Disease progression was documented with MRI, autopsies, and follow-up. Symptoms/signs with potential association with risk of death were analyzed. Among the 288 patients, neurologic symptoms and signs were observed (emotional movement disorders, dyskinesia, involuntary movements, autonomic dysfunction, and disturbance of consciousness). Some of them occurred as initial symptoms. Myoclonic jerks/tremors were observed among >50% of the patients; nearly 40% of patients presented fatigue and 25% were with vomiting. Twenty-eight patients (9.7%) presented poor peripheral perfusion within 53.4 ± 26.1 h; 23 patients (8.0%) presented pulmonary edema and/or hemorrhage within 62.9 ± 28.6 h. Seventeen (5.9%) patients were in a coma. Seven (2.4%) patients died within 62.9 ± 28.6 h. Seventy-seven survivors underwent head and spinal cord MRI and 37.7% (29/77) showed abnormalities. Two fatal cases showed neuronal necrosis, softening, perivascular cuffing, colloid, and neuronophagia phenomenon in the brainstem. Patients with EV-A71 infection showed high complexity of symptoms and onset timing. Death risk may be indicated by autokinetic eyeball, eyeball ataxia, severe coma, respiratory rhythm abnormality, absent pharyngeal reflex, ultrahyperpyrexia, excessive tachycardia, pulmonary edema and/or hemorrhage, and refractory shock and ataxic respiration. Early assessment of these symptoms/signs is important for proper management.

  6. Cytokine Immunopathogenesis of Enterovirus 71 Brain Stem Encephalitis

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    Shih-Min Wang

    2012-01-01

    Full Text Available Enterovirus 71 (EV71 is one of the most important causes of herpangina and hand, foot, and mouth disease. It can also cause severe complications of the central nervous system (CNS. Brain stem encephalitis with pulmonary edema is the severe complication that can lead to death. EV71 replicates in leukocytes, endothelial cells, and dendritic cells resulting in the production of immune and inflammatory mediators that shape innate and acquired immune responses and the complications of disease. Cytokines, as a part of innate immunity, favor the development of antiviral and Th1 immune responses. Cytokines and chemokines play an important role in the pathogenesis EV71 brain stem encephalitis. Both the CNS and the systemic inflammatory responses to infection play important, but distinctly different, roles in the pathogenesis of EV71 pulmonary edema. Administration of intravenous immunoglobulin and milrinone, a phosphodiesterase inhibitor, has been shown to modulate inflammation, to reduce sympathetic overactivity, and to improve survival in patients with EV71 autonomic nervous system dysregulation and pulmonary edema.

  7. EPA Method 1615. Measurement of Enterovirus and Norovirus ...

    Science.gov (United States)

    A standardized method is required when national studies on virus occurrence in environmental and drinking waters utilize multiple analytical laboratories. The U.S Environmental Protection Agency’s (USEPA) Method 1615 was developed with the goal of providing such a standard for measuring Enterovirus and Norovirus in these waters. Virus is concentrated from water using an electropositive filter, eluted from the filter surface with beef extract, and then concentrated further using organic flocculation. Herein we present the protocol from Method 1615 for filter elution, secondary concentration, and measurement of total culturable viruses. A portion of the concentrated eluate from each sample is inoculated onto ten replicate flasks of Buffalo Green Monkey kidney cells. The number of flasks demonstrating cytopathic effects is used to quantify the most probable number (MPN) of infectious units per liter. The method uses a number of quality controls to increase data quality and to reduce interlaboratory and intralaboratory variation. Laboratories must meet defined performance standards. Method 1615 was evaluated by examining virus recovery from reagent-grade and ground waters seeded with Sabin poliovirus type 3. Mean poliovirus recoveries with the total culturable assay were 111% in reagent grade water and 58% in groundwaters. EPA Method 1615 is being used by a number of national and international labs. This paper and the accompanying video will provide training oppo

  8. 2014 outbreak of enterovirus D68 in North America.

    Science.gov (United States)

    Messacar, Kevin; Abzug, Mark J; Dominguez, Samuel R

    2016-05-01

    Enterovirus D68 (EV-D68) is an emerging picornavirus which causes severe respiratory disease, predominantly in children. In 2014, the largest and most widespread outbreak of EV-D68 described to date was reported in North America. Hospitals throughout the United States and Canada reported surges in patient volumes and resource utilization from August to October, 2014. In the US a total of 1,153 infections were confirmed in 49 states, although this is an underestimate of the likely millions of cases that occurred but were not tested. EV-D68 was detected in 14 patients who died; the role of the virus in these deaths is unknown. A possible association between EV-D68 and cases of acute flaccid paralysis with spinal cord gray matter lesions, known as acute flaccid myelitis, was observed during the outbreak and is under investigation. The 2014 outbreak of EV-D68 in North America demonstrates the public health importance of this emerging pathogen. © 2015 Wiley Periodicals, Inc.

  9. Antiviral and Inflammatory Cellular Signaling Associated with Enterovirus 71 Infection

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    Yuefei Jin

    2018-03-01

    Full Text Available Enterovirus 71 (EV71 infection has become a major threat to global public health, especially in infants and young children. Epidemiological studies have indicated that EV71 infection is responsible for severe and even fatal cases of hand, foot, and mouth disease (HFMD. Accumulated evidence indicates that EV71 infection triggers a plethora of interactive signaling pathways, resulting in host immune evasion and inflammatory response. This review mainly covers the effects of EV71 infection on major antiviral and inflammatory cellular signal pathways. EV71 can activate cellular signaling networks including multiple cell surface and intracellular receptors, intracellular kinases, calcium flux, and transcription factors that regulate antiviral innate immunity and inflammatory response. Cellular signaling plays a critical role in the regulation of host innate immune and inflammatory pathogenesis. Elucidation of antiviral and inflammatory cellular signaling pathways initiated by EV71 will not only help uncover the potential mechanisms of EV71 infection-induced pathogenesis, but will also provide clues for the design of therapeutic strategies against EV71 infection.

  10. Animal models of enterovirus 71 infection: applications and limitations

    Science.gov (United States)

    2014-01-01

    Human enterovirus 71 (EV71) has emerged as a neuroinvasive virus that is responsible for several outbreaks in the Asia-Pacific region over the past 15 years. Appropriate animal models are needed to understand EV71 neuropathogenesis better and to facilitate the development of effective vaccines and drugs. Non-human primate models have been used to characterize and evaluate the neurovirulence of EV71 after the early outbreaks in late 1990s. However, these models were not suitable for assessing the neurovirulence level of the virus and were associated with ethical and economic difficulties in terms of broad application. Several strategies have been applied to develop mouse models of EV71 infection, including strategies that employ virus adaption and immunodeficient hosts. Although these mouse models do not closely mimic human disease, they have been applied to determine the pathogenesis of and treatment and prevention of the disease. EV71 receptor-transgenic mouse models have recently been developed and have significantly advanced our understanding of the biological features of the virus and the host-parasite interactions. Overall, each of these models has advantages and disadvantages, and these models are differentially suited for studies of EV71 pathogenesis and/or the pre-clinical testing of antiviral drugs and vaccines. In this paper, we review the characteristics, applications and limitation of these EV71 animal models, including non-human primate and mouse models. PMID:24742252

  11. Recent developments in antiviral agents against enterovirus 71 infection.

    Science.gov (United States)

    Tan, Chee Wah; Lai, Jeffrey Kam Fatt; Sam, I-Ching; Chan, Yoke Fun

    2014-02-12

    Enterovirus 71 (EV-71) is the main etiological agent of hand, foot and mouth disease (HFMD). Recent EV-71 outbreaks in Asia-Pacific were not limited to mild HFMD, but were associated with severe neurological complications such as aseptic meningitis and brainstem encephalitis, which may lead to cardiopulmonary failure and death. The absence of licensed therapeutics for clinical use has intensified research into anti-EV-71 development. This review highlights the potential antiviral agents targeting EV-71 attachment, entry, uncoating, translation, polyprotein processing, virus-induced formation of membranous RNA replication complexes, and RNA-dependent RNA polymerase. The strategies for antiviral development include target-based synthetic compounds, anti-rhinovirus and poliovirus libraries screening, and natural compound libraries screening. Growing knowledge of the EV-71 life cycle will lead to successful development of antivirals. The continued effort to develop antiviral agents for treatment is crucial in the absence of a vaccine. The coupling of antivirals with an effective vaccine will accelerate eradication of the disease.

  12. Leishmania OligoC-TesT as a simple, rapid, and standardized tool for molecular diagnosis of cutaneous leishmaniasis in Peru.

    Science.gov (United States)

    Espinosa, Diego; Boggild, Andrea K; Deborggraeve, Stijn; Laurent, Thierry; Valencia, Cristian; Pacheco, Rosa; Miranda-Verástegui, César; Llanos-Cuentas, Alejandro; Leclipteux, Thierry; Dujardin, Jean-Claude; Büscher, Philippe; Arévalo, Jorge

    2009-08-01

    Molecular methods such as PCR have become attractive tools for diagnosis of cutaneous leishmaniasis (CL), both for their high sensitivity and for their specificity. However, their practical use in routine diagnosis is limited due to the infrastructural requirements and the lack of any standardization. Recently, a simplified and standardized PCR format for molecular detection of Leishmania was developed. The Leishmania OligoC-TesT is based on simple and rapid detection using a dipstick with PCR-amplified Leishmania DNA. In this study, we estimated the diagnostic accuracy of the Leishmania OligoC-TesT for 61 specimens from 44 CL-suspected patients presenting at the leishmaniasis clinic of the Instituto de Medicina Tropical Alexander von Humboldt, Peru. On the basis of parasitological detection and the leishmanin skin test (LST), patients were classified as (i) confirmed CL cases, (ii) LST-positive cases, and (iii) LST-negative cases. The sensitivities of the Leishmania OligoC-TesT was 74% (95% confidence interval (CI), 60.5% to 84.1%) for lesion aspirates and 92% (95% CI, 81.2% to 96.9%) for scrapings. A significantly higher sensitivity was observed with a conventional PCR targeting the kinetoplast DNA on the aspirates (94%) (P = 0.001), while there was no significant difference in sensitivity for the lesion scrapings (88%) (P = 0.317). In addition, the Leishmania OligoC-TesT was evaluated for 13 CL-suspected patients in two different peripheral health centers in the central jungle of Peru. Our findings clearly indicate the high accuracy of the Leishmania OligoC-TesT for lesion scrapings for simple and rapid molecular diagnosis of CL in Peru.

  13. Leishmania OligoC-TesT as a Simple, Rapid, and Standardized Tool for Molecular Diagnosis of Cutaneous Leishmaniasis in Peru▿

    Science.gov (United States)

    Espinosa, Diego; Boggild, Andrea K.; Deborggraeve, Stijn; Laurent, Thierry; Valencia, Cristian; Pacheco, Rosa; Miranda-Verástegui, César; Llanos-Cuentas, Alejandro; Leclipteux, Thierry; Dujardin, Jean-Claude; Büscher, Philippe; Arévalo, Jorge

    2009-01-01

    Molecular methods such as PCR have become attractive tools for diagnosis of cutaneous leishmaniasis (CL), both for their high sensitivity and for their specificity. However, their practical use in routine diagnosis is limited due to the infrastructural requirements and the lack of any standardization. Recently, a simplified and standardized PCR format for molecular detection of Leishmania was developed. The Leishmania OligoC-TesT is based on simple and rapid detection using a dipstick with PCR-amplified Leishmania DNA. In this study, we estimated the diagnostic accuracy of the Leishmania OligoC-TesT for 61 specimens from 44 CL-suspected patients presenting at the leishmaniasis clinic of the Instituto de Medicina Tropical Alexander von Humboldt, Peru. On the basis of parasitological detection and the leishmanin skin test (LST), patients were classified as (i) confirmed CL cases, (ii) LST-positive cases, and (iii) LST-negative cases. The sensitivities of the Leishmania OligoC-TesT was 74% (95% confidence interval (CI), 60.5% to 84.1%) for lesion aspirates and 92% (95% CI, 81.2% to 96.9%) for scrapings. A significantly higher sensitivity was observed with a conventional PCR targeting the kinetoplast DNA on the aspirates (94%) (P = 0.001), while there was no significant difference in sensitivity for the lesion scrapings (88%) (P = 0.317). In addition, the Leishmania OligoC-TesT was evaluated for 13 CL-suspected patients in two different peripheral health centers in the central jungle of Peru. Our findings clearly indicate the high accuracy of the Leishmania OligoC-TesT for lesion scrapings for simple and rapid molecular diagnosis of CL in Peru. PMID:19553579

  14. Tyrosine sulfation of the amino terminus of PSGL-1 is critical for enterovirus 71 infection.

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    Yorihiro Nishimura

    Full Text Available Enterovirus 71 (EV71 is one of the major causative agents of hand, foot, and mouth disease, a common febrile disease in children; however, EV71 has been also associated with various neurological diseases including fatal cases in large EV71 outbreaks particularly in the Asia Pacific region. Recently we identified human P-selectin glycoprotein ligand-1 (PSGL-1 as a cellular receptor for entry and replication of EV71 in leukocytes. PSGL-1 is a sialomucin expressed on the surface of leukocytes, serves as a high affinity counterreceptor for selectins, and mediates leukocyte rolling on the endothelium. The PSGL-1-P-selectin interaction requires sulfation of at least one of three clustered tyrosines and an adjacent O-glycan expressing sialyl Lewis x in an N-terminal region of PSGL-1. To elucidate the molecular basis of the PSGL-1-EV71 interaction, we generated a series of PSGL-1 mutants and identified the post-translational modifications that are critical for binding of PSGL-1 to EV71. We expressed the PSGL-1 mutants in 293T cells and the transfected cells were assayed for their abilities to bind to EV71 by flow cytometry. We found that O-glycosylation on T57, which is critical for PSGL-1-selectin interaction, is not necessary for PSGL-1 binding to EV71. On the other hand, site-directed mutagenesis at one or more potential tyrosine sulfation sites in the N-terminal region of PSGL-1 significantly impaired PSGL-1 binding to EV71. Furthermore, an inhibitor of sulfation, sodium chlorate, blocked the PSGL-1-EV71 interaction and inhibited PSGL-1-mediated viral replication of EV71 in Jurkat T cells in a dose-dependent manner. Thus, the results presented in this study reveal that tyrosine sulfation, but not O-glycosylation, in the N-terminal region of PSGL-1 may facilitate virus entry and replication of EV71 in leukocytes.

  15. A mouse model of paralytic myelitis caused by enterovirus D68.

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    Alison M Hixon

    2017-02-01

    Full Text Available In 2014, the United States experienced an epidemic of acute flaccid myelitis (AFM cases in children coincident with a nationwide outbreak of enterovirus D68 (EV-D68 respiratory disease. Up to half of the 2014 AFM patients had EV-D68 RNA detected by RT-PCR in their respiratory secretions, although EV-D68 was only detected in cerebrospinal fluid (CSF from one 2014 AFM patient. Given previously described molecular and epidemiologic associations between EV-D68 and AFM, we sought to develop an animal model by screening seven EV-D68 strains for the ability to induce neurological disease in neonatal mice. We found that four EV-D68 strains from the 2014 outbreak (out of five tested produced a paralytic disease in mice resembling human AFM. The remaining 2014 strain, as well as 1962 prototype EV-D68 strains Fermon and Rhyne, did not produce, or rarely produced, paralysis in mice. In-depth examination of the paralysis caused by a representative 2014 strain, MO/14-18947, revealed infectious virus, virion particles, and viral genome in the spinal cords of paralyzed mice. Paralysis was elicited in mice following intramuscular, intracerebral, intraperitoneal, and intranasal infection, in descending frequency, and was associated with infection and loss of motor neurons in the anterior horns of spinal cord segments corresponding to paralyzed limbs. Virus isolated from spinal cords of infected mice transmitted disease when injected into naïve mice, fulfilling Koch's postulates in this model. Finally, we found that EV-D68 immune sera, but not normal mouse sera, protected mice from development of paralysis and death when administered prior to viral challenge. These studies establish an experimental model to study EV-D68-induced myelitis and to better understand disease pathogenesis and develop potential therapies.

  16. A Novel Recombinant Enterovirus Type EV-A89 with Low Epidemic Strength in Xinjiang, China.

    Science.gov (United States)

    Fan, Qin; Zhang, Yong; Hu, Lan; Sun, Qiang; Cui, Hui; Yan, Dongmei; Sikandaner, Huerxidan; Tang, Haishu; Wang, Dongyan; Zhu, Zhen; Zhu, Shuangli; Xu, Wenbo

    2015-12-21

    Enterovirus A89 (EV-A89) is a novel member of the EV-A species. To date, only one full-length genome sequence (the prototype strain) has been published. Here, we report the molecular identification and genomic characterization of a Chinese EV-A89 strain, KSYPH-TRMH22F/XJ/CHN/2011, isolated in 2011 from a contact of an acute flaccid paralysis (AFP) patient during AFP case surveillance in Xinjiang China. This was the first report of EV-A89 in China. The VP1 coding sequence of this strain demonstrated 93.2% nucleotide and 99.3% amino acid identity with the EV-A89 prototype strain. In the P2 and P3 regions, the Chinese EV-A89 strain demonstrated markedly higher identity than the prototype strains of EV-A76, EV-A90, and EV-A91, indicating that one or more recombination events between EV-A89 and these EV-A types might have occurred. Long-term evolution of these EV types originated from the same ancestor provides the spatial and temporal circumstances for recombination to occur. An antibody sero-prevalence survey against EV-A89 in two Xinjiang prefectures demonstrated low positive rates and low titres of EV-A89 neutralization antibody, suggesting limited range of transmission and exposure to the population. This study provides a solid foundation for further studies on the biological and pathogenic properties of EV-A89.

  17. Enterovirus D68 detection in respiratory specimens: Association with severe disease.

    Science.gov (United States)

    Engelmann, Ilka; Fatoux, Marie; Lazrek, Mouna; Alidjinou, Enagnon K; Mirand, Audrey; Henquell, Cécile; Dewilde, Anny; Hober, Didier

    2017-07-01

    Molecular techniques increased the number of documented respiratory infections. In a substantial number of cases the causative agent remains undetected. Since August 2014, an increase in Enterovirus(EV)-D68 infections was reported. We aimed to investigate epidemiology and clinical relevance of EV-D68. From June to December 2014 and from September to December 2015, 803 and 847 respiratory specimens, respectively, were tested for respiratory viruses with a multiplex RT-PCR. This multiplex RT-PCR does not detect EV-D68. Therefore, 457 (2014) and 343 (2015) specimens with negative results were submitted to an EV-specific-RT-PCR. EV-positive specimens were tested with an EV-D68-specific-RT-PCR and genotyped. Eleven specimens of 2014 tested positive in the EV-specific-RT-PCR and of these seven were positive in the EV-D68-specific-RT-PCR. Typing confirmed these as EV-D68. Median age of EV-D68-positive patients was 3 years (1 month-91 years). Common symptoms included fever (n = 6, 86%), respiratory distress (n = 5, 71%), and cough (n = 4, 57%). All EV-D68-positive patients were admitted to hospital, 4 (57%) were admitted to intensive care units and 6 (86%) received oxygen. One patient suffered from acute flaccid paralysis. Seven specimens of 2015 were positive in the EV-specific-RT-PCR but negative in the EV-D68-specific-RT-PCR. In conclusion, use of an EV-specific-RT-PCR allowed us to detect EV-D68 circulation in autumn 2014 that was not detected by the multiplex RT-PCR and was associated with severe disease. © 2017 Wiley Periodicals, Inc.

  18. Transmission patterns of human enterovirus 71 to, from and among European countries, 2003 to 2013.

    Science.gov (United States)

    Hassel, Chervin; Mirand, Audrey; Lukashev, Alexander; TerletskaiaLadwig, Elena; Farkas, Agnes; Schuffenecker, Isabelle; Diedrich, Sabine; Huemer, Hartwig P; Archimbaud, Christine; Peigue-Lafeuille, Hélène; Henquell, Cécile; Bailly, Jean-Luc

    2015-01-01

    Enterovirus 71 (EV-71) is involved in epidemics of hand, foot, and mouth disease (HFMD) and has been reported to occur with severe neurological complications in eastern and south-east Asia. In other geographical areas, the transmission of this virus is poorly understood. We used large sequence datasets (of the gene encoding the viral protein 1, VP1) and a Bayesian phylogenetic approach to compare the molecular epidemiology and geographical spread patterns of EV-71 subgenogroups B4, B5, C1, C2, and C4 in Europe relative to other parts of the world. For the study, European countries considered were European Union (EU) Member States and Iceland, Norway and Switzerland. Viruses of the B4, B5, and C4 subgenogroups circulate mainly in eastern and south-east Asia. In Europe sporadic introductions of these subgenogroups are observed, however C1 and C2 viruses predominate. The phylogenies showed evidence of multiple events of spread involving C1 and C2 viruses within Europe since the mid-1990s. Two waves of sporadic C2 infections also occurred in 2010 and 2013. The 2007 Dutch outbreak caused by C2 and the occurrence of B5 and C4 infections in the EU between 2004 and 2013 arose while the circulation of C1 viruses was low. A transmission chain involving a C4 virus was traced from Japan to the EU and then further to Canada between 2001 and 2006. Recent events whereby spread of viruses have occurred from, to, and within Europe appear to be involved in the long term survival of EV-71, highlighting the need for enhanced surveillance of this virus.

  19. Molecularly imprinted polymer cartridges coupled on-line with high performance liquid chromatography for simple and rapid analysis of dextromethorphan in human plasma samples.

    Science.gov (United States)

    Moein, Mohammad Mahdi; Javanbakht, Mehran; Akbari-Adergani, Behrouz

    2011-04-01

    In this paper, a novel method is described for automated determination of dextromethorphan in biological fluids using molecularly imprinted solid-phase extraction (MISPE) as a sample clean-up technique combined with high performance liquid chromatography (HPLC). The water-compatible molecularly imprinted polymers (MIPs) were prepared using methacrylic acid as functional monomer, ethylene glycol dimethacrylate as cross-linker, chloroform as porogen and dextromethorphan as template molecule. These imprinted polymers were used as solid-phase extraction sorbent for the extraction of dextromethorphan from human plasma samples. Various parameters affecting the extraction efficiency of the MIP cartridges were evaluated. The high selectivity of the sorbent coupled to the high performance liquid chromatographic system permitted a simple and rapid analysis of this drug in plasma samples with limits of detection (LOD) and quantification (LOQ) of 0.12 ng/mL and 0.35 ng/mL, respectively. The MIP selectivity was evaluated by analyzing of the dextromethorphan in presence of several substances with similar molecular structures and properties. Results from the HPLC analyses showed that the recoveries of dextromethorphan using MIP cartridges from human plasma samples in the range of 1-50 ng/mL were higher than 87%. Copyright © 2011 Elsevier B.V. All rights reserved.

  20. Molecular epidemiology of human rhinoviruses

    OpenAIRE

    Savolainen-Kopra, Carita

    2006-01-01

    The first part of this work investigates the molecular epidemiology of a human enterovirus (HEV), echovirus 30 (E-30). This project is part of a series of studies performed in our research team analyzing the molecular epidemiology of HEV-B viruses. A total of 129 virus strains had been isolated in different parts of Europe. The sequence analysis was performed in three different genomic regions: 420 nucleotides (nt) in the VP4/VP2 capsid protein coding region, the entire VP1 capsid protein cod...

  1. European surveillance for enterovirus D68 during the emerging North-American outbreak in 2014

    DEFF Research Database (Denmark)

    Poelman, Randy; Schuffenecker, Isabelle; Van Leer-Buter, Coretta

    2015-01-01

    BACKGROUND: In August and September 2014, unexpected clusters of enterovirus-D68 (EV-D68) infections associated with severe respiratory disease emerged from North-America. In September, the European Centre for Disease Prevention and Control (ECDC) asked European countries to strengthen respiratory...... sample screening for enterovirus detection and typing in cases with severe respiratory presentations. OBJECTIVES: To provide a detailed picture of EV-D68 epidemiology in Europe by conducting a retrospective and prospective laboratory analysis of clinical specimens. STUDY DESIGN: An initiative supported...

  2. Shift of Enterovirus species among children in Cameroon--identification of a new enterovirus, EV-A119.

    Science.gov (United States)

    Ayukekbong, James; Kabayiza, Jean-Claude; Lindh, Magnus; Nkuo-Akenji, Theresia; Tah, Ferdinand; Bergström, Tomas; Norder, Helene

    2013-09-01

    Infections caused by human enteroviruses (EVs) are often asymptomatic or mild, although they may cause more severe illnesses as meningitis and acute flaccid paralysis. EVs have globally posed a threat to children, and outbreaks of aseptic meningitis and hand, foot and mouth disease are frequently reported. To identify EV strains circulating among healthy children in a small community in Limbe, Cameroon two years apart. Species and EV types were obtained by partial 5'UTR-VP4 and VP1 sequencing of RNA from stool samples collected in October 2009 and September 2011 from 150 children in Cameroon. In all, 74 children (49%) were infected with 28 different types of EV. There were 29 (54%) infected children in 2009, and 45 (47%) in 2011. There was a significant difference between detected species of EV, with 15 (47%) children infected with EV-A in 2009, and 22 (71%) with EV-B in 2011 (p=0.0001). In 2009, one child was infected by a divergent EV, which was most similar to EV-A90. Based on the complete VP1 sequence, it was shown to be a new EV designated EV-A119. The current study shows a high heterogeneity of circulating EV types among children in Limbe, Cameroon, and a previously not described shift in predominating EV species. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Detection of enterovirus 71 gene from clinical specimens by reverse-transcription loop-mediated isothermal amplification.

    Science.gov (United States)

    Wang, D; Wang, X; Geng, Y; An, C

    2014-01-01

    The objective of this study was to develop a sensitive, specific and rapid approach to diagnose hand foot and mouth disease (HFMD) for an early treatment by using loop-mediated isothermal amplification (LAMP) technique. A reverse-transcription loop-mediated isothermal amplification (RT-LAMP) for detecting EV71 virus was developed, the specificity and sensitivity of RT-LAMP was tested, and the clinical specimens was assayed by the RT-LAMP comparing with conventional reverse-transcription polymerase chain reaction (RT-PCR) and real-time PCR. A total of 116 clinical specimens from the suspected HFMD individual were detected with the RT-LAMP. The detection rate for EV71 was 56.89% by RT-LAMP, 41.38% by real-time PCR and 34.48% by RT-PCR. The minimum detection limit of RT-LAMP was 0.01 PFU, both of RT-PCR and real-time PCR was 0.1PFU. Non-cross-reactive amplification with other enteroviruses was detected in the survey reports. The effectiveness of RT-LAMP is higher than RT-PCR and real-time PCR. The protocol is easy to operate and time saving. It was not an expensive instrument, which was needed; it is an applicable method for rapid diagnosis of the disease, especially in resource-poor countries or in developing countries.

  4. Detection of Enterovirus 71 gene from clinical specimens by reverse-transcription loop-mediated isothermal amplification

    Directory of Open Access Journals (Sweden)

    D Wang

    2014-01-01

    Full Text Available Purpose : The objective of this study was to develop a sensitive, specific and rapid approach to diagnose hand foot and mouth disease (HFMD for an early treatment by using loop-mediated isothermal amplification (LAMP technique. Materials and Methods : A reverse-transcription loop-mediated isothermal amplification (RT-LAMP for detecting EV71 virus was developed, the specificity and sensitivity of RT-LAMP was tested, and the clinical specimens was assayed by the RT-LAMP comparing with conventional reverse-transcription polymerase chain reaction (RT-PCR and real-time PCR. Results : A total of 116 clinical specimens from the suspected HFMD individual were detected with the RT-LAMP. The detection rate for EV71 was 56.89% by RT-LAMP, 41.38% by real-time PCR and 34.48% by RT-PCR. The minimum detection limit of RT-LAMP was 0.01 PFU, both of RT-PCR and real-time PCR was 0.1PFU. Non-cross-reactive amplification with other enteroviruses was detected in the survey reports. Conclusions : The effectiveness of RT-LAMP is higher than RT-PCR and real-time PCR. The protocol is easy to operate and time saving. It was not an expensive instrument, which was needed; it is an applicable method for rapid diagnosis of the disease, especially in resource-poor countries or in developing countries.

  5. Screening and rapid molecular diagnosis of tuberculosis in prisons in Russia and Eastern Europe: a cost-effectiveness analysis.

    Directory of Open Access Journals (Sweden)

    Daniel E Winetsky

    Full Text Available Prisons of the former Soviet Union (FSU have high rates of multidrug-resistant tuberculosis (MDR-TB and are thought to drive general population tuberculosis (TB epidemics. Effective prison case detection, though employing more expensive technologies, may reduce long-term treatment costs and slow MDR-TB transmission.We developed a dynamic transmission model of TB and drug resistance matched to the epidemiology and costs in FSU prisons. We evaluated eight strategies for TB screening and diagnosis involving, alone or in combination, self-referral, symptom screening, mass miniature radiography (MMR, and sputum PCR with probes for rifampin resistance (Xpert MTB/RIF. Over a 10-y horizon, we projected costs, quality-adjusted life years (QALYs, and TB and MDR-TB prevalence. Using sputum PCR as an annual primary screening tool among the general prison population most effectively reduced overall TB prevalence (from 2.78% to 2.31% and MDR-TB prevalence (from 0.74% to 0.63%, and cost US$543/QALY for additional QALYs gained compared to MMR screening with sputum PCR reserved for rapid detection of MDR-TB. Adding sputum PCR to the currently used strategy of annual MMR screening was cost-saving over 10 y compared to MMR screening alone, but produced only a modest reduction in MDR-TB prevalence (from 0.74% to 0.69% and had minimal effect on overall TB prevalence (from 2.78% to 2.74%. Strategies based on symptom screening alone were less effective and more expensive than MMR-based strategies. Study limitations included scarce primary TB time-series data in FSU prisons and uncertainties regarding screening test characteristics.In prisons of the FSU, annual screening of the general inmate population with sputum PCR most effectively reduces TB and MDR-TB prevalence, doing so cost-effectively. If this approach is not feasible, the current strategy of annual MMR is both more effective and less expensive than strategies using self-referral or symptom screening alone

  6. Redundancy and molecular evolution: the rapid Induction of bone formation by the mammalian transforming growth factor-β3 isoform

    Directory of Open Access Journals (Sweden)

    Ugo Ripamonti

    2016-09-01

    Full Text Available The soluble osteogenic molecular signals of the transforming growth factor-β (TGF-β supergene family are the molecular bases of the induction of bone formation and postnatal bone tissue morphogenesis with translation into clinical contexts. The mammalian TGF-β3 isoform, a pleiotropic member of the family, controls a vast array of biological processes including the induction of bone formation. Recombinant hTGF-β3 induces substantial bone formation when implanted with either collagenous bone matrices or coral-derived macroporous bioreactors in the rectus abdominis muscle of the non-human primate Papio ursinus. In marked contrast, the three mammalian TGF-βs do not initiate the induction of bone formation in rodents and lagomorphs. The induction of bone by hTGF-β3/preloaded bioreactors is orchestrated by inducing fibrin-fibronectin rings that structurally organize tissue patterning and morphogenesis within the macroporous spaces. Induced advancing extracellular matrix rings provide the structural anchorage for hyper chromatic cells, interpreted as differentiating osteoblasts re-programmed by hTGF-β3 from invading myoblastic and/or pericytic differentiated cells. Runx2 and Osteocalcin expression are significantly up-regulated correlating to multiple invading cells differentiating into the osteoblastic phenotype. Bioreactors pre-loaded with recombinant human Noggin (hNoggin, a BMPs antagonist, show down-regulation of BMP-2 and other profiled osteogenic proteins’ genes resulting in minimal bone formation. Coral-derived macroporous constructs preloaded with binary applications of hTGF-β3 and hNoggin also show down-regulation of BMP-2 with the induction of limited bone formation. The induction of bone formation by hTGF-β3 is via the BMPs pathway and it is thus blocked by hNoggin. Our systematic studies in Papio ursinus with translational hTGF-β3 in large cranio-mandibulo-facial defects in humans are now requesting the re-evaluation of Bone

  7. Rapid detection of malachite green in fish based on CdTe quantum dots coated with molecularly imprinted silica.

    Science.gov (United States)

    Wu, Le; Lin, Zheng-Zhong; Zhong, Hui-Ping; Peng, Ai-Hong; Chen, Xiao-Mei; Huang, Zhi-Yong

    2017-08-15

    A sensitive fluorescence sensor for the detection of malachite green (MG) was fabricated by grafting molecularly imprinted polymers (MIPs) onto the surface of CdTe quantum dots (QDs). The MIP-coated QDs were synthesized via a reverse microemulsion method using (3-aminopropyl)triethoxysilane (APTES) and tetraethyl orthosilicate (TEOS) as functional monomer and cross-linker, respectively. The optimum molar ratio of MG, functional monomer and cross-linker was 1:3:10. The MIP-coated QDs exhibited uniform spheres with diameter around 49nm and excellent fluorescence emission at λ ex 370nm. A linear relationship with two segments between the relative fluorescence intensities and the MG concentrations ranging from 0.08 to 20μmol·L -1 could be obtained with a detection limit of 12μg·kg -1 . The fluorescent probe was successfully applied to the determination of MG in fish samples with the spiked recoveries ranging from 94.3% to 109.5% which were in accordance with those of the measurement by HPLC-UV. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. A molecular toolbox for rapid generation of viral vectors to up- or down-regulate in vivo neuronal gene expression

    Directory of Open Access Journals (Sweden)

    Melanie D. White

    2011-07-01

    Full Text Available We introduce a molecular toolbox for manipulation of neuronal gene expression in vivo. The toolbox includes promoters, ion channels, optogenetic tools, fluorescent proteins and intronic artificial microRNAs. The components are easily assembled into adeno-associated virus (AAV or lentivirus vectors using recombination cloning. We demonstrate assembly of toolbox components into lentivirus and AAV vectors and use these vectors for in vivo expression of inwardly rectifying potassium channels (Kir2.1, Kir3.1 and Kir3.2 and an artificial microRNA targeted against the ion channel HCN1 (HCN1 miR. We show that AAV assembled to express HCN1 miR produces efficacious and specific in vivo knockdown of HCN1 channels. Comparison of in vivo viral transduction using HCN1 miR with mice containing a germ line deletion of HCN1 reveals similar physiological phenotypes in cerebellar Purkinje cells. The easy assembly and re-usability of the toolbox components, together with the ability to up- or down-regulate neuronal gene expression in vivo, may be useful for applications in many areas of neuroscience.

  9. A novel outbreak enterovirus D68 strain associated with acute flaccid myelitis cases in the USA (2012-14): a retrospective cohort study.

    Science.gov (United States)

    Greninger, Alexander L; Naccache, Samia N; Messacar, Kevin; Clayton, Anna; Yu, Guixia; Somasekar, Sneha; Federman, Scot; Stryke, Doug; Anderson, Christopher; Yagi, Shigeo; Messenger, Sharon; Wadford, Debra; Xia, Dongxiang; Watt, James P; Van Haren, Keith; Dominguez, Samuel R; Glaser, Carol; Aldrovandi, Grace; Chiu, Charles Y

    2015-06-01

    Enterovirus D68 was implicated in a widespread outbreak of severe respiratory illness across the USA in 2014 and has also been reported sporadically in patients with acute flaccid myelitis. We aimed to investigate the association between enterovirus D68 infection and acute flaccid myelitis during the 2014 enterovirus D68 respiratory outbreak in the USA. Patients with acute flaccid myelitis who presented to two hospitals in Colorado and California, USA, between Nov 24, 2013, and Oct 11, 2014, were included in the study. Additional cases identified from Jan 1, 2012, to Oct 4, 2014, via statewide surveillance were provided by the California Department of Public Health. We investigated the cause of these cases by metagenomic next-generation sequencing, viral genome recovery, and enterovirus D68 phylogenetic analysis. We compared patients with acute flaccid myelitis who were positive for enterovirus D68 with those with acute flaccid myelitis but negative for enterovirus D68 using the two-tailed Fisher's exact test, two-sample unpaired t test, and Mann-Whitney U test. 48 patients were included: 25 with acute flaccid myelitis, two with enterovirus-associated encephalitis, five with enterovirus-D68-associated upper respiratory illness, and 16 with aseptic meningitis or encephalitis who tested positive for enterovirus. Enterovirus D68 was detected in respiratory secretions from seven (64%) of 11 patients comprising two temporally and geographically linked acute flaccid myelitis clusters at the height of the 2014 outbreak, and from 12 (48%) of 25 patients with acute flaccid myelitis overall. Phylogenetic analysis revealed that all enterovirus D68 sequences associated with acute flaccid myelitis grouped into a clade B1 strain that emerged in 2010. Of six coding polymorphisms in the clade B1 enterovirus D68 polyprotein, five were present in neuropathogenic poliovirus or enterovirus D70, or both. One child with acute flaccid myelitis and a sibling with only upper respiratory

  10. Coxsackievirus A6 and enterovirus 71 causing hand, foot and mouth disease in Cuba, 2011-2013.

    Science.gov (United States)

    Fonseca, Magilé C; Sarmiento, Luis; Resik, Sonia; Martínez, Yenisleidys; Hung, Lai Heng; Morier, Luis; Piñón, Alexander; Valdéz, Odalys; Kourí, Vivian; González, Guelsys

    2014-09-01

    Hand, foot and mouth disease (HFMD) is usually caused by coxsackievirus A16 or enterovirus 71 (EV71). Between 2011 and 2013, HFMD cases were reported from different Cuban provinces. A total of 42 clinical specimens were obtained from 23 patients. Detection, identification and phylogenetic analysis of enterovirus-associated HFMD were carried out by virus isolation, specific enterovirus PCR and partial VP1 sequences. HEV was detected in 11 HFMD cases. Emerging genetic variants of coxsackievirus A6 and EV71 were identified as the causative agents of the Cuban HFMD cases.

  11. Rapid diagnosis of pyrazinamide-resistant multidrug-resistant tuberculosis using a molecular-based diagnostic algorithm.

    Science.gov (United States)

    Simons, S O; van der Laan, T; Mulder, A; van Ingen, J; Rigouts, L; Dekhuijzen, P N R; Boeree, M J; van Soolingen, D

    2014-10-01

    There is an urgent need for rapid and accurate diagnosis of pyrazinamide-resistant multidrug-resistant tuberculosis (MDR-TB). No diagnostic algorithm has been validated in this population. We hypothesized that pncA sequencing added to rpoB mutation analysis can accurately identify patients with pyrazinamide-resistant MDR-TB. We identified from the Dutch national database (2007-11) patients with a positive Mycobacterium tuberculosis culture containing a mutation in the rpoB gene. In these cases, we prospectively sequenced the pncA gene. Results from the rpoB and pncA mutation analysis (pncA added to rpoB) were compared with phenotypic susceptibility testing results to rifampicin, isoniazid and pyrazinamide (reference standard) using the Mycobacterial Growth Indicator Tube 960 system. We included 83 clinical M. tuberculosis isolates containing rpoB mutations in the primary analysis. Rifampicin resistance was seen in 72 isolates (87%), isoniazid resistance in 73 isolates (88%) and MDR-TB in 65 isolates (78%). Phenotypic reference testing identified pyrazinamide-resistant MDR-TB in 31 isolates (48%). Sensitivity of pncA sequencing added to rpoB mutation analysis for detecting pyrazinamide-resistant MDR-TB was 96.8%, the specificity was 94.2%, the positive predictive value was 90.9%, the negative predictive value was 98.0%, the positive likelihood was 16.8 and the negative likelihood was 0.03. In conclusion, pyrazinamide-resistant MDR-TB can be accurately detected using pncA sequencing added to rpoB mutation analysis. We propose to include pncA sequencing in every isolate with an rpoB mutation, allowing for stratification of MDR-TB treatment according to pyrazinamide susceptibility. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

  12. Rapid visible color change and physical swelling during water exposure in triethanolamine-metalcone films formed by molecular layer deposition

    International Nuclear Information System (INIS)

    Lemaire, Paul C.; Oldham, Christopher J.; Parsons, Gregory N.

    2016-01-01

    Molecular layer deposition (MLD) of “metalcones,” including alucone, zincone, titanicone, and others, involves self-limiting half-reactions between organic and organometallic (or metal-halide) reactants. Studies have typically focused on metal precursors reacting with ethylene glycol or glycerol to form the films' polymeric O-M-O-(CH x ) y -O-M-O repeat units. The authors report new MLD materials that incorporate tertiary amine groups into the organic linkage. Specifically, reacting triethanolamine (TEA) with either trimethylaluminum or titanium tetrachloride produces TEA-alucone (Al-TEA) and TEA-titanicone (Ti-TEA), respectively, and the amine group leads to unique physical and optical properties. Fourier-transform infrared (FTIR) analysis confirms that the films have prominent C-H, C-N, and M-O-C peaks, consistent with the expected bond structure. When exposed to vapors, including water, alcohol, or ammonia, the Ti-TEA films changed their visible color within minutes and increased physical thickness by >35%. The Al-TEA showed significantly less response. X-ray photoelectron spectroscopy and FTIR suggest that HCl generated during MLD coordinates to the amine forming a quaternary ammonium salt that readily binds adsorbates via hydrogen bonding. The visible color change is reversible, and ellipsometry confirms that the color change results from vapor absorption. The unique absorptive and color-changing properties of the TEA-metalcone films point to new possible applications for MLD materials in filtration, chemical absorption, and multifunctional chemical separations/sensing device systems

  13. Rapid visible color change and physical swelling during water exposure in triethanolamine-metalcone films formed by molecular layer deposition

    Energy Technology Data Exchange (ETDEWEB)

    Lemaire, Paul C.; Oldham, Christopher J.; Parsons, Gregory N., E-mail: gnp@ncsu.edu [Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, North Carolina 27695 (United States)

    2016-01-15

    Molecular layer deposition (MLD) of “metalcones,” including alucone, zincone, titanicone, and others, involves self-limiting half-reactions between organic and organometallic (or metal-halide) reactants. Studies have typically focused on metal precursors reacting with ethylene glycol or glycerol to form the films' polymeric O-M-O-(CH{sub x}){sub y}-O-M-O repeat units. The authors report new MLD materials that incorporate tertiary amine groups into the organic linkage. Specifically, reacting triethanolamine (TEA) with either trimethylaluminum or titanium tetrachloride produces TEA-alucone (Al-TEA) and TEA-titanicone (Ti-TEA), respectively, and the amine group leads to unique physical and optical properties. Fourier-transform infrared (FTIR) analysis confirms that the films have prominent C-H, C-N, and M-O-C peaks, consistent with the expected bond structure. When exposed to vapors, including water, alcohol, or ammonia, the Ti-TEA films changed their visible color within minutes and increased physical thickness by >35%. The Al-TEA showed significantly less response. X-ray photoelectron spectroscopy and FTIR suggest that HCl generated during MLD coordinates to the amine forming a quaternary ammonium salt that readily binds adsorbates via hydrogen bonding. The visible color change is reversible, and ellipsometry confirms that the color change results from vapor absorption. The unique absorptive and color-changing properties of the TEA-metalcone films point to new possible applications for MLD materials in filtration, chemical absorption, and multifunctional chemical separations/sensing device systems.

  14. Enterovirus 71 viral capsid protein linear epitopes: Identification and characterization

    Directory of Open Access Journals (Sweden)

    Gao Fan

    2012-01-01

    Full Text Available Abstract Background To characterize the human humoral immune response against enterovirus 71 (EV71 infection and map human epitopes on the viral capsid proteins. Methods A series of 256 peptides spanning the capsid proteins (VP1, VP2, VP3 of BJ08 strain (genomic C4 were synthesized. An indirect enzyme-linked immunosorbent assay (ELISA was carried out to detect anti-EV71 IgM and IgG in sera of infected children in acute or recovery phase. The partially overlapped peptides contained 12 amino acids and were coated in the plate as antigen (0.1 μg/μl. Sera from rabbits immunized with inactivated BJ08 virus were also used to screen the peptide panel. Results A total of 10 human anti-EV71 IgM epitopes (vp1-14 in VP1; vp2-6, 21, 40 and 50 in VP2 and vp3-10, 12, 15, 24 and 75 in VP3 were identified in acute phase sera. In contrast, only one anti-EV71 IgG epitope in VP1 (vp1-15 was identified in sera of recovery stage. Four rabbit anti-EV71 IgG epitopes (vp1-14, 31, 54 and 71 were identified and mapped to VP1. Conclusion These data suggested that human IgM epitopes were mainly mapped to VP2 and VP3 with multi-epitope responses occurred at acute infection, while the only IgG epitope located on protein VP1 was activated in recovery phase sera. The dynamic changes of humoral immune response at different stages of infection may have public health significance in evaluation of EV71 vaccine immunogenicity and the clinical application of diagnostic reagents.

  15. Maternal Enterovirus Infection during Pregnancy as a Risk Factor in Offspring Diagnosed with Type 1 Diabetes between 15 and 30 Years of Age

    Directory of Open Access Journals (Sweden)

    Maria Elfving

    2008-01-01

    Full Text Available Maternal enterovirus infections during pregnancy may increase the risk of offspring developing type 1 diabetes during childhood. The aim of this study was to investigate whether gestational enterovirus infections increase the offspring's risk of type 1 diabetes later in life. Serum samples from 30 mothers without diabetes whose offspring developed type 1 diabetes between 15 and 25 years of age were analyzed for enterovirus-specific immunoglobulin M (IgM antibodies and enterovirus genome (RNA, and compared to a control group. Among the index mothers, 9/30 (30% were enterovirus IgM-positive, and none was positive for enterovirus RNA. In the control group, 14/90 (16% were enterovirus IgM-positive, and 4/90 (4% were positive for enterovirus RNA (n.s.. Boys of enterovirus IgM-positive mothers had approximately 5 times greater risk of developing diabetes (OR 4.63; 95% CI 1.22–17.6, as compared to boys of IgM-negative mothers (P<.025. These results suggest that gestational enterovirus infections may be related to the risk of offspring developing type 1 diabetes in adolescence and young adulthood.

  16. Evaluation of RT-PCR and immunohistochemistry as tools for detection of enterovirus in the human pancreas and islets of Langerhans.

    Science.gov (United States)

    Skog, Oskar; Ingvast, Sofie; Korsgren, Olle

    2014-10-01

    Enteroviruses have been implicated in the etiology of type 1 diabetes, supported by immunoreactivity of enteroviral protein in islets, but presence of enteroviral genome has rarely been reported. Failure to detect enterovirus with RT-PCR has been attributed to the possible presence of PCR inhibitors and that only few cells are infected. The aim of this study was to evaluate strategies for detection of enterovirus in human islets. A scenario was modeled with defined infected islets among a large number of uninfected pancreatic cells and the sensitivity of immunohistochemistry and PCR for detection of enterovirus was evaluated. Enterovirus was detected with PCR when only one single human islet, infected in vitro with a low dose of virus, was mixed with an uninfected pancreatic biopsy. Enterovirus could not be detected by immunohistochemistry under the same conditions, demonstrating the superior sensitivity of PCR also in pancreatic tissue with only a small fraction of infected cells. In addition, we demonstrate that pancreatic cell culture supernatant does not cause degradation of enterovirus at 37°C, indicating that under normal culture conditions released virus is readily detectable. Utilizing PCR, the pancreases of two organ donors that died at onset of type 1 diabetes were found negative for enterovirus genome despite islet cells being positive using immunohistochemistry. These data suggest that PCR should be the preferred screening method for enterovirus in the pancreas and suggest cautious interpretation of immunostaining for enterovirus that cannot be confirmed with PCR. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Broad-range inhibition of enterovirus replication by OSW-1, a natural compound targeting OSBP

    NARCIS (Netherlands)

    Albulescu, Lucian; Strating, Jeroen R. P. M.; Thibaut, Hendrik Jan; van der Linden, Lonneke; Shair, Matthew D.; Neyts, Johan; van Kuppeveld, Frank J. M.

    2015-01-01

    Enteroviruses, e.g., polio-, coxsackie- and rhinoviruses, constitute a large genus within the Picornaviridae family of positive-strand RNA viruses and include many important pathogens linked to a variety of acute and chronic diseases. Despite their huge medical and economic impact, no approved

  18. Emergence of enterovirus D68 in Denmark, June 2014 to February 2015

    DEFF Research Database (Denmark)

    Midgley, S E; Christiansen, C B; Poulsen, M W

    2015-01-01

    From June 2014 through February 2015, respiratory samples from 130 Danish patients were screened for enterovirus D68 (EV-D68). Fourteen EV-D68 cases were detected, of which 12 presented with respiratory symptoms, and eight had known underlying disease. The median age of EV-D68 cases was three years...

  19. Enterovirus 2Apro targets MDA5 and MAVS in infected cells

    NARCIS (Netherlands)

    Feng, Q.; Langereis, M.A.; Lork, M.; Nguyen, M.H.; Hato, S.V.; Lanke, K.H.W.; Emdad, L.; Bhoopathi, P.; Fisher, P.B.; Lloyd, R.E.; Kuppeveld, F.J.M. van

    2014-01-01

    RIG-I-like receptors (RLRs) MDA5 and RIG-I are key players in the innate antiviral response. Upon recognition of viral RNA, they interact with MAVS, eventually inducing type I interferon production. The interferon induction pathway is commonly targeted by viruses. How enteroviruses suppress

  20. Characteristics of pediatric patients with enterovirus meningitis and no cerebral fluid pleocytosis

    NARCIS (Netherlands)

    de Crom, Stephanie C. M.; van Furth, Marceline A. M.; Peeters, Marcel F.; Rossen, John W. A.; Obihara, Charles C.

    UNLABELLED: Human non-polio enterovirus (EV) is the most important cause of aseptic meningitis in children. Only a few studies report the lack of cerobrospinal fluid (CSF) pleocytosis in children with confirmed EV meningitis; however, the characteristics of these children have not been well defined.

  1. Persistent spiking fever in a child with acute myeloid leukemia and disseminated infection with enterovirus

    NARCIS (Netherlands)

    Murk, J. L.; de Vries, A. C.; GeurtsvanKessel, C. H.; Aron, G.; Osterhaus, A. D.; Wolthers, K. C.; Fraaij, P. L.

    2014-01-01

    We here report a 7 year old acute myeloid leukemia patient with persistent spiking fever likely caused by chronic echovirus 20 infection. After immunoglobulin substitution fevers subsided and the virus was cleared. Enterovirus infection should be considered in immunocompromised patients with

  2. Analysis of the synonymous codon usage bias in recently emerged enterovirus D68 strains.

    Science.gov (United States)

    Karniychuk, Uladzimir U

    2016-09-02

    Understanding the codon usage pattern of a pathogen and relationship between pathogen and host's codon usage patterns has fundamental and applied interests. Enterovirus D68 (EV-D68) is an emerging pathogen with a potentially high public health significance. In the present study, the synonymous codon usage bias of 27 recently emerged, and historical EV-D68 strains was analyzed. In contrast to previously studied enteroviruses (enterovirus 71 and poliovirus), EV-D68 and human host have a high discrepancy between favored codons. Analysis of viral synonymous codon usage bias metrics, viral nucleotide/dinucleotide compositional parameters, and viral protein properties showed that mutational pressure is more involved in shaping the synonymous codon usage bias of EV-D68 than translation selection. Computation of codon adaptation indices allowed to estimate expression potential of the EV-D68 genome in several commonly used laboratory animals. This approach requires experimental validation and may provide an auxiliary tool for the rational selection of laboratory animals to model emerging viral diseases. Enterovirus D68 genome compositional and codon usage data can be useful for further pathogenesis, animal model, and vaccine design studies. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Evaluation of methods using celite to concentrate norovirus, adenovirus and enterovirus from wastewater

    Science.gov (United States)

    Enteroviruses, noroviruses and adenoviruses are among the most common viruses infecting humans worldwide. These viruses are shed in the feces of infected individuals and can accumulate in wastewater. Therefore, wastewater is a source of a potentially diverse group of enteric viru...

  4. Poliovirus and other enteroviruses in children infected with intestinal parasites in Nigeria.

    Science.gov (United States)

    Adekolujo, Daniel R; Olayinka, Suraj O; Adeniji, Johnson A; Oyeyemi, Oyetunde T; Odaibo, Alexander B

    2015-10-29

    Poliovirus, an enterovirus, still persists in Nigeria despite the global efforts tailored towards its eradication. This study aimed to assess the impacts of poliovirus and other enteroviruses on the susceptibility of individuals to intestinal parasite infections. A cross-sectional study on the prevalence of intestinal parasites was conducted on two-sample stool specimens of 717 Nigerian children (between 1 and 19 years of age) whose poliovirus/other enteroviruses infection status had been determined. The overall prevalence of Sabin poliovirus and other related enteroviruses infections were 6.6% and 13.8%, respectively. The prevalence of Ascaris lumbricoides was significantly higher than that of other intestinal parasites (p parasitic infection (OR = 11.7, CI = 9.2-15.0). While the prevalence of all species of parasites except S. mansoni showed no significant variations in children with Sabin poliovirus (p > 0.05), the prevalence of hookworms and Taenia spp. was significantly higher in children with other enteroviral infections (p parasites is an indication of possible association of the parasites in a more poliovirus-endemic population. A combined intervention approach for the two infections is advocated.

  5. Epidemiology of Sepsis-like Illness in Young Infants Major Role of Enterovirus and Human Parechovirus

    NARCIS (Netherlands)

    de Jong, Eveline P.; van den Beuken, Monique G. A.; van Elzakker, Erika P. M.; Wolthers, Katja C.; Sprij, Arwen J.; Lopriore, Enrico; Walther, Frans J.; Brus, Frank

    2018-01-01

    Background: Sepsis-like illness is a main cause for hospital admission in young infants. Our aim was to investigate incidence, epidemiology and clinical characteristics of enterovirus (EV) and human parechovirus (HPeV) infections in young infants with sepsis-like illness. Methods: This is a

  6. Acute flaccid myelitis associated with enterovirus-D68 infection in an otherwise healthy child

    NARCIS (Netherlands)

    Esposito, Susanna; Chidini, Giovanna; Cinnante, Claudia; Napolitano, Luisa; Giannini, Alberto; Terranova, Leonardo; Niesters, Hubert; Principi, Nicola; Calderini, Edoardo

    2017-01-01

    Background: Reporting new cases of enterovirus (EV)-D68-associated acute flaccid myelitis (AFM) is essential to understand how the virus causes neurological damage and to characterize EV-D68 strains associated with AFM. Case presentation: A previously healthy 4-year-old boy presented with sudden

  7. Binding of glutathione to enterovirus capsids is essential for virion morphogenesis.

    NARCIS (Netherlands)

    Thibaut, H.J.; Linden, L. van der; Jiang, P.; Thys, B.; Canela, M.D.; Aguado, L.; Rombaut, B.; Wimmer, E.; Paul, A.; Perez-Perez, M.J.; Kuppeveld, F.J.M. van; Neyts, J.

    2014-01-01

    Enteroviruses (family of the Picornaviridae) cover a large group of medically important human pathogens for which no antiviral treatment is approved. Although these viruses have been extensively studied, some aspects of the viral life cycle, in particular morphogenesis, are yet poorly understood. We

  8. Binding of glutathione to enterovirus capsids is essential for virion morphogenesis

    NARCIS (Netherlands)

    Thibaut, Hendrik Jan; van der Linden, Lonneke; Jiang, Ping; Thys, Bert; Canela, María-Dolores; Aguado, Leire; Rombaut, Bart; Wimmer, Eckard; Paul, Aniko; Pérez-Pérez, María-Jesús; van Kuppeveld, Frank J M; Neyts, Johan

    Enteroviruses (family of the Picornaviridae) cover a large group of medically important human pathogens for which no antiviral treatment is approved. Although these viruses have been extensively studied, some aspects of the viral life cycle, in particular morphogenesis, are yet poorly understood. We

  9. Enterovirus 2Apro targets MDA5 and MAVS in infected cells

    NARCIS (Netherlands)

    Feng, Qian; Langereis, Martijn A; Lork, Marie; Nguyen, Mai; Hato, Stanleyson V; Lanke, Kjerstin; Emdad, Luni; Bhoopathi, Praveen; Fisher, Paul B; Lloyd, Richard E; van Kuppeveld, Frank J M

    2014-01-01

    UNLABELLED: RIG-I-like receptors (RLRs) MDA5 and RIG-I are key players in the innate antiviral response. Upon recognition of viral RNA, they interact with MAVS, eventually inducing type I interferon production. The interferon induction pathway is commonly targeted by viruses. How enteroviruses

  10. New introductions of enterovirus 71 subgenogroup C4 strains, France, 2012.

    Science.gov (United States)

    Schuffenecker, Isabelle; Henquell, Cécile; Mirand, Audrey; Coste-Burel, Marianne; Marque-Juillet, Stéphanie; Desbois, Delphine; Lagathu, Gisèle; Bornebusch, Laure; Bailly, Jean-Luc; Lina, Bruno

    2014-08-01

    In France during 2012, human enterovirus 71 (EV-A71) subgenogroup C4 strains were detected in 4 children hospitalized for neonatal fever or meningitis. Phylogenetic analysis showed novel and independent EV-A71 introductions, presumably from China, and suggested circulation of C4 strains throughout France. This observation emphasizes the need for monitoring EV-A71 infections in Europe.

  11. Emergence and epidemic occurrence of enterovirus 68 respiratory infections in The Netherlands in 2010.

    NARCIS (Netherlands)

    Meijer, A.; Sanden, S. van der; Snijders, B.E.P.; Jaramillo-Gutierrez, G.; Bont, L.; Ent, C.K. van der; Overduin, P.; Jenny, S.L.; Jusic, E.; Avoort, H.G.A.M. van der; Smith, G.J.D.; Donker, G.A.; Koopmans, M.P.G.

    2012-01-01

    Following an increase in detection of enterovirus 68 (EV68) in community surveillance of respiratory infections in The Netherlands in 2010, epidemiological and virological analyses were performed to investigate the possible public health impact of EV68 infections. We retrospectively tested specimens

  12. Evolutionary trajectory of the VP1 gene of human enterovirus 71 genogroup B and C viruses

    NARCIS (Netherlands)

    S.M.G. van der Sanden (Sabine); H.G.A.M. van der Avoort (Harrie); P. Lemey (Philippe); G. Uslu (Gökhan); M.P.G. Koopmans D.V.M. (Marion)

    2010-01-01

    textabstractFrom 1963 to 1986, human enterovirus 71 (HEV71) infections in the Netherlands were successively caused by viruses of subgenogroups B0, B1 and B2. A genogroup shift occurred in 1987, after which viruses of subgenogroups C1 and C2 were detected exclusively. This is in line with HEV71

  13. Testing mitochondrial sequences and anonymous nuclear markers for phylogeny reconstruction in a rapidly radiating group: molecular systematics of the Delphininae (Cetacea: Odontoceti: Delphinidae

    Directory of Open Access Journals (Sweden)

    Kingston Sarah E

    2009-10-01

    Full Text Available Abstract Background Many molecular phylogenetic analyses rely on DNA sequence data obtained from single or multiple loci, particularly mitochondrial DNA loci. However, phylogenies for taxa that have undergone recent, rapid radiation events often remain unresolved. Alternative methodologies for discerning evolutionary relationships under these conditions are desirable. The dolphin subfamily Delphininae is a group that has likely resulted from a recent and rapid radiation. Despite several efforts, the evolutionary relationships among the species in the subfamily remain unclear. Results Here, we compare a phylogeny estimated using mitochondrial DNA (mtDNA control region sequences to a multi-locus phylogeny inferred from 418 polymorphic genomic markers obtained from amplified fragment length polymorphism (AFLP analysis. The two sets of phylogenies are largely incongruent, primarily because the mtDNA tree provides very poor resolving power; very few species' nodes in the tree are supported by bootstrap resampling. The AFLP phylogeny is considerably better resolved and more congruent with relationships inferred from morphological data. Both phylogenies support paraphyly for the genera Stenella and Tursiops. The AFLP data indicate a close relationship between the two spotted dolphin species and recent ancestry between Stenella clymene and S. longirostris. The placement of the Lagenodelphis hosei lineage is ambiguous: phenetic analysis of the AFLP data is consistent with morphological expectations but the phylogenetic analysis is not. Conclusion For closely related, recently diverged taxa, a multi-locus genome-wide survey is likely the most comprehensive approach currently available for phylogenetic inference.

  14. Molecular-based rapid inventories of sympatric diversity: a comparison of DNA barcode clustering methods applied to geography-based vs clade-based sampling of amphibians.

    Science.gov (United States)

    Paz, Andrea; Crawford, Andrew J

    2012-11-01

    Molecular markers offer a universal source of data for quantifying biodiversity. DNA barcoding uses a standardized genetic marker and a curated reference database to identify known species and to reveal cryptic diversity within wellsampled clades. Rapid biological inventories, e.g. rapid assessment programs (RAPs), unlike most barcoding campaigns, are focused on particular geographic localities rather than on clades. Because of the potentially sparse phylogenetic sampling, the addition of DNA barcoding to RAPs may present a greater challenge for the identification of named species or for revealing cryptic diversity. In this article we evaluate the use of DNA barcoding for quantifying lineage diversity within a single sampling site as compared to clade-based sampling, and present examples from amphibians. We compared algorithms for identifying DNA barcode clusters (e.g. species, cryptic species or Evolutionary Significant Units) using previously published DNA barcode data obtained from geography-based sampling at a site in Central Panama, and from clade-based sampling in Madagascar. We found that clustering algorithms based on genetic distance performed similarly on sympatric as well as clade-based barcode data, while a promising coalescent-based method performed poorly on sympatric data. The various clustering algorithms were also compared in terms of speed and software implementation. Although each method has its shortcomings in certain contexts, we recommend the use of the ABGD method, which not only performs fairly well under either sampling method, but does so in a few seconds and with a user-friendly Web interface.

  15. Informing Antibiotic Treatment Decisions: Evaluating Rapid Molecular Diagnostics To Identify Susceptibility and Resistance to Carbapenems against Acinetobacter spp. in PRIMERS III.

    Science.gov (United States)

    Evans, Scott R; Hujer, Andrea M; Jiang, Hongyu; Hill, Carol B; Hujer, Kristine M; Mediavilla, Jose R; Manca, Claudia; Tran, Thuy Tien T; Domitrovic, T Nicholas; Higgins, Paul G; Seifert, Harald; Kreiswirth, Barry N; Patel, Robin; Jacobs, Michael R; Chen, Liang; Sampath, Rangarajan; Hall, Thomas; Marzan, Christine; Fowler, Vance G; Chambers, Henry F; Bonomo, Robert A

    2017-01-01

    The widespread dissemination of carbapenem-resistant Acinetobacter spp. has created significant therapeutic challenges. At present, rapid molecular diagnostics (RMDs) that can identify this phenotype are not commercially available. Two RMD platforms, PCR combined with electrospray ionization mass spectrometry (PCR/ESI-MS) and molecular beacons (MB), for detecting genes conferring resistance/susceptibility to carbapenems in Acinetobacter spp. were evaluated. An archived collection of 200 clinical Acinetobacter sp. isolates was tested. Predictive values for susceptibility and resistance were estimated as a function of susceptibility prevalence and were based on the absence or presence of beta-lactamase (bla) NDM, VIM, IMP, KPC, and OXA carbapenemase genes (e.g., bla OXA-23 , bla OXA-24/40 , and bla OXA-58 found in this study) against the reference standard of MIC determinations. According to the interpretation of MICs, 49% (n = 98) of the isolates were carbapenem resistant (as defined by either resistance or intermediate resistance to imipenem). The susceptibility sensitivities (95% confidence interval [CI]) for imipenem were 82% (74%, 89%) and 92% (85%, 97%) for PCR/ESI-MS and MB, respectively. Resistance sensitivities (95% CI) for imipenem were 95% (88%, 98%) and 88% (80%, 94%) for PCR/ESI-MS and MB, respectively. PRIMERS III establishes that RMDs can discriminate between carbapenem resistance and susceptibility in Acinetobacter spp. In the context of a known prevalence of resistance, SPVs and RPVs can inform clinicians regarding the best choice for empiric antimicrobial therapy against this multidrug-resistant pathogen. Copyright © 2016 American Society for Microbiology.

  16. Recent advances in biochemical and molecular diagnostics for the rapid detection of antibiotic-resistant Enterobacteriaceae: a focus on ß-lactam resistance.

    Science.gov (United States)

    Decousser, Jean-Winoc; Poirel, Laurent; Nordmann, Patrice

    2017-04-01

    The rapid detection of resistance is a challenge for clinical microbiologists who wish to prevent deleterious individual and collective consequences such as (i) delaying efficient antibiotic therapy, which worsens the survival rate of the most severely ill patients, or (ii) delaying the isolation of the carriers of multidrug-resistant bacteria and promoting outbreaks; this last consequence is of special concern, and there are an increasing number of approaches and market-based solutions in response. Areas covered: From simple, cheap biochemical tests to whole-genome sequencing, clinical microbiologists must select the most adequate phenotypic and genotypic tools to promptly detect and confirm β-lactam resistance from cultivated bacteria or from clinical specimens. Here, the authors review the published literature from the last 5 years about the primary technical approaches and commercial laboratory reagents for these purposes, including molecular, biochemical and immune assays. Furthermore, the authors discuss their intrinsic and relative performance, and we challenge their putative clinical impact. Expert commentary: Until the availability of fully automated wet and dry whole genome sequencing solutions, microbiologists should focus on inexpensive biochemical tests for cultured isolates or monomicrobial clinical specimen and on using the expensive molecular PCR-based strategies for the targeted screening of complex biological environments.

  17. High-risk Long QT Syndrome Mutations in the Kv7.1 (KCNQ1) Pore Disrupt the Molecular Basis for Rapid K+ Permeation

    Science.gov (United States)

    Burgess, Don E.; Bartos, Daniel C.; Reloj, Allison R.; Campbell, Kenneth S.; Johnson, Jonathan N.; Tester, David J.; Ackerman, Michael J.; Fressart, Véronique; Denjoy, Isabelle; Guicheney, Pascale; Moss, Arthur J.; Ohno, Seiko; Horie, Minoru; Delisle, Brian P.

    2012-01-01

    Type 1 long QT syndrome (LQT1) syndrome is caused by loss-of-function mutations in the KCNQ1, which encodes the K+ channel (Kv7.1) that underlies the slowly activating delayed rectifier K+ current in the heart. Intragenic risk stratification suggests LQT1 mutations that disrupt conserved amino acid residues in the pore are an independent risk factor for LQT1-related cardiac events. The purpose of this study is to determine possible molecular mechanisms that underlie the loss-of-function for these high-risk mutations. Extensive genotype-phenotype analyses of LQT1 patients showed that T322M-, T322A-, or G325R-Kv7.1 confer a high risk for LQT1-related cardiac events. Heterologous expression of these mutations with KCNE1 revealed they generated non-functional channels and caused dominant negative suppression of WT-Kv7.1 current. Molecular dynamic simulations (MDS) of analogous mutations in KcsA (T85M-, T85A-, and G88R-KcsA) demonstrated that they disrupted the symmetrical distribution of the carbonyl oxygen atoms in the selectivity filter, which upset the balance between the strong attractive and K+-K+ repulsive forces required for rapid K+ permeation. We conclude high-risk LQT1 mutations in the pore likely disrupt the architectural and physical properties of the K+ channel selectivity filter. PMID:23092362

  18. High-risk long QT syndrome mutations in the Kv7.1 (KCNQ1) pore disrupt the molecular basis for rapid K(+) permeation.

    Science.gov (United States)

    Burgess, Don E; Bartos, Daniel C; Reloj, Allison R; Campbell, Kenneth S; Johnson, Jonathan N; Tester, David J; Ackerman, Michael J; Fressart, Véronique; Denjoy, Isabelle; Guicheney, Pascale; Moss, Arthur J; Ohno, Seiko; Horie, Minoru; Delisle, Brian P

    2012-11-13

    Type 1 long QT syndrome (LQT1) is caused by loss-of-function mutations in the KCNQ1 gene, which encodes the K(+) channel (Kv7.1) that underlies the slowly activating delayed rectifier K(+) current in the heart. Intragenic risk stratification suggests LQT1 mutations that disrupt conserved amino acid residues in the pore are an independent risk factor for LQT1-related cardiac events. The purpose of this study is to determine possible molecular mechanisms that underlie the loss of function for these high-risk mutations. Extensive genotype-phenotype analyses of LQT1 patients showed that T322M-, T322A-, or G325R-Kv7.1 confers a high risk for LQT1-related cardiac events. Heterologous expression of these mutations with KCNE1 revealed they generated nonfunctional channels and caused dominant negative suppression of WT-Kv7.1 current. Molecular dynamics simulations of analogous mutations in KcsA (T85M-, T85A-, and G88R-KcsA) demonstrated that they disrupted the symmetrical distribution of the carbonyl oxygen atoms in the selectivity filter, which upset the balance between the strong attractive and K(+)-K(+) repulsive forces required for rapid K(+) permeation. We conclude high-risk LQT1 mutations in the pore likely disrupt the architectural and physical properties of the K(+) channel selectivity filter.

  19. Rabi oscillations and rapid-passage effects in the molecular-beam CO2-laser Stark spectroscopy of CH3F

    International Nuclear Information System (INIS)

    Adam, A.G.; Gough, T.E.; Isenor, N.R.; Scoles, G.

    1985-01-01

    sub-Doppler molecular-beam laser Stark spectroscopy has been employed to produce high-contrast Rabi oscillations in the ν 3 band of CH 3 F. By varying the intensity of the cw CO 2 laser, up to five complete oscillations were observed before the phenomenon was washed out by rapid-passage effects and damping mechanisms. Besides being useful in clarifying key features of coherent ir molecular-beam spectroscopy, the observation of Rabi oscillations provides one of the most accurate means of directly measuring transition dipole moments. Analysis of the present data on three rovibrational transitions, Q(1,1) -1reverse arrow0, P(1,0) 0reverse arrow0, and R(1,1) 0reverse arrow1, has yielded a rotationless transition dipole moment of 0.21 +- 0.01 D for the ν 3 = 1reverse arrow0 vibration. This result is in agreement with values estimated from both band-intensity and absorption-coefficient data in the literature

  20. Helicase-dependent isothermal amplification: a novel tool in the development of molecular-based analytical systems for rapid pathogen detection.

    Science.gov (United States)

    Barreda-García, Susana; Miranda-Castro, Rebeca; de-Los-Santos-Álvarez, Noemí; Miranda-Ordieres, Arturo J; Lobo-Castañón, María Jesús

    2018-01-01

    Highly sensitive testing of nucleic acids is essential to improve the detection of pathogens, which pose a major threat for public health worldwide. Currently available molecular assays, mainly based on PCR, have a limited utility in point-of-need control or resource-limited settings. Consequently, there is a strong interest in developing cost-effective, robust, and portable platforms for early detection of these harmful microorganisms. Since its description in 2004, isothermal helicase-dependent amplification (HDA) has been successfully applied in the development of novel molecular-based technologies for rapid, sensitive, and selective detection of viruses and bacteria. In this review, we highlight relevant analytical systems using this simple nucleic acid amplification methodology that takes place at a constant temperature and that is readily compatible with microfluidic technologies. Different strategies for monitoring HDA amplification products are described. In addition, we present technological advances for integrating sample preparation, HDA amplification, and detection. Future perspectives and challenges toward point-of-need use not only for clinical diagnosis but also in food safety testing and environmental monitoring are also discussed. Graphical Abstract Expanding the analytical toolbox for the detection of DNA sequences specific of pathogens with isothermal helicase dependent amplification (HDA).

  1. Rapid and selective screening of melamine in bovine milk using molecularly imprinted matrix solid-phase dispersion coupled with liquid chromatography-ultraviolet detection.

    Science.gov (United States)

    Yan, Hongyuan; Cheng, Xiaoling; Sun, Ning; Cai, Tianyu; Wu, Ruijun; Han, Kun

    2012-11-01

    A simple, convenient and high selective molecularly imprinted matrix solid-phase dispersion (MI-MSPD) using water-compatible cyromazine-imprinted polymer as adsorbent was proposed for the rapid screening of melamine from bovine milk coupled with liquid chromatography-ultraviolet detection. The molecularly imprinted polymers (MIPs) synthesized by cyromazine as dummy template and reformative methanol-water system as reaction medium showed higher affinity and selectivity to melamine, and so they were applied as the specific dispersant of MSPD to extraction of melamine and simultaneously eliminate the effect of template leakage on quantitative analysis. Under the optimized conditions, good linearity was obtained in a range of 0.24-60.0μgg(-1) with the correlation coefficient of 0.9994. The recoveries of melamine at three spiked levels were ranged from 86.0 to 96.2% with the relative standard deviation (RSD)≤4.0%. This proposed MI-MSPD method combined the advantages of MSPD and MIPs, and could be used as an alternative tool for analyzing the residues of melamine in complex milk samples. Copyright © 2012 Elsevier B.V. All rights reserved.

  2. Polio and Measles Down the Drain: Environmental Enterovirus Surveillance in the Netherlands, 2005 to 2015.

    Science.gov (United States)

    Benschop, Kimberley S M; van der Avoort, Harrie G; Jusic, Edin; Vennema, Harry; van Binnendijk, Rob; Duizer, Erwin

    2017-07-01

    Polioviruses (PVs) are members of the genus Enterovirus In the Netherlands, the exclusion of PV circulation is based on clinical enterovirus (EV) surveillance (CEVS) of EV-positive cases and routine environmental EV surveillance (EEVS) conducted on sewage samples collected in the region of the Netherlands where vaccination coverage is low due to religious reasons. We compared the EEVS data to those of the CEVS to gain insight into the relevance of EEVS for poliovirus and nonpolio enterovirus surveillance. Following the polio outbreak in Syria, EEVS was performed at the primary refugee center in Ter Apel in the Netherlands, and data were compared to those of CEVS and EEVS. Furthermore, we assessed the feasibility of poliovirus detection by EEVS using measles virus detection in sewage during a measles outbreak as a proxy. Two Sabin-like PVs were found in routine EEVS, 11 Sabin-like PVs were detected in the CEVS, and one Sabin-like PV was found in the Ter Apel sewage. We observed significant differences between the three programs regarding which EVs were found. In 6 sewage samples collected during the measles outbreak in 2013, measles virus RNA was detected in regions where measles cases were identified. In conclusion, we detected PVs, nonpolio EVs, and measles virus in sewage and showed that environmental surveillance is useful for poliovirus detection in the Netherlands, where live oral poliovirus vaccine is not used and communities with lower vaccination coverage exist. EEVS led to the detection of EV types not seen in the CEVS, showing that EEVS is complementary to CEVS. IMPORTANCE We show that environmental enterovirus surveillance complements clinical enterovirus surveillance for poliovirus detection, or exclusion, and for nonpolio enterovirus surveillance. Even in the presence of adequate surveillance, only a very limited number of Sabin-like poliovirus strains were detected in a 10-year period, and no signs of transmission of oral polio vaccine (OPV) strains

  3. Evaluation of Two Lyophilized Molecular Assays to Rapidly Detect Foot-and-Mouth Disease Virus Directly from Clinical Samples in Field Settings.

    Science.gov (United States)

    Howson, E L A; Armson, B; Madi, M; Kasanga, C J; Kandusi, S; Sallu, R; Chepkwony, E; Siddle, A; Martin, P; Wood, J; Mioulet, V; King, D P; Lembo, T; Cleaveland, S; Fowler, V L

    2017-06-01

    Accurate, timely diagnosis is essential for the control, monitoring and eradication of foot-and-mouth disease (FMD). Clinical samples from suspect cases are normally tested at reference laboratories. However, transport of samples to these centralized facilities can be a lengthy process that can impose delays on critical decision making. These concerns have motivated work to evaluate simple-to-use technologies, including molecular-based diagnostic platforms, that can be deployed closer to suspect cases of FMD. In this context, FMD virus (FMDV)-specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) and real-time RT-PCR (rRT-PCR) assays, compatible with simple sample preparation methods and in situ visualization, have been developed which share equivalent analytical sensitivity with laboratory-based rRT-PCR. However, the lack of robust 'ready-to-use kits' that utilize stabilized reagents limits the deployment of these tests into field settings. To address this gap, this study describes the performance of lyophilized rRT-PCR and RT-LAMP assays to detect FMDV. Both of these assays are compatible with the use of fluorescence to monitor amplification in real-time, and for the RT-LAMP assays end point detection could also be achieved using molecular lateral flow devices. Lyophilization of reagents did not adversely affect the performance of the assays. Importantly, when these assays were deployed into challenging laboratory and field settings within East Africa they proved to be reliable in their ability to detect FMDV in a range of clinical samples from acutely infected as well as convalescent cattle. These data support the use of highly sensitive molecular assays into field settings for simple and rapid detection of FMDV. © 2015 The Authors. Transboundary and Emerging Diseases Published by Blackwell Verlag GmbH.

  4. Automated Online Solid-Phase Derivatization for Sensitive Quantification of Endogenous S-Nitrosoglutathione and Rapid Capture of Other Low-Molecular-Mass S-Nitrosothiols.

    Science.gov (United States)

    Wang, Xin; Garcia, Carlos T; Gong, Guanyu; Wishnok, John S; Tannenbaum, Steven R

    2018-02-06

    S-Nitrosothiols (RSNOs) constitute a circulating endogenous reservoir of nitric oxide and have important biological activities. In this study, an online coupling of solid-phase derivatization (SPD) with liquid chromatography-mass spectrometry (LC-MS) was developed and applied in the analysis of low-molecular-mass RSNOs. A derivatizing-reagent-modified polymer monolithic column was prepared and adapted for online SPD-LC-MS. Analytes from the LC autosampler flowed through the monolithic column for derivatization and then directly into the LC-MS for analysis. This integration of the online derivatization, LC separation, and MS detection facilitated system automation, allowing rapid, laborsaving, and sensitive detection of RSNOs. S-Nitrosoglutathione (GSNO) was quantified using this automated online method with good linearity (R 2 = 0.9994); the limit of detection was 0.015 nM. The online SPD-LC-MS method has been used to determine GSNO levels in mouse samples, 138 ± 13.2 nM of endogenous GSNO was detected in mouse plasma. Besides, the GSNO concentrations in liver (64.8 ± 11.3 pmol/mg protein), kidney (47.2 ± 6.1 pmol/mg protein), heart (8.9 ± 1.8 pmol/mg protein), muscle (1.9 ± 0.3 pmol/mg protein), hippocampus (5.3 ± 0.9 pmol/mg protein), striatum (6.7 ± 0.6 pmol/mg protein), cerebellum (31.4 ± 6.5 pmol/mg protein), and cortex (47.9 ± 4.6 pmol/mg protein) were also successfully quantified. When the derivatization was performed within 8 min, followed by LC-MS detection, samples could be rapidly analyzed compared with the offline manual method. Other low-molecular-mass RSNOs, such as S-nitrosocysteine and S-nitrosocysteinylglycine, were captured by rapid precursor-ion scanning, showing that the proposed method is a potentially powerful tool for capture, identification, and quantification of RSNOs in biological samples.

  5. Variations in cerebrospinal fluid viral loads among enterovirus genotypes in patients hospitalized with laboratory-confirmed meningitis due to enterovirus.

    Science.gov (United States)

    Volle, Romain; Bailly, Jean-Luc; Mirand, Audrey; Pereira, Bruno; Marque-Juillet, Stéphanie; Chambon, Martine; Regagnon, Christel; Brebion, Amélie; Henquell, Cécile; Peigue-Lafeuille, Hélène; Archimbaud, Christine

    2014-08-15

    Acute enterovirus (EV) meningitis is a major cause of hospitalization among adults and children. It is caused by multiple EV genotypes assigned to 4 species (EV-A, EV-B, EV-C, and EV-D). We determined viral loads in the cerebrospinal fluid (CSF) of 156 patients of all ages with EV meningitis during a 5-year observational prospective study. The virus strains were genotyped, and their time origin was determined with Bayesian phylogenetic methods. The CSF viral loads ranged between 3.4 and 7.5 log10 copies/mL (median, 4.9 log10 copies/mL). They were higher in neonates than in infants and children (P = .02) but were comparable in adults. Viral loads were associated with EV genotypes (P < .001). The EV strains were identified in 152 of 156 patients and assigned to 23 genotypes within the EV-A and EV-B species. The most frequent genotypes, echoviruses 6 and 30, were associated with different viral loads (P < .001). The highest viral loads were in meningitis cases caused by coxsackievirus A9, B4, and B5 genotypes. Most patients infected by a same genotype were infected by a major virus variant of recent emergence. The variations in CSF viral loads in patients at the onset of EV meningitis are related to genotypic differences in the virus strains involved. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. Severe Enterovirus Infections in Hospitalized Children in the South of England: Clinical Phenotypes and Causative Genotypes.

    Science.gov (United States)

    de Graaf, Hans; Pelosi, Emanuela; Cooper, Andrea; Pappachan, John; Sykes, Kim; MacIntosh, Iain; Gbesemete, Diane; Clark, Tristan W; Patel, Sanjay V; Faust, Saul N; Tebruegge, Marc

    2016-07-01

    Most enterovirus surveillance studies lack detailed clinical data, which limits their clinical usefulness. This study aimed to describe the clinical spectrum and outcome of severe enterovirus infections in children, and to determine whether there are associations between causative enterovirus genotypes and clinical phenotypes. Retrospective analysis of microbiological and clinical data from a tertiary children's hospital in the South of England over a 17-month period (2012-2013). In total, 30 patients were identified, comprising sepsis (n = 9), myocarditis (n = 8), meningitis (n = 8) and encephalitis (n = 5). Cases with sepsis or myocarditis were significantly younger than those with central nervous system disease (median age 21 and 15 days vs. 79 days; P = 0.0244 and P = 0.0310, respectively). There was considerable diversity in the causative genotypes in each of the clinical phenotypes, with some predominance of echoviruses in the meningitis group, and coxsackie B viruses in the myocarditis group. Thirteen cases required mechanical ventilation, 11 cases inotropic support, 3 cases dialysis and 3 cases extracorporal membrane oxygenation. The overall mortality was 10% (sepsis group, n = 1; myocarditis group, n = 2). Of the survivors, 5 (19%) had long-term sequelae (myocardial dysfunction, n = 2; neurological sequelae, n = 3). Patients with encephalitis had the longest hospital stay (median: 16 days), compared with 9, 6 and 3 days in patients with myocarditis, sepsis and meningitis, respectively (P = 0.005). Enterovirus infections, particularly enteroviral myocarditis and encephalitis, can cause significant morbidity and mortality. The results show that there are currently no strong associations between clinical phenotypes and particular causative enterovirus genotypes in the South of England.

  7. Severe Enterovirus Infections in Hospitalized Children in the South of England

    Science.gov (United States)

    de Graaf, Hans; Pelosi, Emanuela; Cooper, Andrea; Pappachan, John; Sykes, Kim; MacIntosh, Iain; Gbesemete, Diane; Clark, Tristan W.; Patel, Sanjay V.; Faust, Saul N.

    2016-01-01

    Background: Most enterovirus surveillance studies lack detailed clinical data, which limits their clinical usefulness. This study aimed to describe the clinical spectrum and outcome of severe enterovirus infections in children, and to determine whether there are associations between causative enterovirus genotypes and clinical phenotypes. Methods: Retrospective analysis of microbiological and clinical data from a tertiary children’s hospital in the South of England over a 17-month period (2012–2013). Results: In total, 30 patients were identified, comprising sepsis (n = 9), myocarditis (n = 8), meningitis (n = 8) and encephalitis (n = 5). Cases with sepsis or myocarditis were significantly younger than those with central nervous system disease (median age 21 and 15 days vs. 79 days; P = 0.0244 and P = 0.0310, respectively). There was considerable diversity in the causative genotypes in each of the clinical phenotypes, with some predominance of echoviruses in the meningitis group, and coxsackie B viruses in the myocarditis group. Thirteen cases required mechanical ventilation, 11 cases inotropic support, 3 cases dialysis and 3 cases extracorporal membrane oxygenation. The overall mortality was 10% (sepsis group, n = 1; myocarditis group, n = 2). Of the survivors, 5 (19%) had long-term sequelae (myocardial dysfunction, n = 2; neurological sequelae, n = 3). Patients with encephalitis had the longest hospital stay (median: 16 days), compared with 9, 6 and 3 days in patients with myocarditis, sepsis and meningitis, respectively (P = 0.005). Conclusions: Enterovirus infections, particularly enteroviral myocarditis and encephalitis, can cause significant morbidity and mortality. The results show that there are currently no strong associations between clinical phenotypes and particular causative enterovirus genotypes in the South of England. PMID:26882165

  8. Enterovirus infections in Singaporean children: an assessment of neurological manifestations and clinical outcomes.

    Science.gov (United States)

    Thong, Wen Yi; Han, Audrey; Wang, S J Furene; Lin, Jeremy; Isa, Mas Suhaila; Koay, Evelyn Siew Chuan; Tay, Stacey Kiat-Hong

    2017-04-01

    Enterovirus infections in childhood can be associated with significant neurological morbidity. This study aimed to describe the prevalence and range of neurological manifestations, determine the clinical characteristics and assess differences in clinical outcomes for Singaporean children diagnosed with enterovirus infections. In this single-centre, case-control study, clinical data was collected retrospectively from patients admitted to National University Hospital, Singapore, from August 2007 to October 2011 and diagnosed with enterovirus infection, based on the enterovirus polymerase chain reaction test, or cultures from throat and rectal swabs or cerebrospinal fluid samples. The occurrence of neurological manifestations was reviewed and clinical outcomes were assessed. A total of 48 patients (age range: six days-17.8 years) were included in the study. Neurological manifestations were seen in 75.0% of patients, 63.9% of whom presented with aseptic meningitis. Other neurological manifestations included encephalitis, acute cerebellitis, transverse myelitis and autonomic dysfunction. The incidence of neurological manifestations was significantly higher in patients aged > 1 year as compared to younger patients (p = 0.043). In patients without neurological manifestations, a significantly higher proportion presented with hand, foot and mouth disease and poor feeding. Long-term neurological sequelae were seen in 16.7% of patients with neurological manifestations. A wide spectrum of neurological manifestations resulting in a relatively low incidence of long-term neurological sequelae was observed in our study of Singaporean children with enterovirus infections. As some of these neurological morbidities were severe, careful evaluation of children with neurological involvement is therefore necessary. Copyright: © Singapore Medical Association

  9. A novel enterovirus and parechovirus multiplex one-step real-time PCR-validation and clinical experience

    DEFF Research Database (Denmark)

    Nielsen, A. C. Y.; Bottiger, B.; Midgley, S. E.

    2013-01-01

    As the number of new enteroviruses and human parechoviruses seems ever growing, the necessity for updated diagnostics is relevant. We have updated an enterovirus assay and combined it with a previously published assay for human parechovirus resulting in a multiplex one-step RT-PCR assay....... The multiplex assay was validated by analysing the sensitivity and specificity of the assay compared to the respective monoplex assays, and a good concordance was found. Furthermore, the enterovirus assay was able to detect 42 reference strains from all 4 species, and an additional 9 genotypes during panel...... testing and routine usage. During 15 months of routine use, from October 2008 to December 2009, we received and analysed 2187 samples (stool samples, cerebrospinal fluids, blood samples, respiratory samples and autopsy samples) were tested, from 1546 patients and detected enteroviruses and parechoviruses...

  10. Polymorphisms in the innate immune IFIH1 gene, frequency of enterovirus in monthly fecal samples during infancy, and islet autoimmunity

    DEFF Research Database (Denmark)

    Witsø, Elisabet; Tapia, German; Cinek, Ondrej

    2011-01-01

    Interferon induced with helicase C domain 1 (IFIH1) senses and initiates antiviral activity against enteroviruses. Genetic variants of IFIH1, one common and four rare SNPs have been associated with lower risk for type 1 diabetes. Our aim was to test whether these type 1 diabetes-associated IFIH1...... polymorphisms are associated with the occurrence of enterovirus infection in the gut of healthy children, or influence the lack of association between gut enterovirus infection and islet autoimmunity.After testing of 46,939 Norwegian newborns, 421 children carrying the high risk genotype for type 1 diabetes...... (HLA-DR4-DQ8/DR3-DQ2) as well as 375 children without this genotype were included for monthly fecal collections from 3 to 35 months of age, and genotyped for the IFIH1 polymorphisms. A total of 7,793 fecal samples were tested for presence of enterovirus RNA using real time reverse transcriptase PCR...

  11. The evaluation of hollow-fiber ultrafiltration and celite concentration of enteroviruses, adenoviruses and bacteriophage from different water matrices

    Data.gov (United States)

    U.S. Environmental Protection Agency — The data to support the evaluation of hollow-fiber ultrafiltration and celite concentration of enteroviruses, adenoviruses and bacteriophage from different water...

  12. Method 1615: Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR

    Science.gov (United States)

    Version 1.1 - Enteroviruses and noroviruses that may be present in environmental or finished drinking waters are concentrated by passage through electropositive filters. Viruses are eluted from the filters with a beef extract reagent and concentrated using organic flocculation....

  13. Clinical Efficacy of Therapy with Recombinant Human Interferon α1b in Hand, Foot, and Mouth Disease with Enterovirus 71 Infection.

    Directory of Open Access Journals (Sweden)

    Xueyong Huang

    Full Text Available A rapid expansion of HFMD with enterovirus 71 infection outbreaks has occurred and caused deaths in recent years in China, but no vaccine or antiviral drug is currently available for EV71 infection. This study aims to provide treatment programs for HFMD patients. We conducted a randomized, double-blind, controlled trial and evaluated clinical efficacy of therapy with rHuIFN-α1b in HFMD patients with EV71 infection. There were statistical differences in outcomes including the fever clearance time, healing time of typical skin or oral mucosa lesions, and EV71 viral load of the HFMD patients among ultrasonic aerosol inhalation group, intramuscular injection group and control group. rHuIFN-α1b therapy reduced the fever clearance time, healing time of typical skin or oral mucosa lesions, and EV71 viral load in children with HFMD.Chinese Clinical Trial Registry ChiCTR-TRC-14005153.

  14. Rapid molecular screening of black carbon (biochar) thermosequences obtained from chestnut wood and rice straw: A pyrolysis-GC/MS study

    International Nuclear Information System (INIS)

    Kaal, Joeri; Schneider, Maximilian P.W.; Schmidt, Michael W.I.

    2012-01-01

    for rapid assessment of molecular properties of black C. ► Most thermal rearrangements occur between 250 and 450 °C charring temperature. ► Compounds from polysaccharides, lignin, chlorophyll, etc. recognised up to 500 °C. ► Relevant to predict black C degradation/preservation in soil (stability concept).

  15. Prohibitin plays a critical role in Enterovirus 71 neuropathogenesis.

    Directory of Open Access Journals (Sweden)

    Issac Horng Khit Too

    2018-01-01

    Full Text Available A close relative of poliovirus, enterovirus 71 (EV71 is regarded as an important neurotropic virus of serious public health concern. EV71 causes Hand, Foot and Mouth Disease and has been associated with neurological complications in young children. Our limited understanding of the mechanisms involved in its neuropathogenesis has hampered the development of effective therapeutic options. Here, using a two-dimensional proteomics approach combined with mass spectrometry, we have identified a unique panel of host proteins that were differentially and dynamically modulated during EV71 infection of motor-neuron NSC-34 cells, which are found at the neuromuscular junctions where EV71 is believed to enter the central nervous system. Meta-analysis with previously published proteomics studies in neuroblastoma or muscle cell lines revealed minimal overlapping which suggests unique host-pathogen interactions in NSC-34 cells. Among the candidate proteins, we focused our attention on prohibitin (PHB, a protein that is involved in multiple cellular functions and the target of anti-cancer drug Rocaglamide (Roc-A. We demonstrated that cell surface-expressed PHB is involved in EV71 entry into neuronal cells specifically, while membrane-bound mitochondrial PHB associates with the virus replication complex and facilitates viral replication. Furthermore, Roc-A treatment of EV71-infected neuronal cells reduced significantly virus yields. However, the inhibitory effect of Roc-A on PHB in NSC-34 cells was not through blocking the CRAF/MEK/ERK pathway as previously reported. Instead, Roc-A treated NSC-34 cells had lower mitochondria-associated PHB and lower ATP levels that correlated with impaired mitochondria integrity. In vivo, EV71-infected mice treated with Roc-A survived longer than the vehicle-treated animals and had significantly lower virus loads in their spinal cord and brain, whereas virus titers in their limb muscles were comparable to controls. Together, this

  16. Prohibitin plays a critical role in Enterovirus 71 neuropathogenesis

    Science.gov (United States)

    Too, Issac Horng Khit; Bonne, Isabelle; Tan, Eng Lee; Chu, Justin Jang Hann; Alonso, Sylvie

    2018-01-01

    A close relative of poliovirus, enterovirus 71 (EV71) is regarded as an important neurotropic virus of serious public health concern. EV71 causes Hand, Foot and Mouth Disease and has been associated with neurological complications in young children. Our limited understanding of the mechanisms involved in its neuropathogenesis has hampered the development of effective therapeutic options. Here, using a two-dimensional proteomics approach combined with mass spectrometry, we have identified a unique panel of host proteins that were differentially and dynamically modulated during EV71 infection of motor-neuron NSC-34 cells, which are found at the neuromuscular junctions where EV71 is believed to enter the central nervous system. Meta-analysis with previously published proteomics studies in neuroblastoma or muscle cell lines revealed minimal overlapping which suggests unique host-pathogen interactions in NSC-34 cells. Among the candidate proteins, we focused our attention on prohibitin (PHB), a protein that is involved in multiple cellular functions and the target of anti-cancer drug Rocaglamide (Roc-A). We demonstrated that cell surface-expressed PHB is involved in EV71 entry into neuronal cells specifically, while membrane-bound mitochondrial PHB associates with the virus replication complex and facilitates viral replication. Furthermore, Roc-A treatment of EV71-infected neuronal cells reduced significantly virus yields. However, the inhibitory effect of Roc-A on PHB in NSC-34 cells was not through blocking the CRAF/MEK/ERK pathway as previously reported. Instead, Roc-A treated NSC-34 cells had lower mitochondria-associated PHB and lower ATP levels that correlated with impaired mitochondria integrity. In vivo, EV71-infected mice treated with Roc-A survived longer than the vehicle-treated animals and had significantly lower virus loads in their spinal cord and brain, whereas virus titers in their limb muscles were comparable to controls. Together, this study uncovers

  17. Norepinephrine and Epinephrine Enhanced the Infectivity of Enterovirus 71.

    Directory of Open Access Journals (Sweden)

    Yu-Ting Liao

    Full Text Available Enterovirus 71 (EV71 infections may be associated with neurological complications, including brainstem encephalitis (BE. Severe EV71 BE may be complicated with autonomic nervous system (ANS dysregulation and/or pulmonary edema (PE. ANS dysregulation is related to the overactivation of the sympathetic nervous system, which results from catecholamine release.The aims of this study were to explore the effects of catecholamines on severe EV71 infection and to investigate the changes in the percentages of EV71-infected cells, virus titer, and cytokine production on the involvement of catecholamines.Plasma levels of norepinephrine (NE and epinephrine (EP in EV71-infected patients were measured using an enzyme-linked immunoassay. The expression of adrenergic receptors (ADRs on RD, A549, SK-N-SH, THP-1, Jurkat and human peripheral blood mononuclear cells (hPBMCs were detected using flow cytometry. The percentages of EV71-infected cells, virus titer, and cytokine production were investigated after treatment with NE and EP.The plasma levels of NE and EP were significantly higher in EV71-infected patients with ANS dysregulation and PE than in controls. Both α1A- and β2-ADRs were expressed on A549, RD, SK-N-SH, HL-60, THP-1, Jurkat cells and hPBMCs. NE treatment elevated the percentages of EV71-infected cells to 62.9% and 22.7% in THP-1 and Jurkat cells, respectively. Via treatment with EP, the percentages of EV71-infected cells were increased to 64.6% and 26.9% in THP-1 and Jurkat cells. The percentage of EV71-infected cells increased upon NE or EP treatment while the α- and β-blockers reduced the percentages of EV71-infected cells with NE or EP treatment. At least two-fold increase in virus titer was observed in EV71-infected A549, SK-N-SH and hPBMCs after treatment with NE or EP. IL-6 production was enhanced in EV71-infected hPBMCs at a concentration of 102 pg/mL NE.The plasma levels of NE and EP elevated in EV71-infected patients with ANS

  18. Norepinephrine and Epinephrine Enhanced the Infectivity of Enterovirus 71.

    Science.gov (United States)

    Liao, Yu-Ting; Wang, Shih-Min; Wang, Jen-Ren; Yu, Chun-Keung; Liu, Ching-Chuan

    2015-01-01

    Enterovirus 71 (EV71) infections may be associated with neurological complications, including brainstem encephalitis (BE). Severe EV71 BE may be complicated with autonomic nervous system (ANS) dysregulation and/or pulmonary edema (PE). ANS dysregulation is related to the overactivation of the sympathetic nervous system, which results from catecholamine release. The aims of this study were to explore the effects of catecholamines on severe EV71 infection and to investigate the changes in the percentages of EV71-infected cells, virus titer, and cytokine production on the involvement of catecholamines. Plasma levels of norepinephrine (NE) and epinephrine (EP) in EV71-infected patients were measured using an enzyme-linked immunoassay. The expression of adrenergic receptors (ADRs) on RD, A549, SK-N-SH, THP-1, Jurkat and human peripheral blood mononuclear cells (hPBMCs) were detected using flow cytometry. The percentages of EV71-infected cells, virus titer, and cytokine production were investigated after treatment with NE and EP. The plasma levels of NE and EP were significantly higher in EV71-infected patients with ANS dysregulation and PE than in controls. Both α1A- and β2-ADRs were expressed on A549, RD, SK-N-SH, HL-60, THP-1, Jurkat cells and hPBMCs. NE treatment elevated the percentages of EV71-infected cells to 62.9% and 22.7% in THP-1 and Jurkat cells, respectively. Via treatment with EP, the percentages of EV71-infected cells were increased to 64.6% and 26.9% in THP-1 and Jurkat cells. The percentage of EV71-infected cells increased upon NE or EP treatment while the α- and β-blockers reduced the percentages of EV71-infected cells with NE or EP treatment. At least two-fold increase in virus titer was observed in EV71-infected A549, SK-N-SH and hPBMCs after treatment with NE or EP. IL-6 production was enhanced in EV71-infected hPBMCs at a concentration of 102 pg/mL NE. The plasma levels of NE and EP elevated in EV71-infected patients with ANS dysregulation and

  19. A Selective Bottleneck Shapes the Evolutionary Mutant Spectra of Enterovirus A71 during Viral Dissemination in Humans.

    Science.gov (United States)

    Huang, Sheng-Wen; Huang, Yi-Hui; Tsai, Huey-Pin; Kuo, Pin-Hwa; Wang, Shih-Min; Liu, Ching-Chuan; Wang, Jen-Ren

    2017-12-01

    RNA viruses accumulate mutations to rapidly adapt to environmental changes. Enterovirus A71 (EV-A71) causes various clinical manifestations with occasional severe neurological complications. However, the mechanism by which EV-A71 evolves within the human body is unclear. Utilizing deep sequencing and haplotype analyses of viruses from various tissues of an autopsy patient, we sought to define the evolutionary pathway by which enterovirus A71 evolves fitness for invading the central nervous system in humans. Broad mutant spectra with divergent mutations were observed at the initial infection sites in the respiratory and digestive systems. After viral invasion, we identified a haplotype switch and dominant haplotype, with glycine at VP1 residue 31 (VP1-31G) in viral particles disseminated into the integumentary and central nervous systems. In vitro viral growth and fitness analyses indicated that VP1-31G conferred growth and a fitness advantage in human neuronal cells, whereas VP1-31D conferred enhanced replication in human colorectal cells. A higher proportion of VP1-31G was also found among fatal cases, suggesting that it may facilitate central nervous system infection in humans. Our data provide the first glimpse of EV-A71 quasispecies from oral tissues to the central nervous system within humans, showing broad implications for the surveillance and pathogenesis of this reemerging viral pathogen. IMPORTANCE EV-A71 continues to be a worldwide burden to public health. Although EV-A71 is the major etiological agent of hand, foot, and mouth disease, it can also cause neurological pulmonary edema, encephalitis, and even death, especially in children. Understanding selection processes enabling dissemination and accurately estimating EV-A71 diversity during invasion in humans are critical for applications in viral pathogenesis and vaccine studies. Here, we define a selection bottleneck appearing in respiratory and digestive tissues. Glycine substitution at VP1 residue 31

  20. Reaaliaikaisen RT-PCR-menetelmän käyttöönotto ja validointi enterovirus A71:n tunnistamiseksi

    OpenAIRE

    Ruuska, Janika

    2017-01-01

    Vuosina 2011 ja 2012 Kiinassa raportoitiin noin 4 miljoonaa enterorokkoon sairastunutta, joista reilu tuhat johti kuolemaan. Enterorokon tavallisimpia aiheuttajia ovat coxackievirus A16 ja enterovirus A71. Näistä kahdesta enterovirus A71-infektiot saattavat aiheuttaa vaikeitakin neurologisia sairauksia, joita ei tavallisesti aiheudu coxackievirus A16-infektioista. Terveyden ja hyvinvoinnin laitoksessa enterovirus A71:n osoitus on tehty virusviljelyn avulla ja kasvatettu virus on tyypitetty se...

  1. A novel enterovirus and parechovirus multiplex one-step real-time PCR-validation and clinical experience.

    Science.gov (United States)

    Nielsen, Alex Christian Yde; Böttiger, Blenda; Midgley, Sofie Elisabeth; Nielsen, Lars Peter

    2013-11-01

    As the number of new enteroviruses and human parechoviruses seems ever growing, the necessity for updated diagnostics is relevant. We have updated an enterovirus assay and combined it with a previously published assay for human parechovirus resulting in a multiplex one-step RT-PCR assay. The multiplex assay was validated by analysing the sensitivity and specificity of the assay compared to the respective monoplex assays, and a good concordance was found. Furthermore, the enterovirus assay was able to detect 42 reference strains from all 4 species, and an additional 9 genotypes during panel testing and routine usage. During 15 months of routine use, from October 2008 to December 2009, we received and analysed 2187 samples (stool samples, cerebrospinal fluids, blood samples, respiratory samples and autopsy samples) were tested, from 1546 patients and detected enteroviruses and parechoviruses in 171 (8%) and 66 (3%) of the samples, respectively. 180 of the positive samples could be genotyped by PCR and sequencing and the most common genotypes found were human parechovirus type 3, echovirus 9, enterovirus 71, Coxsackievirus A16, and echovirus 25. During 2009 in Denmark, both enterovirus and human parechovirus type 3 had a similar seasonal pattern with a peak during the summer and autumn. Human parechovirus type 3 was almost invariably found in children less than 4 months of age. In conclusion, a multiplex assay was developed allowing simultaneous detection of 2 viruses, which can cause similar clinical symptoms. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. Immunopathogenesis and Virus–Host Interactions of Enterovirus 71 in Patients with Hand, Foot and Mouth Disease

    Science.gov (United States)

    Cox, Jonathan A.; Hiscox, Julian A.; Solomon, Tom; Ooi, Mong-How; Ng, Lisa F. P.

    2017-01-01

    Enterovirus 71 (EV71) is a global infectious disease that affects millions of people. The virus is the main etiological agent for hand, foot, and mouth disease with outbreaks and epidemics being reported globally. Infection can cause severe neurological, cardiac, and respiratory problems in children under the age of 5. Despite on-going efforts, little is known about the pathogenesis of EV71, how the host immune system responds to the virus and the molecular mechanisms behind these responses. Moreover, current animal models remain limited, because they do not recapitulate similar disease patterns and symptoms observed in humans. In this review the role of the host–viral interactions of EV71 are discussed together with the various models available to examine: how EV71 utilizes its proteins to cleave host factors and proteins, aiding virus replication; how EV71 uses its own viral proteins to disrupt host immune responses and aid in its immune evasion. These discoveries along with others, such as the EV71 crystal structure, have provided possible targets for treatment and drug interventions. PMID:29238324

  3. Severe respiratory illness associated with a nationwide outbreak of enterovirus D68 in the USA (2014): a descriptive epidemiological investigation

    Science.gov (United States)

    Midgley, Claire M; Watson, John T; Nix, W Allan; Curns, Aaron T; Rogers, Shannon L; Brown, Betty A; Conover, Craig; Dominguez, Samuel R; Feikin, Daniel R; Gray, Samantha; Hassan, Ferdaus; Hoferka, Stacey; Jackson, Mary Anne; Johnson, Daniel; Leshem, Eyal; Miller, Lisa; Nichols, Janell Bezdek; Nyquist, Ann-Christine; Obringer, Emily; Patel, Ajanta; Patel, Megan; Rha, Brian; Schneider, Eileen; Schuster, Jennifer E; Selvarangan, Rangaraj; Seward, Jane F; Turabelidze, George; Oberste, M Steven; Pallansch, Mark A; Gerber, Susan I

    2016-01-01

    Summary Background Enterovirus D68 (EV-D68) has been infrequently reported historically, and is typically associated with isolated cases or small clusters of respiratory illness. Beginning in August, 2014, increases in severe respiratory illness associated with EV-D68 were reported across the USA. We aimed to describe the clinical, epidemiological, and laboratory features of this outbreak, and to better understand the role of EV-D68 in severe respiratory illness. Methods We collected regional syndromic surveillance data for epidemiological weeks 23 to 44, 2014, (June 1 to Nov 1, 2014) and hospital admissions data for epidemiological weeks 27 to 44, 2014, (June 29 to Nov 1, 2014) from three states: Missouri, Illinois and Colorado. Data were also collected for the same time period of 2013 and 2012. Respiratory specimens from severely ill patients nationwide, who were rhinovirus-positive or enterovirus-positive in hospital testing, were submitted between Aug 1, and Oct 31, 2014, and typed by molecular sequencing. We collected basic clinical and epidemiological characteristics of EV-D68 cases with a standard data collection form submitted with each specimen. We compared patients requiring intensive care with those who did not, and patients requiring ventilator support with those who did not. Mantel-Haenszel χ2 tests were used to test for statistical significance. Findings Regional and hospital-level data from Missouri, Illinois, and Colorado showed increases in respiratory illness between August and September, 2014, compared with in 2013 and 2012. Nationwide, 699 (46%) of 1529 patients tested were confirmed as EV-D68. Among the 614 EV-D68-positive patients admitted to hospital, age ranged from 3 days to 92 years (median 5 years). Common symptoms included dyspnoea (n=513 [84%]), cough (n=500 [81%]), and wheezing (n=427 [70%]); 294 (48%) patients had fever. 338 [59%] of 574 were admitted to intensive care units, and 145 (28%) of 511 received ventilator support; 322 (52

  4. Rapid and simple detection of foot-and-mouth disease virus: Evaluation of a cartridge-based molecular detection system for use in basic laboratories.

    Science.gov (United States)

    Goller, K V; Dill, V; Madi, M; Martin, P; Van der Stede, Y; Vandenberge, V; Haas, B; Van Borm, S; Koenen, F; Kasanga, C J; Ndusilo, N; Beer, M; Liu, L; Mioulet, V; Armson, B; King, D P; Fowler, V L

    2018-04-01

    Highly contagious transboundary animal diseases such as foot-and-mouth disease (FMD) are major threats to the productivity of farm animals. To limit the impact of outbreaks and to take efficient steps towards a timely control and eradication of the disease, rapid and reliable diagnostic systems are of utmost importance. Confirmatory diagnostic assays are typically performed by experienced operators in specialized laboratories, and access to this capability is often limited in the developing countries with the highest disease burden. Advances in molecular technologies allow implementation of modern and reliable techniques for quick and simple pathogen detection either in basic laboratories or even at the pen-side. Here, we report on a study to evaluate a fully automated cartridge-based real-time RT-PCR diagnostic system (Enigma MiniLab ® ) for the detection of FMD virus (FMDV). The modular system integrates both nucleic acid extraction and downstream real-time RT-PCR (rRT-PCR). The analytical sensitivity of this assay was determined using serially diluted culture grown FMDV, and the performance of the assay was evaluated using a selected range of FMDV positive and negative clinical samples of bovine, porcine and ovine origin. The robustness of the assay was evaluated in an international inter-laboratory proficiency test and by deployment into an African laboratory. It was demonstrated that the system is easy to use and can detect FMDV with high sensitivity and specificity, roughly on par with standard laboratory methods. This cartridge-based automated real-time RT-PCR system for the detection of FMDV represents a reliable and easy to use diagnostic tool for the early and rapid disease detection of acutely infected animals even in remote areas. This type of system could be easily deployed for routine surveillance within endemic regions such as Africa or could alternatively be used in the developed world. © 2017 The Authors. Transboundary and Emerging Diseases

  5. Rapid identification of ascomycetous yeasts from clinical specimens by a molecular method based on flow cytometry and comparison with identifications from phenotypic assays.

    Science.gov (United States)

    Page, Brent T; Shields, Christine E; Merz, William G; Kurtzman, Cletus P

    2006-09-01

    This study was designed to compare the identification of ascomycetous yeasts recovered from clinical specimens by using phenotypic assays (PA) and a molecular flow cytometric (FC) method. Large-subunit rRNA domains 1 and 2 (D1/D2) gene sequence analysis was also performed and served as the reference for correct strain identification. A panel of 88 clinical isolates was tested that included representatives of nine commonly encountered species and six infrequently encountered species. The PA included germ tube production, fermentation of seven carbohydrates, morphology on corn meal agar, urease and phenoloxidase activities, and carbohydrate assimilation tests when needed. The FC method (Luminex) employed species-specific oligonucleotides attached to polystyrene beads, which were hybridized with D1/D2 amplicons from the unidentified isolates. The PA identified 81 of 88 strains correctly but misidentified 4 of Candida dubliniensis, 1 of C. bovina, 1 of C. palmioleophila, and 1 of C. bracarensis. The FC method correctly identified 79 of 88 strains and did not misidentify any isolate but did not identify nine isolates because oligonucleotide probes were not available in the current library. The FC assay takes approximately 5 h, whereas the PA takes from 2 h to 5 days for identification. In conclusion, PA did well with the commonly encountered species, was not accurate for uncommon species, and takes significantly longer than the FC method. These data strongly support the potential of FC technology for rapid and accurate identification of medically important yeasts. With the introduction of new antifungals, rapid, accurate identification of pathogenic yeasts is more important than ever for guiding antifungal chemotherapy.

  6. A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax.

    Science.gov (United States)

    Zhang, Xinxin; Ma, Dehua; Zou, Wei; Ding, Yibing; Zhu, Chengchu; Min, Haiyan; Zhang, Bin; Wang, Wei; Chen, Baofu; Ye, Minhua; Cai, Minghui; Pan, Yanqing; Cao, Lei; Wan, Yueming; Jin, Yu; Gao, Qian; Yi, Long

    2016-05-27

    Primary spontaneous pneumothorax (PSP) or pulmonary cysts is one of the manifestations of Birt-Hogg-Dube syndrome (BHDS) that is caused by heterozygous mutations in FLCN gene. Most of the mutations are SNVs and small indels, and there are also approximately 10 % large intragenic deletions and duplications of the mutations. These molecular findings are generally obtained by disparate methods including Sanger sequencing and Multiple Ligation-dependent Probe Amplification in the clinical laboratory. In addition, as a genetically heterogeneous disorder, PSP may be caused by mutations in multiple genes include FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 genes. For differential diagnosis, these genes should also be screened which makes the diagnostic procedure more time-consuming and labor-intensive. Forty PSP patients were divided into 2 groups. Nineteen patients with different pathogenic mutations of FLCN previously identified by conventional Sanger sequencing and MLPA were included in test group, 21 random PSP patients without any genetic screening were included in blinded sample group. 7 PSP genes including FLCN, FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 were designed and enriched by Haloplex system, sequenced on a Miseq platform and analyzed in the 40 patients to evaluate the performance of the targeted-NGS method. We demonstrated that the full spectrum of genes associated with pneumothorax including FLCN gene mutations can be identified simultaneously in multiplexed sequence data. Noteworthy, by our in-house copy number analysis of the sequence data, we could not only detect intragenic deletions, but also determine approximate deletion junctions simultaneously. NGS based Haloplex target enrichment technology is proved to be a rapid and cost-effective screening strategy for the comprehensive molecular diagnosis of BHDS in PSP patients, as it can replace Sanger sequencing and MLPA by simultaneously detecting exonic and intronic SNVs, small indels, large intragenic

  7. Phylodynamics of Enterovirus A71-Associated Hand, Foot, and Mouth Disease in Viet Nam.

    Science.gov (United States)

    Geoghegan, Jemma L; Tan, Le Van; Kühnert, Denise; Halpin, Rebecca A; Lin, Xudong; Simenauer, Ari; Akopov, Asmik; Das, Suman R; Stockwell, Timothy B; Shrivastava, Susmita; Ngoc, Nghiem My; Uyen, Le Thi Tam; Tuyen, Nguyen Thi Kim; Thanh, Tran Tan; Hang, Vu Thi Ty; Qui, Phan Tu; Hung, Nguyen Thanh; Khanh, Truong Huu; Thinh, Le Quoc; Nhan, Le Nguyen Thanh; Van, Hoang Minh Tu; Viet, Do Chau; Tuan, Ha Manh; Viet, Ho Lu; Hien, Tran Tinh; Chau, Nguyen Van Vinh; Thwaites, Guy; Grenfell, Bryan T; Stadler, Tanja; Wentworth, David E; Holmes, Edward C; Van Doorn, H Rogier

    2015-09-01

    Enterovirus A71 (EV-A71) is a major cause of hand, foot, and mouth disease (HFMD) and is particularly prevalent in parts of Southeast Asia, affecting thousands of children and infants each year. Revealing the evolutionary and epidemiological dynamics of EV-A71 through time and space is central to understanding its outbreak potential. We generated the full genome sequences of 200 EV-A71 strains sampled from various locations in Viet Nam between 2011 and 2013 and used these sequence data to determine the evolutionary history and phylodynamics of EV-A71 in Viet Nam, providing estimates of the effective reproduction number (Re) of the infection through time. In addition, we described the phylogeography of EV-A71 throughout Southeast Asia, documenting patterns of viral gene flow. Accordingly, our analysis reveals that a rapid genogroup switch from C4 to B5 likely took place during 2012 in Viet Nam. We show that the Re of subgenogroup C4 decreased during the time frame of sampling, whereas that of B5 increased and remained >1 at the end of 2013, corresponding to a rise in B5 prevalence. Our study reveals that the subgenogroup B5 virus that emerged into Viet Nam is closely related to variants that were responsible for large epidemics in Malaysia and Taiwan and therefore extends our knowledge regarding its associated area of endemicity. Subgenogroup B5 evidently has the potential to cause more widespread outbreaks across Southeast Asia. EV-A71 is one of many viruses that cause HFMD, a common syndrome that largely affects infants and children. HFMD usually causes only mild illness with no long-term consequences. Occasionally, however, severe infection may arise, especially in very young children, causing neurological complications and even death. EV-A71 is highly contagious and is associated with the most severe HFMD cases, with large and frequent epidemics of the virus recorded worldwide. Although major advances have been made in the development of a potential EV-A71

  8. Molecular initiating events of the intersex phenotype: Low-dose exposure to 17α-ethinylestradiol rapidly regulates molecular networks associated with gonad differentiation in the adult fathead minnow testis

    International Nuclear Information System (INIS)

    Feswick, April; Loughery, Jennifer R.; Isaacs, Meghan A.; Munkittrick, Kelly R.; Martyniuk, Christopher J.

    2016-01-01

    testis, despite the fact that dmrt1 itself was not different in expression from control males. Transcriptional networks involving forkhead box L2 (foxl2) (transcript involved in ovarian follicle development) were increased in expression in the testis. Noteworthy was that a gene network associated to granulosa cell development was increased over 100%, suggesting that this transcriptome network may be important for monitoring estrogenic exposures. Other cell processes rapidly downregulated by EE2 at the transcript level included glucose homeostasis, response to heavy metal, amino acid catabolism, and the cyclooxygenase pathway. Conversely, lymphocyte chemotaxis, intermediate filament polymerization, glucocorticoid metabolism, carbohydrate utilization, and anterior/posterior axis specification were increased. These data provide new insight into the transcriptional responses that are perturbed prior to gonadal remodeling and intersex following exposure to estrogens. These data demonstrate that low concentrations of EE2 (1) rapidly suppresses male hormone production, (2) down-regulate molecular networks related to male sex differentiation, and (3) induce transcriptional networks related to granulosa cell development in the adult testis. These responses are hypothesized to be key molecular initiating events that occur prior to the development of the intersex phenotype following estrogenic exposures.

  9. Molecular initiating events of the intersex phenotype: Low-dose exposure to 17α-ethinylestradiol rapidly regulates molecular networks associated with gonad differentiation in the adult fathead minnow testis

    Energy Technology Data Exchange (ETDEWEB)

    Feswick, April; Loughery, Jennifer R.; Isaacs, Meghan A.; Munkittrick, Kelly R.; Martyniuk, Christopher J., E-mail: cmartyni@yahoo.ca

    2016-12-15

    testis, despite the fact that dmrt1 itself was not different in expression from control males. Transcriptional networks involving forkhead box L2 (foxl2) (transcript involved in ovarian follicle development) were increased in expression in the testis. Noteworthy was that a gene network associated to granulosa cell development was increased over 100%, suggesting that this transcriptome network may be important for monitoring estrogenic exposures. Other cell processes rapidly downregulated by EE2 at the transcript level included glucose homeostasis, response to heavy metal, amino acid catabolism, and the cyclooxygenase pathway. Conversely, lymphocyte chemotaxis, intermediate filament polymerization, glucocorticoid metabolism, carbohydrate utilization, and anterior/posterior axis specification were increased. These data provide new insight into the transcriptional responses that are perturbed prior to gonadal remodeling and intersex following exposure to estrogens. These data demonstrate that low concentrations of EE2 (1) rapidly suppresses male hormone production, (2) down-regulate molecular networks related to male sex differentiation, and (3) induce transcriptional networks related to granulosa cell development in the adult testis. These responses are hypothesized to be key molecular initiating events that occur prior to the development of the intersex phenotype following estrogenic exposures.

  10. Antiviral effects of two Ganoderma lucidum triterpenoids against enterovirus 71 infection

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Wenjing; Tao, Junyan; Yang, Xiaoping [State Key Laboratory of Virology and College of Life Sciences, Wuhan University, Wuhan 430072 (China); Yang, Zhuliang [Key Laboratory of Biodiversity and Biogeography, Kunming Institute of Botany, Chinese Academy of Science, Kunming 650201 (China); Zhang, Li; Liu, Hongsheng [Department of Academy of Sciences, Liaoning University, Shenyang 110036 (China); Wu, Kailang [State Key Laboratory of Virology and College of Life Sciences, Wuhan University, Wuhan 430072 (China); Wu, Jianguo, E-mail: jwu@whu.edu.cn [State Key Laboratory of Virology and College of Life Sciences, Wuhan University, Wuhan 430072 (China)

    2014-07-04

    Highlights: • Triterpenoids GLTA and GLTB display anti-EV71 activities without cytotoxicity. • The compounds prevent EV71 infection by blocking adsorption of the virus to the cells. • GLTA and GLTB bind to EV71 capsid at the hydrophobic pocket to block EV71 uncoating. • The two compounds significantly inhibit the replication of EV71 viral RNA. • GLTA and GLTB may be used as potential therapeutic agents to treat EV71 infection. - Abstract: Enterovirus 71 (EV71) is a major causative agent for hand, foot and mouth disease (HFMD), and fatal neurological and systemic complications in children. However, there is currently no clinical approved antiviral drug available for the prevention and treatment of the viral infection. Here, we evaluated the antiviral activities of two Ganoderma lucidum triterpenoids (GLTs), Lanosta-7,9(11),24-trien-3-one,15;26-dihydroxy (GLTA) and Ganoderic acid Y (GLTB), against EV71 infection. The results showed that the two natural compounds display significant anti-EV71 activities without cytotoxicity in human rhabdomyosarcoma (RD) cells as evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay. The mechanisms by which the two compounds affect EV71 infection were further elucidated by three action modes using Ribavirin, a common antiviral drug, as a positive control. The results suggested that GLTA and GLTB prevent EV71 infection through interacting with the viral particle to block the adsorption of virus to the cells. In addition, the interactions between EV71 virion and the compounds were predicated by computer molecular docking, which illustrated that GLTA and GLTB may bind to the viral capsid protein at a hydrophobic pocket (F site), and thus may block uncoating of EV71. Moreover, we demonstrated that GLTA and GLTB significantly inhibit the replication of the viral RNA (vRNA) of EV71 replication through blocking EV71 uncoating. Thus, GLTA and GLTB may represent two potential

  11. Antiviral effects of two Ganoderma lucidum triterpenoids against enterovirus 71 infection

    International Nuclear Information System (INIS)

    Zhang, Wenjing; Tao, Junyan; Yang, Xiaoping; Yang, Zhuliang; Zhang, Li; Liu, Hongsheng; Wu, Kailang; Wu, Jianguo

    2014-01-01

    Highlights: • Triterpenoids GLTA and GLTB display anti-EV71 activities without cytotoxicity. • The compounds prevent EV71 infection by blocking adsorption of the virus to the cells. • GLTA and GLTB bind to EV71 capsid at the hydrophobic pocket to block EV71 uncoating. • The two compounds significantly inhibit the replication of EV71 viral RNA. • GLTA and GLTB may be used as potential therapeutic agents to treat EV71 infection. - Abstract: Enterovirus 71 (EV71) is a major causative agent for hand, foot and mouth disease (HFMD), and fatal neurological and systemic complications in children. However, there is currently no clinical approved antiviral drug available for the prevention and treatment of the viral infection. Here, we evaluated the antiviral activities of two Ganoderma lucidum triterpenoids (GLTs), Lanosta-7,9(11),24-trien-3-one,15;26-dihydroxy (GLTA) and Ganoderic acid Y (GLTB), against EV71 infection. The results showed that the two natural compounds display significant anti-EV71 activities without cytotoxicity in human rhabdomyosarcoma (RD) cells as evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay. The mechanisms by which the two compounds affect EV71 infection were further elucidated by three action modes using Ribavirin, a common antiviral drug, as a positive control. The results suggested that GLTA and GLTB prevent EV71 infection through interacting with the viral particle to block the adsorption of virus to the cells. In addition, the interactions between EV71 virion and the compounds were predicated by computer molecular docking, which illustrated that GLTA and GLTB may bind to the viral capsid protein at a hydrophobic pocket (F site), and thus may block uncoating of EV71. Moreover, we demonstrated that GLTA and GLTB significantly inhibit the replication of the viral RNA (vRNA) of EV71 replication through blocking EV71 uncoating. Thus, GLTA and GLTB may represent two potential

  12. Rapid Convergence of Energy and Free Energy Profiles with Quantum Mechanical Size in Quantum Mechanical-Molecular Mechanical Simulations of Proton Transfer in DNA.

    Science.gov (United States)

    Das, Susanta; Nam, Kwangho; Major, Dan Thomas

    2018-03-13

    In recent years, a number of quantum mechanical-molecular mechanical (QM/MM) enzyme studies have investigated the dependence of reaction energetics on the size of the QM region using energy and free energy calculations. In this study, we revisit the question of QM region size dependence in QM/MM simulations within the context of energy and free energy calculations using a proton transfer in a DNA base pair as a test case. In the simulations, the QM region was treated with a dispersion-corrected AM1/d-PhoT Hamiltonian, which was developed to accurately describe phosphoryl and proton transfer reactions, in conjunction with an electrostatic embedding scheme using the particle-mesh Ewald summation method. With this rigorous QM/MM potential, we performed rather extensive QM/MM sampling, and found that the free energy reaction profiles converge rapidly with respect to the QM region size within ca. ±1 kcal/mol. This finding suggests that the strategy of QM/MM simulations with reasonably sized and selected QM regions, which has been employed for over four decades, is a valid approach for modeling complex biomolecular systems. We point to possible causes for the sensitivity of the energy and free energy calculations to the size of the QM region, and potential implications.

  13. A novel molecularly imprinted material based on magnetic halloysite nanotubes for rapid enrichment of 2,4-dichlorophenoxyacetic acid in water.

    Science.gov (United States)

    Zhong, Shian; Zhou, Chengyun; Zhang, Xiaona; Zhou, Hui; Li, Hui; Zhu, Xiaohong; Wang, Yan

    2014-07-15

    A new type of magnetic halloysite nanotubes molecularly imprinted polymer (MHNTs@MIP) based on halloysite nanotubes (HNTs) with embedded magnetic nanoparticles was introduced in this study. MHNTs@MIP was prepared through surface imprinting technology, using 2,4-dichlorophenoxyacetic acid (2,4-D) as a template, 4-vinylpyridine as the monomer, divinylbenzene as cross-linking agents, and 2,2-azodiisobutyronitrile as initiator. MHNTs@MIP was characterized by Fourier Transform Infrared Spectrometer, transmission electron microscopy, X-ray diffraction, and vibrating sample magnetometer. MHNTs@MIP exhibited rapid and reliable analysis with supermagnetic properties, as well as repeated use and template-specific recognition. The adsorption capacity of magnetic halloysite nanotubes non-imprinted polymer (MHNTs@NIP) and MHNTs@MIP was 10.3mg/g and 35.2mg/g, respectively. In the detailed discussion on specific selectivity, MHNTs@MIP can be applied as an adsorbent for sample pretreatment extraction and obtain high recoveries of about 85-94%. After extraction, high-performance liquid chromatography was used to detect 2,4-D residue in water. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Rapid, High-Throughput, and Direct Molecular Beacon Delivery to Human Cancer Cells Using a Nanowire-Incorporated and Pneumatic Pressure-Driven Microdevice.

    Science.gov (United States)

    Kim, Kyung Hoon; Kim, Jung; Choi, Jong Seob; Bae, Sunwoong; Kwon, Donguk; Park, Inkyu; Kim, Do Hyun; Seo, Tae Seok

    2015-12-01

    Tracking and monitoring the intracellular behavior of mRNA is of paramount importance for understanding real-time gene expression in cell biology. To detect specific mRNA sequences, molecular beacons (MBs) have been widely employed as sensing probes. Although numerous strategies for MB delivery into the target cells have been reported, many issues such as the cytotoxicity of the carriers, dependence on the random probability of MB transfer, and critical cellular damage still need to be overcome. Herein, we have developed a nanowire-incorporated and pneumatic pressure-driven microdevice for rapid, high-throughput, and direct MB delivery to human breast cancer MCF-7 cells to monitor survivin mRNA expression. The proposed microdevice is composed of three layers: a pump-associated glass manifold layer, a monolithic polydimethylsiloxane (PDMS) membrane, and a ZnO nanowire-patterned microchannel layer. The MB is immobilized on the ZnO nanowires by disulfide bonding, and the glass manifold and PDMS membrane serve as a microvalve, so that the cellular attachment and detachment on the MB-coated nanowire array can be manipulated. The combination of the nanowire-mediated MB delivery and the microvalve function enable the transfer of MB into the cells in a controllable way with high cell viability and to detect survivin mRNA expression quantitatively after docetaxel treatment. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Co-Circulation and Genomic Recombination of Coxsackievirus A16 and Enterovirus 71 during a Large Outbreak of Hand, Foot, and Mouth Disease in Central China

    Science.gov (United States)

    Liu, Weiyong; Wu, Shimin; Xiong, Ying; Li, Tongya; Wen, Zhou; Yan, Mingzhe; Qin, Kai; Liu, Yingle; Wu, Jianguo

    2014-01-01

    A total of 1844 patients with hand, foot, and mouth disease (HFMD), most of them were children of age 1–3-year-old, in Central China were hospitalized from 2011 to 2012. Among them, 422 were infected with coxsackievirus A16 (CVA16), 334 were infected with enterovirus 71 (EV71), 38 were co-infected with EV71 and CVA16, and 35 were infected with other enteroviruses. Molecular epidemiology analysis revealed that EV71 and CVA16 were detected year-round, but EV71 circulated mainly in July and CVA16 circulated predominantly in November, and incidence of HFMD was reduced in January and February and increased in March. Clinical data showed that hyperglycemia and neurologic complications were significantly higher in EV71-infected patients, while upper respiratory tract infection and C-reactive protein were significantly higher in CVA16-associated patients. 124 EV71 and 80 CVA16 strains were isolated, among them 56 and 68 EV71 strains were C4a and C4b, while 25 and 55 CVA16 strains were B1a and B1b, respectively. Similarity plots and bootscan analyses based on entire genomic sequences revealed that the three C4a sub-genotype EV71 strains were recombinant with C4b sub-genotype EV71 in 2B–2C region, and the three CVA16 strains were recombinant with EV71 in 2A–2B region. Thus, CVA16 and EV71 were the major causative agents in a large HFMD outbreak in Central China. HFMD incidence was high for children among household contact and was detected year-round, but outbreak was seasonal dependent. CVA16 B1b and EV71 C4b reemerged and caused a large epidemic in China after a quiet period of many years. Moreover, EV71 and CVA16 were co-circulated during the outbreak, which may have contributed to the genomic recombination between the pathogens. It should gain more attention as there may be an upward trend in co-circulation of the two pathogens globally and the new role recombination plays in the emergence of new enterovirus variants. PMID:24776922

  16. Co-circulation and genomic recombination of coxsackievirus A16 and enterovirus 71 during a large outbreak of hand, foot, and mouth disease in Central China.

    Directory of Open Access Journals (Sweden)

    Weiyong Liu

    Full Text Available A total of 1844 patients with hand, foot, and mouth disease (HFMD, most of them were children of age 1-3-year-old, in Central China were hospitalized from 2011 to 2012. Among them, 422 were infected with coxsackievirus A16 (CVA16, 334 were infected with enterovirus 71 (EV71, 38 were co-infected with EV71 and CVA16, and 35 were infected with other enteroviruses. Molecular epidemiology analysis revealed that EV71 and CVA16 were detected year-round, but EV71 circulated mainly in July and CVA16 circulated predominantly in November, and incidence of HFMD was reduced in January and February and increased in March. Clinical data showed that hyperglycemia and neurologic complications were significantly higher in EV71-infected patients, while upper respiratory tract infection and C-reactive protein were significantly higher in CVA16-associated patients. 124 EV71 and 80 CVA16 strains were isolated, among them 56 and 68 EV71 strains were C4a and C4b, while 25 and 55 CVA16 strains were B1a and B1b, respectively. Similarity plots and bootscan analyses based on entire genomic sequences revealed that the three C4a sub-genotype EV71 strains were recombinant with C4b sub-genotype EV71 in 2B-2C region, and the three CVA16 strains were recombinant with EV71 in 2A-2B region. Thus, CVA16 and EV71 were the major causative agents in a large HFMD outbreak in Central China. HFMD incidence was high for children among household contact and was detected year-round, but outbreak was seasonal dependent. CVA16 B1b and EV71 C4b reemerged and caused a large epidemic in China after a quiet period of many years. Moreover, EV71 and CVA16 were co-circulated during the outbreak, which may have contributed to the genomic recombination between the pathogens. It should gain more attention as there may be an upward trend in co-circulation of the two pathogens globally and the new role recombination plays in the emergence of new enterovirus variants.

  17. Innate Immunity Evasion by Enteroviruses: Insights into Virus-Host Interaction

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    Xiaobo Lei

    2016-01-01

    Full Text Available Enterovirus genus includes multiple important human pathogens, such as poliovirus, coxsackievirus, enterovirus (EV A71, EV-D68 and rhinovirus. Infection with EVs can cause numerous clinical conditions including poliomyelitis, meningitis and encephalitis, hand-foot-and-mouth disease, acute flaccid paralysis, diarrhea, myocarditis and respiratory illness. EVs, which are positive-sense single-stranded RNA viruses, trigger activation of the host antiviral innate immune responses through pathogen recognition receptors such as retinoic acid-inducible gene (RIG-I-likeand Toll-like receptors. In turn, EVs have developed sophisticated strategies to evade host antiviral responses. In this review, we discuss the interplay between the host innate immune responses and EV infection, with a primary focus on host immune detection and protection against EV infection and viral strategies to evade these antiviral immune responses.

  18. Large outbreak of herpangina in children caused by enterovirus in summer of 2015 in Hangzhou, China.

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    Li, Wei; Gao, Hui-Hui; Zhang, Qiong; Liu, Yu-Jie; Tao, Ran; Cheng, Yu-Ping; Shu, Qiang; Shang, Shi-Qiang

    2016-10-18

    Herpangina, usually caused by coxsackie virus A, is prevalent in children spreading through the fecal-oral transmission and the respiratory droplets dissemination. Also, it is mostly asymptomatic and self-limiting. In our study, we found that large outbreak of herpangina in children occurred in the summer of 2015 in Hangzhou, China. From May 1th to August 31th, a total of 10 210 children were diagnosed with herpangina in Children's Hospital of Zhejiang University School of Medicine. 2 310 throat swabs were collected and tested for enterovirus detection by real-time RT-PCR, while 1 651 cases were positive with the rate of 71.5%. Based on VP1 gene or 5'UTR region sequences, Coxsackievirus A2, A4, A6, A10, B2, B4 and echovirus 30 were detected in these cases. More importantly, Coxsackievirus A2 may be the major subtype of enterovirus resulting in children with herpangina in hangzhou, China.

  19. Analysis of enterovirus types in patients with symptoms of aseptic meningitis in 2014 in Shandong, China.

    Science.gov (United States)

    Chen, Peng; Lin, Xiaojuan; Liu, Guifang; Wang, Suting; Song, Lizhi; Tao, Zexin; Xu, Aiqiang

    2018-03-01

    We reviewed the epidemiological and clinical characteristics of 927 aseptic meningitis patients in Shandong in 2014, and the phylogeny of predominant enterovirus (EV) types causing this disease was analyzed. A total of 209 patients that were positive for EV were identified by both cell culture and a reverse transcription-seminested PCR in cerebrospinal fluid samples. The positive patients were most likely to be children within 15 years of age, had symptoms such as fever, vomiting and nausea (P< .05). The 209 EV sequences belonged to 11 types, and coxsackievirus B5, echovirus types 6 and 30 were predominant types. VP1 analysis exhibited multiple lineages were co-circulating. The significance of the study could come from the fact that surveillance is important to monitor the prevalence of EV types in population, which shows enterovirus meningitis maintains an important public health problem in China. Copyright © 2018 Elsevier Inc. All rights reserved.

  20. Enterovirus genotypes among patients with severe acute respiratory illness, influenza-like illness, and asymptomatic individuals in South Africa, 2012-2014.

    Science.gov (United States)

    Hellferscee, Orienka; Tempia, Stefano; Walaza, Sibongile; Variava, Ebrahim; Dawood, Halima; Wolter, Nicole; Madhi, Shabir A; du Plessis, Mignon; Cohen, Cheryl; Treurnicht, Florette K

    2017-10-01

    Enteroviruses can cause outbreaks of severe acute respiratory illness (SARI) and EV-A, -B, -C, and -D species have different pathogenic profiles and circulation patterns. We aimed to characterize and determine the prevalence of enterovirus genotypes among South African patients with respiratory illness and controls during June 2012 to July 2014. Syndromic SARI and influenza-like illness (ILI) surveillance was performed at two sentinel sites. At each site nasopharyngeal/oropharyngeal specimens were collected from SARI and ILI patients as well as controls. Specimens were tested for enterovirus by real-time PCR. Positive specimens were further genotyped by sequencing a region of the VP1 gene. The prevalence of enterovirus was 5.8% (87/1494), 3.4% (103/3079), and 3.4% (46/1367) among SARI, ILI, and controls, respectively (SARI/controls, P = 0.002 and ILI/control, P = 0.973). Among the 101/236 (42.8%) enterovirus-positive specimens that could be genotyped, we observed a high diversity of circulating enterovirus genotypes (a total of 33 genotypes) from all four human enterovirus species with high prevalence of Enterovirus-B (60.4%; 61/101) and Enterovirus-A (21.8%; 22/101) compared to Enterovirus-C (10.9%; 11/101) and Enterovirus-D (6.9%; 7/101) (P = 0.477). Of the enterovirus genotypes identified, Echovirus 30 (9.9%, 10/101), Coxsackie virus B5 (7.9%, 8/101) and Enterovirus-D68 (6.9%, 7/101) were most prevalent. There was no difference in disease severity (SARI or ILI compared to controls) between the different enterovirus species (P = 0.167). We observed a high number of enterovirus genotypes in patients with respiratory illness and in controls from South Africa with no disease association of EV species with disease severity. © 2017 Wiley Periodicals, Inc.

  1. Improvement of a real-time RT-PCR assay for the detection of enterovirus RNA

    Directory of Open Access Journals (Sweden)

    Bruynseels Peggy

    2009-07-01

    Full Text Available Abstract We describe an improvement of an earlier reported real-time RT-PCR assay for the detection of enterovirus RNA, based on the 5' exonuclease digestion of a dual-labeled fluorogenic probe by Taq DNA polymerase. A different extraction method, real-time RT-PCR instrument and primer set were evaluated. Our data show that the optimized assay yields a higher sensitivity and reproducibility and resulted in a significant reduced hands-on time per sample.

  2. New Introductions of Enterovirus 71 Subgenogroup C4 Strains, France, 2012

    Science.gov (United States)

    Henquell, Cécile; Mirand, Audrey; Coste-Burel, Marianne; Marque-Juillet, Stéphanie; Desbois, Delphine; Lagathu, Gisèle; Bornebusch, Laure; Bailly, Jean-Luc; Lina, Bruno

    2014-01-01

    In France during 2012, human enterovirus 71 (EV-A71) subgenogroup C4 strains were detected in 4 children hospitalized for neonatal fever or meningitis. Phylogenetic analysis showed novel and independent EV-A71 introductions, presumably from China, and suggested circulation of C4 strains throughout France. This observation emphasizes the need for monitoring EV-A71 infections in Europe. PMID:25061698

  3. Protective effect of an alpha 7 nicotinic acetylcholine receptor agonist against enterovirus 71 infection in neuronal cells.

    Science.gov (United States)

    Song, Feng Xia; Zhao, Lin Qing; Zhu, Ru Nan; Song, Qin Wei; Deng, Jie; Tian, Run; Wang, Fang; Qian, Yuan

    2018-01-01

    Enterovirus 71, as one of the dominant pathogens associated with severe hand, foot, and mouth disease, has been well reported to trigger severe neurological symptoms among young children over the last decade, particularly among children in the Asia-Pacific region. To date, no effective antiviral agent has been developed for the treatment of severe enterovirus 71 infection. PNU-282987, a selective alpha 7 nicotinic acetylcholine receptor (α7nAChR) agonist, has been reported to have a neuroprotective effect by participating in inflammatory regulation in previous studies. Therefore, in the present study, we aimed to assess the cell-protective effect of PNU-282987 against enterovirus 71 infection in neuronal cells, and to discuss potential mechanisms underlying this cell-protective effect in order to elucidate the potential impact of such agonists in the treatment of neurotropic viral infection. We observed that treatment with PNU-282987 improved cell viability and inhibited viral replication in enterovirus 71-infected SH-SY5Y cells. Further investigation revealed that inhibition of enterovirus 71 production by PNU-282987 is likely associated with events of RNA replication, and that increased levels of INF mRNA and its downstream antiviral proteins stimulated by the JAK-STAT2 pathway may contribute to the antiviral effect of PNU-282987. Moreover, our findings suggest that both the antiviral and anti-inflammatory effects of PNU-282987 may contribute to the neural protective effect of the drug in enterovirus 71-infected cells. Taken together, the results suggest that selective α7nAChR agonists may represent viable candidates for future therapeutic treatment of severe enterovirus 71 infection, and for other cases of neurotropic viral infection. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Molecular assays for targeting human and bovine enteric viruses in coastal waters and their application for library-independent source tracking

    Science.gov (United States)

    Fong, T.-T.; Griffin, Dale W.; Lipp, E.K.

    2005-01-01

    Rapid population growth and urban development along waterways and coastal areas have led to decreasing water quality. To examine the effects of upstream anthropogenic activities on microbiological water quality, methods for source-specific testing are required. In this study, molecular assays targeting human enteroviruses (HEV), bovine enteroviruses (BEV), and human adenoviruses (HAdV) were developed and used to identify major sources of fecal contamination in the lower Altamaha River, Georgia. Two-liter grab samples were collected monthly from five tidally influenced stations between July and December 2002. Samples were analyzed by reverse transcription- and nested-PCR. PCR results were confirmed by dot blot hybridization. Eleven and 17 of the 30 surface water samples tested positive for HAdV and HEV, respectively. Two-thirds of the samples tested positive for either HEV or HAdV, and the viruses occurred simultaneously in 26% of samples. BEV were detected in 11 of 30 surface water samples. Binary logistic regression analysis showed that the presence of both human and bovine enteric viruses was not significantly related to either fecal coliform or total coliform levels. The presence of these viruses was directly related to dissolved oxygen and streamflow but inversely related to water temperature, rainfall in the 30 days preceding sampling, and chlorophyll-?? concentrations. The stringent host specificity of enteric viruses makes them good library-independent indicators for identification of water pollution sources. Viral pathogen detection by PCR is a highly sensitive and easy-to-use tool for rapid assessment of water quality and fecal contamination when public health risk characterization is not necessary. Copyright ?? 2005, American Society for Microbiology. All Rights Reserved.

  5. Identification and production of mouse scFv to specific epitope of enterovirus-71 virion protein-2 (VP2).

    Science.gov (United States)

    Thanongsaksrikul, Jeeraphong; Srimanote, Potjanee; Tongtawe, Pongsri; Glab-Ampai, Kittirat; Malik, Aijaz Ahmad; Supasorn, Oratai; Chiawwit, Phatcharaporn; Poovorawan, Yong; Chaicumpa, Wanpen

    2018-05-01

    Enterovirus-71 (EV71) and coxsackievirus-A16 (CA16) frequently cause hand-foot-mouth disease (HFMD) epidemics among infants and young children. CA16 infections are usually mild, while EV71 disease may be fatal due to neurologic complications. As such, the ability to rapidly and specifically recognize EV71 is needed to facilitate proper case management and epidemic control. Accordingly, the aim of this study was to generate antibodies to EV71-virion protein-2 (VP2) by phage display technology for further use in specific detection of EV71. A recombinant peptide sequence of EV71-VP2, carrying a predicted conserved B cell epitope fused to glutathione-S-transferase (GST) (designated GST-EV71-VP2/131-160), was produced. The fusion protein was used as bait in in-solution biopanning to separate protein-bound phages from a murine scFv (MuscFv) phage display library constructed from an immunoglobulin gene repertoire from naïve ICR mice. Three phage-transformed E. coli clones (clones 63, 82, and 83) produced MuscFvs that bound to the GST-EV71-VP2/131-160 peptide. The MuscFv of clone 83 (MuscFv83), which produced the highest ELISA signal to the target antigen, was further tested. MuscFv83 also bound to full-length EV71-VP2 and EV71 particles, but did not bind to GST, full-length EV71-VP1, or the antigenically related CA16. MuscFv83 could be a suitable reagent for rapid antigen-based immunoassay, such as immunochromatography (ICT), for the specific detection and/or diagnosis of EV71 infection as well as epidemic surveillance.

  6. High sensitivity and label-free detection of Enterovirus 71 by nanogold modified electrochemical impedance spectroscopy

    Science.gov (United States)

    Wang, Fang-Yu; Li, Hsing-Yuan; Tseng, Shing-Hua; Cheng, Tsai-Mu; Chu, Hsueh-Liang; Yang, Jyh-Yuan; Chang, Chia-Ching

    2013-03-01

    Enterovirus 71 (EV71), which is the most fulminant and invasive species of enterovirus, can cause children neurologic complications and death within 2-3 days after fever and rash developed. Besides, EV71 has high sequence similarity with Coxsackie A 16 (CA16) that makes differential diagnosis difficult in clinic and laboratory. Since conventional viral diagnostic method cannot diagnose EV71 quickly and EV71 can transmit at low viral titer, the patients might delay in treatment. A quick, high sensitive, and high specific test for EV71 detection is pivotal. Electrochemical impedance spectroscopy (EIS) has been applied for detecting bio-molecules as biosensors recently. In this study, we try to build a detection platform for EV71 detection by nanogold modified EIS probe. The result shows that our probe can detect 3.6 VP1/50 μl (one EV71 particle has 60 VP1) in 3 minutes. The test can also distinguish EV71 from CA16 and lysozyme. Diagnosis of enterovirus 71 by electrochemical impedance spectroscopy has the potential to apply in clinic.

  7. Extracorporeal life support for critical enterovirus 71 rhombencephalomyelitis: long-term neurologic follow-up.

    Science.gov (United States)

    Lee, Hsiu-Fen; Chi, Ching-Shiang; Jan, Sheng-Ling; Fu, Yun-Ching; Huang, Fang-Liang; Chen, Po-Yen; Wang, Chung-Chi; Wei, Hao-Ji

    2012-04-01

    Enterovirus 71 rhombencephalomyelitis with cardiopulmonary dysfunction has become an endemic problem in Taiwan since an epidemic outbreak in 1998. Such cases frequently involve significant morbidity and mortality. From October 2000-June 2008, we collected 10 consecutive patients diagnosed with enterovirus 71 rhombencephalomyelitis complicated by left heart failure, with or without pulmonary edema, and surviving more than 3 months after receiving extracorporeal life support. Follow-up neurologic outcomes were analyzed prospectively. The median duration of neurologic follow-up was 7 years and 2 months. Significant morbidities included bulbar dysfunction, respiratory failure, and flaccid quadriparesis. Eight patients exhibited bulbar dysfunction, and feeding tubes could be removed from four patients (median, 15.5 months). Respiratory failure was observed in seven patients. Three patients were gradually withdrawn from their tracheostomy tube (median period, 30 months). Intelligence tests revealed four patients with normal cognitive function, one with borderline cognitive function, and one with mild mental retardation. Four were bedridden survivors. Extracorporeal life support for critical enterovirus 71 rhombencephalomyelitis demonstrated decreased neurologic sequelae during long-term follow-up, allowing for decannulation of feeding and tracheostomy tubes, and resulting in improved cognitive function. Copyright © 2012 Elsevier Inc. All rights reserved.

  8. Comparison of enterovirus detection in cerebrospinal fluid with Bacterial Meningitis Score in children

    Science.gov (United States)

    Pires, Frederico Ribeiro; Franco, Andréia Christine Bonotto Farias; Gilio, Alfredo Elias; Troster, Eduardo Juan

    2017-01-01

    ABSTRACT Objective To measure the role of enterovirus detection in cerebrospinal fluid compared with the Bacterial Meningitis Score in children with meningitis. Methods A retrospective cohort based on analysis of medical records of pediatric patients diagnosed as meningitis, seen at a private and tertiary hospital in São Paulo, Brazil, between 2011 and 2014. Excluded were patients with critical illness, purpura, ventricular shunt or recent neurosurgery, immunosuppression, concomitant bacterial infection requiring parenteral antibiotic therapy, and those who received antibiotics 72 hours before lumbar puncture. Results The study included 503 patients. Sixty-four patients were excluded and 94 were not submitted to all tests for analysis. Of the remaining 345 patients, 7 were in the Bacterial Meningitis Group and 338 in the Aseptic Meningitis Group. There was no statistical difference between the groups. In the Bacterial Meningitis Score analysis, of the 338 patients with possible aseptic meningitis (negative cultures), 121 of them had one or more points in the Bacterial Meningitis Score, with sensitivity of 100%, specificity of 64.2%, and negative predictive value of 100%. Of the 121 patients with positive Bacterial Meningitis Score, 71% (86 patients) had a positive enterovirus detection in cerebrospinal fluid. Conclusion Enterovirus detection in cerebrospinal fluid was effective to differentiate bacterial from viral meningitis. When the test was analyzed together with the Bacterial Meningitis Score, specificity was higher when compared to Bacterial Meningitis Score alone. PMID:28767914

  9. Indirect rapid prototyping of sol-gel hybrid glass scaffolds for bone regeneration - Effects of organic crosslinker valence, content and molecular weight on mechanical properties.

    Science.gov (United States)

    Hendrikx, Stephan; Kascholke, Christian; Flath, Tobias; Schumann, Dirk; Gressenbuch, Mathias; Schulze, F Peter; Hacker, Michael C; Schulz-Siegmund, Michaela

    2016-04-15

    We present a series of organic/inorganic hybrid sol-gel derived glasses, made from a tetraethoxysilane-derived silica sol (100% SiO2) and oligovalent organic crosslinkers functionalized with 3-isocyanatopropyltriethoxysilane. The material was susceptible to heat sterilization. The hybrids were processed into pore-interconnected scaffolds by an indirect rapid prototyping method, described here for the first time for sol-gel glass materials. A large panel of polyethylene oxide-derived 2- to 4-armed crosslinkers of molecular weights ranging between 170 and 8000Da were incorporated and their effect on scaffold mechanical properties was investigated. By multiple linear regression, 'organic content' and the 'content of ethylene oxide units in the hybrid' were identified as the main factors that determined compressive strength and modulus, respectively. In general, 3- and 4-armed crosslinkers performed better than linear molecules. Compression tests and cell culture experiments with osteoblast-like SaOS-2 cells showed that macroporous scaffolds can be produced with compressive strengths of up to 33±2MPa and with a pore structure that allows cells to grow deep into the scaffolds and form mineral deposits. Compressive moduli between 27±7MPa and 568±98MPa were obtained depending on the hybrid composition and problems associated with the inherent brittleness of sol-gel glass materials could be overcome. SaOS-2 cells showed cytocompatibility on hybrid glass scaffolds and mineral accumulation started as early as day 7. On day 14, we also found mineral accumulation on control hybrid glass scaffolds without cells, indicating a positive effect of the hybrid glass on mineral accumulation. We produced a hybrid sol-gel glass material with significantly improved mechanical properties towards an application in bone regeneration and processed the material into macroporous scaffolds of controlled architecture by indirect rapid prototyping. We were able to produce macroporous materials

  10. Molecular evolution of the Bovini tribe (Bovidae, Bovinae: Is there evidence of rapid evolution or reduced selective constraint in Domestic cattle?

    Directory of Open Access Journals (Sweden)

    McCulloch Alan

    2009-04-01

    Full Text Available Abstract Background If mutation within the coding region of the genome is largely not adaptive, the ratio of nonsynonymous (dN to synonymous substitutions (dS per site (dN/dS should be approximately equal among closely related species. Furthermore, dN/dS in divergence between species should be equivalent to dN/dS in polymorphisms. This hypothesis is of particular interest in closely related members of the Bovini tribe, because domestication has promoted rapid phenotypic divergence through strong artificial selection of some species while others remain undomesticated. We examined a number of genes that may be involved in milk production in Domestic cattle and a number of their wild relatives for evidence that domestication had affected molecular evolution. Elevated rates of dN/dS were further queried to determine if they were the result of positive selection, low effective population size (Ne or reduced selective constraint. Results We have found that the domestication process has contributed to higher dN/dS ratios in cattle, especially in the lineages leading to the Domestic cow (Bos taurus and Mithan (Bos frontalis and within some breeds of Domestic cow. However, the high rates of dN/dS polymorphism within B. taurus when compared to species divergence suggest that positive selection has not elevated evolutionary rates in these genes. Likewise, the low rate of dN/dS in Bison, which has undergone a recent population bottleneck, indicates a reduction in population size alone is not responsible for these observations. Conclusion The effect of selection depends on effective population size and the selection coefficient (Nes. Typically under domestication both selection pressure for traits important in fitness in the wild and Ne are reduced. Therefore, reduced selective constraint could be responsible for the observed elevated evolutionary ratios in domesticated species, especially in B. taurus and B. frontalis, which have the highest dN/dS in the

  11. Sporadic isolation of sabin-like polioviruses and high-level detection of non-polio enteroviruses during sewage surveillance in seven Italian cities, after several years of inactivated poliovirus vaccination.

    Science.gov (United States)

    Battistone, A; Buttinelli, G; Fiore, S; Amato, C; Bonomo, P; Patti, A M; Vulcano, A; Barbi, M; Binda, S; Pellegrinelli, L; Tanzi, M L; Affanni, P; Castiglia, P; Germinario, C; Mercurio, P; Cicala, A; Triassi, M; Pennino, F; Fiore, L

    2014-08-01

    Sewage surveillance in seven Italian cities between 2005 and 2008, after the introduction of inactivated poliovirus vaccination (IPV) in 2002, showed rare polioviruses, none that were wild-type or circulating vaccine-derived poliovirus (cVDPV), and many other enteroviruses among 1,392 samples analyzed. Two of five polioviruses (PV) detected were Sabin-like PV2 and three PV3, based on enzyme-linked immunosorbent assay (ELISA) and PCR results. Neurovirulence-related mutations were found in the 5'noncoding region (5'NCR) of all strains and, for a PV2, also in VP1 region 143 (Ile>Thr). Intertypic recombination in the 3D region was detected in a second PV2 (Sabin 2/Sabin 1) and a PV3 (Sabin 3/Sabin 2). The low mutation rate in VP1 for all PVs suggests limited interhuman virus passages, consistent with efficient polio immunization in Italy. Nonetheless, these findings highlight the risk of wild or Sabin poliovirus reintroduction from abroad. Non-polio enteroviruses (NPEVs) were detected, 448 of which were coxsackievirus B (CVB) and 294 of which were echoviruses (Echo). Fifty-six NPEVs failing serological typing were characterized by sequencing the VP1 region (nucleotides [nt] 2628 to 2976). A total of 448 CVB and 294 Echo strains were identified; among those strains, CVB2, CVB5, and Echo 11 predominated. Environmental CVB5 and CVB2 strains from this study showed high sequence identity with GenBank global strains. The high similarity between environmental NPEVs and clinical strains from the same areas of Italy and the same periods indicates that environmental strains reflect the viruses circulating in the population and highlights the potential risk of inefficient wastewater treatments. This study confirmed that sewage surveillance can be more sensitive than acute flaccid paralysis (AFP) surveillance in monitoring silent poliovirus circulation in the population as well as the suitability of molecular approaches to enterovirus typing.

  12. EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. Part III. Virus Detection by RT-qPCR

    Science.gov (United States)

    EPA Method 1615 measures enteroviruses and noroviruses present in environmental and drinking waters. The viral ribonucleic acid (RNA) from water sample concentrates is extracted and tested for enterovirus and norovirus RNA using reverse transcription-quantitative PCR (RT-qPCR). V...

  13. Prevalence and characterization of enterovirus infections among pediatric patients with hand foot mouth disease, herpangina and influenza like illness in Thailand, 2012.

    Science.gov (United States)

    Puenpa, Jiratchaya; Mauleekoonphairoj, John; Linsuwanon, Piyada; Suwannakarn, Kamol; Chieochansin, Thaweesak; Korkong, Sumeth; Theamboonlers, Apiradee; Poovorawan, Yong

    2014-01-01

    Hand, foot, and mouth disease (HFMD) and herpangina are common infectious diseases caused by several genotypes of human enterovirus species A and frequently occurring in young children. This study was aimed at analyzing enteroviruses from patients with these diseases in Thailand in 2012. Detection and genotype determination of enteroviruses were accomplished by reverse transcription-polymerase chain reaction and sequencing of the VP1 region. Enterovirus-positive samples were differentiated into 17 genotypes (coxsackievirus A4 (CAV4), A5, A6, A8, A9, A10, A12, A16, A21, B1, B2, B4, B5, echovirus 7, 16, 25 and Enterovirus 71). The result showed CAV6 (33.5%), followed by CAV16 (9.4%) and EV71 (8.8%) as the most frequent genotypes in HFMD, CAV8 (19.3%) in herpangina and CAV6 (1.5%) in influenza like illness. Enterovirus infections were most prevalent during July with 34.4% in HFMD, 39.8% in herpangina and 1.6% in ILI. The higher enterovirus infection associated with HFMD and herpangina occurred in infants over one year-old. This represents the first report describing the circulation of multiple enteroviruses in Thailand.

  14. Prevalence and Characterization of Enterovirus Infections among Pediatric Patients with Hand Foot Mouth Disease, Herpangina and Influenza Like Illness in Thailand, 2012

    Science.gov (United States)

    Puenpa, Jiratchaya; Mauleekoonphairoj, John; Linsuwanon, Piyada; Suwannakarn, Kamol; Chieochansin, Thaweesak; Korkong, Sumeth; Theamboonlers, Apiradee; Poovorawan, Yong

    2014-01-01

    Hand, foot, and mouth disease (HFMD) and herpangina are common infectious diseases caused by several genotypes of human enterovirus species A and frequently occurring in young children. This study was aimed at analyzing enteroviruses from patients with these diseases in Thailand in 2012. Detection and genotype determination of enteroviruses were accomplished by reverse transcription-polymerase chain reaction and sequencing of the VP1 region. Enterovirus-positive samples were differentiated into 17 genotypes (coxsackievirus A4 (CAV4), A5, A6, A8, A9, A10, A12, A16, A21, B1, B2, B4, B5, echovirus 7, 16, 25 and Enterovirus 71). The result showed CAV6 (33.5%), followed by CAV16 (9.4%) and EV71 (8.8%) as the most frequent genotypes in HFMD, CAV8 (19.3%) in herpangina and CAV6 (1.5%) in influenza like illness. Enterovirus infections were most prevalent during July with 34.4% in HFMD, 39.8% in herpangina and 1.6% in ILI. The higher enterovirus infection associated with HFMD and herpangina occurred in infants over one year-old. This represents the first report describing the circulation of multiple enteroviruses in Thailand. PMID:24887237

  15. Enterovirus RNA in Peripheral Blood May Be Associated with the Variants of rs1990760, a Common Type 1 Diabetes Associated Polymorphism in IFIH1

    Science.gov (United States)

    Cinek, Ondrej; Tapia, German; Witsø, Elisabet; Kramna, Lenka; Holkova, Katerina; Rasmussen, Trond; Stene, Lars C.; Rønningen, Kjersti S.

    2012-01-01

    Objective Polymorphisms in the IFIH1 (common rs1990760 and four rare rs35667974, rs35337543, rs35744605, rs35732034) have been convincingly associated with type 1 diabetes. The encoded protein (interferon-induced helicase C domain-containing protein 1) senses double-stranded RNA during replication of Picornavirales, including Enterovirus, a genus suspected in the etiology of type 1 diabetes. We therefore investigated whether the polymorphisms are associated with differences in the frequency of enterovirus RNA in blood. Research Design and Methods The study included 1001 blood samples, each from a child participating in the Norwegian ‘Environmental Triggers of Type 1 Diabetes: the MIDIA study’. The enterovirus RNA was tested using qualitative semi-nested real-time reverse transcriptase PCR on RNA extracted from frozen cell packs after removal of plasma. Stool samples previously analyzed for enterovirus RNA were available in 417 children. Results The genotypes of IFIH1 rs1990760 were associated with different frequencies of enterovirus RNA in blood (7.0%, 14.4% and 9.5% bloods were enterovirus positive among children carrying the Ala/Ala, Ala/Thr and Thr/Thr genotypes, respectively, p = 0.012). This association remained essentially unchanged after adjustment for age and calendar year. The presence of enterovirus in the concomitantly sampled stool further increased the likelihood of enterovirus RNA in blood (odds ratio 2.40, CI 95% 1.13–4.70), but did not affect the association with IFIH1 rs1990760. The rare polymorphisms (individually, or pooled) were not significantly associated with enterovirus RNA in blood. Conclusions The common IFIH1 SNP may modify the frequency of enterovirus RNA in blood of healthy children. This effect can help explain the association of IFIH1 with type 1 diabetes. PMID:23144876

  16. Acute Flaccid Paralysis by Enterovirus D68 Infection: First Italian Description in Adult Patient and Role of Electrophysiology

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    Marco Ceccanti

    2017-11-01

    Full Text Available A Peruvian woman was admitted to the Emergency Department, due to an acute flaccid paralysis (AFP of the upper limbs that progressively involved also lower limbs and respiratory muscles. She previously suffered from non-Hodgkin’s lymphoma and had to undergo hematopoietic stem cell transplantation. A magnetic resonance imaging showed a T2 hyperintensity in the anterior and central region of the cervical segment with an elective involvement of gray matter. This finding, combined with other clinical, laboratory, and electrophysiological data, led to a diagnosis of AFP. Enterovirus D68 was isolated in the patient’s cerebrospinal fluid, plasma, and throat swab. To our knowledge, this is the first Italian case of AFP by Enterovirus D68 infection in an adult. The diagnostic assessment and management of AFP by Enterovirus D68 are discussed.

  17. The structure-function relationship of the enterovirus 3'-UTR.

    NARCIS (Netherlands)

    Zoll, J.; Heus, H.A.; Kuppeveld, F.J.M. van; Melchers, W.J.G.

    2009-01-01

    Essential processes in living cells are carried out by large complex assemblies, which typically consist of a large number of proteins and frequently also contain nucleic acids, mostly RNA [Alberts, B., 1998. The cell as a collection of protein machines: preparing the next generation of molecular

  18. Pathways to multidrug-resistant tuberculosis diagnosis and treatment initiation: a qualitative comparison of patients' experiences in the era of rapid molecular diagnostic tests.

    Science.gov (United States)

    Naidoo, Pren; van Niekerk, Margaret; du Toit, Elizabeth; Beyers, Nulda; Leon, Natalie

    2015-10-28

    Although new molecular diagnostic tests such as GenoType MTBDRplus and Xpert® MTB/RIF have reduced multidrug-resistant tuberculosis (MDR-TB) treatment initiation times, patients' experiences of diagnosis and treatment initiation are not known. This study aimed to explore and compare MDR-TB patients' experiences of their diagnostic and treatment initiation pathway in GenoType MTBDRplus and Xpert® MTB/RIF-based diagnostic algorithms. The study was undertaken in Cape Town, South Africa where primary health-care services provided free TB diagnosis and treatment. A smear, culture and GenoType MTBDRplus diagnostic algorithm was used in 2010, with Xpert® MTB/RIF phased in from 2011-2013. Participants diagnosed in each algorithm at four facilities were purposively sampled, stratifying by age, gender and MDR-TB risk profiles. We conducted in-depth qualitative interviews using a semi-structured interview guide. Through constant comparative analysis we induced common and divergent themes related to symptom recognition, health-care access, testing for MDR-TB and treatment initiation within and between groups. Data were triangulated with clinical information and health visit data from a structured questionnaire. We identified both enablers and barriers to early MDR-TB diagnosis and treatment. Half the patients had previously been treated for TB; most recognised recurring symptoms and reported early health-seeking. Those who attributed symptoms to other causes delayed health-seeking. Perceptions of poor public sector services were prevalent and may have contributed both to deferred health-seeking and to patient's use of the private sector, contributing to delays. However, once on treatment, most patients expressed satisfaction with public sector care. Two patients in the Xpert® MTB/RIF-based algorithm exemplified its potential to reduce delays, commencing MDR-TB treatment within a week of their first health contact. However, most patients in both algorithms experienced

  19. Implementation of 3D spatial indexing and compression in a large-scale molecular dynamics simulation database for rapid atomic contact detection.

    Science.gov (United States)

    Toofanny, Rudesh D; Simms, Andrew M; Beck, David A C; Daggett, Valerie

    2011-08-10

    Molecular dynamics (MD) simulations offer the ability to observe the dynamics and interactions of both whole macromolecules and individual atoms as a function of time. Taken in context with experimental data, atomic interactions from simulation provide insight into the mechanics of protein folding, dynamics, and function. The calculation of atomic interactions or contacts from an MD trajectory is computationally demanding and the work required grows exponentially with the size of the simulation system. We describe the implementation of a spatial indexing algorithm in our multi-terabyte MD simulation database that significantly reduces the run-time required for discovery of contacts. The approach is applied to the Dynameomics project data. Spatial indexing, also known as spatial hashing, is a method that divides the simulation space into regular sized bins and attributes an index to each bin. Since, the calculation of contacts is widely employed in the simulation field, we also use this as the basis for testing compression of data tables. We investigate the effects of compression of the trajectory coordinate tables with different options of data and index compression within MS SQL SERVER 2008. Our implementation of spatial indexing speeds up the calculation of contacts over a 1 nanosecond (ns) simulation window by between 14% and 90% (i.e., 1.2 and 10.3 times faster). For a 'full' simulation trajectory (51 ns) spatial indexing reduces the calculation run-time between 31 and 81% (between 1.4 and 5.3 times faster). Compression resulted in reduced table sizes but resulted in no significant difference in the total execution time for neighbour discovery. The greatest compression (~36%) was achieved using page level compression on both the data and indexes. The spatial indexing scheme significantly decreases the time taken to calculate atomic contacts and could be applied to other multidimensional neighbor discovery problems. The speed up enables on-the-fly calculation

  20. Implementation of 3D spatial indexing and compression in a large-scale molecular dynamics simulation database for rapid atomic contact detection

    Directory of Open Access Journals (Sweden)

    Toofanny Rudesh D

    2011-08-01

    Full Text Available Abstract Background Molecular dynamics (MD simulations offer the ability to observe the dynamics and interactions of both whole macromolecules and individual atoms as a function of time. Taken in context with experimental data, atomic interactions from simulation provide insight into the mechanics of protein folding, dynamics, and function. The calculation of atomic interactions or contacts from an MD trajectory is computationally demanding and the work required grows exponentially with the size of the simulation system. We describe the implementation of a spatial indexing algorithm in our multi-terabyte MD simulation database that significantly reduces the run-time required for discovery of contacts. The approach is applied to the Dynameomics project data. Spatial indexing, also known as spatial hashing, is a method that divides the simulation space into regular sized bins and attributes an index to each bin. Since, the calculation of contacts is widely employed in the simulation field, we also use this as the basis for testing compression of data tables. We investigate the effects of compression of the trajectory coordinate tables with different options of data and index compression within MS SQL SERVER 2008. Results Our implementation of spatial indexing speeds up the calculation of contacts over a 1 nanosecond (ns simulation window by between 14% and 90% (i.e., 1.2 and 10.3 times faster. For a 'full' simulation trajectory (51 ns spatial indexing reduces the calculation run-time between 31 and 81% (between 1.4 and 5.3 times faster. Compression resulted in reduced table sizes but resulted in no significant difference in the total execution time for neighbour discovery. The greatest compression (~36% was achieved using page level compression on both the data and indexes. Conclusions The spatial indexing scheme significantly decreases the time taken to calculate atomic contacts and could be applied to other multidimensional neighbor discovery

  1. Genetic Determinants of Enterovirus Infections: Polymorphisms in Type 1 Diabetes and Innate Immune Genes in the MIDIA Study.

    Science.gov (United States)

    Witsø, Elisabet; Cinek, Ondrej; Tapia, German; Brorsson, Caroline A; Stene, Lars C; Gjessing, Håkon K; Rasmussen, Trond; Bergholdt, Regine; Pociot, Flemming M; Rønningen, Kjersti S

    2015-12-01

    Enteroviruses have been suggested as triggers of type 1 diabetes (T1D). We aimed to assess whether established T1D susceptibility single nucleotide polymorphisms (SNPs) and candidate SNPs in innate immune genes were associated with the frequency of enterovirus infection in otherwise healthy children. Fifty-six established T1D SNPs and 97 other candidate immunity SNPs were typed in 419 children carrying the T1D high-risk genotype, HLA-DR4-DQ8/DR3-DQ2 genotype, and 373 children without this genotype. Enteroviral RNA was detected using real-time polymerase chain reaction, with primers detecting essentially all enterovirus serotypes, in 7,393 longitudinal stool samples collected monthly (age range 3-36 months). The most significant association was with two T1D SNPs, rs12150079 (ZPBP2/ORMDL3/GSDMB region) (enterovirus frequency: AA 7.3%, AG 8.7%, GG 9.7%, RR = 0.86, overall p = 1.87E-02) and rs229541 (C1QTNF6/SSTR3/RAC2) (enterovirus frequency: CC 7.8%, CT 9.7%, TT 9.4%, RR = 1.13, overall p = 3.6E-02), followed by TLR8 (rs2407992) (p = 3.8E-02), TLR3 (1914926) (p = 4.9E-02), and two other T1D SNPs (IFIH1 rs3747517, p = 4.9E-02 and PTPN22, rs2476601, p = 5.3E-02). However, the quantile-quantile plot of p-values with confidence intervals for all 153 SNPs did not reveal clear evidence for rejection of the complete null hypothesis. Among a number of SNPs in candidate genes, we found no evidence for strong associations with enterovirus presence in stool samples from Norwegian children.

  2. Human Enterovirus Nonstructural Protein 2CATPase Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

    Science.gov (United States)

    Xia, Hongjie; Wang, Peipei; Wang, Guang-Chuan; Yang, Jie; Sun, Xianlin; Wu, Wenzhe; Qiu, Yang; Shu, Ting; Zhao, Xiaolu; Yin, Lei; Qin, Cheng-Feng; Hu, Yuanyang; Zhou, Xi

    2015-01-01

    RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2CATPase of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3′-to-5′ unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2CATPase middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2CATPase facilitated EV71 RNA synthesis in vitro; when 2CATPase helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2CATPase-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2CATPase are also conserved in coxsackie A virus 16 (CAV16), another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2CATPase, and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings increase our

  3. 2BC Non-Structural Protein of Enterovirus A71 Interacts with SNARE Proteins to Trigger Autolysosome Formation.

    Science.gov (United States)

    Lai, Jeffrey K F; Sam, I-Ching; Verlhac, Pauline; Baguet, Joël; Eskelinen, Eeva-Liisa; Faure, Mathias; Chan, Yoke Fun

    2017-07-04

    Viruses have evolved unique strategies to evade or subvert autophagy machinery. Enterovirus A71 (EV-A71) induces autophagy during infection in vitro and in vivo. In this study, we report that EV-A71 triggers autolysosome formation during infection in human rhabdomyosarcoma (RD) cells to facilitate its replication. Blocking autophagosome-lysosome fusion with chloroquine inhibited virus RNA replication, resulting in lower viral titres, viral RNA copies and viral proteins. Overexpression of the non-structural protein 2BC of EV-A71 induced autolysosome formation. Yeast 2-hybrid and co-affinity purification assays showed that 2BC physically and specifically interacted with a N -ethylmaleimide-sensitive factor attachment receptor (SNARE) protein, syntaxin-17 (STX17). Co-immunoprecipitation assay further showed that 2BC binds to SNARE proteins, STX17 and synaptosome associated protein 29 (SNAP29). Transient knockdown of STX17, SNAP29, and microtubule-associated protein 1 light chain 3B (LC3B), crucial proteins in the fusion between autophagosomes and lysosomes) as well as the lysosomal-associated membrane protein 1 (LAMP1) impaired production of infectious EV-A71 in RD cells. Collectively, these results demonstrate that the generation of autolysosomes triggered by the 2BC non-structural protein is important for EV-A71 replication, revealing a potential molecular pathway targeted by the virus to exploit autophagy. This study opens the possibility for the development of novel antivirals that specifically target 2BC to inhibit formation of autolysosomes during EV-A71 infection.

  4. Antiviral Potential of a Novel Compound CW-33 against Enterovirus A71 via Inhibition of Viral 2A Protease

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    Ching-Ying Wang

    2015-06-01

    Full Text Available Enterovirus A71 (EV-A71 in the Picornaviridae family causes hand-foot-and-mouth disease, aseptic meningitis, severe central nervous system disease, even death. EV-A71 2A protease cleaves Type I interferon (IFN-α/β receptor 1 (IFNAR1 to block IFN-induced Jak/STAT signaling. This study investigated anti-EV-A7l activity and synergistic mechanism(s of a novel furoquinoline alkaloid compound CW-33 alone and in combination with IFN-β Anti-EV-A71 activities of CW-33 alone and in combination with IFN-β were evaluated by inhibitory assays of virus-induced apoptosis, plaque formation, and virus yield. CW-33 showed antiviral activities with an IC50 of near 200 µM in EV-A71 plaque reduction and virus yield inhibition assays. While, anti-EV-A71 activities of CW-33 combined with 100 U/mL IFN-β exhibited a synergistic potency with an IC50 of approximate 1 µM in plaque reduction and virus yield inhibition assays. Molecular docking revealed CW-33 binding to EV-A71 2A protease active sites, correlating with an inhibitory effect of CW33 on in vitro enzymatic activity of recombinant 2A protease IC50 = 53.1 µM. Western blotting demonstrated CW-33 specifically inhibiting 2A protease-mediated cleavage of IFNAR1. CW-33 also recovered Type I IFN-induced Tyk2 and STAT1 phosphorylation as well as 2',5'-OAS upregulation in EV-A71 infected cells. The results demonstrated CW-33 inhibiting viral 2A protease activity to reduce Type I IFN antagonism of EV-A71. Therefore, CW-33 combined with a low-dose of Type I IFN could be applied in developing alternative approaches to treat EV-A71 infection.

  5. Synthesis of Pyrazine-1,3-thiazine Hybrid Analogues as Antiviral Agent Against HIV-1, Influenza A (H1N1), Enterovirus 71 (EV71), and Coxsackievirus B3 (CVB3).

    Science.gov (United States)

    Wu, Hong-Min; Zhou, Kuo; Wu, Tao; Cao, Yin-Guang

    2016-09-01

    A novel series of pyrazine-1,3-thiazine hybrid conjugates were synthesized in excellent yield. These derivatives were subsequently tested against human immunodeficiency virus (HIV-1); hemagglutinin type 1 and neuraminidase type 1-'influenza' A (H1N1) virus; enterovirus 71 (EV71); and coxsackievirus B3. The effect of these conjugates on the key enzymes responsible for the progression of these viral infections was also illustrated via enzyme-based assay, such as HIV-1 reverse transcriptase (RT) and neuraminidase, where entire tested molecules showed considerable inhibition. Particularly, among the tested derivatives, compound 3k was identified as most promising inhibitor of HIV-1 with 94% of inhibition (IC50 3.26 ± 0.2 μm). Moreover, the compound 3d was found to be the most potent analogue to inhibit the H1N1 virus with IC50 of 5.32 ± 0.4 μm together with inhibition of the neuraminidase enzyme (IC50 11.24 ± 1.1 μm). In regard to inhibitory activity against enterovirus 71 (EV71) and coxsackievirus B3 (CVB3), the tested derivatives showed considerable inhibition of infection. Molecular docking studies were also performed for the most promising inhibitors with their corresponding target protein to exemplify the structural requirement for better inhibitory activity. The results of inhibitory assay showed that designed molecules possess considerable inhibitory activity against the virus tested. © 2016 John Wiley & Sons A/S.

  6. Polymorphisms in the Innate Immune IFIH1 Gene, Frequency of Enterovirus in Monthly Fecal Samples during Infancy, and Islet Autoimmunity

    Science.gov (United States)

    Witsø, Elisabet; Tapia, German; Cinek, Ondrej; Pociot, Flemming Michael; Stene, Lars C.; Rønningen, Kjersti S.

    2011-01-01

    Interferon induced with helicase C domain 1 (IFIH1) senses and initiates antiviral activity against enteroviruses. Genetic variants of IFIH1, one common and four rare SNPs have been associated with lower risk for type 1 diabetes. Our aim was to test whether these type 1 diabetes-associated IFIH1 polymorphisms are associated with the occurrence of enterovirus infection in the gut of healthy children, or influence the lack of association between gut enterovirus infection and islet autoimmunity. After testing of 46,939 Norwegian newborns, 421 children carrying the high risk genotype for type 1 diabetes (HLA-DR4-DQ8/DR3-DQ2) as well as 375 children without this genotype were included for monthly fecal collections from 3 to 35 months of age, and genotyped for the IFIH1 polymorphisms. A total of 7,793 fecal samples were tested for presence of enterovirus RNA using real time reverse transcriptase PCR. We found no association with frequency of enterovirus in the gut for the common IFIH1 polymorphism rs1990760, or either of the rare variants of rs35744605, rs35667974, rs35337543, while the enterovirus prevalence marginally differed in samples from the 8 carriers of a rare allele of rs35732034 (26.1%, 18/69 samples) as compared to wild-type homozygotes (12.4%, 955/7724 samples); odds ratio 2.5, p = 0.06. The association was stronger when infections were restricted to those with high viral loads (odds ratio 3.3, 95% CI 1.3–8.4, p = 0.01). The lack of association between enterovirus frequency and islet autoimmunity reported in our previous study was not materially influenced by the IFIH1 SNPs. We conclude that the type 1 diabetes-associated IFIH1 polymorphisms have no, or only minor influence on the occurrence, quantity or duration of enterovirus infection in the gut. Its effect on the risk of diabetes is likely to lie elsewhere in the pathogenic process than in the modification of gut infection. PMID:22110759

  7. Molecular Diagnostics

    OpenAIRE

    Choe, Hyonmin; Deirmengian, Carl A.; Hickok, Noreen J.; Morrison, Tiffany N.; Tuan, Rocky S.

    2015-01-01

    Orthopaedic infections are complex conditions that require immediate diagnosis and accurate identification of the causative organisms to facilitate appropriate management. Conventional methodologies for diagnosis of these infections sometimes lack accuracy or sufficient rapidity. Current molecular diagnostics are an emerging area of bench-to-bedside research in orthopaedic infections. Examples of promising molecular diagnostics include measurement of a specific biomarker in the synovial fluid...

  8. Disease burden of enterovirus infection in Taiwan: Implications for vaccination policy.

    Science.gov (United States)

    Liu, Ding-Ping; Wang, Ting-Ann; Huang, Wan-Ting; Chang, Luan-Yin; Wang, En-Tzu; Cheng, Shou-Hsia; Yang, Ming-Chin

    2016-02-10

    This study aimed to assess the disease burden and economic impacts of human nonpolio enteroviruses (NPEV) and enterovirus A71 (EV-A71) infection in Taiwan. We included children under five years old (n=983,127-1,118,649) with ICD-9-CM codes 0740 (herpangina) or 0743 (hand-foot-and-mouth disease) from the 2006 to 2010 National Health Insurance Database. Severity of enterovirus infection was assessed from outpatient/emergency visits, hospitalization (with/without intensive care unit [ICU] admission), infection with severe complications, and death. We estimated medical costs and indirect costs from the societal perspective. The annual rates of NPEV events for children under five years old ranged from 13.9% to 38.4%, of which 5.1-8.8% were hospitalized. EV-A71 accounted for 7.8% of all NPEV medical costs, but 79.1% of NPEV ICU costs. Travel costs and productivity loss of caregivers were $37.1 (range: $24.5-$64.7) million per year. These costs were not higher in the EV-A71 dominant year ($34.4 million) compared with those in the other years. Productivity losses resulting from premature mortality by NPEV infection were $0.8 (range: $0.0-$2.9) million per year, of which 96.3% were caused by EV-A71. Diseases associated with NPEV other than EV-A71 were responsible for most of the medical expenses. In addition, caregiver productivity loss by high rates of NPEV infection impacted the society much more than medical costs. A multi-valent vaccine that includes EV-A71 and other serotypes, for example coxsackievirus A16, may be beneficial to the health of children in Taiwan. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Clinico-radiological spectrum in enterovirus 71 infection involving the central nervous system in children.

    Science.gov (United States)

    Lee, Kyung Yeon; Lee, Yun-Jin; Kim, Tae Hyoung; Cheon, Doo-Sung; Nam, Sang-Ook

    2014-03-01

    Enterovirus 71 infection causes hand, foot and mouth disease in children, and can produce diverse neurologic complications. Epidemics occurring in Korea between 2009 and 2012 resulted in the death of some patients. The present study aimed to clarify the correlation between clinical features and MRI findings in patients presenting with acute neurologic manifestations related to enterovirus 71 infection. Based on their clinical features, the patients were classified into four clinical groups: (1) brainstem encephalitis (n=17), characterized by myoclonus, tremor, ataxia, and autonomic dysregulation such as pulmonary hemorrhage; (2) aseptic meningitis (n=2); (3) encephalitis (n=2), characterized by decreased consciousness, seizure, and fever without myoclonus, tremor, ataxia, and autonomic dysregulation; and (4) acute flaccid paralysis (n=1). Thirteen of the 17 patients with brainstem encephalitis showed characteristic lesions in the dorsal brainstem and bilateral cerebellar dentate nuclei on brain MRI, whereas three had no abnormality. One of the two patients with meningitis had a small lesion in the left dorsal pons. Two patients with encephalitis had no apparent MRI abnormality. One patient with acute flaccid paralysis of the right leg had contrast-enhancement of the bilateral ventral nerve roots at the lumbar spine level on MRI. Five of 13 patients with lesions in the bilateral dentate nuclei of the cerebellum exhibited no cerebellar symptoms, while two with no cerebellar lesions developed ataxia. Although most patients presenting with neurologic manifestations of enterovirus 71 infection had characteristic clinical features together with typical MRI findings, the clinical features were not necessarily consistent with MRI findings. Copyright © 2013 Elsevier Ltd. All rights reserved.

  10. A cluster of Enterovirus 71 subgenogroup C2 in a nursery school, Italy, 2014.

    Science.gov (United States)

    Medici, Maria Cristina; Tummolo, Fabio; Arcangeletti, Maria Cristina; De Conto, Flora; Chezzi, Carlo; Dodi, Icilio; Calderaro, Adriana

    2016-10-01

    During October 2014, enterovirus (EV) RNA was detected in the stools of four children attending the same class in a nursery school, and hospitalized with mild febrile and vomiting disease in Parma, Italy. Upon sequencing, the viruses were characterized as EV71 subgenogroup C2. Phylogenetic analysis of the four EV71 C2 viruses allowed the distinction of a diverging lineage within subgenogroup C2, containing the Italian EV71 C2 strains and viruses detected in France in 2013. The identification of an outbreak of EV71 C2 in Italy extended information on the geographic diffusion and clinical relevance of these viruses in Europe.

  11. Novel Substituted Heteroaromatic Piperazine and Piperidine Derivatives as Inhibitors of Human Enterovirus 71 and Coxsackievirus A16

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    Xian Zhang

    2013-04-01

    Full Text Available A series of substituted heteroaromatic piperazine and piperidine derivatives were found through virtual screening based on the structure of human enterovirus 71 capsid protein VP1. The preliminary biological evaluation revealed that compounds 8e and 9e have potent activity against EV71 and Coxsackievirus A16 with low cytotoxicity.

  12. PREVALENCE OF HUMAN ENTEROVIRUS AMONG PATIENTS WITH HAND, FOOT, AND MOUTH DISEASE AND HERPANGINA IN THAILAND, 2013.

    Science.gov (United States)

    Mauleekoonphairoj, John; Puenpa, Jiratchaya; Korkong, Sumeth; Vongpunsawad, Sompong; Poovorawan, Yong

    2015-11-01

    Human enterovirus (EV) infection causes hand, foot, and mouth disease (HFMD) and herpangina (HA). We studied the prevalence of enterovirus (EV) among patients with HFMD and HA in Thailand during 2013. We conducted a study in archived specimens of patients sent for screening for enterovirus. A total of 203 clinical specimens from 184 individuals with painful blister in the oropharynx and on the palms, soles, knees, elbows or buttock were examined by semi-nested polymerase chain reaction (PCR) for the 5'UTR and VP1 genes of EV. Eighty-six samples were positive: EV71 was detected in 14 (30%), CV-A8 in 12 (26%) and CV-A16 in 10 (21%). Classification of EV species detected revealed that 46 specimens were EV-A, 14 specimens were EV-B, 1 specimen was EV-D, and 16 specimens were positive for unclassified enterovirus. The majority of individuals with EV infection were aged 2-6 years. Multiple EV-A serotypes were detected among HFMD and HA patients in our study.

  13. Mutations in the Nonstructural Protein 3A Confer Resistance to the Novel Enterovirus Replication Inhibitor TTP-8307

    NARCIS (Netherlands)

    de Palma, Armando M.; Thibaut, Hendrik Jan; van der Linden, Lonneke; Lanke, Kjerstin; Heggermont, Ward; Ireland, Stephen; Andrews, Robert; Arimilli, Murty; Al-tel, Taleb H.; de Clercq, Erik; van Kuppeveld, Frank; Neyts, Johan

    2009-01-01

    A novel compound, TTP-8307, was identified as a potent inhibitor of the replication of several rhino- and enteroviruses. TTP-8307 inhibits viral RNA synthesis in a dose-dependent manner, without affecting polyprotein synthesis and/or processing. Drug-resistant variants of coxsackievirus B3 were all

  14. Prospective assessment of clinical symptoms associated with enterovirus and parechovirus genotypes in a multicenter study in Dutch children

    NARCIS (Netherlands)

    de Crom, S. C. M.; Rossen, J. W. A.; de Moor, R. A.; Veldkamp, E. J. M.; van Furth, A. M.; Obihara, C. C.

    Background: Human non-polio enterovirus (EV) and human parechovirus (HPeV) are important pathogens viral infection and aseptic meningitis in children. The aim of this study is to prospectively compare the incidence, clinical signs, blood and cerebrospinal fluid in EV and HPeV infected children.

  15. Enterovirus 71 Neutralizing Antibodies Seroepidemiological Research among Children in Guangzhou, China between 2014 and 2015: A Cross-Sectional Study.

    Science.gov (United States)

    Zhang, Dingmei; Chen, Yan; Chen, Xiashi; He, Zhenjian; Zhu, Xun; Hao, Yuantao

    2017-03-20

    A hand-foot-mouth disease outbreak occurred in 2014 around Guangdong. The purpose of this study was investigating the status and susceptibility of infectious neutralizing antibodies to enterovirus 71 among children so as to provide scientific evidence for the population immunity level of hand-foot-mouth disease and prepare for enterovirus 71 vaccination implementation. Serum specimens were collected from children in communities from January 2014 to March 2015 in Guangzhou. A total of 197 serum samples from children 1-5 years old were collected for this cross-sectional study via non-probabilistic sampling from the database of Chinese National Science and Technique Major Project. Neutralization activity was measured via micro neutralization test in vitro. The positive rate of enterovirus 71 neutralizing antibodies was 59.4%, whereas the geometric mean titre was 1:12.7. A statistically significant difference in true positive rates was found between different age groups but not between different genders. Being the most susceptible population of hand-foot-mouth disease, children under 3 years of age are more likely to be infected with enterovirus 71, and the immunity of children increases with increasing age. Further cohort studies should be conducted, and measures for prevention and vaccination should be taken.

  16. Enterovirus 71 Neutralizing Antibodies Seroepidemiological Research among Children in Guangzhou, China between 2014 and 2015: A Cross-Sectional Study

    Directory of Open Access Journals (Sweden)

    Dingmei Zhang

    2017-03-01

    Full Text Available A hand-foot-mouth disease outbreak occurred in 2014 around Guangdong. The purpose of this study was investigating the status and susceptibility of infectious neutralizing antibodies to enterovirus 71 among children so as to provide scientific evidence for the population immunity level of hand-foot-mouth disease and prepare for enterovirus 71 vaccination implementation. Serum specimens were collected from children in communities from January 2014 to March 2015 in Guangzhou. A total of 197 serum samples from children 1–5 years old were collected for this cross-sectional study via non-probabilistic sampling from the database of Chinese National Science and Technique Major Project. Neutralization activity was measured via micro neutralization test in vitro. The positive rate of enterovirus 71 neutralizing antibodies was 59.4%, whereas the geometric mean titre was 1:12.7. A statistically significant difference in true positive rates was found between different age groups but not between different genders. Being the most susceptible population of hand–foot–mouth disease, children under 3 years of age are more likely to be infected with enterovirus 71, and the immunity of children increases with increasing age. Further cohort studies should be conducted, and measures for prevention and vaccination should be taken.

  17. A dominant EV71-specific CD4+ T cell epitope is highly conserved among human enteroviruses.

    Directory of Open Access Journals (Sweden)

    Ruicheng Wei

    Full Text Available CD4+ T cell-mediated immunity plays a central role in determining the immunopathogenesis of viral infections. However, the role of CD4+ T cells in EV71 infection, which causes hand, foot and mouth disease (HFMD, has yet to be elucidated. We applied a sophisticated method to identify promiscuous CD4+ T cell epitopes contained within the sequence of the EV71 polyprotein. Fifteen epitopes were identified, and three of them are dominant ones. The most dominant epitope is highly conserved among enterovirus species, including HFMD-related coxsackieviruses, HFMD-unrelated echoviruses and polioviruses. Furthermore, the CD4+ T cells specific to the epitope indeed cross-reacted with the homolog of poliovirus 3 Sabin. Our findings imply that CD4+ T cell responses to poliovirus following vaccination, or to other enteroviruses to which individuals may be exposed in early childhood, may have a modulating effect on subsequent CD4+ T cell response to EV71 infection or vaccine.

  18. Enterovirus infections and type 1 diabetes mellitus: is there any relationship?

    Directory of Open Access Journals (Sweden)

    RG Tavares

    2012-01-01

    Full Text Available Several health organizations have classified diabetes mellitus, a metabolic syndrome, as the epidemic of the century, since it affects millions of people worldwide and is one of the top ten causes of death. Type 1 diabetes is considered to be an autoimmune disease, in which autoaggressive T cells infiltrate the islets of Langerhans in the pancreas, leading to the destruction of insulin producing beta cells. The risk of the disease is modulated by genetic factors, mainly genes coding for human leukocyte antigens (HLA. However, the incidence of this disease has increased significantly during the recent decades, which cannot be explained only by genetic factors. Environmental perturbations have also been associated to the development of diabetes. Among these factors, viral triggers have been implicated; particularly enteroviruses, which have been associated to the induction of the disease. Supporting the hypothesis, numerous lines of evidence coming from mouse models and patients with this type of diabetes have shown the association. The present review aims to provide some understanding of how type 1 diabetes occurs and the possible role of enterovirus in this pathology.

  19. Expression and immunogenicity of novel subunit enterovirus 71 VP1 antigens

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Juan [China-US Vaccine Research Center, The First Affiliated Hospital, Nanjing Medical University (China); Department of Microbiology and Immunology, Nanjing Medical University (China); Wang, Shixia [China-US Vaccine Research Center, The First Affiliated Hospital, Nanjing Medical University (China); Department of Medicine, University of Massachusetts Medical School (United States); Gan, Weihua [Department of Pediatrics, The Second Affiliated Hospital, Nanjing Medical University (China); Zhang, Wenhong [Department of Infectious Diseases, Huashan Hospital, Fudan University (China); Ju, Liwen [School of Public Health, Fudan University (China); Huang, Zuhu [Department of Infectious Diseases, The First Affiliated Hospital, Nanjing Medical University (China); China-US Vaccine Research Center, The First Affiliated Hospital, Nanjing Medical University (China); Lu, Shan, E-mail: shan.lu@umassmed.edu [Department of Infectious Diseases, The First Affiliated Hospital, Nanjing Medical University (China); China-US Vaccine Research Center, The First Affiliated Hospital, Nanjing Medical University (China); Department of Medicine, University of Massachusetts Medical School (United States)

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer EV71 is a major emerging infectious disease in many Asian countries. Black-Right-Pointing-Pointer Inactivated EV71 vaccines are in clinical studies but their safety and efficacy are unknown. Black-Right-Pointing-Pointer Developing subunit based EV71 vaccines is significant and novel antigen design is needed. Black-Right-Pointing-Pointer DNA immunization is an efficient tool to test the immunogenicity of VP1 based EV71 vaccines. Black-Right-Pointing-Pointer Multiple VP1 antigens are developed showing immunogenic potential. -- Abstract: Hand, foot, and mouth disease (HFMD) is a common viral illness in young children. HFMD is caused by viruses belonging to the enterovirus genus of the picornavirus family. Recently, enterovirus 71 (EV71) has emerged as a virulent agent for HFMD with severe clinical outcomes. In the current report, we conducted a pilot antigen engineering study to optimize the expression and immunogenicity of subunit VP1 antigen for the design of EV71 vaccines. DNA immunization was adopted as a simple technical approach to test different designs of VP1 antigens without the need to express VP1 protein in vitro first. Our studies indicated that the expression and immunogenicity of VP1 protein can be improved with alternated VP1 antigen designs. Data presented in the current report revealed novel pathways to optimize the design of VP1 antigen-based EV71 vaccines.

  20. Expression and immunogenicity of novel subunit enterovirus 71 VP1 antigens

    International Nuclear Information System (INIS)

    Xu, Juan; Wang, Shixia; Gan, Weihua; Zhang, Wenhong; Ju, Liwen; Huang, Zuhu; Lu, Shan

    2012-01-01

    Highlights: ► EV71 is a major emerging infectious disease in many Asian countries. ► Inactivated EV71 vaccines are in clinical studies but their safety and efficacy are unknown. ► Developing subunit based EV71 vaccines is significant and novel antigen design is needed. ► DNA immunization is an efficient tool to test the immunogenicity of VP1 based EV71 vaccines. ► Multiple VP1 antigens are developed showing immunogenic potential. -- Abstract: Hand, foot, and mouth disease (HFMD) is a common viral illness in young children. HFMD is caused by viruses belonging to the enterovirus genus of the picornavirus family. Recently, enterovirus 71 (EV71) has emerged as a virulent agent for HFMD with severe clinical outcomes. In the current report, we conducted a pilot antigen engineering study to optimize the expression and immunogenicity of subunit VP1 antigen for the design of EV71 vaccines. DNA immunization was adopted as a simple technical approach to test different designs of VP1 antigens without the need to express VP1 protein in vitro first. Our studies indicated that the expression and immunogenicity of VP1 protein can be improved with alternated VP1 antigen designs. Data presented in the current report revealed novel pathways to optimize the design of VP1 antigen-based EV71 vaccines.

  1. Evaluation of the enterovirus laboratory surveillance system in Denmark, 2010 to 2013

    DEFF Research Database (Denmark)

    Condell, Orla; Midgley, Sofie E; Christiansen, Claus Bohn

    2016-01-01

    The primary aim of the Danish enterovirus (EV) surveillance system is to document absence of poliovirus infection. The conflict in Syria has left many children unvaccinated and movement from areas with polio cases to Europe calls for increased awareness to detect and respond to virus-transmission......The primary aim of the Danish enterovirus (EV) surveillance system is to document absence of poliovirus infection. The conflict in Syria has left many children unvaccinated and movement from areas with polio cases to Europe calls for increased awareness to detect and respond to virus......-transmission in a timely manner. We evaluate the national EV laboratory surveillance, to generate recommendations for system strengthening. The system was analysed for completeness of viral typing analysis and clinical information and timeliness of specimen collection, laboratory results and reporting of clinical...... information. Of 23,720 specimens screened, 2,202 (9.3%) were EV-positive. Submission of cerebrospinal fluid and faecal specimens from primary diagnostic laboratories was 79.5% complete (845/1,063), and varied by laboratory and patient age. EV genotypes were determined in 68.5% (979/1,430) of laboratory...

  2. CLINICAL AND LABORATORY CHARACTERISTICS OF THE INFECTION CAUSED BY ENTEROVIRUS-71

    Directory of Open Access Journals (Sweden)

    E. N. Simovanyan

    2014-01-01

    Full Text Available Adverse course of enterovirus-71 infection (EVI-71 in children, frequent development of the nervous system pathology determine the need for early disease diagnosis. The study included 139 children aged 6 months to 13 years. Modern EVI-71 features is a frequent defeat of children aged from 3 to 7 years, attending organized groups, the priority development of combined moderate severity forms without nervous system pathology. EVI-71 is characterized by cyclical course, appearance in the first two disease days fever, murrain-like, catarrhal, lymphoproliferative syndromes, conjunctivitis, headaches. The second stage of the disease (3— 6 days in 37.9% children accompanied by attaching of meningitis and meningoencephalitis symptoms (common cerebral, meningeal and encephalic syndromes, changes in cerebrospinal fluid. In the EVI-71 diagnosis must be observed epidemiological history, clinical and laboratory parameters, detection of enterovirus-71 and its RNA from feces, oropharyngeal mucus and cerebrospinal fluid. Patients with EVI-71 need for combined treatment, including causal agents and pathogenetic therapy. 

  3. Detection of enteroviruses in cases of neurological disorders in the State of Pará, Brazil

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    GOMES Maria de Lourdes Contente

    2001-01-01

    Full Text Available Eighty-one cerebrospinal fluid (CSF samples mainly from cases of aseptic meningitis and motor deficiency syndrome were sent to the Virology Section of Evandro Chagas Institute, Belém Pará, in the period of January 1995 to January 1996 in order to isolate viruses. All samples were inoculated onto HEp-2 cell culture and newborn mice, with negative results. The probability of isolating viruses by these methods is reduced because of the low concentration of viral particles in these specimens. In order to obtain more information about the etiology of these cases, a group of 23 samples were selected to be tested by a more sensitive technique than the virus isolation - the reverse transcription polymerase chain reaction (RT-PCR. Specific primers directed to conserved regions in the enterovirus genome were used, considering that this group of viruses is frequently associated with these neurological disorder. The age of the patients ranged from 1 to 55 years and nearly all of them lived in Belém, State of Pará, North of Brazil. Of 15 samples analyzed by RT PCR nine (60% were positive; of these, 6 (66.6% had motor deficiency and 3 (33.3% developed aseptic meningitis. These results show that it is important to investigate enterovirus as cause of these syndromes.

  4. Neutralizing Antibodies to Enterovirus 71 in Belém, Brazil

    Directory of Open Access Journals (Sweden)

    Maria de Lourdes C Gomes

    2002-01-01

    Full Text Available Non-polio enteroviruses (Coxsackievirus A, Coxsackievirus B, Echovirus and EV 68-72 which belong to the enterovirus (EV genus, Picornaviridae family, may be responsible for acute flaccid paralysis, aseptic meningitis, myocarditis, hepatitis, pleurodinia, neonatal sepsis, hand, foot and mouth disease (HFMD even though 50-80% of infections are asymptomatic. EV 71 has been responsible for outbreaks and epidemics of HFMD and acute neurologic disease justifying its study in our country. The aim of this study was to detect neutralizing antibodies (NtAb to EV 71 in individuals up to 15 years of age living in Belém, State of Pará, northern Brazil. Serum samples from 238 patients attending the Virology Sector of Evandro Chagas Institute in Belém, Brazil, were analyzed using microneutralization tests that included RD cells and BrCr strain. Overall 40.8% (97/238 of tested samples had NtAb to EV 71. Regarding the distribution per age group, 85.2% (92/108 of patients aged 0-3 years had no NtAb to this virus and 69.2% of those 12 to15 years of age were seropositive. These results confirm that EV 71 infection occurs in the city of Belém; and that a high rate of individuals in this study were infected aged 3 years and over and, when aged 15 years nearly 70% had EV 71 NtAb.

  5. Sequence analysis of the first C2 subgenogroup strain of enterovirus 71 isolated in Korea.

    Science.gov (United States)

    Lee, Jung-Ah; Yoon, Young Sil; Hyeon, Ji-Yeon; Yoo, Jung Sik; Lee, Sang-Won; Lee, June-Woo; Lee, Sang-Won

    2016-12-01

    Enterovirus 71 (EV71) is an important causative agent of hand-foot-and-mouth disease with severe neurological complications, which may lead to death in children. Large outbreaks caused by EV71 have frequently occurred in Asia-Pacific region. In Korea, the outbreaks have been caused by EV71 subgenogroups C3, and C4. Only genogroup C, especially subgenogroup C1, C3, C4, and C5, has been detected by the national enterovirus surveillance system in Korea. This study reports the first isolation of EV71 A1451 strain, which belongs to subgenogroup C2. EV71 was isolated from a Korean patient with meningoencephalitis. Complete genome analysis and phylogenetic analysis was performed to identify the characteristics of the strain. Comparative genome analysis of the A1451 strain indicated that this novel C2 strain is associated with the Taiwan strains, which are recombinant virus combined with subgenogroup C2 and B3. Because the subgenogroup B3 was not previously detected in Korea, the A1451 strain is regarded as an imported recombinant virus. Periodic surveillance of EV71 is required to control the spread of this disease and its introduction from overseas. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. [Research progress on seroepidemiological study of enterovirus 71 and coxsackievirus A16 infection among children].

    Science.gov (United States)

    Luo, Li; Xing, Weijia; Liao, Qiaohong; Yu, Hongjie

    2015-02-01

    Most common causative agents for hand, foot and mouth disease (HFMD) are enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16). The symptomatic and asymptomatic cases could transmit the disease in population. Many sero-epidemiological surveys were launched to estimate the sero-incidence of EV-A71 and CV-A16 enterovirus, the susceptibility of different sub-population, and to observe the dynamics of neutralizing antibody. A literature search of sero-epidemiological study focused on EV-A71 or CV-A16 was conducted via PubMed and China Hospital Knowledge Database. Based on the 20 selected studies, the different age groups' antibody level, the susceptibility, the dynamics of antibody and sero-incidence of EV-A71 or CV-A16 were analyzed. From our results, the antibody level against EV-A71 or CV-A16 in neonates was associated with their mothers, which was similar with that of adults. The antibody level against EV-A71 or CV-A16 in neonates dropped to lowest level at one years-old, and started to dramatically increase until four years-old, and reached a plateau at five years-old. In conclusion, the infants aged 6-12 months were the priority group to receive vaccination when the EV-A71 vaccine is licensed in the future.

  7. Studies on Inhibition of Proliferation of Enterovirus-71 by Compound YZ-LY-0.

    Science.gov (United States)

    Yang, Qingzhan; Jie, Qing; Shaw, Neil; Li, Lei; Rao, Zihe; Yin, Zheng; Lou, Zhiyong

    2015-01-01

    In recent years, hand-foot-and-mouth disease (HFMD), which is caused by Enteroviruses, has emerged as a serious illness. It affects mainly children under the age of five and results in high fatality rates. Enterovirus 71 (EV71) is the main causative agent of HFMD in China and currently there are no effective anti-viral drugs available to treat HFMD. In the present study, we screened compounds for inhibition of proliferation of EV71. Compound YZ-LY-0 stalled the life cycle of EV71. The inhibitor exhibited EC50 value of 0.29 μm against SK-EV006 strain of EV71. Notably, YZ-LY-0 had low cytotoxicity (CC50 > 100 μM) and a high selectivity index (over 300) in Vero and RD cells. YZ-LY-0 in combination with an EV71 RdRp inhibitor or an entry inhibitor showed an antagonistic effect at very low concentrations. However, at higher concentrations the inhibitors exhibited a synergistic effect in inhibiting viral replication. Preliminary results on investigation of the mechanism of inhibition indicate that YZ-LY-0 does not block the entry of the virus in the host cell, but instead inhibits an early stage of EV71 replication. Our studies provide a potential clinical therapeutic option against EV71 infections and suggest that a combined application of YZ-LY-0 with other inhibitors could be more effective in the treatment of HFMD.

  8. A rapid molecular diagnosis of cutaneous leishmaniasis bycolorimetric malachite green-loop-mediated isothermal amplification(LAMP) combined with an FTA card as a direct sampling tool

    OpenAIRE

    Nzelu, Chukwunonso O.; Cáceres, Abraham G.; Guerrero-Quincho, Silvia; Tineo-Villafuerte, Edwin; Rodriquez-Delfin, Luis; Mimori, Tatsuyuki; Uezato, Hiroshi; Katakura, Ken; Gomez, Eduardo A.; Guevara, Angel G.; Hashiguchi, Yoshihisa; Kato, Hirotomo

    2015-01-01

    Leishmaniasis remains one of the world's most neglected diseases, and early detection of the infectious agent, especially in developing countries, will require a simple and rapid test. In this study, we established a quick, one-step, single-tube, highly sensitive loop-mediated isothermal amplification (LAMP) assay for rapid detection of Leishmania DNA from tissue materials spotted on an FTA card. An FTA-LAMP with pre-added malachite green was performed at 64 degrees C for 60 mm using a heatin...

  9. A Randomized, Double-Blind, Placebo-Controlled Trial of Pleconaril for the Treatment of Neonates With Enterovirus Sepsis.

    Science.gov (United States)

    Abzug, Mark J; Michaels, Marian G; Wald, Ellen; Jacobs, Richard F; Romero, José R; Sánchez, Pablo J; Wilson, Gregory; Krogstad, Paul; Storch, Gregory A; Lawrence, Robert; Shelton, Mark; Palmer, April; Robinson, Joan; Dennehy, Penelope; Sood, Sunil K; Cloud, Gretchen; Jester, Penelope; Acosta, Edward P; Whitley, Richard; Kimberlin, David

    2016-03-01

    Neonatal enterovirus sepsis has high mortality. Antiviral therapy is not available. Neonates with suspected enterovirus sepsis (hepatitis, coagulopathy, and/or myocarditis) with onset at ≤15 days of life were randomized 2:1 to receive oral pleconaril or placebo for 7 days. Serial virologic (oropharynx, rectum, urine, serum), clinical, pharmacokinetic, and safety evaluations were performed. Sixty-one subjects were enrolled (43 treatment, 18 placebo), of whom 43 were confirmed enterovirus infected (31 treatment, 12 placebo). There was no difference in day 5 oropharyngeal culture positivity (primary endpoint; 0% in both groups). However, enterovirus-infected subjects in the treatment group became culture negative from all anatomic sites combined faster than placebo group subjects (median 4.0 versus 7.0 days, P = .08), and fewer subjects in the treatment group remained polymerase chain reaction (PCR)-positive from the oropharynx when last sampled (23% versus 58%, P = .02; median, 14.0 days). By intent to treat, 10/43 (23%) subjects in the treatment group and 8/18 (44%) in the placebo group died (P = .02 for 2-month survival difference); among enterovirus-confirmed subjects, 7/31 (23%) in the treatment group died versus 5/12 (42%) in the placebo group (P = .26). All pleconaril recipients attained concentrations greater than the IC90 after the first study day, but 38% were less than the IC90 during the first day of treatment. One subject in the treatment group and three in the placebo group had treatment-related adverse events. Shorter times to culture and PCR negativity and greater survival among pleconaril recipients support potential efficacy and warrant further evaluation. © The Author 2015. Published by Oxford University Press on behalf of the Pediatric Infectious Diseases Society. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  10. Enterovirus-related diarrhoea in Guangdong, China: clinical features and implications in hand, foot and mouth disease and herpangina.

    Science.gov (United States)

    Zhou, Hong-Tao; Yi, Hai-Su; Guo, Yong-Hui; Pan, Yu-Xian; Tao, Shao-Hua; Wang, Bin; Chen, Man-Jun; Yang, Mei; Yu, Nan

    2016-03-16

    A series of complications caused by enteroviruses, including meningitis, encephalitis, acute flaccid paralysis, acute cardiopulmonary failure, respiratory infection, and myocardial injury have been reported in hand, foot and mouth disease/herpangina (HFMD/HA). However, the complication of diarrhoea caused by enteroviruses has been neglected, and a summary of its clinical features and impact on HFMD/HA is unavailable. We included inpatients with HFMD/HA admitted to the Paediatric Department of Zhujiang Hospital during 2009-2012. We summarised and compared clinical data for cases with and without diarrhoea, and determined enterovirus serotypes by reverse transcriptase polymerase chain reaction and genotyping based on a partial-length fragment of viral protein 1 or the 5'-untranslated region. There were 804 inpatients with HFMD/HA and 28 (3.5%) presented with diarrhoea. Gastrointestinal symptoms were mild in most cases of diarrhoea (82.1%), with high prevalence of no dehydration (82.1%), short duration of diarrhoea (78.6%) and watery stools (75.0%). The prevalence of multi-organ dysfunction syndrome (10.7 vs 0.40%) (p = 0.001), hepatic injury (14.3 vs 3.4%) (p = 0.019), myocardial injury (21.4 vs 6.1%) (p = 0.002) and convulsion (21.4 vs 7.2%) (p = 0.016) was significantly higher in the diarrhoea than no diarrhoea group. There was no significant difference between the two groups regarding prevalence of death, altered consciousness, paralysis, central nervous system involvement, or acute respiratory infection. Most patients with diarrhoea caused by enteroviruses circulating in Guangdong Province in 2009-2012 had mild or moderate gastrointestinal symptoms. Although enterovirus-related diarrhoea caused additional multi-organ dysfunction syndrome, hepatic injury and myocardial injury in children with HFMD/HA, timely intervention efficiently reduced disease severity and improved outcome.

  11. Detection of human enterovirus 71 and coxsackievirus A16 in children with hand, foot and mouth disease in China.

    Science.gov (United States)

    Chen, Ling; Mou, Xiaozhou; Zhang, Qiong; Li, Yifei; Lin, Jian; Liu, Fanlong; Yuan, Li; Tang, Yiming; Xiang, Charlie

    2012-04-01

    The aims of the present study were to investigate the genetic characteristics of enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) strains in China and to evaluate the relationship between the genotypes of CVA16 and EV71 and their geographical distribution. A total of 399 stool specimens were collected from children with symptoms of hand, foot and mouth disease (HFMD) in Zhejiang Province. The presence of enteroviruses was determined using reverse transcription-semi-nested PCR targeted to the VP1 gene of all human enteroviruses and DNA sequencing. EV71 and CVA16, the major etiological agents of HFMD, were detected in 38.4% (38/99) and 35.4% (35/99) of HEV-A species-positive cases, respectively. Based on the phylogenetic analysis of the VP1 gene, EV71 strains identified in this study belong to subgenotype C4, and CVA16 strains herein were classified into clusters B2a and B2b within the genotype B2. Taking into consideration other published data, we conclude that the genetic characteristics of enteroviruses in China reflect the pattern of the endemic circulation of the subgenotype C4 to EV71 and clusters B2a and B2b within genotype B2 to CVA16, which have been continuously circulating in China since 1997. This observation indicates that the genetic characteristics of enteroviruses in China seem to depend on their special geographical and climatical features allowing them to be sustained with little external effect.

  12. Possible Internalization of an Enterovirus in Hydroponically Grown Lettuce.

    Science.gov (United States)

    Carducci, Annalaura; Caponi, Elisa; Ciurli, Adriana; Verani, Marco

    2015-07-17

    Several studies have shown that enteric viruses can be transferred onto the surface of vegetables and fruits through spray irrigation, but, recently, reports have suggested viral contamination of vegetables sub-irrigated with reused wastewater. Hydroponic cultures, used to grow ready to eat fresh lettuce, have also been used to study the possibility of viral absorption through roots. This study was conducted to assess a possible risk of viral contamination in lettuce from contaminated water. The leaves of lettuce plants grown in hydroponic cultures where the roots were exposed to water containing Coxsakievirus B2, were analysed for evidence of the virus. The plants and water were sampled at different times and virus was measured using quantitative RT-PCR and infectivity assay. In leaf samples, the lowest observed infective data were lower than the qRT-PCR detection limits, suggesting that free viral RNA or damaged viruses are eliminated rapidly while infectious particles remain stable for a longer time. The obtained data revealed that the leaves were contaminated at a water concentration of 4.11 ± 1 Log Most Probable Number/L (8.03 ± 1 Log GC/L) a concentration observed in contaminated untreated water of wastewater treatment plants. However, the absorption dynamics and whether the virus is inactive in the leaves still remains to be clarified. Nevertheless, this work has practical implications for risk management in using reclaimed water for agricultural use; when irrigated vegetables are destined for raw consumption, virological contamination in water sources should be evaluated.

  13. QUANTIFICATION OF ENTEROVIRUS AND HEPATITIS A VIRUSES IN WELLS AND SPRINGS IN EAST TENNESSEE USING REAL-TIME REVERSE TRANSCIPTION PCR

    Science.gov (United States)

    This project involves development, validation testing and application of a fast, efficient method of quantitatively measuring occurrence and concentration of common human viral pathogens, enterovirus and hepatitis A virus, in ground water samples using real-time reverse transcrip...

  14. Possible Internalization of an Enterovirus in Hydroponically Grown Lettuce

    Directory of Open Access Journals (Sweden)

    Annalaura Carducci

    2015-07-01

    Full Text Available Several studies have shown that enteric viruses can be transferred onto the surface of vegetables and fruits through spray irrigation, but, recently, reports have suggested viral contamination of vegetables sub-irrigated with reused wastewater. Hydroponic cultures, used to grow ready to eat fresh lettuce, have also been used to study the possibility of viral absorption through roots. This study was conducted to assess a possible risk of viral contamination in lettuce from contaminated water. The leaves of lettuce plants grown in hydroponic cultures where the roots were exposed to water containing Coxsakievirus B2, were analysed for evidence of the virus. The plants and water were sampled at different times and virus was measured using quantitative RT-PCR and infectivity assay. In leaf samples, the lowest observed infective data were lower than the qRT-PCR detection limits, suggesting that free viral RNA or damaged viruses are eliminated rapidly while infectious particles remain stable for a longer time. The obtained data revealed that the leaves were contaminated at a water concentration of 4.11 ± 1 Log Most Probable Number/L (8.03 ± 1 Log GC/L a concentration observed in contaminated untreated water of wastewater treatment plants. However, the absorption dynamics and whether the virus is inactive in the leaves still remains to be clarified. Nevertheless, this work has practical implications for risk management in using reclaimed water for agricultural use; when irrigated vegetables are destined for raw consumption, virological contamination in water sources should be evaluated.

  15. A rapid molecular diagnosis of cutaneous leishmaniasis by colorimetric malachite green-loop-mediated isothermal amplification (LAMP) combined with an FTA card as a direct sampling tool.

    Science.gov (United States)

    Nzelu, Chukwunonso O; Cáceres, Abraham G; Guerrero-Quincho, Silvia; Tineo-Villafuerte, Edwin; Rodriquez-Delfin, Luis; Mimori, Tatsuyuki; Uezato, Hiroshi; Katakura, Ken; Gomez, Eduardo A; Guevara, Angel G; Hashiguchi, Yoshihisa; Kato, Hirotomo

    2016-01-01

    Leishmaniasis remains one of the world's most neglected diseases, and early detection of the infectious agent, especially in developing countries, will require a simple and rapid test. In this study, we established a quick, one-step, single-tube, highly sensitive loop-mediated isothermal amplification (LAMP) assay for rapid detection of Leishmania DNA from tissue materials spotted on an FTA card. An FTA-LAMP with pre-added malachite green was performed at 64°C for 60min using a heating block and/or water bath and DNA amplification was detected immediately after incubation. The LAMP assay had high detection sensitivity down to a level of 0.01 parasites per μl. The field- and clinic-applicability of the colorimetric FTA-LAMP assay was demonstrated with 122 clinical samples collected from patients suspected of having cutaneous leishmaniasis in Peru, from which 71 positives were detected. The LAMP assay in combination with an FTA card described here is rapid and sensitive, as well as simple to perform, and has great potential usefulness for diagnosis and surveillance of leishmaniasis in endemic areas. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Enterovirus Exposure Uniquely Discriminates Type 1 Diabetes Patients with a Homozygous from a Heterozygous Melanoma Differentiation-Associated Protein 5/Interferon Induced with Helicase C Domain 1 A946T Genotype.

    Science.gov (United States)

    Schulte, Barbara M; Gielen, Paul R; Kers-Rebel, Esther D; Prosser, Amy C; Lind, Katharina; Flodström-Tullberg, Malin; Tack, Cees J; Elving, Lammy D; Adema, Gosse J

    2016-09-01

    In children at risk for type 1 diabetes, innate immune activity is detected before seroconversion. Enterovirus infections have been linked to diabetes development, and a polymorphism (A946T) in the innate immune sensor recognizing enterovirus RNA, interferon-induced with helicase C domain 1/melanoma differentiation-associated protein 5, predisposes to disease. We hypothesized that the strength of innate antienteroviral responses is affected in autoimmune type 1 diabetes patients and linked to the A946T polymorphism. We compared induction of interferon-stimulated genes (ISGs) in peripheral blood mononuclear cells (PBMCs) and dendritic cells (DCs) in healthy individuals and diabetes patients upon stimulation with enterovirus, enterovirus-antibody complexes, or ligands mimicking infection in relation to the A946T polymorphism. Overall, PBMCs of diabetes patients and healthy donors showed comparable ISG induction upon stimulation. No differences were observed in DCs. Interestingly, the data imply that the magnitude of responses to enterovirus and enterovirus-antibody complexes in PBMCs is critically influenced by the A946T polymorphism and elevated in heterozygotes compared to TT homozygous individuals in autoimmune diabetes patients, but not healthy controls. These data imply an intrinsic difference in the responses to enterovirus and enterovirus-antibody complexes in diabetes patients carrying a TT risk genotype compared to heterozygotes that may influence control of enterovirus clearance.

  17. Nuclear Protein Sam68 Interacts with the Enterovirus 71 Internal Ribosome Entry Site and Positively Regulates Viral Protein Translation.

    Science.gov (United States)

    Zhang, Hua; Song, Lei; Cong, Haolong; Tien, Po

    2015-10-01

    Enterovirus 71 (EV71) recruits various cellular factors to assist in the replication and translation of its genome. Identification of the host factors involved in the EV71 life cycle not only will enable a better understanding of the infection mechanism but also has the potential to be of use in the development of antiviral therapeutics. In this study, we demonstrated that the cellular factor 68-kDa Src-associated protein in mitosis (Sam68) acts as an internal ribosome entry site (IRES) trans-acting factor (ITAF) that binds specifically to the EV71 5' untranslated region (5'UTR). Interaction sites in both the viral IRES (stem-loops IV and V) and the heterogeneous nuclear ribonucleoprotein K homology (KH) domain of Sam68 protein were further mapped using an electrophoretic mobility shift assay (EMSA) and biotin RNA pulldown assay. More importantly, dual-luciferase (firefly) reporter analysis suggested that overexpression of Sam68 positively regulated IRES-dependent translation of virus proteins. In contrast, both IRES activity and viral protein translation significantly decreased in Sam68 knockdown cells compared with the negative-control cells treated with short hairpin RNA (shRNA). However, downregulation of Sam68 did not have a significant inhibitory effect on the accumulation of the EV71 genome. Moreover, Sam68 was redistributed from the nucleus to the cytoplasm and interacts with cellular factors, such as poly(rC)-binding protein 2 (PCBP2) and poly(A)-binding protein (PABP), during EV71 infection. The cytoplasmic relocalization of Sam68 in EV71-infected cells may be involved in the enhancement of EV71 IRES-mediated translation. Since Sam68 is known to be a RNA-binding protein, these results provide direct evidence that Sam68 is a novel ITAF that interacts with EV71 IRES and positively regulates viral protein translation. The nuclear protein Sam68 is found as an additional new host factor that interacts with the EV71 IRES during infection and could potentially

  18. TIA-1 and TIAR interact with 5'-UTR of enterovirus 71 genome and facilitate viral replication.

    Science.gov (United States)

    Wang, Xiaohui; Wang, Huanru; Li, Yixuan; Jin, Yu; Chu, Ying; Su, Airong; Wu, Zhiwei

    2015-10-16

    Enterovirus 71 is one of the major causative pathogens of HFMD in children. Upon infection, the viral RNA is translated in an IRES-dependent manner and requires several host factors for effective replication. Here, we found that T-cell-restricted intracellular antigen 1 (TIA-1), and TIA-1 related protein (TIAR) were translocated from nucleus to cytoplasm after EV71 infection and localized to the sites of viral replication. We found that TIA-1 and TIAR can facilitate EV71 replication by enhancing the viral genome synthesis in host cells. We demonstrated that both proteins bound to the stem-loop I of 5'-UTR of viral genome and improved the stability of viral genomic RNA. Our results suggest that TIA-1 and TIAR are two new host factors that interact with 5-UTR of EV71 genome and positively regulate viral replication. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Detection and genotyping of enteroviruses in cerebrospinal fluid in patients in Victoria, Australia, 2007-2013.

    Science.gov (United States)

    Papadakis, Georgina; Chibo, Doris; Druce, Julian; Catton, Michael; Birch, Chris

    2014-09-01

    Genotyping by VP1 fragment polymerase chain reaction (PCR) and nucleic acid sequencing to detect enterovirus (EV) genotypes was performed directly on 729 EV PCR positive cerebrospinal fluid (CSF) samples collected between 2007 and 2012 from Victorian hospital inpatients. The overall genotype identification rate from CSF-positive material was 43%. The four most common genotypes identified were Echovirus 6 (24%), Echovirus 30 (17%), Echovirus 25 (10%), and Coxsackievirus A9 (10%), together comprising 61% of all EVs typed. The seasonal distribution of all EVs identified followed the recognized pattern of mainly summer epidemics. Three of the four predominant genotypes were present in each of the 6 years in which the study was conducted, with 20 other EV genotypes also detected, often in only a single year. Genotyping of EVs directly in CSF is faster, simpler and more sensitive than traditional virus neutralization assays performed on EV positive samples. © 2014 Wiley Periodicals, Inc.

  20. Cyclical Patterns of Hand, Foot and Mouth Disease Caused by Enterovirus A71 in Malaysia

    Science.gov (United States)

    NikNadia, NMN; Sam, I-Ching; Rampal, Sanjay; WanNorAmalina, WMZ; NurAtifah, Ghazali; Verasahib, Khebir; Ong, Chia Ching; MohdAdib, MohdAidinniza; Chan, Yoke Fun

    2016-01-01

    Enterovirus A71 (EV-A71) is an important emerging pathogen causing large epidemics of hand, foot and mouth disease (HFMD) in children. In Malaysia, since the first EV-A71 epidemic in 1997, recurrent cyclical epidemics have occurred every 2–3 years for reasons that remain unclear. We hypothesize that this cyclical pattern is due to changes in population immunity in children (measured as seroprevalence). Neutralizing antibody titers against EV-A71 were measured in 2,141 residual serum samples collected from children ≤12 years old between 1995 and 2012 to determine the seroprevalence of EV-A71. Reported national HFMD incidence was highest in children Malaysia is mainly due to the fall of population immunity accompanying the accumulation of susceptible children between epidemics. This study will impact the future planning, timing and target populations for vaccine programs. PMID:27010319

  1. Enterovirus co-infections and onychomadesis after hand, foot, and mouth disease, Spain, 2008.

    Science.gov (United States)

    Bracho, Maria A; González-Candelas, Fernando; Valero, Ana; Córdoba, Juan; Salazar, Antonio

    2011-12-01

    Hand, foot, and mouth disease (HFMD), a common disease caused by enteroviruses (EVs), usually affects children. Clustered and sporadic HFMD cases, followed by onychomadesis (nail shedding), occurred during summer and fall 2008 in Valencia, Spain. Fecal samples from onychomadesis patients, who did or did not have previous HFMD, and from healthy children exposed to onychomadesis patients tested positive for EV. The complete viral protein 1 capsid gene sequence was obtained for typing and phylogenetic analysis. Two EV serotypes, coxsackievirus A10 and coxsackievirus B1 (CVB1), were mainly detected as a monoinfection or co-infection in a childcare center where an onychomadesis outbreak occurred. On the basis of our results, and detection of CVB1 in 2 other contemporary onychomadesis outbreaks in childcare centers in Spain, we propose that mixed infection of an EV serotype that causes HFMD, plus the serotype CVB1, could explain the emergence after HFMD of onychomadesis, a rare and late complication.

  2. Lycorine reduces mortality of human enterovirus 71-infected mice by inhibiting virus replication

    Directory of Open Access Journals (Sweden)

    Qin Chuan

    2011-10-01

    Full Text Available Abstract Human enterovirus 71 (EV71 infection causes hand, foot and mouth disease in children under 6 years old and this infection occasionally induces severe neurological complications. No vaccines or drugs are clinical available to control EV71 epidemics. In present study, we show that treatment with lycorine reduced the viral cytopathic effect (CPE on rhabdomyosarcoma (RD cells by inhibiting virus replication. Analysis of this inhibitory effect of lycorine on viral proteins synthesis suggests that lycorine blocks the elongation of the viral polyprotein during translation. Lycorine treatment of mice challenged with a lethal dose of EV71 resulted in reduction of mortality, clinical scores and pathological changes in the muscles of mice, which were achieved through inhibition of viral replication. When mice were infected with a moderate dose of EV71, lycorine treatment was able to protect them from paralysis. Lycorine may be a potential drug candidate for the clinical treatment of EV71-infected patients.

  3. Phage Display-Derived Cross-Reactive Neutralizing Antibody against Enterovirus 71 and Coxsackievirus A16.

    Science.gov (United States)

    Zhang, Xiao; Sun, Chunyun; Xiao, Xiangqian; Pang, Lin; Shen, Sisi; Zhang, Jie; Cen, Shan; Yang, Burton B; Huang, Yuming; Sheng, Wang; Zeng, Yi

    2016-01-01

    Enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) are members of the Picornaviridae family and are considered the main causative agents of hand, foot and mouth disease (HFMD). In recent decades large HFMD outbreaks caused by EV71 and CVA16 have become significant public health concerns in the Asia-Pacific region. Vaccines and antiviral drugs are unavailable to prevent EV71 and CVA16 infection. In the current study, a chimeric antibody targeting a highly conserved peptide in the EV71 VP4 protein was isolated by using a phage display technique. The antibody showed cross-neutralizing capability against EV71 and CVA16 in vitro. The results suggest that this phage display-derived antibody will have great potential as a broad neutralizing antibody against EV71 and CVA16 after affinity maturation and humanization.

  4. Enterovirus D68-associated community-acquired pneumonia in children living in Milan, Italy.

    Science.gov (United States)

    Esposito, Susanna; Zampiero, Alberto; Ruggiero, Luca; Madini, Barbara; Niesters, Hubert; Principi, Nicola

    2015-07-01

    An increasing number of children infected by enterovirus D68 (EV-D68) and affected by severe respiratory illness, muscle weakness and paralysis were described in the USA and Canada in 2014 OBJECTIVES: To investigate the potential involvement of EV-D68 in determining community-acquired pneumonia (CAP) in hospitalised children in order to acquire information concerning the clinical problems associated with EV-D68 in Italy. This prospective study of children hospitalised for CAP in the largest Pediatric Department in Milan, Italy, was carried out between 1 June and 31 December 2014. All of the children's admission nasopharyngeal swabs were investigated for the presence of EV-D68. One hundred and seventy-six children with radiographically confirmed CAP were hospitalised during the 7-month study period: 97 (55.1%) had enterovirus/rhinovirus-positive nasopharyngeal samples, including four (2.3%) positive for EV-D68. These four samples were collected between 9 and 21 October, a month in which 21 cases of CAP were recorded. Phylogenetic analysis showed that all of the sequences fell into clade B. The most severe case was diagnosed in a 14-year-old girl with mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS syndrome), who died after 12 days of hospitalisation. EV-D68 was detected in few children with usually mild-to-moderate lower respiratory tract infection, although the disease lead to the death of a girl with a severe chronic underlying disease. Further studies capable of better defining the epidemiological, genetic and pathogenetic characteristics of the virus are required in order to be able to prepare appropriate preventive and therapeutic measures. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Expression of enterovirus 71 capsid protein VP1 in Escherichia coli and its clinical application

    Directory of Open Access Journals (Sweden)

    Mei Shi

    2013-12-01

    Full Text Available The VPl gene of enterovirus 71 (EV71 was synthesized, construct a recombinant plasmid pET15b/VP1 and expressed in E. coli BL21. The recombinant VP1 protein could specifically react with EV71-infected patient sera without the cross-reaction with serum antibodies of coxsackievirus A16 (CA16, A4, A5, B3 and B5 as well as echovirus 6. In acute and convalescent phases, IgM and IgG antibodies of 182 serum samples were detected by ELISA with recombinant VP1 protein as a coated antigen. The results showed that the sensitivity, specificity, positive predictive value (PPV and negative predictive value (NPV of IgM antibodies in serum samples for the diagnosis of EV71 infection were 90.1, 98.4, 98.8 and 88.7%, respectively; similarly, those of IgG antibodies in serum samples were 82.4, 89.1, 91.5 and 78.1%, respectively. Five of 80 samples (6.25% from CA16infected patients were detected positive by ELISA with recombinant VP1 protein in which indicated the cross reactions and 0 of 5 samples from patients infected with other enteroviruses including CA4, CA5, CB3, CB5 and echovirus 6. Therefore, the recombinant VP1 protein of EV7l may provide a theoretical reference for establishing an effective antibody screening of IgM for EV71-infected patients with clinically suspected hand, foot, and mouth disease (HFMD.

  6. Monitoring antigenic variations of enterovirus 71: implications for virus surveillance and vaccine development.

    Directory of Open Access Journals (Sweden)

    Min-Yuan Chia

    2014-07-01

    Full Text Available Enterovirus 71 (EV71 causes life-threatening epidemics in Asia and can be phylogenetically classified into three major genogroups (A ∼ C including 11 genotypes (A, B1 ∼ B5, and C1 ∼ C5. Recently, EV71 epidemics occurred cyclically in Taiwan with different genotypes. In recent years, human studies using post-infection sera obtained from children have detected antigenic variations among different EV71 strains. Therefore, surveillance of enterovirus 71 should include phylogenetic and antigenic analysis. Due to limitation of sera available from children with EV71 primary infection, suitable animal models should be developed to generate a panel of antisera for monitoring EV71 antigenic variations. Twelve reference strains representing the 11 EV71 genotypes were grown in rhabdomyosarcoma cells. Infectious EV71 particles were purified and collected to immunize rabbits. The rabbit antisera were then employed to measure neutralizing antibody titers against the 12 reference strains and 5 recent strains. Rabbits immunized with genogroup B and C viruses consistently have a lower neutralizing antibody titers against genogroup A (≧ 8-fold difference and antigenic variations between genogroup B and C viruses can be detected but did not have a clear pattern, which are consistent with previous human studies. Comparison between human and rabbit neutralizing antibody profiles, the results showed that ≧ 8-fold difference in rabbit cross-reactive antibody ratios could be used to screen EV71 isolates for identifying potential antigenic variants. In conclusion, a rabbit model was developed to monitor antigenic variations of EV71, which are critical to select vaccine strains and predict epidemics.

  7. From Lab to Lake - Evaluation of Current Molecular Methods for the Detection of Infectious Enteric Viruses in Complex Water Matrices in an Urban Area.

    Directory of Open Access Journals (Sweden)

    Mats Leifels

    Full Text Available Quantitative PCR methods are commonly used to monitor enteric viruses in the aquatic environment because of their high sensitivity, short reaction times and relatively low operational cost. However, conclusions for public health drawn from results of such molecular techniques are limited due to their inability to determine viral infectivity. Ethidium monoazide (EMA and propidium monoazide (PMA are capable to penetrate the damaged or compromised capsid of the inactivated viruses and bind to the viral nucleic acids. We assessed whether dye treatment is a suitable approach to improve the ability of qPCR to distinguish between infectious and non-infectious human adenovirus, enterovirus and rotavirus A in surface water of an urban river and sewage before and after UV disinfection. Like the gold standard of cell culture assays, pretreatment EMA-/PMA-qPCR succeeded in removing false positive results which would lead to an overestimation of the viral load if only qPCR of the environmental samples was considered. A dye pretreatment could therefore provide a rapid and relatively inexpensive tool to improve the efficacy of molecular quantification methods in regards to viral infectivity.

  8. Molecularly imprinted polymer cartridges coupled to high performance liquid chromatography (HPLC-UV) for simple and rapid analysis of fenthion in olive oil.

    Science.gov (United States)

    Bakas, Idriss; Ben Oujji, Najwa; Istamboulié, Georges; Piletsky, Sergey; Piletska, Elena; Ait-Addi, Elhabib; Ait-Ichou, Ihya; Noguer, Thierry; Rouillon, Régis

    2014-07-01

    A combination of molecular modelling and a screening of the library of non-imprinted polymers (NIPs) was used to identify acrylamide as a functional monomer with high affinity towards fenthion, organophosphate insecticide, which is frequently used in the treatment of olives. A good correlation was found between the screening tests and modelling of monomer-template interactions performed using a computational approach. Acrylamide-based molecularly imprinted polymer (MIP) and non-imprinted polymer (NIP) were thermally synthesised in dimethyl formamide (porogen) using ethylene glycol dimethacrylate as a cross-linker and 1,1-azo-bis (isobutyronitrile) as an initiator. The chemical and physical properties of the prepared polymers were characterised. The binding of fenthion by the polymers was studied using solvents with different polarities. The developed MIP showed a high selectivity towards fenthion, compared to other organophosphates (dimethoate, methidathion malalthion), and allowed extraction of fenthion from olive oil samples with a recovery rate of about 96%. The extraction of fenthion using MIPs was much more effective than traditional C18 reverse-phase solid phase extraction and allowed to achieve a low detection limit (LOD) (5 µg L(-1)). Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Ultrahigh Molecular Weight Linear Block Copolymers: Rapid Access by Reversible-Deactivation Radical Polymerization and Self- Assembly into Large Domain Nanostructures

    Energy Technology Data Exchange (ETDEWEB)

    Mapas, Jose Kenneth D.; Thomay, Tim; Cartwright, Alexander N.; Ilavsky, Jan; Rzayev, Javid

    2016-05-05

    Block copolymer (BCP) derived periodic nanostructures with domain sizes larger than 150 nm present a versatile platform for the fabrication of photonic materials. So far, the access to such materials has been limited to highly synthetically involved protocols. Herein, we report a simple, “user-friendly” method for the preparation of ultrahigh molecular weight linear poly(solketal methacrylate-b-styrene) block copolymers by a combination of Cu-wire-mediated ATRP and RAFT polymerizations. The synthesized copolymers with molecular weights up to 1.6 million g/mol and moderate dispersities readily assemble into highly ordered cylindrical or lamella microstructures with domain sizes as large as 292 nm, as determined by ultra-small-angle x-ray scattering and scanning electron microscopy analyses. Solvent cast films of the synthesized block copolymers exhibit stop bands in the visible spectrum correlated to their domain spacings. The described method opens new avenues for facilitated fabrication and the advancement of fundamental understanding of BCP-derived photonic nanomaterials for a variety of applications.

  10. A one-step, triplex, real-time RT-PCR assay for the simultaneous detection of enterovirus 71, coxsackie A16 and pan-enterovirus in a single tube.

    Directory of Open Access Journals (Sweden)

    Shiyin Zhang

    Full Text Available The recent, ongoing epidemic of hand, foot, and mouth disease (HFMD, which is caused by enterovirus infection, has affected millions of children and resulted in thousands of deaths in China. Enterovirus 71 (EV71 and coxsackie A16 (CA16 are the two major distinct pathogens for HFMD. However, EV71 is more commonly associated with neurologic complications and even fatalities. Therefore, simultaneously detecting and differentiating EV71 and CA16 specifically from other enteroviruses for diagnosing HFMD is important. Here, we developed a one-step, triplex, real-time RT-PCR assay for the simultaneous detection of EV71, CA16, and pan-enterovirus (EVs in a single tube with an internal amplification control. The detection results for the serially diluted viruses indicate that the lower limit of detection for this assay is 0.001-0.04 TCID50/ml, 0.02 TCID50/ml, and 0.001 TCID50/ml for EVs, EV71, and CA16, respectively. After evaluating known HFMD virus stocks of 17 strains of 16 different serotypes, this assay showed a favorable detection spectrum and no obvious cross-reactivity. The results for 141 clinical throat swabs from HFMD-suspected patients demonstrated sensitivities of 98.4%, 98.7%, and 100% for EVs, EV71, and CA16, respectively, and 100% specificity for each virus. The application of this one-step, triplex, real-time RT-PCR assay in clinical units will contribute to HFMD surveillance and help to identify causative pathogen in patients with suspected HFMD.

  11. A one-step, triplex, real-time RT-PCR assay for the simultaneous detection of enterovirus 71, coxsackie A16 and pan-enterovirus in a single tube.

    Science.gov (United States)

    Zhang, Shiyin; Wang, Jin; Yan, Qiang; He, Shuizhen; Zhou, Wenbin; Ge, Shengxiang; Xia, Ningshao

    2014-01-01

    The recent, ongoing epidemic of hand, foot, and mouth disease (HFMD), which is caused by enterovirus infection, has affected millions of children and resulted in thousands of deaths in China. Enterovirus 71 (EV71) and coxsackie A16 (CA16) are the two major distinct pathogens for HFMD. However, EV71 is more commonly associated with neurologic complications and even fatalities. Therefore, simultaneously detecting and differentiating EV71 and CA16 specifically from other enteroviruses for diagnosing HFMD is important. Here, we developed a one-step, triplex, real-time RT-PCR assay for the simultaneous detection of EV71, CA16, and pan-enterovirus (EVs) in a single tube with an internal amplification control. The detection results for the serially diluted viruses indicate that the lower limit of detection for this assay is 0.001-0.04 TCID50/ml, 0.02 TCID50/ml, and 0.001 TCID50/ml for EVs, EV71, and CA16, respectively. After evaluating known HFMD virus stocks of 17 strains of 16 different serotypes, this assay showed a favorable detection spectrum and no obvious cross-reactivity. The results for 141 clinical throat swabs from HFMD-suspected patients demonstrated sensitivities of 98.4%, 98.7%, and 100% for EVs, EV71, and CA16, respectively, and 100% specificity for each virus. The application of this one-step, triplex, real-time RT-PCR assay in clinical units will contribute to HFMD surveillance and help to identify causative pathogen in patients with suspected HFMD.

  12. [Analysis on the change of genotype of enteroviruses associated hand, foot and mouth disease in Beijing during 2013 to 2014].

    Science.gov (United States)

    Song, Qinwei; Huang, Hui; Deng, Jie; Zhao, Linqing; Deng, Li; Sun, Yu; Wang, Fang; Oian, Yuan; Zhu, Runan

    2015-08-01

    To analyze the genotype, epidemic pattern and the characteristics of the disease of enteroviruses during the epidemic season of hand, foot and mouth disease (HMFD) in children from 2013 to 2014 in Beijing to provide the scientific evidence for prevention and treatment of HFMD. During April to September in 2013 and March to October in 2014, a total of 977 throat swabs were collected from children who visited the Children's Hospital Affiliated to Capital Institute of Pediatrics, including 147 from patients with HFMD in 2013, 343 with HFMD, 201 with atypical HFMD, 83 with herpangina, 25 with fever with convulsions, 64 fever with rash and 114 with rash in 2014. Enteroviruses universal type (EV), Enteroviruses type 71 (EV71) and Coxsackievirus group A 16 (CA16) were detected by real-time RT-PCR respectively. The nucleic acid of specimens which were identified with non-EV71, non-CA16 was tested by nested PCR and analyzed by VP1 sequencing. The detection rate and epidemic pattern of different genotypes of enterovirus were analyzed among different age groups and between 2013 and 2014. Of 977 throat swabs, 80. 1% samples were detected positive for enteroviruses. The positive rates of CA16, EV71, CA6, CA10, CA4 and other EVs were 25. 6% (250/977), 18. 9% (185/977), 20. 0% (195/977), 5. 0% (49/977), 1.5% (15/977) and 9.1% (89/977), respectively. Twenty six of the 89 other EVs included CA2, CA5, CA8, CA9, CA12, CA14, CB2, CB5, E6, E9 and E25, each genotype of which was no more than 3. The nucleotide homologies shared among CA6, CA10 and CA4 strains between 2013 and 2014 were 94. 3% - 100%, 93. 8% - 99. 1% and 92.7% - 99. 8%, respectively. The positive rates of ≤1 year group were 71. 1% (106/149), which was lower than that of other age groups (all P 5 year group (χ2 =1. 181,P = 0. 277). In 2013, the positive rate of EV was 85. 7% (126/147) and the predominant genotype was CA6 54. 8% (69/126), followed by CA16 20. 6% (26/126) and EV71 11. 9% (15/126). In 2014, the positive

  13. [Association between S100B gene polymorphisms and hand, foot and mouth disease caused by enterovirus 71 infection].

    Science.gov (United States)

    Li, Jing; Shan, Ruo-Bing; Liu, Rui-Hai; Xu, Ying-Jun; Qu, Ni-Yan; Pan, Gui-Mei; Zhang, Na; Yang, Na; Chen, Zhen-Zhen; Zhang, Wen-Xiang; Li, Zi-Pu

    2017-08-01

    To investigate the association between rs9722 polymorphisms in the S100B gene and hand, foot and mouth disease (HFMD) caused by enterovirus 71. A total of 124 HFMD children with enterovirus 71 infection were enrolled as subjects, and 56 healthy children were enrolled as control group. The rs9722 polymorphisms in the S100B gene were detected for both groups, and the serum level of S100B protein was measured for 74 HFMD children. The rs9722 locus of the S100B gene had three genotypes, CC, CT, and TT, and the genotype frequencies were in accordance with Hardy-Weinberg equilibrium. Compared with the control group, the HFMD group had significant increases in the frequencies of TT genotype and T allele (Penterovirus 71 infection had significantly higher frequencies of TT genotype and T allele than those with moderate or mild HFMD (Penterovirus 71 infection.

  14. Development of a One-Step Probe Based Molecular Assay for Rapid Immunodiagnosis of Infection with M. tuberculosis Using Dried Blood Spots

    DEFF Research Database (Denmark)

    Blauenfeldt, Thomas; Heyckendorf, Jan; Graff Jensen, Sidse

    2014-01-01

    BACKGROUND: Antigen specific release of IP-10 is the most promising alternative marker to IFN-γ for infection with M. tuberculosis. Compared to Interferon-γ release assays (IGRA), IP-10 is released in high levels enabling novel approaches such as field friendly dried blood spots (DBS) and molecular...... was validated and applied for the diagnosis of M. tuberculosis infection in DBS samples from 43 patients with confirmed TB, 13 patients with latent TB and 96 presumed uninfected controls. In parallel, IP-10 and INF-γ levels were measured in Quantiferon (QFT-TB) plasma supernatants. RESULTS: IP-10 m......RNA upregulation was detectable at 4 hours after stimulation (6 fold upregulation) peaking at 8 hours (108 fold upregulation). IFN-γ expression occurred in concert but levels were lower (peak 6.7 fold upregulation). IP-10 gene expression level was significantly higher in patients with tuberculosis (median 31...

  15. A Novel, Broad-Spectrum Inhibitor of Enterovirus Replication That Targets Host Cell Factor Phosphatidylinositol 4-Kinase IIIβ

    Science.gov (United States)

    van der Schaar, Hilde M.; Leyssen, Pieter; Thibaut, Hendrik J.; de Palma, Armando; van der Linden, Lonneke; Lanke, Kjerstin H. W.; Lacroix, Céline; Verbeken, Erik; Conrath, Katja; MacLeod, Angus M.; Mitchell, Dale R.; Palmer, Nicholas J.; van de Poël, Hervé; Andrews, Martin

    2013-01-01

    Despite their high clinical and socioeconomic impacts, there is currently no approved antiviral therapy for the prophylaxis or treatment of enterovirus infections. Here we report on a novel inhibitor of enterovirus replication, compound 1, 2-fluoro-4-(2-methyl-8-(3-(methylsulfonyl)benzylamino)imidazo[1,2-a]pyrazin-3-yl)phenol. This compound exhibited a broad spectrum of antiviral activity, as it inhibited all tested species of enteroviruses and rhinoviruses, with 50% effective concentrations ranging between 4 and 71 nM. After a lengthy resistance selection process, coxsackievirus mutants resistant to compound 1 were isolated that carried substitutions in their 3A protein. Remarkably, the same substitutions were recently shown to provide resistance to inhibitors of phosphatidylinositol 4-kinase IIIβ (PI4KIIIβ), a lipid kinase that is essential for enterovirus replication, suggesting that compound 1 may also target this host factor. Accordingly, compound 1 directly inhibited PI4KIIIβ in an in vitro kinase activity assay. Furthermore, the compound strongly reduced the PI 4-phosphate levels of the Golgi complex in cells. Rescue of coxsackievirus replication in the presence of compound 1 by a mutant PI4KIIIβ carrying a substitution in its ATP-binding pocket revealed that the compound directly binds the kinase at this site. Finally, we determined that an analogue of compound 1, 3-(3-fluoro-4-methoxyphenyl)-2-methyl-N-(pyridin-4-ylmethyl)imidazo[1,2-a]pyrazin-8-amine, is well tolerated in mice and has a dose-dependent protective activity in a coxsackievirus serotype B4-induced pancreatitis model. PMID:23896472

  16. Comparative study of the cytokine/chemokine response in children with differing disease severity in enterovirus 71-induced hand, foot, and mouth disease.

    Directory of Open Access Journals (Sweden)

    Yan Zhang

    Full Text Available BACKGROUND: Enterovirus 71 (EV71 infection can lead to a rapidly progressing, life-threatening, and severe neurological disease in young children, including the development of human hand, foot, and mouth disease (HFMD. This study aims to further characterize the specific immunological features in EV71-mediated HFMD patients presenting with differing degrees of disease severity. METHODOLOGY: Comprehensive cytokine and chemokine expression were broadly evaluated by cytokine antibody array in EV71-infected patients hospitalized for HFMD compared to Coxsackievirus A16-infected patients and age-matched healthy controls. More detailed analysis using Luminex-based cytokine bead array was performed in EV71-infected patients stratified into diverse clinic outcomes. Additionally, immune cell frequencies in peripheral blood and EV71-specific antibodies in plasma were also examined. PRINCIPAL FINDINGS: Expression of several cytokines and chemokines were significantly increased in plasma from EV71-infected patients compared to healthy controls, which further indicated that: (1 GM-CSF, MIP-1β, IL-2, IL-33, and IL-23 secretion was elevated in patients who rapidly developed disease and presented with uncomplicated neurological damage; (2 G-CSF and MCP-1 were distinguishably secreted in EV71 infected very severe patients presenting with acute respiratory failure; (3 IP-10, MCP-1, IL-6, IL-8, and G-CSF levels were much higher in cerebrospinal fluid than in plasma from patients with neurological damage; (4 FACS analysis revealed that the frequency of CD19(+HLADR(+ mature B cells dynamically changed over time during the course of hospitalization and was accompanied by dramatically increased EV71-specific antibodies. Our data provide a panoramic view of specific immune mediator and cellular immune responses of HFMD and may provide useful immunological profiles for monitoring the progress of EV71-induced fatal neurological symptoms with acute respiratory failure.

  17. Coexistence of two clades of enterovirus D68 in pediatric Swedish patients in the summer and fall of 2014.

    Science.gov (United States)

    Dyrdak, Robert; Rotzén-Östlund, Maria; Samuelson, Agneta; Eriksson, Margareta; Albert, Jan

    2015-01-01

    In 2014, an outbreak of enterovirus D68 (EV-D68) was observed in North America, with cases of severe respiratory illness and a possible etiological link to cases of acute flaccid paralysis. EV-D68 has also been reported from European countries, but no data from Sweden are available. This study investigated respiratory specimens collected during July-October 2014 from 30 Swedish children aged 0-9 years who were positive for enterovirus and/or rhinovirus in routine clinical PCR. Seven samples were typed as EV-D68 by VP4/VP2 sequencing. Two genetically distinct EV-D68 variants coexisted. Six viruses belonged to clade B, the variant involved in the North American outbreak, and one virus belonged to clade A. Respiratory illness was the major symptom among EV-D68 infected patients and all fully recovered. This is the first report of EV-D68 in Sweden. Considering the current epidemiological situation, genotyping and specific EV-D68 testing should be considered in patients with severe respiratory illness who test positive for enterovirus or rhinovirus in routine diagnostics.

  18. Characterization of enterovirus 71 and coxsackievirus A16 isolated in hand, foot, and mouth disease patients in Guangdong, 2010.

    Science.gov (United States)

    He, Si-Jie; Han, Jian-Feng; Ding, Xi-Xia; Wang, Ya-Di; Qin, Cheng-Feng

    2013-11-01

    Hand, foot, and mouth disease (HFMD) is an acute viral disease caused by human enteroviruses, especially human enterovirus 71 (HEV71) and coxsackievirus A16 (CVA16), and mainly affects infants and young children. After the outbreak in 2008 in Fuyang, China, HFMD was classified as a category C notifiable infectious disease by the Ministry of Health of China. In this study, we report the epidemiologic and clinical manifestations of HFMD in Guangdong Province, China in 2010, and characterize HEV71 and CVA16 isolated from clinical specimens. Among the 542 HFMD patients, 495 (91.3%) were positive for enterovirus as detected by real-time reverse transcriptase PCR; 243 were positive for HEV71 (49.1%, 243/495) and 114 were positive for CVA16 (23.0%, 114/495). Most of the affected children were aged 5 years or under (93.7%, 508/542). Phylogenetic analyses of VP1 gene sequences showed that the HEV71 isolates belonged to C4a subgenotype, and CVA16 isolates belonged to B1 genotype. Our results demonstrate that HEV71 and CVA16 are the primary causative agents responsible for HFMD in Guangdong Province, and their co-circulation poses a potential risk to public health. Copyright © 2013 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  19. Epidemiological and genetic analysis concerning the non-enterovirus 71 and non-coxsackievirus A16 causative agents related to hand, foot and mouth disease in Anyang city, Henan Province, China, from 2011 to 2015.

    Science.gov (United States)

    Li, Yang; Bao, Honghong; Zhang, Xiangping; Zhai, Mingqiang; Bao, Xiaobing; Wang, Demin; Zhang, Shuanhu

    2017-10-01

    Enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16) are major pathogens of hand, foot, and mouth disease (HFMD) and have been associated with consecutive outbreaks of HFMD in China over the past years. Although several other human enteroviruses (HEVs) have also acted as causative agents of HFMD, published information on their roles in the prevalence of HFMD is limited. This study was conducted to reveal the characteristics of the pathogenic spectrum and molecular epidemiology of the non-EV-71 and -CV-A16 HEVs in Anyang City, which is located in north-central China and has a population of five million. From 2011 to 2015, 2270 samples were collected from HFMD patients (3.89 ± 1.06 years of age), and 1863 HEV-positive samples, including 524 samples with 23 non-EV-71 and non-CV-A16 serotypes, were identified. Based on the nucleotide sequence of the VP1 gene, 6 common non-EV-71 and non-CV-A16 HEVs, including coxsackievirus A2, A6, A10, A14, B2, and B5, were studied to determine their phylogenies and selective pressures. Phylogenetic analyses revealed a high level of genetic divergence and a pattern of lineage replacement over time in Mainland China. Selective pressure analyses showed that purifying selection was predominant in the evolution of the VP1 gene, whereas positive selection acted on individual codons. Overall, non-EV-71 and non-CV-A16 HEVs were important constituents of the pathogenic spectrum of HFMD in Anyang City during 2011-2015. Some of these HEVs with complex and active phylogenies represent a potential threat to public health, suggesting that long-term monitoring of these pathogens should be implemented to prevent HFMD outbreaks. © 2017 Wiley Periodicals, Inc.

  20. Discussion on the structural origins of the fracture toughness and hardness changes in rapidly quenched borosilicate glasses: a molecular dynamics study

    International Nuclear Information System (INIS)

    Kieu, L.H.; Kilymis, D.; Delaye, J.M.; Peuget, S.

    2014-01-01

    Understanding radiation effects in nuclear glasses is important in order to assess their long term behaviour in a geological repository. In this paper, the results obtained by classical molecular dynamics about the mechanical properties of a simplified SiO 2 - B 2 O 3 - Na 2 O glass with the same molar proportions as in the real French nuclear glass were reviewed. A pristine and a disordered structure, representative of the irradiated one, were prepared in order to address the question of the radiation effects on the fracture toughness and hardness. In a first step, fracture simulations were performed to compare the atomistic mechanisms in the pristine and disordered glasses. In a second step, nano indentation was simulated using a diamond tip and again the pristine and disordered glass behaviors were compared. The depolymerisation associated to the glass disordering, in particular the increase of the tri-coordinated boron concentration relative to the tetra-coordinated boron concentration improves the fracture resistance by favouring the plastic flows. On the contrary, the swelling of the disordered glass limits the fracture resistance by facilitating the cavity formation. The first effect is dominant in the case of the glass investigated here because, experimentally, the fracture toughness increases after irradiation. Concerning the hardness, both the decrease of the boron coordination in parallel with non-bridging oxygen formation and free volume accumulation lead to a hardness decrease because the more disordered and open structure facilitates the tip penetration. (authors)

  1. Enterovirus Exposure Uniquely Discriminates Type 1 Diabetes Patients with a Homozygous from a Heterozygous Melanoma Differentiation-Associated Protein 5/Interferon Induced with Helicase C Domain 1 A946T Genotype

    NARCIS (Netherlands)

    Schulte, B.M.; Gielen, P.R.; Kers-Rebel, E.D.; Prosser, A.C.; Lind, K.; Flodstrom-Tullberg, M.; Tack, C.J.J.; Elving, L.D.; Adema, G.J.

    2016-01-01

    In children at risk for type 1 diabetes, innate immune activity is detected before seroconversion. Enterovirus infections have been linked to diabetes development, and a polymorphism (A946T) in the innate immune sensor recognizing enterovirus RNA, interferon-induced with helicase C domain 1/melanoma

  2. Development and Evaluation of a Molecular Diagnostic Method for Rapid Detection of Histoplasma capsulatum var. farciminosum, the Causative Agent of Epizootic Lymphangitis, in Equine Clinical Samples.

    Science.gov (United States)

    Scantlebury, C E; Pinchbeck, G L; Loughnane, P; Aklilu, N; Ashine, T; Stringer, A P; Gordon, L; Marshall, M; Christley, R M; McCarthy, A J

    2016-12-01

    Histoplasma capsulatum var. farciminosum, the causative agent of epizootic lymphangitis (EZL), is endemic in parts of Africa. Diagnosis based on clinical signs and microscopy lacks specificity and is a barrier to further understanding this neglected disease. Here, a nested PCR method targeting the internal transcribed spacer (ITS) region of the rRNA operon was validated for application to equine clinical samples. Twenty-nine horses with signs of EZL from different climatic regions of Ethiopia were clinically examined. Blood samples and aspirates of pus from cutaneous nodules were taken, along with blood from a further 20 horses with no cutaneous EZL lesions. Among the 29 horses with suspected cases of EZL, H. capsulatum var. farciminosum was confirmed by extraction of DNA from pus and blood samples from 25 and 17 horses, respectively. Positive PCR results were also obtained with heat-inactivated pus (24 horses) and blood (23 horses) spotted onto Whatman FTA cards. Two positive results were obtained among blood samples from 20 horses that did not exhibit clinical signs of EZL. These are the first reports of the direct detection of H. capsulatum var. farciminosum in equine blood and at high frequency among horses exhibiting cutaneous lesions. The nested PCR outperformed conventional microscopic diagnosis, as characteristic yeast cells could be observed only in 14 pus samples. The presence of H. capsulatum var. farciminosum DNA was confirmed by sequencing the cloned PCR products, and while alignment of the ITS amplicons showed very little sequence variation, there was preliminary single nucleotide polymorphism-based evidence for the existence of two subgroups of H. capsulatum var. farciminosum This molecular diagnostic method now permits investigation of the epidemiology of EZL. Copyright © 2016 Scantlebury et al.

  3. Development and Evaluation of a Molecular Diagnostic Method for Rapid Detection of Histoplasma capsulatum var. farciminosum, the Causative Agent of Epizootic Lymphangitis, in Equine Clinical Samples

    Science.gov (United States)

    Pinchbeck, G. L.; Loughnane, P.; Aklilu, N.; Ashine, T.; Stringer, A. P.; Gordon, L.; Marshall, M.; Christley, R. M.

    2016-01-01

    Histoplasma capsulatum var. farciminosum, the causative agent of epizootic lymphangitis (EZL), is endemic in parts of Africa. Diagnosis based on clinical signs and microscopy lacks specificity and is a barrier to further understanding this neglected disease. Here, a nested PCR method targeting the internal transcribed spacer (ITS) region of the rRNA operon was validated for application to equine clinical samples. Twenty-nine horses with signs of EZL from different climatic regions of Ethiopia were clinically examined. Blood samples and aspirates of pus from cutaneous nodules were taken, along with blood from a further 20 horses with no cutaneous EZL lesions. Among the 29 horses with suspected cases of EZL, H. capsulatum var. farciminosum was confirmed by extraction of DNA from pus and blood samples from 25 and 17 horses, respectively. Positive PCR results were also obtained with heat-inactivated pus (24 horses) and blood (23 horses) spotted onto Whatman FTA cards. Two positive results were obtained among blood samples from 20 horses that did not exhibit clinical signs of EZL. These are the first reports of the direct detection of H. capsulatum var. farciminosum in equine blood and at high frequency among horses exhibiting cutaneous lesions. The nested PCR outperformed conventional microscopic diagnosis, as characteristic yeast cells could be observed only in 14 pus samples. The presence of H. capsulatum var. farciminosum DNA was confirmed by sequencing the cloned PCR products, and while alignment of the ITS amplicons showed very little sequence variation, there was preliminary single nucleotide polymorphism-based evidence for the existence of two subgroups of H. capsulatum var. farciminosum. This molecular diagnostic method now permits investigation of the epidemiology of EZL. PMID:27707938

  4. Detection, quantitation and identification of enteroviruses from surface waters and sponge tissue from the Florida Keys using real-time RT-PCR

    Science.gov (United States)

    Donaldson, K.A.; Griffin, Dale W.; Paul, J.H.

    2002-01-01

    A method was developed for the quantitative detection of pathogenic human enteroviruses from surface waters in the Florida Keys using Taqman (R) one-step Reverse transcription (RT)-PCR with the Model 7700 ABI Prism (R) Sequence Detection System. Viruses were directly extracted from unconcentrated grab samples of seawater, from seawater concentrated by vortex flow filtration using a 100kD filter and from sponge tissue. Total RNA was extracted from the samples, purified and concentrated using spin-column chromatography. A 192-196 base pair portion of the 5??? untranscribed region was amplified from these extracts. Enterovirus concentrations were estimated using real-time RT-PCR technology. Nine of 15 sample sites or 60% were positive for the presence of pathogenic human enteroviruses. Considering only near-shore sites, 69% were positive with viral concentrations ranging from 9.3viruses/ml to 83viruses/g of sponge tissue (uncorrected for extraction efficiency). Certain amplicons were selected for cloning and sequencing for identification. Three strains of waterborne enteroviruses were identified as Coxsackievirus A9, Coxsackievirus A16, and Poliovirus Sabin type 1. Time and cost efficiency of this one-step real-time RT-PCR methodology makes this an ideal technique to detect, quantitate and identify pathogenic enteroviruses in recreational waters. Copyright ?? 2002 Elsevier Science Ltd.

  5. Dasatinib rapidly induces deep molecular response in chronic-phase chronic myeloid leukemia patients who achieved major molecular response with detectable levels of BCR-ABL1 transcripts by imatinib therapy.

    Science.gov (United States)

    Shiseki, Masayuki; Yoshida, Chikashi; Takezako, Naoki; Ohwada, Akira; Kumagai, Takashi; Nishiwaki, Kaichi; Horikoshi, Akira; Fukuda, Tetsuya; Takano, Hina; Kouzai, Yasuji; Tanaka, Junji; Morita, Satoshi; Sakamoto, Junichi; Sakamaki, Hisashi; Inokuchi, Koiti

    2017-10-01

    With the introduction of imatinib, a first-generation tyrosine kinase inhibitor (TKI) to inhibit BCR-ABL1 kinase, the outcome of chronic-phase chronic myeloid leukemia (CP-CML) has improved dramatically. However, only a small proportion of CP-CML patients subsequently achieve a deep molecular response (DMR) with imatinib. Dasatinib, a second-generation TKI, is more potent than imatinib in the inhibition of BCR-ABL1 tyrosine kinase in vitro and more effective in CP-CML patients who do not achieve an optimal response with imatinib treatment. In the present study, we attempted to investigate whether switching the treatment from imatinib to dasatinib can induce DMR in 16 CP-CML patients treated with imatinib for at least two years who achieved a major molecular response (MMR) with detectable levels of BCR-ABL1 transcripts. The rates of achievement of DMR at 1, 3, 6 and 12 months after switching to dasatinib treatment in the 16 patients were 44% (7/16), 56% (9/16), 63% (10/16) and 75% (12/16), respectively. The cumulative rate of achieving DMR at 12 months from initiation of dasatinib therapy was 93.8% (15/16). The proportion of natural killer cells and cytotoxic T cells in peripheral lymphocytes increased after switching to dasatinib. In contrast, the proportion of regulatory T cells decreased during treatment. The safety profile of dasatinib was consistent with previous studies. Switching to dasatinib would be a therapeutic option for CP-CML patients who achieved MMR but not DMR by imatinib, especially for patients who wish to discontinue TKI therapy.

  6. Comparison of enterovirus detection in cerebrospinal fluid with Bacterial Meningitis Score in children.

    Science.gov (United States)

    Pires, Frederico Ribeiro; Franco, Andréia Christine Bonotto Farias; Gilio, Alfredo Elias; Troster, Eduardo Juan

    2017-01-01

    To measure the role of enterovirus detection in cerebrospinal fluid compared with the Bacterial Meningitis Score in children with meningitis. A retrospective cohort based on analysis of medical records of pediatric patients diagnosed as meningitis, seen at a private and tertiary hospital in São Paulo, Brazil, between 2011 and 2014. Excluded were patients with critical illness, purpura, ventricular shunt or recent neurosurgery, immunosuppression, concomitant bacterial infection requiring parenteral antibiotic therapy, and those who received antibiotics 72 hours before lumbar puncture. The study included 503 patients. Sixty-four patients were excluded and 94 were not submitted to all tests for analysis. Of the remaining 345 patients, 7 were in the Bacterial Meningitis Group and 338 in the Aseptic Meningitis Group. There was no statistical difference between the groups. In the Bacterial Meningitis Score analysis, of the 338 patients with possible aseptic meningitis (negative cultures), 121 of them had one or more points in the Bacterial Meningitis Score, with sensitivity of 100%, specificity of 64.2%, and negative predictive value of 100%. Of the 121 patients with positive Bacterial Meningitis Score, 71% (86 patients) had a positive enterovirus detection in cerebrospinal fluid. Enterovirus detection in cerebrospinal fluid was effective to differentiate bacterial from viral meningitis. When the test was analyzed together with the Bacterial Meningitis Score, specificity was higher when compared to Bacterial Meningitis Score alone. Avaliar o papel da pesquisa de enterovírus no líquido cefalorraquidiano em comparação com o Escore de Meningite Bacteriana em crianças com meningite. Coorte retrospectiva, realizada pela análise de prontuários, incluindo pacientes pediátricos, com diagnóstico de meningite e atendidos em um hospital privado e terciário, localizado em São Paulo, entre 2011 e 2014. Foram excluídos os pacientes com doença crítica, púrpura, deriva

  7. Magnetic molecularly imprinted polymers based on silica modified by deep eutectic solvents for the rapid simultaneous magnetic-based solid-phase extraction of Salvia miltiorrhiza bunge, Glycine max (Linn.) Merr and green tea.

    Science.gov (United States)

    Li, Guizhen; Wang, Xiaoqin; Row, Kyung Ho

    2018-04-01

    Novel magnetic molecularly imprinted polymers (MMIPs) with multiple-template based on silica were modified by four types of deep eutectic solvents (DESs) for the rapid simultaneous magnetic solid-phase extraction (MSPE) of tanshinone Ⅰ, tanshinone ⅡA, and cryptotanshinone from Salvia miltiorrhiza bunge; glycitein, genistein, and daidzein from Glycine max (Linn.) Merr; and epicatechin, epigallocatechin gallate, and epicatechin gallate from green tea, respectively. The synthesized materials were characterized by Fourier transform infrared spectroscopy and field emission scanning electron microscopy. Single factor experiments were to explore the relationship between the extraction efficiency and four factors (the sample solution pH, amount of DESs for modification, amount of adsorbent, and extraction time). It was showed that the DES4-MMIPs have better extraction ability than the MMIPs without DESs and the other three DESs-modified MMIPs. The best extraction recoveries with DES4-MMIP were tanshinone Ⅰ (85.57%), tanshinone ⅡA (80.58%), cryptotanshinone (92.12%), glycitein (81.65%), genistein (87.72%), daidzein (92.24%), epicatechin (86.43%), epigallocatechin gallate (80.92%), and epicatechin gallate (93.64%), respectively. The novel multiple-template MMIPs materials modified by DES for the rapid simultaneous MSPE of active compounds were proved to reduce the experimental steps than single-template technique, and increase the extraction efficiency. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Inhibition of enterovirus 71 infection by antisense octaguanidinium dendrimer-conjugated morpholino oligomers.

    Science.gov (United States)

    Tan, Chee Wah; Chan, Yoke Fun; Quah, Yi Wan; Poh, Chit Laa

    2014-07-01

    Enterovirus 71 (EV-71) infections are generally manifested as mild hand, foot and mouth disease, but have been reported to cause severe neurological complications with high mortality rates. Treatment options remain limited due to the lack of antivirals. Octaguanidinium-conjugated morpholino oligomers (vivo-MOs) are single-stranded DNA-like antisense agents that can readily penetrate cells and reduce gene expression by steric blocking of complementary RNA sequences. In this study, inhibitory effects of three vivo-MOs that are complementary to the EV-71 internal ribosome entry site (IRES) and the RNA-dependent RNA polymerase (RdRP) were tested in RD cells. Vivo-MO-1 and vivo-MO-2 targeting the EV-71 IRES showed significant viral plaque reductions of 2.5 and 3.5 log10PFU/ml, respectively. Both vivo-MOs reduced viral RNA copies and viral capsid expression in RD cells in a dose-dependent manner. In contrast, vivo-MO-3 targeting the EV-71 RdRP exhibited less antiviral activity. Both vivo-MO-1 and 2 remained active when administered either 4h before or within 6h after EV-71 infection. Vivo-MO-2 exhibited antiviral activities against poliovirus (PV) and coxsackievirus A16 but vivo-MO-1 showed no antiviral activities against PV. Both the IRES-targeting vivo-MO-1 and vivo-MO-2 inhibit EV-71 RNA translation. Resistant mutants arose after serial passages in the presence of vivo-MO-1, but none were isolated against vivo-MO-2. A single T to C substitution at nucleotide position 533 was sufficient to confer resistance to vivo-MO-1. Our findings suggest that IRES-targeting vivo-MOs are good antiviral candidates for treating early EV-71 infection, and vivo-MO-2 is a more favorable candidate with broader antiviral spectrum against enteroviruses and are refractory to antiviral resistance. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Risk Factors for Enterovirus A71 Seropositivity in Rural Indigenous Populations in West Malaysia.

    Science.gov (United States)

    NikNadia, Nmn; Sam, I-Ching; Khaidir, Nasibah; Ngui, Romano; Lim, Yvonne A L; Goh, Xiang Ting; Choy, Seow Huey; Chan, Yoke Fun

    2016-01-01

    Enterovirus A71 (EV-A71), which is transmitted by the fecal-oral route, causes hand, foot and mouth disease and, rarely, severe neurological complications. In Malaysia, the indigenous rural community (Orang Asli) has a high prevalence of parasitic diseases due to poor sanitation, water supply and hygiene practices. This cross-sectional study compared the seroepidemiology of EV-A71 among rural Orang Asli and urban Kuala Lumpur populations in West Malaysia, and determined the risk factors associated with EV-A71 seropositivity in rural Orang Asli. Seropositive rates were determined by neutralization assay. EV-A71 seropositivity was strongly associated with increasing age in both populations. Rural Orang Asli children ≤12 years had significantly higher EV-A71 seropositivity rates than urban Kuala Lumpur children (95.5% vs 57.6%, P water (adjusted OR 6.2, 95% CI 2.3-16.6, P water may reduce the risk of EV-A71 infection. With significantly higher EV-A71 seropositive rates, younger rural children should be a priority target for future vaccination programs in Malaysia.

  10. The clinical correlation of regulatory T cells and cyclic adenosine monophosphate in enterovirus 71 infection.

    Directory of Open Access Journals (Sweden)

    Shih-Min Wang

    Full Text Available Brainstem encephalitis (BE and pulmonary edema (PE are notable complications of enterovirus 71 (EV71 infection.This study investigated the immunoregulatory characterizations of EV71 neurological complications by disease severity and milrinone treatment.Patients <18 years with virologically confirmed EV71 infections were enrolled and divided into 2 groups: the hand, foot, and mouth disease (HFMD or BE group, and the autonomic nervous system (ANS dysregulation or PE group. Cytokine and cyclic adenosine monophosphate (cAMP levels, and the regulatory T cell (Tregs profiles of the patients were determined.Patients with ANS dysregulation or PE exhibited significantly low frequency of CD4(+CD25(+Foxp3+ and CD4(+Foxp3(+ T cells compared with patients with HFMD or BE. The expression frequency of CD4-CD8- was also significantly decreased in patients with ANS dysregulation or PE. Among patients with ANS dysregulation or PE, the expression frequency of CD4+Foxp3+ increased markedly after milrinone treatment, and was associated with reduction of plasma levels IL-6, IL-8 and IL-10. Plasma concentrations of cAMP were significantly decreased in patients with ANS dysregulation or PE compared with patients with HFMD or BE; however, cAMP levels increased after milrinone treatment.These findings suggested decreased different regulatory T populations and cAMP expression correlate with increased EV71 disease severity. Improved outcome after milrinone treatment may associate with increased regulatory T populations, cAMP expression and modulation of cytokines levels.

  11. Antiviral Ability of Kalanchoe gracilis Leaf Extract against Enterovirus 71 and Coxsackievirus A16

    Directory of Open Access Journals (Sweden)

    Ching-Ying Wang

    2012-01-01

    Full Text Available Pandemic infection or reemergence of Enterovirus 71 (EV71 and coxsackievirus A16 (CVA16 occurs in tropical and subtropical regions, being associated with hand-foot-and-mouth disease, herpangina, aseptic meningitis, brain stem encephalitis, pulmonary edema, and paralysis. However, effective therapeutic drugs against EV71 and CVA16 are rare. Kalanchoe gracilis (L. DC is used for the treatment of injuries, pain, and inflammation. This study investigated antiviral effects of K. gracilis leaf extract on EV71 and CVA16 replications. HPLC analysis with a C-18 reverse phase column showed fingerprint profiles of K. gracilis leaf extract had 15 chromatographic peaks. UV/vis absorption spectra revealed peaks 5, 12, and 15 as ferulic acid, quercetin, and kaempferol, respectively. K. gracilis leaf extract showed little cytotoxicity, but exhibited concentration-dependent antiviral activities including cytopathic effect, plaque, and virus yield reductions. K. gracilis leaf extract was shown to be more potent in antiviral activity than ferulic acid, quercetin, and kaempferol, significantly inhibiting in vitro replication of EV71 (IC50=35.88 μg/mL and CVA16 (IC50=42.91 μg/mL. Moreover, K. gracilis leaf extract is a safe antienteroviral agent with the inactivation of viral 2A protease and reduction of IL-6 and RANTES expressions.

  12. Enterovirus type 71 2A protease functions as a transcriptional activator in yeast

    Directory of Open Access Journals (Sweden)

    Lai Meng-Jiun

    2010-08-01

    Full Text Available Abstract Enterovirus type 71 (EV71 2A protease exhibited strong transcriptional activity in yeast cells. The transcriptional activity of 2A protease was independent of its protease activity. EV71 2A protease retained its transcriptional activity after truncation of 40 amino acids at the N-terminus but lost this activity after truncation of 60 amino acids at the N-terminus or deletion of 20 amino acids at the C-terminus. Thus, the acidic domain at the C-terminus of this protein is essential for its transcriptional activity. Indeed, deletion of amino acids from 146 to 149 (EAME in this acidic domain lost the transcriptional activity of EV71 2A protein though still retained its protease activity. EV71 2A protease was detected both in the cytoplasm and nucleus using confocal microscopy analysis. Coxsackie virus B3 2A protease also exhibited transcriptional activity in yeast cells. As expected, an acidic domain in the C-terminus of Coxsackie virus B3 2A protease was also identified. Truncation of this acidic domain resulted in the loss of transcriptional activity. Interestingly, this acidic region of poliovirus 2A protease is critical for viral RNA replication. The transcriptional activity of the EV71 or Coxsackie virus B3 2A protease should play a role in viral replication and/or pathogenesis.

  13. MRI findings of neurologic complications in the enterovirus 71-infected hand-foot-mouth disease

    International Nuclear Information System (INIS)

    Chen Feng; Li Jianjun; Liu Tao; Xiang Wei; Wen Guoqiang

    2010-01-01

    Objective: To explore the imaging characteristics of neurologic complications associated with the enterovirus 71 (EV71) epidemic by analyzing 25 cases and reviewing the literature. Methods: Twenty-five cases of hand-foot-mouth disease with neurologic complications during the recent EV71 outbreaks of Hainan province were studied for the clinical features and imaging findings, and literature were reviewed. Results: In 5 cases, acute flaccid paralysis associated with EV71-infected hand-foot-mouth disease was related to the linear high signal in the spinal cord on sagittal images. Two cases showed symmetrical, well- defined hyperintense lesions in the spinal cord on T 2 WI transverse. Strong enhancement of the ventral horns and root was seen on the contrast-enhanced axial T 1 WI. In brainstem encephalitis, all lesions presented with significant hyperintensity on T 2 WI and hypointense on T 1 WI in the posterior portions of the medulla oblongata, midbrain, and pons. The manifestations of aseptic meningitis (AM) on MRI have no characteristics, but subdural effusion, meningeal enhancement and hydrocephalus can be the indirect signs of AM. Conclusions: MRI is an effective method to investigate neurologic complications associated with the EV71 epidemic. Posterior portions of the medulla oblongata and pons, bilateral ventral horns of spinal involvement are characteristic findings of enteroviral encephalomyelitis. (authors)

  14. Identification of a Novel Enterovirus Species in Rhesus Macaque in China.

    Science.gov (United States)

    Ao, Yuan-Yun; Yu, Jie-Mei; Zhang, Cui-Yuan; Xin, Yun-Yun; Li, Li-Li; Duan, Zhao-Jun

    2016-06-22

    Recent studies of Enterovirus (EV) in nonhuman primates (NHPs), which could act as a source of future emerging human viral diseases, have boosted interest in the search for novel EVs. Here, a highly divergent strain of EV, tentatively named SEV-gx, was identified by viral metagenomic analysis from stool samples of rhesus macaques in China. In total, 27 of 280 (9.6%) faecal samples from rhesus macaques were positive for SEV-gx. Its complete genomic sequence is 7,367 nucleotide (nt). Genomic analyses showed that it has a standard genomic organisation for EVs, being more closely related to EV-J strains (approximately 54.0%, 43.0-44.1%, 52.3-55.2%, 61.1-62.7% and 64.0% amino acids identity in polyprotein, P1, P2 and P3 and combined 2C/3CD regions, respectively). It was also shown to have genome characteristics typical of EVs. Phylogenetic analysis of P1, 2C and 3CD aa indicated that SEV-gx can be classified as a distinct cluster in the EVs. All of this evidence demonstrates SEV-gx is a novel species (tentatively named EV-K) in the EV genus, which contributes to our understanding of the genetic diversity and evolution of EVs. Further studies are needed to investigate the potential pathogenicity of SEV-gx in NHPs and humans.

  15. Enterovirus infections in hospitals of Ile de France region over 2013.

    Science.gov (United States)

    Molet, Lucie; Saloum, Kenda; Marque-Juillet, Stéphanie; Garbarg-Chenon, Antoine; Henquell, Cécile; Schuffenecker, Isabelle; Peigue-Lafeuille, Hélène; Rozenberg, Flore; Mirand, Audrey

    2016-01-01

    The monitoring and genotyping of Enterovirus (EV) infections can help to associate particular or severe clinical manifestations with specific EV types and to identify the aetiology of infectious outbreaks. To describe the epidemiological features of EV infections diagnosed during the year 2013 in the Greater Paris area (Ile de France). During 2013, 2497 samples taken from 470 patients in 33 hospitals of Ile-de France were tested for EV genome by RT-PCR. EV genotyping was performed by the National Reference Centre (NRC) laboratories. EV infections were retrospectively reviewed by retrieving clinical and genotyping data from the NRC database. Of the 2497 samples, 490 (19.6%) was positive for EV genome detection. These EV infections represented 88.7% and 24.1%, respectively, of all reported regional and national infections. Twenty-seven different genotypes were identified. Echovirus 30 (E-30) accounted for 54.1% of all characterized strains and caused a large outbreak. Four severe neonatal infections were reported, of which two were caused by EV-A71. Respiratory infections involving EV-D68 were observed in two adults. One fatal case of Coxsackievirus A2-associated myocarditis was reported. Monitoring EV infections in combination with EV genotyping via the French EV network characterized the epidemiology of EV infections in the Ile de France region in 2013 and documented severe EV infections associated with EV-A71 or CV-A2. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Evaluation of the enterovirus laboratory surveillance system in Denmark, 2010 to 2013.

    Science.gov (United States)

    Condell, Orla; Midgley, Sofie; Christiansen, Claus Bohn; Chen, Ming; Chen Nielsen, Xiaohui; Ellermann-Eriksen, Svend; Mølvadgaard, Mette; Schønning, Kristian; Vermedal Hoegh, Silje; Andersen, Peter Henrik; Voldstedlund, Marianne; Fischer, Thea Kølsen

    2016-05-05

    The primary aim of the Danish enterovirus (EV) surveillance system is to document absence of poliovirus infection. The conflict in Syria has left many children unvaccinated and movement from areas with polio cases to Europe calls for increased awareness to detect and respond to virus-transmission in a timely manner. We evaluate the national EV laboratory surveillance, to generate recommendations for system strengthening. The system was analysed for completeness of viral typing analysis and clinical information and timeliness of specimen collection, laboratory results and reporting of clinica