دورية أكاديمية
أعرض في EDS

Development and evaluation of a non-ribosomal random PCR and next-generation sequencing based assay for detection and sequencing of hand, foot and mouth disease pathogens.

التفاصيل البيبلوغرافية
العنوان: Development and evaluation of a non-ribosomal random PCR and next-generation sequencing based assay for detection and sequencing of hand, foot and mouth disease pathogens.
المؤلفون: Nguyen AT; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam. anhnt@oucru.org., Tran TT; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam., Hoang VM; Children's Hospital 2, Ho Chi Minh City, Vietnam., Nghiem NM; Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam., Le NN; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam., Le TT; Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam., Phan QT; Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam., Truong KH; Children's Hospital 1, Ho Chi Minh City, Vietnam., Le NN; Children's Hospital 1, Ho Chi Minh City, Vietnam., Ho VL; Children's Hospital 2, Ho Chi Minh City, Vietnam., Do VC; Children's Hospital 2, Ho Chi Minh City, Vietnam., Ha TM; Children's Hospital 2, Ho Chi Minh City, Vietnam., Nguyen HT; Children's Hospital 1, Ho Chi Minh City, Vietnam., Nguyen CV; Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam., Thwaites G; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam.; Centre for Tropical Medicine, Nuffield Department of Medicine, University of Oxford, Oxford, UK., van Doorn HR; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam.; Centre for Tropical Medicine, Nuffield Department of Medicine, University of Oxford, Oxford, UK., Le TV; Oxford University Clinical Research Unit, 764 Vo Van Kiet Street, Ward 1, District 5, Ho Chi Minh City, Vietnam.
المصدر: Virology journal [Virol J] 2016 Jul 07; Vol. 13, pp. 125. Date of Electronic Publication: 2016 Jul 07.
نوع المنشور: Evaluation Study; Journal Article
اللغة: English
بيانات الدورية: Publisher: BioMed Central Country of Publication: England NLM ID: 101231645 Publication Model: Electronic Cited Medium: Internet ISSN: 1743-422X (Electronic) Linking ISSN: 1743422X NLM ISO Abbreviation: Virol J Subsets: MEDLINE
أسماء مطبوعة: Original Publication: [London] : BioMed Central, 2004-
مواضيع طبية MeSH: Enterovirus A, Human/*isolation & purification , Hand, Foot and Mouth Disease/*virology , High-Throughput Nucleotide Sequencing/*methods , Polymerase Chain Reaction/*methods, Enterovirus A, Human/classification ; Enterovirus A, Human/genetics ; Genotype ; Hand, Foot and Mouth Disease/diagnosis ; Humans ; Phylogeny ; Serotyping
مستخلص: Background: Hand, foot and mouth disease (HFMD) has become a major public health problem across the Asia-Pacific region, and is commonly caused by enterovirus A71 (EV-A71) and coxsackievirus A6 (CV-A6), CV-A10 and CV-A16. Generating pathogen whole-genome sequences is essential for understanding their evolutionary biology. The frequent replacements among EV serotypes and a limited numbers of available whole-genome sequences hinder the development of overlapping PCRs for whole-genome sequencing. We developed and evaluated a non-ribosomal random PCR (rPCR) and next-generation sequencing based assay for sequence-independent whole-genome amplification and sequencing of HFMD pathogens. A total of 16 EV-A71/CV-A6/CV-A10/CV-A16 PCR positive rectal/throat swabs (Cp values: 20.9-33.3) were used for assay evaluation.
Results: Our assay evidently outperformed the conventional rPCR in terms of the total number of EV-A71 reads and the percentage of EV-A71 reads: 2.6 % (1275/50,000 reads) vs. 0.1 % (31/50,000) and 6 % (3008/50,000) vs. 0.9 % (433/50,000) for two samples with Cp values of 30 and 26, respectively. Additionally the assay could generate genome sequences with the percentages of coverage of 94-100 % of 4 different enterovirus serotypes in 73 % of the tested samples, representing the first whole-genome sequences of CV-A6/10/16 from Vietnam, and could assign correctly serotyping results in 100 % of 24 tested specimens. In all but three the obtained consensuses of two replicates from the same sample were 100 % identical, suggesting that our assay is highly reproducible.
Conclusions: In conclusion, we have successfully developed a non-ribosomal rPCR and next-generation sequencing based assay for sensitive detection and direct whole-genome sequencing of HFMD pathogens from clinical samples.
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معلومات مُعتمدة: United Kingdom WT_ Wellcome Trust
فهرسة مساهمة: Keywords: Enterovirus A; FR26RV-Endoh primer; Hand, foot and mouth disease; Next-generation sequencing; Random PCR
تواريخ الأحداث: Date Created: 20160709 Date Completed: 20170801 Latest Revision: 20230723
رمز التحديث: 20230723
مُعرف محوري في PubMed: PMC4937578
DOI: 10.1186/s12985-016-0580-9
PMID: 27388326
قاعدة البيانات: MEDLINE
الوصف
تدمد:1743-422X
DOI:10.1186/s12985-016-0580-9