{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2024,3,6]],"date-time":"2024-03-06T09:57:21Z","timestamp":1709719041281},"reference-count":12,"publisher":"Wiley","issue":"10","license":[{"start":{"date-parts":[[2010,9,1]],"date-time":"2010-09-01T00:00:00Z","timestamp":1283299200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Journal of Medical Virology"],"published-print":{"date-parts":[[2010,10]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>A large number of human infections are caused by different dengue virus strains, mainly in the tropical and subtropical parts of the world, but also outside the endemic regions. RT\u2010PCR methods are used widely for detection of dengue virus RNA in acute\u2010phase serum samples; however, new sequence variation can inhibit these methods. An assay was developed integrating an anchored Pan Dengue RT\u2010PCR with a new Fast Sanger sequencing protocol. For broad detection and identification of dengue virus RNA, including new strains of all serotypes, the conserved 3\u2032 genome end was targeted for highly specific cDNA synthesis. A combination of degenerated primers was used for second strand synthesis, followed by tag primed amplification. The mixture of generated amplicons was identified directly by the Fast Sanger sequencing from the anchored 3\u2032 genome end. Evaluating the assay on human serum RNA spiked with viral RNA representing the four dengue serotypes demonstrated a detection limit of 44\u2013124 copies viral RNA per reaction for a two\u2010step format of the anchored Pan Dengue RT\u2010PCR and 100\u2013500 copies for a one\u2010step protocol, respectively. The different serotypes were clearly identified from the generated sequences. Further, the 5\u2010hr procedure was evaluated and compared to standard real\u2010time RT\u2010PCR protocols on acute\u2010phase serum samples from patients with confirmed dengue infections. This assay demonstrates a strategy for virus detection, which combines nucleic acid amplification adapted for dengue virus RNA with direct and rapid sequencing. It provides a tolerance for new sequence variation and the strategy should be applicable for other RNA viruses. J. Med. Virol. 82:1701\u20131710, 2010. \u00a9 2010 Wiley\u2010Liss, Inc.<\/jats:p>","DOI":"10.1002\/jmv.21882","type":"journal-article","created":{"date-parts":[[2010,9,14]],"date-time":"2010-09-14T05:48:27Z","timestamp":1284443307000},"page":"1701-1710","source":"Crossref","is-referenced-by-count":4,"title":["Anchored Pan Dengue RT\u2010PCR and Fast Sanger sequencing for detection of Dengue RNA in human serum"],"prefix":"10.1002","volume":"82","author":[{"given":"Zhe","family":"Hu","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Henrik","family":"Nordstr\u00f6m","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Norbert","family":"Nowotny","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kerstin I.","family":"Falk","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Gunnar","family":"Sandstr\u00f6m","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2010,9]]},"reference":[{"key":"e_1_2_6_2_1","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.11.3.480"},{"key":"e_1_2_6_3_1","doi-asserted-by":"publisher","DOI":"10.1016\/S1473-3099(01)00171-2"},{"key":"e_1_2_6_4_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jviromet.2006.08.003"},{"key":"e_1_2_6_5_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01258-06"},{"key":"e_1_2_6_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jcv.2009.02.010"},{"key":"e_1_2_6_7_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.43.6.2850-2855.2005"},{"key":"e_1_2_6_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.diagmicrobio.2009.01.002"},{"key":"e_1_2_6_9_1","doi-asserted-by":"publisher","DOI":"10.1002\/jmv.20489"},{"key":"e_1_2_6_10_1","doi-asserted-by":"publisher","DOI":"10.1006\/viro.2000.0748"},{"key":"e_1_2_6_11_1","doi-asserted-by":"publisher","DOI":"10.2353\/jmoldx.2006.060019"},{"key":"e_1_2_6_12_1","first-page":"53","volume":"79","author":"World Health Organization","year":"2004","journal-title":"Wkly Epidemiol Rec"},{"key":"e_1_2_6_13_1","unstructured":"World Health Organization.2009.Dengue and dengue haemorrhagic fever. (Fact sheet No 117)."}],"container-title":["Journal of Medical Virology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1002%2Fjmv.21882","content-type":"unspecified","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/jmv.21882","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,10,9]],"date-time":"2023-10-09T21:18:02Z","timestamp":1696886282000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1002\/jmv.21882"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2010,9]]},"references-count":12,"journal-issue":{"issue":"10","published-print":{"date-parts":[[2010,10]]}},"alternative-id":["10.1002\/jmv.21882"],"URL":"https:\/\/doi.org\/10.1002\/jmv.21882","archive":["Portico"],"relation":{},"ISSN":["0146-6615","1096-9071"],"issn-type":[{"value":"0146-6615","type":"print"},{"value":"1096-9071","type":"electronic"}],"subject":[],"published":{"date-parts":[[2010,9]]}}}