{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,26]],"date-time":"2026-03-26T18:10:17Z","timestamp":1774548617452,"version":"3.50.1"},"reference-count":48,"publisher":"MDPI AG","issue":"21","license":[{"start":{"date-parts":[[2020,11,5]],"date-time":"2020-11-05T00:00:00Z","timestamp":1604534400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The goal of the study was to develop a specific, sensitive, and cost-effective molecular RT-PCR diagnostic assay for the rapid and simultaneous detection of the serotypes of dengue virus (DENV) and Chikungunya virus (CHIKV) from sera of suspected febrile patients. A single-tube, single-step multiplex RT-PCR (mRT-PCR) assay was designed for the detection of viral genomes from clinical and field samples. Specificity and sensitivity of the mRT-PCR assay were evaluated against six different combinations using two reverse transcriptases (AMV-RT and RT-Ace) and three DNA polymerases (LA-Taq, rTaq, and Tth). Among the six combinations, the AMV-RT and LA-Taq combination was more specific and sensitive than other enzyme combinations for detecting viral genomes of DENV-1, DENV-2, DENV-3, and DENV-4 (p &lt; 0.01), and for detecting viral genomes of CHIKV (p &lt; 0.05). The detection limits of the mRT-PCR were 10 focus forming units (FFU) for CHIKV and 1 FFU, 20 FFU, 0.1 FFU, and 10 FFU for DENV-1, DENV-2, DENV-3, and DENV-4, respectively. The primers used for the mRT-PCR did not show any cross-reactivity among the serotypes of DENV or CHIKV. Specificity and sensitivity of the newly developed mRT-PCR were validated using serum samples collected from febrile patients during dengue outbreaks in Bangladesh. The sensitivity for serotype detection of DENV and CHIKV was superior to the virus isolation method and the antigen detection method using the Dengue NS1-Ag assay. This novel mRT-PCR method can be used for molecular epidemiological surveillance of DENV and CHIKV in epidemic and endemic countries.<\/jats:p>","DOI":"10.3390\/ijms21218281","type":"journal-article","created":{"date-parts":[[2020,11,5]],"date-time":"2020-11-05T09:04:34Z","timestamp":1604567074000},"page":"8281","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":18,"title":["A Novel Multiplex RT-PCR Assay for Simultaneous Detection of Dengue and Chikungunya Viruses"],"prefix":"10.3390","volume":"21","author":[{"given":"Mohammed Alimul","family":"Islam","sequence":"first","affiliation":[{"name":"Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1056-4761","authenticated-orcid":false,"given":"Mohamed E.","family":"El Zowalaty","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, College of Medicine, University of Sharjah, Sharjah 27272, UAE"},{"name":"Zoonosis Science Center, Department of Medical Biochemistry and Microbiology, Uppsala University, SE 75123 Uppsala, Sweden"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sumaiya","family":"Islam","sequence":"additional","affiliation":[{"name":"Department of Medicine, Bangladesh Medical College and Hospital, Dhaka 1209, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5580-2823","authenticated-orcid":false,"given":"Mohiuddin","family":"Sharif","sequence":"additional","affiliation":[{"name":"Department of Medicine, Dhaka Medical College and Hospital, Dhaka 1000, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Md. Rajibur","family":"Rahman","sequence":"additional","affiliation":[{"name":"Department of Medicine, Dhaka Medical College and Hospital, Dhaka 1000, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5500-5103","authenticated-orcid":false,"given":"Mohammad Robed","family":"Amin","sequence":"additional","affiliation":[{"name":"Department of Medicine, Dhaka Medical College and Hospital, Dhaka 1000, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Md. Mortuza","family":"Ali","sequence":"additional","affiliation":[{"name":"Department of Community Medicine, Monno Medical College and Hospital, Manikganj 1800, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5432-480X","authenticated-orcid":false,"given":"Md. Tanvir","family":"Rahman","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh 2202, Bangladesh"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kouichi","family":"Morita","sequence":"additional","affiliation":[{"name":"Department of Virology, Institute of Tropical Medicine, Nagasaki University, 1-12-4, Sakamoto, Nagasaki 852-8523, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hossam M.","family":"Ashour","sequence":"additional","affiliation":[{"name":"Department of Integrative Biology, College of Arts and Sciences, University of South Florida, St. Petersburg, FL 33701, USA"},{"name":"Department of Microbiology and Immunology, Faculty of Pharmacy, Cairo University, Cairo 11562, Egypt"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2020,11,5]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"371","DOI":"10.1016\/j.cll.2017.01.008","article-title":"Chikungunya Virus","volume":"37","author":"Vu","year":"2019","journal-title":"Clin. 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