{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,9,28]],"date-time":"2025-09-28T12:50:54Z","timestamp":1759063854613,"version":"3.37.3"},"reference-count":37,"publisher":"Wiley","issue":"10","license":[{"start":{"date-parts":[[2018,10,21]],"date-time":"2018-10-21T00:00:00Z","timestamp":1540080000000},"content-version":"vor","delay-in-days":20,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"name":"JSPS KAKENHI","award":["17K10185 and 17H04346"],"award-info":[{"award-number":["17K10185 and 17H04346"]}]},{"DOI":"10.13039\/100009619","name":"AMED","doi-asserted-by":"crossref","award":["17gk0110021h0002"],"award-info":[{"award-number":["17gk0110021h0002"]}],"id":[{"id":"10.13039\/100009619","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Microbiology and Immunology"],"published-print":{"date-parts":[[2018,10]]},"abstract":"<jats:title>ABSTRACT<\/jats:title><jats:sec><jats:label\/><jats:p>Quantification of human cytomegalovirus (HCMV) replication by plaque assay reflects viral infectivity but has several drawbacks. Recombinant virus expressing reporter genes can facilitate quantification of HCMV replication. In this study, a recombinant virus, Towne\u2010BAC(dTT)\u2010MetLuc, was constructed and the secretable <jats:italic>Metridia<\/jats:italic> luciferase (MetLuc) gene inserted into it under <jats:italic>UL146<\/jats:italic> promoter. In addition, the <jats:italic>UL130<\/jats:italic> gene was repaired to allow growth of the recombinant virus in both fibroblasts and epithelial cells. As predicted, luciferase activity was secreted into the culture medium and correlated with virus replication in infected fibroblasts and epithelial cells. Furthermore, secreted luciferase activity was correlated with the size of the recombinant virus inoculum with a dynamic range of 3 logs. This recombinant virus was used in a two\u2010step culture protocol for detection of the anti\u2010HCMV activity of compounds; that is, the supernatant of a first\u2010step culture with anti\u2010viral compounds was collected and inoculated into uninfected cells to create a second\u2010step culture. Although secreted luciferase activity leaked in the first\u2010step culture supernatant in the presence of some antiviral compounds, luciferase activity in the second\u2010step culture supernatant reflected the virus yield in the first\u2010step culture. Therefore, comparison of luciferase activity in the first\u2010 and second\u2010step cultures indicated the anti\u2010viral activity of the compounds. This two\u2010step culture protocol facilitates screening of anti\u2010viral compounds.<\/jats:p><\/jats:sec>","DOI":"10.1111\/1348-0421.12645","type":"journal-article","created":{"date-parts":[[2018,9,6]],"date-time":"2018-09-06T11:33:26Z","timestamp":1536233606000},"page":"651-658","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["A two\u2010step culture method utilizing secreted luciferase recombinant virus for detection of anti\u2010cytomegalovirus compounds"],"prefix":"10.1111","volume":"62","author":[{"given":"Tetsuo","family":"Koshizuka","sequence":"first","affiliation":[{"name":"Department of Microbiology Fukushima Medical University School of Medicine 1 Hikarigaoka Fukushima 960\u20101295 Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuko","family":"Sato","sequence":"additional","affiliation":[{"name":"Department of Microbiology Fukushima Medical University School of Medicine 1 Hikarigaoka Fukushima 960\u20101295 Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shoe","family":"Kobiyama","sequence":"additional","affiliation":[{"name":"Department of Microbiology Fukushima Medical University School of Medicine 1 Hikarigaoka Fukushima 960\u20101295 Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mami","family":"Oshima","sequence":"additional","affiliation":[{"name":"Department of Microbiology Fukushima Medical University School of Medicine 1 Hikarigaoka Fukushima 960\u20101295 Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tatsuo","family":"Suzutani","sequence":"additional","affiliation":[{"name":"Department of Microbiology Fukushima Medical University School of Medicine 1 Hikarigaoka Fukushima 960\u20101295 Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2018,10,21]]},"reference":[{"volume-title":"Fields Virology, 6th ed","year":"2013","author":"Knipe D.M.","key":"e_1_2_8_2_1"},{"key":"e_1_2_8_3_1","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2014.00389"},{"key":"e_1_2_8_4_1","doi-asserted-by":"publisher","DOI":"10.1172\/JCI45449"},{"key":"e_1_2_8_5_1","doi-asserted-by":"publisher","DOI":"10.1002\/path.4437"},{"key":"e_1_2_8_6_1","doi-asserted-by":"publisher","DOI":"10.1586\/14787210.5.3.427"},{"key":"e_1_2_8_7_1","doi-asserted-by":"publisher","DOI":"10.1086\/598506"},{"key":"e_1_2_8_8_1","doi-asserted-by":"publisher","DOI":"10.1086\/511981"},{"key":"e_1_2_8_9_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.78.18.10023-10033.2004"},{"key":"e_1_2_8_10_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.80.2.710-722.2006"},{"key":"e_1_2_8_11_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.79.13.8361-8373.2005"},{"key":"e_1_2_8_12_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.79.16.10330-10338.2005"},{"key":"e_1_2_8_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.antiviral.2006.05.002"},{"key":"e_1_2_8_14_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.transproceed.2009.02.093"},{"key":"e_1_2_8_15_1","doi-asserted-by":"publisher","DOI":"10.1517\/14656560802678138"},{"key":"e_1_2_8_16_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.05265-11"},{"key":"e_1_2_8_17_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.01596-09"},{"key":"e_1_2_8_18_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.01794-13"},{"key":"e_1_2_8_19_1","doi-asserted-by":"publisher","DOI":"10.1093\/infdis\/jiv352"},{"key":"e_1_2_8_20_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.234.4778.856"},{"key":"e_1_2_8_21_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M309639200"},{"key":"e_1_2_8_22_1","doi-asserted-by":"publisher","DOI":"10.1021\/ac025742k"},{"key":"e_1_2_8_23_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.279.5349.349"},{"key":"e_1_2_8_24_1","doi-asserted-by":"publisher","DOI":"10.1006\/exer.1996.0020"},{"key":"e_1_2_8_25_1","doi-asserted-by":"publisher","DOI":"10.2144\/000112096"},{"key":"e_1_2_8_26_1","doi-asserted-by":"publisher","DOI":"10.1099\/jgv.0.000336"},{"key":"e_1_2_8_27_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.96.17.9839"},{"key":"e_1_2_8_28_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.antiviral.2013.07.016"},{"key":"e_1_2_8_29_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01275-13"},{"key":"e_1_2_8_30_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.00214-13"},{"key":"e_1_2_8_31_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.03274-15"},{"key":"e_1_2_8_32_1","doi-asserted-by":"publisher","DOI":"10.1093\/bmb\/ldt023"},{"key":"e_1_2_8_33_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.44.6.1588-1597.2000"},{"key":"e_1_2_8_34_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jviromet.2016.06.008"},{"key":"e_1_2_8_35_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1002172"},{"key":"e_1_2_8_36_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1001256"},{"key":"e_1_2_8_37_1","doi-asserted-by":"publisher","DOI":"10.1186\/1743-422X-8-40"},{"key":"e_1_2_8_38_1","doi-asserted-by":"publisher","DOI":"10.2174\/187152609787847758"}],"container-title":["Microbiology and Immunology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1111%2F1348-0421.12645","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/1348-0421.12645","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,9,16]],"date-time":"2023-09-16T17:13:25Z","timestamp":1694884405000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1111\/1348-0421.12645"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2018,10]]},"references-count":37,"journal-issue":{"issue":"10","published-print":{"date-parts":[[2018,10]]}},"alternative-id":["10.1111\/1348-0421.12645"],"URL":"https:\/\/doi.org\/10.1111\/1348-0421.12645","archive":["Portico"],"relation":{},"ISSN":["0385-5600","1348-0421"],"issn-type":[{"type":"print","value":"0385-5600"},{"type":"electronic","value":"1348-0421"}],"subject":[],"published":{"date-parts":[[2018,10]]},"assertion":[{"value":"2018-06-22","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2018-08-30","order":1,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2018-10-21","order":2,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}]}}