{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,27]],"date-time":"2025-12-27T21:10:28Z","timestamp":1766869828460,"version":"3.41.2"},"reference-count":29,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,11,25]],"date-time":"2021-11-25T00:00:00Z","timestamp":1637798400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100009619","name":"Japan Agency for Medical Research and Development","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100009619","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Microbiol."],"abstract":"<jats:p>The 3\u2032 untranslated region (UTR) of the hepatitis C virus (HCV) genome plays a significant role in replication including the poly(U) tract (<jats:xref>You and Rice, 2008<\/jats:xref>). Here we established an HCV clone that is infectious <jats:italic>in vitro<\/jats:italic> and <jats:italic>in vivo<\/jats:italic>, from an Egyptian patient with chronic HCV infection and hepatocellular carcinoma (HCC). First, we inoculated the patient plasma into a humanized chimeric mouse and passaged. We observed HCV genotype 4a propagation in the chimeric mouse sera at 1.7 \u00d7 10<jats:sup>7<\/jats:sup> copies\/mL after 6 weeks. Next, we cloned the entire HCV sequence from the HCV-infected chimeric mouse sera using RT-PCR, and 5\u2032 and 3\u2032 RACE methodologies. We obtained first a shorter clone (HCV-G4 KM short, GenBank: AB795432.1), which contained 9,545 nucleotides with 341 nucleotides of the 5\u2032UTR and 177 nucleotides of the 3\u2032UTR, and this was frequently obtained for unknown reasons. We also obtained a longer clone by dividing the HCV genome into three fragments and the poly (U) sequences. We obtained a longer 3\u2032UTR sequence than that of the HCV-G4 KM short clone, which contained 9,617 nucleotides. This longer clone possessed a 3\u2032-UTR of 249 nucleotides (HCV-G4 KM long, GenBank: AB795432.2), because of a 71-nucleotide longer poly (U) stretch. The HCV-G4-KM long clone, but not the HCV-G4-KM short clone, could establish infection in human hepatoma HuH-7 cells. HCV RNAs carrying a nanoluciferase (NL) reporter were also constructed and higher replication activity was observed with G4-KM long-NL <jats:italic>in vitro<\/jats:italic>. Next, both short and long RNAs were intra-hepatically injected into humanized chimeric mice. Viral propagation was only observed for the chimeric mouse injected with the HCV-G4 KM long RNA in the sera after 21 days (1.64 \u00d7 10<jats:sup>6<\/jats:sup> copies\/mL) and continued until 10 weeks post inoculation (wpi; 1.45\u20134.74 \u00d7 10<jats:sup>7<\/jats:sup> copies\/mL). Moreover, sequencing of the HCV genome in mouse sera at 6 wpi revealed the sequence of the HCV-G4-KM long clone. Thus, the <jats:italic>in vitro<\/jats:italic> and <jats:italic>in vivo<\/jats:italic> results of this study indicate that the sequence of the HCV-G4-KM long RNA is that of an infectious clone.<\/jats:p>","DOI":"10.3389\/fmicb.2021.764816","type":"journal-article","created":{"date-parts":[[2021,12,1]],"date-time":"2021-12-01T02:29:48Z","timestamp":1638325788000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":4,"title":["Longer Poly(U) Stretches in the 3\u2032UTR Are Essential for Replication of the Hepatitis C Virus Genotype 4a Clone in in vitro and in vivo"],"prefix":"10.3389","volume":"12","author":[{"given":"Asako","family":"Takagi","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yutaka","family":"Amako","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Daisuke","family":"Yamane","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bouchra","family":"Kitab","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuko","family":"Tokunaga","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ahmed","family":"El-Gohary","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Michinori","family":"Kohara","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kyoko","family":"Tsukiyama-Kohara","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1965","published-online":{"date-parts":[[2021,11,25]]},"reference":[{"key":"B1","doi-asserted-by":"crossref","first-page":"1722","DOI":"10.1093\/infdis\/jiy401","article-title":"Identification of a novel hepatitis C virus genotype from Punjab, India: expanding classification of hepatitis C virus into 8 genotypes.","volume":"218","author":"Borgia","year":"2018","journal-title":"J. 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