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The HBx protein, encoded by <jats:italic>HBX<\/jats:italic>, has a key role in viral replication. Here, we analyzed the viral quasispecies at the 5\u2032 end of <jats:italic>HBX<\/jats:italic> in CI patients and compared it with that of patients in other clinical stages. Fifty-eight HBeAg-negative patients were included: 16 CI, 19 chronic hepatitis B, 16 hepatocellular carcinoma and 6 liver cirrhosis. Quasispecies complexity and conservation were determined in the region between nucleotides 1255 and 1611. Amino acid changes detected were tested in vitro. CI patients showed higher complexity in terms of mutation frequency and nucleotide diversity and higher quasispecies conservation (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.05). A genotype D-specific pattern of mutations (A12S\/P33S\/P46S\/T36D-G) was identified in CI (median frequency, 81.7%), which determined a reduction in HBV DNA release of up to 1.5 log in vitro. CI patients showed a more complex and conserved viral quasispecies than the other groups. The genotype-specific pattern of mutations could partially explain the low viremia observed in these patients.<\/jats:p>","DOI":"10.1038\/s41598-021-83762-4","type":"journal-article","created":{"date-parts":[[2021,2,20]],"date-time":"2021-02-20T09:14:59Z","timestamp":1613812499000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Sophisticated viral quasispecies with a genotype-related pattern of mutations in the hepatitis B X gene of HBeAg-ve chronically infected patients"],"prefix":"10.1038","volume":"11","author":[{"given":"Maria Francesca","family":"Cortese","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carolina","family":"Gonz\u00e1lez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Josep","family":"Gregori","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rosario","family":"Casillas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luca","family":"Carioti","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mercedes","family":"Guerrero-Murillo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar","family":"Riveiro-Barciela","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cristina","family":"Godoy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sara","family":"Sopena","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00e7al","family":"Yll","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Josep","family":"Quer","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ariadna","family":"Rando","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rosa","family":"Lopez-Martinez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Beatriz","family":"Pac\u00edn Ruiz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Selene","family":"Garc\u00eda-Garc\u00eda","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rafael","family":"Esteban-Mur","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Tabernero","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria","family":"Buti","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francisco","family":"Rodr\u00edguez-Fr\u00edas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2021,2,18]]},"reference":[{"key":"83762_CR1","doi-asserted-by":"publisher","first-page":"2575","DOI":"10.1099\/0022-1317-69-10-2575","volume":"69","author":"H Okamoto","year":"1988","unstructured":"Okamoto, H. et al. 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