Single-step real-time PCR to quantify hepatitis B virus and distinguish genotype D from non-D genotypes
Hepatitis B virus (HBV) viral load and its genotype play important roles in clinical outcome, management of disease and response to antiviral therapy. In many parts of the world such as Europe or the Middle East, distinguishing HBV genotype D from non‐D is most relevant for treatment decisions, beca...
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| Veröffentlicht in: | Journal of viral hepatitis 2011-04, Vol.18 (4), p.300-304 |
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| container_title | Journal of viral hepatitis |
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| creator | Amini-Bavil-Olyaee, S. Pourkarim, M. R. Schaefer, S. Mahboudi, F. Van Ranst, M. Adeli, A. Trautwein, C. Tacke, F. |
| description | Hepatitis B virus (HBV) viral load and its genotype play important roles in clinical outcome, management of disease and response to antiviral therapy. In many parts of the world such as Europe or the Middle East, distinguishing HBV genotype D from non‐D is most relevant for treatment decisions, because genotype D‐infected patients respond poorly to interferon‐based therapeutic regimens. Here, we developed an in‐house real‐time PCR to concordantly assess HBV genotype (D vs non‐D) based on melt curve analysis and quantify the viral load. Genotype distinction was established with control plasmids of all HBV genotypes and validated with 57 clinical samples from patients infected with six different HBV genotypes. Our in‐house real‐time PCR assay could discriminate HBV genotype D from non‐D using single‐step melt curve analysis with a 2 °C difference in the melt curve temperature in all samples tested. Viral load quantification was calibrated with the WHO HBV international standard, demonstrating an excellent correlation with a commercial kit (r = 0.852; P |
| doi_str_mv | 10.1111/j.1365-2893.2010.01308.x |
| format | Article |
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R. ; Schaefer, S. ; Mahboudi, F. ; Van Ranst, M. ; Adeli, A. ; Trautwein, C. ; Tacke, F.</creator><creatorcontrib>Amini-Bavil-Olyaee, S. ; Pourkarim, M. R. ; Schaefer, S. ; Mahboudi, F. ; Van Ranst, M. ; Adeli, A. ; Trautwein, C. ; Tacke, F.</creatorcontrib><description>Hepatitis B virus (HBV) viral load and its genotype play important roles in clinical outcome, management of disease and response to antiviral therapy. In many parts of the world such as Europe or the Middle East, distinguishing HBV genotype D from non‐D is most relevant for treatment decisions, because genotype D‐infected patients respond poorly to interferon‐based therapeutic regimens. Here, we developed an in‐house real‐time PCR to concordantly assess HBV genotype (D vs non‐D) based on melt curve analysis and quantify the viral load. Genotype distinction was established with control plasmids of all HBV genotypes and validated with 57 clinical samples from patients infected with six different HBV genotypes. Our in‐house real‐time PCR assay could discriminate HBV genotype D from non‐D using single‐step melt curve analysis with a 2 °C difference in the melt curve temperature in all samples tested. Viral load quantification was calibrated with the WHO HBV international standard, demonstrating an excellent correlation with a commercial kit (r = 0.852; P < 0.0001) in a linear range from 3.2 × 102 to 3.2 × 1010 IU/mL. In conclusion, we developed a rapid, simple and cost‐effective method to simultaneously quantify and distinguish HBV genotypes D from non‐D with a single‐step PCR run and melt curve analysis. This assay should be a useful diagnostic alternative to aid clinical decisions about initiation and choice of antiviral therapy, especially in geographical regions with a high prevalence of HBV genotype D.</description><identifier>ISSN: 1352-0504</identifier><identifier>EISSN: 1365-2893</identifier><identifier>DOI: 10.1111/j.1365-2893.2010.01308.x</identifier><identifier>PMID: 20367802</identifier><language>eng</language><publisher>Oxford, UK: Blackwell Publishing Ltd</publisher><subject>copy number ; DNA, Viral ; DNA, Viral - genetics ; Europe ; Genotype ; Genotypes ; Hepatitis B virus ; Hepatitis B virus - classification ; Hepatitis B virus - genetics ; Hepatitis B virus - isolation & purification ; Humans ; Immunology ; interferon ; Life Sciences ; Middle East ; pegylated interferon ; Polymerase Chain Reaction ; Polymerase Chain Reaction - methods ; quantitative real-time PCR ; Viral Load ; Viral Load - methods</subject><ispartof>Journal of viral hepatitis, 2011-04, Vol.18 (4), p.300-304</ispartof><rights>2010 Blackwell Publishing Ltd</rights><rights>2010 Blackwell Publishing Ltd.</rights><rights>Distributed under a Creative Commons Attribution 4.0 International License</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4768-df887a430e7981f4235cb6b411af35a5de7c8f843490606f4a841929f18280963</citedby><cites>FETCH-LOGICAL-c4768-df887a430e7981f4235cb6b411af35a5de7c8f843490606f4a841929f18280963</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1111%2Fj.1365-2893.2010.01308.x$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1111%2Fj.1365-2893.2010.01308.x$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>230,314,776,780,881,1411,27901,27902,45550,45551</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/20367802$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://riip.hal.science/pasteur-00829137$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Amini-Bavil-Olyaee, S.</creatorcontrib><creatorcontrib>Pourkarim, M. 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Here, we developed an in‐house real‐time PCR to concordantly assess HBV genotype (D vs non‐D) based on melt curve analysis and quantify the viral load. Genotype distinction was established with control plasmids of all HBV genotypes and validated with 57 clinical samples from patients infected with six different HBV genotypes. Our in‐house real‐time PCR assay could discriminate HBV genotype D from non‐D using single‐step melt curve analysis with a 2 °C difference in the melt curve temperature in all samples tested. Viral load quantification was calibrated with the WHO HBV international standard, demonstrating an excellent correlation with a commercial kit (r = 0.852; P < 0.0001) in a linear range from 3.2 × 102 to 3.2 × 1010 IU/mL. In conclusion, we developed a rapid, simple and cost‐effective method to simultaneously quantify and distinguish HBV genotypes D from non‐D with a single‐step PCR run and melt curve analysis. This assay should be a useful diagnostic alternative to aid clinical decisions about initiation and choice of antiviral therapy, especially in geographical regions with a high prevalence of HBV genotype D.</description><subject>copy number</subject><subject>DNA, Viral</subject><subject>DNA, Viral - genetics</subject><subject>Europe</subject><subject>Genotype</subject><subject>Genotypes</subject><subject>Hepatitis B virus</subject><subject>Hepatitis B virus - classification</subject><subject>Hepatitis B virus - genetics</subject><subject>Hepatitis B virus - isolation & purification</subject><subject>Humans</subject><subject>Immunology</subject><subject>interferon</subject><subject>Life Sciences</subject><subject>Middle East</subject><subject>pegylated interferon</subject><subject>Polymerase Chain Reaction</subject><subject>Polymerase Chain Reaction - methods</subject><subject>quantitative real-time PCR</subject><subject>Viral Load</subject><subject>Viral Load - methods</subject><issn>1352-0504</issn><issn>1365-2893</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2011</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNkUtz0zAUhT0MDC2Fv8Box8pBT0tesCgpJGXCY8Jrhs0dxZYSBb8q2W3y75FJMUvQRneuvnMlnZMkiOAZievlfkZYJlKqcjajOHYxYVjNDg-S8-ng4VgLmmKB-VnyJIQ9jhQV5HFyRjHLpML0PNl-ds22MmnoTYe80VXau9qgT_M16lt0M-imd_aIdqbTvetdQK_RrfNDQLopUelCH-WDCzu0NU3bHzuDrpD1bY2atkmvpm54mjyyugrm2f1-kXx9--bLfJmuPi6u55ertOAyU2lplZKaM2xkrojllIlik204IdoyoUVpZKGs4oznOMOZ5VpxktPcEkUVzjN2kaSnuTtdQeddrf0RWu1gebmCTsdvDh4wVjQnTN6SyL848Z1vbwYTeqhdKExV6ca0QwAlaXSURsP-SYpMyui4jKQ6kYVvQ_DGTg8hGMb8YA9jTDDGBGN-8Ds_OETp8_tLhk1tykn4J7AIvDoBd64yx_8eDO--Lcfqrz8xOnOY9Nr_hEwyKeD7hwXINeeL9_QHrNkvCY62TQ</recordid><startdate>201104</startdate><enddate>201104</enddate><creator>Amini-Bavil-Olyaee, S.</creator><creator>Pourkarim, M. R.</creator><creator>Schaefer, S.</creator><creator>Mahboudi, F.</creator><creator>Van Ranst, M.</creator><creator>Adeli, A.</creator><creator>Trautwein, C.</creator><creator>Tacke, F.</creator><general>Blackwell Publishing Ltd</general><general>Wiley-Blackwell</general><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>7U9</scope><scope>H94</scope><scope>1XC</scope><scope>VOOES</scope></search><sort><creationdate>201104</creationdate><title>Single-step real-time PCR to quantify hepatitis B virus and distinguish genotype D from non-D genotypes</title><author>Amini-Bavil-Olyaee, S. ; Pourkarim, M. 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Here, we developed an in‐house real‐time PCR to concordantly assess HBV genotype (D vs non‐D) based on melt curve analysis and quantify the viral load. Genotype distinction was established with control plasmids of all HBV genotypes and validated with 57 clinical samples from patients infected with six different HBV genotypes. Our in‐house real‐time PCR assay could discriminate HBV genotype D from non‐D using single‐step melt curve analysis with a 2 °C difference in the melt curve temperature in all samples tested. Viral load quantification was calibrated with the WHO HBV international standard, demonstrating an excellent correlation with a commercial kit (r = 0.852; P < 0.0001) in a linear range from 3.2 × 102 to 3.2 × 1010 IU/mL. In conclusion, we developed a rapid, simple and cost‐effective method to simultaneously quantify and distinguish HBV genotypes D from non‐D with a single‐step PCR run and melt curve analysis. 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| ispartof | Journal of viral hepatitis, 2011-04, Vol.18 (4), p.300-304 |
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| source | MEDLINE; Wiley Online Library |
| subjects | copy number DNA, Viral DNA, Viral - genetics Europe Genotype Genotypes Hepatitis B virus Hepatitis B virus - classification Hepatitis B virus - genetics Hepatitis B virus - isolation & purification Humans Immunology interferon Life Sciences Middle East pegylated interferon Polymerase Chain Reaction Polymerase Chain Reaction - methods quantitative real-time PCR Viral Load Viral Load - methods |
| title | Single-step real-time PCR to quantify hepatitis B virus and distinguish genotype D from non-D genotypes |
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