A universal primer-independent next-generation sequencing approach for investigations of norovirus outbreaks and novel variants
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A universal primer-independent next-generation sequencing approach for investigations of norovirus outbreaks and novel variants. / Fonager, Jannik; Stegger, Marc; Rasmussen, Lasse Dam; Poulsen, Mille Weismann; Ronn, Jesper; Andersen, Paal Skytt; Fischer, Thea Kolsen.
I: Scientific Reports, Bind 7, 813, 2017.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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T1 - A universal primer-independent next-generation sequencing approach for investigations of norovirus outbreaks and novel variants
AU - Fonager, Jannik
AU - Stegger, Marc
AU - Rasmussen, Lasse Dam
AU - Poulsen, Mille Weismann
AU - Ronn, Jesper
AU - Andersen, Paal Skytt
AU - Fischer, Thea Kolsen
PY - 2017
Y1 - 2017
N2 - Norovirus (NoV) is the most common cause of non-bacterial gastroenteritis and is a major agent associated with outbreaks of gastroenteritis. Conventional molecular genotyping analysis of NoV, used for the identification of transmission routes, relies on standard typing methods (STM) by Sanger-sequencing of only a limited part of the NoV genome, which could lead to wrong conclusions. Here, we combined a NoV capture method with next generation sequencing (NGS), which increased the proportion of norovirus reads by ~40 fold compared to NGS without prior capture. Of 15 NoV samples from 6 single-genotype outbreaks, near full-genome coverage (>90%) was obtained from 9 samples. Fourteen polymerase (RdRp) and 15 capsid (cap) genotypes were identified compared to 12 and 13 for the STM, respectively. Analysis of 9 samples from two mixed-genotype outbreaks identified 6 RdRp and 6 cap genotypes (two at >90% NoV genome coverage) compared to 4 and 2 for the STM, respectively. Furthermore, complete or partial sequences from the P2 hypervariable region were obtained from 7 of 8 outbreaks and a new NoV recombinant was identified. This approach could therefore strengthen outbreak investigations and could be applied to other important viruses in stool samples such as hepatitis A and enterovirus.
AB - Norovirus (NoV) is the most common cause of non-bacterial gastroenteritis and is a major agent associated with outbreaks of gastroenteritis. Conventional molecular genotyping analysis of NoV, used for the identification of transmission routes, relies on standard typing methods (STM) by Sanger-sequencing of only a limited part of the NoV genome, which could lead to wrong conclusions. Here, we combined a NoV capture method with next generation sequencing (NGS), which increased the proportion of norovirus reads by ~40 fold compared to NGS without prior capture. Of 15 NoV samples from 6 single-genotype outbreaks, near full-genome coverage (>90%) was obtained from 9 samples. Fourteen polymerase (RdRp) and 15 capsid (cap) genotypes were identified compared to 12 and 13 for the STM, respectively. Analysis of 9 samples from two mixed-genotype outbreaks identified 6 RdRp and 6 cap genotypes (two at >90% NoV genome coverage) compared to 4 and 2 for the STM, respectively. Furthermore, complete or partial sequences from the P2 hypervariable region were obtained from 7 of 8 outbreaks and a new NoV recombinant was identified. This approach could therefore strengthen outbreak investigations and could be applied to other important viruses in stool samples such as hepatitis A and enterovirus.
U2 - 10.1038/s41598-017-00926-x
DO - 10.1038/s41598-017-00926-x
M3 - Journal article
C2 - 28400558
VL - 7
JO - Scientific Reports
JF - Scientific Reports
SN - 2045-2322
M1 - 813
ER -
ID: 179526193