title: Determination of cut-off cycle threshold values in routine RT-PCR assays to assist differential diagnosis of norovirus in children hospitalized for acute gastroenteritis.
abstract: Norovirus (NV) is an important cause of acute gastroenteritis in children, but is also frequently detected in asymptomatic children, which complicates the interpretation of NV detection results in both the clinical setting and population prevalence studies. A total of 807 faecal samples from children aged <5 years hospitalized for acute gastroenteritis were collected in Thai Binh, Vietnam, from January 2011 to September 2012. Real-time RT-PCR was used to detect and quantify NV-RNA in clinical samples. A bimodal distribution of cycle threshold (Ct) values was observed in which the lower peak was assumed to represent cases for which NV was the causal agent of diarrhoea, whereas the higher peak was assumed to represent cases involving an alternative pathogen other than NV. Under these assumptions, we applied finite-mixture modelling to estimate a threshold of Ct <21·36 (95% confidence interval 20·29-22·46) to distinguish NV-positive patients for which NV was the likely cause of diarrhoea. We evaluated the validity of the threshold through comparisons with NV antigen ELISA results, and comparisons of Ct values in patients co-infected with rotavirus. We conclude that the use of an appropriate cut-off value in the interpretation of NV real-time RT-PCR results may improve differential diagnosis of enteric infections, and could contribute to improved estimates of the burden of NV disease.
authors.affiliation: The National Institute of Hygiene and Epidemiology,Hanoi,Vietnam.
authors.forename: N V
authors.initials: NV
authors.lastname: Trang
authors.affiliation: MIVEGEC (UM1-UM2-CNRS 5290-IRD 224),Centre de Recherche IRD,Montpellier,France.
authors.forename: M
authors.initials: M
authors.lastname: Choisy
authors.affiliation: Division of Molecular Epidemiology,Graduate School of Biomedical Science,Nagasaki University,Japan.
authors.forename: T
authors.initials: T
authors.lastname: Nakagomi
authors.affiliation: Thai Binh Paediatric Hospital,Vietnam.
authors.forename: N T M
authors.initials: NT
authors.lastname: Chinh
authors.affiliation: Division of Molecular Epidemiology,Graduate School of Biomedical Science,Nagasaki University,Japan.
authors.forename: Y H
authors.initials: YH
authors.lastname: Doan
authors.affiliation: Centre for Infectious Disease Research in Asia and Africa,Institute of Tropical Medicine,Nagasaki University,Japan.
authors.forename: T
authors.initials: T
authors.lastname: Yamashiro
authors.affiliation: Oxford University Clinical Research Unit,Hanoi,Vietnam.
authors.forename: J E
authors.initials: JE
authors.lastname: Bryant
authors.affiliation: Division of Molecular Epidemiology,Graduate School of Biomedical Science,Nagasaki University,Japan.
authors.forename: O
authors.initials: O
authors.lastname: Nakagomi
authors.affiliation: The National Institute of Hygiene and Epidemiology,Hanoi,Vietnam.
authors.forename: D D
authors.initials: DD
authors.lastname: Anh
mesh_terms: D000956:Antigens, Viral / Q000032:analysis*; D017250:Caliciviridae Infections / Q000175:diagnosis* / Q000453:epidemiology / Q000821:virology; D002675:Child, Preschool; D003937:Diagnosis, Differential; D004797:Enzyme-Linked Immunosorbent Assay; D005243:Feces / Q000821:virology; D005260:Female; D005759:Gastroenteritis / Q000175:diagnosis* / Q000453:epidemiology / Q000821:virology; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D029322:Norovirus / Q000235:genetics* / Q000276:immunology; D015995:Prevalence; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D012016:Reference Values; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012401:Rotavirus / Q000235:genetics* / Q000276:immunology; D012400:Rotavirus Infections / Q000175:diagnosis* / Q000453:epidemiology / Q000821:virology; D014744:Vietnam / Q000453:epidemiology
chemical_list: D000956:Antigens, Viral
keywords: Caliciviruses; diarrhoea; estimating disease prevalence; gastrointestinal infections; modelling