Abstract：
:Hepatitis A virus (HAV) is mainly transmitted via contaminated food or water or through person-to-person contact. Here, we describe development and evaluation of a reverse transcription droplet digital PCR (RT-ddPCR) and reverse transcription real-time PCR (RT-qPCR) assay for detection of HAV in food and clinical specimens. The assay was evaluated by assessing limit of detection, precision, matrix effects, sensitivity and quantitative agreement. The 95 % limit of detection (LOD95 %) was 10 % higher for RT-ddPCR than for RT-qPCR. A Bayesian model was used to estimate precision on different target concentrations. From this, we found that RT-ddPCR had somewhat greater precision than RT-qPCR within runs and markedly greater precision between runs. By analysing serum from naturally infected persons and a naturally contaminated food sample, we found that the two methods agreed well in quantification and had comparable sensitivities. Tests with artificially contaminated food samples revealed that neither RT-ddPCR nor RT-qPCR was severely inhibited by presence of oysters, raspberries, blueberries or leafy-green vegetables. For this assay, we conclude that RT-qPCR should be considered if rapid, qualitative detection is the main interest and that RT-ddPCR should be considered if precise quantification is the main interest. The high precision of RT-ddPCR allows for detection of small changes in viral concentration over time, which has direct implications for both food control and clinical studies.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
Persson S,Alm E,Karlsson M,Enkirch T,Norder H,Eriksson R,Simonsson M,Ellström Pdoi
10.1016/j.jviromet.2020.114010subject
Has Abstractpub_date
2021-02-01 00:00:00pages
114010eissn
0166-0934issn
1879-0984pii
S0166-0934(20)30262-7journal_volume
288pub_type
杂志文章abstract：:The human coronavirus NL63 was identified in 2004 and subsequent studies showed its worldwide distribution. Infection with this pathogen is associated with upper and lower respiratory tract diseases of mild to moderate severity. Furthermore, HCoV-NL63 is the main cause of croup in children. Within this study an optima...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.04.001
更新日期：2014-09-01 00:00:00
abstract：:Receptor-mediated endocytosis was employed for a highly efficient transport of oligodeoxynucleotides into hepatoma cell line PLC/PRF/5. The oligodeoxynucleotides were bound to the asialofetuin-poly-L-lysine conjugate and this complex was internalized by the cells via asialoglycoprotein receptor, an endocytic receptor ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(93)90181-p
更新日期：1993-04-01 00:00:00
abstract：:Representatives from several different virus families (Baculoviridae, Herpesviridae, Myoviridae, Phycodnaviridae, Picornaviridae, Podoviridae, Retroviridae, and Siphoviridae) were stained using a variety of highly fluorescent nucleic acid specific dyes (SYBR Green I, SYBR Green II, OliGreen, PicoGreen) and examined us...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00167-6
更新日期：2000-03-01 00:00:00
abstract：:Tomato necrotic stunt virus (ToNStV) is an emerging potyvirus that causes severe stunting to infected tomato plants. A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for sensitive detection of ToNStV. The sensitivity of RT-LAMP was comparable to that of conventional RT-PCR, ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.01.017
更新日期：2014-05-01 00:00:00
abstract：:Atypical bovine pestiviruses are related antigenically and phylogenetically to bovine viral diarrhea viruses (BVDV-1 and BVDV-2), and may cause the same clinical manifestations in animals. Glycoprotein E(rns) of an atypical bovine pestivirus Th/04_KhonKaen was produced in a baculovirus expression system and was purifi...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2012.06.017
更新日期：2012-11-01 00:00:00
abstract：:Adaptation through fixation of spontaneous mutations in the viral genome is considered to be one of the important factors that enable recurrent West Nile virus (WNV) outbreaks in the U.S. Genetic variations can alter viral phenotype and virulence, and degrade the performance of diagnostic and screening assays, vaccine...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2008.09.015
更新日期：2008-12-01 00:00:00
abstract：:A TaqMan real-time RT-PCR was developed to detect and quantify RNA-targets from the non-circulative, non-persistently transmitted Plum pox virus (PPV) in individual fresh or aphids captured previously and squashed on paper. Reliable quantitation ranged from 40 up to 4 x 10(8) copies of control transcripts. This techni...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2005.05.011
更新日期：2005-09-01 00:00:00
abstract：:All human herpesviruses cause chronic infections in which latent virus is periodically reactivated. Persistently active infections are uncommon, however, and occur exclusively in individuals whose immune systems fail to control virus multiplication and spread. This paper summarizes the management of these unusual infe...
journal_title：Journal of virological methods
pub_type： 杂志文章,评审
doi：10.1016/0166-0934(88)90076-6
更新日期：1988-09-01 00:00:00
abstract：:Ultra-deep pyrosequencing (UDPS) of targeted amplicons allows to determine a large number of individual sequence reads from a single PCR product, and this approach is thus extremely valuable for analysis of mixed viral populations. A mixture of genetically distinct DNA templates, however, may lead to the generation of...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2010.07.040
更新日期：2010-10-01 00:00:00
abstract：:Research progress on the pleiotropic effects of interferons (IFN) has thus far required detecting responses by weak IFN signals. The activity of 2',5'-oligoadenylate synthetase (2-5OAS) is a valuable indicator in the prognosis and IFN treatment of patients with viral diseases such as hepatitis B and C. Although serum ...
journal_title：Journal of virological methods
pub_type： 临床试验,杂志文章
doi：10.1016/j.jviromet.2006.05.005
更新日期：2006-09-01 00:00:00
abstract：:Lujo virus is an emerging arenavirus circulating in Southern Africa. Although to date there has only been a single outbreak of the novel haemorrhagic disease resulting from human infection with this virus, the case-fatality rate of exposed individuals, including nosocomial transmission, was 80%. The ability to identif...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2013.09.006
更新日期：2014-01-01 00:00:00
abstract：:In order to improve the accuracy for detecting human foamy virus (HFV), an indicator cell line was established by co-transfecting baby hamster kidney-21 cells with two plasmids: one containing a G418 antibiotic resistance marker and the other including the luc gene which was placed downstream of the inducible HFV long...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(01)00285-3
更新日期：2001-05-01 00:00:00
abstract：:A genetically modified cell line (Vero-ICP10-EGFP) was constructed for detection of herpes simplex virus (HSV) by a simple, rapid and direct method. The cell line was developed by stable transfection of Vero cell with a plasmid encoding the green fluorescent protein (GFP) driven by the promoter of the HSV-2 ICP10 gene...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(00)00234-2
更新日期：2000-11-01 00:00:00
abstract：:HIV-1 RNA viral load is the preferred tool to monitor virological failure during antiretroviral therapy (ART) exposure. Timely detection of virological failure can reduce the prevalence and complexity of HIV-1 drug resistance. This field evaluation further characterizes a two-step approach to identify virological fail...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2013.08.015
更新日期：2013-12-01 00:00:00
abstract：:IgM antibody capture radioimmunoassay (MACRIA) and enzyme immunoassay (MACEIA) were compared with immunofluorescence (IF) for detecting specific IgM antibody in 99 sera from 76 infants with confirmed congenital rubella, and 61 sera from a comparative group of 59 infants who had miscellaneous abnormalities but in whom ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(82)90055-6
更新日期：1982-08-01 00:00:00
abstract：:An efficient method for isolation of virus mutants with antigenically altered proteins is described. The method is based on the separation of viruses with wild-type and antigenically altered proteins by affinity chromatography using monoclonal antibodies (MAbs). A nonessential glycoprotein E (gE) of Aujeszky's disease...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(98)00137-2
更新日期：1999-01-01 00:00:00
abstract：:Maize chlorotic mottle virus (MCMV) causes corn lethal necrosis disease, and can be transmitted through infected maize seeds. It remains a challenge to detect this virus in the seeds to prevent its introduction and infection. For this purpose, a real-time TaqMan RT-PCR procedure for efficient detection of MCMV was dev...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2010.11.002
更新日期：2011-01-01 00:00:00
abstract：:A quantitative PCR assay for the detection of HIV-1 nucleic acids is described. The assay is based on a competitive internal standard nucleic acid which can be discriminated from target sequences by the presence of a new restriction enzyme site. The method was used to quantitate plasmid molecules containing HIV-1 sequ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(91)90095-h
更新日期：1991-09-01 00:00:00
abstract：:Arboviruses in the genus Flavivirus are major causes of human disease worldwide, and yet they are widely under reported. This is partially due to the classical methods of detection, using virologic or immunologic techniques, which typically lack sensitivity and specificity or require biological containment facilities ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2007.05.001
更新日期：2007-10-01 00:00:00
abstract：:A real-time RT-PCR assay using SYBR Green was developed for specific and reliable quantitative detection of Citrus tristeza virus (CTV) in infected plants. A general primer set designed from conserved sequences in ORFs 1b and 2 enabled amplification of the genomic RNA (gRNA) while excluding most subgenomic and defecti...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2007.05.011
更新日期：2007-11-01 00:00:00
abstract：:A rapid and sensitive method for detecting HIV-1 DNA sequences amplified by polymerase chain reaction (PCR) is described. The method uses solution phase hybridization for rapid and efficient annealing between digoxigenin-labeled targets and biotinylated capture probes. Hybrids containing biotin are captured onto strep...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(92)90174-c
更新日期：1992-07-01 00:00:00
abstract：:During a 12-month period, two methods for detection of cytomegalovirus (CMV) in 1624 clinical specimens were compared: (1) centrifugal inoculation of MRC-5 cells on coverslips in 24-well plates and staining with a monoclonal antibody to CMV early nuclear antigen after incubation for 40 h (EA assay), and (2) convention...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(87)90082-6
更新日期：1987-12-01 00:00:00
abstract：:A mRNA in situ hybridization method was developed to detect mRNA of Epstein--Barr virus-determined nuclear antigen-2 (EBNA2). Strong in situ hybridization signals were detected in EBNA2-transfected cells with the antisense probe but not with the sense probe of this mRNA. Hybridization signals were also found in formal...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(00)00271-8
更新日期：2001-04-01 00:00:00
abstract：:Expression of recombinant beak and feather disease virus (BFDV) capsid-associated protein (Cap) has relied on inefficient techniques that typically produce low yields or use specialized expression systems, which greatly increase the cost and expertise required for mass production. An Escherichia coli system was used t...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2013.01.020
更新日期：2013-04-01 00:00:00
abstract：:Cervical carcinoma is one of the major consequences of human papillomavirus (HPV) infections. Although HPV infections of cervix do not always progress to cancer, 90% cases of cervical cancer have been found associated with high risk HPV (hrHPV) infection. Usually, HPV infection is asymptomatic; however, this asymptoma...
journal_title：Journal of virological methods
pub_type： 杂志文章,评审
doi：10.1016/j.jviromet.2015.06.015
更新日期：2015-09-15 00:00:00
abstract：:Commercially marketed kits are now available for rapid viral assay of clinical specimens. This study was conducted to determine the suitability of these kits for use in environmental testing. Eight rotavirus kits and one enteric adenovirus kit were screened for sensitivity using simian rotavirus SA11, human rotavirus ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(93)90098-c
更新日期：1993-12-15 00:00:00
abstract：:Hepatitis B virus (HBV) is hyperendemic to southern Africa, with genotype A of HBV being the predominant genotype, and subgenotype A1 prevailing. Infection with this subgenotype is associated with rapid disease progression, and high frequency of hepatocellular carcinoma development. The objectives of our study was to ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.03.015
更新日期：2014-07-01 00:00:00
abstract：:An international collaborative study was performed to evaluate a set of PCR reference reagents for HIV diagnosis. Twenty-six laboratories from 9 countries analysed a proficiency panel of 10 coded DNA samples using the PCR reference reagents and protocols. For comparison, these coded samples were then assessed using a ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(92)90018-9
更新日期：1992-04-01 00:00:00
abstract：:We assessed the performance of Abbott RealTime CMV assay (ARC) compared to Roche Cobas Amplicor CMV Monitor Test (RCM) for quantification of CMV in plasma of transplant patients. Commercial panels were used to test linearity, precision and interference and 83 clinical samples were used for the accuracy and precision a...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2015.08.010
更新日期：2015-12-01 00:00:00
abstract：:Plum pox virus (PPV) causes the most destructive viral disease known as plum pox or Sharka disease in stone fruit trees. As an important regulated pathogen, detection of PPV is thus of critical importance to quarantine and eradication of the spreading disease. In this study, the innovative development of two AmplifyRP...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.06.026
更新日期：2014-10-01 00:00:00
