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      SYBR Green based real-time RT-PCR assay for detection and genotype prediction of bovine noroviruses and assessment of clinical significance in Norway


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          A novel SYBR Green based real-time RT-PCR assay for detection of genogroup III bovine noroviruses (BoNoV) was developed and the assay applied to 419 faecal samples from calves with and without diarrhoea. The samples were obtained from 190 Norwegian dairy and beef herds. BoNoV was detected in 49.6% of the samples from 61.1% of the herds indicating that BoNoV is ubiquitous in Norway. The overall prevalence was not significantly different in diarrhoea and non-diarrhoea samples.

          Analyses of polymerase gene sequences revealed both genotype III/1 and III/2 with genotype III/2 (Newbury2-like) being the most prevalent. Detected capsid sequences were restricted to Newbury2-like and the chimeric Bo/Thirsk10/00/UK strain.

          The RNA polymerase genotypes of the circulating BoNoVs in Norway were predicted by melting temperature analysis.

          Additional data from a challenge experiment suggest that a high proportion of young calves are shedding low levels of BoNoV for a prolonged time after recovering from the associated diarrhoea. The findings may explain some of the discrepancies in detection rates from previous studies and explain why some studies have failed to detect significant prevalence differences between calves with and without diarrhoea. It may also shed new light on some epidemiological aspects of norovirus infections.

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          Most cited references43

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          SplitsTree: analyzing and visualizing evolutionary data.

          D Huson (1998)
          Real evolutionary data often contain a number of different and sometimes conflicting phylogenetic signals, and thus do not always clearly support a unique tree. To address this problem, Bandelt and Dress (Adv. Math., 92, 47-05, 1992) developed the method of split decomposition. For ideal data, this method gives rise to a tree, whereas less ideal data are represented by a tree-like network that may indicate evidence for different and conflicting phylogenies. SplitsTree is an interactive program, for analyzing and visualizing evolutionary data, that implements this approach. It also supports a number of distances transformations, the computation of parsimony splits, spectral analysis and bootstrapping.
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            Is Open Access


            GenBank (R) is a comprehensive database that contains publicly available nucleotide sequences for more than 240 000 named organisms, obtained primarily through submissions from individual laboratories and batch submissions from large-scale sequencing projects. Most submissions are made using the web-based BankIt or standalone Sequin programs and accession numbers are assigned by GenBank staff upon receipt. Daily data exchange with the EMBL Data Library in Europe and the DNA Data Bank of Japan ensures worldwide coverage. GenBank is accessible through NCBI's retrieval system, Entrez, which integrates data from the major DNA and protein sequence databases along with taxonomy, genome, mapping, protein structure and domain information, and the biomedical journal literature via PubMed. BLAST provides sequence similarity searches of GenBank and other sequence databases. Complete bimonthly releases and daily updates of the GenBank database are available by FTP. To access GenBank and its related retrieval and analysis services, begin at the NCBI Homepage ().
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              Norovirus recombination.

              RNA recombination is a significant driving force in viral evolution. Increased awareness of recombination within the genus Norovirus of the family Calicivirus has led to a rise in the identification of norovirus (NoV) recombinants and they are now reported at high frequency. Currently, there is no classification system for recombinant NoVs and a widely accepted recombinant genotyping system is still needed. Consequently, there is duplication in reporting of novel recombinants. This has led to difficulties in defining the number and types of recombinants in circulation. In this study, 120 NoV nucleotide sequences were compiled from the current GenBank database and published literature. NoV recombinants and their recombination breakpoints were identified using three methods: phylogenetic analysis, SimPlot analysis and the maximum chi2 method. A total of 20 NoV recombinant types were identified in circulation worldwide. The recombination point is the ORF1/2 overlap in all isolates except one, which demonstrated a double recombination event within the polymerase region.

                Author and article information

                J Virol Methods
                J. Virol. Methods
                Journal of Virological Methods
                Elsevier B.V.
                8 April 2010
                October 2010
                8 April 2010
                : 169
                : 1
                : 1-7
                [a ]Department of Animal Health, National Veterinary Institute, PO Box 750, Sentrum, NO-0106 Oslo, Norway
                [b ]Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, PO Box 8146 Dep., NO-0033 Oslo, Norway
                [c ]Center for Laboratory Medicine, Akershus University Hospital, NO-1478 Lørenskog, Norway
                Author notes
                [* ]Corresponding author. Tel.: +47 23216326/45661911; fax: +47 23216301. evert.jor@ 123456vetinst.no evert.jor@ 123456gmail.com
                Copyright © 2010 Elsevier B.V. All rights reserved.

                Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

                : 12 October 2009
                : 17 March 2010
                : 30 March 2010

                Microbiology & Virology
                sybr green real-time rt-pcr,genotype prediction,norovirus excretion,challenge experiment


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