Broadly Reactive and Highly Sensitive Assay for Norwalk-Like Viruses Based on Real-Time Quantitative Reverse Transcription-PCR

Kageyama, Tsutomu; Kojima, Saburo; Shinohara, Michiyo; Uchida, Kenzo; Fukushi, Shuetsu; Hoshino, Fuminori B.; Takeda, Naokazu; Katayama, Kazuhiko

doi:10.1128/jcm.41.4.1548-1557.2003

Cited by 1,271 publications

(904 citation statements)

References 38 publications

Supporting

Mentioning

883

Contrasting

Unclassified

Order By: Relevance

“…Briefly, cDNA synthesis was carried out with 10 l of the RNA in 20 l of the reaction mixture containing 50 pmol random hexamer (Takara, Tokyo, Japan), 1ϫ Superscript III reverse transcriptase buffer (Invitrogen, Carlsbad, CA), 10 mM dithiothreitol (Invitrogen), 0.4 mM each deoxynucleoside triphosphate (Roche, Mannheim, Germany), 1 U RNase inhibitor (Toyobo, Tokyo, Japan), and 10 U Superscript reverse transcriptase III (Invitrogen). RT was performed at 37°C for 15 min, followed by 50°C for 1 h. Real-time RT-PCR was performed as previously described, and the cutoff for positive norovirus specimens was set at Ͼ10 copies per well (13).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Norovirus Infections in Symptomatic and Asymptomatic Food Handlers in Japan

Ozawa¹,

et al. 2007

Self Cite

View full text Add to dashboard Cite

Noroviruses are the leading cause of outbreaks of gastroenteritis in the world. At present, norovirus genogroup II, genotype 4 (GII/4), strains are the most prevalent in many countries. In this study we investigated 55 outbreaks and 35 sporadic cases of norovirus-associated gastroenteritis in food handlers in foodcatering settings between 10 November 2005 and 9 December 2006 in Japan. Stool specimens were collected from both symptomatic and asymptomatic individuals and were examined for norovirus by real-time reverse transcription-PCR; the results were then confirmed by sequence analysis. Norovirus was detected in 449 of 2,376 (19%) specimens. Four genogroup I (GI) genotypes and 12 GII genotypes, including one new GII genotype, were detected. The GII/4 sequences were predominant, accounting for 19 of 55 (35%) outbreaks and 16 of 35 (46%) sporadic cases. Our results also showed that a large number of asymptomatic food handlers were infected with norovirus GII/4 strains. Norovirus GII had a slightly higher mean viral load (1 log unit higher) than norovirus GI, i.e., 3.81 ؋ 10 8 versus 2.79 ؋ 10 7 copies/g of stool. Among norovirus GI strains, GI/4 had the highest mean viral load, whereas among GII strains, GII/4 had the highest mean viral load (2.02 ؋ 10 8 and 7.96 ؋ 10 9 copies/g of stool, respectively). Importantly, we found that asymptomatic individuals had mean viral loads similar to those of symptomatic individuals, which may account for the increased number of infections and the predominance of an asymptomatic transmission route.The positive-sense polyadenylated single-stranded RNA virus family Caliciviridae contains four genera: Norovirus, Sapovirus, Lagovirus, and Vesivirus (1). The prototype strain of human norovirus is the Norwalk virus (NV/Human/US/1968), which was first discovered in an outbreak of gastroenteritis in an elementary school in Norwalk, OH, in 1968 (15). Noroviruses are the leading cause of outbreaks of gastroenteritis in the world; they cause outbreaks in various settings, including hospitals, cruise ships, schools, and restaurants (2,9,12,15,23,24,29). In addition, noroviruses have been detected in environmental samples (e.g., treated and untreated sewage) as well as in contaminated foods such as oysters, shellfish, sandwiches, salads, raspberries, and even ice (7,18,19,26). Numerous molecular epidemiological studies have revealed a global distribution of these viruses (25,27,31).The most widely used method of detecting noroviruses is reverse transcription-PCR (RT-PCR), which has high sensitivity; also, the products can be used for further genetic analysis. Real-time RT-PCR assays have also been developed; they are sensitive, broadly reactive, and rapid for the detection of human noroviruses in clinical stool specimens and environmental samples (13,14,21).As the detection methods become more and more sensitive, the numbers of genogroups and genotypes are expected to increase. One emerging characteristic is that strains have been found to persist in one geographical region, only to disapp...

show abstract

Section: Methodsmentioning

confidence: 99%

“…Real-time RT-PCR assays have also been developed; they are sensitive, broadly reactive, and rapid for the detection of human noroviruses in clinical stool specimens and environmental samples (13,14,21).…”

mentioning

confidence: 99%

Norovirus Infections in Symptomatic and Asymptomatic Food Handlers in Japan

Ozawa¹,

et al. 2007

Self Cite

View full text Add to dashboard Cite

show abstract

“…SybrGreen and TaqMan, have been used for detection of human NoVs from fecal and shellfish samples [69,70,[82][83][84][85][86][87]. These may be used for detection of porcine NoVs and SaVs in the near future.…”

Section: Real-time Rt-pcr-in the Past 2-3 Years Various Real-time Rtmentioning

confidence: 99%

Porcine enteric caliciviruses: Genetic and antigenic relatedness to human caliciviruses, diagnosis and epidemiology

Wang

Costantini

Saif

2007

Vaccine

View full text Add to dashboard Cite

Porcine enteric caliciviruses include sapoviruses and noroviruses. Porcine sapoviruses infect pigs of all ages and cause diarrhea in young pigs, whereas porcine noroviruses were detected exclusively from adult pigs without clinical signs. Importantly, certain porcine norovirus strains were genetically and antigenically related to human noroviruses. This raises public health concerns that pigs may be reservoirs for emergence of epidemic human norovirus strains. This article reviews the discovery of porcine noroviruses and sapoviruses, their classification, diagnosis, epidemiology and genetic and antigenic relatedness to human caliciviruses.

show abstract

“…After incubation at 428C for 1 hr, 5 ml of cDNA were amplified in a semi-nested PCR targeting a 344 bp size of the ORF1/ORF2 genome (region C). The first round of PCR was carried out using COG2F and G2SKR primers, the second round of PCR using primers G2SKF and G2SKR Kageyama et al, 2003]. The region D of VP1 gene was amplified in a single round of PCR according to Vinjé et al [2004].…”

Section: Rt-pcr Amplificationmentioning

confidence: 99%

“…During the outbreak event, NoV was detected by amplification of 344 bp size of the ORF1/ORF2 genome (region C) Kageyama et al, 2003]. Based on the NCBI data bank, all of the NoV GII strains detected were genotyped as GII.4 with a sequence homology of 98% with New Orleans-2009 strain (accession number: GU445325) [Donia et al, 2011].…”

Section: Characterization Of the Outbreak In Ballshmentioning

confidence: 99%

Molecular characterization of norovirus GII strains identified in Albania

Donia

Cenko²,

Divizia

2013

Journal of Medical Virology

View full text Add to dashboard Cite

Noroviruses (NoVs) are considered as the leading cause of diarrheal diseases in all groups of age. In the last decade the number of NoV outbreaks worldwide is increasing. Data published by the systems of NoV surveillance show the GII.4 strain as the human predominant genotype circulating worldwide and new genetic variants of GII.4 were associated with epidemic events. In Albania the economy transformation has damaged significantly the environment and a large circulation of enteric viruses was reported in the past with the presence of NoV among the genotyped strains. This study aimed to characterize, by molecular analysis, the NoV GII strains detected in Albania during two time periods: in 2010 from the outbreak occurred in Ballsh and in 2002–2003 from sporadic cases of diarrhoea. A total of 21 Nov GII strains were characterized. The NoV GII.4 was genotyped more frequently and it was related closely to the pandemic variants recorded in GenBank. During 2002–2003, six NoV GII recombinant strains have been characterized

show abstract

Broadly Reactive and Highly Sensitive Assay for Norwalk-Like Viruses Based on Real-Time Quantitative Reverse Transcription-PCR

Cited by 1,271 publications

References 38 publications

Norovirus Infections in Symptomatic and Asymptomatic Food Handlers in Japan

Norovirus Infections in Symptomatic and Asymptomatic Food Handlers in Japan

Porcine enteric caliciviruses: Genetic and antigenic relatedness to human caliciviruses, diagnosis and epidemiology

Molecular characterization of norovirus GII strains identified in Albania

Contact Info

Product

Resources

About