Objective: To analyze the genetic characterization of norovirus isolated in an outbreak of gastroenteritis in Jiangsu province. Methods: Extracted viral RNA from the swab samples of cases of acute gastroenteritis outbreak in Jiangsu province on December 16-27, 2016 was reversely transcribed to cDNA, and partial RNA-dependent RNA polymerase sequence and complete capsid sequence (VP1) were amplified by RT-PCR. Amplification products were sequenced for the analysis of genetic characteristics. Results: Based on sequence alignment, the variant shared a high level of identity with the strain GⅡ.g isolated in Spain and Finland (98.7%) in the RNA-dependent RNA polymerase region, and with the strain GⅡ.1 isolated in American (99.4%) in the VP1. The recombination was determined by using software Simplot, and the breakpoint of recombination was located in the ORF1/2 overlap region at position 5 106 of VP1. The result of amino acids alignment in capsid region showed that there were no mutations in the amino acids of the predicted epitopes and receptor binding site Ⅰ-Ⅲ, but a unique amino acid change was detected at position 132 (N-S). Conclusion: The norovirus isolated in the outbreak of gastroenteritis in Jiangsu province was a rare recombinant norovirus variant GⅡ.g-GⅡ.1.
Caliciviridae Infections/epidemiology* ; Capsid Proteins ; Disease Outbreaks ; Gastroenteritis/epidemiology* ; Genotype ; Humans ; Norovirus/isolation & purification* ; Phylogeny ; RNA, Viral/genetics* ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA

Objective: To understand the epidemiologic characteristics of outbreaks, caused by norovirus-GⅡ.2、GⅡ.17 and GⅡ.4/Sydney in Guangdong Province from 2013 to 2017 and to provide scientific evidence for epidemic prevention and control. Methods: Incidence data of norovirus outbreaks in Guangdong from January 1(st) 2013 to November 30(th) 2017 were collected from Public Health Emergency Management Information System. RT-PCR was performed for every case of each outbreak to detect norovirus nucleic acid and gene sequencing was conducted to identify the genotype of norovirus. Characteristics of norovirus GⅡ.2, GⅡ.17 and GⅡ.4/Sydney outbreaks were analyzed. Directly standardized method was used to calculate the proportion of symtoms as diarrhea and vomitting. Results: From January 1(st) 2013 to November 30(th) 2017, a total of 167 norovirus outbreaks were reported in Guangdong, and 115 outbreaks were caused by norovirus GⅡ.2, GⅡ.17 and GⅡ.4/Sydney respectively. The outbreaks caused by norovirus GⅡ.2 accounted for 39.68% (25/63) in primary schools, 28.57% (18/63) in child care settings, 25.40% (16/63) in middle schools and 6.35% (4/63) in universities. Outbreaks caused by norovirus GⅡ.17 accounted for 41.03% (16/39) in middle schools, 20.51% (8/39) at workplaces, 15.38% (6/39) in primary schools, 12.82% (5/39) in universities, 5.13% (2/39) in communities and child care settings respectively. The outbreaks caused by norovirus GⅡ.4/Sydney accounted for 53.85% (7/13) in universities, 15.38% (2/13) in child care settings and at workplaces respectively, 7.69%(1/13) in primary schools and middle schools respectively. The outbreaks caused by norovirus GⅡ.2 had 77.78% (49/63) of contact transmission, 17.46% (11/63) of food-borne transmission. The outbreaks caused by norovirus GⅡ.17 showed 53.85% (21/39) of food-borne transmission, 15.38% (6/39) of contract transmission, 12.82% (5/39) of water-borne transmission. The outbreaks caused by norovirus GⅡ.4/Sydney had 53.85% (7/13) of food-borne transmission, 38.46% (5/13) of the contact transmission. In terms of the clinical manifestations, the standardized proportion of vomit was 73.76% and the proportion of diarrhea was 42.85% in cases infected with norovirus GⅡ.2, the proportion of standardized of vomit was 76.37% and the proportion of diarrhea was 51.40% in cases infected with norovirus GⅡ.17, with the standardized proportion of vomit was 54.10% and the proportion of diarrhea was 55.95% in cases infected with norovirus GⅡ.4/Sydney. Conclusions: The outbreaks caused by norovirus GⅡ.2 through contact transmission mainly occurred in primary schools, child care settings and middle schools. The outbreaks caused by norovirus GⅡ.17 through food-borne transmission mainly occurred in middle schools and at workplaces. The outbreaks caused by norovirus GⅡ.4/Sydney food-borne transmission and contact mainly occurred in universities.
Adolescent ; Caliciviridae Infections/epidemiology* ; Child ; Child, Preschool ; Diarrhea/etiology* ; Disease Outbreaks ; Epidemics ; Gastroenteritis/epidemiology* ; Genotype ; Humans ; Norovirus/isolation & purification* ; Reverse Transcriptase Polymerase Chain Reaction ; Sentinel Surveillance ; Vomiting/etiology*

Objective: To understand the epidemiological and molecular characteristics of a norovirus- borne outbreak caused by GⅡ.4 Sydney 2012 in a university of Guangzhou to provide evidence for the prevention and control strategy on norovirus-caused epidemics. Methods: A self-designed questionnaire was used to collect clinical information from the patients as well as other data related to the epidemic. Pathogen detections were performed through anal swab specimens from the patients, kitchen workers and samples from the environment. Positive samples were further sequenced for phylogenetic analysis. A case-control study was employed to identify the risk factors related to this outbreak. Results: A total of 226 cases of norovirus-borne infection were identified between September 17 and 21, 2017, including 223 students, with an attack rate of 0.73% (223/30 711), and 3 kitchen workers. Students staying in the A dormitory area had the highest attack rate (1.73%, 164/9 459). No clustering was found in different colleges or classes. Results from the case-control study revealed that people who ate at the canteen in A dormitory area during September 18 to 20 was at risk for the onset of illness (OR=10.75, 95%CI: 5.56-20.79). The highest risk was related to the dinner on September 18. Another significant risk factor (OR=3.65, 95%CI: 1.92-6.94) was close personal contact in the same room of the dorm. The 3 norovirus infected kitchen workers were all from the canteen in A dormitory area where the positive rate of norovirus identified in kitchen workers was 26.67% (12/45). Positive samples were sequenced and sub-typed with results showing that the GⅡ.4 Sydney 2012 variant and the nucleotide sequences of cases and kitchen workers were 100% identical. Conclusions: The outbreak was caused by norovirus GⅡ.4 Sydney 2012 variant at campus. Similar outbreaks had been seen since 2013, with the routes of transmission most likely due to food-borne or personal contact.
Adolescent ; Adult ; Caliciviridae Infections/epidemiology* ; Case-Control Studies ; China/epidemiology* ; Disease Outbreaks ; Female ; Foodborne Diseases/virology* ; Gastroenteritis/virology* ; Humans ; Male ; Norovirus/isolation & purification* ; Phylogeny ; Surveys and Questionnaires

Noroviruses are the leading cause of epidemic gastroenteritis, including foodborne outbreak, in Korea. The prevalence of human noroviruses was studied in diarrheal stool samples of patients with acute gastroenteritis by conventional duplex reverse transcription (RT)-PCR. Diarrheal stool samples were collected from 1,685 patients from the local hospitals in Seoul. The prevalence of the noroviruses was 22.8% (222/972 patients) in 2012 and 11.2% (80/713 patients) in 2013, with a total of 17.9% (302/1,685 patients). Genotyping was performed on 302 norovirus-positive stool samples to reveal 5.6% prevalence of genogroup I (GI) (17/302) and 94.4% prevalence of genogroup II (GII) (285/302). The patients with norovirus-associated acute gastroenteritis mostly showed prevalence of GII norovirus, especially GII.4 (64.6%; 195/302).
Acute Disease ; Feces/virology ; Gastroenteritis/*diagnosis/epidemiology/virology ; Genotype ; Humans ; Norovirus/*genetics/isolation & purification ; Prevalence ; RNA, Viral/genetics/metabolism ; Real-Time Polymerase Chain Reaction

Because of limited viral replication and lack of cytopathic effect in cell culture, a new PCR-based rapid seroneutralization assay for detection of GII.4 norovirus neutralized antibodies was developed with serum samples from acute-phase patients, convalescent-phase patients and healthy controls. According to this study, neutralizing antibodies were detected in 100% of convalescent-phase sera, and in 2.5% of healthy controls sera. However, all of the acute-phase serum samples could not neutralize virus efficiently. Compared to the results from ELISA (96.2% at sensitivity and 80% at specificity), the present in vitro neutralization assay is more specific and more sensitive.
Antibodies, Neutralizing ; immunology ; Base Sequence ; Caliciviridae Infections ; diagnosis ; virology ; Case-Control Studies ; Cell Line ; DNA Primers ; Enzyme-Linked Immunosorbent Assay ; Gastroenteritis ; diagnosis ; virology ; Humans ; Norovirus ; immunology ; isolation & purification ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

This article reviewed the researches proceeding on the contamination and detection of the foodborne pathogen noroviruses (NoVs) in fresh produce, which involved the NoVs contaminations in fresh produce, the special attachment of NoVs in fresh produce, the NoVs outbreaks associated with fresh produce and the NoVs detection in fresh produce. There had been an increase in reported infectious disease risks associated with the consumptions of fresh produce for recent 30 years. Because the NoVs, as a primary cause of viral gastroenteritis thoughout the world, were highly contagious, had a low infectious dose, and were persistent in the environment. And also the methods for NoVs detection in food had significantly developed over the last 15 years. Currently NoVs were the most common pathogen accounting for 40% of outbreaks associated with fresh produce (i. e., fruits and vegetables). Data from outbreaks investigations verified fresh produce as the high risk food products for NoVs. The fresh produce were typically eaten raw with no thermal processing, can be contaminated at any step during production and processing from faecally polluted water and fertilizers, the poor hygiene practices by food handlers and the cross-contamination. The attachment of NoVs to the fresh produce was due to the physio-chemical factors of virus protein coat, the special attachment to different fresh produce, and the possibility for internalization of NoVs. It might provide answers to why those high risk foods were more frequently implicated (i. e., lettuce and raspberries). According to the data of foodborne NoVs outbreaks which were associated with fresh produce from EU countries and the USA, the outbreaks in EU countries were mainly associated with NoVs contaminated raspberries and lettuce, while in USA which were associated with NoVs contaminated lettuce. Unfortunately, there were no NoVs detection methods for fresh produce or the data of foodborne NoVs outbreaks which were associated with fresh produce in China. That made it difficult to analyze the NoVs contamination situation in China. The heterogeneous distributions of presumably low levels of virus, which presented in contaminated fresh produce, also made it difficult to detect NoVs. To solve this problem, different sampling methods, viral elution methods and RT-qPCR methods were chosen. For example, according to the isoelectric point of NoVs particles, high pH and high ionic strength solution could be used as means for releasing NoVs. For the elution from acidic fruit, the buffer capacity and the virus recovery could be increased by the addition of tris-HCl. When analyzing pectin containing raspberries or strawberries, the viral elution usually incubated with pectinase at neutral pH to avoid from foaming jelly. In this paper, the latest ISO standard for NoV detection in food and the new approaches for NoV detection were also reviewed to provide references for domestic researches. It was necessary to establish and develop domestic methods for NoV detection in fresh produce, especially the different NoV conventional molecular detection methods with corresponding NoV extraction methods, which targeted to the different adsorption characteristics of different fruits and vegetables, in order to strengthen the national food safety monitoring.
Food Analysis ; methods ; Food Contamination ; analysis ; Foodborne Diseases ; virology ; Fruit ; virology ; Gastroenteritis ; virology ; Humans ; Norovirus ; classification ; genetics ; isolation & purification ; physiology ; Vegetables ; virology

Noroviruses (NoVs) are one of the most important foodborne viral pathogens worldwide. Shellfish are the most common carriers of NoVs as they can concentrate and accumulate large amounts of the virus through filter feeding from seawater. Shellfish may selectively accumulate NoVs with different genotypes, and this bioaccumulation may depend on the season and location. Our previous studies found various histo-blood group antigens (HBGAs) in shellfish tissues. While HBGAs might be the main reason that NoVs are accumulated in shellfish, the detailed mechanism behind NoV concentration and bioaccumulation in shellfish is not clear. Here we review current research into NoV bioaccumulation, tissue distribution, seasonal variation, and binding mechanism in shellfish. This paper may provide insight into controlling NoV transmission and decreasing the risks associated with shellfish consumption.
Animals ; Caliciviridae Infections ; transmission ; virology ; Food Contamination ; analysis ; Foodborne Diseases ; virology ; Humans ; Norovirus ; classification ; genetics ; isolation & purification ; physiology ; Shellfish ; virology

OBJECTIVETo monitor the epidemiology of norovirus infection in diarrheal children in Shanghai between 2009 and 2011 and characterize the genotypes of norovirus strains.

METHODThe stool samples were collected from children visiting outpatient clinic for acute non-dysenteric diarrhea between 2009 and 2011.One step real-time RT-PCR was used for screening norovirus genogroups GI and GII. The genotypes of norovirus genogroup GII were classified based on the nucleotide sequences of both partial capsid and polymerase fragments.

RESULTA total of 2 288 outpatient children with acute diarrhea were included in this study, out of whom, 531 (23.1%) were positive for norovirus in the fecal specimens based on real-time RT-PCR test.Norovirus was prevalent throughout the year and an increased activity of norovirus infection was usually observed between July and October. Children <4 years of age accounted for 95.2% of norovirus-infected cases, and the detection rate of norovirus was significantly higher in diarrheal children <4 years than in those ≥ 4 years (24.4% vs. 10.7%,χ(2) = 10.66, P < 0.05).Of 531 norovirus-positive specimens, 4 (1.7%) were positive for genogroup GI and 527 (98.3%) positive for genogroup GII. Seven distinct capsid genotypes were identified in 234 norovirus strains, including 153 (64.4%) GII.4 (9 belonging to 2010 variants and 145 belonging to 2006b variants), 66 (27.6%) GII.3, 7 (2.9%) GII.2, 6 (2.5%) GII.6, 4 (1.7%) GII.12, 1 (0.4%) GII.7 and GII.14 in each. Seven polymerase genotypes were identified in 244 norovirus strains, including 189 (77.5%) GII.4 (14 belonging to 2010 variants and 175 belonging to 2006b variants), 47 (19.3%) GII.12, 2 (0.8%) GII.16, GII.b and GII.g in each, 1 (0.4%) GII.2 and GII.6 in each. A new GII.4-2010 (New Orleans) variant was first detected in June 2010 and sporadically circulated afterwards.Of 198 norovirus strains in which both polymerase and capsid genotypes were determined, 56 showed discordant results, indicating potential norovirus recombinants. The common discordant combinations of the polymerase and capsid genotypes were GII.12/GII.3 (69.6%) and GII.4/GII.3 (8.9%).

CONCLUSIONNorovirus is a common causative agent responsible for diarrhea in Shanghai children over the three years and norovirus-associated diarrhea was epidemic year round with high activity in late summer and autumn in Shanghai.Infants and young children are susceptible to norovirus infection. The circulating norovirus showed genetic diversity. The GII.4-2006b variant continued to predominate in Shanghai during the period of 2009-2011 despite the emergence of the novel GII.4-2010 (New Orleans) variant.

Adolescent ; Caliciviridae Infections ; epidemiology ; virology ; Capsid Proteins ; genetics ; Child ; Child, Preschool ; China ; epidemiology ; Diarrhea ; epidemiology ; virology ; Feces ; virology ; Female ; Gastroenteritis ; epidemiology ; virology ; Genetic Variation ; Genotype ; Humans ; Infant ; Male ; Molecular Epidemiology ; Norovirus ; classification ; genetics ; isolation & purification ; Prevalence ; RNA, Viral ; genetics ; isolation & purification ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA

Murine norovirus (MNV) was first discovered in mice in 2003. MNV is a member of the genus Norovirus in the family Caliciviridae. It is one of the most important and prevalent pathogens of laboratory mice, and almost all mouse strains are susceptible to MNV infection. In this study, a MNV strain was isolated from the cecal contents of infected mice and identified by the cytopathic effect (CPE) assay, virus plaque assay, 50% tissue culture infectious dose (TCID50) assay, electron microscopy, indirect immunofluorescence assay (IFA) and nucleotide sequencing. On infection, the RAW264.7 cell line showed obvious cytopathic effects within 24 to 48 hours post-inoculation, as infected cells became rounded, bright and shrunken, with ultimate disintegration of the cell sheet. After the isolation of the MNV virus, the virus was plaque-purified in RAW264.7 cells. The TCID50 of the virus was 10(5.25/0.1 mL. Electron microscopic observations of the purified virus showed the presence of spherical and non-enveloped viral particles that were 30 to 35 nm in diameter. According to the identification results, the isolate was named as MNV Guangzhou/K162/09/CHN. Thereafter, five overlapping gene fragments that covered the entire open reading frame (ORF) were amplified by RT-PCR, and the 3'-untranslated region (UTR) and 5'-UTR were amplified using the 3'-rapid amplification of cDNA ends (RACE) and the 5'-RACE method, respectively. Each of the gene fragments were cloned and sequenced, and whole genome sequences of the strain were obtained by assembling the cDNA fragment sequences. The results showed that the length of the complete genome was 7 380 nucleotides (GenBank accession number: HQ317203). The comparison of nucleotide and deduced amino acid sequences of the isolate was performed against other MNV strains in the GenBank database. A phylogenetic tree based on VP1 nucleotide sequences was constructed using MEGA5.0 software. The homology of nucleotides between the MNV Guangzhou/K162/09/CHN strain and other MNV isolates ranged from 87.4% to 89.7%. Phylogenetic analysis showed that there was a close genetic relationship between the Guangzhou/K162/09/CHN strain and MNV strains isolated from Japan (S7-P2 and S7-PP3 isolates), Korea (K4 isolate), and Germany (Berlin/04/06/DE and Berlin/05/06/DE isolates). This is the first report of the isolation and identification of MNV in China, and the first report of the genetic analysis of its complete genome.
Animals ; Caliciviridae Infections ; veterinary ; virology ; Mice ; Molecular Sequence Data ; Norovirus ; classification ; genetics ; isolation & purification ; physiology ; Open Reading Frames ; Phylogeny ; Rodent Diseases ; virology ; Sequence Homology, Amino Acid ; Viral Proteins ; chemistry ; genetics

To describe the epidemiological characteristics of norovirus (NOV) associated acute gastroenteritis in Shanghai and characterize the evolution pattern of circulating strains. From March 2012 to February 2013, 502 stool specimens were collected from adult (> or = 16 years) outpatients who visited either of the two sentinel hospitals in Shanghai for acute gastroenteritis. Molecular detection and genotyping of NoV were performed and the phylogenetic relationship of the circulating strains has also been comprehensively analyzed. The epidemics level of GI NoV was low throughout the surveillance period, with the positive rate of 3.78% (19 cases), and no seasonality of GI NoV infection could be distinguished. For GII genogroup, higher epidemics in adults in Shanghai, with the detection rate of 17.13% (86 cases), were observed. And relatively high epidemics of GII NoV infection were spotted between October and December in 2012. The frequency of NoV associated acute gastroenteritis in older people is significantly higher than that in young individuals (P < 0.05). Sequencing and genotyping analysis revealed that the high epidemics of GII NoV infection between October and December in 2012 is associated with the emergence of a novel GII.4 norovirus strain, termed Sydney_2012. Sequence analysis also demonstrated that this was a recombinant virus between a GII.e polymerase and GII.4 capsid, which has also been the dominant circulating strain in Shanghai. In 2012, a new GII.4 variant, termed Sydney_2012, emerged in Shanghai and caused high epidemics of acute gastroenteritis during late autumn and winter.
Adolescent ; Adult ; Aged ; Caliciviridae Infections ; epidemiology ; virology ; China ; epidemiology ; Disease Outbreaks ; Female ; Gastroenteritis ; epidemiology ; virology ; Genotype ; Humans ; Male ; Middle Aged ; Norovirus ; chemistry ; classification ; genetics ; isolation & purification ; Phylogeny ; Viral Envelope Proteins ; chemistry ; genetics ; Young Adult