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Corrigendum: Updated classification of norovirus genogroups and genotypes
Updated classification of norovirus genogroups and genotypes
Noroviruses are genetically diverse RNA viruses associated with acute gastroenteritis in mammalian hosts. Phylogenetically they can be segregated into different genogroups as well as P (polymerase)-groups and further into genotypes and P-types based on amino acid diversity of the complete VP1 gene and nucleotide diversity of the RNA-dependent RNA polymerase (RdRp) region of ORF1 respectively. In recent years several new noroviruses have been reported that warrant an update of the existing classification scheme. Using previously described 2× standard deviation (sd) criteria to group sequences into separate clusters we expanded the number of genogroups to 10 (GI-GX) and the number of genotypes to 49 (9 GI 27 GII 3 GIII 2 GIV 2 GV 2 GVI and 1 genotype each for GVII GVIII GIX [formerly GII.15] and GX). Viruses for which currently only one sequence is available in public databases were classified into tentative new genogroups (GNA1 and GNA2) and genotypes (GII.NA1 GII.NA2 and GIV.NA1) with their definitive assignment awaiting additional related sequences. Based on nucleotide diversity in the RdRp region noroviruses can be divided into 60 P-types (14 GI 37 GII 2 GIII 1 GIV 2 GV 2 GVI 1 GVII and 1 GX) 2 tentative P-groups and 14 tentative P-types. Future classification and nomenclature updates will be based on complete genome sequences and will be coordinated and disseminated by the international norovirus classification-working group.
Comparison of norovirus genogroup I, II and IV seroprevalence among children in the Netherlands, 1963, 1983 and 2006
Noroviruses are a major cause of acute gastroenteritis worldwide and are a genetically diverse group of viruses. Since 2002 an increasing number of norovirus outbreaks have been reported globally but it is not clear whether this increase has been caused by a higher awareness or reflects the emergence of new genogroup II genotype 4 (GII.4) variants. The hypothesis that norovirus prevalence has increased post-2002 and is related to the emergence of GII.4 is tested in this study. Sera collected from children aged <5 years of three Dutch cross-sectional population based cohorts in 1963 1983 and 2006/2007 (n=143 n=130 and n=376 respectively) were tested for specific serum IgG by protein array using antigens to GII.4 and a range of other antigens representing norovirus GI GII and GIV genotypes. The protein array was validated by paired sera of norovirus infected patients and supernatants of B-cell cultures with single epitope specificity. Evidence for norovirus infection was found to be common among Dutch children in each cohort but the prevalence towards different genotypes changed over time. At the genogroup level GI seroprevalence decreased significantly between 1963 and 2006/2007 while a significant increase of GII and in particular specific antibodies of the genotype GII.4 was detected in the 2006/2007 cohort. There were no children with only GII.4 antibodies in the 1963 cohort. This study shows that the high GII.4 norovirus incidence in very young children is a recent phenomenon. These findings are of importance for vaccine development and trials that are currently focusing mostly on GII.4 viruses.
Systemic immune response after rotavirus inoculation of neonatal mice depends on source and level of purification of the virus: implications for the use of heterologous vaccine candidates
Rotavirus is an important cause of morbidity and mortality worldwide and vaccines are currently under development with clinical trails conducted in humans worldwide. The immune responses in infant BALB/c mice were examined following oral inoculation with murine rotavirus EDIM (2×104 focus-forming units) and with three CsCl gradient-purified fractions of heterologous simian rotavirus SA11 (standardized at 2×106 CCID50) that differed in antigen composition: fraction 1 was enriched for double-layered rotavirus particles fraction 2 for triple-layered particles and fraction 3 consisted mainly of cell components. Diarrhoea and high IgG responses but marginal IgA responses were observed after inoculation with all three SA11 fractions. Virus shedding was observed in all EDIM-inoculated mice but in none of the SA11-inoculated mice. Rotavirus-specific IgG1 : 2a ratios were similar in mice inoculated with EDIM and SA11 fraction 1 but higher for SA11 fraction 3- and lower for SA11 fraction 2-inoculated mice. A higher IgG1 : 2a ratio indicates a more Th2-like immune response. This undesirable response is apparently mostly induced by inoculation with heterologous rotavirus in the presence of abundant cell-associated and soluble rotavirus proteins compared with infection with a more purified preparation or with homologous virus. These data show that following inoculation with a standardized amount of infectious virus the composition of the fraction influences the outcome of the immune responses significantly.
Laboratory efforts to cultivate noroviruses
Noroviruses (NoVs) are a leading cause of gastroenteritis worldwide and are recognized as the foremost cause of foodborne illness. Despite numerous efforts routine cell cultures have failed to yield replicating NoV. This paper describes methods used to try to grow NoV in vitro in two laboratories. Cells (A549 AGS Caco-2 CCD-18 CRFK CR-PEC Detroit 551 Detroit 562 FRhK-4 HCT-8 HeLa HEC HEp-2 Ht-29 HuTu-80 I-407 IEC-6 IEC-18 Kato-3 L20B MA104 MDBK MDCK RD TMK Vero and 293) were cultured on solid or permeable surfaces. Differentiation was induced using cell culture supplements such as insulin DMSO and butyric acid. In some cases the cells and the NoV-containing stool samples were treated with bioactive digestive additives. Variables evaluated in cultivation experiments included the method of preparation of the virus inoculum the genotype of the virus conditions for maintenance of cell monolayers additives in the maintenance medium and the method of inoculation of the cells. Serial blind passage studies were performed routinely. In addition to evaluation for CPE evidence of virus replication was sought using immunofluorescent assays to detect newly produced viral capsid antigen and RT-PCR assays to detect the viral genome. Although some infected cultures remained NoV positive by RT-PCR for up to five passages and an occasional cell in a monolayer showed evidence of specific immunofluorescence no reproducible NoV-induced CPE was observed and all RT-PCR results that were positive initially were negative following continued passaging. Thus attempts to develop a method for the cultivation of NoV were unsuccessful.
A Sabin vaccine-derived field isolate of poliovirus type 1 displaying aberrant phenotypic and genetic features, including a deletion in antigenic site 1.
Poliovirus strains derived from the oral poliovirus vaccine (Sabin) can be differentiated from wild-type poliovirus by tests based on either immunological or genetic properties of the strains. The characterization of a recently identified poliovirus type 1 isolate with exceptional properties is described. Initial phenotypic analysis of the virus by use of polyclonal absorbed antisera suggested a wild-type character. However the different genomic analyses all confirmed the Sabin-derived character of the virus. All 17 plaques isolated from the strain shared these properties thus excluding the possibility of a mixture of a wild-type and a Sabin-derived strain. To elucidate the properties of this virus further the nucleotide sequences of the P1 region and most of the 5′ non-coding region were established. Although the nucleotide identity with Sabin 1 was more than 99.4% mutations were observed in regions encoding three major antigenic sites; the deduced amino acid substitutions confirmed the aberrant results of micro-neutralization assays with site-specific monoclonal antibodies. The most striking feature was the existence of a hexanucleotide deletion in the VP1 gene which gave rise to a two amino acid deletion in the BC loop. In spite of these antigenic changes the strain was readily serotyped as poliovirus type 1 under standard conditions. Likewise replication of the virus under cell culture conditions was not affected by these mutations or by the deletion. Standard polio vaccination protects against this aberrant virus and its epidemiological significance remains open.