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      Molecular characterisation of the NSP4 gene of group A human rotavirus G2P[4] strains circulating in São Paulo, Brazil, from 1994 and 2006 to 2010

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          Abstract

          Group A human rotaviruses (HuRVA) are causative agents of acute gastroenteritis. Six viral structural proteins (VPs) and six nonstructural proteins (NSPs) are produced in RV-infected cells. NSP4 is a diarrhoea-inducing viral enterotoxin and NSP4 gene analysis revealed at least 15 (E1-E15) genotypes. This study analysed the NSP4 genetic diversity of HuRVA G2P[4] strains collected in the state of São Paulo (SP) from 1994 and 2006-2010 using reverse transcription-polymerase chain reaction, sequencing and phylogenetic analysis. Forty (97.6%) G2P[4] strains displayed genotype E2; one strain (2.4%) displayed genotype E1. These results are consistent with the proposed linkage between VP4/VP7 (G2P[4]) and the NSP4 (E2) genotype of HuRVA. NSP4 phylogenetic analysis showed distinct clusters, with grouping of most strains by their genotype and collection year, and most strains from SP were clustered together with strains from other Brazilian states. A deduced amino acid sequence alignment for E2 showed many variations in the C-terminal region, including the VP4-binding domain. Considering the ability of NSP4 to generate host immunity, monitoring NSP4 variations, along with those in the VP4 or VP7 protein, is important for evaluating the circulation and pathogenesis of RV. Finally, the presence of one G2P[4]E1 strain reinforces the idea that new genotype combinations emerge through reassortment and independent segregation.

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          Uniformity of rotavirus strain nomenclature proposed by the Rotavirus Classification Working Group (RCWG).

          Jelle Matthijnssens, Max Ciarlet, Sarah McDonald … (2011)
          In April 2008, a nucleotide-sequence-based, complete genome classification system was developed for group A rotaviruses (RVs). This system assigns a specific genotype to each of the 11 genome segments of a particular RV strain according to established nucleotide percent cutoff values. Using this approach, the genome of individual RV strains are given the complete descriptor of Gx-P[x]-Ix-Rx-Cx-Mx-Ax-Nx-Tx-Ex-Hx. The Rotavirus Classification Working Group (RCWG) was formed by scientists in the field to maintain, evaluate and develop the RV genotype classification system, in particular to aid in the designation of new genotypes. Since its conception, the group has ratified 51 new genotypes: as of April 2011, new genotypes for VP7 (G20-G27), VP4 (P[28]-P[35]), VP6 (I12-I16), VP1 (R5-R9), VP2 (C6-C9), VP3 (M7-M8), NSP1 (A15-A16), NSP2 (N6-N9), NSP3 (T8-T12), NSP4 (E12-E14) and NSP5/6 (H7-H11) have been defined for RV strains recovered from humans, cows, pigs, horses, mice, South American camelids (guanaco), chickens, turkeys, pheasants, bats and a sugar glider. With increasing numbers of complete RV genome sequences becoming available, a standardized RV strain nomenclature system is needed, and the RCWG proposes that individual RV strains are named as follows: RV group/species of origin/country of identification/common name/year of identification/G- and P-type. In collaboration with the National Center for Biotechnology Information (NCBI), the RCWG is also working on developing a RV-specific resource for the deposition of nucleotide sequences. This resource will provide useful information regarding RV strains, including, but not limited to, the individual gene genotypes and epidemiological and clinical information. Together, the proposed nomenclature system and the NCBI RV resource will offer highly useful tools for investigators to search for, retrieve, and analyze the ever-growing volume of RV genomic data.
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            Full genome-based classification of rotaviruses reveals a common origin between human Wa-Like and porcine rotavirus strains and human DS-1-like and bovine rotavirus strains.

            Jelle Matthijnssens, Max Ciarlet, Erica Heiman … (2008)
            Group A rotavirus classification is currently based on the molecular properties of the two outer layer proteins, VP7 and VP4, and the middle layer protein, VP6. As reassortment of all the 11 rotavirus gene segments plays a key role in generating rotavirus diversity in nature, a classification system that is based on all the rotavirus gene segments is desirable for determining which genes influence rotavirus host range restriction, replication, and virulence, as well as for studying rotavirus epidemiology and evolution. Toward establishing such a classification system, gene sequences encoding VP1 to VP3, VP6, and NSP1 to NSP5 were determined for human and animal rotavirus strains belonging to different G and P genotypes in addition to those available in databases, and they were used to define phylogenetic relationships among all rotavirus genes. Based on these phylogenetic analyses, appropriate identity cutoff values were determined for each gene. For the VP4 gene, a nucleotide identity cutoff value of 80% completely correlated with the 27 established P genotypes. For the VP7 gene, a nucleotide identity cutoff value of 80% largely coincided with the established G genotypes but identified four additional distinct genotypes comprised of murine or avian rotavirus strains. Phylogenetic analyses of the VP1 to VP3, VP6, and NSP1 to NSP5 genes showed the existence of 4, 5, 6, 11, 14, 5, 7, 11, and 6 genotypes, respectively, based on nucleotide identity cutoff values of 83%, 84%, 81%, 85%, 79%, 85%, 85%, 85%, and 91%, respectively. In accordance with these data, a revised nomenclature of rotavirus strains is proposed. The novel classification system allows the identification of (i) distinct genotypes, which probably followed separate evolutionary paths; (ii) interspecies transmissions and a plethora of reassortment events; and (iii) certain gene constellations that revealed (a) a common origin between human Wa-like rotavirus strains and porcine rotavirus strains and (b) a common origin between human DS-1-like rotavirus strains and bovine rotaviruses. These close evolutionary links between human and animal rotaviruses emphasize the need for close simultaneous monitoring of rotaviruses in animals and humans.
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              Recommendations for the classification of group A rotaviruses using all 11 genomic RNA segments.

              Krisztián Bányai, Koki Taniguchi, Andrej Steyer … (2007)
              Recently, a classification system was proposed for rotaviruses in which all the 11 genomic RNA segments are used (Matthijnssens et al. in J Virol 82:3204-3219, 2008). Based on nucleotide identity cut-off percentages, different genotypes were defined for each genome segment. A nomenclature for the comparison of complete rotavirus genomes was considered in which the notations Gx-P[x]-Ix-Rx-Cx-Mx-Ax-Nx-Tx-Ex-Hx are used for the VP7-VP4-VP6-VP1-VP2-VP3-NSP1-NSP2-NSP3-NSP4-NSP5/6 encoding genes, respectively. This classification system is an extension of the previously applied genotype-based system which made use of the rotavirus gene segments encoding VP4, VP7, VP6, and NSP4. In order to assign rotavirus strains to one of the established genotypes or a new genotype, a standard procedure is proposed in this report. As more human and animal rotavirus genomes will be completely sequenced, new genotypes for each of the 11 gene segments may be identified. A Rotavirus Classification Working Group (RCWG) including specialists in molecular virology, infectious diseases, epidemiology, and public health was formed, which can assist in the appropriate delineation of new genotypes, thus avoiding duplications and helping minimize errors. Scientists discovering a potentially new rotavirus genotype for any of the 11 gene segments are invited to send the novel sequence to the RCWG, where the sequence will be analyzed, and a new nomenclature will be advised as appropriate. The RCWG will update the list of classified strains regularly and make this accessible on a website. Close collaboration with the Study Group Reoviridae of the International Committee on the Taxonomy of Viruses will be maintained.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Mem Inst Oswaldo Cruz
                Journal ID (iso-abbrev): Mem. Inst. Oswaldo Cruz
                Journal ID (publisher-id): mioc
                Title: Memórias do Instituto Oswaldo Cruz
                Publisher: Instituto Oswaldo Cruz, Ministério da Saúde
                ISSN (Print): 0074-0276
                ISSN (Electronic): 1678-8060
                Publication date (Print and electronic): September 2015
                Publication date (Print): September 2015
                Volume: 110
                Issue: 6
                Pages: 786-792
                Affiliations
                [1 ]Universidade Estadual de Campinas, Instituto de Biologia, Departamento de Genética, Evolução e Bioagentes, Campinas, SP, Brasil
                [2 ]Universidade da Cidade de São Paulo, São Paulo, SP, Brasil
                [3 ]Universidade Estadual de Campinas, Centro de Biologia Molecular e Engenharia Genética, Campinas, SP, Brasil
                [4 ]Universidade de São Paulo, Instituto de Ciências Biomédicas, Departamento de Microbiologia, São Paulo, SP, Brasil
                [5 ]Faculdade de Ciências Médicas da Santa Casa de São Paulo, São Paulo, SP, Brasil
                Author notes
                [+ ] Corresponding author: jessicabertol@ 123456hotmail.com
                Article
                DOI: 10.1590/0074-02760150199
                PMC ID: 4667582
                PubMed ID: 26517658
                SO-VID: a7169f3c-a437-4328-9cad-036c041383f9
                License:

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                Date received : 19 May 2015
                Date accepted : 20 August 2015
                Page count
                Figures: 2, Tables: 1, Equations: 0, References: 51, Pages: 7
                Categories
                Subject: Articles

                Keywords: human rotavirus,nsp4 genotypes,phylogenetic analysis,vp4/vp7/nsp4 linkage,brazil
                Data availability:
                Keywords: human rotavirus, nsp4 genotypes, phylogenetic analysis, vp4/vp7/nsp4 linkage, brazil

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