ORCID Profile
0000-0001-8735-4618
Current Organisation
University of Tokyo
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Publisher: Elsevier BV
Date: 2021
Publisher: Springer Science and Business Media LLC
Date: 17-03-2005
DOI: 10.1007/S00705-005-0506-0
Abstract: Sapovirus (SaV), a member of the genus Sapovirus in the family Caliciviridae, is an agent of human and porcine gastroenteritis. SaV strains were recently ided into five genogroups (GI to GV). We characterized novel polyclonal antibodies raised against SaV GV virus-like particles (VLPs) by Western blot analysis, and both antibody and antigen enzyme-linked immunosorbent assays (ELISAs). Our results have indicated SaV GI and GV VLPs were antigenically distinct by Western blotting and ELISAs. These reagents may be useful for genogroup specific detection of SaV.
Publisher: American Society for Microbiology
Date: 04-2007
DOI: 10.1128/JCM.01854-06
Abstract: An outbreak of gastroenteritis occurred at a kindergarten in Yokote City, Japan, between February 2006 and March 2006. Sapovirus was identified in 19 of 26 stool specimens by reverse transcription-PCR. A high viral shedding pattern was found for this strain, which was shown to be antigenically distinct from other genogroups.
Publisher: American Society for Microbiology
Date: 16-06-2020
DOI: 10.1128/JVI.00660-20
Abstract: This research discovered and analyzed two different types of MNV-neutralizing nanobodies. The top-binding nanobodies sterically inhibited the receptor-binding site, whereas the dimeric-binding nanobodies interfered with a structural modification associated with cofactor binding. Moreover, we found that the capsid contained a number of vulnerable regions that were essential for viral replication. In fact, the capsid appeared to be organized in a state of flux, which could be important for cofactor/receptor-binding functions. Blocking these capsid-binding events with nanobodies directly inhibited essential capsid functions. Moreover, a number of MNV-specific nanobody binding epitopes were comparable to human norovirus-specific nanobody inhibitors. Therefore, this additional structural and inhibition information could be further exploited in the development of human norovirus antivirals.
Publisher: Elsevier BV
Date: 2001
Publisher: American Society for Microbiology
Date: 26-04-2017
Abstract: The norovirus genogroup II genotype 4 (GII.4) variants have approximately 5% ergence in capsid amino acid identity and have dominated over the past decade. The precise reason(s) for the GII.4 emergence and persistence in the human population is still unknown, but some studies have suggested that chronically infected patients might generate novel variants that can cause new epidemics. We examined GII.4 noroviruses isolated from an immunocompromised patient with a long-term infection. Numerous norovirus capsid quasi-species were isolated during the 13-month study. The capsid quasi-species clustered into two genetic and antigenic types. However, the HBGA binding profiles were similar between the two antigenic clusters, indicating that the amino acid substitutions did not alter the HBGA binding interactions. The isolated sequences represented two new GII.4 variants, but similar sequences were not found in the database. These results indicated that chronically infected patients might not generate novel noroviruses that cause outbreaks.
Publisher: Wiley
Date: 03-2018
Abstract: Human milk oligosaccharides (HMOs) are erse unconjugated carbohydrates that are highly abundant in human breast milk. These glycans are investigated in the context of exhibiting multiple functions in infant growth and development. They seem to provide protection against infectious diseases, including a number of poorly manageable viral infections. Although the potential mechanism of the HMO antiviral protection is rather broad, much of the current experimental work has focused on studying of HMO antiadhesive properties. HMOs may mimic structures of viral receptors and block adherence to target cells, thus preventing infection. Still, the potential of HMOs as a source for new antiviral drugs is relatively unexploited. This can be partly attributed to the extreme complexity of the virus-carbohydrate interactions and technical difficulties in HMO isolation, characterization, and manufacturing procedures. Fortunately, we are currently entering a period of major technological advances that have enabled deeper insights into carbohydrate mediated viral entry, rational selection of HMOs as anti-entry inhibitors, and even evaluation of in idual synthetic HMO structures. Here, we provide an up-to-date review on glycan binding studies for rotaviruses, noroviruses, influenza viruses, and human immunodeficiency viruses. We also discuss the preventive and therapeutic potential of HMOs as anti-entry inhibitors and address challenges on the route from fundamental studies to clinical trials.
Publisher: American Society for Microbiology
Date: 03-2019
DOI: 10.1128/JVI.02174-18
Abstract: The emergence of new epidemic GII.4 norovirus variants is thought to be associated with changes in antigenicity and HBGA binding capacity. Here, we show that HBGA binding profiles remain unchanged between the 1974 and 2012 GII.4 variants, whereas these variants showed various levels of reactivity against a panel of GII.4 MAbs. We identified a MAb that bound at the HBGA pocket, blocked norovirus VLPs from binding to HBGAs, and neutralized norovirus virions in the cell culture system. Raised against a GII.4 2006 strain, this MAb was unreactive to a GII.4 1974 isolate but was able to neutralize the newer 2012 strain, which has important implications for vaccine design. Altogether, these new findings suggest that the amino acid variations surrounding the HBGA pocket lead to temporal changes in antigenicity without affecting the ability of GII.4 variants to bind HBGAs, which are known cofactors for infection.
Publisher: American Society for Microbiology
Date: 27-04-2023
DOI: 10.1128/JVI.01833-22
Abstract: Human noroviruses are highly contagious and a major problem in closed institutions, such as schools, hospitals, and cruise ships. Reducing norovirus infections is challenging on multiple levels and includes the frequent emergence of antigenic variants, which complicates designing effective, broadly reactive capsid therapeutics.
Publisher: WORLD SCIENTIFIC
Date: 2008
Publisher: MyJove Corporation
Date: 19-04-2016
DOI: 10.3791/53845
Publisher: MDPI AG
Date: 05-01-2023
DOI: 10.3390/NU15020268
Abstract: Garcinia dulcis (GD) extract has been found to have anti-hypertensive properties in animal studies. GD can also alter the colonic microbiota of rats. However, the effects of GD on changes in the gut microbiota and metabolomic profiles of normotensive and hypertensive rats are currently unknown. The purpose of this study was to evaluate changes in the gut microbiota and metabolomic profiles of 2-kidneys-1 clip (2K1C) hypertensive rats after feeding with GD flower extract. Rats were randomly ided into the following 4 groups: sham operation (SO) receiving corn oil (CO) (SO + CO), SO receiving GD (SO + GD), 2K1C receiving corn oil (2K1C + CO) and 2K1C receiving GD (2K1C + GD). Body weight (BW) and systolic blood pressure (SBP) were measured weekly throughout the study. Gut microbiota and fecal metabolites were measured from fresh fecal contents. Alpha ersity results demonstrated a similar microbial richness and ersity between groups. Linear discriminant analysis (LDA) effect size (LEfSe) suggested that GD treatment affected gut microbial community structure in both hypertensive and normotensive rats. Feeding rats with GD caused metabolic alterations that rendered 2K1C + GD rats similar to SO + CO and SO + GD rats. Findings suggest that the impact of GD on gut microbiota and metabolite profiles may be related to its anti-hypertensive properties.
Publisher: CRC Press
Date: 14-01-2009
Publisher: Wiley
Date: 2007
DOI: 10.1002/RMV.552
Publisher: American Society for Microbiology
Date: 2012
DOI: 10.1128/JVI.05909-11
Abstract: Human noroviruses bind with their capsid-protruding domains to histo-blood-group antigens (HBGAs), an interaction thought to direct their entry into cells. Although human noroviruses are the major cause of gastroenteritis outbreaks, development of antivirals has been lacking, mainly because human noroviruses cannot be cultivated. Here we use X-ray crystallography and saturation transfer difference nuclear magnetic resonance (STD NMR) to analyze the interaction of citrate with genogroup II (GII) noroviruses. Crystals of citrate in complex with the protruding domain from norovirus GII.10 Vietnam026 diffracted to 1.4 Å and showed a single citrate bound at the site of HBGA interaction. The citrate interaction was coordinated with a set of capsid interactions almost identical to that involved in recognizing the terminal HBGA fucose, the saccharide which forms the primary conserved interaction between HBGAs and GII noroviruses. Citrate and a water molecule formed a ring-like structure that mimicked the pyranoside ring of fucose. STD NMR showed the protruding domain to have weak affinity for citrate (460 μM). This affinity, however, was similar to the affinities of the protruding domain for fucose (460 μM) and H type 2 trisaccharide (390 μM), an HBGA shown previously to be specifically recognized by human noroviruses. Importantly, competition STD NMR showed that citrate could compete with HBGA for norovirus binding. Together, the results suggest that citrate and other glycomimetics have the potential to block human noroviruses from binding to HBGAs.
Publisher: Elsevier BV
Date: 08-2019
DOI: 10.1016/J.ANTIVIRAL.2019.05.010
Abstract: Human noroviruses are a leading cause of acute gastroenteritis, yet there are still no vaccines or antivirals available. Expression of the norovirus capsid protein (VP1) in insect cells typically results in the formation of virus-like particles (VLPs) that are morphologically and antigenically comparable to native virions. Indeed, several different norovirus VLP candidates are currently used in clinical trials. So far, structural analysis of norovirus VLPs showed that the capsid has a T = 3 icosahedral symmetry and is composed of 180 copies of VP1 that are folded into three quasi-equivalent subunits (A, B, and C). In this study, the VLP structures of two norovirus GII.4 genetic variants that were identified in 1974 and 2012 were determined using cryo-EM. Surprisingly, we found that greater than 95% of these GII.4 VLPs were larger than virions and 3D reconstruction showed that these VLPs exhibited T = 4 icosahedral symmetry. We also discovered that the T = 4 VLPs presented several novel structural features. The T = 4 particles assembled from 240 copies of VP1 that adopted four quasi-equivalent conformations (A, B, C, and D) and formed two distinct dimers, A/B and C/D. The protruding domains were elevated ∼21 Å off the capsid shell, which was ∼7 Å more than in the previously studied GII.10 T = 3 VLPs. A small cavity and flap-like structure at the icosahedral two-fold axis disrupted the contiguous T = 4 shell. Overall, our findings indicated that GII.4 VP1 sequences assemble into T = 4 VLPs and these larger particles might have important consequences for VLP-based vaccine development.
Publisher: Springer Science and Business Media LLC
Date: 07-02-2006
DOI: 10.1007/S00705-005-0717-4
Abstract: Noroviruses are a major health burden and are responsible for the majority of outbreaks of gastroenteritis in the world. Human noroviruses can be genetically ided into two main genogroups (GI and GII) and sub ided into many genotypes. In this study, stool specimens collected from 12 outbreaks of gastroenteritis in Taiwan were screened for viral agents between the 23rd of November 2004 and 9th of March 2005. Noroviruses were detected in all outbreaks. We detected six different norovirus genotypes: GI/11, GI/14, GII/3, GII/4, GII/6, and GII/18. Noroviruses belonging to GII/4 were dominant, 50 of 60 (83%) sequences, and were detected in 10 of 12 outbreaks. Furthermore, the norovirus GII/4 strains were detected throughout Taiwan, demonstrating their widespread distribution. We also found that three outbreaks had noroviruses from multiple genotypes. Our results have shown for the first time that noroviruses are an important cause of gastroenteritis in Taiwan.
Publisher: American Society for Microbiology
Date: 06-2018
DOI: 10.1128/JVI.00413-18
Abstract: Human noroviruses are the leading cause of acute gastroenteritis in humans. Noroviruses also infect animals, such as cows, mice, cats, and dogs. How noroviruses bind and enter host cells is still incompletely understood. Recently, the type I transmembrane protein CD300lf was identified as the murine norovirus receptor, yet it is unclear how the virus capsid and receptor interact at the molecular level. In this study, we determined the X-ray crystal structure of the soluble CD300lf (sCD300lf) and the murine norovirus capsid protruding domain complex at a 2.05-Å resolution. We found that the sCD300lf-binding site is located on the topside of the protruding domain and involves a network of hydrophilic and hydrophobic interactions. sCD300lf locked nicely into a complementary cavity on the protruding domain that is additionally coordinated with a positive surface charge on sCD300lf and a negative surface charge on the protruding domain. Five of six protruding domain residues interacting with sCD300lf were maintained between different murine norovirus strains, suggesting that sCD300lf was capable of binding to a highly conserved pocket. Moreover, a sequence alignment with other CD300 paralogs showed that the sCD300lf-interacting residues were partially conserved in CD300ld but variable in other CD300 family members, consistent with previously reported infection selectivity. Overall, these data provide insights into how a norovirus engages a protein receptor and will be important for a better understanding of selective recognition and norovirus attachment and entry mechanisms. IMPORTANCE Noroviruses exhibit exquisite host range specificity due to species-specific interactions between the norovirus capsid protein and host molecules. Given this strict host range restriction, it has been unclear how the viruses are maintained within a species between relatively sporadic epidemics. While much data demonstrate that noroviruses can interact with carbohydrates, recent work has shown that expression of the protein CD300lf is both necessary and sufficient for murine norovirus infection of mice and binding of the virus to permissive cells. Importantly, the expression of this murine protein by human cells renders them fully permissive for murine norovirus infection, indicating that at least in this case, host range restriction is determined by molecular events that control receptor binding and entry. Defining the atomic-resolution interactions between the norovirus capsid protein and its cognate receptor is essential for a molecular understanding of host-range restriction and norovirus tropism.
Publisher: Springer Science and Business Media LLC
Date: 25-04-2013
Publisher: Springer Science and Business Media LLC
Date: 20-10-2011
DOI: 10.1007/S11262-010-0535-Y
Abstract: The human caliciviruses (CV), norovirus (NoV) and sapovirus (SaV), are major causes of outbreak gastroenteritis worldwide. To date, the investigation of human NoV and SaV replication cycles has been impeded as neither is culturable. Consequently, the recently discovered murine NoV (MNV) has been adopted as a surrogate replication model for the human CVs. In this study, we sought to compare the biochemical properties of the MNV RNA-dependent RNA polymerase (RdRp) with related human NoV and SaV-RdRps to address the suitability of MNV as a model for the human CVs. Three human NoV-RdRps (GII.b, GII.4 and GII.7), an MNV-RdRp and two human SaV-RdRps (GI and GII) were overexpressed in Escherichia coli, purified and their enzymatic activity and fidelity compared. Despite ~70% amino acid variation between the RdRp from the two different CV genera, the majority of the physiological characteristics of the RdRps were similar. All RdRps exhibited co-operative dimerisation and had optimal activity at 25°C, a pH range between 7 and 8, required 2-5 mM MnCl(2) and were inhibited with increasing NaCl concentrations. We observed RdRp activity at temperatures as low as 5°C and as high as 65°C. Using an in vitro fidelity assay, similar mutation rates were observed for the separate RdRps (1 × 10(-4)-1 × 10(-5)). This is the first report to compare the physiological, biochemical and mutational properties of the MNV-RdRp to those of the human CV-RdRps and it suggests that MNV may be directly applicable to the study of human NoV.
Publisher: American Society for Microbiology
Date: 31-05-2023
DOI: 10.1128/JVI.00451-23
Publisher: Microbiology Society
Date: 04-2006
Abstract: Human norovirus (NoV) strains cause a considerable number of outbreaks of gastroenteritis worldwide. Based on their capsid gene (VP1) sequence, human NoV strains can be grouped into two genogroups (GI and GII) and at least 14 GI and 17 GII genotypes (GI/1–14 and GII/1–17). Human NoV strains cannot be propagated in cell-culture systems, but expression of recombinant VP1 in insect cells results in the formation of virus-like particles (VLPs). In order to understand NoV antigenic relationships better, cross-reactivity among 26 different NoV VLPs was analysed. Phylogenetic analyses grouped these NoV strains into six GI and 12 GII genotypes. An antibody ELISA using polyclonal antisera raised against these VLPs was used to determine cross-reactivity. Antisera reacted strongly with homologous VLPs however, a number of novel cross-reactivities among different genotypes was observed. For ex le, GI/11 antiserum showed a broad-range cross-reactivity, detecting two GI and 10 GII genotypes. Likewise, GII/1, GII/10 and GII/12 antisera showed a broad-range cross-reactivity, detecting several other distinct GII genotypes. Alignment of VP1 amino acid sequences suggested that these broad-range cross-reactivities were due to conserved amino acid residues located within the shell and/or P1-1 domains. However, unusual cross-reactivities among different GII/3 antisera were found, with the results indicating that both conserved amino acid residues and VP1 secondary structures influence antigenicity.
Publisher: American Society for Microbiology
Date: 26-10-2016
Abstract: We previously identified a Nanobody (termed Nano-85) that bound to a highly conserved region on the norovirus capsid. In this study, the Nanobody was biotinylated and gold conjugated for a lateral flow immunoassay (termed Nano-IC). We showed that the Nano-IC assay was capable of detecting at least four antigenically distinct GII genotypes, including the newly emerging GII.17. In the clinical setting, the Nano-IC assay had sensitivities equivalent to other commercially available lateral flow systems. The Nano-IC method was capable of producing results in ~5 min, which makes this method useful in settings that require rapid diagnosis, such as cruise ship outbreaks and elder care facilities. The Nano-IC assay has several advantages over antibody-based IC methods: for ex le, Nanobodies can be readily produced in large quantities, they are generally more stable than conventional antibodies, and the Nanobody binding sites can be easily obtained by X-ray crystallography.
Publisher: American Society for Microbiology
Date: 23-11-2022
DOI: 10.1128/JVI.01217-22
Abstract: Isolated RHDV antibodies have been used for decades to distinguish between antigenic variants, monitor temporal capsid evolution, and examine neutralizing capacities. In this study, we provided the structural basis for an RHDV GI.2 specific diagnostic antibody (2D9) binding and reveal that a small number of amino acid substitutions at the binding site could differentiate between RHDV GI.2 and GI.1b.
Publisher: Cold Spring Harbor Laboratory
Date: 25-01-2019
DOI: 10.1101/528240
Abstract: Human noroviruses are a major cause of gastroenteritis, yet there are still no vaccines or antivirals available. Nevertheless, a number of vaccine candidates that are currently in clinical trials are composed of norovirus virus-like particles (VLPs). These VLPs are recognized as morphologically and antigenically similar to norovirus virions. An X-ray crystal structure of the prototype (GI.1) VLPs showed that the norovirus capsid has a T=3 icosahedral symmetry and is composed of 180 copies of the major capsid protein (VP1) that folds into three quasi-equivalent subunits (A, B, and C). In this study, we determined the cryo-EM structure of VLPs for two GII.4 noroviruses that were detected in 1974 and 2012. We showed that these VLPs had a T=4 symmetry and were composed of 240 copies of VP1. The VP1 on the T=4 VLPs adapted four quasi-equivalent subunits (termed A, B, C, and D), which formed two distinct dimers (A/B and C/D). We found that the T=4 protruding domain was elevated ~21 Å off the capsid shell, which was ~7 Å more than the previously determined for the T=3 GII.10 norovirus. Another interesting feature of the T=4 VLPs was a small cavity and flaplike structure located at the twofold axis. This structural feature was associated with the shell domain (D subunit) and disrupted the contiguous shell. Altogether, we showed that the T=4 VLPs had a number of structural similarities and differences with other noroviruses, but how these structural changes associate with norovirus virions could be important for vaccine studies. The discovery that the GII.4 VLPs (identified in 1974 and 2012, termed CHDC-1974 and NSW-2012, respectively) have a T=4 symmetry is of major significance, since the NSW-2012 is clinically important and previous structural and biochemical studies assumed noroviruses have a T=3 symmetry and are composed of 180 copies of VP1. More importantly, NSW-2012 norovirus shared 96% amino acid identity with a GII.4 vaccine candidate and our data suggests that this vaccine might also have a T=4 symmetry. Although it is not clear if the T=4 VLPs were an artifact of the insect cell expression system, the T=4 VLP vaccines might not recognize equivalent epitopes on T=3 virions, which will be important for future neutralization studies. Finally, further studies with other norovirus genotypes and virions are clearly needed in order to determine the level of this structural ersity.
Publisher: American Society for Microbiology
Date: 22-06-2023
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 07-2005
Abstract: Norovirus (NoV) genogroups I and II (GI and GII) are now recognized as the predominant worldwide cause of outbreaks of acute gastroenteritis in humans. Three recombinant NoV GII isolates were identified and characterized, 2 of which are unrelated to any previously published recombinant NoV. Using data from the current study, published sequences, database searches, and molecular techniques, we identified 23 recombinant NoV GII and 1 recombinant NoV GI isolates. Analysis of the genetic relationships among the recombinant NoV GII isolates identified 9 independent recombinant sequences the other 14 strains were close relatives. Two of the 9 independent recombinant NoV were closely related to other recombinants only in the polymerase region, and in a similar fashion 1 recombinant NoV was closely related to another only in the capsid region. Breakpoint analysis of recombinant NoV showed that recombination occurred in the open reading frame (ORF)1/ORF2 overlap. We provide evidence to support the theory of the role of subgenomic RNA promoters as recombination hotspots and describe a simple mechanism of how recombination might occur in NoV.
Publisher: Springer Science and Business Media LLC
Date: 21-04-2005
DOI: 10.1007/S00705-005-0540-Y
Abstract: A total of 921 fecal specimens collected from 44 infants in a day care center (DCC) in Tokyo, Japan during June 1999 to July 2000 were tested for the presence of rotavirus, norovirus, sapovirus, astrovirus and adenovirus by reverse-transcription-multiplex polymerase chain reaction (RT-multiplex PCR) and sequence analysis. Of 88 fecal specimens from infants with acute gastroenteritis, 51.1% (45) were found to be positive for diarrheal viruses. Astrovirus was the most prevalent (15.9%, 14 of 88), followed by norovirus GII (14.8%, 13 of 88), adenovirus (12.5%, 11 of 88), and sapovirus (2.3%, 2 of 88). Viral mixed infection accounted for 5.7% (5 of 88). Interestingly, 230 of 833 (27.6%) fecal specimens collected from asymptomatic infants were also infected with diarrheal viruses. Of these, astrovirus, norovirus GII, adenovirus and sapovirus were identified in 53, 46, 96 and 22 fecal specimens (23%, 20%, 41.7%, and 9.6%, respectively). Moreover, 13 of 833 (1.6%) normal specimens showed mixed viral infections. Surprisingly, no rotavirus (known as the most common causative agent of acute gastroenteritis in DCCs) was detected in those subjects. Another interesting feature was the demonstration of five separate outbreaks of acute gastroenteritis identified in a single DCC. Outbreak A was associated with both astrovirus serotype 1 and norovirus GII/3 (known as Toronto virus cluster) Outbreak B with adenovirus 12 Outbreak C with norovirus GII/4 (Lordsdale virus cluster) Outbreak D with sapovirus GIV and Outbreak E with astrovirus serotype 1. To our knowledge, this is the first proof of multiple outbreaks of viral gastroenteritis in Japanese infants in a single DCC. Our results confirm the presence as well as the importance of these viruses and warn of the threat they pose.
Publisher: S. Karger AG
Date: 2009
DOI: 10.1159/000205527
Abstract: Twelve outbreaks of food handler-associated gastroenteritis between November 2002 and March 2006 in Japan were examined for norovirus (NoV) using RT-PCR and sequence analysis. NoV was detected in 77 of 81 customers and 45 of 104 food handlers. Identical NoV sequences were detected in patients and food handlers in each outbreak.
Publisher: Oxford University Press (OUP)
Date: 27-09-2017
Abstract: Human noroviruses (HuNoV), members of the family Caliciviridae, are the major cause of acute viral gastroenteritis worldwide. Successful infection is linked to the ability of the protruding (P) domain of the viral capsid to bind histo-blood group antigens (HBGA). Binding to gangliosides plays a major role for many nonhuman calici- and noroviruses. Increasing evidence points to a broader role of sialylated carbohydrates such as gangliosides in norovirus infection. Here, we compare HBGA and ganglioside binding of a GII.4 HuNoV variant (MI001), previously shown to be infectious in a HuNoV mouse model. Saturation transfer difference nuclear magnetic resonance spectroscopy, native mass spectrometry (MS) and surface plasmon resonance spectroscopy were used to characterize binding epitopes, affinities, stoichiometry and dynamics, focusing on 3'-sialyllactose, the GM3 ganglioside saccharide and B antigen. Binding was observed for 3'-sialyllactose and various HBGAs following a multistep binding process. Intrinsic affinities (Kd) of fucose, 3'-sialyllactose and B antigen were determined for the in idual binding steps. Stronger affinities were observed for B antigen over 3'-sialyllactose and fucose, which bound in the mM range. Binding stoichiometry was analyzed by native MS showing the presence of four B antigens or two 3'-sialyllactose in the complex. Epitope mapping of 3'-sialyllactose revealed direct interaction of α2,3-linked sialic acid with the P domain. The ability of HuNoV to engage multiple carbohydrates emphasizes the multivalent nature of norovirus glycan-specificity. Our findings reveal direct binding of a GII.4 HuNoV P dimer to α2,3-linked sialic acid and support a broader role of ganglioside binding in norovirus infection.
Publisher: Microbiology Society
Date: 11-2006
Abstract: Sapovirus (SaV), a member of the family Caliciviridae , is a causative agent of acute gastroenteritis in humans and swine and is currently ided into five genogroups, GI–GV. The proteolytic processing of the SaV open reading frame 1 (ORF1) polyprotein with a human GII SaV Mc10 strain has recently been determined and the products are arranged in the following order: NH 2 –p11–p28–p35 (NTPase)–p32–p14 (VPg)–p70 (Pro–Pol)–p60 (VP1)–COOH. The cleavage site between p14 (VPg) and p70 (Pro–Pol) was identified as E 1055 /A 1056 by N-terminal amino acid sequencing. To identify other cleavage sites, a series of GII SaV Mc10 full-length clones containing disrupted potential cleavage sites in the ORF1 polyprotein were constructed and used to generate linear DNA templates for in vitro coupled transcription–translation. The translation products were analysed by SDS-PAGE or by immunoprecipitation with region-specific antibodies. N-terminal amino acid sequencing with Escherichia coli -expressed recombinant proteins was also used to identify the cleavage site between p32 and p14. These approaches enabled identification of the six cleavage sites of the Mc10 ORF1 polyprotein as E 69 /G 70 , Q 325 /G 326 , Q 666 /G 667 , E 940 /A 941 , E 1055 /A 1056 and E 1722 /G 1723 . The alignment of the SaV full-length ORF1 amino acid sequences indicated that the dipeptides used for the cleavage sites were either E or Q at the P1 position and A, G or S at the P1′ position, which were conserved in the GI, GII, GIII, GIV and GV SaV ORF1 polyprotein.
Publisher: Cold Spring Harbor Laboratory
Date: 21-03-2019
DOI: 10.1101/583385
Abstract: Human noroviruses are a leading cause of acute gastroenteritis, yet there are still no vaccines or antivirals available. Expression of the norovirus capsid protein (VP1) in insect cells typically results in the formation of virus-like particles (VLPs) that are morphologically and antigenically comparable to native virions. Previous structural analysis of norovirus VLPs showed that the capsid has a T=3 icosahedral symmetry and is composed of 180 copies of VP1 that are folded into three quasi-equivalent subunits (A, B, and C). In this study, we determined the cryo-EM VLP structures of two GII.4 variants, termed CHDC-1974 and NSW-2012. Surprisingly, we found that greater than 95% of these GII.4 VLPs were larger than virions and 3D reconstruction showed that these VLPs exhibited T=4 icosahedral symmetry. We found that the T=4 VLPs showed several structural differences to the T=3 VLPs. The T=4 particles assemble from 240 copies of VP1 that adopt four quasi-equivalent conformations (A, B, C, and D) that form two distinct dimers, A/B and C/D. The T=4 protruding domains were elevated ∼21-Å off the capsid shell, which was ∼7-Å more than the previously studied GII.10 T=3 VLPs. A small cavity and flap-like structure at the icosahedral twofold axis disrupted the contiguous T=4 shell, a consequence of the D-subunit S-domains having smaller contact interfaces with neighboring dimers. Overall, our findings that old and new GII.4 VP1 sequences assemble T=4 VLPs might have implications for the design of potential future vaccines. The discovery that the GII.4 VLPs have a T=4 symmetry is of significance, since this represents the first known T=4 calicivirus structure. Interestingly, the GII.4 2012 variant shares 96% amino acid identity with a current GII.4 VLP vaccine candidate sequence, which suggests that this vaccine might also have a T=4 symmetry. Our previous results with these GII.4 VLPs showed functional binding properties to antibodies and Nanobodies that were raised against T=3 (GII.10) VLPs. This suggests that the T=4 VLPs were antigenically comparable to T=3 particles, despite the obvious structural and size differences. On the other hand, these larger T=4 VLPs with novel structural features and possibly new epitopes might elicit antibodies that do not recognize equivalent epitopes on the T=3 VLPs. Further structural and binding studies using a library of GII.4-specific Nanobodies are planned in order to precisely investigate whether new epitopes are formed.
Publisher: Wiley
Date: 2007
DOI: 10.1002/RMV.533
Abstract: The family Caliciviridae contains four genera Sapovirus, Norovirus, Lagovirus and Vesivirus, which include Sapporo virus (SaV), Norwalk virus (NoV), Rabbit hemorrhagic disease virus (RHDV) and Feline calicivirus (FCV), respectively. SaV is a causative agent of gastroenteritis in children and adults. SaV can be ided into five genogroups (GI-GV), among which GI, GII, GIV and GV are known to infect humans, whereas SaV GIII infects porcine species. Detection methods include ELISA, RT-PCR and real-time RT-PCR. Since few SaV studies have been conducted, it is difficult to draw correlations between or conclusions about rates of incidence, detection and overall prevalence. Nevertheless, most studies agree that SaV infection is more frequent in young children than adults and that infection in children almost always occurs by 5 years of age. In addition, children at day-care centres and institutions are at greatest risk of SaV-associated infection and transmission. Recently, a number of important findings concerning human SaV were discovered. SaV strains were detected in water s les, which included untreated wastewater specimens, treated wastewater s les and river s les. SaV strains were also detected in shellfish s les destined for human consumption, and recombinant SaV strains were identified in a number of different countries. The purpose of this review was to highlight the current knowledge of human SaV, which appears to be an increasingly important virus causing gastroenteritis in humans.
Publisher: Elsevier BV
Date: 06-2023
Publisher: American Society for Microbiology
Date: 12-2007
DOI: 10.1128/JCM.01516-07
Abstract: 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 food-catering settings between 10 November 2005 and 9 December 2006 in Japan. Stool specimens were collected from both symptomatic and asymptomatic in iduals 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 in iduals had mean viral loads similar to those of symptomatic in iduals, which may account for the increased number of infections and the predominance of an asymptomatic transmission route.
Publisher: Springer Science and Business Media LLC
Date: 23-02-2005
Abstract: Human sapovirus (SaV), an agent of human gastroenteritis, cannot be grown in cell culture, but expression of the recombinant capsid protein (rVP1) in a baculovirus expression system results in the formation of virus-like particles (VLPs). In this study we compared the time-course expression of two different SaV rVP1 constructs. One construct had the native sequence (Wt construct), whereas the other had two nucleotide point mutations in which one mutation caused an amino acid substitution and one was silent (MEG-1076 construct). While both constructs formed VLPs morphologically similar to native SaV, Northern blot analysis indicated that the MEG-1076 rVP1 mRNA had increased steady-state levels. Furthermore, Western blot analysis and an antigen enzyme-linked immunosorbent assay showed that the MEG-1076 construct had increased expression levels of rVP1 and yields of VLPs. Interestingly, the position of the mutated residue was strictly conserved residue among other human SaV strains, suggesting an important role for rVP1 expression.
Publisher: International Union of Crystallography (IUCr)
Date: 29-04-2019
DOI: 10.1107/S2059798319003991
Abstract: Bovine meat and milk factors (BMMFs) are circular, single-stranded episomal DNAs that have been detected in bovine meat and milk products. BMMFs are thought to have roles in human malignant and degenerative diseases. BMMFs encode a replication initiator protein (Rep) that is actively transcribed and translated in human cells. In this study, a Rep WH1 domain encoded on a BMMF (MSBI1.176) isolated from a multiple sclerosis human brain s le was determined to 1.53 Å resolution using X-ray crystallography. The overall structure of the MSBI1.176 WH1 domain was remarkably similar to other Rep structures, despite having a low (28%) amino-acid sequence identity. The MSBI1.176 WH1 domain contained elements common to other Reps, including five α-helices, five β-strands and a hydrophobic pocket. These new findings suggest that the MSBI1.176 Rep might have comparable roles and functions to other known Reps of different origins.
Publisher: CRC Press
Date: 19-04-2016
DOI: 10.1201/B13590
Publisher: Elsevier BV
Date: 09-2015
DOI: 10.1016/J.VIROL.2015.04.006
Abstract: Human noroviruses bind histo-blood group antigens (HBGAs) and this interaction is thought to be important for an infection. We identified two additional fucose-binding pockets (termed fucose-3/4 sites) on a genogroup II human (GII.10) norovirus-protruding (P) dimer using X-ray crystallography. Fucose-3/4 sites were located between two previously determined HBGA binding pockets (termed fucose-1/2 sites). We found that four fucose molecules were capable of binding altogether at fucose-1/2/3/4 sites on the P dimer, though the fucose molecules bound in a dose-dependent and step-wise manner. We also showed that HBGA B-trisaccharide molecules bound in a similar way at the fucose-1/2 sites. Interestingly, we discovered that the monomers of the P dimer were asymmetrical in an unliganded state and when a single B-trisaccharide molecule bound, but were symmetrical when two B-trisaccharide molecules bound. We postulate that the symmetrical dimers might favor HBGA binding interactions at fucose-1/2 sites.
Publisher: Korea Genome Organization
Date: 30-09-2023
DOI: 10.5808/GI.23030
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 10-2007
Publisher: American Society for Microbiology
Date: 03-2015
DOI: 10.1128/JVI.03176-14
Abstract: Human noroviruses are icosahedral single-stranded RNA viruses. The capsid protein is ided into shell (S) and protruding (P) domains, which are connected by a flexible hinge region. There are numerous genetically and antigenically distinct noroviruses, and the dominant strains evolve every other year. Vaccine and antiviral development is h ered by the difficulties in growing human norovirus in cell culture and the continually evolving strains. Here, we show the X-ray crystal structures of human norovirus P domains in complex with two different nanobodies. One nanobody, Nano-85, was broadly reactive, while the other, Nano-25, was strain specific. We showed that both nanobodies bound to the lower region on the P domain and had nanomolar affinities. The Nano-85 binding site mainly comprised highly conserved amino acids among the genetically distinct genogroup II noroviruses. Several of the conserved residues also were recognized by a broadly reactive monoclonal antibody, which suggested this region contained a dominant epitope. Superposition of the P domain nanobody complex structures into a cryoelectron microscopy particle structure revealed that both nanobodies bound at occluded sites on the particles. The flexible hinge region, which contained ∼10 to 12 amino acids, likely permitted a certain degree of P domain movement on the particles in order to accommodate the nanobodies. Interestingly, the Nano-85 binding interaction with intact particles caused the particles to disassemble in vitro . Altogether, these results suggested that the highly conserved Nano-85 binding epitope contained a trigger mechanism for particle disassembly. Principally, this epitope represents a potential site of norovirus vulnerability. IMPORTANCE We characterized two different nanobodies (Nano-85 and Nano-25) that bind to human noroviruses. Both nanobodies bound with high affinities to the lower region of the P domain, which was occluded on intact particles. Nano-25 was specific for GII.10, whereas Nano-85 bound several different GII genotypes, including GII.4, GII.10, and GII.12. We showed that Nano-85 was able to detect norovirus virions in clinical stool specimens using a sandwich enzyme-linked immunosorbent assay. Importantly, we found that Nano-85 binding to intact particles caused the particles to disassemble. We believe that with further testing, Nano-85 not only will work as a diagnostic reagent in norovirus detection systems but also could function as a broadly reactive GII norovirus antiviral.
Publisher: American Society for Microbiology
Date: 04-2013
DOI: 10.1128/JVI.06868-11
Abstract: Human noroviruses are genetically and antigenically highly ergent. Monoclonal antibodies raised in mice against one kind of norovirus virus-like particle (VLP), however, were found to have broad recognition. In this study, we present the crystal structure of the antigen-binding fragment (Fab) for one of these broadly reactive monoclonal antibodies, 5B18, in complex with the capsid-protruding domain from a genogroup II genotype 10 (GII.10) norovirus at 3.3-Å resolution and, also, the cryo-electron microscopy structure of the GII.10 VLP at ∼10-Å resolution. The GII.10 VLP structure was more similar in overall architecture to the GV.1 murine norovirus virion than to the prototype GI.1 human norovirus VLP, with the GII.10 protruding domain raised ∼15 Å off the shell domain and rotated ∼40° relative to the GI.1 protruding domain. In the crystal structure, the 5B18 Fab bound to a highly conserved region of the protruding domain. Based on the VLP structure, this region is involved in interactions with other regions of the capsid and is buried in the virus particle. Despite the occluded nature of the recognized epitope in the VLP structure, enzyme-linked immunosorbent assay (ELISA) binding suggested that the 5B18 antibody was able to capture intact VLPs. Together, the results provide evidence that the norovirus particle is capable of extreme conformational flexibility, which may allow for antibody recognition of conserved surfaces that would otherwise be buried on intact particles.
Publisher: American Society for Microbiology
Date: 2008
DOI: 10.1128/JCM.02108-07
Publisher: Springer Science and Business Media LLC
Date: 23-08-2005
DOI: 10.1007/S00705-005-0613-Y
Abstract: Sapovirus (SaV) is an etiological agent of acute gastroenteritis in human and swine. SaV can be ided into five genogroups, GI to GV. Virus-like particles (VLPs) morphologically similar to native SaV have been expressed for GI, GII, GIII and GV strains in insect cells, although only low expression levels were observed for GII strains. In this study, we report the successful expression of SaV GII VLPs using cultured mammalian COS-7 and 293T cells. Our results demonstrated that this mammalian expression system was able to express and form SaV VLPs.
Publisher: Springer Science and Business Media LLC
Date: 10-05-2004
DOI: 10.1007/S00705-004-0345-4
Abstract: This report describes norovirus (NoV) and sapovirus (SaV) infections in hospitalized children with acute sporadic gastroenteritis in Ho Chi Minh City, Vietnam. Stool specimens collected between December 1999 and November 2000 were examined for NoV and SaV using reverse transcription-PCR and phylogenetic analysis. NoVs were detected in 72 of 448 rotavirus-negative specimens, counted as part of an overall annual detection rate of 5.4% (72 of 1,339 children). This included four NoV genogroup I (GI) strains and 68 NoV GII strains. Only one SaV GI strain was detected in the rotavirus-negative specimens. Over 73% of the NoV sequences belonged to GII/4 (Lordsdale cluster) and were detected in all months except March. We also detected GII/3 strains (Saitama U201 cluster), a naturally occurring recombinant NoV, between January 2000 and March 2000 but not after this period. Other NoV strains belonging to GI/4, GI/8, GII/1, and GII/7 were also detected but were infrequent. In addition, two almost identical NoV GII strains (strains 026 and 0703) collected six months apart were classified into a new genotype that includes the Mc37 strain, which was previously shown to be a recombinant NoV. During this one-year study, the NoV prevailed at the end of the rainy season and the beginning of the dry season. Further epidemiological studies may be necessary to determine whether the GII/4 strains continue to dominant in this region.
Publisher: Public Library of Science (PLoS)
Date: 27-10-2017
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 05-2007
Publisher: Wiley
Date: 24-07-2002
DOI: 10.1002/JMV.10177
Abstract: Norwalk-like viruses (NLVs) were detected using a nested reverse transcriptase-polymerase chain reaction (RT-PCR) with primers directed to the RNA polymerase region. S les were examined from 11 separate outbreaks of gastroenteritis and five sporadic cases of childhood gastroenteritis between 1997 and 2000. Phylogenetic analysis of the 298 bp sequences showed that all strains belong to NLV genogroup II and the majority of the sequenced isolates (30/36) were members of the 95/96-US subset of strains associated with outbreaks recorded worldwide between 1995 and 1996. This was confirmed by analysis of the full length capsid region of a representative Australian isolate. This study demonstrates the usefulness of targeting primers for NLVs to the predominant circulating genotype(s) and confirms the spread of this subtype globally, including the Southern Hemisphere.
Publisher: American Society for Microbiology
Date: 07-2016
DOI: 10.1128/JVI.00317-16
Abstract: Human noroviruses interact with both human histo-blood group antigens (HBGAs) and human milk oligosaccharides (HMOs). The former are believed to be important for a virus infection, while the latter might act as natural decoys in the host during an infection. However, certain noroviruses are known to bind poorly to HBGAs and yet still cause infections some interact with numerous HBGA types but are nonprevalent and yet others bind HBGAs and seem to be increasing in prevalence. HBGAs and HMOs can be found as soluble antigens in humans, can be structurally alike, and can interact with equivalent residues at identical binding pockets on the capsid. In this Gem, we discuss HBGA and HMO binding studies for human noroviruses, concentrating on the clinically important genogroup II noroviruses. In short, the roles of HBGA and HMO interactions in norovirus infections are still unclear.
Publisher: Elsevier BV
Date: 2015
DOI: 10.1016/J.VIROL.2014.10.028
Abstract: Norovirus infects different animals, including humans, mice, dogs, and cats. Here, we show an X-ray crystal structure of a feline GIV.2 norovirus capsid-protruding (P) domain to 2.35Å resolution. The feline GIV.2 P domain was reminiscent of human norovirus P domains, except for a novel P2 subdomain α-helix and an extended P1 subdomain interface loop. These new structural features likely obstructed histo-blood group antigens, which are attachment factors for human norovirus, from binding at the equivalent sites on the feline GIV.2 P domain. Additionally, an ELISA showed that the feline GIV.2 was antigenically distinct from a human GII.10 norovirus.
Publisher: American Society for Microbiology
Date: 03-2004
DOI: 10.1128/JCM.42.3.1305-1307.2004
Abstract: Stool specimens from hospitalized infants with sporadic gastroenteritis in Chiang Mai, Thailand, between July 2000 and July 2001 were examined for norovirus and sapovirus by reverse transcription-PCR and sequence analysis. These viruses were identified in 13 of 105 (12%) specimens. One strain was found to be a recombinant norovirus.
Publisher: Elsevier BV
Date: 12-2023
Publisher: CRC Press
Date: 19-04-2016
DOI: 10.1201/B13590
Publisher: American Society for Microbiology
Date: 03-2016
DOI: 10.1128/JVI.03119-15
Abstract: Recent reports suggest that human genogroup II genotype 17 (GII.17) noroviruses are increasing in prevalence. We analyzed the evolutionary changes of three GII.17 capsid protruding (P) domains. We found that the GII.17 P domains had little cross-reactivity with antisera raised against the dominant GII.4 strains. X-ray structural analysis of GII.17 P domains from 2002 to 2014 and 2015 suggested that surface-exposed substitutions on the uppermost part of the P domain might have generated a novel 2014-2015 GII.17 variant.
Publisher: WORLD SCIENTIFIC
Date: 2008
DOI: 10.1142/6324
Publisher: American Society for Microbiology
Date: 07-2007
DOI: 10.1128/JVI.02840-06
Abstract: A common feature of caliciviruses is the proteolytic processing of the viral polyprotein catalyzed by the viral 3C-like protease encoded in open reading frame 1 (ORF1). Here we report the identification and structural characterization of the protease domains and amino acid residues in sapovirus (SaV) and feline calicivirus (FCV). The in vitro expression and processing of a panel of truncated ORF1 polyproteins and corresponding mutant forms showed that the functional protease domain is 146 amino acids (aa) in SaV and 154 aa in FCV. Site-directed mutagenesis of the protease domains identified four amino acid residues essential to protease activities: H 31 , E 52 , C 116 , and H 131 in SaV and H 39 , E 60 , C 122 , and H 137 in FCV. A computer-assisted structural analysis showed that despite high levels of ersity in the primary structures of the protease domains in the family Caliciviridae , the configurations of the H, E, C, and H residues are highly conserved, with these residues positioned closely along the inner surface of the potential binding cleft for the substrate. These results strongly suggest that the H, E, C, and H residues are involved in the formation of a conserved catalytic surface of the SaV and FCV 3C-like proteases.
Publisher: Springer Science and Business Media LLC
Date: 20-02-2018
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 07-2007
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 07-2008
Publisher: Wiley
Date: 23-06-2006
DOI: 10.1016/J.FEBSLET.2006.06.040
Abstract: Human sapovirus (SaV) is uncultivable, but expression of the recombinant capsid protein (rVP1) in insect cells results in the formation of virus-like particles (VLPs) that are morphologically similar to the native viruses. However, the SaV rVP1 expression levels are considerably low. We have found that inclusions of short foreign nucleotide sequences inserted directly upstream from the predicted rVP1 AUG start codon lead to increased yield of VLPs. This method allowed us to express a SaV rVP1, which could not have been expressed to measurable or practical levels otherwise.
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 04-2007
Abstract: Human sapovirus was detected in 4 of 57 clam packages by reverse transcription-PCR and sequence analysis. This represents the first finding of sapovirus contamination in food. Closely matching sequences have been detected in stool specimens from patients with gastroenteritis in Japan, which indicates a possible food-to-human transmission link.
Publisher: Springer Science and Business Media LLC
Date: 24-09-2004
DOI: 10.1007/S00705-004-0406-8
Abstract: Sapovirus (SaV), a member of the genus Sapovirus in the family Caliciviridae, is an agent of human and porcine gastroenteritis. SaV strains are ided into five genogroups (GI-GV) based on their capsid (VP1) sequences. Human SaV strains are noncultivable, but expression of the recombinant capsid protein (rVP1) in a baculovirus expression system results in the self-assembly of virus-like particles (VLPs) that are morphologically similar to native SaV. In this study, rVP1 constructs of SaV GI, GII, and GV strains were expressed in a baculovirus expression system. The structures of the GI, GII, and GV VLPs, with diameters of 41-48 nm, were morphologically similar to those of native SaV. However a fraction of GV VLPs were smaller, with diameters of 26-31 nm and spikes on the outline. This is the first report of GII and GV VLP formation and the first identification of small VLPs. To examine the cross-reactivities among GI, GII, and GV rVP1, hyperimmune rabbit antisera were raised against Escherichia coli-expressed GI, GII, and GV N- and C-terminal VP1. Western blotting showed the GI antisera cross-reacted with GV rVP1 but not GII rVP1 GII antisera cross-reacted weakly with GI rVP1 but did not cross-react with GV rVP1 and GV antisera reacted only with GV rVP1. Also, hyperimmune rabbit and guinea pig antisera raised against purified GI VLPs were used to examine the cross-reactivities among GI, GII, and GV VLPs by an antigen enzyme-linked immunosorbent assay (ELISA). The ELISA showed that the GI VLPs were antigenically distinct from GII and GV VLPs.
Publisher: Elsevier BV
Date: 11-2015
DOI: 10.1016/J.VIROL.2015.07.009
Abstract: Human norovirus is a dominant cause of acute gastroenteritis around the world. Several norovirus disinfectants label citric acid as an active ingredient. In this study, we showed that norovirus virus-like particles (VLPs) treated with citrate buffer caused the particles to alter their morphology, including increased diameters associated with a new ring-like structure. We also found that epitopes on the protruding (P) domain on these particles were more readily accessible to antibodies after the citrate treatment. These results suggested that citrate had a direct effect on the norovirus particles. Using X-ray crystallography, we showed that the P domain bound citrate from lemon juice and a disinfectant containing citric acid. Importantly, citrate binds at the histo-blood group antigen binding pocket, which are attachment factors for norovirus infections. Taken together, these new findings suggested that it might be possible to treat/reduce norovirus infections with citrate, although further studies are needed.
Publisher: American Society for Microbiology
Date: 15-06-2005
DOI: 10.1128/JVI.79.12.7283-7290.2005
Abstract: The genome of Sapovirus (SaV), a causative agent of gastroenteritis in humans and swine, contains either two or three open reading frames (ORFs). Functional motifs characteristic to the 2C-like NTPase (NTPase), VPg, 3C-like protease (Pro), 3D-like RNA-dependent RNA polymerase (Pol), and capsid protein (VP1) are encoded in the ORF1 polyprotein, which is afterwards cleaved into the nonstructural and structural proteins. We recently determined the complete genome sequence of a novel human SaV strain, Mc10, which has two ORFs. To investigate the proteolytic cleavage of SaV ORF1 and the function of protease on the cleavage, both full-length and truncated forms of the ORF1 polyprotein either with or without mutation in 1171 Cys to Ala of the GDCG motif were expressed in an in vitro coupled transcription-translation system. The translation products were analyzed directly by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or by immunoprecipitation with region-specific antibodies. The ORF1 polyprotein was processed into at least 10 major proteins: p11, p28, p35, p32, p14, p70, p60, p66, p46, and p120. Seven of these products were arranged in the following order: NH 2 -p11-p28-p35(NTPase)-p32-p14(VPg)-p70(Pro-Pol)-p60(VP1)-COOH. p66, p46 and p120 were precursors of p28-p35 (NTPase), p32-p14 (VPg), and p32-p14 (VPg)-p70 (Pro-Pol), respectively. Mutagenesis in the 3C-like protease motif fully abolished the proteolytic activity. The cleavage map of SaV ORF1 is similar to those of other heretofore known members of the family Caliciviridae , especially to rabbit hemorrhagic disease virus, a member of the genus Lagovirus .
Publisher: American Society for Microbiology
Date: 27-04-2016
Abstract: Human norovirus interacts with the polymorphic human histo-blood group antigens (HBGAs), and this interaction is thought to be important for infection. The genogroup II genotype 4 (GII.4) noroviruses are the dominant cluster, evolve every other year, and are thought to modify their binding interactions with different HBGA types. Most human noroviruses bind HBGAs, while some strains were found to have minimal or no HBGA interactions. Here, we explain some possible structural constraints for several noroviruses that were found to bind poorly to HBGAs by using X-ray crystallography. We showed that one aspartic acid was flexible or positioned away from the fucose moiety of the HBGAs and this likely hindered binding, although other fucose-interacting residues were perfectly oriented. Interestingly, a neighboring loop also appeared to influence the loop hosting the aspartic acid. These new findings might explain why some human noroviruses bound HBGAs poorly, although further studies are required.
Publisher: American Society for Microbiology
Date: 15-01-2019
DOI: 10.1128/JVI.01581-18
Abstract: Given that human norovirus virions likely interact with bile acid during a natural infection, our evidence that an HBGA nonbinder (GII.1) can be converted to an HBGA binder after bile acid binding is of major significance. Our data provide direct evidence that, like HBGAs, bile acid interaction on the capsid is an important cofactor for certain genotypes. However, more unanswered questions seem to arise from these new discoveries. For ex le, is there an association between the bile acid requirement and the prevalence of certain genotypes? That is, the GII.1 and GII.10 (bile acid binders) genotypes rarely caused outbreaks, whereas the GII.4 and GII.17 genotypes (bile acid nonbinders) were responsible for large epidemics. Therefore, it seems plausible that certain genotypes require bile acids, whereas others have modified their bile acid requirements on the capsid.
Publisher: Elsevier BV
Date: 11-2008
DOI: 10.1016/J.VIRUSRES.2008.07.002
Abstract: We expressed full-length sapovirus genome constructs in insect cells and analyzed their products. The capsid protein was cleaved from the ORF1 polyprotein from a native-like genome construct and two full-length genome constructs with mutations in an active polymerase motif, whereas the capsid protein was not cleaved from a full-length genome construct with a mutation in an active protease motif. Our results showed that the sapovirus protease-polymerase precursor protein cleaved the capsid protein from the polyprotein at the putative conserved capsid start. Importantly, the cleaved capsid protein formed empty virus-like particles that were morphologically and antigenically similar to native sapovirus.
Publisher: IOP Publishing
Date: 17-01-2018
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 2007
Abstract: Sapoviruses are etiologic agents of human gastroenteritis. We detected sapovirus in untreated wastewater, treated wastewater, and a river in Japan. A total of 7 of 69 water s les were positive by reverse transcription-PCR. Phylogenetic analysis of the viral capsid gene grouped these strains into 4 genetic clusters.
Publisher: American Society for Microbiology
Date: 03-2016
DOI: 10.1128/JVI.02916-15
Abstract: The capsid protein (VP1) of all caliciviruses forms an icosahedral particle with two principal domains, shell (S) and protruding (P) domains, which are connected via a flexible hinge region. The S domain forms a scaffold surrounding the nucleic acid, while the P domains form a homodimer that interacts with receptors. The P domain is further sub ided into two subdomains, termed P1 and P2. The P2 subdomain is likely an insertion in the P1 subdomain consequently, the P domain is ided into the P1-1, P2, and P1-2 subdomains. In order to investigate capsid antigenicity, N-terminal (N-term)/S/P1-1 and P2/P1-2 were switched between two sapovirus genotypes GI.1 and GI.5. The chimeric VP1 constructs were expressed in insect cells and were shown to self-assemble into virus-like particles (VLPs) morphologically similar to the parental VLPs. Interestingly, the chimeric VLPs had higher levels of cross-reactivities to heterogeneous antisera than the parental VLPs. In order to better understand the antigenicity from a structural perspective, we determined an intermediate-resolution (8.5-Å) cryo-electron microscopy (cryo-EM) structure of a chimeric VLP and developed a VP1 homology model. The cryo-EM structure revealed that the P domain dimers were raised slightly (∼5 Å) above the S domain. The VP1 homology model allowed us predict the S domain (67–229) and P1-1 (229–280), P2 (281–447), and P1-2 (448–567) subdomains. Our results suggested that the raised P dimers might expose immunoreactive S/P1-1 subdomain epitopes. Consequently, the higher levels of cross-reactivities with the chimeric VLPs resulted from a combination of GI.1 and GI.5 epitopes. IMPORTANCE We developed sapovirus chimeric VP1 constructs and produced the chimeric VLPs in insect cells. We found that both chimeric VLPs had a higher level of cross-reactivity against heterogeneous VLP antisera than the parental VLPs. The cryo-EM structure of one chimeric VLP (Yokote/Mc114) was solved to 8.5-Å resolution. A homology model of the VP1 indicated for the first time the putative S and P (P1-1, P2, and P1-2) domains. The overall structure of Yokote/Mc114 contained features common among other caliciviruses. We showed that the P2 subdomain was mainly involved in the homodimeric interface, whereas a large gap between the P1 subdomains had fewer interactions.
Publisher: American Society for Microbiology
Date: 27-06-2023
Publisher: CRC Press
Date: 28-07-2009
Publisher: CRC Press
Date: 25-11-2016
Publisher: American Society for Microbiology
Date: 07-2011
DOI: 10.1128/JVI.00246-11
Abstract: Noroviruses are the dominant cause of outbreaks of gastroenteritis worldwide, and interactions with human histo-blood group antigens (HBGAs) are thought to play a critical role in their entry mechanism. Structures of noroviruses from genogroups GI and GII in complex with HBGAs, however, reveal different modes of interaction. To gain insight into norovirus recognition of HBGAs, we determined crystal structures of norovirus protruding domains from two rarely detected GII genotypes, GII.10 and GII.12, alone and in complex with a panel of HBGAs, and analyzed structure-function implications related to conservation of the HBGA binding pocket. The GII.10- and GII.12-apo structures as well as the previously solved GII.4-apo structure resembled each other more closely than the GI.1-derived structure, and all three GII structures showed similar modes of HBGA recognition. The primary GII norovirus-HBGA interaction involved six hydrogen bonds between a terminal αfucose1-2 of the HBGAs and a dimeric capsid interface, which was composed of elements from two protruding subdomains. Norovirus interactions with other saccharide units of the HBGAs were variable and involved fewer hydrogen bonds. Sequence analysis revealed a site of GII norovirus sequence conservation to reside under the critical αfucose1-2 and to be one of the few patches of conserved residues on the outer virion-capsid surface. The site was smaller than that involved in full HBGA recognition, a consequence of variable recognition of peripheral saccharides. Despite this evasion tactic, the HBGA site of viral vulnerability may provide a viable target for small molecule- and antibody-mediated neutralization of GII norovirus.
Publisher: Wiley
Date: 2007
DOI: 10.1002/JMV.20906
Abstract: Norovirus (NoV) capsid proteins were expressed as virus-like particles (VLPs) by using recombinant baculovirus in insect cells, which had 5 genotypes in genogroup I and 11 genotypes in genogroup II, and the VLPs were used as immunogens. Polyclonal antibody against the VLP of GII/3 genotype showed broad-range cross-reactivity, reacting not only with intra-genogroup strains, but also inter-genogroup strains, by antibody-ELISA using 16 kinds of VLPs. Furthermore, antigen-ELISA was conducted in sandwich enzyme-linked immunosorbent assay (ELISA) using the polyclonal antibody for capturing antigens, and three kinds of monoclonal antibodies against the VLP of GII/4 genotype for detecting antigens. This format successfully detected eight genotypes of NoV from clinical specimens and proved that polyclonal antibody, which has broad-range cross-reactivity, was capable of detecting various types of genotypes from clinical specimens.
Publisher: American Society for Microbiology
Date: 15-11-2008
DOI: 10.1128/JVI.00897-08
Abstract: Our norovirus (NoV) surveillance group reported a -fold increase in NoV infection in Japan during the winter of 2006-2007 compared to the previous winter. Because the increase was not linked to changes in the surveillance system, we suspected the emergence of new NoV GII/4 epidemic variants. To obtain information on viral changes, we conducted full-length genomic analysis. Stool specimens from 55 acute gastroenteritis patients of various ages were collected at 11 sites in Japan between May 2006 and January 2007. Direct sequencing of long PCR products revealed 37 GII/4 genome sequences. Phylogenetic study of viral genome and partial sequences showed that the two new GII/4 variants in Europe, termed 2006a and 2006b, initially coexisted as minorities in early 2006 in Japan and that 2006b alone had dominated over the resident GII/4 variants during 2006. A combination of phylogenetic and entropy analyses revealed for the first time the unique amino acid substitutions in all eight proteins of the new epidemic strains. These data and computer-assisted structural study of the NoV capsid protein are compatible with a model of antigenic drift with tuning of the structure and functions of multiple proteins for the global outgrowth of new GII/4 variants. The availability of comprehensive information on genome sequences and unique protein changes of the recent global epidemic variants will allow studies of diagnostic assays, molecular epidemiology, molecular biology, and adaptive changes of NoV in nature.
Publisher: American Society for Microbiology
Date: 15-02-2015
DOI: 10.1128/JVI.02968-14
Abstract: Human noroviruses are the dominant cause of outbreaks of gastroenteritis around the world. Human noroviruses interact with the polymorphic human histo-blood group antigens (HBGAs), and this interaction is thought to be important for infection. Indeed, synthetic HBGAs or HBGA-expressing enteric bacteria were shown to enhance norovirus infection in B cells. A number of studies have found a possible relationship between HBGA type and norovirus susceptibility. The genogroup II, genotype 4 (GII.4) noroviruses are the dominant cluster, evolve every other year, and are thought to modify their binding interactions with different HBGA types. Here we show high-resolution X-ray crystal structures of the capsid protruding (P) domains from epidemic GII.4 variants from 2004, 2006, and 2012, cocrystallized with a panel of HBGA types (H type 2, Lewis Y, Lewis B, Lewis A, Lewis X, A type, and B type). Many of the HBGA binding interactions were found to be complex, involving capsid loop movements, alternative HBGA conformations, and HBGA rotations. We showed that a loop (residues 391 to 395) was elegantly repositioned to allow for Lewis Y binding. This loop was also slightly shifted to provide direct hydrogen- and water-mediated bonds with Lewis B. We considered that the flexible loop modulated Lewis HBGA binding. The GII.4 noroviruses have dominated outbreaks over the past decade, which may be explained by their exquisite HBGA binding mechanisms, their fondness for Lewis HBGAs, and their temporal amino acid modifications. IMPORTANCE Our data provide a comprehensive picture of GII.4 P domain and HBGA binding interactions. The exceptionally high resolutions of our X-ray crystal structures allowed us to accurately recognize novel GII.4 P domain interactions with numerous HBGA types. We showed that the GII.4 P domain-HBGA interactions involved complex binding mechanisms that were not previously observed in norovirus structural studies. Many of the GII.4 P domain-HBGA interactions we identified were negative in earlier enzyme-linked immunosorbent assay (ELISA)-based studies. Altogether, our data show that the GII.4 norovirus P domains can accommodate numerous HBGA types.
Publisher: American Society for Microbiology
Date: 15-03-2019
DOI: 10.1128/JVI.02005-18
Abstract: The discovery of vulnerable regions on norovirus particles is instrumental in the development of effective inhibitors, particularly for GI noroviruses that are genetically erse. Analysis of these GI.1-specific Nanobodies has shown that similar to GII norovirus particles, the GI particles have vulnerable regions. The only known cofactor region, the HBGA binding pocket, represents the main target for inhibition. With a combination treatment, i.e., the addition of Nano-7 or Nano-94 with 2′FL, the effect of inhibition was increased. Therefore, combination drug treatments might offer a better approach to combat norovirus infections, especially since the GI genotypes are highly erse and are continually changing the capsid landscape, and few conserved epitopes have so far been identified.
Publisher: Springer Science and Business Media LLC
Date: 08-2005
DOI: 10.1007/S00705-005-0591-0
Abstract: We recently determined the ORF1 cleavage map of Mc10, a human sapovirus (SaV) strain, as follows: NH2-p11-p28-p35(NTPase)-p32-p14(VPg)-p70(Pro-Pol)-p60(VP1)-COOH. This cleavage was dependent on the viral encoded 3C-like protease. To identify the cleavage site of SaV ORF1, putative p70 (Pro-Pol) and p14-p70 (VPg-Pro-Pol) were expressed as N-terminal GST and C-terminal 6 x His-tag fusion proteins in Escherichia coli, and the expressed products were analyzed by SDS-PAGE and Western blotting. Our results indicated that the efficient proteolytic cleavage occurred between p14 (VPg) and p70 (Pro-Pol), and N-terminal amino acid sequencing revealed that the cleavage site was between E(1055) and A(1056). In contrast, the p70 (Pro-Pol) was not further cleaved. We also found that SaV protease cleaved the Q/G site within the rhinovirus 3C protease recognition site. Site-directed mutagenesis in a conserved GDCG motif of the protease completely abolished these proteolytic activities. This is the first report to identify the cleavage site of the SaV ORF1 polyprotein.
Publisher: Elsevier BV
Date: 08-2017
DOI: 10.1016/J.VIROL.2017.04.032
Abstract: Human noroviruses are the leading cause of outbreaks of acute gastroenteritis. Norovirus interactions with histo-blood group antigens (HBGAs) are known to be important for an infection. In this study, we identified the HBGA binding pocket for an emerging GII genotype 17 (GII.17) variant using X-ray crystallography. The GII.17 variant bound the HBGA with an equivalent set of residues as the leading pandemic GII.4 variants. These structural data highlights the conserved nature of HBGA binding site between prevalent GII noroviruses. Noroviruses also interact with human milk oligosaccharides (HMOs), which mimic HBGAs and may function as receptor decoys. We previously showed that HMOs inhibited the binding of rarely detected GII.10 norovirus to HBGAs. We now found that an HMO, 2'-fucosyllactose (2'FL), additionally blocked both the GI.1 and GII.17 noroviruses from binding to HBGAs. Together, these findings provide evidence that 2'FL might function as a broadly reactive antiviral against multiple norovirus genogroups.
Publisher: Elsevier BV
Date: 08-2008
DOI: 10.4315/0362-028X-71.8.1689
Abstract: A total of 57 clam packages that were collected from supermarkets and fish markets from 11 different sites in western Japan between 8 December 2005 and 6 September 2006 were examined for human enteric viruses (i.e., norovirus, Aichi virus, rotavirus, adenovirus, hepatitis A virus, and astrovirus), using PCR and reverse transcription PCR. Sixty-one percent of the packages were contaminated with one type of virus, 9% had two different types of viruses, 28% had three different types of viruses, and 9% had at least four different types of viruses. Thirty-one (54%) of 57 packages were contaminated with noroviruses. Norovirus genogroup I and genogroup II sequences were detected in 24 and 23 packages, respectively, and these sequences belonged to nine genogroup I and eight genogroup II genotypes. Aichi viruses were found in 19 (33%) of 57 packages, and these belonged to genogroup A. Rotaviruses (group A) were detected in 14 (42%) of 33 of packages and 9 of 14 rotavirus-positive packages contained two or more rotavirus genogroup types. Adenoviruses (Ad40 and Ad41) were detected in 17 (52%) of 33 packages. One of the 57 (2%) packages was positive with hepatitis A virus (subtype IA). Astrovirus was not detected in any of the packages. This is the first study to detect such a high level of contamination in Japanese clams. These results represent an important finding because the Japanese clams were considered suitable for human consumption. Further studies are needed to determine the health risks associated with eating these highly contaminated clams.
Publisher: Springer Science and Business Media LLC
Date: 08-2005
DOI: 10.1007/S00705-005-0599-5
Abstract: Human sapovirus (SaV) strains are agents of gastroenteritis. They cannot be grown in cell culture. In this study, constructs containing SaV N- and C-terminal-deleted recombinant capsid proteins (rVP1) were expressed in a baculovirus expression system to allow us to better understand the sequence requirements for the formation of virus-like particles (VLPs). Only proteins derived from N-terminal-deleted rVP1 constructs that began 49 nucleotides downstream assembled into VLPs, which included both small and native-size VLPs. Our results were similar to those reported in a rabbit hemorrhagic disease virus (RHDV) N- and C-terminal-deleted rVP1 expression study but were distinct from those reported in a norovirus N- and C-terminal-deleted rVP1 expression study, suggesting that SaV and RHDV may have similar expression requirements.
Publisher: Elsevier BV
Date: 2016
DOI: 10.1016/J.VIROL.2015.10.022
Abstract: We determined a structure of a bovine (genogroup III, GIII) norovirus capsid protruding (P) domain using X-ray crystallography. The bovine P domain was reminiscent of other norovirus genogroups (GI, GII, GIV, and GV), but closely matched the human GI P domain. We also identified a monoclonal antibody that was capable of binding the five different (GI-GV) P domains. Our data suggests that genetically erse noroviruses still contain common epitopes.
Publisher: Oxford University Press (OUP)
Date: 03-08-2016
Abstract: Recently, combined nuclear magnetic resonance (NMR), native mass spectrometry (MS) and X-ray crystallographic studies have demonstrated that binding of histo-blood group antigens (HBGAs) to norovirus capsid protein (P-dimers) is a cooperative process involving four binding pockets. Here, we show that binding to norovirus virus-like particles (VLPs) is even more complex. We performed saturation transfer difference (STD) NMR titration experiments with two representative genotypes of norovirus VLPs using l-fucose as a minimal HBGA. Compared to titrations with P-dimers, the corresponding binding isotherms reflect at least six distinct binding events.
Publisher: Wiley
Date: 19-08-2015
Abstract: Human noroviruses recognize histo blood group antigens (HBGAs) as cellular attachment factors. Recently, it has been discovered that norovirus infection can be significantly enhanced by HBGA binding. Yet the attachment process and how it promotes host-cell entry is only poorly understood. The binding of a norovirus protruding (P) domain of a predominant GII.4 Saga strain to HBGAs at atomic resolution was studied. So far, independent and equivalent multiple binding sites were held responsible for attachment. Using NMR experiments we show that norovirus-HBGA binding is a cooperative multi-step process, and native mass spectrometry reveals four instead of two HBGA binding sites per P-dimer. An accompanying crystallographic study has disclosed four instead of two L-fucose binding sites per P-dimer of a related GII.10 strain1 further supporting our findings. We have uncovered a novel paradigm for norovirus-HBGA recognition that will inspire further studies into norovirus-host interactions.
Publisher: CRC Press
Date: 14-01-2009
Publisher: American Chemical Society (ACS)
Date: 02-08-2018
DOI: 10.1021/ACS.BIOMAC.8B00829
Abstract: Norovirus infection is the major cause of nonbacterial gastroenteritis in humans and has been the subject of numerous studies investigating the virus's biophysical properties and biochemical function with the aim of deriving novel and highly potent entry inhibitors to prevent infection. Recently, it has been shown that the protruding P domain dimer (P-dimer) of a GII.10 Norovirus strain exhibits two new binding sites for l-fucose in addition to the canonical binding sites. Thus, these sites provide a novel target for the design of multivalent fucose ligands as entry inhibitors of norovirus infections. In this current study, a first generation of multivalent fucose-functionalized glycomacromolecules was synthesized and applied as model structures to investigate the potential targeting of fucose binding sites in human norovirus P-dimer. Following previously established solid phase polymer synthesis, eight precision glycomacromolecules varying in number and position of fucose ligands along an oligo(amidoamine) backbone were obtained and then used in a series of binding studies applying native MS, NMR, and X-ray crystallography. We observed only one fucose per glycomacromolecule binding to one P-dimer resulting in similar binding affinities for all fucose-functionalized glycomacromolecules, which based on our current findings we attribute to the overall size of macromolecular ligands and possibly to steric hindrance.
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 2005
Publisher: Elsevier BV
Date: 02-2014
DOI: 10.1016/J.VIROL.2013.12.007
Abstract: Global surveillance for norovirus identified in 2012 the emergence of a novel pandemic GII.4 variant, termed Sydney 2012. In Italy, the novel pandemic variant was identified as early as November 2011 but became predominant only in the winter season 2012-2013. Upon sequencing and comparison with strains of global origin, the early Sydney 2012 strains were found to differ from those spreading in 2012-2013 in the capsid (ORF2) putative epitopes B, C and D, segregating into a distinct phylogenetic clade. At least three residues (333, 340 and 393, in epitopes B, C and D, respectively) of the VP1 varied among Sydney 2012 strains of different clades. These findings suggest that the spread of the pandemic variant in Italy during the winter season 2012-2013 was due to the introduction of strains distinct from those circulating at low frequency in the former winter season and that similar strains were also circulating elsewhere worldwide.
Publisher: CRC Press
Date: 28-07-2009
Publisher: Public Library of Science (PLoS)
Date: 02-11-2017
Publisher: Springer Science and Business Media LLC
Date: 26-02-2006
DOI: 10.1007/S00705-005-0720-9
Abstract: Noroviruses (NoVs) belong to the genus Norovirus and are members of the family Caliciviridae. NoVs are the dominant cause of outbreaks of gastroenteritis, but progress in understanding the molecular characteristics of NoV and its replication strategies have been h ered by the lack of a cell culture system or a practical animal model, except for murine NoVs. To elucidate the transcription and replication of the NoV genome, a complete genome of a human NoV genogroup II strain was cloned downstream of a T7 RNA polymerase promoter and expressed in human embryonic kidney (HEK) 293T/17 cells using a T7 vaccinia virus expression system. Bands for a 7.6-kb negative-strand RNA, a 7.6-kb positive-strand genomic RNA, and a 2.6-kb positive-strand subgenomic-like RNA were found in the infected cells. However, recombinant capsid protein (rVP1) and rVP2 were not detected by Western blotting. When a construct containing VP1 and VP2 genes was co-transfected with a full-length construct, the expression of virus-like particles (VLPs) with a buoyant density of 1.271 g/cm3 was observed. We also observed round particles, 20 to 80 nm in diameter, with a buoyant density of 1.318 g/cm3. Our results indicated that NoV RNA was incorporated into the heavier particles. However, further studies are needed to investigate the antigenicity of these particles and to determine if they represent undeveloped VLPs.
Publisher: American Society for Microbiology
Date: 05-2016
DOI: 10.1128/JVI.03223-15
Abstract: Histo-blood group antigens (HBGAs) are important binding factors for norovirus infections. We show that two human milk oligosaccharides, 2′-fucosyllactose (2′FL) and 3-fucosyllactose (3FL), could block norovirus from binding to surrogate HBGA s les. We found that 2′FL and 3FL bound at the equivalent HBGA pockets on the norovirus capsid using X-ray crystallography. Our data revealed that 2′FL and 3FL structurally mimic HBGAs. These results suggest that 2′FL and 3FL might act as naturally occurring decoys in humans.
Publisher: American Society for Microbiology
Date: 15-02-2015
DOI: 10.1128/JVI.02832-14
Abstract: Rabbit hemorrhagic disease virus (RHDV) is a member of the Caliciviridae family ( Lagovirus genus). RHDV is highly contagious and attaches to epithelial cells in the digestive or respiratory tract, leading to massive lesions with high mortality rates. A new variant of RHDV (termed RHDVb) recently has emerged, and previously vaccinated rabbits appear to have little protection against this new strain. Similar to human norovirus ( Caliciviridae , Norovirus genus), RHDV binds histo-blood group antigens (HBGAs), and this is thought to be important for infection. Here, we report the HBGA binding site on the RHDVb capsid-protruding domain (P domain) using X-ray crystallography. The HBGA binding pocket was located in a negatively charged patch on the side of the P domain and at a dimeric interface. Residues from both monomers contributed to the HBGA binding and involved a network of direct hydrogen bonds and water-mediated interactions. An amino acid sequence alignment of different RHDV strains indicated that the residues directly interacting with the ABH-fucose of the HBGAs (Asp472, Asn474, and Ser479) were highly conserved. This result suggested that different RHDV strains also could bind HBGAs at the equivalent pocket. Moreover, several HBGA binding characteristics between RHDVb and human genogroup II norovirus were similar, which indicated a possible convergent evolution of HBGA binding interactions. Further structural studies with other RHDV strains are needed in order to better understand the HBGA binding mechanisms among the erse RHDV strains. IMPORTANCE We identified, for the first time, the HBGA binding site on an RHDVb P domain using X-ray crystallography. Our results showed that RHDVb and human genogroup II noroviruses had similar HBGA binding interactions. Recently, it was discovered that synthetic HBGAs or HBGA-expressing enteric bacteria could enhance human genogroup II norovirus infection in B cells. Considering that RHDVb and genogroup II norovirus similarly interacted with HBGAs, it may be possible that a comparable cell culture system also could work with RHDVb. Taken together, these new findings will extend our understanding of calicivirus HBGA interactions and may help to elucidate the specific roles of HBGAs in calicivirus infections.
Publisher: Springer Science and Business Media LLC
Date: 17-11-2009
DOI: 10.1038/NGEO689
Publisher: Wiley
Date: 2006
DOI: 10.1002/JMV.20699
Abstract: Sapovirus (SaV) is an agent of gastroenteritis for humans and swine, and is ided into five distinct genogroups (GI-GV) based on its capsid gene sequences. Typical methods of SaV detection include electron microscopy (EM), enzyme-linked immunosorbent assay (ELISA), and reverse transcription-polymerase chain reaction (RT-PCR). A novel TaqMan-based real-time RT-PCR assay was developed that is sensitive and has the ability to detect the broad range of genetically erse human SaV strains. A nucleotide alignment of 10 full-length SaV genome sequences was subjected to similarity plot analysis, which indicated that the most conserved site was the polymerase-capsid junction in open reading frame 1 (ORF1). Based on multiple alignments of the 27 available sequences encoding this junction, we designed sets of primers and TaqMan MGB probes that detect human SaV GI, GII, GIV, and GV sequences in a single tube. The reactivity was confirmed with SaV GI, GII, GIV, and GV control plasmids, and the efficiency ranged from 2.5 x 10(7) to 2.5 x 10(1) copies per tube. Analysis using clinical stool specimens revealed that the present system was capable of detecting SaV GI, GII, GIV, and GV sequences, and no cross-reactivity was observed against other enteric viruses, including norovirus (NoV), rotavirus, astrovirus, and adenovirus. This is the first real-time RT-PCR system that could detect all genogroups of human sapoviruses.
Publisher: Cold Spring Harbor Laboratory
Date: 07-12-2018
DOI: 10.1101/489906
Abstract: Temporal changes in the GII.4 human norovirus capsid sequences occasionally result in the emergence of genetic variants capable of causing new epidemics. The GII.4 persistence is believed to be associated with the recognition of numerous histo-blood group antigen (HBGA) types and antigenic drift. We found that one of the earliest known GII.4 isolate (1974) and a more recent epidemic GII.4 variant (2012) had varied norovirus-specific monoclonal antibody (MAb) reactivities, yet similar HBGA binding profiles. To better understand the binding interaction of one MAb (10E9) that had varied reactivities with these GII.4 variants, we determined the X-ray crystal structure of the NSW-2012 GII.4 P domain 10E9 Fab complex. We showed that the 10E9 Fab interacted with conserved and variable residues, which could be associated with antigenic drift. Interestingly, the 10E9 Fab binding pocket partially overlapped the HBGA pocket and haddirect competition for conserved HBGA binding residues (i.e., Arg345 and Tyr444). Indeed, the 10E9 MAb blocked norovirus VLPs from binding to several sources of HBGAs. Moreover, the 10E9 antibody completely abolished virus replication in the human norovirus intestinal enteroid cell culture system. Our new findings provide first direct evidence that competition for GII.4 HBGA binding residues and steric obstruction could lead to norovirus neutralization. On the other hand, the 10E9 MAb recognized residues flanking the HBGA pocket, which are often substituted as the virus evolves. This mechanism of antigenic drift likely influences herd immunityand impedes the possibility of acquiring broadly reactive HBGA-blocking antibodies.
Publisher: Wiley
Date: 2006
DOI: 10.1002/JMV.20696
Abstract: Norovirus (NoV) is recognized as one of the most common causative agent of diarrheal disease in young children worldwide. The current study was undertaken to determine the distribution of NoV genotypes in Japan. A total of 3,864 fecal specimens from children with acute gastroenteritis in five regions (Tokyo, Maizuru, Saga, Sapporo, and Osaka) of Japan from July 1995 to June 2001 were collected and then tested for the presence of NoV by RT-PCR. Three hundred sixty four were found to be positive for NoV, accounting for 11%. The highest prevalence of NoV infection was in November, December, and January as the early winter months in Japan. NoV was subjected to be further characterized to sequencing analysis. All NoVs belonged to two different genogroups I and II and these represented 3% and 97%, respectively. This finding indicated that NoV genogroup II was the dominant group causing acute gastroenteritis in Japan. Interestingly, NoV strains were classified into 16 distinct genotypes including genogroup II genotype 9 that was firstly identified in Japan. Of these, NoV genogroup II genotypes 3 and 4 dominated over other genotypes and became the leading strains in Japanese pediatric population. In conclusion, diarrhea due to NoV infection is still a health burden in Japan. This report also stresses the great genetic ersity as well as the importance of NoV causing the diarrhea in Japan.
Publisher: Elsevier BV
Date: 07-2018
Publisher: Springer Science and Business Media LLC
Date: 12-09-2005
DOI: 10.1007/S00705-005-0630-X
Abstract: Human sapovirus (SaV) strains are etiological agents of mild and/or acute gastroenteritis in children and adults. In this study, we describe the development of a novel antigen enzyme-linked immunosorbent assay (ELISA) detection system that was based on hyperimmune rabbit and guinea pig antisera raised against SaV genogroup I (GI) virus-like particles. The ELISA had 100% specificity, and sensitivities of 60% and 25% when compared to single-round PCR and nested PCR, respectively. Our results have shown the ELISA was useful in detecting SaV GI antigens in clinical stool specimens collected two days after the onset of illness.
Publisher: Springer Science and Business Media LLC
Date: 27-11-2006
DOI: 10.1007/S00705-006-0883-Z
Abstract: Noroviruses (NoVs) and sapoviruses (SaVs) are causative agents of human gastroenteritis. There is increasing evidence that certain human NoV strains bind to histo-blood group antigens (HBGAs). We found that several NoV virus-like particles (VLPs) showed binding activity to HBGAs, while neither SaV genogroup I (GI) VLP nor SaV GV VLP showed such activity.
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 10-2004
Abstract: We determined the complete genome sequences of two sapovirus strains isolated in Thailand and Japan. One of these strains represented a novel, naturally occurring recombinant sapovirus. Evidence suggested the recombination site was at the polymerase-capsid junction within open reading frame one.
Publisher: Microbiology Society
Date: 2017
DOI: 10.1099/JGV.0.000658
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 2006
Abstract: Sapovirus was detected in 7 of 95 stool specimens from children with gastroenteritis of unknown etiology in Sydney, Australia, from August 2001 to August 2002 and from February 2004 to August 2004, by using reverse transcription-polymerase chain reaction. Sequence analysis of the N-terminal capsid region showed all human sapovirus genogroups.
Publisher: Informa UK Limited
Date: 15-09-2023
Start Date: 2017
End Date: 2020
Funder: Baden-Württemberg Stiftung
View Funded ActivityStart Date: 2012
End Date: 2016
Funder: Deutsche Forschungsgemeinschaft
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