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Globally, noroviruses are associated with [approximately equal to] 20% of acute gastroenteritis (AGE) cases, causing an estimated 685 million episodes and 210,000 deaths each year (2,2). By 2 years of [...]

We investigated the molecular epidemiology of human norovirus (HuNoV) in all age groups using samples from April 2019 to March 2023, before and after the COVID-19 countermeasures were implemented. GII.2[P16] and GII.4[P31], the prevalent strains in Japan before COVID-19 countermeasures, remained prevalent during the COVID-19 pandemic, except from April to November 2020; in 2021, the prevalence of GII.2[P16] increased among children. Furthermore, there was an increase in the prevalence of GII.4[P16] after December 2022. Phylogenetic analysis of GII.P31 RdRp showed that some strains detected in 2022 belonged to a different cluster of other strains obtained during the present study period, suggesting that HuNoV strains will evolve differently even if they have the same type of RdRp. An analysis of the amino acid sequence of VP1 showed that some antigenic sites of GII.4[P16] were different from those of GII.4[P31]. The present study showed high infectivity of HuNoV despite the COVID-19 countermeasures and revealed changes in the prevalent genotypes and mutations of each genotype. In the future, we will investigate whether GII.4[P16] becomes more prevalent, providing new insights by comparing the new data with those analyzed in the present study. [ABSTRACT FROM AUTHOR]

Seven foodborne norovirus outbreaks were reported in remote areas across Japan during January-February 2017, causing illness in 2,094 persons (Figure). In all outbreaks, norovirus GII.P17-GII.17 was detected in stool specimens [...]

Norovirus is a major cause of acute gastroenteritis in humans and is genetically classified into 7 genogroups (GI-GVII). Among them, norovirus GI, GII, and GIV infect humans, and human norovirus [...]

Norovirus infections are a leading cause of acute gastroenteritis worldwide in persons of all age groups. Despite the broad genetic diversity, norovirus GII.4 has predominated during the past 20 years [...]

Sapoviruses, like noroviruses, are single-stranded positive-sense RNA viruses classified in the family Caliciviridae and are recognized as a causative pathogen of diarrhea in infants and the elderly. Like human norovirus, human sapovirus (HuSaV) has long been difficult to replicate in vitro. Recently, it has been reported that HuSaV can be replicated in vitro by using intestinal epithelial cells (IECs) derived from human tissues and cell lines derived from testicular and duodenal cancers. In this study, we report that multiple genotypes of HuSaV can sufficiently infect and replicate in human-induced pluripotent stem cell-derived IECs. We also show that this HuSaV replication system can be used to investigate the conditions for inactivation of HuSaV by heat and alcohol, and the effects of virus neutralization of antisera obtained by immunization with vaccine antigens, under conditions closer to the living environment. The results of this study confirm that HuSaV can also infect and replicate in human normal IECs regardless of their origin and are expected to contribute to future virological studies. [ABSTRACT FROM AUTHOR]

Passive administration of neutralizing antibodies (Abs) is an attractive strategy for the control of gastrointestinal infections. However, an unanswered practical concern is the need to assure the stability of sufficient amounts of orally administered neutralizing Abs against intestinal pathogens (e.g., norovirus) in the harsh environment of the gastrointestinal tract. To this end, we expressed a single-domain Ab (VHH, nanobody) against norovirus on the cell surface of Lactobacillus, a natural and beneficial commensal component of the gut microbiome. First, we used intestinal epithelial cells generated from human induced pluripotent stem cells to confirm that VHH 1E4 showed neutralizing activity against GII.17 norovirus. We then expressed VHH 1E4 as a cell-wall–anchored form in Lactobacillus paracasei BL23. Flow cytometry confirmed the expression of VHH 1E4 on the surface of lactobacilli, and L. paracasei that expressed VHH 1E4 inhibited the replication of GII.17 norovirus in vitro. We then orally administered VHH 1E4-expressing L. paracasei BL23 to germ-free BALB/c mice and confirmed the presence of lactobacilli with neutralizing activity in the intestine for at least 10 days after administration. Thus, cell-wall-anchored VHH-displaying lactobacilli are attractive oral nanobody deliver vectors for passive immunization against norovirus infection. [ABSTRACT FROM AUTHOR]

(1) Background: General infection control measures have been implemented at the societal level against COVID-19 since the middle of 2020, namely, hand hygiene, universal masking, and social distancing. The suppressive effect of the social implementation of general infection control measures on pediatric infections has not been systematically assessed. (2) Methods: We addressed this issue based on publicly available data on 11 pediatric infections reported weekly by sentinel sites in Osaka and Iwate prefectures in Japan since 2010. We obtained the 5-year average for 2015–2019 and compared it to the weekly report for 2020–2021. (3) Results: The rate of 6 of the 11 pediatric infections decreased significantly during 2020–2021, regardless of the magnitude of the prevalence of COVID-19 in both areas. However, only RSV infection, one of the six infections, was endemic in 2021. Exanthem subitum was not as affected by COVID-19 countermeasures as other diseases. (4) Conclusions: The social implementation of infectious disease control measures was effective in controling certain infectious diseases in younger age groups, where compliance with the countermeasures should not be as high as that of adults. [ABSTRACT FROM AUTHOR]

Sapovirus (SaV) is a causative agent of gastroenteritis. On the basis of capsid protein (VP1) nucleotide sequences, SaV can be divided into 5 genogroups (GI-GV), of which the GI, GII, [...]

Human norovirus is the leading cause of acute nonbacterial gastroenteritis in people of all ages worldwide. Currently, no licensed norovirus vaccine, pharmaceutical drug, or therapy is available for the control of norovirus infection. Here, we used a rice transgenic system, MucoRice, to produce a variable domain of a llama heavy-chain antibody fragment (VHH) specific for human norovirus (MucoRice-VHH). VHH is a small heat- and acid-stable protein that resembles a monoclonal antibody. Consequently, VHHs have become attractive and useful antibodies (Abs) for oral immunotherapy against intestinal infectious diseases. MucoRice-VHH constructs were generated at high yields in rice seeds by using an overexpression system with RNA interference to suppress the production of the major rice endogenous storage proteins. The average production levels of monomeric VHH (7C6) to GII.4 norovirus and heterodimeric VHH (7C6-1E4) to GII.4 and GII.17 noroviruses in rice seed were 0.54 and 0.28% (w/w), respectively, as phosphate buffered saline (PBS)-soluble VHHs. By using a human norovirus propagation system in human induced pluripotent stem-cell-derived intestinal epithelial cells (IECs), we demonstrated the high neutralizing activity of MucoRice expressing monomeric VHH (7C6) against GII.4 norovirus and of heterodimeric VHH (7C6-1E4) against both GII.4 and GII.17 noroviruses. In addition, MucoRice-VHH (7C6-1E4) retained neutralizing activity even after heat treatment at 90°C for 20 min. These results build a fundamental platform for the continued development of MucoRice-VHH heterodimer as a candidate for oral immunotherapy and for prophylaxis against GII.4 and GII.17 noroviruses in not only healthy adults and children but also immunocompromised patients and the elderly. [ABSTRACT FROM AUTHOR]

Human noroviruses cause an estimated 685 million infections and 200 000 deaths annually worldwide. Although vaccines against GII.4 and GI.1 genotypes are under development, no information is available regarding vaccines or monoclonal antibodies to other noroviral genotypes. Here, we developed 2 variable-domain llama heavy-chain antibody fragment (VHHs) clones, 7C6 and 1E4, against GII.4 and GII.17 human noroviruses, respectively. Although 7C6 cross-reacted with virus-like particles (VLPs) of GII.17, GII.6, GII.3, and GII.4, it neutralized only GII.4 norovirus. In contrast, 1E4 reacted with and neutralized only GII.17 VLPs. Both VHHs blocked VLP binding to human induced pluripotent stem cell-derived intestinal epithelial cells and carbohydrate attachment factors. Using these 2 VHHs, we produced a heterodimeric VHH fragment that neutralized both GII.4 and GII.17 noroviruses. Because VHH fragments are heat- and acid-stable recombinant monoclonal antibodies, the heterodimer likely will be useful for oral immunotherapy and prophylaxis against GII.4 and GII.17 noroviruses in young, elderly, or immunocompromised persons. [ABSTRACT FROM AUTHOR]

Some phthalates, which are used mainly as plasticizers, are suspected to be endocrine disruptors. In the present study, daily intakes of phthalates by Japanese children through all exposure pathways and the contribution of indoor air quality to the intakes were examined by measuring urinary phthalate metabolites in the children and the airborne phthalates in their residences. Spot urine samples excreted first after waking up in the morning were collected from the subjects aged 6 to 15 years (n = 132), and airborne phthalates were sampled in the subjects' bedrooms for 24 h. Eight airborne phthalates and their urinary metabolites were determined by gas chromatography/mass spectrometry. The daily intakes of the phthalates estimated were as follows (median, μg/kg b.w./day): dimethyl phthalate (DMP), 0.15; diethyl phthalate (DEP), 0.42; diisobutyl phthalate (DiBP), 1.1; di-n-butyl phthalate (DnBP), 2.2; dicyclohexyl phthalate (DcHP), 0.026; benzylbutyl phthalate (BBzP), 0.032; di(2-ethylhexyl) phthalate (DEHP), 6.3. The 95th percentile (21 μg/kg b.w./day) of the DEHP intakes exceeded the reference doses (RfD, 20 μg/kg b.w./day) of the US Environmental Protection Agency (EPA). DEHP was suggested to be the most notable phthalate from the perspective of adverse effects on the health of Japanese children. On the other hand, DcHP and di-n-hexyl phthalate were not considered to be very important as indoor air pollutants and as internal contaminants for the children. The contribution rates of the amounts absorbed by inhalation to the amounts absorbed via all of the exposure pathways were only 7.9, 4.4, 6.6, 3.2, 0.22, and 1.0% as the median for DMP, DEP, DiBP, DnBP, BBzP, and DEHP, respectively. Therefore, inhalation did not seem to contribute very much as an absorption pathway of the phthalates for Japanese children while at home. [ABSTRACT FROM AUTHOR]

Human norovirus (HuNoV) GII.P17-GII.17 (Kawasaki2014 variant) reportedly emerged in 2014 and caused gastroenteritis outbreaks worldwide. To clarify the evolution of both VP1 and RNA-dependent RNA polymerase (RdRp) regions of GII.P17-GII.17, we analyzed both global and novel Japanese strains detected during 2013–2017. Time-scaled phylogenetic trees revealed that the ancestral GII.17 VP1 region diverged around 1949, while the ancestral GII.P17 RdRp region diverged around 2010. The evolutionary rates of the VP1 and RdRp regions were estimated at ~2.7 × 10−3 and ~2.3 × 10−3 substitutions/site/year, respectively. The phylogenetic distances of the VP1 region exhibited no overlaps between intra-cluster and inter-cluster peaks in the GII.17 strains, whereas those of the RdRp region exhibited a unimodal distribution in the GII.P17 strains. Conformational epitope positions in the VP1 protein of the GII.P17-GII.17 strains were similar, although some substitutions, insertions and deletions had occurred. Strains belonging to the same cluster also harbored substitutions around the binding sites for the histo-blood group antigens of the VP1 protein. Moreover, some amino acid substitutions were estimated to be near the interface between monomers and the active site of the RdRp protein. These results suggest that the GII.P17-GII.17 virus has produced variants with the potential to alter viral antigenicity, host-binding capability, and replication property over the past 10 years. [ABSTRACT FROM AUTHOR]

Seven foodborne norovirus outbreaks attributable to the GII.P17-GII.17 strain were reported across Japan in 2017, causing illness in a total of 2,094 persons. Nori (dried shredded seaweed) was implicated in all outbreaks and tested positive for norovirus. Our data highlight the stability of norovirus in dehydrated food products. [ABSTRACT FROM AUTHOR]

In the 2016/2017 winter season in Japan, HuNoV GII.P16-GII.2 strains (2016 strains) emerged and caused large outbreaks of acute gastroenteritis. To better understand the outbreaks, we examined the molecular evolution of the VP1 gene and RdRp region in 2016 strains from patients by studying their time-scale evolutionary phylogeny, positive/negative selection, conformational epitopes, and phylodynamics. The time-scale phylogeny suggested that the common ancestors of the 2016 strains VP1 gene and RdRp region diverged in 2006 and 1999, respectively, and that the 2016 strain was the progeny of a pre-2016 GII.2. The evolutionary rates of the VP1 gene and RdRp region were around 10-3 substitutions/site/year. Amino acid substitutions (position 341) in an epitope in the P2 domain of 2016 strains were not found in pre-2016 GII.2 strains. Bayesian skyline plot analyses showed that the effective population size of the VP1 gene in GII.2 strains was almost constant for those 50 years, although the number of patients with NoV GII.2 increased in 2016. The 2016 strain may be involved in future outbreaks in Japan and elsewhere. [ABSTRACT FROM AUTHOR]

With the severe acute respiratory syndrome epidemic of 2003 and renewed attention on avian influenza viral pandemics, new surveillance systems are needed for the earlier detection of emerging infectious diseases. We applied a ''nextgeneration'' parallel sequencing platform for viral detection in nasopharyngeal and fecal samples collected during seasonal influenza virus (Flu) infections and norovirus outbreaks from 2005 to 2007 in Osaka, Japan. Random RT-PCR was performed to amplify RNA extracted from 0.1-0.25 ml of nasopharyngeal aspirates (N = 3) and fecal specimens (N = 5), and more than 10 μg of cDNA was synthesized. Unbiased high-throughput sequencing of these 8 samples yielded 15,298-32,335 (average 24,738) reads in a single 7.5 h run. In nasopharyngeal samples, although whole genome analysis was not available because the majority (>90%) of reads were host genome-derived, 20-460 Flu-reads were detected, which was sufficient for subtype identification. In fecal samples, bacteria and host cells were removed by centrifugation, resulting in gain of 484-15,260 reads of norovirus sequence (78-98% of the whole genome was covered), except for one specimen that was under-detectable by RT-PCR. These results suggest that our unbiased high-throughput sequencing approach is useful for directly detecting pathogenic viruses without advance genetic information. Although its cost and technological availability make it unlikely that this system will very soon be the diagnostic standard worldwide, this system could be useful for the earlier discovery of novel emerging viruses and bioterrorism, which are difficult to detect with conventional procedures. [ABSTRACT FROM AUTHOR]

Background: Human norovirus (HuNoV) is a major cause of enteric infectious gastroenteritis and is classified into several genotypes based on its capsid protein amino acid sequence and nucleotide sequence of the polymerase gene. Among these, GII.4 is the major genotype worldwide. Epidemiological studies have highlighted the prevalence of GII.2. Although recent advances using human tissue- and induced pluripotent stem cell (iPSC)-derived intestinal epithelial cells (IECs) have enabled in vitro replication of multiple HuNoV genotypes, GII.2 HuNoV could replicate only in tissue-derived IECs and not in iPSC-derived IECs., Methods: We investigated the factors influencing GII.2 HuNoV replication in IECs, focusing on histo-blood group antigens. We also assessed the immunogenicity of GII.2 virus-like particles (VLPs) and their ability to induce neutralizing antibodies. Antibody cross-reactivity was tested to determine whether GII.2 VLPs could neutralize other HuNoV genotypes, including GII.4, GII.3, GII.6, and GII.17., Results: Our findings indicated that GII.2 HuNoV replication in vitro requires the presence of both H and B antigens. Moreover, GII.2 VLPs generated neutralizing antibodies effective against both GII.2 and GII.4 but not against GII.3, GII.6, or GII.17. Comparatively, GII.2 and GII.17 VLPs induced broader neutralizing responses than GII.4 VLPs., Conclusions: The findings of this study suggests that GII.2 and GII.17 VLPs may be advantageous as HuNoV vaccine candidates because they elicit neutralizing antibodies against the predominant GII.4 genotype, which could be particularly beneficial for infants without prior HuNoV exposure. These insights will contribute to the development of effective HuNoV vaccines., Competing Interests: Potential conflicts of interest. Y. Y. and H. K. are the cofounders and directors of HanaVax, which provided research funding to S. S. Y. Y., H. K., and S. S. have filed a patent application related to the content of this article. All other authors report no potential conflicts., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Sapovirus (SaV) infections are a public health problem because they cause acute gastroenteritis in humans of all ages, both sporadically and as outbreaks. However, only a limited amount of SaV sequence information, especially whole-genome sequences for all the SaV genotypes, is publicly available. Therefore, in this study, we determined the full/near-full-length genomic sequences of 138 SaVs from the 2001 to 2015 seasons in 13 prefectures across Japan. The genogroup GI was predominant (67%, n = 92), followed by genogroups GII (18%, n = 25), GIV (9%, n = 12), and GV (6%, n = 9). Within the GI genogroup, four different genotypes were identified: GI.1 (n = 44), GI.2 (n = 40), GI.3 (n = 7), and GI.5 (n = 1). We then compared these Japanese SaV sequences with 3,119 publicly available human SaV sequences collected from 49 countries over the last 46 years. The results indicated that GI.1, and GI.2 have been the predominant genotypes in Japan, as well as in other countries, over at least four decades. The 138 newly determined Japanese SaV sequences together with the currently available SaV sequences, could facilitate a better understanding of the evolutionary patterns of SaV genotypes.

The number of person-to-person transmitted norovirus cases (n = 4,712) in school children in Osaka, Japan, during 2016/17 was the largest since 2012/13. Norovirus outbreaks were reported by 101 schools including 53 nursery schools (1,927 cases), 18 kindergartens (1,086 cases) and 30 elementary schools (1,699 cases). The dominant genotype among outbreaks was GII.P16-GII.2 (57.4%; 58/101), followed by GII.P2-GII.2 (8.9%; 9/101) and GII.P7-GII.6 (5.9%; 6/101). GII.4 was not detected despite dominance in previous years.

The RNA-dependent RNA polymerase ( RdRp ) and capsid ( VP1 ) genes of 51 GII.2 human norovirus (HuNoV) strains collected during the period of 2004-2015 in Japan were analyzed. Full-length analyses of the genes were performed using next-generation sequencing. Based on the gene sequences, we constructed the time-scale evolutionary trees by Bayesian Markov chain Monte Carlo methods. Time-scale phylogenies showed that the RdRp and VP1 genes evolved uniquely and independently. Four genotypes of GII.2 (major types: GII.P2-GII.2 and GII.P16-GII.2) were detected. A common ancestor of the GII.2 VP1 gene existed until about 1956. The evolutionary rates of the genes were high (over 10 -3 substitutions/site/year). Moreover, the VP1 gene evolution may depend on the RdRp gene. Based on these results, we hypothesized that transfer of the RdRp gene accelerated the VP1 gene evolution of HuNoV genotype GII.2. Consequently, recombination between ORF1 (polymerase) and ORF2 (capsid) might promote changes of GII.2 antigenicity.

A total of 1,141 rotavirus-negative stool specimens collected from diarrheic children in 4 distinct regions under sentinel surveillance in Thailand between 2006 and 2008 were examined by reverse-transcription (RT)-PCR for norovirus (NoV) and sapovirus (SaV). Three hundred 3 specimens (26.6%) were positive for NoV, with 34 and 269 belonging to genogroup I (GI) and genogroup II (GII), respectively. Twelve specimens (1.1%) were positive for SaV. Mixed infections were found in 5 specimens: 3 samples indicated the presence of both NoV GI and GII, and 2 samples indicated the presence of both NoV GII and SaV. Analysis of the monthly distribution of NoV and SaV revealed that NoV GII was clustered between September and February, while NoV GI was detected mainly in June and July; SaV was found in May, June, and July. In addition, 3 outbreaks of acute gastroenteritis at 2 junior high schools in Phichit and Bangkok, and at a university in Phitsanulok, Thailand in 2006 were found to have been caused by NoV infection. Sequence analysis of NoVs from sporadic cases and outbreaks showed them to be genotypes GII.4 and GII.6.

We have been characterizing monoclonal antibodies against Norovirus (Norwalk-like virus). In the course of our study, two monoclonal antibodies generated against Norovirus genogroup II capsid protein were found to react not only to genogroup II but also to genogroup I recombinant capsid proteins. In addition, we showed that these two monoclonal antibodies reacted to a 40-amino-acid-fragment located close to the N-terminal region of genogroup II Norovirus. Similar reactivity was observed with the equivalent region of genogroup I Norovirus. In this study, we confirmed that the epitopes of the two monoclonal antibodies existed within an 11-amino-acid peptide. To obtain an idea of the reactive ranges of the two monoclonal antibodies toward different strains of Norovirus, their reactivities were investigated using 16 types of peptide constructed according to the data in GenBank and 8 recombinant capsid proteins (7 whole capsid proteins and 1 short [80-amino-acid] protein fragment). A characteristic broad reactivity of the two monoclonal antibodies is clearly shown by the results of this study. Thus, these monoclonal antibodies could be useful tools for detecting a broad range of Norovirus strains.