EHEC O111:H8 strain and norovirus GII.4 Sydney [P16] causing an outbreak in a daycare center, Brazil, 2019.

BACKGROUND: This study describes the investigation of an outbreak of diarrhea, hemorrhagic colitis (HC), and hemolytic uremic syndrome (HUS) at a daycare center in southeastern Brazil, involving fourteen children, six staff members, six family members, and one nurse. All bacterial and viral pathogens detected were genetically characterized. RESULTS: Two isolates of a strain of enterohemorrhagic Escherichia coli (EHEC) serotype O111:H8 were recovered, one implicated in a case of HUS and the other in a case of uncomplicated diarrhea. These isolates had a clonal relationship of 94% and carried the stx2a and eae virulence genes and the OI-122 pathogenicity island. The EHEC strain was determined to be a single-locus variant of sequence type (ST) 327. EHEC isolates were resistant to ofloxacin, doxycycline, tetracycline, ampicillin, and trimethoprim-sulfamethoxazole and intermediately resistant to levofloxacin and ciprofloxacin. Rotavirus was not detected in any samples, and norovirus was detected in 46.7% (14/30) of the stool samples, three of which were from asymptomatic staff members. The noroviruses were classified as the recombinant GII.4 Sydney [P16] by gene sequencing. CONCLUSION: In this outbreak, it was possible to identify an uncommon stx2a + EHEC O111:H8 strain, and the most recent pandemic norovirus strain GII.4 Sydney [P16]. Our findings reinforce the need for surveillance and diagnosis of multiple enteric pathogens by public health authorities, especially during outbreaks.

The Role of Host Glycobiology and Gut Microbiota in Rotavirus and Norovirus Infection, an Update.

Rotavirus (RV) and norovirus (NoV) are the leading causes of acute gastroenteritis (AGE) worldwide. Several studies have demonstrated that histo-blood group antigens (HBGAs) have a role in NoV and RV infections since their presence on the gut epithelial surfaces is essential for the susceptibility to many NoV and RV genotypes. Polymorphisms in genes that code for enzymes required for HBGAs synthesis lead to secretor or non-secretor and Lewis positive or Lewis negative individuals. While secretor individuals appear to be more susceptible to RV infections, regarding NoVs infections, there are too many discrepancies that prevent the ability to draw conclusions. A second factor that influences enteric viral infections is the gut microbiota of the host. In vitro and animal studies have determined that the gut microbiota limits, but in some cases enhances enteric viral infection. The ways that microbiota can enhance NoV or RV infection include virion stabilization and promotion of virus attachment to host cells, whereas experiments with microbiota-depleted and germ-free animals point to immunoregulation as the mechanism by which the microbiota restrict infection. Human trials with live, attenuated RV vaccines and analysis of the microbiota in responder and non-responder individuals also allowed the identification of bacterial taxa linked to vaccine efficacy. As more information is gained on the complex relationships that are established between the host (glycobiology and immune system), the gut microbiota and intestinal viruses, new avenues will open for the development of novel anti-NoV and anti-RV therapies.

Contrasting Results from Two Commercial Kits Testing for the Presence of Clostridium perfringens Enterotoxin in Feces from Norovirus-Infected Human Patients.

BACKGROUND: Detection of Clostridium perfringens enterotoxin (CPE) is critical for disease surveillance; however, commercial testing kits produce contrasting results. METHODS: We examined the cause of the differing results from a reversed passive latex agglutination (RPLA) assay (PET-RPLA Toxin Detection Kit) and an enzyme-linked immunosorbent assay (C. perfringens Enterotoxin ELISA Kit) using 73 human norovirus-positive fecal samples from gastroenteritis patients across 22 episodes in Japan. RESULTS: CPE was detected in 39/73 samples using the RPLA method; however, ELISA-based examination of 10 RPLA-positive samples produced negative results. Moreover, cpe was not detected in any of the RPLA-positive (n = 32) or -negative (n = 5) samples, and C. perfringens was only isolated from one RPLA-positive sample. CONCLUSIONS: An ELISA-based testing approach may be more reliable than RPLA assays for CPE detection from human fecal samples. These findings may also be applicable to the detection of other foodborne diseases.

HOIL1 Is Essential for the Induction of Type I and III Interferons by MDA5 and Regulates Persistent Murine Norovirus Infection.

The linear ubiquitin chain assembly complex (LUBAC), composed of heme-oxidized IRP2 ubiquitin ligase 1 (HOIL1), HOIL1-interacting protein (HOIP), and SHANK-associated RH domain-interacting protein (SHARPIN), is a crucial regulator of multiple immune signaling pathways. In humans, HOIL1 or HOIP deficiency is associated with an immune disorder involving autoinflammation, immunodeficiency, and inflammatory bowel disease (IBD)-like symptoms. During viral infection, LUBAC is reported to inhibit the induction of interferon (IFN) by the cytosolic RNA sensor retinoic acid-inducible gene I (RIG-I). Surprisingly, we found that HOIL1 is essential for the induction of both type I and type III IFNs, as well as the phosphorylation of IFN regulatory factor 3 (IRF3), during murine norovirus (MNoV) infection in cultured dendritic cells. The RIG-I-like receptor, melanoma differentiation-associated protein 5 (MDA5), is also required for IFN induction and IRF3 phosphorylation during MNoV infection. Furthermore, HOIL1 and MDA5 were required for IFN induction after Theiler's murine encephalomyelitis virus infection and poly(I·C) transfection, but not Sendai virus or vesicular stomatitis virus infection, indicating that HOIL1 and LUBAC are required selectively for MDA5 signaling. Moreover, Hoil1-/- mice exhibited defective control of acute and persistent murine norovirus infection and defective regulation of MNoV persistence by the microbiome as also observed previously for mice deficient in interferon lambda (IFN-λ) receptor, signal transducer and activator of transcription factor 1 (STAT1), and IRF3. These data indicate that LUBAC plays a critical role in IFN induction to control RNA viruses sensed by MDA5.IMPORTANCE Human noroviruses are a leading cause of gastroenteritis throughout the world but are challenging to study in vivo and in vitro Murine norovirus (MNoV) provides a tractable genetic and small-animal model to study norovirus biology and immune responses. Interferons are critical mediators of antiviral immunity, but excessive expression can dysregulate the immune system. IFN-λ plays an important role at mucosal surfaces, including the gastrointestinal tract, and both IFN-λ and commensal enteric bacteria are important modulators of persistent MNoV infection. LUBAC, of which HOIL1 is a component, is reported to inhibit type I IFN induction after RIG-I stimulation. We show, in contrast, that HOIL1 is critical for type I and III IFN induction during infection with MNoV, a virus that preferentially activates MDA5. Moreover, HOIL1 regulates MNoV infection in vivo These data reveal distinct functions for LUBAC in these closely related signaling pathways and in modulation of IFN expression.

Waterborne outbreak of gastroenteritis on the KwaZulu-Natal Coast, South Africa, December 2016/January 2017.

An unexpected increase in gastroenteritis cases was reported by healthcare workers on the KwaZulu-Natal Coast, South Africa, January 2017 with >600 cases seen over a 3-week period. A case-control study was conducted to identify the source and risk factors associated with the outbreak so as to recommend control and prevention measures. Record review identified cases and controls and structured-telephonic interviews were conducted to obtain exposure history. Stool specimens were collected from 20 cases along with environmental samples and both screened for enteric pathogens. A total of 126 cases and 62 controls were included in the analysis. The odds of developing gastroenteritis were 6.0 times greater among holiday makers than residents (95% confidence interval (CI) 2.0-17.7). Swimming in the lagoon increased the odds of developing gastroenteritis by 3.3 times (95% CI 1.06-10.38). Lagoon water samples tested positive for norovirus (NoV) GI.6, GII.3 and GII.6, astrovirus and rotavirus. Eleven (55%) stool specimens were positive for NoV with eight genotyped as GI.1 (n = 2), GI.5 (n = 3), GI.6 (n = 2), and GI.7 (n = 1). A reported sewage contamination event impacting the lagoon was the likely source with person-to-person spread perpetuating the outbreak. Restriction to swimming in the lagoon was apparently ineffective at preventing the outbreak, possibly due to inadequate enforcement, communication and signage strategies.

Genetic and epidemiological analysis of norovirus from children with gastroenteritis in Botswana, 2013-2015.

BACKGROUND: Norovirus is a leading cause of viral gastroenteritis worldwide with a peak of disease seen in children. The epidemiological analysis regarding the virus strains in Africa is limited. The first report of norovirus in Botswana was in 2010 and currently, the prevalence and circulating genotypes of norovirus are unknown, as the country has no systems to report the norovirus cases. This study investigated the prevalence, patterns and molecular characteristics of norovirus infections among children ≤5 years of age admitted with acute gastroenteritis at four hospitals in Botswana. METHODS: A total of 484 faecal samples were collected from children who were admitted with acute gastroenteritis during the rotavirus vaccine impact survey between July 2013 and December 2015. Norovirus was detected using real-time RT-PCR. Positive samples were genotyped using conventional RT-PCR followed by partial sequencing of the capsid and RdRp genes. Norovirus strains were determined by nucleotide sequence analysis using the online Norovirus Genotyping Tool Version 1.0, and confirmed using maximum likelihood tree construction as implemented in MEGA 6.0. RESULTS: The prevalence of norovirus was 9.3% (95% CI 6.7-11.9). The genotype diversity was dominated by the GII.4 strain at 69.7%. This was followed by GII.2, GII.12 each at 9.1%, GI.9 at 6.6% and GII.6, GII.10 each at 3.0%. The most common combined RdRp/Capsid genotype was the GII.Pe/GII.4 Sydney 2012. Norovirus was detected during most part of the year; however, there was a preponderance of cases in the wet season (December to March). CONCLUSION: The study showed a possible decline of norovirus infections in the last 10 years since the first report. An upward trend seen between 2013 and 2015 may be attributable to the success of rotavirus vaccine introductions in 2012. Knowledge of circulating genotypes, seasonal trends and overall prevalence is critical for prevention programming and possible future vaccine design implications.

[Norovirus infection surveillance in early warning of outbreaks in population in oyster farming areas].

OBJECTIVE: To have an insight into the prevalence tendency caused by Noroviruses( NoVs) in population of oyster farming areas, and evaluate the early warning signification of acute gastroenteritis( AGE) monitoring. METHODS: Between January and December in 2014, a retrospective household survey of AGE during past four weeks was conducted to residents using multi-stage sampling in oyster farming areas. Norovirus capsid genome was detected using real-time RT-PCR and semi-nested PCR, sequenced and phylogenetic analyzed. RESULTS: A total of 75 cases of AGE were reported. The incidence rate was 0. 10 per person-year, the detection rate of norovirus was 20. 0%( 15 /75). The dominant epidemic No Vs strain in oyster farming was still GII. 4 Sydney 2012. Furthermore, a newly GII. 17 variant was detected and early in March 2014. CONCLUSION: The GII. 4 Sydney 2012 genotype is still prevalent strain that caused AGE currently in the community residents of oyster farming areas. NoVs GII. 17 variant was found in sporadic patients of sentinel hospital AGE monitoring in March 2014 that have an early warning effect to the outbreaks of winter in 2014.

Recent advances in understanding norovirus pathogenesis.

Noroviruses constitute a family of ubiquitous and highly efficient human pathogens. In spite of decades of dedicated research, human noroviruses remain a major cause of gastroenteritis and severe diarrheal disease around the world. Recent findings have begun to unravel the complex mechanisms that regulate norovirus pathogenesis and persistent infection, including the important interplay between the virus, the host immune system, and commensal bacteria. Herein, we will summarize recent research developments regarding norovirus cell tropism, the use of M cells, and commensal bacteria to facilitate norovirus infection, and virus, host, and bacterial determinants of persistent norovirus infections. J. Med. Virol. 88:1837-1843, 2016. © 2016 Wiley Periodicals, Inc.

Outbreak of acute gastroenteritis caused by contamination of drinking water in a factory, the Basque Country.

On 18 September 2013, the Gipuzkoa Epidemiology Unit was notified of an outbreak of acute gastroenteritis (AGE) among employees at a domestic appliance factory. The first signs of the outbreak had emerged at the end of June and at the time of the notification 30 workers were on sick leave for gastroenteritis. Some employees had had more than one episode and the main symptoms were diarrhoea and vomiting. An investigation began to identify the causative agent, assess exposure and determine the route of transmission. Data collected by a questionnaire identified 302 episodes of AGE among 238 people affected between June and September 2013. The source of water consumed was found to be a risk factor associated with the appearance of symptoms both in the crude and the adjusted analysis: odds ratio 1.8 (0.8-4.2) and 6.4 (4.2-9.8), respectively. Microbiological analysis of stool samples and of water confirmed the presence of norovirus and rotavirus. The environmental study detected a connection between an industrial use water system and drinking water at the factory. It was concluded that the outbreak was caused by mixed viral infections, due to contamination of drinking water.

Investigation of gut microbiota disorders in norovirus infected children patients based on 16s rRNA sequencing.

BACKGROUND: Norovirus is the leading cause of sporadic viral gastroenteritis cases and outbreaks. Gut microbiota plays a key role in maintaining immune homeostasis. We aimed to investigate the composition and functional effects of gut microbiota in children infected with norovirus. METHODS: Stool samples were collected from 31 children infected with norovirus and 25 healthy children. The gut microbiota was analyzed by 16S rRNA gene sequencing, followed by composition, correlation network, functional and phenotype prediction analyses. RESULTS: Gut microbiota in children infected with norovirus was characterized by lower species richness and diversity. Veillonella is the dominant gut microbiota specie in norovirus infection. Blautia was significantly lower in norovirus infection. There was a positive correlation between Faecalibacterium, Blautia, Subdoligranulum, Eubacterium_hallii_group, Fusicatenibacter, Agathobacter, Roseburia and Dorea. Functionally, secondary metabolites biosynthesis, transport and catabolism, selenocysteine lyase and peroxiredoxin were the most significantly higher functional compositions of gut microbiota in norovirus infection. However, sn-glycerol-1-phosphate dehydrogenase and fermentation were the most significantly lower functional compositions in norovirus infection group. Phenotype analysis showed that Contains_Mobile_Elements had the highest level of phenotypes in the gut microbiota of norovirus infection. CONCLUSION: Norovirus infection may lead to dysregulation of the gut microbiome in children.

[A mechanism of norovirus pandemic based on comprehensive genome analysis].

Norovirus GII.4 is a major etiological agent of acute viral gastroenteritis worldwide. We examined GII.4 evolution using 277 near-full-length GII.4 genome sequences from human stool specimens collected at 20 sites in Japan between May 2006 and March 2010. We found outbreaks of 8 monophyletic GII.4 subtypes, among which a single subtype, termed 2006b, had continually predominated (222/277: 80.7%). Four of the 8 GII.4 subtypes were chimera viruses of recently prevalent GII.4 subtypes. Notably, single putative recombination breakpoints with the highest statistical significance were constantly located around the border of open reading frame 1 (ORF) 1 and ORF 2 (P<0.0001), suggesting outgrowth of specific recombinant viruses in the outbreaks. The GII.4 subtypes had many unique amino acids at the time of their outbreaks, especially in the N-term, 3A-like, and capsid proteins. Unique amino acids in the capsid were preferentially positioned on the outer surface loops of the protruding P2 domain. These data and computer-assisted structural study of NoV capsid protein are compatible with a model of antigenic drift with tuning of the structure-functions of multiple proteins for the survival strategy of GII.4 2006b variant.

[Infection prevention and control for foodborne infections].

Patients' care for foodborne infections is sometimes very critical, since these patients exerting high copy numbers of contagious pathogens. Recently, Norovirus infection became the most frequent pathogen for large outbreaks in the community and the hospital around the world. Norovirus is alcohol-resistant and highly contagious. For preventing outbreaks of foodborne infections, standard precaution(and contact precaution for diaper changing patients) is required by the CDC's isolation precaution guideline revised at 2007. We need to provide for infection prevention and control in the epidemic winter period not only in healthcare facilities but also for communities.

Norovirus diarrhea is significantly associated with higher counts of fecal histo-blood group antigen expressing Enterobacter cloacae among black South African infants.

The study tested the hypothesis that harboring high levels of histo-blood group antigen-expressing Enerobactero cloacae is a risk factor for norovirus diarrhea. The fecal E. cloacae abundance in diarrheic norovirus positive (DNP), non-diarrheic norovirus negative (NDNN), diarrhea norovirus negative (DNN), and non-diarrhea norovirus positive (NDNP) infants was determined by qPCR, and the risk of norovirus diarrhea was assessed by logistical regression. DNP infants contained significantly higher counts of E. cloacae than NDNN and DNN infants, p = .0294, and 0.0001, respectively. The risk of norovirus diarrhea was significantly high in infants with higher counts of E. cloacae than those with lower counts, p = .009. Harboring higher counts of E. cloacae is a risk factor for norovirus diarrhea.

Norovirus: Facts and Reflections from Past, Present, and Future.

Human Norovirus is currently the main viral cause of acute gastroenteritis (AGEs) in most countries worldwide. Nearly 50 years after the discovery of the "Norwalk virus" by Kapikian and colleagues, the scientific and medical community continue to generate new knowledge on the full biological and disease spectrum of Norovirus infection. Nevertheless, several areas remain incompletely understood due to the serious constraints to effectively replicate and propagate the virus. Here, we present a narrated historic perspective and summarize our current knowledge, including insights and reflections on current points of interest for a broad medical community, including clinical and molecular epidemiology, viral-host-microbiota interactions, antivirals, and vaccine prototypes. We also include a reflection on the present and future impacts of the COVID-19 pandemic on Norovirus infection and disease.

Tracking environmental norovirus contamination in a pediatric primary immunodeficiency unit.

Norovirus strains were detected in two patients and in environmental swabs from a pediatric primary immunodeficiency unit in London, United Kingdom, during an infection control incident in November and December 2007. Detailed analyses of the gene encoding the P2 domain demonstrated that the majority of the strains were not related to the patients and that the environmental contamination was most likely due to secondary transfer by the hands of staff or visitors.

Norovirus outbreaks linked to oyster consumption in the United Kingdom, Norway, France, Sweden and Denmark, 2010.

This paper reports on several simultaneous outbreaks of norovirus infection linked to the consumption of raw oysters. Since January 2010, 334 cases in 65 clusters were reported from five European countries: the United Kingdom, Norway, France, Sweden and Denmark. The article describes the available epidemiological and microbiological evidence of these outbreaks.

The scenario of norovirus contamination in food and food handlers.

Recently, many cases related to viral gastroenteritis outbreaks have been reported all over the world. Noroviruses are found to be leading as the major cause of outbreaks of acute gastroenteritis. Patients with the acute gastroenteritis normally found to be positive with norovirus when stools and vomit were analyzed. This paper reviews various activities and previous reports that describe norovirus contaminated in various food matrixes and relationship between food handlers. Lately, a numbers of norovirus outbreaks have been reported which are involved fresh produce (such as vegetables, fruits), shellfish and prepared food. Food produces by infected food handlers may therefore easily contaminated. In addition, food that required much handling and have been eaten without heat treatment gave the high risk for getting foodborne illnesses. The standard method for detection of norovirus has already been available for stool samples. However, only few methods for detection of norovirus in food samples have been developed until now.

The intestinal regionalization of acute norovirus infection is regulated by the microbiota via bile acid-mediated priming of type III interferon.

Evidence has accumulated to demonstrate that the intestinal microbiota enhances mammalian enteric virus infections1. For example, we and others previously reported that commensal bacteria stimulate acute and persistent murine norovirus infections2-4. However, in apparent contradiction of these results, the virulence of murine norovirus infection was unaffected by antibiotic treatment. This prompted us to perform a detailed investigation of murine norovirus infection in microbially deplete mice, revealing a more complex picture in which commensal bacteria inhibit viral infection of the proximal small intestine while simultaneously stimulating the infection of distal regions of the gut. Thus, commensal bacteria can regulate viral regionalization along the intestinal tract. We further show that the mechanism underlying bacteria-dependent inhibition of norovirus infection in the proximal gut involves bile acid priming of type III interferon. Finally, the regional effects of the microbiota on norovirus infection may result from distinct regional expression profiles of key bile acid receptors that regulate the type III interferon response. Overall, these findings reveal that the biotransformation of host metabolites by the intestinal microbiota directly and regionally impacts infection by a pathogenic enteric virus.

Attach Me If You Can: Murine Norovirus Binds to Commensal Bacteria and Fungi.

The presence of commensal bacteria enhances both acute and persistent infection of murine noroviruses. For several enteric viral pathogens, mechanisms by which these bacteria enhance infection involve direct interactions between the virus and bacteria. While it has been demonstrated that human noroviruses bind to a variety of commensal bacteria, it is not known if this is also true for murine noroviruses. The goal of this study was to characterize interactions between murine noroviruses and commensal bacteria and determine the impact of bacterial growth conditions, incubation temperature and time, on murine norovirus attachment to microbes that comprise the mammalian microbiome. We show that murine noroviruses bind directly to commensal bacteria and show similar patterns of attachment as human norovirus VLPs examined under the same conditions. Furthermore, while binding levels are not impacted by the growth phase of the bacteria, they do change with time and incubation temperature. We also found that murine norovirus can bind to a commensal fungal species, Candidaalbicans.