[Norovirus nosocomial outbreak : an hospital experience]. / Epidémie nosocomiale à norovirus : à propos d'une expérience hospitalière.

Human noroviruses (NoV) are the main pathogenic agents worldwide responsible for viral sporadic and epidemic gastroenteritis worldwide. A gastroenteritis outbreak broke out in patients hospitalized in several wards located in two different floors of a hospital in Liege, Belgium. The objective was to determine whether a same NoV strain would be involved in the two different floors, and to explore how this outbreak would have spread from a floor to the other. Stool samples from patients and healthcare workers were collected, as well as data from medical files. NoV detection, quantification and characterization were performed using molecular biology methods. A same NoV strain, from genotype GII.4, was detected in two patients hospitalized on the two different floors. This finding allowed to conclude that a same outbreak spread in the two floors, probably due to movements of common healthcare workers. A rapid NoV detection during outbreak is important in the aim to rapidly implement hygiene measures to limit the size of the outbreak.

Les norovirus humains (NoV) sont reconnus mondialement comme les principaux agents étiologiques de gastro-entérites virales sporadiques et épidémiques au niveau mondial. Une épidémie de gastro-entérites s'est déclarée chez des patients hospitalisés dans plusieurs salles d'un hôpital de la région liégeoise, situées à deux étages différents. L'objectif était de déterminer si une même souche de NoV était impliquée aux deux étages, et d'investiguer la manière dont l'épidémie se serait propagée d'un étage à l'autre. Des prélèvements ont été collectés chez les patients et le personnel soignant. Les dossiers médicaux ont été examinés. La détection, la quantification et la caractérisation des souches de NoV ont été réalisées par des méthodes de biologie moléculaire. Une même souche de NoV, du génotype GII.4, a été mise en évidence chez deux patients hospitalisés aux deux étages différents. Ce résultat indique qu'il s'agit de la même épidémie qui s'est étendue à deux étages, probablement transmise par l'intermédiaire du personnel soignant commun. L'identification précoce des NoV lors des épidémies est primordiale afin de mettre en place rapidement les mesures d'hygiène permettant de limiter leur propagation.

[Noroviruses, these unknown worldwide epidemic enteropathogens]. / Les norovirus, ces entéro-pathogènes épidémiques mondiaux méconnus.

Discovered in the 1970s, human noroviruses (NoV) are the leading cause of foodborne disease and gastroenteritis outbreaks worldwide. NoV affect people of all ages. In children less than 5 years old, despite rotavirus remains the main enteropathogen responsible for viral gastroenteritis, NoV become the first etiological virus in countries where the rotavirus vaccine was introduced. Treatment of viral gastroenteritis is symptomatic. The key element in front of NoV infection is limiting their transmission. A rapid NoV detection during outbreak is important in the aim to rapidly implement hygiene measures to limit the size of the outbreak. Prevention of NoV infections relies on the use of adequate hand hygiene measures and disinfection of contaminated environmental surfaces. In face of an acute gastroenteritis outbreak, the early NoV identification with rapid laboratory tests or molecular biology methods is needed in the aim to implement as soon as possible hygiene measures to limit the size of the NoV outbreak. Due to antigenically diverse NoV strains and the lack of long term immunity, the development of an effective vaccine is difficult.

Découverts dans les années 1970, les norovirus humains (NoV) sont reconnus comme les principaux agents pathogènes responsables de toxi-infections d'origine alimentaire et d'épidémies de gastro-entérites au niveau mondial. Ils infectent toutes les tranches d'âge. Chez les enfants de moins de 5 ans, bien que le rotavirus reste actuellement la première cause de gastro-entérites virales, force est de constater que les NoV sont en passe d'en devenir la première cause dans les pays où la vaccination contre le rotavirus a été introduite. Le traitement des gastro-entérites virales est symptomatique. L'élément clé face aux infections à NoV est de limiter leur transmission. La prévention des infections à NoV repose principalement sur l'application de mesures d'hygiène des mains adéquates et la désinfection de l'environnement contaminé. Lors des épidémies de gastro-entérites aiguës, l'identification précoce des NoV par des méthodes de laboratoire rapides ou de biologie moléculaire est primordiale afin de mettre en place rapidement les mesures d'hygiène permettant de limiter leur propagation. La diversité antigénique des NoV et le manque d'immunité protectrice à long terme rendent la mise au point de vaccins difficile.

Global computation of phase-amplitude reduction for limit-cycle dynamics.

Recent years have witnessed increasing interest in phase-amplitude reduction of limit-cycle dynamics. Adding an amplitude coordinate to the phase coordinate allows us to take into account the dynamics transversal to the limit cycle and thereby overcome the main limitations of classic phase reduction (strong convergence to the limit cycle and weak inputs). While previous studies, mostly focus on local quantities such as infinitesimal responses, a major and limiting challenge of phase-amplitude reduction is to compute amplitude coordinates globally, in the basin of attraction of the limit cycle. In this paper, we propose a method to compute the full set of phase-amplitude coordinates in the large. Our method is based on the so-called Koopman (composition) operator and aims at computing the eigenfunctions of the operator through Laplace averages (in combination with the harmonic balance method). This yields a forward integration method that is not limited to two-dimensional systems. We illustrate the method by computing the so-called isostables of limit cycles in two-, three-, and four-dimensional state spaces, as well as their responses to strong external inputs.

Hepatitis E Virus and Related Viruses in Animals.

Hepatitis E is an acute human liver disease in healthy individuals which may eventually become chronic. It is caused by the hepatitis E virus (HEV) and can have a zoonotic origin. Nearly 57,000 people die yearly from hepatitis E-related conditions. The disease is endemic in both developing and developed countries with distinct epidemiologic profiles. In developing countries, the disease is associated with inadequate water treatment, while in developed countries, transmission is associated with animal contact and the ingestion of raw or uncooked meat, especially liver. All human HEV are grouped into at least four genotypes, while HEV or HEV-related viruses have been identified in an increasing number of domestic and wild animal species. Despite a high genetic diversity, only one single HEV serotype has been described to date for HEV genotypes 1-4. The discovery of new HEV or HEV-related viruses leads to a continuing increase in the number of genotypes. In addition, the genome organization of all these viruses is variable with overlapping open reading frames (ORF) and differences in the location of ORF3. In spite of the role of some domestic and wild animals as reservoir, the origin of HEV and HEV-related viruses in humans and animals is still unclear. This review discusses aspects of the detection, molecular virology, zoonotic transmission and origin of HEV and HEV-related viruses in the context of 'One Health' and establishes a link between the previous and the new taxonomy of this growing virus family.

Belgian Wildlife as Potential Zoonotic Reservoir of Hepatitis E Virus.

Hepatitis E is an acute human liver disease in healthy individuals but may become chronic in immunocompromised patients. It is caused by the hepatitis E virus (HEV) and can have a zoonotic origin, particularly in high-income countries. In this study, 383 sera from wild boars were selected for serology; for virological analyses, 69 sera and 61 livers from young wild boars were used. A total of 189 and 235 sera of, respectively, red deer and roe deer were collected for serological analysis. For virological analyses, 84 and 68 sera and 29 and 27 livers from, respectively, red and roe deer were sampled. An apparent seroprevalence of 34% (95% CI 29.71-39.46) was found in wild boars, of 1% (95% CI 0-2.4) in red deer and 3% (95% CI 0.8-4.2) in roe deer. To assess the ELISA screening prevalence, Western blot (WB) analyses were carried out, a receiver operating characteristic curve analysis was performed and different scenarios with varying ELISA specificities relative to WB were analysed. Seroprevalence remained high whatever the scenario in the wild boar population. In wild boar, 4 of 69 sera and 4 of 61 livers were detected as positive for HEV RNA. All sequences obtained from sera belonged to genotype HEV-3. HEV RNA, belonging to genotype HEV-3, was detected in one of 29 red deer livers. Wild boar can be considered as a host reservoir of the virus in Belgium. However, in contrast to the epidemiological role played by them in other countries, the low prevalence in deer makes these species an unlikely reservoir. This evidence needs further investigation to determine in which situation deer can serve as reservoir. These results also raise the question of the dynamics of HEV infection between wild fauna, domestic pigs and humans.

Susceptibility of Pigs to Zoonotic Hepatitis E Virus Genotype 3 Isolated from a Wild Boar.

In Europe, zoonotic hepatitis E virus (HEV) genotype 3 strains mainly circulate in humans, swine and wild boar. The aim of this study was to investigate the potential transmission of a wild boar originating HEV strain (WbHEV) to swine by intravenous or oral inoculation and to study the consequences of infection of a WbHEV strain, a WbHEV strain previously passaged in a pig and a swine HEV strain after oral inoculation. Firstly, an intravenous infection was performed for which five piglets were divided into two groups with three pigs inoculated with a WbHEV field strain and two pigs inoculated with a HEV-negative swine liver homogenate. All pigs were necropsied 8, 9 and 10 days post-inoculation. Secondly, an oral infection of 56 days was performed on 12 piglets divided into four groups inoculated with a WbHEV strain, a WbHEV strain previously passaged in swine, a swine HEV strain or a HEV-negative swine liver homogenate. After intravenous inoculation, HEV RNA was detected in serum, bile, liver, spleen, duodenum, jejunum, colon, lung, gastro-hepatic lymph nodes and faeces in all infected piglets. After oral inoculation, HEV RNA was detected in serum, bile, liver, gastro-hepatic lymph nodes and faeces. Most of HEV-inoculated pigs became seropositive at day 15. This study provides experimental evidence of early viral spread throughout the organism after intravenous infection with a WbHEV strain and supports the notion that such a zoonotic strain could be transmitted via the natural faecal-oral route of infection between wild boar and pigs but also between pigs.

Cattle are a potential reservoir of bubaline herpesvirus 1 (BuHV1).

In the present work, controlled experimental infection and transmission studies in domestic cattle (Bos taurus) and water buffaloes (Bubalus bubalis) were carried out to study the in vivo behaviour of bubaline herpesvirus 1 (BuHV1). Two bovine and two buffalo calves were infected with BuHV1 (20287N isolate) by intranasal aerosolisation. Two sentinel cattle did not receive the virus challenge, but were housed with infected buffaloes to evaluate horizontal transmission. All experimentally inoculated animals showed viral infection and respiratory clinical signs. BuHV1 experimentally infected calves showed intermittent viral excretion between 2 days and 18 days postinfection (dpi) with a maximum titre of excretion of 106 TCID50/ml and moderate rhinitis between 2 dpi and 20 dpi. BuHV1 experimentally inoculated buffaloes showed mild respiratory signs, which consisted mainly of serous nasal secretions during the infection period. Sentinel calves showed mucosal specific IgG1 antibodies at seven days postcontact. Viral DNA was detected by PCR and sequencing in both buffaloes and sentinel calves, which could be associated with latency. In conclusion, this study showed the susceptibility of cattle to BuHV1 after both experimental infection and contact with infected buffaloes. These data increase the scarce knowledge on the pathogenesis in natural host and the susceptibility of cattle to BuHV1 experimental infection.

Genetic and evolutionary perspectives on genogroup III, genotype 2 bovine noroviruses.

Bovine noroviruses are enteric pathogens that are detected in stool samples from cattle. Five genogroups are currently described in the genus Norovirus (family Caliciviridae), and within the genogroups, sequences are further divided into genotypes according to genetic homology and phylogenetic relationships. In this study, stool specimens from Belgian cattle were screened by RT-PCR. All of the sequences that were detected were phylogenetically related to genogroup III genotype 2 bovine noroviruses, confirming their higher prevalence in comparison with strains from genotype 1. When other sequences from around the world were introduced, phylogenetic inferences allowed neither the determination of phylogenetic lineages over time nor the deduction of topotypes for genotype 2 bovine noroviruses. Three complete genotype 2 bovine norovirus sequences were also compared genetically (Newbury2/1976 /UK, Dumfries/1994/UK and B309/2003/BE). Interestingly, the genetic divergence of the complete genomes of these three strains was relatively low, but a region of the N-terminal protein encoded by ORF1, the hypervariable region of the capsid gene encoded by ORF2, and a region of the minor structural protein encoded by ORF3 seem to be the most exposed to genetic evolution. Bayesian inference also showed that genetic evolution of genogroup III, genotype 2 bovine noroviruses over a 30-year period seemed to be lower than that already reported for noroviruses from the genotypes 3 and 4 in genogroup II.

Clinical pattern characterization of cattle naturally infected by BTV-8.

Forty-one cattle from seven Belgian farms and two French farms confirmed as infected with bluetongue virus serotype 8 (BTV-8) were monitored from the onset of clinical signs to describe the disease pattern and estimate the duration of blood RT-qPCR and competitiveELISA positivity under field conditions. On each visit, blood samples were taken, and a standardized clinical form was filled in for each animal. A clinical score was calculated for every week until the end of clinical signs. A classification and regression tree (CART) analysis was conducted to determine the most important clinical signs every week for the first 7 weeks. The highest scores were recorded within 2 weeks of clinical onset. The first recorded clinical signs were quite obviously visible (lethargy, conjunctivitis, lesions of nasal mucosa, nasal discharge). Skin lesions, a drop in milk production and weight loss appeared later in the course of the disease. A biphasic pattern regarding nasal lesions was noticed: the first peak concerned mainly congestive and ulcerative lesions, whereas the second peak mainly concerned crusty lesions. The median time estimated by survival analysis to obtain negative RT-qPCR results from the onset of clinical signs was 195 days (range 166-213 days) in the 23 cattle included in the analysis. Serological results remained strongly positive until the end of the study. These results should ensure more accurate detection of an emerging infectious disease and are of prime importance in improving the modelling of BTV-8 persistence in Europe.

Complete genome sequence of a novel bovine norovirus: evidence for slow genetic evolution in genogroup III genotype 2 noroviruses.

A new genogroup III genotype 2 bovine norovirus, B309/2003/BE, was entirely sequenced and genetically compared to the original Newbury2/1976/UK strain and to Dumfries/1994/UK, detected in 1976 and 1994, respectively. Interestingly, except in well-defined coding regions (N-terminal protein, 3A-like protease, hypervariable region of the capsid protein, and C-terminal part of the minor structural protein), very low genetic differences were noted between the entire genomes of these three strains along a 30-year-long period. It allowed some hypotheses of hotspots of genetic evolution through a low genetic evolution background in genotype 2 genogroup III bovine noroviruses.

On the use of Fourier averages to compute the global isochrons of (quasi)periodic dynamics.

The concept of isochrons is crucial for the analysis of asymptotically periodic systems. Roughly, isochrons are sets of points that partition the basin of attraction of a limit cycle according to the asymptotic behavior of the trajectories. The computation of global isochrons (in the whole basin of attraction) is however difficult, and the existing methods are inefficient in high-dimensional spaces. In this context, we present a novel (forward integration) algorithm for computing the global isochrons of high-dimensional dynamics, which is based on the notion of Fourier time averages evaluated along the trajectories. Such Fourier averages in fact produce eigenfunctions of the Koopman semigroup associated with the system, and isochrons are obtained as level sets of those eigenfunctions. The method is supported by theoretical results and validated by several examples of increasing complexity, including the 4-dimensional Hodgkin-Huxley model. In addition, the framework is naturally extended to the study of quasiperiodic systems and motivates the definition of generalized isochrons of the torus. This situation is illustrated in the case of two coupled Van der Pol oscillators.

Animal noroviruses.

Among enteric caliciviruses, noroviruses belong to the genus Norovirus, one of the four accepted genera in the family Caliciviridae. These single-stranded, positive-sense RNA viruses are highly variable both genetically and antigenically. Several animal enteric caliciviruses that are morphologically indistinguishable and genetically closely related to human noroviruses have been identified. The first bovine enteric noroviruses were described in Great Britain and are known as Newbury Agent 2. At least three genetic clusters of porcine noroviruses join together within genogroup II noroviruses. Human noroviruses are the most important cause of acute gastroenteritis illness in people of all ages. In the USA, they are associated with approximately 30-50% of all food-borne outbreaks. Until now, noroviruses have not been associated with gastroenteritis outbreaks in immunocompetent animals. Neither bovine nor porcine noroviruses can replicate in cell culture, although human norovirus can grow in a complex 3D culture system. However, the recently discovered murine noroviruses can replicate in cell culture and are therefore used as model viruses to study human noroviruses. This review focusses on virus classification, virion structure, pathogenesis, epidemiology, immune response and diagnosis of animal noroviruses in comparison with human noroviruses. The classification of animal enteric caliciviruses within the Norovirus genus raises the question of whether transmission from an animal reservoir to humans could occur. Answering this question is important in determining the risk of cross-species infections affecting the epidemiology and evolution of these viruses and so complicating the control of human norovirus infections.