The immune system is known to be involved in the early phase of scrapie pathogenesis. However, the infection route of naturally occurring scrapie and its spread within the host are not entirely known. In this study, the pathogenesis of scrapie was investigated in sheep of three PrP genotypes, from 2 to 9 months of age, which were born and raised together in a naturally scrapie-affected Romanov flock. The kinetics of PrP Sc accumulation in sheep organs were determined by immunohistochemistry. PrP Sc was detected only in susceptible VRQ/VRQ sheep, from 2 months of age, with an apparent entry site at the ileal Peyer's patch as well as its draining mesenteric lymph node. At the cellular level, PrP Sc deposits were associated with CD68-positive cells of the dome area and B follicles before being detected in follicular dendritic cells. In 3-to 6-month-old sheep, PrP Sc was detected in most of the gut-associated lymphoid tissues (GALT) and to a lesser extent in more systemic lymphoid formations such as the spleen or the mediastinal lymph node. All secondary lymphoid organs showed a similar intensity of PrP Sc -immunolabelling at 9 months of age. At this time-point, PrPSc was also detected in the autonomic myenteric nervous plexus and in the nucleus parasympathicus nervi X of the brain stem. These data suggest that natural scrapie infection occurs by the oral route via infection of the Peyer's patches followed by replication in the GALT. It may then spread to the central nervous system through the autonomic nervous fibres innervating the digestive tract.

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Most viruses express one or several proteins that counter the antiviral defences of the host cell. This is the task of non-structural protein NS1 in influenza viruses. Absent in the viral particle, but highly expressed in the infected cell, NS1 dramatically inhibits cellular gene expression and prevents the activation of key players in the IFN system. In addition, NS1 selectively enhances the translation of viral mRNAs and may regulate the synthesis of viral RNAs. Our knowledge of the virus and of NS1 has increased dramatically during the last 15 years. The atomic structure of NS1 has been determined, many cellular partners have been identified and its multiple activities have been studied in depth. This review presents our current knowledge, and attempts to establish relationships between the RNA sequence, the structure of the protein, its ligands, its activities and the pathogenicity of the virus. A better understanding of NS1 could help in elaborating novel antiviral strategies, based on either live vaccines with altered NS1 or on small-compound inhibitors of NS1. IntroductionInfluenza viruses are enveloped viruses whose genome, 13 kb in size, is made up of eight ssRNAs of negative polarity. Transcription and replication of viral RNAs take place in the nucleus of the infected cell, generating at least 18 different species of positive-strand viral RNAs. These include (i) the eight complementary RNAs that act as templates for the synthesis of new genomic RNAs, (ii) eight unspliced mRNAs and (iii) at least two spliced mRNAs. The viral mRNas collectively encode up to 15 proteins, which schematically fall in three classes: (i) the four membraneassociated proteins comprise the two major surface glycoproteins haemagglutinin (HA) and neuraminidase (NA), the ion channel M2, and the matrix protein M1, which coats the inner face of the viral envelope; (ii) the three subunits of the viral polymerase (PA, PB1 and PB2) are tightly associated with the viral ribonucleoproteins (RNPs), in which the viral RNAs are bound to the nucleoprotein (NP); and, finally, (iii) nuclear export protein (NEP, formerly known as NS2) is also present in the virion, whilst NS1, PB1-F2, PA-X, PA-N155, PA-N182 and N40 are expressed in the infected cells, but absent from the viral particle. The unspliced viral mRNAs encode eight full-length products, and three PB1-and PArelated N-terminally truncated products that are not uniformly expressed among different strains (PB1-N40, PA-N155 and PA-N182) (Muramoto et al., 2013;Wise et al., 2009). In addition, PB1-F2 is encoded in the (+1) reading frame of PB1 and PA-X is produced from the PA mRNA through a ribosomal frameshift (Jagger et al., 2012). Finally,~10-15 % of the M-and NS-specific mRNAs are spliced , thus encoding M2 and NEP, respectively.Amongst the viral proteins, non-structural protein NS1 is probably that which is most involved in the interactions between the virus and the host cell, notably in antagonizing the antiviral response. It is therefore a key player in the viral cycle (Hale et al., 2...

A deletion of about 20 amino acids in the stalk of the neuraminidase (NA) is frequently detected upon transmission of influenza A viruses from waterfowl to domestic poultry. Using reverse genetics, a recombinant virus derived from a wild duck influenza virus isolate, A/Mallard/Marquenterre/Z237/83 (MZ), and an NA stalk deletion variant (MZ-delNA) were produced. Compared to the wild type, the MZ-delNA virus showed a moderate growth advantage on avian cultured cells. In 4-week-old chickens inoculated intratracheally with the MZ-delNA virus, viral replication in the lungs, liver, and kidneys was enhanced and interstitial pneumonia lesions were more severe than with the wild-type virus. The MZ-delNA-inoculated chickens showed significantly increased levels of mRNAs encoding interleukin-6 (IL-6), transforming growth factor-␤4 (TGF-␤4), and CCL5 in the lungs and a higher frequency of apoptotic cells in the liver than did their MZ-inoculated counterparts. Molecular mechanisms possibly underlying the growth advantage of the MZ-delNA virus were explored. The measured enzymatic activities toward a small substrate were similar for the wild-type and deleted NA, but the MZ-delNA virus eluted from chicken erythrocytes at reduced rates. Pseudoviral particles expressing the MZ hemagglutinin in combination with the MZ-NA or MZ-delNA protein were produced from avian cultured cells with similar efficiencies, suggesting that the deletion in the NA stalk does not enhance the release of progeny virions and probably affects an earlier step of the viral cycle. Overall, our data indicate that a shortened NA stalk is a strong determinant of adaptation and virulence of waterfowl influenza viruses in chickens.