The molecular processes that determine the outcome of influenza virus infection in humans are multifactorial and involve a complex interplay between host, viral and bacterial factors. However, it is generally accepted that a strong innate immune dysregulation known as 'cytokine storm' contributes to the pathology of infections with the 1918 H1N1 pandemic or the highly pathogenic avian influenza viruses of the H5N1 subtype. The RNA sensor retinoic acid-inducible gene I (RIG-I) plays an important role in sensing viral infection and initiating a signalling cascade that leads to interferon expression. Here, we show that short aberrant RNAs (mini viral RNAs (mvRNAs)), produced by the viral RNA polymerase during the replication of the viral RNA genome, bind to and activate RIG-I and lead to the expression of interferon-β. We find that erroneous polymerase activity, dysregulation of viral RNA replication or the presence of avian-specific amino acids underlie mvRNA generation and cytokine expression in mammalian cells. By deep sequencing RNA samples from the lungs of ferrets infected with influenza viruses, we show that mvRNAs are generated during infection in vivo. We propose that mvRNAs act as the main agonists of RIG-I during influenza virus infection.

The mechanism by which ZIKV causes a range of neurological complications, especially congenital microcephaly, is not well understood. The fact that congenital microcephaly is associated with Asian lineage ZIKV strains raises the question of why this was not discovered earlier. One possible explanation is that Asian and African ZIKV strains differ in their abilities to infect cells of the CNS and to cause neurodevelopmental problems. Here, we show that Asian ZIKV strains infect and induce cell death in human neural progenitor cells—which are important target cells in the development of congenital microcephaly—less efficiently than African ZIKV strains. These features of Asian ZIKV strains likely contribute to their ability to cause chronic infections, often observed in congenital microcephaly cases. It is therefore likely that phenotypic differences between ZIKV strains could be, at least in part, responsible for the ability of Asian ZIKV strains to cause congenital microcephaly.

1918 H1N1 virus spread to and induced cytokine responses in tissues outside the respiratory tract, which likely contributed to the severity of infection. Moreover, our data support the suggested link between 1918 H1N1 infection and central nervous system disease.

Influenza A viruses from animal reservoirs have the capacity to adapt to humans and cause influenza pandemics. The occurrence of an influenza pandemic requires efficient virus transmission among humans, which is associated with virus attachment to the upper respiratory tract. Pandemic severity depends on virus ability to cause pneumonia, which is associated with virus attachment to the lower respiratory tract. Recently, a novel avian-origin H7N9 influenza A virus with unknown pandemic potential emerged in humans. We determined the pattern of attachment of two genetically engineered viruses containing the hemagglutinin of either influenza virus A/Shanghai/1/13 or A/Anhui/1/13 to formalin-fixed human respiratory tract tissues using histochemical analysis. Our results show that the emerging H7N9 virus attached moderately or abundantly to both upper and lower respiratory tract, a pattern not seen before for avian influenza A viruses. With the caveat that virus attachment is only the first step in the virus replication cycle, these results suggest that the emerging H7N9 virus has the potential both to transmit efficiently among humans and to cause severe pneumonia.

β-Thalassemia is associated with several abnormalities of the innate immune system. Neutrophils in particular are defective, predisposing patients to life-threatening bacterial infections. The molecular and cellular mechanisms involved in impaired neutrophil function remain incompletely defined. We used the Hbb β-thalassemia mouse and hemoglobin E (HbE)/β-thalassemia patients to investigate dysregulated neutrophil activity. Mature neutrophils from Hbb mice displayed a significant reduction in chemotaxis, opsonophagocytosis, and production of reactive oxygen species, closely mimicking the defective immune functions observed in β-thalassemia patients. In Hbb mice, the expression of neutrophil CXCR2, CD11b, and reduced NAD phosphate oxidase components (p22phox, p67phox, and gp91phox) were significantly reduced. Morphological analysis of Hbb neutrophils showed that a large percentage of mature phenotype neutrophils (Ly6GLy6C) appeared as band form cells, and a striking expansion of immature (Ly6GLy6C) hyposegmented neutrophils, consisting mainly of myelocytes and metamyelocytes, was noted. Intriguingly, expression of an essential mediator of neutrophil terminal differentiation, the ets transcription factor PU.1, was significantly decreased in Hbb neutrophils. In addition, in vivo infection with failed to induce PU.1 expression or upregulate neutrophil effector functions in Hbb mice. Similar changes to neutrophil morphology and PU.1 expression were observed in splenectomized and nonsplenectomized HbE/β-thalassemia patients. This study provides a mechanistic insight into defective neutrophil maturation in β-thalassemia patients, which contributes to deficiencies in neutrophil effector functions.

and propose an intramolecular copy-choice mechanism for mvRNA generation. 43 By deep-sequencing RNA samples from lungs of ferrets infected with influenza 44viruses we show that mvRNAs are generated during infection of animal models. 45 We propose that mvRNAs act as main agonists of RIG-I during influenza virus 46infection and the ability of influenza virus strains to generate mvRNAs should be 47 considered when assessing their virulence potential. 48The negative sense viral RNA (vRNA) genome segments of influenza A viruses, 49 as well as the complementary RNA (cRNA) replicative intermediates, contain 5ʹ 50 triphosphates and partially complementary 5ʹ and 3ʹ termini that serve as the viral 51 promoter for replication and transcription of the viral RNA genome 6 . RIG-I has been 52 shown to bind and be activated by the dsRNA structure formed by the termini of 53 influenza virus RNAs 7,8 . However, it remains unclear how RIG-I gains access to this 54 dsRNA structure. Both vRNA and cRNA are assembled into ribonucleoprotein 55 complexes (vRNP and cRNP, respectively) in which the viral RNA polymerase, a 56 heterotrimeric complex of the viral proteins PB1, PB2 and PA, associates with the 57 partially complementary termini, while the rest of the RNA is bound by oligomeric 58 nucleoprotein (NP) 6 (Fig. 1a). The tight binding of the 5ʹ and 3ʹ termini of vRNA and 59for use under a CC0 license.This article is a US Government work. It is not subject to copyright under 17 USC 105 and is also made available (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint . http://dx.doi.org/10.1101/385716 doi: bioRxiv preprint first posted online Aug. 6, 2018; 3 cRNA by the RNA polymerase 9 is likely to preclude an interaction with RIG-I. 60Moreover, it has been demonstrated that IFN expression is triggered only in a fraction 61 of influenza virus infected cells 10,11 , suggesting that influenza viruses efficiently hide 62 their genome segments during infection by replicating them in the context of RNPs 11 . 63This led to the proposal that an aberrant RNA replication product might be binding to 64 RIG-I and triggering IFN expression 12 . Indeed, the influenza virus polymerase is known 65 to generate defective interfering (DI) RNAs, which are ≥178 nt long subgenomic RNAs 66 generated during high multiplicity infections 13 , and small viral RNAs (svRNAs), which 67 are 22-27 nt long and correspond to the 5ʹ end of vRNA segments. However, svRNAs 68 have been shown not to be involved in the induction of antiviral cellular defences 14 and 69 DI RNAs assemble into RNP structures (Fig. 1a) NP and a luciferase reporter to measure the activation of the IFN-b promoter (Fig. 1b). 82We found that the expression of mvRNAs induced significantly higher IFN expression 83 than full-length vRNA or DI RNA, comparable to the levels induced by transfection of 84 2 µg of poly(I:C), a known activator of IFN expression 18 . Similar results were ob...

Influenza A viruses (IAV) and SARS-CoV-2 can spread via liquid droplets and aerosols. Face masks and other personal protective equipment (PPE) can act as barriers that prevent the spread of these viruses. However, IAV and SARS-CoV-2 are stable for hours on various materials, which makes frequent and correct disposal of these PPE important. Metal ions embedded into PPE may inactivate respiratory viruses, but confounding factors such as adsorption of viruses make measuring and optimizing the inactivation characteristics difficult. Here, we used polyamide 6.6 (PA66) fibers containing embedded zinc ions and systematically investigated if these fibers can adsorb and inactivate SARS-CoV-2 and IAV H1N1 when woven into a fabric. We found that our PA66-based fabric decreased the IAV H1N1 and SARS-CoV-2 titer by approximately 100-fold. Moreover, we found that the zinc content and the virus inactivating property of the fabric remained stable over 50 standardized washes. Overall, these results provide insights into the development of reusable PPE that offer protection against RNA virus spread.

Central nervous system (CNS) disease is one of the most common extrarespiratory tract complications of influenza A virus infections. Remarkably, zoonotic H5N1 virus infections are more frequently associated with CNS disease than seasonal or pandemic influenza viruses. Little is known about the interaction between influenza A viruses and cells of the CNS; therefore, it is currently unknown which viral factors are important for efficient replication. Here, we determined the replication kinetics of a seasonal, pandemic, zoonotic, and lab-adapted influenza A virus in human neuron-like (SK-N-SH) and astrocyte-like (U87-MG) cells and primary mouse cortex neurons. In general, highly pathogenic avian influenza (HPAI) H5N1 virus replicated most efficiently in all cells, which was associated with efficient attachment and infection. Seasonal H3N2 and to a lesser extent pandemic H1N1 virus replicated in a trypsin-dependent manner in SK-N-SH but not in U87-MG cells. In the absence of trypsin, only HPAI H5N1 and WSN viruses replicated. Removal of the multibasic cleavage site (MBCS) from HPAI H5N1 virus attenuated, but did not abrogate, replication. Taken together, our results showed that the MBCS and, to a lesser extent, the ability to attach are important determinants for efficient replication of HPAI H5N1 virus in cells of the CNS. This suggests that both an alternative hemagglutinin (HA) cleavage mechanism and preference for α-2,3-linked sialic acids allowing efficient attachment contribute to the ability of influenza A viruses to replicate efficiently in cells of the CNS. This study further improves our knowledge on potential viral factors important for the neurotropic potential of influenza A viruses. IMPORTANCE Central nervous system (CNS) disease is one of the most common extrarespiratory tract complications of influenza A virus infections, and the frequency and severity differ between seasonal, pandemic, and zoonotic influenza viruses. However, little is known about the interaction of these viruses with cells of the CNS. Differences among seasonal, pandemic, and zoonotic influenza viruses in replication efficacy in CNS cells, in vitro, suggest that the presence of an alternative HA cleavage mechanism and ability to attach are important viral factors. Identifying these viral factors and detailed knowledge of the interaction between influenza virus and CNS cells are important to prevent and treat this potentially lethal CNS disease.