Phenotypic effects of mutations observed in the neuraminidase of human origin H5N1 influenza A viruses

Global spread and regional endemicity of H5Nx Goose/Guangdong avian influenza viruses (AIV) pose a continuous threat for poultry production and zoonotic, potentially pre-pandemic, transmission to humans. Little is known about the role of mutations in the viral neuraminidase (NA) that accompanied bird-to-human transmission to support AIV infection of mammals. Here, after detailed analysis of the NA sequence of human H5N1 viruses, we studied the role of A46D, L204M, S319F and S430G mutations in virus fitness in vitro and in vivo. Although H5N1 AIV carrying avian- or human-like NAs had similar replication efficiency in avian cells, human-like NA enhanced virus replication in human airway epithelia. The L204M substitution consistently reduced NA activity of H5N1 and nine other influenza viruses carrying NA of groups 1 and 2, indicating a universal effect. Compared to the avian ancestor, human-like H5N1 virus has less NA incorporated in the virion, reduced levels of viral NA RNA replication and NA expression. We also demonstrate increased accumulation of NA at the plasma membrane, reduced virus release and enhanced cell-to-cell spread. Furthermore, NA mutations increased virus binding to human-type receptors. While not affecting high virulence of H5N1 in chickens, the studied NA mutations modulated virulence and replication of H5N1 AIV in mice and to a lesser extent in ferrets. Together, mutations in the NA of human H5N1 viruses play different roles in infection of mammals without affecting virulence or transmission in chickens. These results are important to understand the genetic determinants for replication of AIV in mammals and should assist in the prediction of AIV with zoonotic potential

Host Cell Restriction Factors of Bunyaviruses and Viral Countermeasures

The Bunyavirales order comprises more than 500 viruses (generally defined as bunyaviruses) classified into 12 families. Some of these are highly pathogenic viruses infecting different hosts, including humans, mammals, reptiles, arthropods, birds, and/or plants. Host cell sensing of infection activates the innate immune system that aims at inhibiting viral replication and propagation. Upon recognition of pathogen-associated molecular patterns (PAMPs) by cellular pattern recognition receptors (PRRs), numerous signaling cascades are activated, leading to the production of interferons (IFNs). IFNs act in an autocrine and paracrine manner to establish an antiviral state by inducing the expression of hundreds of IFN-stimulated genes (ISGs). Some of these ISGs are known to restrict bunyavirus infection. Along with other constitutively expressed host cellular factors with antiviral activity, these proteins (hereafter referred to as “restriction factors”) target different steps of the viral cycle, including viral entry, genome transcription and replication, and virion egress. In reaction to this, bunyaviruses have developed strategies to circumvent this antiviral response, by avoiding cellular recognition of PAMPs, inhibiting IFN production or interfering with the IFN-mediated response. Herein, we review the current knowledge on host cellular factors that were shown to restrict infections by bunyaviruses. Moreover, we focus on the strategies developed by bunyaviruses in order to escape the antiviral state developed by the infected cells

Host Cell Restriction Factors of Bunyaviruses and Viral Countermeasures

International audienceThe Bunyavirales order comprises more than 500 viruses (generally defined as bunyaviruses) classified into 12 families. Some of these are highly pathogenic viruses infecting different hosts, including humans, mammals, reptiles, arthropods, birds, and/or plants. Host cell sensing of infection activates the innate immune system that aims at inhibiting viral replication and propagation. Upon recognition of pathogen-associated molecular patterns (PAMPs) by cellular pattern recognition receptors (PRRs), numerous signaling cascades are activated, leading to the production of interferons (IFNs). IFNs act in an autocrine and paracrine manner to establish an antiviral state by inducing the expression of hundreds of IFN-stimulated genes (ISGs). Some of these ISGs are known to restrict bunyavirus infection. Along with other constitutively expressed host cellular factors with antiviral activity, these proteins (hereafter referred to as “restriction factors”) target different steps of the viral cycle, including viral entry, genome transcription and replication, and virion egress. In reaction to this, bunyaviruses have developed strategies to circumvent this antiviral response, by avoiding cellular recognition of PAMPs, inhibiting IFN production or interfering with the IFN-mediated response. Herein, we review the current knowledge on host cellular factors that were shown to restrict infections by bunyaviruses. Moreover, we focus on the strategies developed by bunyaviruses in order to escape the antiviral state developed by the infected cells

The interplays between Crimean-Congo hemorrhagic fever virus (CCHFV) M segment-encoded accessory proteins and structural proteins promote virus assembly and infectivity

International audienceCrimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne orthonairovirus that has become a serious threat to the public health. CCHFV has a single-stranded, tripartite RNA genome composed of L, M, and S segments. Cleavage of the M polyprotein precursor generates the two envelope glycoproteins (GPs) as well as three secreted nonstructural proteins GP38 and GP85 or GP160, representing GP38 only or GP38 linked to a mucin-like protein (MLD), and a double-membrane-spanning protein called NSm. Here, we examined the relevance of each M-segment non-structural proteins in virus assembly, egress and infectivity using a well-established CCHFV virus-like-particle system (tc-VLP). Deletion of MLD protein had no impact on infectivity although it reduced by 60% incorporation of GPs into particles. Additional deletion of GP38 abolished production of infectious tc-VLPs. The loss of infectivity was associated with impaired Gc maturation and exclusion from the Golgi, showing that Gn is not sufficient to target CCHFV GPs to the site of assembly. Consistent with this, efficient complementation was achieved in cells expressing MLD-GP38 in trans with increased levels of preGc to Gc conversion, co-targeting to the Golgi, resulting in particle incorporation and restored infectivity. Contrastingly, a MLD-GP38 variant retained in the ER allowed preGc cleavage but failed to rescue miss-localization or infectivity. NSm deletion, conversely, did not affect trafficking of Gc but interfered with Gc processing, particle formation and secretion. NSm expression affected N-glycosylation of different viral proteins most likely due to increased speed of trafficking through the secretory pathway. This highlights a potential role of NSm in overcoming Golgi retention and facilitating CCHFV egress. Thus, deletions of GP38 or NSm demonstrate their important role on CCHFV particle production and infectivity. GP85 is an essential viral factor for preGc cleavage, trafficking and Gc incorporation into particles, whereas NSm protein is involved in CCHFV assembly and virion secretion

A highly virulent variant of HIV-1 circulating in the Netherlands

We discovered a highly virulent variant of subtype-B HIV-1 in the Netherlands. One hundred nine individuals with this variant had a 0.54 to 0.74 log10 increase (i.e., a ~3.5-fold to 5.5-fold increase) in viral load compared with, and exhibited CD4 cell decline twice as fast as, 6604 individuals with other subtype-B strains. Without treatment, advanced HIV-CD4 cell counts below 350 cells per cubic millimeter, with long-term clinical consequences-is expected to be reached, on average, 9 months after diagnosis for individuals in their thirties with this variant. Age, sex, suspected mode of transmission, and place of birth for the aforementioned 109 individuals were typical for HIV-positive people in the Netherlands, which suggests that the increased virulence is attributable to the viral strain. Genetic sequence analysis suggests that this variant arose in the 1990s from de novo mutation, not recombination, with increased transmissibility and an unfamiliar molecular mechanism of virulence