Imaging Bunyavirus Infections by Transmission Electron Microscopy: Conventional Sample Preparation vs High-Pressure Freezing and Freeze-Substitution.

Transmission electron microscopy significantly contributed to unveil the course of virus entry, replication, morphogenesis, and egress. For these studies, the most widely used approach is imaging ultrathin sections of virus-infected cells embedded in a plastic resin that is transparent to electrons. Before infiltration in a resin, cells must be processed to stabilize their components under the observation conditions in an electron microscope, such as high vacuum and irradiation with electrons.

Molecular mapping of virus-infected cells with immunogold and metal-tagging transmission electron microscopy.

Transmission electron microscopy (TEM) has been essential to study virus-cell interactions. The architecture of viral replication factories, the principles of virus assembly and the components of virus egress pathways are known thanks to the contribution of TEM methods. Specially, when studying viruses in cells, methodologies for labeling proteins and other macromolecules are important tools to correlate morphology with function.

β-Cyclodextrins as affordable antivirals to treat coronavirus infection.

The SARS-CoV-2 pandemic made evident that there are only a few drugs against coronavirus. Here we aimed to identify a cost-effective antiviral with broad spectrum activity and high safety profile. Starting from a list of 116 drug candidates, we used molecular modelling tools to rank the 44 most promising inhibitors.

Unraveling the antiviral activity of plitidepsin against SARS-CoV-2 by subcellular and morphological analysis.

The pandemic caused by the new coronavirus SARS-CoV-2 has made evident the need for broad-spectrum, efficient antiviral treatments to combat emerging and re-emerging viruses. Plitidepsin is an antitumor agent of marine origin that has also shown a potent pre-clinical efficacy against SARS-CoV-2. Plitidepsin targets the host protein eEF1A (eukaryotic translation elongation factor 1 alpha) and affects viral infection at an early, post-entry step.

Reovirus Nonstructural Protein σNS Recruits Viral RNA to Replication Organelles.

The function of the mammalian orthoreovirus (reovirus) σNS nonstructural protein is enigmatic. σNS is an RNA-binding protein that forms oligomers and enhances the stability of bound RNAs, but the mechanisms by which it contributes to reovirus replication are unknown. To determine the function of σNS-RNA binding in reovirus replication, we engineered σNS mutants deficient in RNA-binding capacity.

Ins and Outs of Reovirus: Vesicular Trafficking in Viral Entry and Egress.

Cell entry and egress are essential steps in the viral life cycle that govern pathogenesis and spread. Mammalian orthoreoviruses (reoviruses) are nonenveloped viruses implicated in human disease that serve as tractable models for studies of pathogen-host interactions. In this review we discuss the function of intracellular vesicular transport systems in reovirus entry, trafficking, and egress and comment on shared themes for diverse viruses.

A modified lysosomal organelle mediates nonlytic egress of reovirus.

Mammalian orthoreoviruses (reoviruses) are nonenveloped viruses that replicate in cytoplasmic membranous organelles called viral inclusions (VIs) where progeny virions are assembled. To better understand cellular routes of nonlytic reovirus exit, we imaged sites of virus egress in infected, nonpolarized human brain microvascular endothelial cells (HBMECs) and observed one or two distinct egress zones per cell at the basal surface. Transmission electron microscopy and 3D electron tomography (ET) of the egress zones revealed clusters of virions within membrane-bound structures, which we term membranous carriers (MCs), approaching and fusing with the plasma membrane.

The viral replication organelles within cells studied by electron microscopy.

Transmission electron microscopy (TEM) has been crucial to study viral infections. As a result of recent advances in light and electron microscopy, we are starting to be aware of the variety of structures that viruses assemble inside cells. Viruses often remodel cellular compartments to build their replication factories.

Function, Architecture, and Biogenesis of Reovirus Replication Neoorganelles.

Most viruses that replicate in the cytoplasm of host cells form neoorganelles that serve as sites of viral genome replication and particle assembly. These highly specialized structures concentrate viral proteins and nucleic acids, prevent the activation of cell-intrinsic defenses, and coordinate the release of progeny particles. Reoviruses are common pathogens of mammals that have been linked to celiac disease and show promise for oncolytic applications.

Reovirus σNS and μNS Proteins Remodel the Endoplasmic Reticulum to Build Replication Neo-Organelles.

Like most viruses that replicate in the cytoplasm, mammalian reoviruses assemble membranous neo-organelles called inclusions that serve as sites of viral genome replication and particle morphogenesis. Viral inclusion formation is essential for viral infection, but how these organelles form is not well understood. We investigated the biogenesis of reovirus inclusions.

Virus assembly factories in a lipid world.

Many viruses build specialized structures known as viral factories, a protected environment in which viral genome replication and morphogenesis take place. Recent findings show that viruses manipulate lipid flows to assemble these replication platforms. Viruses are thus able to create new membranes by interfering with lipid metabolism, targeting and transport; they make use of specific lipid transfer proteins (LTP) at membrane contact sites, and frequently recruit endoplasmic reticulum (ER), ER export sites, and mitochondria.

Heterogeneous virulence of pandemic 2009 influenza H1N1 virus in mice.

Background: Understanding the pathogenesis of influenza infection is a key factor leading to the prevention and control of future outbreaks. Pandemic 2009 Influenza H1N1 infection, although frequently mild, led to a severe and fatal form of disease in certain cases that make its virulence nature debatable. Much effort has been made toward explaining the determinants of disease severity; however, no absolute reason has been established.

Double stranded RNA activated EIF2 alpha kinase (EIF2AK2; PKR) is associated with Alzheimer's disease.

Sporadic Alzheimer's disease (AD) appears to be the consequence of the interaction between combinations of genes and environmental factors (for example virus infections). To test this hypothesis, we are examining human genes relevant to herpes simplex virus type 1 (HSV-1) infection via genetic association studies in AD case-control samples. Recently, we found that a variant in TAP2, a major target used by HSV-1 to evade immune surveillance, is associated with AD.

Association of DSC1, a gene modulated by adrenergic stimulation, with Alzheimer's disease.

Alzheimer's disease (AD) is a complex multifactorial disorder involving a number of genetic and environmental factors, with severe head injury consistently reported as a major non-genetic risk factor. The adrenergic activation that occurs during major trauma increases cAMP levels, therefore the cAMP signaling pathway might be involved in AD pathogenesis. Time course of candidate gene expression following adrenergic stimulation with isoproterenol was assayed in neuroblastoma cells by quantitative reverse transcription (RT)-PCR.

Hippocampal volume in childhood complex partial seizures.

Purpose: This study compared hippocampal volume in children with cryptogenic epilepsy, all of whom had complex partial seizures (CPS), and age and gender matched normal children controlling for between group differences in IQ and demographic variables (e.g., age, gender, ethnicity, socioeconomic status).