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International audience; The incidence of arbovirus infections has increased dramatically in recent decades, affecting hundreds of millions of people each year. The Togaviridae family includes the chikungunya virus (CHIKV), which is typically transmitted by Aedes mosquitoes and causes a wide range of symptoms from flu-like fever to severe arthralgia. Although conventional diagnostic tests can provide early diagnosis of CHIKV infections, access to these tests is often limited in developing countries. Consequently, there is an urgent need to develop efficient, affordable, simple, rapid, and robust diagnostic tools that can be used in point-of-care settings. Early diagnosis is crucial to improve patient management and to reduce the risk of complications. A glass-fiber laser-cut microfluidic device (paper-based analytical device [PAD]) was designed and evaluated in a proof of principle context, for the analysis of 30 µL of patient serum. Biological raw materials used for the functionalization of the PAD were first screened by MAC-ELISA (IgM capture enzyme-linked immunosorbent assay) for CHIKV Immunoglobulin M (IgM) capture and then evaluated on the PAD using various human samples. Compared with viral lysate traditionally used for chikungunya (CHIK) serology, CHIKV pseudo-particles (PPs) have proven to be powerful antigens for specific IgM capture. The PAD was able to detect CHIKV IgM in human sera in less than 10 minutes. Results obtained in patient sera showed a sensitivity of 70.6% and a specificity of around 98%. The PAD showed few cross-reactions with other tropical viral diseases. The PAD could help health workers in the early diagnosis of tropical diseases such as CHIK, which require specific management protocols in at-risk populations.

Viral pathogens are a serious health threat around the world, particularly in resource limited settings, where current sensing approaches are often insufficient and slow, compounding the spread and burden of these pathogens. Here, we describe a label-free, point-of-care approach toward detection of virus particles, based on a microfluidic paper-based analytical device with integrated microwire Au electrodes. The device is initially characterized through capturing of streptavidin modified nanoparticles by biotin-modified microwires. An order of magnitude improvement in detection limits is achieved through use of a microfluidic device over a classical static paper-based device, due to enhanced mass transport and capturing of particles on the modified electrodes. Electrochemical impedance spectroscopy detection or West Nile virus particles was carried out using antibody runctionalized Au microwires, achieving a detection limit of 10.2 particles in 50 μL of cell culture media. No increase in signal is found on addition of an excess of a nonspecific target (Sindbis). This detection motif is significantly cheaper (~$1 per test) and faster (~30 min) than current methods, while achieving the desired selectivity and sensitivity. This sensing motif represents a general platform for trace detection of a wide range of biological pathogens.

The capsids of certain Archaea-infecting viruses undergo large shape changes, while maintaining their integrity against rupture by osmotic pressure. We propose that these capsids are in a smectic liquid crystalline state, with the capsid proteins assembling along spirals. We show that smectic capsids are intrinsically stabilized against the formation of localized bulges with non-zero Gauss curvature while still allowing for large-scale cooperative shape transformation that involves global changes in the Gauss curvature.

Under many in vitro conditions, some small viruses spontaneously encapsidate a single stranded (ss) RNA into a protein shell called the capsid. While viral RNAs are found to be compact and highly branched because of long distance base-pairing between nucleotides, recent experiments reveal that in a head-to-head competition between an ssRNA with no secondary or higher order structure and a viral RNA, the capsid proteins preferentially encapsulate the linear polymer! In this paper, we study the impact of genome stiffness on the encapsidation free energy of the complex of RNA and capsid proteins. We show that an increase in effective chain stiffness because of base-pairing could be the reason why under certain conditions linear chains have an advantage over branched chains when it comes to encapsidation efficiency. While branching makes the genome more compact, RNA base-pairing increases the effective Kuhn length of the RNA molecule, which could result in an increase of the free energy of RNA confinement, that is, the work required to encapsidate RNA, and thus less efficient packaging.

Hollow protein containers are widespread in nature, and include virus capsids as well as eukaryotic and bacterial complexes. Protein cages are studied extensively for applications in nanotechnology, nanomedicine and materials science. Their inner and outer surfaces can be modified chemically or genetically, and the internal cavity can be used to template, store and/or arrange molecular cargos. Virus capsids and virus-like particles (VLP, noninfectious particles) provide versatile platforms for nanoscale bioengineering. Study of capsid protein self-assembly into monodispersed particles, and of VLP structure and biophysics is necessary not only to understand natural processes, but also to infer how these platforms can be redesigned to furnish novel functional VLP. Here we address the assembly dynamics of infectious bursal disease virus (IBDV), a complex icosahedral virus. IBDV has a ~70 nm-diameter T = 13 capsid with VP2 trimers as the only structural subunits. During capsid assembly, VP2 is synthesized as a precursor (pVP2) whose C terminus is cleaved. The pVP2 C terminus has an amphipathic helix that controls VP2 polymorphism. In the absence of the VP3 scaffolding protein, necessary for control of assembly, 466/456-residue pVP2 intermediates bearing this helix assemble into VLP only when expressed with an N-terminal His6 tag (the HT-VP2-466 protein). HT-VP2-466 capsids are optimal for genetic insertion of proteins (cargo space ~78 000 nm3). We established an in vitro assembly/disassembly system of HT-VP2-466-based VLP for heterologous nucleic acid packaging and/or encapsulation of drugs and other molecules. HT-VP2-466 (empty) capsids were disassembled and reassembled by dialysis against low-salt/basic pH and high-salt/acid pH buffers, respectively, thus illustrating the reversibility in vitro of IBDV capsid assembly. HT-VP2-466 VLP also packed heterologous DNA by non-specific confinement during assembly. These and previous results establish the bases for biotechnological applications based on the IBDV capsid and its ability to incorporate exogenous proteins and nucleic acids.

We consider self-assembly of proteins into a virus capsid by the methods of molecular dynamics. The capsid corresponds either to SPMV or CCMV and is studied with and without the RNA molecule inside. The proteins are flexible and described by the structure-based coarse-grained model augmented by electrostatic interactions. Previous studies of the capsid self-assembly involved solid objects of a supramolecular scale, e.g. corresponding to capsomeres, with engineered couplings and stochastic movements. In our approach, a single capsid is dissociated by an application of a high temperature for a variable period and then the system is cooled down to allow for self-assembly. The restoration of the capsid proceeds to various extent, depending on the nature of the dissociated state, but is rarely complete because some proteins depart too far unless the process takes place in a confined space., 13 figures