DNA-origami-directed virus capsid polymorphism

Viral capsids can adopt various geometries, most iconically characterized by icosahedral or helical symmetries. Importantly, precise control over the size and shape of virus capsids would have advantages in the development of new vaccines and delivery systems. However, current tools to direct the assembly process in a programmable manner are exceedingly elusive. Here we introduce a modular approach by demonstrating DNA-origami-directed polymorphism of single-protein subunit capsids. We achieve control over the capsid shape, size and topology by employing user-defined DNA origami nanostructures as binding and assembly platforms, which are efficiently encapsulated within the capsid. Furthermore, the obtained viral capsid coatings can shield the encapsulated DNA origami from degradation. Our approach is, moreover, not limited to a single type of capsomers and can also be applied to RNA–DNA origami structures to pave way for next-generation cargo protection and targeting strategies.</p

Protein Cargo Encapsulation by Virus-Like Particles: Strategies and Applications

Viruses and the recombinant protein cages assembled from their structural proteins, known as virus-like particles (VLPs), have gained wide interest as tools in biotechnology and nanotechnology. Detailed structural information and their amenability to genetic and chemical modification make them attractive systems for further engineering. This review describes the range of non-enveloped viruses that have been co-opted for heterologous protein cargo encapsulation and the strategies that have been developed to drive encapsulation. Spherical capsids of a range of sizes have been used as platforms for protein cargo encapsulation. Various approaches, based on native and non-native interactions between the cargo proteins and inner surface of VLP capsids, have been devised to drive encapsulation. Here we outline the evolution of these approaches, discussing their benefits and limitations. Like the viruses from which they are derived, VLPs are of interest in both biomedical and materials applications. The encapsulation of protein cargo inside VLPs leads to numerous uses in both fundamental and applied biocatalysis and biomedicine, some of which are discussed herein. The applied science of protein encapsulating VLPs is emerging as a research field with great potential. Developments in loading control, higher order assembly, and capsid optimization are poised to realize this potential in the near future

HPeV-3 predominated among Parechovirus A positive infants during an outbreak in 2013-2014 in Queensland, Australia

Parechoviruses (HPeV) are endemic seasonal pathogens detected from the respiratory tract, gut, blood and central nervous system (CNS) of children and adults, sometimes in conjunction with a range of acute illnesses. HPeV CNS infection may lead to neurodevelopmental sequelae, especially following infection by HPeV-3, hence screening and genotyping are important to inform epidemiology, aetiology and prognosis.To identify and characterise HPeVs circulating during an outbreak between November 2013 and April 2014 in Queensland, Australia.To perform PCR-based screening and comparative nucleotide sequence analysis on samples from children with clinically suspected infections submitted to a research laboratory for HPeV investigations.HPeVs were detected among 25/62 samples, identified as HPeV-3 from 23 that could be genotyped. These variants closely matched those which have occurred worldwide and in other States of Australia.The inclusion of PCR-based HPeV testing is not systematically applied but should be considered essential for children under 3 months of age with CNS symptoms as should long-term follow-up of severe sepsis-like cases

Tunable In Vivo Co-localisation of Enzymes Within P22 Capsid-Based Nanoreactors

The spatial organisation of enzymatic pathways through compartmentalisation is a mechanism used in nature for the regulation of multi-step biocatalytic processes. Virus-like particles (VLPs) derived from Bacteriophage P22 have been explored as biomimetic catalytic compartments. The in vivo co-encapsulation of enzymes is typically achieved via sequential fusion to the scaffold protein (SP), which results in an equimolar ratio of enzyme monomers. However, control over enzyme stoichiometry, which has been shown to influence pathway flux, is key to realising the full potential of P22 VLPs as artificial metabolons. Here we present a strategy for the stoichiometrically controlled in vivo co-encapsulation of cargo proteins within P22-based VLPs. Co-encapsulation was achieved via co-expression of cargo proteins with individual SP fusions using a dual plasmid system and verified for fluorescent protein cargo by Förster resonance energy transfer. This strategy was subsequently applied to a two-enzyme reaction cascade. L-homoalanine, an unnatural amino acid and chiral precursor to several drugs, can be synthesised from the readily available L-threonine by the sequential activity of threonine dehydratase and glutamate dehydrogenase. We find that scaffolding by this system has a profound impact on the activity of each enzyme and, using a purification strategy designed to isolate the range of particle forms that exist in vivo, that scaffolding of multimeric enzymes can be at unexpectedly high densities. This work demonstrates the controlled co-localisation of multiple heterologous cargo proteins in a P22-based nanoreactor and shows that careful consideration of loading densities of individual enzymes in an enzymatic cascade is required for the optimal design of synthetic metabolons

DNA-origami-directed virus capsid polymorphism

| openaire: EC/H2020/101002258/EU//ProCrystal Funding Information: The authors acknowledge financial support from the European Research Council (ERC) and ERA Chair MATTER under the European Union’s Horizon 2020 research and innovation programme (grant agreement numbers 101002258 (M.A.K.) and 856705 (V. Linko)), the Emil Aaltonen Foundation (V. Linko), the Sigrid Jusélius Foundation (V. Linko), the Academy of Finland (grant numbers 341057 (E.A.-P.) and 314671 (M.A.K)), the Finnish Foundation for Technology Promotion (V. Lampinen), and the Jane and Aatos Erkko Foundation (V. Linko and M.A.K.). This work was carried out under the Academy of Finland Centers of Excellence Program (2022-2029) in Life-Inspired Hybrid Materials (LIBER), project number number 346110 (M.A.K.). K. M. Nguyen and A. Kuzyk are acknowledged for providing the staple strands for the 13HR sample, P. Laurinmäki and B. Löflund for technical assistance with cryo-EM, M. Hankaniemi for support with NoV production, and M. Sammalkorpi and A. Scacchi for technical discussions. The facilities and expertise of the HiLIFE cryo-EM unit at the University of Helsinki, a member of Instruct-ERIC Centre Finland, FINStruct and Biocenter Finland are gratefully acknowledged. The authors also acknowledge CSC–IT Center of Science, Finland, for computational resources, Biocenter Finland for support of protein production and characterization infrastructure, and the provision of facilities and technical support by Aalto University Bioeconomy Facilities, OtaNanoNanomicroscopy Center (Aalto-NMC) and Micronova Nanofabrication Center. Publisher Copyright: © 2023, The Author(s).Viral capsids can adopt various geometries, most iconically characterized by icosahedral or helical symmetries. Importantly, precise control over the size and shape of virus capsids would have advantages in the development of new vaccines and delivery systems. However, current tools to direct the assembly process in a programmable manner are exceedingly elusive. Here we introduce a modular approach by demonstrating DNA-origami-directed polymorphism of single-protein subunit capsids. We achieve control over the capsid shape, size and topology by employing user-defined DNA origami nanostructures as binding and assembly platforms, which are efficiently encapsulated within the capsid. Furthermore, the obtained viral capsid coatings can shield the encapsulated DNA origami from degradation. Our approach is, moreover, not limited to a single type of capsomers and can also be applied to RNA–DNA origami structures to pave way for next-generation cargo protection and targeting strategies.Peer reviewe

Particle and bioaerosol characteristics in a paediatric intensive care unit

The paediatric intensive care unit (PICU) provides care to critically ill neonates, infants and children. These patients are vulnerable and susceptible to the environment surrounding them, yet there is little information available on indoor air quality and factors affecting it within a PICU. To address this gap in knowledge we conducted continuous indoor and outdoor airborne particle concentration measurements over a two-week period at the Royal Children's Hospital PICU in Brisbane, Australia, and we also collected 82 bioaerosol samples to test for the presence of bacterial and viral pathogens. Our results showed that both 24-hour average indoor particle mass (PM10) (0.6-2.2 mu g m(-3), median: 0.9 mu g m(-3)) and submicrometer particle number (PN) (0.1-2.8 x10(3) p cm(-3), median: 0.67 x10(3) p cm(-3)) concentrations were significantly lower (p < 0.01) than the outdoor concentrations (6.7-10.2 mu g m(-3), median: 8.0 mu g m(-3) for PM10 and 12.1-22.2 x10(3) p cm(-3), median: 16.4 x10(3) p cm(-3) for PN). In general, we found that indoor particle concentrations in the PICU were mainly affected by indoor particle sources, with outdoor particles providing a negligible background. We identified strong indoor particle sources in the PICU, which occasionally increased indoor PN and PM10 concentrations from 0.1 x10(3) to 100 x10(3) p cm(-3), and from 2 mu g m(-3) to 70 mu g m(-3), respectively. The most substantial indoor particle sources were nebulization therapy, tracheal suction and cleaning activities. The average PM10 and PN emission rates of nebulization therapy ranged from 1.29 to 7.41 mg min(-1) and from 1.20 to 3.96 p min(-1)x 10(11), respectively. Based on multipoint measurement data, it was found that particles generated at each location could be quickly transported to other locations, even when originating from isolated single-bed rooms. The most commonly isolated bacterial genera from both primary and broth cultures were skin commensals while viruses were rarely identified. Based on the findings from the study, we developed a set of practical recommendations for PICU design, as well as for medical and cleaning staff to mitigate aerosol generation and transmission to minimize infection risk to PICU patients

Statement in Support of: “Virology under the Microscope—a Call for Rational Discourse”

[Extract] We, members of the Australasian Virology Society, agree with and support the statement entitled “Virology under the Microscope—a Call for Rational Discourse” (1). Like virologists everywhere, we have worked with scientist and clinician colleagues worldwide to develop knowledge, tests, and interventions which collectively have reduced the number of deaths due to COVID-19 and curtailed its economic impact. Such work adds to the extraordinary achievements resulting from virology research that have delivered vaccines and/or antivirals against a long list of diseases and global scourges, including AIDS, smallpox, and polio (1). We believe the question of the origin of SARS-CoV-2 should be approached with an open mind and in consideration of the best scientific evidence available. We concur with the view that the zoonosis hypothesis has the strongest supporting evidence (2–4), and this is a scenario that has been observed repeatedly in the past (5), including in Australia (6). Recent data strongly support the zoonosis hypothesis (7). We share the concern that emotive and fear-based dialogues in this area add to public confusion and can lead to ill-informed condemnation of virology research