SNAP-tagged Chikungunya Virus Replicons Improve Visualisation of Non-Structural Protein 3 by Fluorescence Microscopy

Chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes febrile disease, muscle and joint pain, which can become chronic in some individuals. The non-structural protein 3 (nsP3) plays essential roles during infection, but a complete understanding of its function is lacking. Here we used a microscopy-based approach to image CHIKV nsP3 inside human cells. The SNAP system consists of a self-labelling enzyme tag, which catalyses the covalent linking of exogenously supplemented synthetic ligands. Genetic insertion of this tag resulted in viable replicons and specific labelling while preserving the effect of nsP3 on stress granule responses and co-localisation with GTPase Activating Protein (SH3 domain) Binding Proteins (G3BPs). With sub-diffraction, three-dimensional, optical imaging, we visualised nsP3-positive structures with variable density and morphology, including high-density rod-like structures, large spherical granules, and small, low-density structures. Next, we confirmed the utility of the SNAP tag for studying protein turnover by pulse-chase labelling. We also revealed an association of nsP3 with cellular lipid droplets and examined the spatial relationships between nsP3 and the non-structural protein 1 (nsP1). Together, our study provides a sensitive, specific, and versatile system for fundamental research into the individual functions of a viral non-structural protein during infection with a medically important arthropod-borne virus (arbovirus)

A systematic review of participatory integrated assessment at the catchment scale: Lessons learned from practice

Participatory integrated assessment (PIA) emerged as a response to conventional integrated assessment methods in the mid-to-late 1990s. PIA is based on the tenet that more inclusive stakeholder involvement may lead to increased accountability and legitimacy in decision-making, greater levels of trust and social learning between participants, and improved quality and relevancy of knowledge outputs. In this paper, we conduct a systematic literature review to update and deepen our understanding of the approaches, methods, opportunities, and challenges associated with PIA as applied at the catchment scale. Of the total 278 studies identified in our literature search, only 37 catchment-level cases presented a clear PIA application. From our review, lessons learnt were drawn in relation to the integration of less-easily quantified areas of social science, entry and exit planning in PIA, boundary work on issue cycles and accounting for the human dimension. We conclude that PIA is a potentially useful approach for navigating the dual social-ecological dimensions of current environmental and resource management issues, especially when projects include tailored objectives and methods, user-friendly outputs, and early and consistent stakeholder involvement. However, we also highlight gaps in the field concerning the integrative reach of PIA, PIA's real-world impact, and the relationship between PIA processes and outcomes along stages of environmental issue cycles. We conclude that further work is therefore still needed to help advance the field of PIA in both research and boundary work practice

On the Tibur road; a freshman's Horace,

Partly reprinted from various periodicals.First published 1911.Mode of access: Internet