Plaque assay free method for Semliki Forest virus quantification and gene expression analysis during autophagy

Alphaviruses are an emerging public health threat, causing epidemics of human infection across Europe, Africa, Asia, Australia, and America. Autophagy is an autonomous, tightly regulated, cellular process where cytoplasmic cargo and cellular debris are delivered to lysosomes for degradation in double membrane vesicles (autophagosomes). We showed Semliki Forest virus (SFV), a model for pathogenic alphaviruses, suppresses autophagy to enhance replication during the early stages of infection. There are no treatments for these alphaviruses, and autophagy has promise for therapeutic development. This project develops RT-qPCR as a plaque assay free method for virus quantification that will also facilitate gene expression analysis. We currently harvest virus and quantify via plaque assay, which is time-consuming (4 day duration) and does not allow analysis of gene expression. Removal of free RNA using RNAase A, and RNA extraction from virus samples will allow RT-qPCR quantification of virus and cDNA production for gene expression analysis. The RT-qPCR quantification method will be correlated with the plaque assay quantification method to produce a less expensive, more rapid assay that also permits analysis of virus gene expression. Students will need to organise their own accommodation and be expected to present their findings orally at a research day in York in September 2026.