Recent Developments in Single-Virus Fusion Assay

Viral infection is a global health hazard. A crucial step in the infection cycle of enveloped viruses is the fusion of viral and host cellular membranes, which permits the transfer of the viral genome to the host cells. Membrane fusion is a ubiquitous process involved in sperm-egg fusion, exocytosis, vesicular trafficking, and viral entry to host cells. While different protein machineries catalyze the diverse fusion processes, the essential step, i.e., merging of two lipid bilayers against a kinetic energy barrier, is the same. Therefore, viral fusion machineries/pathways are not only the sites for antiviral drug development but also serve as model fusogens. Ensemble-based spectroscopic approaches or bulk fusion assays have yielded valuable insights regarding the fusion processes. However, due to the stochastic nature of the fusion events, ensemble-based assays do not permit synchronization of all the fusion events, and the molecular steps leading to fusion pore opening cannot be resolved entirely and correlated with the structural changes in viral fusion proteins. Several single-virus fusion assays have been developed to circumvent these issues. The review describes the recent advancements in single-virus/particle fusion assays using the Influenza virus as a paradigm.