Limitations While this study provides novel results on the viability of viruses in evaporating droplets of different compositions over a range of RHs, it does not examine how the surface material upon which viruses are deposited might affect viability. Previous studies have reported that the persistence of viruses in droplets depends on the type of material (e.g., plastic vs. steel) [74]. It is possible that material exchange between surfaces and droplets (e.g., dissolution of metal ions into droplets) leads to accumulation of surface materials in droplets and inactivates viruses. It will be interesting to investigate the effects of the interplay among surface materials, droplet composition, and environment on the survival of viruses in droplets. Additionally, we have focused on the biological inactivation of viruses in droplets but not on their physical behavior, which likely depends on physicochemical characteristics of the droplets. Future studies should be conducted in this area, although pinpointing viruses within droplets is challenging. Results might help explain the protective or inactivating effect of certain media components we observed in this study. Previous studies have demonstrated differences in the persistence of viruses on surfaces as a function of initial viral titer in the inoculum [75, 76]. Investigating the role of viral titer, which might affect aggregation and other characteristics, on virus survival is another interesting research question. Lastly, this study reported findings from two bacteriophage models, which may not fully represent human viruses. Human viruses, such as influenza virus and coronavirus, should be used in future studies to elucidate the effects of droplets’ chemical composition and RH on virus survival and transmission of viruses. To conclude, we demonstrated that both the chemical composition of droplets and RH strongly affect the viability of non-enveloped and enveloped viruses. The effects of sodium chloride and SDS varied by RH level and virus type. pH did not affect the viability of MS2 but effectively inactivated Φ6 in solutions at pHs of 4 and 10. BSA generally preserved the viability of MS2 and Φ6 in droplets. We also found that the viability of viruses in droplets of certain compositions was RH-dependent at most conditions. Our results reveal that two factors contribute to the inactivation of viruses in droplets: (1) droplet evaporation kinetics, which are controlled by RH; and (2) inactivation or protective effects induced by chemicals. Additionally, the physical behavior of viruses, such as forming aggregates and partitioning to the air-liquid interface, resulting from changes in droplets’ characteristics may also affect inactivation. Results from our study are meaningful in predicting the persistence of viruses in droplets of various compositions in the environment and infectious disease transmission.