The susceptibility of waterborne viruses to different inactivating treatments is acknowledged to vary between viruses and even between closely related strains, yet the extent of this variation, or the underlying mechanisms, are not known. Here, different enteroviruses (six strains of coxsackievirus B5 (CVB5), two strains of coxsackievirus B4 (CVB4) and one strain of coxackievirus B1 (CVB1)) were isolated from wastewater. The different viruses were then exposed to disinfectants used in water and wastewater treatment (UV254, free chlorine (FC), chlorine dioxide (ClO2)) and to stressors encountered in the environment (sunlight, temperature). Inactivation kinetics of the environmental isolates were compared with those of laboratory enterovirus strains (CVB5 Faulkner and echovirus 11 Gregory) and MS2 bacteriophage. FC exhibited the greatest variability in inactivation kinetics between different strains, whereas inactivation by UV254 differed only subtly. The variability in inactivation kinetics was greater between serotypes than it was among the seven strains of the CVB5 serotype. MS2 was a conservative surrogate of enterovirus inactivation by UV254, sunlight or heat, but frequently underestimated the disinfection requirements for FC and ClO2. To assess the mechanisms underlying the differing susceptibilities of these viruses to inactivation, we focused on thermal inactivation. Specifically, we extensively analyzed the inactivation of these viruses at 30 and 55°C, and under different conditions of pH and NaCl concentrations. At 30°C, inactivation at neutral pH was slow, but both acidic and alkaline pH enhanced inactivation, and the addition of 1 M NaCl exerted a synergistic inactivating effect. These findings are consistent with RNA cleavage being the main mechanism of inactivation, and genome degradation was experimentally confirmed. At 55°C, salt had a protective effect on all viruses. This was rationalized by calculations of the different protein interaction forces, which demonstrated that increasing concentrations of salt resulted in increasing attractive forces at the capsid pentamer interfaces. At this temperature, major differences in thermoresistance between the viruses were observed, with CVB4 and E11 displaying the lowest thermoresistance, and the CVB5 laboratory strain being less thermoresistant than the CVB5 isolates. These differences could not be explained by a shift in capsid pentamer interaction forces, but likely resulted from mutations located in VP1 pocket region. The importance of the VP1 pocket region was further confirmed by adapting CVB5 to two different temperatures (50 and 55 °C). The thermo-adapted strains exhibited a competitive fitness trade-off compared to control strains, but were significantly more resistant to thermal inactivation. This resistance coincided with the appearance of one or several of four mutations in the VP1 region of the structural proteins. These mutations did not affect the interaction forces at the pentamer interface. Instead, they were located in the VP1 pocket region, confirming the importance of this region in the acquisition of thermotolerance. Overall, these data indicate that the thermostability of a virus can be enhanced by external (matrix) factors, in particular salinity, or by intrinsic (structural) modifications in the VP1 pocket region.