Inactivation of collected viral aerosols is important for preventing a filter medium’s serving as a fomite. The focus of this study was to evaluate the inactivation efficiency (IE) achieved through filtration coupled with microwave irradiation. MS2 aerosolized through a Collison nebulizer was fed into the system and collected onto the filter. For in-flight microwave decontamination, microwave irradiation was applied to an HVAC (heating, ventilation and air conditioning) filter supported on a SiC disk for three cycles of selected irradiation times per 10 min (i.e., 1, 2.5, 5, and 10 min/10 min) at power levels ranging from 125 W to 375 W. The survival fraction (SF) on the substrate and the IE through the entire system were investigated to determine the efficacy of this approach. SF decreased and IE increased as microwave power level was increased (p = 0.02 and p < 0.01, respectively) or the application time was extended (p = 0.03 and p < 0.01, respectively). Both measures changed sharply above a threshold temperature of around 90°C and reached 2 logs at 116 and 109°C, respectively. The log SF and IE of –2.59 and 3.62, respectively, were observed when the operating condition of 375 W for 10 min/cycle was used and the SiC disk facilitated microwave absorption. When a quartz frit was used as a support instead of the SiC disk, log inactivation efficiencies of 0.8, 1.0, and 1.3 were measured at relative humidities of 30%, 60% and 90%, respectively, under the same irradiation conditions. Relative humidity is a significant parameter from 50–80°C (p = 0.01). The results demonstrate that microwave-assisted filtration systems can be used as an effective means for inactivating viruses.