Assessing microbial health risks related to floodwater is a complex undertaking, hindered by factors like the intricate nature of the microorganisms involved and the susceptibility of those exposed to them. This research contributes a framework and application that combines health risk assessment, disease burden calculation, and hydrodynamic modeling to estimate adverse health consequences of microbial pathogens in floodwater through traffic activities which is a common factor of exposure during floods. The case study is Ninh Kieu District (Can Tho City, Vietnam) located on the western side of Hau River, a Mekong tributary. The study focuses on the health risk and disease burden due to rotavirus A in floodwater in Ninh Kieu through traffic activities, especially for motorcyclist. This research is one of the first to consider the input parameter concentrations and the number of exposed people to reduce the health impact of flood risk. It reveals that mitigation measures should not only focus on reducing urban floods but also on raising awareness of the local people of microbial health risks in floodwater. The disease burden is considered the prime variable of the health indicator to represent the social dimension in the assessment of flood vulnerability.