Groundwater table fluctuation during natural and anthropogenic processes can facilitate the interaction between oxygen (O2) from the unsaturated zone and ferrous iron (Fe2+) from the saturated zone. In light of previous findings that Fe(III)-reducing bacteria can be killed by the reactive oxidants produced from Fe2+ oxidation under static oxic conditions, we hypothesize that Fe(III)-reducing bacteria will be attenuated during groundwater table fluctuations. To test this hypothesis, this study explored the variations of cell numbers of Shewanella oneidensis strain MR-1 (MR-1), a typical strain of Fe(III)-reducing bacteria, together with dissolved oxygen (DO) and Fe2+, at different points during controlled groundwater table fluctuations in a sand column. The results showed that, during the rise of the water table, O2 in the pore air was entrapped by the deoxygenated groundwater, and Fe2+ in the groundwater was oxidized by the entrapped O2. In this process, 1.40-2.42 orders of magnitude of viable MR-1 cells were killed at different points in the column. Further investigation proposed that the death of MR-1 is caused by the production of intracellular reactive oxidants, such as O2•- and OH•, from the oxidation of adsorbed/absorbed Fe2+ instead of by bulk reactive oxidants, such as OH• and Fe(IV), produced from the oxidation of aqueous Fe2+. The findings here provide new insights for Fe biogeochemical cycling in the redox-dynamic zone.