In tests with Hele–Shaw cells containing silica sand, O2 profiles were measured after formation of a capillary fringe (CF) with a nutrient broth, that was inoculated with Pseudomonas putida. After 7 d, the CF became anaerobic up to 2 cm above the water level. Within the next 3 cm at decreasing water saturation, the O2 concentration approached its maximum value. After growth, most P. putida cells were found at the 4- to 5-cm height of the CF, whereas cell densities decreased significantly toward the top of the CF due to water-limited respiratory activity. This was shown in separate batch experiments by measuring respiration rates of P. putida for defined water saturations. In silica sand (rough surface) or glass beads (smooth surface), respiration of P. putida was significantly reduced at a water saturation of ≤7.5% but was always higher for growth on sand than on glass beads, presumably due to different water bioavailability at the surface. The highest O2 consumption rates of P. putida in both media were measured at 60 to 85% water saturation, which represents the saturated–unsaturated interface region in a CF. To find the minimum required water availability for respiration, batch assays with increasing agar concentrations in LB medium were conducted. The O2 consumption rates of P. putida decreased to zero above an agar concentration of 18% (w/v), equivalent to a water activity <0.978. The results indicate that limited bacterial respiration due to water potential stress would only occur in the top few millimeters of a CF in sand.