The objective of this study was to investigate Escherichia coli infiltration in saturated soil columns, at a constant temperature of 35°C and compare three different E. coli detection methods. Four soil columns were prepared with loamy clay soil using the wet packing method. All columns consisted of a 10-cm soil column with a water head of 10 cm above the soil surface. Three columns were inoculated with autoclaved ultrapure water containing 1.74 × 10⁷ colony-forming units per mL of E. coli K12. The fourth column was used as control and therefore only autoclaved ultrapure water was used as input. The infiltration experiment in all columns lasted for 278 h after E. coli inoculation. The three E. coli inoculated columns showed similar behavior, with an initial decrease of the infiltration rate during the first 10–15 flushed pore volumes of water containing E. coli. This decrease rate was attributed to the clogging of the soil by the E. coli. However, after reaching the minimum infiltration rate, it was followed by recovery of up to 97% of the initial infiltration rate. A recovery of the infiltration rate was not observed in the control soil column. These results are similar to the results of a previous experiment conducted at a constant temperature of 25°C. Additionally, the E. coli output from each soil column was measured by three methods: spectrometry, plate counting, and quantitative polymerase chain reaction. Additional supplementary experiments were conducted to assess the effects of temperature on the performance of each method. Due to the increased temperature, bacteria decay subsequently led to the inability to detect E. coli by plate counting and qPCR methods. From the results of this research, spectrometry was judged as the most suitable for measuring E. coli in the experiments.