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PurposeIncreased microbial respiration (Rm) usually decreases with warming time in incubation and field manipulative experiments, and substrate depletion and/or microbial thermal acclimation to warming is hypothesized to be responsible for this decrease. However, few studies have been conducted to determine the mechanism for this decrease, especially in alpine regions.Materials and methodsFirst, soils taken from an alpine meadow on the Tibetan Plateau were incubated at three different temperatures (i.e., 5, 15, and 25 degrees C) with two soil moistures (i.e., 30 and 60% water holding capacity (WHC)) for 58days, then another two experiments were conducted with incubation at high-low-high temperature (i.e., 25-15-25 degrees C) for 2weeks and glucose-induced respiration (GIR) for 4h.Results and discussionRm increased with soil temperature increase at 60% WHC, but there were no significant differences between incubation at 5 and 15 degrees C with 30% WHC. Higher Rm was found at 60% WHC than at 30% WHC only when incubated at 15 and 25 degrees C. Both the high-low-high temperature incubation and GIR experiments indicated that decreased soil substrate availability weakened the responses of Rm to high temperature at 60% WHC, and there was no microbial thermal acclimation to temperature over the incubation period. Temperature sensitivity of Rm (Q(10)) was not affected by soil moisture, high-low-high temperature incubation, or glucose addition.ConclusionsOur findings suggest that the increase in Rm at higher temperature disappeared over the incubation period, and that this could be due to depletion of soil substrate availability.