The interaction between soil property and soil microbial community in karst area still remains an open question. The characteristics of soil physicochemical properties and microbial community structure and their relationship under five vegetation succession stages (grassland, shrub land, secondary forest, plantation forest, and natural forest) at two soil depths (0-10 cm and 10-20 cm) were explored in a karst mountain ecosystem. We found that soil moisture content (SMC) and pH increased with soil depth across vegetation succession. The highest content of soil nutrients was found in the natural forest stage at both soil depths. The total PLFAs, the abundance of Gram-positive (GP) bacteria, actinomycetes (ACT), fungi, and arbuscular mycorrhizal fungi (AMF) were significantly (P < 0.05) related to variations with soil total carbon (TC) and total nitrogen (TN). Furthermore, the distribution of soil microbial community distinctly differed in vegetation succession both at two soil layers which was demonstrated by Principal-coordinates analysis. Redundancy analyses patterns indicated that soil TC and TN were positively related to cy19:0 and 10Me 16:0, but an opposite relationship with a15:0. Changes of soil microbial communities were significantly determined by vegetation succession, and soil microbial community structure can be a sensitive indicator to reflect the stabilization of karst mountain ecosystem, southwest of China.