Among karst mountain agricultural fertilization measures, the partial replacement of chemical fertilizers with organic fertilizers is important for protecting its vulnerable mountain environment and developing ecologically-friendly agriculture. In this study, we investigated the effects of short-term organic fertilizer application on the bacterial diversity of maize soil in karst areas and the potential of using organic fertilizer as a partial substitute for chemical fertilizers. Two maize fields with different parent materials in a karst region were selected for a short-term field control experiment using chemical fertilizer and organic fertilizer treatment, combined with the high-throughput sequencing method of 16S rDNA gene amplicons. (i) The soil physicochemical properties and bacterial diversity of different parent material soil are different, but the main dominant bacterial types are similar. (ii) Short-term organic fertilizer treatment, rather than chemical fertilizer treatment, increased the bacterial richness significantly, especially for some functional bacteria (such asNitrospira,Gemmatimonas). (iii) Analysis of the correlation between environmental factors and bacterial diversity indicated that soil pH and total P had the most significant effects on bacterial community structure (r = 0.91,p = 0.001;r = 0.33,p = 0.001). This study showed that it is an effective method to maintain a richer bacterial community and increasing the abundance of some functional bacteria by increasing organic fertilizers and reducing chemical fertilizers in the farm soil in karst regions, which could also be applied to other fragile agricultural ecosystems in the world.