Although understory vegetation is known to play an important role in driving the processes and functions of forest ecosystems, little is known about how understory vegetation affects the composition and function of soil microbial communities in forest ecosystems, especially in subtropical and tropical forests. This study used the experimental removal of understory fern ( Dicranopteris dichotoma ) to investigate the effect of the fern on substrate utilization patterns of culturable soil bacterial communities in two subtropical Eucalyptus plantations. One year after treatment, the removal of understory fern significantly increased soil temperature by 2–3<ce:hsp sp="0.25"/>°C and retarded litter decomposition by 5.6–23.1%. However, understory fern removal did not affect the substrate utilization pattern of soil bacterial communities. Our study provides evidence that, although understory fern removal significantly alters soil temperature and litter decomposition rate, the disturbance caused by understory removal one year after treatment is too weak to cause detectable changes in substrate utilization pattern of culturable soil bacterial communities in subtropical Eucalyptus plantations.