Although many studies have reported the negative effects of elevated O on plant physiological characteristics, the influence of elevated O on below-ground processes and soil microbial functioning is less studied. In this study, we examined the effects of elevated O on soil properties, soil microbial biomass, as well as microbial community composition using high-throughput sequencing. Throughout one growing season, one-year old seedlings of two important endemic trees in subtropical China: Taxus chinensis (Pilger) Rehd. var. chinensis, and Machilus ichangensis Rehd. Et Wils, were exposed to charcoal-filtered air (CF as control), 100 nl l (E100) or 150 nl l (E150) O -enriched air, in open top chambers (OTCs). We found that only higher O exposure (E150) significantly decreased soil microbial biomass carbon and nitrogen in M. ichangensis, and the contents of organic matter were significantly decreased by E150 in both tree species. Although both levels of O exposure decreased NO -N in T. chinensis, only E150 increased NO -N in M. ichangensis, and there were no effects of O on NH -N. Moreover, elevated O elicited changes in soil microbial community structure and decreased fungal diversity in both M. ichangensis and T. chinensis. However, even though O exposure reduced bacterial diversity in M. ichangensis, no effect of O exposure on bacterial diversity was detected in soil grown with T. chinensis. Our results showed that elevated O altered the abundance of bacteria and fungi in general, and in particular reduced nitrifiers and increased the relative abundance of some fungal taxa capable of denitrification, which may stimulate N O emissions. Overall, our findings indicate that elevated O not only impacts the soil microbial community structure, but may also exert an influence on the functioning of microbial communities.