Establishment of improved perennial pasture systems for livestock production has been demonstrated to improve soil health in the semiarid Texas Southern High Plains (SHP). Use of waste products from other components of the animal production chain, such as applying composted manure from feedlot operations to grazed pastures, may provide an additional means of improving soil health and sustainability of these systems. However, few studies have examined the impact of composted animal manure on the changes in soil microbial communities, although they are important indicators of soil health. We investigated the effects of a one-time composted cattle manure application at 3.36 Mg ha−1 on soil microbial community size and structure, soil organic C (SOC), total N (TN), and soil organic matter (SOM) up to 1.5 yr after application. We compared pastures consisting of either WW-B. Dahl Old World bluestem grass [Bothriochloa bladhii (Retz.) S.T. Blake] alone or in mix with alfalfa (Medicago sativa L.). Results showed that compost application significantly increased SOC by about 4.4 g kg−1 soil within 1.5 yr after application. In N-fertilized grass-only pastures, compost addition increased microbial phospholipid fatty acid abundance, SOC, TN, and SOM, but no significant differences were observed between compost treatments in unfertilized pastures containing legumes. Our results show that even a one-time compost addition at a relatively low rate can augment SOC in this semiarid pasture system after 1.5 yr, and that stimulation of soil microbial abundance and soil C and N from compost is more pronounced in fertilized grass-only pastures compared with those with legume establishment.