Increasing organic matter input and phosphorus fertilization are employed extensively to restore degraded grasslands. Nevertheless, little is known about their effects on microbes, especially on active microbial populations. Therefore, this study is aimed at examining the short-term influences of litter and phosphorus addition on microbes in degraded grassland soils. Materials and methods: A microcosm experiment was established using soils sampled from a heavily degraded Tibetan alpine meadow. The experiment used a two-way factorial design with grass litter and phosphorus addition as the main factors. Microbial abundance and rDNA transcriptional activity were assessed through quantitative PCR. Total and active microbial community profiles were measured using DNA- and RNA-based MiSeq sequencing, respectively. Results and discussion: As shown in this study, litter addition significantly increased microbial rDNA transcriptional activity and fungal abundance, but it decreased microbial α-diversity. However, prokaryote abundance was unaffected by the litter addition. Total and active soil microbial community profiles and interaction patterns were also significantly altered by litter addition. The relative abundance of copiotrophic and oligotrophic microbial lineages significantly increased and decreased, respectively, in the soils with litter addition. Functional predictions suggested that litter addition might significantly increase the abundance of pathogens, as well as microbes related to nitrogen fixation, denitrification, and chitinolysis, while decreasing nitrifier abundance. In contrast, no significant effects of the phosphorus addition on soil microbes were observed. Conclusions: These findings highlight the significant effects of increasing litter input on total and active soil microbial communities and suggest that microbial responses should be considered when restoring degraded grasslands by increasing organic matter input.