Most studies on the effects of biochar and fertilizer on soil carbon (C) and nitrogen (N) mineralization, and microbial C and N content, are restricted to a single soil type, limiting our understanding of the interactions between these factors and microbial functions. To address this paucity in knowledge, we undertook a 3-year experiment using four contrasting soils to assess the role of peanut shell biochar and fertilizer on C and N mineralization, microbial C and N, and N stoichiometry. Across all four soils, biochar significantly (P < 0.05) increased soil carbon mineralization (Cmin) and nitrogen mineralization (Nmin) over three years compared to fertilizer and the control. Biochar also increased total C (Csoil) across the four soils in year 1, with the Fluvisol recording greater total C in year 2 and Phaeozem having greater total C in year 3. Biochar resulted in a higher microbial biomass C (Cmic), total N (Nsoil) and microbial biomass N (Nmic); the degree of change was closely related to Csoil and Nsoil. There was a positive correlation between Cmic:Nmic and Csoil:Nsoil; while Csoil and Cmic increased following amendment with biochar, which reduced the soil C and N stoichiometric imbalance (Nimb) caused by the increase in the C to N ratio. However, fertilizer exacerbated the imbalance of soil C and N stoichiometry. Fertilizer also reduced the Csoil:Nsoil and Cmic:Nmic ratios. Soil pH had a positive correlation with Csoil, Cmic, Nmic, Cmin, Nmin, Csoil:Nsoil, Cmic:Nmic, and biochar increases this correlation. The soil pH was negatively correlated with Cimb:Nimb and Nsoil. Fertilizer was positively correlated Cimb:Nimb and Nsoil. In contrast, fertilizer N application lowered microbial biomass C:N. We conclude that biochar reduces the imbalance of soil C and N stoichiometry, whereas fertilizer increased this imbalance. Biochar had a greater impact on C and N in soils with a lower pH.