This study aimed to: (1) determine whether soil microbial communities along a gradient from intensive (fertilized) to low-input (unfertilized) grassland management, shift in their composition as shown by an increase in the abundance of fungi relative to bacteria and (2) whether these shifts in soil microbial communities vary depending on season. At all sample dates soil microbial biomass-C and -N, and the total abundance of phospholipid fatty acids (PLFA) were highest in unfertilized, undrained treatments and lowest in fertilized and drained grassland. Similarly, microbial activity, measured as CO2-C respiration, was found to be at its lowest in the most intensively managed grassland. Measures of microbial biomass showed a high degree of seasonality, having summer maxima and winter minima. In contrast, PLFA measures had spring maxima and autumn minima. Seasonal and management differences were also observed within the microbial community. PLFA profiles revealed that most individual fatty acids were highest in the unfertilized treatments, and lowest in fertilized grassland. The fungal-to-bacterial biomass ratio was also highest in the unfertilized and lowest in the fertilized soils, suggesting that higher microbial biomass in former were more due to the growth of fungi than bacteria. As with total PLFA, the abundance of individual fatty acids showed a spring maximum and an autumn minimum. Seasonal differences in PLFA patterns were shown to be related to soil mineral-N and soil moisture contents. Factors controlling shifts in microbial community structure between sample dates and sites are discussed in relation to other studies. A critical assessment of the different measures of microbial biomass is also given. Overall, the findings of this study support the thesis that fungi play a more significant role in soil biological processes of low-input, unfertilized grasslands, than in intensively managed systems.