The present study was carried out to investigate the influence of crop rotation and fertilization on the number of different groups of microorganisms. Soil samples were taken from the following treatments: 1. Monoculture of maize, 2. Maize-wheat (without fertilization), 3. Maize-wheat (with fertilization), 4. Maize-soybean-wheat (with fertilization), 5. Pasture soil, 6. Forest soil under oak, from three depths: 0-10 cm, 10-20 cm, and 20-30 cm in May and October 2012. Microbiological analyses included the determination of total number of bacteria, number of fungi, actinomycetes, aminoheterotrophs, and azotobacter. The number of investigated groups of microorganisms was higher in May on average. Fertilization did not affect the number of investigated groups of microorganisms. Crop rotation positively affected the total number of bacteria and the number of aminoheterotrophs in soil under maize.