Among environmental factors affecting life - history traits of birds breeding in nest boxes, an influence of microbial communities is relatively poorly understood. In this study, nest boxes used for breeding by great tit (Parus major) and blue tit (Cyanistes caeruleus) were sampled before the start of the breeding season to assess the bacterial loads of the nest box. Samples from the entrance hole and the interior of the nest box were taken at two different study sites: an urban parkland and a natural forest. Nest boxes were sampled to check if their bacterial loads differed between habitats. The second objective of this study was to check whether the occupancy of the nest boxes during the previous season would influence the bacterial load of the nest box. To verify this prediction, two categories of nest boxes were sampled at both study sites: nest boxes occupied by any of the two tit species in the previous season for breeding and nest boxes that had remained empty that year. The bacterial load of the nest box was significantly higher in the forest study area in both the occupied and unoccupied nest boxes. The nest boxes used for breeding in the previous season had significantly higher bacterial loads, but only in the forest area. Our results suggest that the bacterial load of the nest box can vary between habitats and may be positively related to the presence of the nests in the previous breeding season.