The assemblage of bacteria and fungi from black (Solenopsis richteri)/hybrid imported fire ant (BIFA/HIFA) mounds were obtained from four counties in northeast Mississippi. These locations were selected due to high concentrations of BIFA/HIFA that were free from red imported fire ants (RIFA). Mound samples were obtained during October, November, and December in 2003 and January 2004. Patterns of species composition and diversity (species richness) were evaluated from mound soil, mound plant debris, and ant bodies. A total of 5742 isolates consisting of 58 bacterial and 35 fungal taxa were obtained. The most common bacteria identified included Chryseobacterium indolegenes, Stenotrophomonas maltophilia, Actinomadura yumaensis, and Arcanobacterium haemolyticum. Approximately 66% of the fungi cultured belonged to the artificial assemblage Fungi Imperfecti, including Curvularia geniculata, Penicillium spp., Nigrospora sphaerica, and Monoacrosporium leptosporium. The insect pathogen Beauveria bassiana was obtained from mound soil, mound plant debris, and ant bodies, with the greatest percentage from ant bodies. Species richness for bacteria and fungi were both highest from mound soil at 53 and 30 taxa, respectively, and lowest, with 8 bacteria and 25 fungal taxa, from mound plant debris. Species diversity for bacteria was also highest from mound soil, and highest for fungi from ant bodies than the other two isolation conditions. Evenness values for bacteria (0.72–0.80) and fungi (0.74–0.77) during each sampling date had moderate to high relative abundance (1.0 = highest level possible), indicating similarity of taxa among bacteria and among fungi from the four sampling dates. Coefficient of community values comparing sampling dates for bacteria and fungi were greatest between the first and last sampling date (October and January). Temperatures during those dates ranged from 14.4 °C to 28.9 °C in October and −2.8 °C to 10.0 °C in January. As a continuation of this research, cultures of the different bacteria and fungi obtained in this study are currently being evaluated for their potential as biological control agents of BIFA/HIFA and RIFA that occur in Mississippi.