Abstarct: Abstract Edible mushrooms, including button mushrooms, are considered an important part of the diet in many countries due to their nutritional value. Button mushrooms require a casing laxyer with specific physical, chemical, and microbial properties to transition from the vegetative phase to the reproductive phase and form the pinhead structure. This phase transition is particularly facilitated by a reduction in temperature and carbon dioxide concentration in the presence of bacteria found in the casing soil. The growing substrate for button mushrooms is rich in active microorganisms, which have both positive and negative effects on mushroom growth and yield. Bacteria are active microorganisms that influence the growth of button mushrooms through their activities. Therefore, this study investigates the bacteria in the substrate of white button mushrooms to identify the bacterial species present. The initial phase of the experiments showed that the isolated bacteria grow well at ambient temperature and are either aerobic or facultative aerobic. Staining and light microscopy results indicated that the bacteria are rod-shaped, gram-positive, and spore-forming. Scanning electron microscopy (FESEM) analysis revealed that these bacteria appear as single rods or in short chains. In the final phase, DNA extraction and 16S region sequencing largely confirmed the presence of Bacillus cereus strengthening the findings of previous experiments.