Microbial dynamics on commercially grown white button mushrooms is of importance in terms of food safety assurance and quality control. The purpose of this study was to establish the microbial profile of fresh white button mushrooms. The total microbial load was determined through standard viable counts. Presence and isolation of Gram-negative bacteria including coagulase-positive Staphylococci were performed using a selective enrichment approach. Dominant and presumptive organisms were confirmed using molecular methods. Total mushroom microbial counts ranged from 5·2 to 12·4 log CFU per g, with the genus Pseudomonas being most frequently isolated (45·37% of all isolations). In total, 91 different microbial species were isolated and identified using Matrix-assisted laser desorption ionization-time of flight mass spectrophotometry, PCR and sequencing. Considering current food safety guidelines in South Africa for ready-to-eat fresh produce, coliform counts exceeded the guidance specifications for fresh fruit and vegetables. Based on our research and similar studies, it is proposed that specifications for microbial loads on fresh, healthy mushrooms reflect a more natural microbiome at the point-of-harvest and point-of-sale.Presence and persistence of micro-organisms within the microbiome of fresh produce is important when identifying a potential niche for foodborne pathogens. Most foodborne outbreaks can be attributed to microbial imbalances or lack of diversity within the associated host surface and residing microbial population. Agaricus bisporus samples analysed during this study showed a higher microbial load (5·2 up to 12·4 log CFU per g) compared to known values for other fresh produce. These mushrooms were considered to carry microbial loads representing a healthy and safe product, fit for consumption, despite showing a high indicator incidence. Although foodborne pathogens may be associated on occasion with fresh mushrooms, it remains a low-risk commodity; therefore, this study provides insight and experimental evidence identifying microbial population dynamics of fresh and packed mushrooms.