The ability of a soil to reduce waxes that cause water repellency depends, in part, on the presence of populations of wax-degrading microorganisms. Therefore, estimating the size of populations of wax-degrading bacteria in water repellent soils is likely to be a useful indicator of the potential for bioremediation of water repellency. A Most Probable Number (MPN) method that selects specifically for the function of wax degradation in bacteria was developed and tested using both pure cultures and natural populations of wax-degrading bacteria. Two carbon sources (coconut oil and hexadecane) that contain fatty acids or hydrocarbons implicated as causative agents of water repellency were compared in terms of growth and emulsification by a range of known wax-degrading bacteria. Emulsification resulting from surfactant production, a direct measure of wax degradation, was the criterion for a positive test. All the bacteria tested emulsified coconut oil, which was chosen as the carbon source in subsequent testing of the MPN assay. Population estimates of pure cultures of wax-degrading bacteria (liquid suspensions and inoculated into sterile soils) made using the MPN and a plating technique showed good agreement between the 2 methods, indicating that the MPN method gives an accurate measure of size of the wax-degrading population. Extensive replication of MPN counts of known wax-degrading bacteria inoculated into sterile soils or populations in soils collected from a range of natural habitats showed little variation within samples, indicating that the new method was reproducible. Based on its accuracy and reproducibility the MPN method was deemed suitable for estimating populations in a range of soils. Monitoring wax-degrading populations under different agricultural managements and at different times of the year, i.e. seasonal dynamics, could provide clues for the development of new managements that minimise the risk of water repellency in agricultural soils.