Hydrophobicity is a vital parameter for initial cell adhesion that ultimately leads to biofouling of surfaces and loss of system performance and health issues. The efficiency of a number of biological systems could be improved by increasing the hydrophobicity of concerned bacteria. Here we used ammonium sulfate (salt) to enhance the bacterial hydrophobicity, as measured by a commonly used liquid-liquid partitioning based hydrophobicity assessment assay - the MATH test. We observed successive increases in bacterial hydrophobicity with incremental increase in salt concentration for Gram-negative bacteria. Upon addition of 2 M salt, three closely related E. coli strains were easily distinguishable from one another. Gram-positive bacteria exhibited different trends than Gram-negative strains, with no change in the hydrophobicity of S. salivarius HB cells and a sharp decline followed by an increase in hydrophobicity for D. radiodurans. Cell size measurements revealed that Gram-positive cells exhibited a change in cell size on hydrocarbon exposure, while the Gram-negative cultures remained mostly unaffected. Overall, salt addition was observed to enhance the hydrophobicity of different test strains, especially at the higher concentrations used here of 1.5 and 2 M. Salt addition in conjunction with the MATH test successfully differentiated and quantified otherwise weakly hydrophobic bacteria, thus enhancing the range of this laboratory assay. Our results demonstrate the effectiveness of salt addition in increasing the bacterial hydrophobicity, which could potentially be used in diverse areas, ranging from applied microbiology and engineering to oral care. (C) Koninklijke Brill NV, Leiden, 2011