Effects of biocides and other metal removal fluid constituents on Mycobacterium immunogenum

Cells of Mycobacterium immunogenum are significantly more resistant to biocides and antimicrobial compounds used for disinfection of metal removal fluids (MRFs) than cells of the gram-negative bacterium Pseudomonas pseudoalcaligenes. To ensure accurate measurement of M. immunogenum susceptibility, a biocide inactivation step must be included to prevent an overestimation of killing for short-term exposures. Cell suspensions in 5% MRF for 30 min rather than direct plating following biocide exposure increased the killing of M. immunogenum cells, suggesting a heretofore undiscovered interaction between MRF components and biocides. Biocide killing was reduced at high M. immunogenum cell densities (>10(6)/ml). Among the chemicals tested, only Synergex Premier, Preventol CMK, and dicyclohexylamine killed greater than 90% of M. immunogenum cells in 6 h. M. immunogenum cells adhered to and grew rapidly on glass, copper, and galvanized surfaces, forming biofilms of 10(6) cells/cm(2) within 2 weeks and suggesting it is likely that the majority of M. immunogenum cells in MRF systems are on surfaces. M. immunogenum cells grown on surfaces in biofilms were three- to 100-fold more resistant to the biocides Synergex Premier and Preventol CMK than cells grown in suspension, compounding the problem of eradication of M. immunogenum cells from MRF systems.