Mutagenesis of Strains of Propionibacterium to Produce Cold-Sensitive Mutants

A mutagenesis procedure that involved exposures to N-metbyl-N '-nitro-N-nitrosoguanidine followed by penicillin counterselection was developed to obtain cold-sensitive mutants of strains of Propionibacterium commonly used for production of Swiss cheese. Optimum conditions for mutagenesis were exposure to 150 gg/ml N-methyl-N'-nitro-N-nitrosoguanidine for 30 min at 32C to kill approximately 99.0% of the cells, followed by exposure to 2000/ag/ml penicillin G at 14C for 48 h to select for those organisms unable to grow at low temperature. Seven cold-sensitive mutants were found that grew at rates similar to that of the parent at 32C but grew much more slowly than the parent 14C. All mutants behaved identically to the parent strain in tests to determine sugar fermentations and nitrate reduction. Three of the cold-sensitive mutants differed from the parent strain in sensitivity to streptomycin, which suggests they may contain altered ribosomes.