USE OF ANTIBIOTICS TO SELECT AUXOTROPHIC MUTANTS OF NEISSERIA-MENINGITIDIS

Antibiotic selection of auxotrophs has been a powerful tool in the elucidation of bacterial meta belie pathways, but it has been difficult to adapt th is method to Neisseria spp. We describe a procedure by which a population of mutagenized N. meningitidis is enriched for mutants with specific growth phenotypes. These experiments used a simple defined medium (modified from that described in J Bacteriol 1962; 83: 470-4) in which meningococci grow well on a variety of carbon sources. Nitrosoguanidine-treated meningococci were incubated with an antibiotic (cefotaxime, streptomycin or nalidixic acid) in a defined medium that was nonpermissive for the desired phenotype. The survivors were grown for several generations in a permissive defined medium to reduce the proportion of mutants with phenotypes other than that desired, then subjected to a second antibiotic treatment in nonpermissive medium. Survivors of the second antibiotic treatment were plated, and colonies were screened to identify auxotrophs. This procedure has allowed the isolation of meningococcal mutants with amino acid or vitamin requirements or with altered utilization of carbon sources.