Use of antibiotics to select auxotrophic mutants of Neisseria meningitidis

• Published in: Microbial pathogenesis 1995-04, Vol.18 (4), p.289-296
• Main Authors: Erwin, Alice L., Gill, Martin J., Gotschlich, Emil C.
• Format: Article
• Language: English
• Subjects: Anti-Bacterial Agents - pharmacology; antibiotic selection; auxotroph; Bacteriological methods and techniques used in bacteriology; Bacteriology; Biological and medical sciences; Cell Division - drug effects; Fundamental and applied biological sciences. Psychology; Microbiology; mutagenesis; Mutation; Neisseria meningitidis; Neisseria meningitidis - drug effects; Neisseria meningitidis - genetics; Neisseria meningitidis - isolation & purification
• ISSN: 0882-4010; 1096-1208
• DOI: 10.1016/S0882-4010(05)80005-1

Antibiotic selection of auxotrophs has been a powerful tool in the elucidation of bacterial metabolic pathways, but it has been difficultto adaptthis method to Neisseriaspp. 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: 470–474) 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.