DETECTION OF MUTATOR SUBPOPULATIONS IN SALMONELLA-TYPHIMURIUM LT2 BY REVERSION OF HIS ALLELES

Defects in the methyl-directed mismatch repair lead to both the hyperm utability phenotype and removal of a barrier to genetic exchange betwe en species. Mutator bacteria carrying such defects occur frequently am ong bacterial pathogens, suggesting that subpopulations of mutators ar e contained within pathogen clones and give rise to the genetic varian ts that are acted upon by selective forces to allow survival or succes sful infection. We report here on the detection of the mutator subpopu lation in Salmonella typhimurium and determination of its frequency in laboratory cultures. The analysis involved screening for mutators amo ng revertants of S. typhimurium histidine auxotrophs selected for the His(+) phenotype, since the frequency of mutators is expected to be in creased in the selected mutant population they helped to spawn. The in creases in spontaneous reversion of histidine mutations were first mea sured in isogenic strains carrying mismatch repair-defective mutH, mut L, mutS, or uvrD alleles, relative to their mismatch repair-proficient counterparts. Screening for the mutator phenotype in nearly 12,000 re vertants of repair-proficient strains carrying his mutations highly st imulated for reversion in mutator backgrounds, the base substitution i n hisG428 and frameshift in hisC3076, yielded five mutator strains (0. 04%). The his(+) reversion mutations contained within the newly-arisen mutator strains were characteristic of the predominant nucleotide cha nges expected in such mutators, as assessed by comparison with the spe ctra for reversion events in wild-type and mismatch correction-defecti ve backgrounds. The results show that subpopulations of mutators, resi ding in normal populations at a finite frequency, can be culled from t he culture by strong selection for a required phenotype. We calculate that the frequency of mutators in the unselected population of S. typh imurium is 1-4 x 10(-6), an incidence 10-fold lower than that expected based on studies of laboratory cultures of Escherichia coli. (C) 1998 Elsevier Science B.V. All rights reserved.