LeClerc J E, Payne W L, Kupchella E, Cebula T A
Molecular Biology Branch (HFS-237), Center for Food Safety and Applied Nutrition, US Food and Drug Administration, 200 C St. S.W., Washington, DC 20204, USA.
Mutat Res. 1998 May 25;400(1-2):89-97. doi: 10.1016/s0027-5107(98)00069-4.
Defects in the methyl-directed mismatch repair lead to both the hypermutability phenotype and removal of a barrier to genetic exchange between species. Mutator bacteria carrying such defects occur frequently among bacterial pathogens, suggesting that subpopulations of mutators are contained within pathogen clones and give rise to the genetic variants that are acted upon by selective forces to allow survival or successful infection. We report here on the detection of the mutator subpopulation in Salmonella typhimurium and determination of its frequency in laboratory cultures. The analysis involved screening for mutators among revertants of S. typhimurium histidine auxotrophs selected for the His+ phenotype, since the frequency of mutators is expected to be increased in the selected mutant population they helped to spawn. The increases in spontaneous reversion of histidine mutations were first measured in isogenic strains carrying mismatch repair-defective mutH, mutL, mutS, or uvrD alleles, relative to their mismatch repair-proficient counterparts. Screening for the mutator phenotype in nearly 12,000 revertants of repair-proficient strains carrying his mutations highly stimulated for reversion in mutator backgrounds, the base substitution in 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 changes expected in such mutators, as assessed by comparison with the spectra for reversion events in wild-type and mismatch correction-defective backgrounds. The results show that subpopulations of mutators, residing in normal populations at a finite frequency, can be culled from the culture by strong selection for a required phenotype. We calculate that the frequency of mutators in the unselected population of S. typhimurium is 1-4x10-6, an incidence 10-fold lower than that expected based on studies of laboratory cultures of Escherichia coli.
甲基导向错配修复缺陷会导致超突变表型以及消除物种间基因交换的障碍。携带此类缺陷的突变细菌在细菌病原体中频繁出现，这表明突变细菌亚群存在于病原体克隆中，并产生遗传变异，这些变异会受到选择压力的作用，以实现生存或成功感染。我们在此报告鼠伤寒沙门氏菌中突变细菌亚群的检测及其在实验室培养物中的频率测定。该分析涉及在为获得His+表型而选择的鼠伤寒沙门氏菌组氨酸营养缺陷型回复突变体中筛选突变细菌，因为预计在它们所产生的选定突变群体中，突变细菌的频率会增加。首先在携带错配修复缺陷的mutH、mutL、mutS或uvrD等位基因的同基因菌株中，相对于错配修复功能正常的对应菌株，测量组氨酸突变的自发回复率。在携带his突变且在突变背景中高度刺激回复的修复功能正常菌株的近12,000个回复突变体中筛选突变体表型，hisG428中的碱基替换和hisC3076中的移码突变，产生了5个突变菌株（0.04%）。通过与野生型和错配校正缺陷背景中的回复事件谱进行比较评估，新出现的突变菌株中包含的his+回复突变具有此类突变体预期的主要核苷酸变化特征。结果表明，以有限频率存在于正常群体中的突变细菌亚群，可以通过对所需表型的强烈选择从培养物中筛选出来。我们计算出未选择的鼠伤寒沙门氏菌群体中突变细菌的频率为1-4×10-⁶，这一发生率比基于大肠杆菌实验室培养研究预期的低10倍。
