title: Rates and mechanisms of bacterial mutagenesis from maximum-depth sequencing.
abstract: In 1943, Luria and Delbrück used a phage-resistance assay to establish spontaneous mutation as a driving force of microbial diversity. Mutation rates are still studied using such assays, but these can only be used to examine the small minority of mutations conferring survival in a particular condition. Newer approaches, such as long-term evolution followed by whole-genome sequencing, may be skewed by mutational ‘hot’ or ‘cold’ spots. Both approaches are affected by numerous caveats. Here we devise a method, maximum-depth sequencing (MDS), to detect extremely rare variants in a population of cells through error-corrected, high-throughput sequencing. We directly measure locus-specific mutation rates in Escherichia coli and show that they vary across the genome by at least an order of magnitude. Our data suggest that certain types of nucleotide misincorporation occur 10(4)-fold more frequently than the basal rate of mutations, but are repaired in vivo. Our data also suggest specific mechanisms of antibiotic-induced mutagenesis, including downregulation of mismatch repair via oxidative stress, transcription–replication conflicts, and, in the case of fluoroquinolones, direct damage to DNA.
authors.forename: Justin
authors.initials: J
authors.lastname: Jee
authors.forename: Aviram
authors.initials: A
authors.lastname: Rasouly
authors.forename: Ilya
authors.initials: I
authors.lastname: Shamovsky
authors.forename: Yonatan
authors.initials: Y
authors.lastname: Akivis
authors.forename: Susan R
authors.initials: SR
authors.lastname: Steinman
authors.forename: Bud
authors.initials: B
authors.lastname: Mishra
authors.forename: Evgeny
authors.initials: E
authors.lastname: Nudler
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D004249:DNA Damage / Q000235:genetics; D053843:DNA Mismatch Repair / Q000187:drug effects / Q000235:genetics; D004261:DNA Replication / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D019143:Evolution, Molecular; D024841:Fluoroquinolones / Q000494:pharmacology; D056426:Genetic Loci / Q000187:drug effects / Q000235:genetics; D014644:Genetic Variation / Q000187:drug effects / Q000235:genetics*; D016680:Genome, Bacterial / Q000187:drug effects / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D054643:INDEL Mutation / Q000235:genetics; D016296:Mutagenesis / Q000187:drug effects / Q000235:genetics*; D059645:Mutation Rate; D009711:Nucleotides / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D024841:Fluoroquinolones; D009711:Nucleotides