Epistatic Roles for Pseudomonas aeruginosa MutS and DinB (DNA Pol IV) in Coping with Reactive Oxygen Species-Induced DNA Damage

Abstract

Pseudomonas aeruginosa is especially adept at colonizing the airways of individuals afflicted with the autosomal recessive disease cystic fibrosis (CF). CF patients suffer from chronic airway inflammation, which contributes to lung deterioration. Once established in the airways, P. aeruginosa continuously adapts to the changing environment, in part through acquisition of beneficial mutations via a process termed pathoadaptation. MutS and DinB are proposed to play opposing roles in P. aeruginosa pathoadaptation: MutS acts in replication-coupled mismatch repair, which acts to limit spontaneous mutations; in contrast, DinB (DNA polymerase IV) catalyzes error-prone bypass of DNA lesions, contributing to mutations. As part of an ongoing effort to understand mechanisms underlying P. aeruginosa pathoadaptation, we characterized hydrogen peroxide (H2O2)-induced phenotypes of isogenic P. aeruginosa strains bearing different combinations of mutS and dinB alleles. Our results demonstrate an unexpected epistatic relationship between mutS and dinB with respect to H2O2-induced cell killing involving error-prone repair and/or tolerance of oxidized DNA lesions. In striking contrast to these error-prone roles, both MutS and DinB played largely accurate roles in coping with DNA lesions induced by ultraviolet light, mitomycin C, or 4-nitroquinilone 1-oxide. Models discussing roles for MutS and DinB functionality in DNA damage-induced mutagenesis, particularly during CF airway colonization and subsequent P. aeruginosa pathoadaptation are discussed.

Pharmacokinetics and Pharmacodynamics of Aerosolized Antibacterial Agents in Chronically Infected Cystic Fibrosis Patients

“…Such data are considered to provide important information about the risk of resistance emerging in a treated patient. However, pathoadaptive processes favor the emergence of bacteria displaying a hypermutator phenotype (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27), exhibiting significantly increased rates of mutation, or a small-colony morphotype (30)(31)(32)(33)(34), facilitating recurrent and persistent infections. Both adaptive responses confer antibiotic resistance, which, however, is not analyzed by routine procedures.…”

“…In addition to the poor penetration of antibacterials into the focus of infection, antibacterial therapy of CF patients is aggravated by several factors, such as biofilm formation, which impairs drug penetration and protects the pathogens from immune defense (7)(8)(9)(10)(11)(12)(13)(14); the development of hypermutator strains, characterized by high mutation rates, which result in pronounced resistance against antimicrobials and pathoadaptation ( [15][16][17][18][19][20][21][22][23][24][25][26][27]; drug adsorption to sputum components (28,29), which limits antibiotic activities; and slow growth as small-colony morphotypes (30)(31)(32)(33)(34), which facilitates recurrent and persistent infections and reduces the antibacterial activities of several antimicrobials (35,36). The result of the multiplicity of factors impairing the activities of antibacterials is that the pathogen cannot be eliminated, and infection persists in a chronic state.…”

Optimization of Polymyxin B in Combination with Doripenem To Combat Mutator Pseudomonas aeruginosa

“…Li et al determined that deactivation of DNA mismatch repair also helps to prolong bacterial survival (7). Although mutations in DNA repair mechanisms that lead to the mutator phenotype have been well characterized in species such as Escherichia coli and P. aeruginosa (10,34), there is a scarcity of data regarding antibiotic pharmacodynamics against these pathogens.…”

“…Wild-type and five isogenic strains with a deficiency in the MMR DNA repair system (34) or the GO BER system (10) were used in our studies. The genotypes and antibiotic MICs are listed in Table 1.…”

Emergence of Polymyxin B Resistance Influences Pathogenicity in Pseudomonas aeruginosa Mutators

“…Time-kill experiments were performed as previously described (15). Polymyxin B concentrations of 2, 4, and 8 mg/liter were evaluated against a starting bacterial inoculum of ϳ10 8 CFU/ml, and samples were obtained after 0 (predose), 1,2,4,6,8, 24, 28, 32, and 48 h. Experiments were performed in duplicate, and data are plotted as the average Ϯ standard deviation (SD). For the virulence experiments, bacterial suspensions were removed from each time-kill experiment at 0 (predose) and 48 h (for the polymyxin B 8-mg/liter arm).…”