The food-borne pathogen Campylobacter jejuni depends on the AddAB DNA repair system to defend against bile in the intestinal environment

Edwards

2021

Preprint

Antibiotics inhibit essential bacterial processes, resulting in arrest of growth and in some cases cell death. Many antibiotics are also reported to trigger endogenous production of reactive oxygen species (ROS), which damage DNA and other macromolecules. However, the type of DNA damage that arises and the mechanisms used by bacteria to repair it are largely unclear. We found that several different classes of antibiotic triggered dose-dependent DNA damage in Staphylococcus aureus, including some bacteriostatic drugs. Damage was heterogenous and varied in magnitude between strains. However, antibiotic-triggered DNA damage led to double strand breaks, the processing of which by the RexAB helicase/nuclease complex triggered the SOS response and reduced staphylococcal susceptibility to most of the antibacterials tested. In most cases, DNA DSBs occurred under aerobic but not anaerobic conditions, suggesting a role for ROS. We conclude that DNA double strand breaks are a common occurrence during bacterial exposure to several different antibiotic classes and that repair of this damage by the RexAB complex promotes bacterial survival.

Section: Discussionsupporting

confidence: 61%

Multiple classes of bactericidal antibiotics cause DNA double strand breaks inStaphylococcus aureus

Edwards

2021

Preprint

“…Previous work demonstrated the requirement of DNA DSB repair for the survival of Gram-negative pathogens in vivo . For example, AddAB was shown to be required for the infection of chickens and mice by Campylobacter jejuni and Helicobacter pylori , respectively ( 44 , 45 ). Furthermore, the virulence of Salmonella enterica in a murine model of bacteremia was dependent upon the RecBCD DNA DSB repair complex ( 46 ).…”

Section: Discussionmentioning

confidence: 99%

Staphylococcal DNA Repair Is Required for Infection

Kim

et al. 2020

mBio

To cause infection, Staphylococcus aureus must withstand damage caused by host immune defenses. However, the mechanisms by which staphylococcal DNA is damaged and repaired during infection are poorly understood. Using a panel of transposon mutants, we identified the rexBA operon as being important for the survival of Staphylococcus aureus in whole human blood. Mutants lacking rexB were also attenuated for virulence in murine models of both systemic and skin infections. We then demonstrated that RexAB is a member of the AddAB family of helicase/nuclease complexes responsible for initiating the repair of DNA double-strand breaks. Using a fluorescent reporter system, we were able to show that neutrophils cause staphylococcal DNA double-strand breaks through reactive oxygen species (ROS) generated by the respiratory burst, which are repaired by RexAB, leading to the induction of the mutagenic SOS response. We found that RexAB homologues in Enterococcus faecalis and Streptococcus gordonii also promoted the survival of these pathogens in human blood, suggesting that DNA double-strand break repair is required for Gram-positive bacteria to survive in host tissues. Together, these data demonstrate that DNA is a target of host immune cells, leading to double-strand breaks, and that the repair of this damage by an AddAB-family enzyme enables the survival of Gram-positive pathogens during infection. IMPORTANCE To cause infection, bacteria must survive attack by the host immune system. For many bacteria, including the major human pathogen Staphylococcus aureus, the greatest threat is posed by neutrophils. These immune cells ingest the invading organisms and try to kill them with a cocktail of chemicals that includes reactive oxygen species (ROS). The ability of S. aureus to survive this attack is crucial for the progression of infection. However, it was not clear how the ROS damaged S. aureus and how the bacterium repaired this damage. In this work, we show that ROS cause breaks in the staphylococcal DNA, which must be repaired by a two-protein complex known as RexAB; otherwise, the bacterium is killed, and it cannot sustain infection. This provides information on the type of damage that neutrophils cause S. aureus and the mechanism by which this damage is repaired, enabling infection.

“…Previous work has demonstrated the requirement of DNA DSB repair for the survival of Gram-negative pathogens in vivo . For example, AddAB was shown to be required for the infection of chickens and mice by Campylobacter jejuni and Helicobacter pylori respectively 41,42 . Furthermore, the virulence of Salmonella enterica in a murine model of bacteraemia was dependent upon the RecBCD DNA DSB repair complex 43 .…”

Section: Discussionmentioning

confidence: 99%

Staphylococcal DNA repair is required for infection

Kim

et al. 2020

Preprint

35Staphylococcus aureus is a leading cause of chronic and recurrent infections of the skin, bones, joints 36 and bloodstream. During infection, S. aureus faces the twin threats of the host immune system and 37 therapeutic antibiotics. Since both host immune cells and antibiotics generate reactive oxygen species, 38we hypothesised that S. aureus may employ a common mechanism to repair damage caused by either 39 threat. We found that staphylococcal DNA is damaged during exposure to both human neutrophils 40 and most bactericidal antibiotics. To understand the nature of this damage and how S. aureus repairs 41 it, we screened a panel of transposon mutants defective for various DNA repair processes. This 42 revealed that loss of the rexBA operon significantly reduced staphylococcal survival in human blood, 43 during incubation with purified neutrophils, in the peritoneal cavity of mice and during exposure to a 44 large panel of bactericidal antibiotics. We then used biochemical assays to demonstrate that RexAB is 45 a member of the AddAB family of ATP-dependent helicase/nucleases that are required for the repair 46 of DNA double strand breaks. Finally, we found that RexAB homologues in Enterococcus faecalis and 47Streptococcus gordonii also promoted survival of these pathogens in human blood, suggesting that 48 DNA repair constitutes a broadly conserved defence against neutrophils. Together, these data 49 demonstrate that DNA is a target of host immune cells and several antibiotics, leading to double-50 strand breaks, and that repair of this damage by an AddAB-family enzyme enables the survival of 51Gram-positive pathogens during infection. 52 53 54