How the Anaerobic Enteropathogen Clostridioides difficile Tolerates Low O₂ Tensions

Area of Science:

• Microbiology
• Bacterial Physiology
• Oxygen Metabolism

Background:

• Clostridioides difficile causes antibiotic-associated diarrhea.
• C. difficile spores germinate in the small intestine, exposing vegetative cells to low oxygen.
• The bacterium tolerates low oxygen levels, suggesting detoxification mechanisms.

Purpose of the Study:

• To investigate the role of specific proteins in C. difficile oxygen tolerance.
• To identify key enzymes involved in oxygen detoxification.

Main Methods:

• Studied the alternative sigma factor σB and its regulated genes.
• Purified and characterized flavodiiron proteins (FdpA, FdpF) and reverse rubrerythrins (revRbr1, revRbr2).
• Assessed the growth of C. difficile mutants under varying low oxygen conditions.

Main Results:

• σB controls genes encoding FdpA, FdpF, revRbr1, and revRbr2.
• Purified revRbr1 and revRbr2 exhibit NADH-linked O2- and H2O2-reductase activities.
• Purified FdpA primarily functions as an O2-reductase.
• Mutants lacking these proteins showed impaired growth at low oxygen levels (0.1-0.4% O2).
• The quadruple mutant exhibited a more severe growth defect than triple mutants, indicating additive effects.

Conclusions:

• Reverse rubrerythrins (revRbrs), FdpF, and FdpA are essential for C. difficile growth in physiological oxygen tensions found in the colon.
• These O2-reductase enzymes are critical for protecting C. difficile vegetative cells from oxygen-induced damage.
• C. difficile employs complex detoxification pathways involving these enzymes to survive fluctuating oxygen levels in the gastrointestinal tract.