Nutrient Acquisition in the Gram-Positive Pathogens Clostridioides Difficile and Staphylococcus Aureus

Abstract

Bacterial infections are a leading cause of death globally, with the Gram-positive bacterial pathogens Staphylococcus aureus and Clostridioides difficile posing serious threats to human health. The discovery and implementation of antibiotics has increased the global life expectancy. However, several unintended consequences of antibiotic usage have arisen including antibiotic resistance and antibiotic-associated infections. S. aureus is capable of developing resistance to all classes of clinically-relevant antibiotics, and C. difficile is the leading cause of antibiotic-associated infections. These facts highlight the necessity of developing improved therapeutic strategies to treat antibiotic-resistant infections, and reduce infections associated with antibiotic usage. We hypothesized that strategies to obtain essential nutrients serve as a promising target to treat infections. Here we investigated a mechanism used by C. difficile to metabolize a poisonous uracil analog, 4-thiouracil (4-TU) to acquire essential pyrimidine nucleotides. A thiouracil desulfurase protects C. difficile RNA from incorporation of 4-TU, and confers a fitness advantage against gut commensal bacteria that are unable to grow in the presence of 4-TU. Due to the antibacterial properties of 4-TU, we hypothesized 4-TU inhibits the growth of drug-resistant S. aureus. Indeed, 4-TU inhibits the growth of drug-resistant S. aureus, and analysis of RNA from 4-TU-treated S. aureus revealed that 4-TU hijacks the pyrimidine salvage pathway for incorporation into RNA. In addition to acquiring essential pyrimidine nucleotides, bacteria must acquire nutrient metals during infection. The host actively restricts metals from pathogens, and microbes maintain mechanisms to circumvent host-imposed metal limitation. Calprotectin is a pro-inflammatory antimicrobial protein that inhibits bacterial growth by sequestering metals. Calprotectin plays a critical role in the host response to C. difficile infection, and investigation of the cell types responsible for delivering calprotectin to the gut during infection revealed myeloid-derived cells are critical contributors of calprotectin. Analysis of C. difficile genes required for fitness in the presence of calprotectin suggest that high-affinity metal and siderophore transporters are important for competing with calprotectin for metals. Collectively, this Thesis proposes that bacterial mechanisms to obtain essential nutrients may serve as a promising target for therapeutic development.