The Klebsiella pneumoniae citrate synthase gene, gltA, influences site specific fitness during infection

Abstract

Klebsiella pneumoniae (Kp), one of the most common causes of healthcare-associated infections, increases patient morbidity, mortality and hospitalization costs. Kp must acquire nutrients from the host for successful infection. However, the host is able to prevent bacterial nutrient acquisition through multiple systems, including the innate immune protein lipocalin 2 (Lcn2) that prevents Kp iron acquisition by sequestering the siderophore enterobactin. To identify novel Kp factors that mediate evasion of nutritional immunity during lung infection, we undertook an InSeq study using a pool of >20,000 transposon mutants administered to Lcn2+/+ and Lcn2-/- mice. Comparing mutant frequencies between mouse genotypes, this genome-wide screen identified the Kp citrate synthase GltA as potentially interacting with Lcn2, and this novel finding was independently validated. Interestingly, in vitro studies suggest that this interaction is not direct. Given that GltA is involved in oxidative metabolism, we screened the ability of this mutant to use a variety of carbon and nitrogen sources. The results indicated that the gltA mutant has a distinct amino acid auxotrophy and is unable to use a variety of carbon sources. Specifically, we show that gltA is necessary for growth in bronchioloalveolar lavage fluid, which is amino acid-limited, but dispensable in serum, which is amino acid rich. Deletion of Lcn2 from the host leads to increased amino acid levels in bronchioloalveolar lavage fluid, and thus abrogates the loss of gltA during pneumonia in the Lcn2-/- background. GltA was also required for gut colonization, but dispensable in the bloodstream in a bacteremia model, demonstrating that deletion of gltA leads to an organ-specific fitness defect. Together, this study is the first to mechanistically describe a role for gltA in Kp infection and provide unique insight into how metabolic flexibility impacts bacterial fitness during infection.\n\nAuthor SummaryThe bacteria Klebsiella pneumoniae (Kp) is an important cause of infection in healthcare settings. These infections can be difficult to treat, as they frequently occur in chronically ill patients and the bacteria has the ability to acquire multiple antibiotic resistance markers. Kp is a common colonizer of the intestinal tract in hospitalized patients, and can progress to infections of the bloodstream, respiratory and urinary tract. However, the bacterial factors that allow Kp to replicate in these different body sites is unclear. In this study, we found that the Kp citrate synthase, GltA, enables bacterial replication in the lung and intestine by enhancing the ability of Kp to use diverse nutrients, in a mechanism known as metabolic flexibility. Kp lacking GltA require specific amino acids that are abundant in blood, but not other body sites. The work in this study provides novel insight into why Kp is a successful hospital pathogen that can colonize and infect multiple body sites.