Faculty Bibliography

Methicillin-resistant Staphylococcus aureus (MRSA) strains of the pulsed-field type USA300 are primarily responsible for the current community-associated epidemic of MRSA infections in the United States. The success of USA300 is partly attributed to the ability of the pathogen to avoid destruction by human neutrophils (polymorphonuclear leukocytes [PMNs]), which are crucial to the host immune response to S. aureus infection. In this work, we investigated the contribution of bicomponent pore-forming toxins to the ability of USA300 to withstand attack from primary human PMNs. We demonstrate that in vitro growth conditions influence the expression, production, and availability of leukotoxins by USA300, which in turn impact the cytotoxic potential of this clone toward PMNs. Interestingly, we also found that upon exposure to PMNs, USA300 preferentially activates the promoter of the lukAB operon, which encodes the recently identified leukocidin AB (LukAB). LukAB elaborated by extracellular S. aureus forms pores in the plasma membrane of PMNs, leading to PMN lysis, highlighting a contribution of LukAB to USA300 virulence. We now show that LukAB also facilitates the escape of bacteria engulfed within PMNs, in turn enabling the replication and outgrowth of S. aureus. Together, these results suggest that upon encountering PMNs S. aureus induces the production of LukAB, which serves as an extra- and intracellular weapon to protect the bacterium from destruction by human PMNs.

The success of Staphylococcus aureus as a leading cause of deadly hospital-acquired and community-acquired infections is attributed to its high-level resistance to most antibiotics, and the multitude of virulence factors it elaborates. Most clinical isolates produce up to four bi-component pore-forming toxins capable of lysing cells of the immune system. Subtle differences in activity and target range of each leukotoxin suggest that these toxins are not redundant, but instead may have specialized functions in attacking and/or evading host defenses. In turn, the host has developed countermeasures recognizing sublytic levels of leukotoxins as signals to activate protective immune defenses. The opposing cytotoxic and immune-activating effects of leukotoxins on host cells make for a complex dynamic between S. aureus and the host.

Staphylococcus aureus epidemic strain USA300 is a highly successful pathogen. However, the underlying basis of this success is not clear. Now, Thurlow and colleagues (2013) provide evidence linking the bacterial arginine catabolic mobile element (ACME) to the dominance of USA300 as a pathogen of the skin.

Pore-forming toxins are critical virulence factors for many bacterial pathogens and are central to Staphylococcus aureus-mediated killing of host cells. S. aureus encodes pore-forming bi-component leukotoxins that are toxic towards neutrophils, but also specifically target other immune cells. Despite decades since the first description of staphylococcal leukocidal activity, the host factors responsible for the selectivity of leukotoxins towards different immune cells remain unknown. Here we identify the human immunodeficiency virus (HIV) co-receptor CCR5 as a cellular determinant required for cytotoxic targeting of subsets of myeloid cells and T lymphocytes by the S. aureus leukotoxin ED (LukED). We further demonstrate that LukED-dependent cell killing is blocked by CCR5 receptor antagonists, including the HIV drug maraviroc. Remarkably, CCR5-deficient mice are largely resistant to lethal S. aureus infection, highlighting the importance of CCR5 targeting in S. aureus pathogenesis. Thus, depletion of CCR5(+) leukocytes by LukED suggests a new immune evasion mechanism of S. aureus that can be therapeutically targeted.

Staphylococcus aureus is a significant human pathogen that is capable of infecting a wide range of host tissues. This bacterium is able to evade the host immune response by utilizing a repertoire of virulence factors. These factors are tightly regulated by various two-component systems (TCS) and transcription factors. Previous studies have suggested that transcriptional regulation of a subset of immunomodulators, known as the staphylococcal superantigen-like proteins (Ssls), is mediated by the master regulators accessory gene regulator (Agr) TCS, S. aureus exoprotein expression (Sae) TCS, and Rot. Here we demonstrate that Rot and SaeR, the response regulator of the Sae TCS, synergize to coordinate the activation of the ssl promoters. We have determined that both transcription factors are required, but that neither is sufficient, for promoter activation. This regulatory scheme is mediated by direct binding of both transcription factors to the ssl promoters. We also demonstrate that clinically relevant methicillin-resistant S. aureus (MRSA) strains respond to neutrophils via the Sae TCS to upregulate the expression of ssls. Until now, Rot and the Sae TCS have been proposed to work in opposition of one another on their target genes. This is the first example of these two regulators working in concert to activate promoters.

Staphylococcus aureus is a major human pathogen that is capable of producing an expansive repertoire of cell surface-associated and extracellular virulence factors. Herein we describe an S. aureus regulatory RNA, SSR42, which modulates the expression of approximately 80 mRNA species, including several virulence factors, in S. aureus strains UAMS-1 and USA300 (LAC) during stationary-phase growth. Mutagenesis studies revealed that SSR42 codes for an 891-nucleotide RNA molecule and that the molecule's regulatory effects are mediated by the full-length transcript. Western blotting and functional assays indicated that the regulatory effects of SSR42 correlate with biologically significant changes in corresponding protein abundances. Further, in S. aureus strain LAC, SSR42 is required for wild-type levels of erythrocyte lysis, resistance to human polymorphonuclear leukocyte killing, and pathogenesis in a murine model of skin and soft tissue infection. Taken together, our results indicate that SSR42 is a novel S. aureus regulatory RNA molecule that contributes to the organism's ability to cause disease.

We investigated the antimicrobial activities of N-substituted glycine "peptoid" oligomers incorporating cationic and hydrophobic side chains. Head-to-tail macrocyclization was employed to enhance antimicrobial activity. Both linear and cyclic peptoids, ranging from six to ten residues, demonstrate potent antimicrobial activity against Gram-positive and Gram-negative bacteria. These peptoids do not cause significant lysis of human erythrocytes, indicating selective antimicrobial activity. Conformational ordering established upon macrocyclization is generally associated with an enhanced capacity to inhibit bacterial cell growth. Moreover, increased hydrophobic surface area also plays a role in improving antimicrobial activity. We demonstrate the potency of a cyclic peptoid in exerting antimicrobial activity against clinical strains of S. aureus while deterring the emergence of antimicrobial resistance.

Bloodstream infection with Staphylococcus aureus is common and can be fatal. However, virulence factors that contribute to lethality in S. aureus bloodstream infection are poorly defined. We discovered that LukED, a commonly overlooked leucotoxin, is critical for S. aureus bloodstream infection in mice. We also determined that LukED promotes S. aureus replication in vivo by directly killing phagocytes recruited to sites of haematogenously seeded tissue. Furthermore, we established that murine neutrophils are the primary target of LukED, as the greater virulence of wild-type S. aureus compared with a lukED mutant was abrogated by depleting neutrophils. The in vivo toxicity of LukED towards murine phagocytes is unique among S. aureus leucotoxins, implying its crucial role in pathogenesis. Moreover, the tropism of LukED for murine phagocytes highlights the utility of murine models to study LukED pathobiology, including development and testing of strategies to inhibit toxin activity and control bacterial infection