Staphylococcus aureus Elaborates Leukocidin AB To Mediate Escape from within Human Neutrophils

Abstract

bMethicillin-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 poreforming 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.

Staphylococcus aureusLukAB cytotoxin kills human neutrophils by targeting the CD11b subunit of the integrin Mac-1

“…Despite the lack of overlap in cellular targets identified thus far, the staphylococcal leukotoxins have been shown to exhibit synergistic effects (26). The identification of CD11b as a cellular target for LukAB provides an explanation for the previously reported synergism between LukAB and PVL toward human PMNs (10,11), as sublytic concentrations of PVL increase CD11b surface levels on these cells (27). From an evolutionary standpoint, the use of multiple, nonoverlapping cellular targets provides an explanation for why S. aureus has such a large arsenal of cytotoxins and is such a welladapted pathogen when it comes to evading host PMNs.…”

“…For these experiments, PMNs were pretreated with the LM2/1 antibody or an isotype control before infection with GFP-LAC WT, isogenic ΔlukAB, or isogenic ΔlukAB chromosomally complemented with lukAB (11). These experiments were performed in the presence of lysostaphin and antiLukA to eliminate the potential influence of extracellular bacteria and LukAB, as well as the fluorescent dye ethidum bromide (EtBr) to measure membrane damage (11). Of note, pretreatment with LM2/1 before infection does not block phagocytosis of S. aureus as the amount of GFP-LAC observed within PMNs was similar regardless of LM2/1 treatment (Fig.…”

“…We recently established that LukAB-mediated cell damage postphagocytosis promotes the early escape of LAC from within PMNs and subsequent bacterial outgrowth (11). To determine if CD11b contributes to the intracellular cytotoxic activity of LukAB, the NT and CD11b shRNA PMN-HL60 cells were infected with opsonized USA300 and synchronized to promote phagocytosis (11).…”

“…LukAB is the most recently identified member of the bicomponent leukocidin family (9,10). This toxin contributes to the cytotoxicity of clinical isolates toward innate immune cells and has been shown to play an important role in the success of S. aureus in both ex vivo and in vivo models of infection (9,11). LukAB specifically targets phagocytes such as polymorphonuclear cells (PMNs or neutrophils) (9)(10)(11)(12), which are an integral part of the host innate immune response to S. aureus (13).…”

“…This toxin contributes to the cytotoxicity of clinical isolates toward innate immune cells and has been shown to play an important role in the success of S. aureus in both ex vivo and in vivo models of infection (9,11). LukAB specifically targets phagocytes such as polymorphonuclear cells (PMNs or neutrophils) (9)(10)(11)(12), which are an integral part of the host innate immune response to S. aureus (13). The capacity to kill PMNs is conserved among all leukocidins (4,5), although the actions of LukED (14) and Hlg (15) are not limited to phagocytes (5).…”