A Novel Bacterial Resistance Mechanism against Human Group IIA-Secreted Phospholipase A2: Role of Streptococcus pyogenes Sortase A

Abstract

Human group IIA-secreted phospholipase A2 (sPLA2-IIA) is a bactericidal molecule important for the innate immune defense against Gram-positive bacteria. In this study, we analyzed its role in the host defense against Streptococcus pyogenes, a major human pathogen, and demonstrated that this bacterium has evolved a previously unidentified mechanism to resist killing by sPLA2-IIA. Analysis of a set of clinical isolates demonstrated that an ∼500-fold higher concentration of sPLA2-IIA was required to kill S. pyogenes compared with strains of the group B Streptococcus, which previously were shown to be sensitive to sPLA2-IIA, indicating that S. pyogenes exhibits a high degree of resistance to sPLA2-IIA. We found that an S. pyogenes mutant lacking sortase A, a transpeptidase responsible for anchoring LPXTG proteins to the cell wall in Gram-positive bacteria, was significantly more sensitive (∼30-fold) to sPLA2-IIA compared with the parental strain, indicating that one or more LPXTG surface proteins protect S. pyogenes against sPLA2-IIA. Importantly, using transgenic mice expressing human sPLA2-IIA, we showed that the sortase A-mediated sPLA2-IIA resistance mechanism in S. pyogenes also occurs in vivo. Moreover, in this mouse model, we also showed that human sPLA2-IIA is important for the defense against lethal S. pyogenes infection. Thus, we demonstrated a novel mechanism by which a pathogenic bacterium can evade the bactericidal action of sPLA2-IIA and we showed that sPLA2-IIA contributes to the host defense against S. pyogenes infection.

Pseudomonas aeruginosa eradicates Staphylococcus aureus by manipulating the host immunity

“…We propose that PA colonization improves this elimination by increasing sPLA2-IIA expression of airways cells. In agreement, previous studies showed that sPLA2-IIA is present in human and animal biological fluids or cell supernatants at sufficient levels to kill bacteria 21,[53][54][55][56] . AMPs such as LL-37 have also been shown to play a role in pulmonary host defense toward SA and PA in CF lungs 57 , although the antimicrobial activity of these AMPs is impaired in CF airways [57][58][59] .…”

“…NET289, NEN Radiochemicals, MA, USA). This allowed labelling of bacterial membrane phospholipids 55 . After centrifugation (3,000 g, 25°C, 15 min), bacterial pellets were suspended in 5 ml of LB medium and incubated for additional 30 min to allow binding oleic acid on the surface of bacteria to be incorporated, followed by rinsing once using 0.15 M NaCl to remove non-incorporated 3 H-OA.…”

Molecular determinants of bacterial sensitivity and resistance to mammalian Group IIA phospholipase A2

“…Bacillus subtilis is among the most sensitive to sPLA 2 -IIA, with as little as 0.1–1 nM (1.5–15 ng/ml) of human sPLA 2 -IIA sufficient to produce 1–3 logs killing of 10 6 bacteria/ml within 1 hour of incubation. Similar effects on S. aureus and Streptococcus agalactiae (20) (Group B streptococci; GBS) generally (but see below) require 10–100× higher sPLA2-IIA concentrations. Other Gram-positive bacterial species (e.g., Staphylococcus epidermidis, Streptococcus pyogenes , Enterococcus faecalis ) can also be killed by sPLA 2 -IIA but require even higher (10–100-fold) doses (17, 20).…”

Sortase A Induces Th17-Mediated and Antibody-Independent Immunity to Heterologous Serotypes of Group A Streptococci

“…Up to 12 GAS cell wall-anchored proteins contain LPXTG-like motifs [6], such as M proteins, protein F, C5a peptidase (ScpA), protein G-related α2-macroglobin-binding protein (GRAB), and pili [7]. Mutation of SrtA in most human Gram-positive pathogens, including GAS, dramatically reduced their ability to infect animals [8]–[13]. Analysis of 24 isolates of 12 GAS serotypes indicates that SrtA is present in all strains examined [14], indicating that SrtA is highly conserved and for that reason is a potential target for vaccine candidate.…”