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Streptolysin O Rapidly Impairs Neutrophil Oxidative Burst and Antibacterial Responses to Group A Streptococcus.

Uchiyama S, Döhrmann S, Timmer AM, Dixit N, Ghochani M, Bhandari T, Timmer JC, Sprague K, Bubeck-Wardenburg J, Simon SI, Nizet V - Front Immunol (2015)

Bottom Line: Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome.The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination.We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of California San Diego , La Jolla, CA , USA.

ABSTRACT
Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome. A globally disseminated clone of M1T1 GAS has been associated with an increase in severe, invasive GAS infections in recent decades. The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination. SLO is known to promote GAS resistance to phagocytic clearance by neutrophils, a critical first element of host defense against invasive bacterial infection. Here, we examine the role of SLO in modulating specific neutrophil functions during their early interaction with GAS. We find that SLO at subcytotoxic concentrations and early time points is necessary and sufficient to suppress neutrophil oxidative burst, in a manner reversed by free cholesterol and anti-SLO blocking antibodies. In addition, SLO at subcytotoxic concentrations blocked neutrophil degranulation, interleukin-8 secretion and responsiveness, and elaboration of DNA-based neutrophil extracellular traps, cumulatively supporting a key role for SLO in GAS resistance to immediate neutrophil killing. A non-toxic SLO derivate elicits protective immunity against lethal GAS challenge in a murine infection model. We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance.

No MeSH data available.


Related in: MedlinePlus

Group A Streptococcus SLO suppresses neutrophil oxidative burst. (A) GAS WT serotype M1 and M3 suppress oxidative burst response in human neutrophils compared to representative WT strains of Streptococcus pneumoniae (D39) and Staphylococcus aureus (Newman) and media only control. (B) An isogenic GAS ΔSLO mutant fails to suppress oxidative burst in neutrophils compared to isogenic ΔM1, ΔSpeB, ΔNAD, and ΔSLS mutants and GAS WT. (C) The complemented GAS ΔSLO mutant (ΔSLO + pSLO) restores the suppression of oxidative burst. (D) Recombinant, active SLO protein suppresses oxidative burst in a dose-dependent manner in PMA-stimulated neutrophils, including media alone as a negative control. (E) SLO protects against killing by neutrophils at an MOI of 10 at 30 min post infection. Results are given in average ± SEM and analyzed by Student’s t-test (n.s., not significant).
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Figure 1: Group A Streptococcus SLO suppresses neutrophil oxidative burst. (A) GAS WT serotype M1 and M3 suppress oxidative burst response in human neutrophils compared to representative WT strains of Streptococcus pneumoniae (D39) and Staphylococcus aureus (Newman) and media only control. (B) An isogenic GAS ΔSLO mutant fails to suppress oxidative burst in neutrophils compared to isogenic ΔM1, ΔSpeB, ΔNAD, and ΔSLS mutants and GAS WT. (C) The complemented GAS ΔSLO mutant (ΔSLO + pSLO) restores the suppression of oxidative burst. (D) Recombinant, active SLO protein suppresses oxidative burst in a dose-dependent manner in PMA-stimulated neutrophils, including media alone as a negative control. (E) SLO protects against killing by neutrophils at an MOI of 10 at 30 min post infection. Results are given in average ± SEM and analyzed by Student’s t-test (n.s., not significant).

Mentions: A select group of Gram-positive human pathogens including GAS, S. aureus, and S. pneumoniae are leading causes of invasive, potentially life-threatening infections worldwide (1, 33, 34). Neutrophils are a critical first line of defense against such invading pathogens, and the rapid production of bactericidal ROS is key element of their effectiveness. Compared to S. aureus and S. pneumoniae, GAS of the M1 and M3 serotypes elicited a reduced level of oxidative burst from freshly isolated human neutrophils 20 min post infection (Figure 1A). As each bacterial species is replete with pathogen-associated molecular patterns (e.g., lipoteichoic acid, peptidoglycan, and formyl peptides) to stimulate neutrophil activation, we hypothesized that GAS produced factor(s) that suppressed the full host oxidative burst potential. By screening a panel of isogenic M1T1 GAS mutants lacking defined virulence factors, we identified a higher neutrophil oxidative burst in response to a GAS ΔSLO mutant, suggesting that the toxin could play a key role in oxidative burst suppression (Figure 1B).


Streptolysin O Rapidly Impairs Neutrophil Oxidative Burst and Antibacterial Responses to Group A Streptococcus.

Uchiyama S, Döhrmann S, Timmer AM, Dixit N, Ghochani M, Bhandari T, Timmer JC, Sprague K, Bubeck-Wardenburg J, Simon SI, Nizet V - Front Immunol (2015)

Group A Streptococcus SLO suppresses neutrophil oxidative burst. (A) GAS WT serotype M1 and M3 suppress oxidative burst response in human neutrophils compared to representative WT strains of Streptococcus pneumoniae (D39) and Staphylococcus aureus (Newman) and media only control. (B) An isogenic GAS ΔSLO mutant fails to suppress oxidative burst in neutrophils compared to isogenic ΔM1, ΔSpeB, ΔNAD, and ΔSLS mutants and GAS WT. (C) The complemented GAS ΔSLO mutant (ΔSLO + pSLO) restores the suppression of oxidative burst. (D) Recombinant, active SLO protein suppresses oxidative burst in a dose-dependent manner in PMA-stimulated neutrophils, including media alone as a negative control. (E) SLO protects against killing by neutrophils at an MOI of 10 at 30 min post infection. Results are given in average ± SEM and analyzed by Student’s t-test (n.s., not significant).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4644796&req=5

Figure 1: Group A Streptococcus SLO suppresses neutrophil oxidative burst. (A) GAS WT serotype M1 and M3 suppress oxidative burst response in human neutrophils compared to representative WT strains of Streptococcus pneumoniae (D39) and Staphylococcus aureus (Newman) and media only control. (B) An isogenic GAS ΔSLO mutant fails to suppress oxidative burst in neutrophils compared to isogenic ΔM1, ΔSpeB, ΔNAD, and ΔSLS mutants and GAS WT. (C) The complemented GAS ΔSLO mutant (ΔSLO + pSLO) restores the suppression of oxidative burst. (D) Recombinant, active SLO protein suppresses oxidative burst in a dose-dependent manner in PMA-stimulated neutrophils, including media alone as a negative control. (E) SLO protects against killing by neutrophils at an MOI of 10 at 30 min post infection. Results are given in average ± SEM and analyzed by Student’s t-test (n.s., not significant).
Mentions: A select group of Gram-positive human pathogens including GAS, S. aureus, and S. pneumoniae are leading causes of invasive, potentially life-threatening infections worldwide (1, 33, 34). Neutrophils are a critical first line of defense against such invading pathogens, and the rapid production of bactericidal ROS is key element of their effectiveness. Compared to S. aureus and S. pneumoniae, GAS of the M1 and M3 serotypes elicited a reduced level of oxidative burst from freshly isolated human neutrophils 20 min post infection (Figure 1A). As each bacterial species is replete with pathogen-associated molecular patterns (e.g., lipoteichoic acid, peptidoglycan, and formyl peptides) to stimulate neutrophil activation, we hypothesized that GAS produced factor(s) that suppressed the full host oxidative burst potential. By screening a panel of isogenic M1T1 GAS mutants lacking defined virulence factors, we identified a higher neutrophil oxidative burst in response to a GAS ΔSLO mutant, suggesting that the toxin could play a key role in oxidative burst suppression (Figure 1B).

Bottom Line: Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome.The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination.We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of California San Diego , La Jolla, CA , USA.

ABSTRACT
Group A Streptococcus (GAS) causes a wide range of human infections, ranging from simple pharyngitis to life-threatening necrotizing fasciitis and toxic shock syndrome. A globally disseminated clone of M1T1 GAS has been associated with an increase in severe, invasive GAS infections in recent decades. The secreted GAS pore-forming toxin streptolysin O (SLO), which induces eukaryotic cell lysis in a cholesterol-dependent manner, is highly upregulated in the GAS M1T1 clone during bloodstream dissemination. SLO is known to promote GAS resistance to phagocytic clearance by neutrophils, a critical first element of host defense against invasive bacterial infection. Here, we examine the role of SLO in modulating specific neutrophil functions during their early interaction with GAS. We find that SLO at subcytotoxic concentrations and early time points is necessary and sufficient to suppress neutrophil oxidative burst, in a manner reversed by free cholesterol and anti-SLO blocking antibodies. In addition, SLO at subcytotoxic concentrations blocked neutrophil degranulation, interleukin-8 secretion and responsiveness, and elaboration of DNA-based neutrophil extracellular traps, cumulatively supporting a key role for SLO in GAS resistance to immediate neutrophil killing. A non-toxic SLO derivate elicits protective immunity against lethal GAS challenge in a murine infection model. We conclude that SLO exerts a novel cytotoxic-independent function at early stages of invasive infections (<30 min), contributing to GAS escape from neutrophil clearance.

No MeSH data available.


Related in: MedlinePlus