Neutrophil interactions with Streptococcus pyogenes and Staphylococcus aureus
Author: Uhlmann, Julia
Date: 2017-05-05
Location: Lecture Hall 4Z, Alfred Nobels Allé 8, Karolinska University Hospital, Huddinge
Time: 09.30
Department: Inst för medicin, Huddinge / Dept of Medicine, Huddinge
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Thesis (3.498Mb)
Abstract
Streptococcus pyogenes and Staphylococcus aureus are Gram-positive bacteria that share
many features, including clinical presentations and pathogenic mechanisms, and yet, they
still have unique properties. They both cause diseases ranging from uncomplicated
infections to severe invasive diseases, such as sepsis and toxic shock syndrome, as well as
severe tissue infections including necrotizing fasciitis and necrotizing pneumonia. While
S. pyogenes is susceptible to penicillin, the public health concern regarding
staphylococcal infections is enhanced by the increasing prevalence of methicillin-resistant
S. aureus. Neutrophils have a central role as the first line of defense against bacteria, by
killing invaders through phagocytosis, neutrophil extracellular trap formation or
degranulation. Neutrophil granules contain a variety of proteins that contribute to
antimicrobial defense. However, degranulation and release of these proteolytic and
inflammatory factors in the tissue milieu can also be harmful to the host. This thesis
project explored interactions between neutrophils and both Gram-positive species, with a
particular focus on neutrophils as potential contributors to the pathogenesis of these
infections.
In paper I we investigated the neutrophil response towards streptococcal factors secreted by different clinical isolates. We identified phosphoglycerate kinase (PGK), a moonlighting protein with glycolytic function, as a novel, potent neutrophil activator with the ability to trigger degranulation. PGK was found to be susceptible to proteolytic degradation by SpeB and consequently SpeB-negative strains elicited stronger neutrophil responses. This finding is of interest, as hypervirulent SpeB-negative strains are associated with invasive streptococcal infections.
In paper II we studied the effect of sub-inhibitory concentrations of LL-37 on S. pyogenes, as these concentrations have been reported to alter virulence gene expression. We showed that LL-37 induced the release of extracellular vesicle-like structures, which contained several virulence factors with immunostimulatory properties. This is the first report of vesicle-like structures release by S. pyogenes in response to LL-37. The presence of virulence factors in these vesicles and the pro-inflammatory effect towards neutrophils implicates a potential role for LL-37 in S. pyogenes pathogenesis.
In paper III we focused on bi-component leukocidins from S. aureus and their effect on neutrophil degranulation. PVL and LukED had a dose-dependent effect on cytotoxicity. However, neutrophil degranulation showed a different pattern. While PVL triggered even at sub-lytic concentrations neutrophil granule exocytosis, this event was only seen when lytic concentrations of LukED were used. Determination of the whole neutrophil secretomes triggered by lytic and sub-lytic concentrations of PVL and LukED revealed significantly different response profiles. This study demonstrates that neutrophil activation and degranulation in response to S. aureus pore-forming toxins depends on both the type and concentration of toxin. PVL was found to be a potent activator even at sublytic concentrations.
In summary, we identified the streptococcal factor PGK, as well as the pore-forming toxins from S. aureus as potent triggers of neutrophil activation and degranulation. In addition, we show that the antimicrobial peptide LL-37 induces the release of proinflammatory extracellular vesicle-like structures from the surface of S. pyogenes. Together, these studies demonstrate specific neutrophil responses triggered by Grampositive bacterial virulence factors, which is in line with a likely contribution to disease pathogenesis
In paper I we investigated the neutrophil response towards streptococcal factors secreted by different clinical isolates. We identified phosphoglycerate kinase (PGK), a moonlighting protein with glycolytic function, as a novel, potent neutrophil activator with the ability to trigger degranulation. PGK was found to be susceptible to proteolytic degradation by SpeB and consequently SpeB-negative strains elicited stronger neutrophil responses. This finding is of interest, as hypervirulent SpeB-negative strains are associated with invasive streptococcal infections.
In paper II we studied the effect of sub-inhibitory concentrations of LL-37 on S. pyogenes, as these concentrations have been reported to alter virulence gene expression. We showed that LL-37 induced the release of extracellular vesicle-like structures, which contained several virulence factors with immunostimulatory properties. This is the first report of vesicle-like structures release by S. pyogenes in response to LL-37. The presence of virulence factors in these vesicles and the pro-inflammatory effect towards neutrophils implicates a potential role for LL-37 in S. pyogenes pathogenesis.
In paper III we focused on bi-component leukocidins from S. aureus and their effect on neutrophil degranulation. PVL and LukED had a dose-dependent effect on cytotoxicity. However, neutrophil degranulation showed a different pattern. While PVL triggered even at sub-lytic concentrations neutrophil granule exocytosis, this event was only seen when lytic concentrations of LukED were used. Determination of the whole neutrophil secretomes triggered by lytic and sub-lytic concentrations of PVL and LukED revealed significantly different response profiles. This study demonstrates that neutrophil activation and degranulation in response to S. aureus pore-forming toxins depends on both the type and concentration of toxin. PVL was found to be a potent activator even at sublytic concentrations.
In summary, we identified the streptococcal factor PGK, as well as the pore-forming toxins from S. aureus as potent triggers of neutrophil activation and degranulation. In addition, we show that the antimicrobial peptide LL-37 induces the release of proinflammatory extracellular vesicle-like structures from the surface of S. pyogenes. Together, these studies demonstrate specific neutrophil responses triggered by Grampositive bacterial virulence factors, which is in line with a likely contribution to disease pathogenesis
List of papers:
I. Uhlmann J, Siemens N, Kai-Larsen Y, Fiedler T, Bergman P, Johansson L, Norrby-Teglund A. Phosphoglycerate kinase – a novel streptococcal factor involved in neutrophil activation and degranulation. Journal of Infectious Diseases (2016) 214: 1876-83.
Fulltext (DOI)
Pubmed
View record in Web of Science®
II. Uhlmann J, Rohde M, Siemens N, Kreikemeyer B, Bergman P, Johansson L, Norrby-Teglund A. LL-37 triggers formation of Streptococcus pyogenes extracellular vesicle-like structures with immune stimulatory properties. Journal of Innate Immunity (2016) 8: 243-57.
Fulltext (DOI)
Pubmed
View record in Web of Science®
III. Uhlmann J, Snäll J, Wajima T, Mairpady Shambat S, Vandenesch F, Bergman P, Siemens N, Norrby-Teglund A. Dissection of neutrophil cytotoxicity and exocytosis caused by Staphylococcus aureus leukotoxins. [Manuscript]
I. Uhlmann J, Siemens N, Kai-Larsen Y, Fiedler T, Bergman P, Johansson L, Norrby-Teglund A. Phosphoglycerate kinase – a novel streptococcal factor involved in neutrophil activation and degranulation. Journal of Infectious Diseases (2016) 214: 1876-83.
Fulltext (DOI)
Pubmed
View record in Web of Science®
II. Uhlmann J, Rohde M, Siemens N, Kreikemeyer B, Bergman P, Johansson L, Norrby-Teglund A. LL-37 triggers formation of Streptococcus pyogenes extracellular vesicle-like structures with immune stimulatory properties. Journal of Innate Immunity (2016) 8: 243-57.
Fulltext (DOI)
Pubmed
View record in Web of Science®
III. Uhlmann J, Snäll J, Wajima T, Mairpady Shambat S, Vandenesch F, Bergman P, Siemens N, Norrby-Teglund A. Dissection of neutrophil cytotoxicity and exocytosis caused by Staphylococcus aureus leukotoxins. [Manuscript]
Institution: Karolinska Institutet
Supervisor: Norrby-Teglund, Anna
Co-supervisor: Jervelius, Linda; Bergman, Peter; Siemens, Nikolai
Issue date: 2017-04-03
Rights:
Publication year: 2017
ISBN: 978-91-7676-581-4
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