Pneumococcal Neuraminidases Increase Platelet Killing by Pneumolysin

Abstract

Background Platelets prevent extravasation of capillary fluids into the pulmonary interstitial tissue by sealing gaps in inflamed endothelium. This reduces respiratory distress associated with pneumonia. Streptococcus pneumoniae is the leading cause of severe community-acquired pneumonia. Pneumococci produce pneumolysin (PLY), which forms pores in membranes of eukaryotic cells including platelets. Additionally, pneumococci express neuraminidases, which cleave sialic acid residues from eukaryotic glycoproteins. In this study, we investigated the effect of desialylation on PLY binding and pore formation on platelets.

Materials and Methods We incubated human platelets with purified neuraminidases and PLY, or nonencapsulated S. pneumoniae D39/TIGR4 and isogenic mutants deficient in PLY and/or NanA. We assessed platelet desialylation, PLY binding, and pore formation by flow cytometry. We also analyzed the inhibitory potential of therapeutic immunoglobulin G preparations (IVIG [intravenous immunoglobulin]).

Results Wild-type pneumococci cause desialylation of platelet glycoproteins by neuraminidases, which is reduced by 90 to 100% in NanA-deficient mutants. NanC, cleaving only α2,3-linked sialic acid, induced platelet desialylation. PLY binding to platelets then x2doubled (p = 0.0166) and pore formation tripled (p = 0.0373). A neuraminidase cleaving α2,3-, α2,6-, and α2,8-linked sialic acid like NanA was even more efficient. Addition of polyvalent IVIG (5 mg/mL) decreased platelet desialylation induced by NanC up to 90% (p = 0.263) and reduced pore formation >95% (p < 0.0001) when incubated with pneumococci.

Conclusion Neuraminidases are key virulence factors of pneumococci and desialylate platelet glycoproteins, thereby unmasking PLY-binding sites. This enhances binding of PLY and pore formation showing that pneumococcal neuraminidases and PLY act in concert to kill platelets. However, human polyvalent immunoglobulin G preparations are promising agents for therapeutic intervention during severe pneumococcal pneumonia.

Keywords

Streptococcus pneumoniae - neuraminidase - pneumolysin - platelets - immunoglobulins

Authors' Contribution

K.J.F. performed the experiments with purified toxins, evaluated the data, and wrote the manuscript. L.K. performed the experiments with whole bacteria, evaluated the data, and wrote the manuscript. S.H. and T.P.K. planned the experiments, evaluated the data, and edited the manuscript. K.J. and J.W. contributed to the experiments and edited the manuscript. A.G. and Sv.H. designed the project, supervised the project, evaluated the data, and edited the manuscript. All authors reviewed the final version of the manuscript.

^* These authors contributed equally to the study.

Publication History

Received: 11 November 2023

Accepted: 18 July 2024

Accepted Manuscript online:
19 July 2024

Article published online:
20 August 2024

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