https://orcid.org/0000-0002-2452-965X
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ABSTRACT
Streptococcus suis (S. suis) causes meningitis, arthritis and endocarditis in piglets. The aim of this study was to characterize the IgM degrading enzyme of S. suis (IdeSsuis) and to investigate the role of IgM cleavage in evasion of the classical complement pathway and pathogenesis. Targeted mutagenesis of a cysteine in the putative active center of IdeSsuis abrogated IgM cleavage completely. In contrast to wt rIdeSsuis, point mutated rIdeSsuis_C195S did not reduce complement-mediated hemolysis indicating that complement inhibition by rIdeSsuis depends on the IgM proteolytic activity. A S. suis mutant expressing IdeSsuis_C195S did not reduce IgM labeling, whereas the wt and complemented mutant showed less IgM F(ab’)2 and IgM Fc antigen on the surface. IgM cleavage increased survival of S. suis in porcine blood ex vivo and mediated complement evasion as demonstrated by blood survival and C3 deposition assays including the comparative addition of rIdeSsuis and rIdeSsuis_C195S. However, experimental infection of piglets disclosed no significant differences in virulence between S. suis wt and isogenic mutants without IgM cleavage activity. This work revealed for the first time in vivo labeling of S. suis with IgM in the cerebrospinal fluid of piglets with meningitis. In conclusion, this study classifies IdeSsuis as a cysteine protease and emphasizes the role of IgM cleavage for bacterial survival in porcine blood and complement evasion though IgM cleavage is not crucial for the pathogenesis of serotype 2 meningitis.
Abbreviations
| αEry | = | porcine anti-sheep erythrocyte serum |
| BCR | = | B-cell receptor |
| CDS | = | colostrum-deprived piglet serum |
| CFU | = | Colony forming units |
| CSF | = | cerebrospinal fluid |
| FITC | = | fluorescein isothiocyanate |
| IdeE | = | Immunoglobulin G degrading enzyme of Streptococcus equi subsp. equi |
| IdeP | = | Immunoglobulin G degrading enzyme of Streptococcus phocae |
| IdeS | = | Immunoglobulin G degrading enzyme of Streptococcus pyogenes |
| IdeSsuis | = | Immunoglobulin M degrading enzyme of Streptococcus suis |
| IdeZ | = | Immunoglobulin G degrading enzyme of Streptococcus equi subsp. zooepidemicus |
| LB-broth | = | Luria Bertani Broth |
| MFI | = | geometric mean fluorescence intensity |
| MRP | = | Muramidase-released protein |
| OPA | = | Opsonophagocytosis assay |
| PBS | = | Phosphate buffered saline |
| PCR | = | Polymerase chain reaction |
| PE | = | phycoerythrin |
| POD | = | horseradish peroxidase |
| RT | = | Room temperature |
| SD | = | Standard deviation |
| SF | = | survival factor |
| S. suis | = | Streptococcus suis |
| ST | = | Streptococcus suis sequence type |
| TBST | = | Tris buffered saline Tween20 |
| THB | = | Todd-Hewitt Broth |
| VCP | = | vaccinia virus complement control protein |
| wt | = | wild type |
Acknowledgments
We thank H. Smith (DLO-Lelystad, Netherlands) for S. suis strain 10. Daisuke Takamatsu (National Institute of Animal Health, Japan) kindly provided the plasmid pSET5s. Ulrich von Pawel-Rammingen (Department of Molecular Biology, Umeå University, Umeå, Sweden) is acknowledged for providing the plasmid pET45bideSsuis_C195S. Renate Rutz (Institute for Immunology, University of Heidelberg, Heidelberg, Germany) is acknowledged for the execution of the sC5b-9 ELISA. Flow cytometry was performed at the core unit for flow cytometry, CUDZ, of the Veterinary Faculty of the University of Leipzig, Germany. This study was financially supported by the German Research Foundation (DFG BA 4730/3-1).
Disclosure statement
No potential conflict of interest was reported by the authors.
Supplemental material
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