Monoclonal antibody-based localization of major diagnostic antigens in metacestode tissue, excretory/secretory products, and extracellular vesicles of Echinococcus species

Philipp A Kronenberg; Michael Reinehr; Ramon Marc Eichenberger; Sina Hasler; Teivi Laurimäe; Achim Weber; Ansgar Deibel; Beat Müllhaupt; Bruno Gottstein; Norbert Müller; Andrew Hemphill; Peter Deplazes

doi:10.3389/fcimb.2023.1162530

Monoclonal antibody-based localization of major diagnostic antigens in metacestode tissue, excretory/secretory products, and extracellular vesicles of Echinococcus species

Front Cell Infect Microbiol. 2023 Mar 16:13:1162530. doi: 10.3389/fcimb.2023.1162530. eCollection 2023.

Authors

Philipp A Kronenberg^{1

2}, Michael Reinehr³, Ramon Marc Eichenberger^{1

4}, Sina Hasler¹, Teivi Laurimäe¹, Achim Weber³, Ansgar Deibel⁵, Beat Müllhaupt⁵, Bruno Gottstein⁶, Norbert Müller^{6

7}, Andrew Hemphill⁷, Peter Deplazes^{1

5}

Affiliations

¹ Institute of Parasitology, Vetsuisse and Medical Faculty, University of Zurich, Zurich, Switzerland.
² Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
³ Department of Pathology and Molecular Pathology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.
⁴ Microbiology and Molecular Biology, Institute of Chemistry and Biotechnology, Zurich University of Applied Sciences, Zurich University of Applied Sciences' (ZHAW), Wädenswil, Switzerland.
⁵ Department of Gastroenterology and Hepatology and Swiss HPB and Transplant Center, University Hospital Zurich, University of Zurich, Zurich, Switzerland.
⁶ Institute for Infectious Diseases, Medical Faculty, University of Bern, Bern, Switzerland.
⁷ Institute of Parasitology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.

Abstract

Alveolar (AE) and cystic echinococcosis (CE) are severe parasitic zoonoses caused by the larval stages of Echinococcus multilocularis and E. granulosus sensu lato, respectively. A panel of 7 monoclonal antibodies (mAbs) was selected against major diagnostic epitopes of both species. The binding capacity of the mAbs to Echinococcus spp. excretory/secretory products (ESP) was analyzed by sandwich-ELISA, where mAb Em2G11 and mAb EmG3 detected in vitro extravesicular ESP of both E. multilocularis and E. granulosus s.s. These findings were subsequently confirmed by the detection of circulating ESP in a subset of serum samples from infected hosts including humans. Extracellular vesicles (EVs) were purified, and the binding to mAbs was analyzed by sandwich-ELISA. Transmission electron microscopy (TEM) was used to confirm the binding of mAb EmG3 to EVs from intravesicular fluid of Echinococcus spp. vesicles. The specificity of the mAbs in ELISA corresponded to the immunohistochemical staining (IHC-S) patterns performed on human AE and CE liver sections. Antigenic small particles designated as ''spems'' for E. multilocularis and ''spegs'' for E. granulosus s.l. were stained by the mAb EmG3_IgM, mAb EmG3_IgG1, mAb AgB, and mAb 2B2, while mAb Em2G11 reacted with spems and mAb Eg2 with spegs only. The laminated layer (LL) of both species was strongly visualized by using mAb EmG3_IgM, mAb EmG3_IgG1, mAb AgB, and mAb 2B2. The LL was specifically stained by mAb Em2G11 in E. multilocularis and by mAb Eg2 in E. granulosus s.l. In the germinal layer (GL), including the protoscoleces, a wide staining pattern with all structures of both species was observed with mAb EmG3_IgG1, mAb EmG3_IgM, mAb AgB, mAb 2B2, and mAb Em18. In the GL and protoscoleces, the mAb Eg2 displayed a strong E. granulosus s.l. specific binding, while mAb Em2G11 exhibited a weak granular E. multilocularis specific reaction. The most notable staining pattern in IHC-S was found with mAb Em18, which solely bound to the GL and protoscoleces of Echinococcus species and potentially to primary cells. To conclude, mAbs represent valuable tools for the visualization of major antigens in the most important Echinococcus species, as well as providing insights into parasite-host interactions and pathogenesis.

Keywords: 2B2; Echinococcus granulosus sensu lato; Echinococcus multilocularis; Em18; Em2; EmG3; II/3-10; antigen B.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antibodies, Monoclonal
Antigens, Helminth
Echinococcosis* / diagnosis
Echinococcus multilocularis*
Extracellular Vesicles*
Humans
Immunoglobulin M

Substances

Antibodies, Monoclonal
Antigens, Helminth
Immunoglobulin M

Grants and funding

Sources of funding included grant number: 173131 – “Transmission modelling of emergent echinococcosis in Kyrgyzstan” from the Swiss National Science Foundation (http://www.snf.ch). Co-applicant PD (main applicant Paul R. Torgerson, University of Zurich, Switzerland).