Development and characterization of monoclonal antibodies against Besnoitia besnoiti tachyzoites

Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
García-Lunar, P;
GND
1019642998
Affiliation
Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Epidemiology,Greifswald-Insel Riems,Germany.
Schares, Gereon;
Affiliation
Inmunología y Genética Aplicada, SA (INGENASA),Calle de los Hermanos García Noblejas,41,28037,Madrid,Spain.
Sanz-Fernández, A;
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Jiménez-Meléndez, A;
Affiliation
Inmunología y Genética Aplicada, SA (INGENASA),Calle de los Hermanos García Noblejas,41,28037,Madrid,Spain.
García-Soto, I;
ORCID
0000-0001-5831-6970
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Regidor-Cerrillo, J;
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Pastor-Fernández, I;
ORCID
0000-0002-0622-2128
Affiliation
Institute of Parasitology,Vetsuisse Faculty,University of Bern,Länggass-Strasse 122,CH-3012 Bern,Switzerland.
Hemphill, A;
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Fernández-Álvarez, M;
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Ortega-Mora, L M;
ORCID
0000-0002-6627-7577
Affiliation
SALUVET,Animal Health Department,Faculty of Veterinary Sciences,Complutense University of Madrid,Ciudad Universitaria s/n,28040 Madrid,Spain.
Álvarez-García, G

This is the first report on the development and characterization of eight monoclonal antibodies (MABs) generated against whole- and membrane-enriched tachyzoite extracts of the apicomplexan parasite Besnoitia besnoiti. Confocal laser scanning immunofluorescence microscopy was used to localize respective epitopes in B. besnoiti tachyzoites along the lytic cycle. A pattern compatible with dense granule staining was observed with MABs 2.A.12, 2.F.3 and 2.G.4, which could be confirmed by immunogold electron microscopy for MABs 2.A.12 and 2.F.3. In particular, MABs 2.F.3 and 2.G.4 were secreted during early invasion, proliferation and egress phases. MABs 3.10.8 and 5.5.11 labelled the tachyzoite surface, whilst MABs 1.17.8, 8.9.2 and 2.G.A recognized the apical tip, which is reminiscent for microneme localization. Besides, the epitopes recognized by the latter two (MABs 8.9.2 and 2.G.A) exhibited a redistribution from the anterior part across the parasite surface towards the posterior end during invasion. Most MABs developed were genus-specific. Indeed, the MABs cross-reacted neither with T. gondii nor with N. caninum tachyzoites. In summary, we have generated MABs that will be useful to study the key processes in the lytic cycle of the parasite and with additional promising diagnostic value. However, the molecular identity of the antigens recognized remains to be elucidated.

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