title: A monoallelic deletion of the TcCRT gene increases the attenuation of a cultured Trypanosoma cruzi strain, protecting against an in vivo virulent challenge.
abstract: Trypanosoma cruzi calreticulin (TcCRT) is a virulence factor that binds complement C1, thus inhibiting the activation of the classical complement pathway and generating pro-phagocytic signals that increase parasite infectivity. In a previous work, we characterized a clonal cell line lacking one TcCRT allele (TcCRT+/-) and another overexpressing it (TcCRT+), both derived from the attenuated TCC T. cruzi strain. The TcCRT+/- mutant was highly susceptible to killing by the complement machinery and presented a remarkable reduced propagation and differentiation rate both in vitro and in vivo. In this report, we have extended these studies to assess, in a mouse model of disease, the virulence, immunogenicity and safety of the mutant as an experimental vaccine. Balb/c mice were inoculated with TcCRT+/- parasites and followed-up during a 6-month period. Mutant parasites were not detected by sensitive techniques, even after mice immune suppression. Total anti-T. cruzi IgG levels were undetectable in TcCRT+/- inoculated mice and the genetic alteration was stable after long-term infection and it did not revert back to wild type form. Most importantly, immunization with TcCRT+/- parasites induces a highly protective response after challenge with a virulent T. cruzi strain, as evidenced by lower parasite density, mortality, spleen index and tissue inflammatory response. TcCRT+/- clones are restricted in two important properties conferred by TcCRT and indirectly by C1q: their ability to evade the host immune response and their virulence. Therefore, deletion of one copy of the TcCRT gene in the attenuated TCC strain generated a safe and irreversibly gene-deleted live attenuated parasite with high immunoprotective properties. Our results also contribute to endorse the important role of TcCRT as a T. cruzi virulence factor.
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Fernando J
authors.initials: FJ
authors.lastname: Sánchez-Valdéz
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Cecilia
authors.initials: C
authors.lastname: Pérez Brandán
authors.affiliation: Departamento de Medicina Preventiva Animal, Facultad de Ciencias Veterinarias y Pecuarias, Universidad de Chile, Santiago, Chile.
authors.forename: Galia
authors.initials: G
authors.lastname: Ramírez
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Alejandro D
authors.initials: AD
authors.lastname: Uncos
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: M Paola
authors.initials: MP
authors.lastname: Zago
authors.affiliation: Cátedra de Química Biológica, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Rubén O
authors.initials: RO
authors.lastname: Cimino
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Rubén M
authors.initials: RM
authors.lastname: Cardozo
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Jorge D
authors.initials: JD
authors.lastname: Marco
authors.affiliation: Programa Disciplinario de Inmunología, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Arturo
authors.initials: A
authors.lastname: Ferreira
authors.affiliation: Instituto de Patología Experimental-CONICET, Universidad Nacional de Salta, Salta, Argentina.
authors.forename: Miguel Ángel
authors.initials: MÁ
authors.lastname: Basombrío
mesh_terms: D000818:Animals; D037282:Calreticulin / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D006790:Host-Parasite Interactions / Q000235:genetics; D008297:Male; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D014774:Virulence / Q000235:genetics
chemical_list: D037282:Calreticulin; D015800:Protozoan Proteins