A Monoallelic Deletion of the TcCRT Gene Increases the Attenuation of a Cultured Trypanosoma cruzi Strain,Protecting against an in Vivo Virulent Challenge

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. © 2014 sánchez-valdéz et al.