A monoallelic deletion of the TcCRT gene increases the attenuation of a cultured Trypanosoma cruzi strain, protecting against an in vivo virulent challenge.

A monoallelic deletion of the TcCRT gene increases the attenuation of a cultured Trypanosoma cruzi strain, protecting against an in vivo virulent challenge.

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Trypanosoma cruzi calreticulin (TcCRT) is a virulence factor that binds complement C1, thus Vase (Set of 2) 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 Swimwear 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.