O uso de sistemas de edição genômica para avaliação de potenciais fatores de virulência dos protozoários neospora caninum e toxoplasma gondii

Abstract

Neospora caninum is a parasite of veterinary relevance, inducing severe disease in dogs and reproductive disorders in ruminants, mainly cattle, leading to significant economic losses. The close phylogenetic relationship with Toxoplasma gondii and the lack of pathogenicity in humans drive the interest of the scientific community in using N. caninum as a model to study the virulence of T. gondii. To enable this comparison, it is important to develop efficient molecular tools for N. caninum, to gain precision and save time in genetic manipulation protocols. The development of molecular genetics has greatly improved the study of the biology and pathology associated with parasites of the phylum Apicomplexa. Although molecular tools are highly developed for T. gondii, N. caninum lacks efficient tools for genetic manipulation. Here, we developed basic strains and protocols using the genomic reference strain of N. caninum to allow efficient knockout and knockdown assays in this model. Using these tools, we mapped the protein content of N. caninum dense granules, described proteins exclusive to N. caninum, and compared different targets with orthologs in T. gondii, applying genetically altered parasites in established models to measure the host-parasite relationship, with the intention of observing the immune responses induced in vitro and in vivo and to search for virulence factors that may be amenable to common intervention.