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Eimeria species are pathogenic protozoa with a wide range of hosts and the cause of poultry coccidiosis, which results in huge economic losses to the poultry industry. These parasites encode a genome of ∼8000 genes that control a highly coordinated life cycle of asexual replication and sexual differentiation, transmission, and virulence. However, the function and physiological importance of the large majority of these genes remain unknown mostly due to the lack of tools for systematic analysis of gene functions. Here, we report the first application of CRISPR-Cas9 gene editing technology in Eimeria tenella for analysis of gene function at a single gene level as well as for systematic functional analysis of an entire gene family. Using a transgenic line constitutively expressing Cas9, we demonstrated successful and efficient loss of function through non-homologous end joining as well as guided homologous recombination. Application of this approach to the study of the localization of EtGRA9 revealed that the gene encodes a secreted protein whose cellular distribution varied during the life cycle. Systematic disruption of the ApiAp2 transcription factor gene family using this approach revealed that 23 of the 33 factors expressed by this parasite are essential for development and survival in the host. Our data thus establish CRISPR-Cas9 as a powerful technology for gene editing in Eimeria and will set the stage for systematic functional analysis of its genome to understand its biology and pathogenesis, and will make it possible to identify and validate new targets for coccidiosis therapy.

Apicomplexan parasites are protozoan organisms that are characterised by complex life cycles and they include medically important species, such as the malaria parasite Plasmodium and the causative agents of toxoplasmosis (Toxoplasma gondii) and cryptosporidiosis (Cryptosporidium spp.). Apicomplexan parasites can infect one or more hosts, in which they differentiate into several morphologically and metabolically distinct life cycle stages. These developmental transitions rely on changes in gene expression. In the last few years, the important roles of different members of the ApiAP2 transcription factor family in regulating life cycle transitions and other aspects of parasite biology have become apparent. Here, we review recent progress in our understanding of the different members of the ApiAP2 transcription factor family in apicomplexan parasites.

Abstract: Malaria is caused by protozoan parasites of the genus Plasmodium and involves infection of multiple hosts and cell types during the course of an infection. To complete its complex life cycle the parasite requires strict control of gene regulation for survival and successful propagation. Thus far, the Apicomplexan AP2 (ApiAP2) family of DNA-binding proteins is the sole family of proteins to have surfaced as candidate transcription factors in all apicomplexan species. Work from several laboratories is beginning to shed light on how the ApiAP2 proteins from Plasmodium spp. contribute to the regulation of gene expression at various stages of parasite development. Here we highlight recent progress toward understanding the role of Plasmodium ApiAP2 proteins in DNA related regulatory processes including transcriptional regulation and gene silencing. [ABSTRACT FROM AUTHOR]

Apicomplexan parasites are protozoan organisms that are characterised by complex life cycles and they include medically important species, such as the malaria parasite Plasmodium and the causative agents of toxoplasmosis (Toxoplasma gondii) and cryptosporidiosis (Cryptosporidium spp.). Apicomplexan parasites can infect one or more hosts, in which they differentiate into several morphologically and metabolically distinct life cycle stages. These developmental transitions rely on changes in gene expression. In the last few years, the important roles of different members of the ApiAP2 transcription factor family in regulating life cycle transitions and other aspects of parasite biology have become apparent. Here, we review recent progress in our understanding of the different members of the ApiAP2 transcription factor family in apicomplexan parasites. [ABSTRACT FROM AUTHOR]