The apicomplexan Cryptosporidium parvum is an intestinal parasite that affects healthy humans and animals, and causes an unrelenting infection in immunocompromised individuals such as AIDS patients. We report the complete genome sequence of C. parvum, type II isolate. Genome analysis identifies extremely streamlined metabolic pathways and a reliance on the host for nutrients. In contrast to Plasmodium and Toxoplasma, the parasite lacks an apicoplast and its genome, and possesses a degenerate mitochondrion that has lost its genome. Several novel classes of cell-surface and secreted proteins with a potential role in host interactions and pathogenesis were also detected. Elucidation of the core metabolism, including enzymes with high similarities to bacterial and plant counterparts, opens new avenues for drug development.
We have expanded the livestock gene editing toolbox to include transcription activator-like (TAL) effector nuclease (TALEN)-and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-stimulated homology-directed repair (HDR) using plasmid, rAAV, and oligonucleotide templates. Toward the genetic dehorning of dairy cattle, we introgressed a bovine POLLED allele into horned bull fibroblasts. Single nucleotide alterations or small indels were introduced into 14 additional genes in pig, goat, and cattle fibroblasts using TALEN mRNA and oligonucleotide transfection with efficiencies of 10-50% in populations. Several of the chosen edits mimic naturally occurring performance-enhancing or disease-resistance alleles, including alteration of single base pairs. Up to 70% of the fibroblast colonies propagated without selection harbored the intended edits, of which more than onehalf were homozygous. Edited fibroblasts were used to generate pigs with knockout alleles in the DAZL and APC genes to model infertility and colon cancer. Our methods enable unprecedented meiosis-free intraspecific and interspecific introgression of select alleles in livestock for agricultural and biomedical applications.
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