Cas9HF1 enhanced specificity in Ustilago maydis.

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system is widely used as a tool to precisely manipulate genomic sequence targeted by sgRNA (single guide RNA) and is adapted in different species for genome editing. One of the major concerns of CRISPR-Cas9 is the possibility of off-target effects, which can be remedied by the deployment of high fidelity Cas9 variants. Ustilago maydis is a maize fungal pathogen, which has served as a model organism for biotrophic pathogens for decades. The successful adaption of CRISPR-Cas9 in U. maydis greatly facilitated effector biology studies. Here, we constructed an U. maydis reporter strain that allows in vivo quantification of efficiency and target specificity of three high fidelity Cas9 variants, Cas9HF1, Cas9esp1.1 and Cas9hypa. This approach identified Cas9HF1 as most specific Cas9 variant in U. maydis. Furthermore, whole genome sequencing showed absence of off-target effects in U. maydis by CRISPR-Cas9 editing.

Interspecific hybridization impacts host range and pathogenicity of filamentous microbes.

Interspecific hybridization is widely observed within diverse eukaryotic taxa, and is considered an important driver for genome evolution. As hybridization fuels genomic and transcriptional alterations, hybrids are adept to respond to environmental changes or to invade novel niches. This may be particularly relevant for organisms that establish symbiotic relationships with host organisms, such as mutualistic symbionts, endophytes and pathogens. The latter group is especially well-known for engaging in everlasting arms races with their hosts. Illustrated by the increased identification of hybrid pathogens with altered virulence or host ranges when compared with their parental lineages, it appears that hybridization is a strong driver for pathogen evolution, and may thus significantly impact agriculture and natural ecosystems.