Compact CRISPR/Cas9 systems that can be delivered by AAV for in vivo genome editing hold great promise for clinical applications. Brevibacillus laterosporus Cas9 (BlatCas9) is a compact Cas9 nuclease that has been identified for plant genome editing. Here, we characterize BlatCas9 as an alternative tool for mammalian genome editing. We demonstrate that BlatCas9 prefers a N4CNAA protospacer adjacent motif (PAM), but N4C PAM is also editable in mammalian cells. We next demonstrate that BlatCas9 enables genome editing in a variety of cell types. Furthermore, BlatCas9 can be packaged into AAV for genome editing. Finally, we characterize the specificity of BlatCas9. In summary, BlatCas9 offers an alternative tool for both basic research and clinical applications.

Root exudates play a critical role in root–microbe interactions. Agrobacterium tumefaciens causes crown gall disease in multiple plant species, but rose root exudate-mediated inhibition of Agrobacterium in the rhizosphere is poorly understood. In this study, the influence of preinoculation with beneficial bacteria or pathogens on root exudates and subsequent colonization by A. tumefaciens was investigated in a split-root system. We found that preinoculation of rose plants in a split-root system with Bacillus velezensis CLA178 or A. tumefaciens C58 inhibited subsequent colonization by C58. Root secretion of valine had positive effects on the chemotaxis, biofilm formation, colonization of C58, and crown gall disease severity, but valine secretion decreased significantly when Rosa multiflora plants were preinoculated with CLA178 or C58. These results indicated that rose plants reduced root secretion of valine in response to microbial colonization, thereby reducing the colonization of Agrobacterium colonization and disease severity. This study provides new insights into the root exudate-mediated interactions of rose plants, B. velezensis, and A. tumefaciens and proposes a potential way to control crown gall disease.

The pine shoot beetle, Tomicus yunnanensis (Curculionidae: Scolytinae), is a significant trunk-boring pest of Pinus yunnanensis forests, causing extensive ecological degradation through direct feeding and its role as a vector for destructive pathogenic fungi. The scarcity of a high-quality reference genome has hindered in-depth studies of its biology, particularly the mechanisms underlying its pathogen transmission and host adaptation. Here, we report a chromosome-level genome assembly of T. yunnanensis. Using a combination of PacBio long-read sequencing, Illumina short-read sequencing, and Hi-C chromatin interaction mapping, we generated a final genome assembly of 379.3 Mb, which was anchored into 14 linkage pseudo-chromosomes, with scaffold N50 value of 21.6 Mb. A total of 12, 941 protein coding genes were predicted and annotated. BUSCO completeness assessment indicated that the completeness of the assembly and annotation reached 96.4% and 95.3%, respectively. This high-quality genome provides a critical resource for future research on the pest's biology, ecological adaptation, management and will ultimately support the development of targeted strategies to control its damage and disrupt its role in pathogen dissemination.