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Research on Cas9 nucleases from different organisms holds great promise for advancing genome engineering and gene therapy tools, as it could provide novel structural insights into CRISPR editing mechanisms, expanding its application area in biology and medicine. The subclass of thermophilic Cas9 nucleases is actively expanding due to the advances in genome sequencing allowing for the meticulous examination of various microorganisms' genomes in search of the novel CRISPR systems. The most prominent thermophilic Cas9 effectors known to date are GeoCas9, ThermoCas9, IgnaviCas9, AceCas9, and others. These nucleases are characterized by a varying temperature range of the activity and stringent PAM preferences; thus, further diversification of the naturally occurring thermophilic Cas9 subclass presents an intriguing task. This study focuses on generating a construct to express a compact Cas9 nuclease (AnoCas9) from the thermophilic microorganism Anoxybacillus flavithermus displaying the nuclease activity in the 37–60 °C range and the PAM preference of 5′-NNNNCDAA-3′ in vitro. Here, we highlight the close relation of AnoCas9 to the GeoCas9 family of compact thermophilic Cas9 effectors. AnoCas9, beyond broadening the repertoire of Cas9 nucleases, suggests application in areas requiring the presence of thermostable CRISPR/Cas systems in vitro, such as sequencing libraries' enrichment, allele-specific isothermal PCR, and others. [ABSTRACT FROM AUTHOR]

Cuscuta campestris Yunck. is a stem holoparasitic flowering plant, wide-spread and currently considered invasive worldwide. It has both ecological and agricultural significance. However, little is known on how the conditions of the environment, and abiotic stresses in particular, could affect the success of this parasite. A total of 22 potential host plants, belonging to 10 species and 4 families, were tested as C. campestris hosts under different saline regimes. The changes in survival rate (number of successful infections) and growth (as cm day−1) of the parasite were studied. Salinity did significantly change the survival and growth rate of C. campestris , which effect was species- and cultivar-dependent. While generally the survival of the parasite decreased with increasing salinization, the opposite effect was observed on several host plants. However, survival rate did not always coincide with higher growth rate. This suggests that salinity produces different effects on the parasite depending on the host plant and these depend strongly not only on the species, but also on the cultivar. These results strongly suggest that under salt stress the suitability of various crop plants as hosts for C. campestris may change significantly, thus affecting the overall success of the parasite. In the light of the increased salinization of soils, this may have a significant effect on the spread of C. campestris and its agricultural impact. [ABSTRACT FROM AUTHOR]