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CRISPR-Cas systems defend prokaryotic cells from invasive DNA of viruses, plasmids and other mobile genetic elements. Here, we show using metagenomics, metatranscriptomics and single-cell genomics that CRISPR systems of widespread, uncultivated archaea can also target chromosomal DNA of archaeal episymbionts of the DPANN superphylum. Using meta-omics datasets from Crystal Geyser and Horonobe Underground Research Laboratory, we find that CRISPR spacers of the hosts Candidatus Altiarchaeum crystalense and Ca. A. horonobense, respectively, match putative essential genes in their episymbionts' genomes of the genus Ca. Huberiarchaeum and that some of these spacers are expressed in situ. Metabolic interaction modelling also reveals complementation between host-episymbiont systems, on the basis of which we propose that episymbionts are either parasitic or mutualistic depending on the genotype of the host. By expanding our analysis to 7,012 archaeal genomes, we suggest that CRISPR-Cas targeting of genomes associated with symbiotic archaea evolved independently in various archaeal lineages.

Abstract Many CRISPR-Cas immune systems generate guide (g)RNAs using trans-activating CRISPR RNAs (tracrRNAs). Recent work revealed that Cas9 tracrRNAs could be reprogrammed to convert any RNA-of-interest into a gRNA, linking the RNA’s presence to Cas9-mediated cleavage of double-stranded (ds)DNA. Here, we reprogram tracrRNAs from diverse Cas12 nucleases, linking the presence of an RNA-of-interest to dsDNA cleavage and subsequent collateral single-stranded DNA cleavage—all without the RNA necessarily encoding a protospacer-adjacent motif (PAM). After elucidating nuclease-specific design rules, we demonstrate PAM-independent RNA detection with Cas12b, Cas12e, and Cas12f nucleases. Furthermore, rationally truncating the dsDNA target boosts collateral cleavage activity, while the absence of a gRNA reduces background collateral activity and enhances sensitivity. Finally, we apply this platform to detect 16 S rRNA sequences from five different bacterial pathogens using a universal reprogrammed tracrRNA. These findings extend tracrRNA reprogramming to diverse dsDNA-targeting Cas12 nucleases, expanding the flexibility and versatility of CRISPR-based RNA detection.

Multiple peptide resistance factor (MprF) confers resistance to cationic antimicrobial peptides (AMPs) in several pathogens, thereby enabling evasion of the host immune response. The role of MprF in commensals remains, however, uncharacterized. To close this knowledge gap, we used a common gut commensal of animals, Lactiplantibacillus plantarum, and its natural host, the fruit fly Drosophila melanogaster, as an experimental model to investigate the role of MprF in commensal-host interactions. The L. plantarum ΔmprF mutant that we generated exhibited deficiency in the synthesis of lysyl-phosphatidylglycerol (Lys-PG), resulting in increased negative cell surface charge and increased susceptibility to AMPs. Susceptibility to AMPs had no effect on ΔmprF mutant's ability to colonize guts of uninfected flies. However, we observed significantly reduced abundance of the ΔmprF mutant after infection-induced inflammation in the guts of wild-type flies but not of flies lacking AMPs. Additionally, we found that the ΔmprF mutant compared to wild-type L. plantarum induces a stronger intestinal immune response in flies due to the increased release of immunostimulatory peptidoglycan fragments, indicating an important role of MprF in promoting host tolerance to commensals. Our further analysis suggests that MprF-mediated lipoteichoic acid modifications are involved in host immunomodulation. Overall, our results demonstrate that MprF, besides its well-characterized role in pathogen immune evasion and virulence, is also an important commensal resilience factor.

Abstract Alkaline polymer membrane electrochemical energy conversion devices offer the prospect of using non-platinum group catalysts. However, their cationic functionalities are currently not sufficiently stable for vapor-phase applications, such as fuel cells. Herein, we report 1,6-diazabicyclo[4.4.4]tetradecan-1,6-ium (in-DBD), a cationic proton cage, that is orders of magnitude more resistant to hydroxide-promoted degradation than state-of-the-art organic cations under ultra-dry conditions and elevated temperature, and the first organic cation-hydroxide to persist at critically low hydration levels (

Abstract Understanding how tropical forests respond to abiotic environmental changes is critical for preserving biodiversity, mitigating climate change, and maintaining ecosystem services in the coming century. To evaluate the relative roles of the abiotic environment and human disturbance on Central African tree community composition, we employ tree inventory data, remotely sensed climatic data, and soil nutrient data collected from 30 1‐ha plots distributed across a large‐scale observational experiment in forests that had been differently impacted by logging and hunting in northern Republic of Congo. We show that the composition of Afrotropical plant communities at this scale responds to human disturbance more than to climate, with particular sensitivities to hunting and distance to the nearest village (a proxy for other human activities, including tree‐cutting and gathering). These findings contrast neotropical predictions, highlighting the unique ecological, evolutionary, and anthropogenic history of Afrotropical forests.

Abstract CRISPR interference (CRISPRi) is the leading technique to silence gene expression in bacteria; however, design rules remain poorly defined. We develop a best-in-class prediction algorithm for guide silencing efficiency by systematically investigating factors influencing guide depletion in genome-wide essentiality screens, with the surprising discovery that gene-specific features substantially impact prediction. We develop a mixed-effect random forest regression model that provides better estimates of guide efficiency. We further apply methods from explainable AI to extract interpretable design rules from the model. This study provides a blueprint for predictive models for CRISPR technologies where only indirect measurements of guide activity are available.

CRISPR technologies comprising a Cas nuclease and a guide RNA (gRNA) can utilize multiple gRNAs to enact multi-site editing or regulation in the same cell. Nature devised a highly compact means of encoding gRNAs in the form of CRISPR arrays composed of conserved repeats separated by targeting spacers. However, the capacity to acquire new spacers keeps the arrays longer than necessary for CRISPR technologies. Here, we show that CRISPR arrays utilized by the Cas9 nuclease can be shortened without compromising and sometimes even enhancing targeting activity. Using multiplexed gene repression in E. coli, we found that each region could be systematically shortened to varying degrees before severely compromising targeting activity. Surprisingly, shortening some spacers yielded enhanced targeting activity, which was linked to folding of the transcribed array prior to processing. Overall, shortened CRISPR-Cas9 arrays can facilitate multiplexed editing and gene regulation from a smaller DNA footprint across many bacterial applications of CRISPR technologies.

Despite its importance for forest regeneration, food webs, and human economies, changes in tree fecundity with tree size and age remain largely unknown. The allometric increase with tree diameter assumed in ecological models would substantially overestimate seed contributions from large trees if fecundity eventually declines with size. Current estimates are dominated by overrepresentation of small trees in regression models. We combined global fecundity data, including a substantial representation of large trees. We compared size-fecundity relationships against traditional allometric scaling with diameter and two models based on crown architecture. All allometric models fail to describe the declining rate of increase in fecundity with diameter found for 80% of 597 species in our analysis. The strong evidence of declining fecundity, beyond what can be explained by crown architectural change, is consistent with physiological decline. A downward revision of projected fecundity of large trees can improve the next generation of forest dynamic models.

Abstract CRISPR-based gene perturbation enables unbiased investigations of single and combinatorial genotype-to-phenotype associations. In light of efforts to map combinatorial gene dependencies at scale, choosing an efficient and robust CRISPR-associated (Cas) nuclease is of utmost importance. Even though SpCas9 and AsCas12a are widely used for single, combinatorial, and orthogonal screenings, side-by-side comparisons remain sparse. Here, we systematically compared combinatorial SpCas9, AsCas12a, and CHyMErA in hTERT-immortalized retinal pigment epithelial cells and extracted performance-critical parameters for combinatorial and orthogonal CRISPR screens. Our analyses identified SpCas9 to be superior to enhanced and optimized AsCas12a, with CHyMErA being largely inactive in the tested conditions. Since AsCas12a contains RNA processing activity, we used arrayed dual-gRNAs to improve AsCas12a and CHyMErA applications. While this negatively influenced the effect size range of combinatorial AsCas12a applications, it enhanced the performance of CHyMErA. This improved performance, however, was limited to AsCas12a dual-gRNAs, as SpCas9 gRNAs remained largely inactive. To avoid the use of hybrid gRNAs for orthogonal applications, we engineered the multiplex SpCas9-enAsCas12a approach (multiSPAS) that avoids RNA processing for efficient orthogonal gene editing.

Genome editing in bacteria normally requires efficient recombination and high transformation efficiencies, which often isn’t. Here the authors report that systematically attenuating DNA targeting activity enables RecA-mediated repair in different bacteria, allowing chromosomal cleavage to drive editing.

Onchocerca lupi (Rodonaja, 1967) is an understudied, vector-borne, filarioid nematode that causes ocular onchocercosis in dogs, cats, coyotes, wolves, and is also capable of infecting humans. Onchocercosis in dogs has been reported with increasing incidence worldwide. However, despite the growing number of reports describing canine O. lupi cases as well as zoonotic infections globally, the disease prevalence in endemic areas and vector species of this parasite remains largely unknown. Here, our study aimed to identify the occurrence of O. lupi infected dogs in northern Arizona, New Mexico, and Utah, United States and identify the vector of this nematode. A total of 532 skin samples from randomly selected companion animals with known geographic locations within the Navajo Reservation were collected and molecularly surveyed by PCR for the presence of O. lupi DNA (September 2019–June 2022) using previously published nematode primers (COI) and DNA sequencing. O. lupi DNA was detected in 50 (9.4%) sampled animals throughout the reservation. Using positive animal samples to target geographic locations, pointed hematophagous insect trapping was performed to identify potential O. lupi vectors. Out of 1,922 insects screened, 38 individual insects and 19 insect pools tested positive for the presence of O. lupi, all of which belong to the Diptera family. This increased surveillance of definitive host and biological vector/intermediate host is the first large scale prevalence study of O. lupi in companion animals in an endemic area of the United States, and identified an overall prevalence of 9.4% in companion animals as well as multiple likely biological vector and putative vector species in the southwestern United States. Furthermore, the identification of these putative vectors in close proximity to human populations coupled with multiple, local zoonotic cases highlight the One Health importance of O. lupi.

Wearable, disposable, implantable and lightweight electronic devices all require low power consumption, typically less than 3 V. Polymerized ionic liquids represent an emerging class of dielectric materials which can enable low voltage organic thin film transistor operation (OTFT) while in the solid state, reducing the potential for toxic or flammable electrolyte leaks. Here we show that bottom gate bottom contact OTFTs were successfully fabricated using a library of 12 poly-(styrene)-b-poly(1-(4-vinylbenzyl)-3-butylimidazolium-r-poly(ethylene glycol) methyl ether methacrylate) block copolymers with varying VBBI+ and PEGMA ratios, and can achieve OTFTs that turn on between 0 and 1 V. We demonstrate that the poly(ionic liquid) microstructure dictates the self-assembly which correlates to OTFT performance. Notably, we indicate that changing the surface chemistry of the substrate the poly(ionic liquid) is cast on is less significant in driving block copolymer self-assembly compared to block ratios and the counter ion. We also report that the semiconductor crystalline domains are similar when deposited on the poly(ionic liquids), however the films differ in terms of semiconductor molecular order in the amorphous domains. Overall, we demonstrate that device performance is primarily linked to the poly(ionic liquid) self-assembly, while also highlighting the importance of both the crystalline and amorphous domains in the semiconductor structure of thin films. [ABSTRACT FROM AUTHOR]

The spin–orbit alignment of binary stars traces their formation and accretion history. Previous studies of spin–orbit alignment have been limited to small samples, slowly rotating solar-type stars, and/or wide visual binaries that not surprisingly manifest random spin–orbit orientations. We analyze 917 Gaia astrometric binaries across periods P = 100–3000 days ( a = 0.5–5 au) that have B8-F1 IV/V primaries ( M _1 = 1.5–3 M _⊙ ) and measured projected rotational velocities v sin i . The primary stars in face-on orbits exhibit substantially smaller v sin i compared to those in edge-on orbits at the 6 σ level, demonstrating significant spin–orbit alignment. The primaries in our astrometric binaries are rotating more slowly than their single-star or wide-binary counterparts and therefore comprise the slow-rotator population in the observed bimodal rotational velocity distribution of early-type stars. We discuss formation models of close binaries where some of the disk angular momentum is transferred to the orbit and/or secondary spin, quenching angular momentum flow to the primary spin. The primaries in astrometric binaries with small mass ratios q = M _2 / M _1 < 0.3 possess even smaller v sin i , consistent with model predictions. Meanwhile, astrometric binaries with large eccentricities e > 0.4 do not display spin–orbit alignment or spin reduction. Using a Monte Carlo technique, we measure a spin–orbit alignment fraction of F _align = 75% ± 5% and an average spin reduction factor of 〈 S _align 〉 = 0.43 ± 0.04. We conclude that 75% of close A-type binaries likely experienced circumbinary disk accretion and probably formed via disk fragmentation and inward disk migration. The remaining 25%, mostly those with e > 0.4, likely formed via core fragmentation and orbital decay via dynamical friction.

Organic solar cells (OSCs) have long promised to provide renewable energy in a scalable, cost-effective way; however, for years, their relatively low efficiency has been a significant barrier to commercialization. Recent progress on cell efficiency means that OSCs are now much more competitive with other established technologies. These key advancements have come from better understanding and controlling the molecular structure, solid-state packing, and film morphology of the light absorbing layer. This focused review will explore the different ways that the solid-state structure and film morphology of the light absorbing layer can be controlled. It will examine the key features of an efficient light absorbing layer and present guiding principles for creating efficient OSCs. The future directions and remaining research questions of this field will be briefly discussed. Key words: organic photovoltaics, organic solar cells, non-fullerene acceptors, film morphology, organic electronics. Les cellules solaires organiques (CSO) ont longtemps représenté la promesse d'une énergie renouvelable qui soit rentable et exploitable à grande échelle. Cependant, pendant des années, leur faible efficacité a été un frein important à leur commercialisation. Les récents progrès sur le plan de l'efficacité des CSO montrent qu'elles sont maintenant en bien meilleure position pour concurrencer d'autres technologies établies. Ces importants progrès tiennent à une meilleure compréhension et à une meilleure maîtrise de la structure moléculaire, de l'empilement à l'état solide et de la morphologie des films de la couche absorbant la lumière. Dans le cadre de cette revue ciblée, nous explorerons les différentes façons de diriger la structure à l'état solide et la morphologie des films de la couche absorbant la lumière. Nous examinerons les caractéristiques clés d'une couche absorbant efficacement la lumière et nous présenterons les principes directeurs pour créer des CSO efficaces. Nous discuterons brièvement des orientations futures et des questions de recherche qui restent à élucider dans ce domaine. [Traduit par la Rédaction] Mots-clés: photovoltaïque organique, cellules solaires organiques, accepteurs non-fullerène, morphologie des films, électronique organique., Introduction Organic solar cells (OSCs) are an established technology that is recently experiencing a renaissance due to the development of new classes of materials and a deeper understanding of device [...]

Technologies that can halt the spread of gene drives would be highly useful in controlling or reverting their effect. Here the authors use the anti-CRISPR protein AcrIIA4 to inactivate drives in A. gambiae.