CRISPR technologies increasingly require spatiotemporal and dosage control of nuclease activity. One promising strategy involves linking nuclease activity to a cell’s transcriptional state by engineering guide RNAs (gRNAs) to function only after complexing with a ‘trigger’ RNA. However, standard gRNA switch designs do not allow independent selection of trigger and guide sequences, limiting gRNA switch application. Here, we demonstrate the modular design of Cas12a gRNA switches that decouples selection of these sequences. The 5 end of the Cas12a gRNA is fused to two distinct and nonoverlapping domains: one base pairs with the gRNA repeat, blocking formation of a hairpin required for Cas12a recognition; the other hybridizes to the RNA trigger, stimulating refolding of the gRNA repeat and subsequent gRNA-dependent Cas12a activity. Using a cell-free transcription-translation system and Escherichia coli, we show that designed gRNA switches can respond to different triggers and target different DNA sequences. Modulating the length and composition of the sensory domain altered gRNA switch performance. Finally, gRNA switches could be designed to sense endogenous RNAs expressed only under specific growth conditions, rendering Cas12a targeting activity dependent on cellular metabolism and stress. Our design framework thus further enables tethering of CRISPR activities to cellular states.

The detection of pathogens in clinical and environmental samples using high-throughput sequencing (HTS) is often hampered by large amounts of background information, which is especially true for viruses with small genomes. Enormous sequencing depth can be necessary to compile sufficient information for identification of a certain pathogen. Generic HTS combining with in-solution capture enrichment can markedly increase the sensitivity for virus detection in complex diagnostic samples.

Some tropical plant families, such as the Sapotaceae, have a complex taxonomy, which can be resolved using Next Generation Sequencing (NGS). For most groups however, methodological protocols are still missing. Here we identified 531 monocopy genes and 227 Short Tandem Repeats (STR) markers and tested them on Sapotaceae using target capture and NGS. The probes were designed using two genome skimming samples from Capurodendron delphinense and Bemangidia lowryi, both from the Tseboneae tribe, as well as the published Manilkara zapota transcriptome from the Sapotoideae tribe. We combined our probes with 261 additional ones previously published and designed for the entire angiosperm group. On a total of 792 low-copy genes, 638 showed no signs of paralogy and were used to build a phylogeny of the family with 231 individuals from all main lineages. A highly supported topology was obtained at high taxonomic ranks but also at the species level. This phylogeny revealed the existence of more than 20 putative new species. Single nucleotide polymorphisms (SNPs) extracted from the 638 genes were able to distinguish lineages within a species complex and to highlight geographical structuration. STR were recovered efficiently for the species used as reference (C. delphinense) but the recovery rate decreased dramatically with the phylogenetic distance to the focal species. Altogether, the new loci will help reaching a sound taxonomic understanding of the family Sapotaceae for which many circumscriptions and relationships are still debated, at the species, genus and tribe levels.

The Opiliones superfamily Triaenonychoidea currently includes two families, the monogeneric New Zealand–endemic Synthetonychiidae Forster, 1954 and Triaenonychidae Sørensen, 1886, a diverse family distributed mostly throughout the temperate Gondwanan terranes, with ~110 genera and ~500 species and subspecies currently described. Traditionally, Triaenonychidae has been divided into subfamilies diagnosed by very few morphological characters largely derived from the troublesome ‘Roewerian system’ of morphology, and classifications based on this system led to many complications. Recent research within Triaenonychoidea using morphology and traditional multilocus data has shown multiple deeply divergent lineages, non-monophyly of Triaenonychidae, and non-monophyly of subfamilies, necessitating a revision based on phylogenomic data. We used sequence capture of ultraconserved elements across 164 samples to create a 50% taxon occupancy matrix with 704 loci. Using phylogenomic and morphological examinations, we explored family-level relationships within Triaenonychoidea, including describing two new families: (1) Lomanellidae Mendes & Derkarabetian, fam. nov., consisting of Lomanella Pocock, 1903, and a newly described genus Abaddon Derkarabetian & Baker, gen. nov. with one species, A. despoliator Derkarabetian, sp. nov.; and (2) the elevation to family of Buemarinoidae Karaman, 2019, consisting of Buemarinoa Roewer, 1956, Fumontana Shear, 1977, Flavonuncia Lawrence, 1959, and a newly described genus Turonychus Derkarabetian, Prieto & Giribet, gen. nov., with one species, T. fadriquei Derkarabetian, Prieto & Giribet, sp. nov. With our dataset we also explored phylogenomic relationships within Triaenonychidae with an extensive taxon set including samples representing ~80% of the genus-level diversity. Based on our results we (1) discuss systematics of this family including the historical use of subfamilies, (2) reassess morphology in the context of our phylogeny, (3) hypothesise placement for all unsampled genera, (4) highlight lineages most in need of taxonomic revision, and (5) provide an updated species-level checklist. Aside from describing new taxa, our study provides the phylogenomic context necessary for future evolutionary and systematic research across this diverse lineage. ZooBank Registration: urn:lsid:zoobank.org:pub:81683834-98AB-43AA-B25A-C28C6A404F41

Genomic resources are under-developed for rodents, including well-studied species such as prairie dogs (Cynomys spp.). We conducted whole-genome resequencing on 10 Gunnison’s prairie dogs (C. gunnisoni, GUPD) and identified 12,842,055 high-quality SNPs, from which four sets of bait sequences were created. We designed two sets each (containing either 20k or 60k baits) for projects using contemporary (120 bp baits) or historical (100 bp baits) DNA. These bait sets can be used to study a variety of ecological and evolutionary questions in prairie dogs and other ground squirrel taxa.

Meiotic recombination is a critical process for plant breeding, as it creates novel allele combinations that can be exploited for crop improvement. In wheat, a complex allohexaploid that has a diploid-like behaviour, meiotic recombination between homoeologous or alien chromosomes is suppressed through the action of several loci. Here, we report positional cloning of Pairing homoeologous 2 (Ph2) and functional validation of the wheat DNA mismatch repair protein MSH7-3D as a key inhibitor of homoeologous recombination, thus solving a half-century-old question. Similar to ph2 mutant phenotype, we show that mutating MSH7-3D induces a substantial increase in homoeologous recombination (up to 5.5 fold) in wheat-wild relative hybrids, which is also associated with a reduction in homologous recombination. These data reveal a role for MSH7-3D in meiotic stabilisation of allopolyploidy and provides an opportunity to improve wheat’s genetic diversity through alien gene introgression, a major bottleneck facing crop improvement.

Understanding diversity has been a pursuit in evolutionary biology since its inception. A challenge arises when sexual selection has played a role in diversification. Questions of what constitutes a ‘species’, homoplasy vs. synapomorphy, and whether sexually selected traits show phylogenetic signal have hampered work on many systems. Peacock spiders are famous for sexually selected male courtship dances and peacock-like abdominal ornamentation. This lineage of jumping spiders currently includes over 90 species classified into two genera, Maratus and Saratus. Most Maratus species have been placed into groups based on secondary sexual characters, but evolutionary relationships remain unresolved. Here we assess relationships in peacock spiders using phylogenomic data (ultraconserved elements and RAD-sequencing). Analyses reveal that Maratus and the related genus Saitis are paraphyletic. Many, but not all, morphological groups within a ‘core Maratus’ clade are recovered as genetic clades but we find evidence for undocumented speciation. Based on original observations of male courtship, our comparative analyses suggest that courtship behaviour and peacock-like abdominal ornamentation have evolved sequentially, with some traits inherited from ancestors and others evolving repeatedly and independently from ‘simple’ forms. Our results have important implications for the taxonomy of these spiders, and provide a much-needed evolutionary framework for comparative studies of the evolution of sexual signal characters.

In climates with seasonally limited precipitation, terrestrial animals congregate at high densities at scarce water sources. We hypothesize that viruses can exploit the recurrence of these diverse animal congregations to spread. In this paper, we test the central prediction of this hypothesis — that viruses employing this transmission strategy remain stable and infectious in water. Equid herpesviruses (EHVs) were chosen as a model as they have been shown to remain stable and infectious in water for weeks under laboratory conditions. Using fecal data from wild equids from a previous study, we establish that EHVs are shed more frequently by their hosts during the dry season, increasing the probability of water source contamination with EHV. We document the presence of several strains of EHVs present in high genome copy number from the surface water and sediments of waterholes sampled across a variety of mammalian assemblages, locations, temperatures and pH. Phylogenetic analysis reveals that the different EHV strains found exhibit little divergence despite representing ancient lineages. We employed molecular approaches to show that EHVs shed remain stable in waterholes with detection decreasing with increasing temperature in sediments. Infectivity experiments using cell culture reveals that EHVs remain infectious in water derived from waterholes. The results are supportive of water as an abiotic viral vector for EHV.

Sex determination directs development as male or female in sexually reproducing organisms. Evolutionary transitions in sex determination have occurred frequently, suggesting simple mechanisms behind the transitions, yet their detail remains elusive. Here we explore the links between mechanisms of transitions in sex determination and sex chromosome evolution at both recent and deeper temporal scales (<1 Myr; ~79 Myr). We studied a rare example of a species with intraspecific variation in sex determination, Carinascincus ocellatus, and a relative, Liopholis whitii, using c-banding and mapping of repeat motifs and a custom Y chromosome probe set to identify the sex chromosomes. We identified both unique and conserved regions of the Y chromosome among C. ocellatus populations differing in sex determination. There was no evidence for homology of sex chromosomes between C. ocellatus and L. whitii, suggesting independent evolutionary origins. We discuss sex chromosome homology between members of the subfamily Lygosominae and propose links between sex chromosome evolution, sex determination transitions, and karyotype evolution.