Abstract Mammalian olfactory receptor genes (ORs) are a diverse family of genes encoding proteins that directly interact with environmental chemical cues. ORs evolve via gene duplication in a birth-death fashion, neofunctionalizing and pseudogenizing over time. Olfaction is a primary sense used for food detection in plant-visiting bats, but the relationship between dietary specialization and OR repertoire diversity is unclear. Within neotropical Leaf-nosed bats (Phyllostomidae), many lineages are plant specialists, and some have a distinct OR repertoire compared to insectivorous species. Yet, whether specialization on particular plant genera is associated with the evolution of specialized, less diverse OR repertoires has never been tested. Using targeted sequence capture, we sequenced the OR repertoires of three sympatric species of short-tailed fruit bats (Carollia), which vary in their degree of specialization on the fruits of Piper plants. We characterized orthologous vs duplicated receptors among Carollia species, and explored the diversity and redundancy of the receptor gene repertoire. At the species level, the most dedicated Piper specialist, Carollia castanea, had lower OR diversity compared to the two generalists (C. sowelli and C. perspicillata), but we discovered a few unique sets of ORs within C. castanea with high redundancy of similar gene duplicates. These unique receptors potentially enable C. castanea to detect Piper fruit odorants better than its two congeners. Carollia perspicillata, the species with the most generalist diet, had a higher diversity of intact receptors, suggesting the ability to detect a wider range of odorant molecules. Variation among ORs may be a factor in the coexistence of these sympatric species, facilitating the exploitation of different plant resources. Our study sheds light on how gene duplication and changes in OR diversity may play a role in dietary adaptations and underlie ecological interactions between bats and plants.

Transcriptome-based exon capture approaches, along with next-generation sequencing, are allowing for the rapid and cost-effective production of extensive and informative phylogenomic datasets from non-model organisms for phylogenetics and population genetics research. These approaches generally employ a reference genome to infer the intron-exon structure of targeted loci and preferentially select longer exons. However, in the absence of an existing and well-annotated genome, we applied this exon capture method directly, without initially identifying intron-exon boundaries for bait design, to a group of highly diverse Haloniscus (Philosciidae), paraplatyarthrid and armadillid isopods, and examined the performance of our methods and bait design for phylogenetic inference. Here, we identified an isopod-specific set of single-copy protein-coding loci, and a custom bait design to capture targeted regions from 469 genes, and analysed the resulting sequence data with a mapping approach and newly-created post-processing scripts. We effectively recovered a large and informative dataset comprising both short (<100 bp) and longer (>300 bp) exons, with high uniformity in sequencing depth. We were also able to successfully capture exon data from up to 16-year-old museum specimens along with more distantly related outgroup taxa, and efficiently pool multiple samples prior to capture. Our well-resolved phylogenies highlight the overall utility of this methodological approach and custom bait design, which offer enormous potential for application to future isopod, as well as broader crustacean, molecular studies.

The compounding challenges of low signal, high background, and uncertain targets plague many metagenomic sequencing efforts. One solution has been DNA capture, wherein probes are designed to hybridize with target sequences, enriching them in relation to their background. However, balancing probe depth with breadth of capture is challenging for diverse targets. To find this balance, we have developed the HUBDesign pipeline, which makes use of sequence homology to design probes at multiple taxonomic levels. This creates an efficient probe set capable of simultaneously and specifically capturing known and related sequences. We validated HUBDesign by generating probe sets targeting the breadth of coronavirus diversity, as well as a suite of bacterial pathogens often underlying sepsis. In separate experiments demonstrating significant, simultaneous enrichment, we captured SARS-CoV-2 and HCoV-NL63 in a human RNA background and seven bacterial strains in human blood. HUBDesign (https://github.com/zacherydickson/HUBDesign) has broad applicability wherever there are multiple organisms of interest.

Seven years after the declaration of the first epidemic of Ebola virus disease in Guinea, the country faced a new outbreak—between 14 February and 19 June 2021—near the epicentre of the previous epidemic1,2. Here we use next-generation sequencing to generate complete or near-complete genomes of Zaire ebolavirus from samples obtained from 12 different patients. These genomes form a well-supported phylogenetic cluster with genomes from the previous outbreak, which indicates that the new outbreak was not the result of a new spillover event from an animal reservoir. The 2021 lineage shows considerably lower divergence than would be expected during sustained human-to-human transmission, which suggests a persistent infection with reduced replication or a period of latency. The resurgence of Zaire ebolavirus from humans five years after the end of the previous outbreak of Ebola virus disease reinforces the need for long-term medical and social care for patients who survive the disease, to reduce the risk of re-emergence and to prevent further stigmatization.

Significance Since its emergence in China in December 2019, SARS-CoV-2 has caused a global pandemic. Repurposing of FDA-approved drugs is a promising strategy for identifying rapidly deployable treatments for COVID-19. Herein, we developed a pipeline for quantitative, high-throughput, image-based screening of SARS-CoV-2 infection in human cells that led to the identification of several FDA-approved drugs and clinical candidates with in vitro antiviral activity. , The global spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and the associated disease COVID-19, requires therapeutic interventions that can be rapidly identified and translated to clinical care. Traditional drug discovery methods have a >90% failure rate and can take 10 to 15 y from target identification to clinical use. In contrast, drug repurposing can significantly accelerate translation. We developed a quantitative high-throughput screen to identify efficacious agents against SARS-CoV-2. From a library of 1,425 US Food and Drug Administration (FDA)-approved compounds and clinical candidates, we identified 17 hits that inhibited SARS-CoV-2 infection and analyzed their antiviral activity across multiple cell lines, including lymph node carcinoma of the prostate (LNCaP) cells and a physiologically relevant model of alveolar epithelial type 2 cells (iAEC2s). Additionally, we found that inhibitors of the Ras/Raf/MEK/ERK signaling pathway exacerbate SARS-CoV-2 infection in vitro. Notably, we discovered that lactoferrin, a glycoprotein found in secretory fluids including mammalian milk, inhibits SARS-CoV-2 infection in the nanomolar range in all cell models with multiple modes of action, including blockage of virus attachment to cellular heparan sulfate and enhancement of interferon responses. Given its safety profile, lactoferrin is a readily translatable therapeutic option for the management of COVID-19.

Alpine plant radiations are common across all major mountain systems of the world, and have been regarded as the main explanation for the species diversity found within these areas. To study the mechanisms behind the origin of this diversity, it is necessary to determine phylogenetic relationships and species boundaries in radiating alpine groups. The genus Dendrosenecio (Asteraceae) is an iconic example of a tropical-alpine plant radiation in the East African high mountains. To this date, limited sampling of molecular markers has resulted in insufficient phylogenetic resolution and infrageneric classification, hindering a comprehensive understanding of the drivers of diversification. Here, we used Hyb-Seq and the Compositae1061 probe set to generate targeted nuclear and off-target plastid DNA data for 42 samples representing all currently accepted 11 species. We combined coalescent methods and paralogy analysis to infer phylogenetic relationships, estimate divergence times and evaluate species boundaries. Lineage differentiation in Dendrosenecio seems to have occurred between the Late Miocene and the Pleistocene, starting when the first high elevation habitats became available in East Africa. We retrieved four major clades corresponding to four geographically distant mountain groups, testifying the importance of allopatric speciation in the early diversification of the group. Cytonuclear discordance suggested the occurrence of historical hybridization following occasional long-distance dispersal between mountain groups. The species delimitation analysis favored 10 species, but only five were fully supported, suggesting that population-level studies addressing processes such as ecological speciation and hybridization after secondary contact are needed to determine the current diversity found in the genus.

The tribe Senecioneae is one of the largest tribes in Asteraceae, with a nearly cosmopolitan distribution. Despite great efforts devoted to elucidate the evolution of Senecioneae, many questions still remain concerning the systematics of this group, from the tribal circumscription and position to species relationships in many genera. The hybridization-based target enrichment method of next-generation sequencing has been accepted as a promising approach to resolve phylogenetic problems. We herein develop a set of single-/low-copy genes for Senecioneae, and test their phylogenetic utilities. Our results demonstrate that these genes work highly efficiently for Senecioneae, with a high average gene recovery of 98.8% across the tribe and recovering robust phylogenetic hypotheses at different levels. In particular, the delimitation of the Senecioneae has been confirmed to include Abrotanella and exclude Doronicum, with the former sister to core Senecioneae and the latter shown to be more closely related to Calenduleae. Moreover, Doronicum and Calenduleae are inferred to be the closest relatives of Senecioneae, which is a new hypothesis well supported by statistical topology tests, morphological evidence, and the profile of pyrrolizidine alkaloids, a special kind of chemical characters generally used to define Senecioneae. Furthermore, this study suggests a complex reticulation history in the diversification of Senecioneae, accounting for the prevalence of polyploid groups in the tribe. With subtribe Tussilagininae s.str. as a case study showing a more evident pattern of gene duplication, we further explored reconstructing the phylogeny in the groups with high ploidy levels. Our results also demonstrate that tree topologies based on sorted paralogous copies are stable across different methods of phylogenetic inference, and more congruent with the morphological evidence and the results of previous phylogenetic studies.

Although least chipmunks (Neotamias minimus) are a widely distributed North American species of least concern, the southernmost population, N. m. atristriatus (Peñasco least chipmunk), is imperiled and a candidate for federal listing as a subspecies. We conducted a phylogeographic analysis across the N. minimus range to assess genomic differentiation and distinctiveness of the N. m. atristriatus population. Additionally, we leveraged the historical component of sampling to conduct a temporal analysis of N. minimus genetic diversity and also considered climate change effects on range persistence probability by projecting a species distribution model into the IPCC5 RCP 2.6 and 8.5 scenarios. We identified three geographically structured groups (West, North, and South) that were supported by both mitochondrial and nuclear data. N. m. atristriatus grouped within a unique South subclade but were not reciprocally monophyletic from N. m. operarius, and nuclear genome analyses did not separate N. m. atristriatus, N. m. caryi, and N. m. operarius. Thus, while least chipmunks in the Southwest represent an evolutionary significant unit, subspecies distinctions were not supported and listing of the Peñasco population as a Distinct Population Segment of N. m. operarius may be warranted. Our results also support consideration of populations with North and West mitogenomes as two additional evolutionary significant units. We found that N. minimus genetic diversity declined by 87% over the last century, and our models predicted substantial future habitat contraction, including the loss of the full contemporary ranges of N. m. atristriatus, N. m. arizonensis, and N. m. chuskaensis.

The red wolf (Canis rufus) of the eastern US was driven to near-extinction by colonial-era persecution and habitat conversion, which facilitated coyote (C. latrans) range expansion and widespread hybridization with red wolves. The observation of some gray wolf (C. lupus) ancestry within red wolves sparked controversy over whether it was historically a subspecies of gray wolf with its predominant “coyote-like” ancestry obtained from post-colonial coyote hybridization (2-species hypothesis) versus a distinct species closely related to the coyote that hybridized with gray wolf (3-species hypothesis). We analyzed mitogenomes sourced from before the 20th century bottleneck and coyote invasion, along with hundreds of modern amplicons, which led us to reject the 2-species model and to investigate a broader phylogeographic 3-species model suggested by the fossil record. Our findings broadly support this model, in which red wolves ranged the width of the American continent prior to arrival of the gray wolf to the mid-continent 60–80 ka; red wolves subsequently disappeared from the mid-continent, relegated to California and the eastern forests, which ushered in emergence of the coyote in their place (50–30 ka); by the early Holocene (12–10 ka), coyotes had expanded into California, where they admixed with and phenotypically replaced western red wolves in a process analogous to the 20th century coyote invasion of the eastern forests. Findings indicate that the red wolf pre-dated not only European colonization but human, and possibly coyote, presence in North America . These findings highlight the urgency of expanding conservation efforts for the red wolf.