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Gene expression can be highly plastic in response to environmental variation. However, we know little about how expression plasticity is shaped by natural selection and evolves in wild and domesticated species. We used genotypic selection analysis to characterize selection on drought-induced plasticity of over 7,500 leaf transcripts of 118 rice accessions (genotypes) from different environmental conditions grown in a field experiment. Gene expression plasticity was neutral for most gradually plastic transcripts, but transcripts with discrete patterns of expression showed stronger selection on expression plasticity. Whether plasticity was adaptive and co-gradient or maladaptive and counter-gradient varied among varietal groups. No transcripts that experienced selection for plasticity across environments showed selection against plasticity within environments, indicating a lack of evidence for costs of adaptive plasticity that may constrain its evolution. Selection on expression plasticity was influenced by degree of plasticity, transcript length and gene body methylation. We observed positive selection on plasticity of co-expression modules containing transcripts involved in photosynthesis, translation and responsiveness to abiotic stress. Taken together, these results indicate that patterns of selection on expression plasticity were context-dependent and likely associated with environmental conditions of varietal groups, but that the evolution of adaptive plasticity would likely not be constrained by opposing patterns of selection on plasticity within compared to across environments. These results offer a genome-wide view of patterns of selection and ecological constraints on gene expression plasticity and provide insights into the interplay between plastic and evolutionary responses to drought at the molecular level.

The American black bear, Ursus americanus, is a widespread and ecologically important species in North America. In California, the black bear plays an important role in a variety of ecosystems and serves as an important species for recreational hunting. While research suggests that the populations in California are currently healthy, continued monitoring is critical, with genomic analyses providing an important surveillance tool. Here we report a high-quality, near chromosome-level genome assembly from a U. americanus sample from California. The primary assembly has a total length of 2.5 Gb contained in 316 scaffolds, a contig N50 of 58.9 Mb, a scaffold N50 of 67.6 Mb, and a BUSCO completeness score of 96%. This U. americanus genome assembly will provide an important resource for the targeted management of black bear populations in California, with the goal of achieving an appropriate balance between the recreational value of black bears and the maintenance of viable populations. The high quality of this genome assembly will also make it a valuable resource for comparative genomic analyses among black bear populations and among bear species.

The California Pipevine, Aristolochia californica Torr., is the only endemic California species within the cosmopolitan birthwort family Aristolochiaceae. It occurs as an understory vine in riparian and chaparral areas and in forest edges and windrows. The geographic range of this plant species almost entirely overlaps with that of its major specialized herbivore, the California Pipevine Swallowtail Butterfly Battus philenor hirsuta. While this species pair is a useful, ecologically well-understood system to study co-evolution, until recently, genomic resources for both have been lacking. Here, we report a new, chromosome-level assembly of A. californica as part of the California Conservation Genomics Project (CCGP). Following the sequencing and assembly strategy of the CCGP, we used Pacific Biosciences HiFi long reads and Hi-C chromatin proximity sequencing technology to produce a de novo assembled genome. Our genome assembly, the first for any species in the genus, contains 531 scaffolds spanning 661 megabase (Mb) pairs, with a contig N50 of 6.53 Mb, a scaffold N50 of 42.2 Mb, and BUSCO complete score of 98%. In combination with the recently published B. philenor hirsuta reference genome assembly, the A. californica reference genome assembly will be a powerful tool for studying co-evolution in a rapidly changing California landscape.

Dissecting plant responses to the environment is key to understanding whether and how plants adapt to anthropogenic climate change. Stomata, plants' pores for gas exchange, are expected to decrease in density following increased CO2 concentrations, a trend already observed in multiple plant species. However, it is unclear whether such responses are based on genetic changes and evolutionary adaptation. Here we make use of extensive knowledge of 43 genes in the stomatal development pathway and newly generated genome information of 191 Arabidopsis thaliana historical herbarium specimens collected over 193 years to directly link genetic variation with climate change. While we find that the essential transcription factors SPCH, MUTE and FAMA, central to stomatal development, are under strong evolutionary constraints, several regulators of stomatal development show signs of local adaptation in contemporary samples from different geographic regions. We then develop a functional score based on known effects of gene knock-out on stomatal development that recovers a classic pattern of stomatal density decrease over the past centuries, suggesting a genetic component contributing to this change. This approach combining historical genomics with functional experimental knowledge could allow further investigations of how different, even in historical samples unmeasurable, cellular plant phenotypes may have already responded to climate change through adaptive evolution.

San Fernando Valley spineflower (Chorizanthe parryi var. fernandina [S. Watson] Jeps.) (Polygonaceae) is an herbaceous annual plant, endemic to California, and until rediscovered in 1999 had been thought to be extinct for almost seven decades. Historically documented at 10 locations, it currently persists at 2, separated by approximately 27 km. State listed as endangered, a description of its genetic diversity and structure is of conservation interest. After determining a lack of variation in ploidy, we examined genetic variation from samples within both populations: a common garden study for potentially adaptive genetic variation in selected growth and phenological traits and analysis of single nucleotide polymorphisms identified through restriction-site associated DNA sequencing. Both measures indicated that this highly restricted taxon nevertheless harbors substantial levels of genetic diversity and has significant between- and within-population genetic structure. Combining approaches from population genomics and common garden studies provided more insight into the patterns and basis of genetic diversity than is typical for studies of non-model species. Although local adaptation was not specifically studied (i.e., via reciprocal transplant studies), the differences determined from these two independent lines of evidence indicate that mixing gene pools between populations is not recommended at this time. Further, with significant differences revealed among subpopulations, we caution against mixing genotypes across subpopulations for the most part, and without much more evidence that this would not pose a risk of outbreeding depression. The importance of supporting pollinator health and diversity is highlighted. With genetic diversity—particularly with an annual species—being dynamic, fluctuating with the usual processes and with contributions from the soil seedbank, we recommend periodic resampling to monitor genetic diversity and structure. Climate change is anticipated to contribute to this variability.

Pigmentation is an excellent trait to examine patterns of evolutionary change because it is often under natural selection. Benthic and limnetic threespine stickleback (Gasterosteus aculeatus) exhibit distinct pigmentation phenotypes, likely an adaptation to occupation of divergent niches. The genetic architecture of pigmentation in vertebrates appears to be complex. Prior QTL mapping of threespine stickleback pigmentation phenotypes has identified several candidate loci. However-relative to other morphological phenotypes (e.g., spines or lateral plates)-the genetic architecture of threespine stickleback pigmentation remains understudied. Here, we performed QTL mapping for two melanic pigmentation traits (melanophore density and lateral barring) using benthic-limnetic F2 crosses. The two traits mapped to different chromosomes, suggesting a distinct genetic basis. The resulting QTLs were additive, but explained a relatively small fraction of the total variance (~6%). QTLs maps differed by F1 family, suggesting variation in genetic architecture or ability to detect loci of small effect. Functional analysis identified enriched pathways for candidate loci. Several of the resulting candidate loci for pigmentation, including three loci in enriched pathways (bco1, sulf1, and tyms) have been previously indicated to affect pigmentation in other vertebrates. These findings add to a growing body of evidence suggesting pigmentation is often polygenic.

The jaguar (Panthera onca) is the largest living cat species native to the Americas and one of few large American carnivorans to have survived into the Holocene. However, the extent to which jaguar diversity declined during the end-Pleistocene extinction event remains unclear. For example, Pleistocene jaguar fossils from North America are notably larger than the average extant jaguar, leading to hypotheses that jaguars from this continent represent a now-extinct subspecies (Panthera onca augusta) or species (Panthera augusta). Here, we used a hybridization capture approach to recover an ancient mitochondrial genome from a large, late Pleistocene jaguar from Kingston Saltpeter Cave, Georgia, United States, which we sequenced to 26-fold coverage. We then estimated the evolutionary relationship between the ancient jaguar mitogenome and those from other extinct and living large felids, including multiple jaguars sampled across the species' current range. The ancient mitogenome falls within the diversity of living jaguars. All sampled jaguar mitogenomes share a common mitochondrial ancestor ~400 thousand years ago, indicating that the lineage represented by the ancient specimen dispersed into North America from the south at least once during the late Pleistocene. While genomic data from additional and older specimens will continue to improve understanding of Pleistocene jaguar diversity in the Americas, our results suggest that this specimen falls within the variation of extant jaguars despite the relatively larger size and geographic location and does not represent a distinct taxon.

Segregation distorters (SDs) are genetic elements that distort the Mendelian segregation ratio to favor their own transmission and are able to spread even when they incur fitness costs on organisms carrying them. Depending on the biology of the host organisms and the genetic architecture of the SDs, the population dynamics of SDs can be highly variable. Inbreeding is considered an effective mechanism for inhibiting the spread of SDs in populations, and can evolve as a defense mechanism against SDs in some systems. However, we show that inbreeding in the form of selfing in fact promotes the spread of SDs acting as pollen killers in a toxin-antidote system in hermaphroditic plants by two mechanisms: (i) By reducing the effective recombination rate between killer and antidote loci in the two-locus system and (ii) by increasing the proportion of SD alleles in individual flowers, rather than in the general gene-pool. We also show that in rice (Oryza sativa L.), a typical hermaphroditic plant, all molecularly characterized SDs associated with pollen killing were involved in population hybridization and have introgressed across different species. Paradoxically, these loci, which are associated with hybrid incompatibility and can be thought of as Bateson-Dobzhansky-Muller incompatibility loci are expected to reduce gene-flow between species, in fact cross species boundaries more frequently than random loci, and may act as important drivers of introgression.

Aposematic organisms rely on their conspicuous appearance to signal that they are defended and unpalatable. Such phenotypes are strongly tied to survival and reproduction. Aposematic colors and patterns are highly variable; however, the genetic, biochemical, and physiological mechanisms producing this conspicuous coloration remain largely unidentified. Here, we identify genes potentially affecting color variation in two color morphs of Ranitomeya imitator: the orange-banded Sauce and the redheaded Varadero morphs. We examine gene expression in black and orange skin patches from the Sauce morph and black and red skin patches from the Varadero morph. We identified genes differentially expressed between skin patches, including those that are involved in melanin synthesis (e.g. mlana, pmel, tyrp1), iridophore development (e.g. paics, ppat, ak1), pteridine synthesis (e.g. gch1, pax3-a, xdh), and carotenoid metabolism (e.g. dgat2, rbp1, scarb2). In addition, using weighted correlation network analysis, we identified the top 50 genes with high connectivity from the most significant network associated with gene expression differences between color morphs. Of these 50 genes, 13 were known to be related to color production (gch1, gmps, gpr143, impdh1, mc1r, pax3-a, pax7, ppat, rab27a, rlbp1, tfec, trpm1, xdh).

Increasing evidence suggests that urbanization is associated with higher mutation rates, which can affect the health and evolution of organisms that inhabit cities. Elevated pollution levels in urban areas can induce DNA damage, leading to de novo mutations. Studies on mutations induced by urban pollution are most prevalent in humans and microorganisms, whereas studies of non-human eukaryotes are rare, even though increased mutation rates have the potential to affect organisms and their populations in contemporary time. Our Perspective explores how higher mutation rates in urban environments could impact the fitness, ecology and evolution of populations. Most mutations will be neutral or deleterious, and higher mutation rates associated with elevated pollution in urban populations can increase the risk of cancer in humans and potentially other species. We highlight the potential for urban-driven increased deleterious mutational loads in some organisms, which could lead to a decline in population growth of a wide diversity of organisms. Although beneficial mutations are expected to be rare, we argue that higher mutation rates in urban areas could influence adaptive evolution, especially in organisms with short generation times. Finally, we explore avenues for future research to better understand the effects of urban-induced mutations on the fitness, ecology and evolution of city-dwelling organisms.

The Yellow Warbler (Setophaga petechia) is a small songbird in the wood-warbler family (Parulidae) that exhibits phenotypic and ecological differences across a widespread distribution and is important to California's riparian habitat conservation. Here, we present a high-quality de novo genome assembly of a vouchered female Yellow Warbler from southern California. Using HiFi long-read and Omni-C proximity sequencing technologies, we generated a 1.22 Gb assembly including 687 scaffolds with a contig N50 of 6.80 Mb, scaffold N50 of 21.18 Mb, and a BUSCO completeness score of 96.0%. This highly contiguous genome assembly provides an essential resource for understanding the history of gene flow, divergence, and local adaptation in Yellow Warblers and can inform conservation management of this charismatic bird species.

The coppery titi monkey (Plecturocebus cupreus) is an emerging nonhuman primate model system for behavioral and neurobiological research. At the same time, the almost entire absence of genomic resources for the species has hampered insights into the genetic underpinnings of the phenotypic traits of interest. To facilitate future genotype-to-phenotype studies, we here present a high-quality, fully annotated de novo genome assembly for the species with chromosome-length scaffolds spanning the autosomes and chromosome X (scaffold N50 = 130.8 Mb), constructed using data obtained from several orthologous short- and long-read sequencing and scaffolding techniques. With a base-level accuracy of ∼99.99% in chromosome-length scaffolds as well as benchmarking universal single-copy ortholog and k-mer completeness scores of >99.0% and 95.1% at the genome level, this assembly represents one of the most complete Pitheciidae genomes to date, making it an invaluable resource for comparative evolutionary genomics research to improve our understanding of lineage-specific changes underlying adaptive traits as well as deleterious mutations associated with disease.

How do chemically defended animals resist their own toxins? This intriguing question on the concept of autotoxicity is at the heart of how species interactions evolve. In this issue of Molecular Ecology (Molecular Ecology, 2024, 33), Bodawatta and colleagues report on how Papua New Guinean birds coopted deadly neurotoxins to create lethal mantles that protect against predators and parasites. Combining chemical screening of the plumage of a diverse collection of passerine birds with genome sequencing, the researchers unlocked a deeper understanding of how some birds sequester deadly batrachotoxin (BTX) from their food without poisoning themselves. They identified that birds impervious to BTX bear amino acid substitutions in the toxin-binding site of the voltage-gated sodium channel Nav1.4, whose function is essential for proper contraction and relaxation of vertebrate muscles. Comparative genetic and molecular docking analyses show that several of the substitutions associated with insensitivity to BTX may have become prevalent among toxic birds through positive selection. Intriguingly, poison dart frogs that also co-opted BTX in their lethal mantles were found to harbour similar toxin insensitivity substitutions in their Nav1.4 channels. Taken together, this sets up a powerful model system for studying the mechanisms behind convergent molecular evolution and how it may drive biological diversity.

The genus Xanthomonas has been primarily studied for pathogenic interactions with plants. However, besides host and tissue-specific pathogenic strains, this genus also comprises nonpathogenic strains isolated from a broad range of hosts, sometimes in association with pathogenic strains, and other environments, including rainwater. Based on their incapacity or limited capacity to cause symptoms on the host of isolation, nonpathogenic xanthomonads can be further characterized as commensal and weakly pathogenic. This study aimed to understand the diversity and evolution of nonpathogenic xanthomonads compared to their pathogenic counterparts based on their cooccurrence and phylogenetic relationship and to identify genomic traits that form the basis of a life history framework that groups xanthomonads by ecological strategies. We sequenced genomes of 83 strains spanning the genus phylogeny and identified eight novel species, indicating unexplored diversity. While some nonpathogenic species have experienced a recent loss of a type III secretion system, specifically the hrp2 cluster, we observed an apparent lack of association of the hrp2 cluster with lifestyles of diverse species. We performed association analysis on a large data set of 337 Xanthomonas strains to explain how xanthomonads may have established association with the plants across the continuum of lifestyles from commensals to weak pathogens to pathogens. Presence of distinct transcriptional regulators, distinct nutrient utilization and assimilation genes, transcriptional regulators, and chemotaxis genes may explain lifestyle-specific adaptations of xanthomonads.

Transposable elements (TE) play critical roles in shaping genome evolution. Highly repetitive TE sequences are also a major source of assembly gaps making it difficult to fully understand the impact of these elements on host genomes. The increased capacity of long-read sequencing technologies to span highly repetitive regions promises to provide new insights into patterns of TE activity across diverse taxa. Here we report the generation of highly contiguous reference genomes using PacBio long-read and Omni-C technologies for three species of Passerellidae sparrow. We compared these assemblies to three chromosome-level sparrow assemblies and nine other sparrow assemblies generated using a variety of short- and long-read technologies. All long-read based assemblies were longer (range: 1.12 to 1.41 Gb) than short-read assemblies (0.91 to 1.08 Gb) and assembly length was strongly correlated with the amount of repeat content. Repeat content for Bell's sparrow (31.2% of genome) was the highest level ever reported within the order Passeriformes, which comprises over half of avian diversity. The highest levels of repeat content (79.2% to 93.7%) were found on the W chromosome relative to other regions of the genome. Finally, we show that proliferation of different TE classes varied even among species with similar levels of repeat content. These patterns support a dynamic model of TE expansion and contraction even in a clade where TEs were once thought to be fairly depauperate and static. Our work highlights how the resolution of difficult-to-assemble regions of the genome with new sequencing technologies promises to transform our understanding of avian genome evolution.

Genomic evidence supports an important role for selection in shaping patterns of introgression along the genome, but frameworks for understanding the evolutionary dynamics within hybrid populations that underlie these patterns have been lacking. Due to the clock-like effect of recombination in hybrids breaking up parental haplotypes, drift and selection produce predictable patterns of ancestry variation at varying spatial genomic scales through time. Here, we develop methods based on the Discrete Wavelet Transform to study the genomic scale of local ancestry variation and its association with recombination rates and show that these methods capture temporal dynamics of drift and genome-wide selection after hybridization. We apply these methods to published datasets from hybrid populations of swordtail fish (Xiphophorus) and baboons (Papio) and to inferred Neanderthal introgression in modern humans. Across systems, upward of 20% of variation in local ancestry at the broadest genomic scales can be attributed to systematic selection against introgressed alleles, consistent with strong selection acting on early-generation hybrids. Signatures of selection at fine genomic scales suggest selection over longer time scales; however, we suggest that our ability to confidently infer selection at fine scales is likely limited by inherent biases in current methods for estimating local ancestry from contiguous segments of genomic similarity. Wavelet approaches will become widely applicable as genomic data from systems with introgression become increasingly available and can help shed light on generalities of the genomic consequences of interspecific hybridization.

The entomopathogenic fungus Conoideocrella luteorostrata has recently been implicated in natural epizootics among exotic elongate hemlock scale (EHS) insects in Christmas tree farms in the eastern United States. Since 1913, C. luteorostrata has been reported from various plant feeding Hemiptera in the southeastern United States, but comprehensive morphological and phylogenetic studies of U.S. populations are lacking. The recovery of multiple strains of C. luteorostrata from mycosed EHS in North Carolina provided an opportunity to conduct pathogenicity assays and morphological and phylogenetic studies to investigate genus- and species-level boundaries among the Clavicipitaceae. Pathogenicity assays confirmed that C. luteorostrata causes mortality of EHS crawlers, an essential first step in developing this fungus as a biocontrol. Morphological studies revealed that conidia aligned with previous measurements of the Paecilomyces-like asexual state of C. luteorostrata, with conidiophore morphology consistent with historical observations. Additionally, a Hirsutella-like synanamorph was observed in select C. luteorostrata strains. In both a four-locus, 54-taxon Clavicipitaceae-wide phylogenetic analysis including D1-D2 domains of the nuclear 28S rRNA region (28S), elongation factor 1 alpha (EF1-α), DNA-directed RNA polymerase II subunit 1 (RPB1), and DNA-directed RNA polymerase II subunit 2 (RPB2) and a two-locus, 38-taxon (28S and EF1-α) phylogenetic analysis, all three Conoideocrella species were resolved as strongly supported monophyletic lineages across all loci and both methods (maximum likelihood and Bayesian inference) of phylogenetic inference except for 28S for C. tenuis. Despite the strong support for individual Conoideocrella species, none of the analyses supported the monophyly of Conoideocrella with the inclusion of Dussiella. Due to the paucity of RPB1 and RPB2 sequence data, EF1-α provided superior delimitation of intraspecies groupings for Conoideocrella and should be used in future studies. Further development of C. luteorostrata as a biocontrol against EHS will require additional surveys across diverse Hemiptera and expanded pathogenicity testing to clarify host range and efficacy of this fungus.

Populations composed of individuals descended from multiple distinct genetic lineages often feature significant differences in phenotypic frequencies. We considered hatchery production of steelhead, the migratory anadromous form of the salmonid species Oncorhynchus mykiss, and investigated how differences among genetic lineages and environmental variation impacted life history traits. We genotyped 23,670 steelhead returning to the four California Central Valley hatcheries over 9 years from 2011 to 2019, confidently assigning parentage to 13,576 individuals to determine age and date of spawning and rates of iteroparity and repeat spawning within each year. We found steelhead from different genetic lineages showed significant differences in adult life history traits despite inhabiting similar environments. Differences between coastal and Central Valley steelhead lineages contributed to significant differences in age at return, timing of spawning, and rates of iteroparity among programs. In addition, adaptive genomic variation associated with life history development in this species varied among hatchery programs and was associated with the age of steelhead spawners only in the coastal lineage population. Environmental variation likely contributed to variations in phenotypic patterns observed over time, as our study period spanned both a marine heatwave and a serious drought in California. Our results highlight evidence of a strong genetic component underlying known phenotypic differences in life history traits between two steelhead lineages.

Our understanding on the interplay between gene functionality and gene arrangement at different chromosome scales relies on a few Diptera and the honeybee, species with quality-reference genome assemblies, accurate gene annotations, and abundant transcriptome data. Using recently generated 'omics resources in the monarch butterfly D. plexippus, a species with many more and smaller chromosomes relative to Drosophila species and the honeybee, we examined the organization of genes preferentially expressed at broadly defined developmental stages (larva, pupa, adult males, and adult females) at both fine and whole-chromosome scales. We found that developmental stage-regulated genes do not form more clusters, but do form larger clusters, than expected by chance, a pattern consistent across the gene categories examined. Notably, out of the thirty chromosomes in the monarch genome, twelve of them, plus the fraction of the chromosome Z that corresponds to the ancestral Z in other Lepidoptera, were found enriched for developmental stage-regulated genes. These two levels of nonrandom gene organization are not independent as enriched chromosomes for developmental stage-regulated genes tend to harbor disproportionately large clusters of these genes. Further, although paralogous genes were overrepresented in gene clusters, their presence is not enough to explain two-thirds of the documented cases of whole-chromosome enrichment. The composition of the largest clusters often included paralogs from more than one multigene family as well as unrelated single-copy genes. Our results reveal intriguing patterns at the whole-chromosome scale in D. plexippus while shedding light on the interplay between gene expression and chromosome organization beyond Diptera and Hymenoptera.

Carpenter ants in the genus Camponotus are large, conspicuous ants that are abundant and ecologically influential in many terrestrial ecosystems. The bicolored carpenter ant, Camponotus vicinus Mayr, is distributed across a wide range of elevations and latitudes in western North America, where it is a prominent scavenger and predator. Here, we present a high-quality genome assembly of C. vicinus from a sample collected in Sonoma County, California, near the type locality of the species. This genome assembly consists of 38 scaffolds spanning 302.74 Mb, with contig N50 of 15.9 Mb, scaffold N50 of 19.9 Mb, and BUSCO completeness of 99.2%. This genome sequence will be a valuable resource for exploring the evolutionary ecology of C. vicinus and carpenter ants generally. It also provides an important tool for clarifying cryptic diversity within the C. vicinus species complex, a genetically diverse set of populations, some of which are quite localized and of conservation interest.

The little pocket mouse, Perognathus longimembris, and its nine congeners are small heteromyid rodents found in arid and seasonally arid regions of Western North America. The genus is characterized by behavioral and physiological adaptations to dry and often harsh environments, including nocturnality, seasonal torpor, food caching, enhanced osmoregulation, and a well-developed sense of hearing. Here we present a genome assembly of Perognathus longimembris longimembris generated from PacBio HiFi long read and Omni-C chromatin-proximity sequencing as part of the California Conservation Genomics Project. The assembly has a length of 2.35 Gb, contig N50 of 11.6 Mb, scaffold N50 of 73.2 Mb, and includes 93.8% of the BUSCO Glires genes. Interspersed repetitive elements constitute 41.2% of the genome. A comparison with the highly endangered Pacific pocket mouse, P. l. pacificus, reveals broad synteny. These new resources will enable studies of local adaptation, genetic diversity, and conservation of threatened taxa.

Smoky rubyspot damselflies (Hetaerina titia Drury, 1773) are one of the most commonly encountered odonates along streams and rivers on both slopes of Central America and the Atlantic drainages in the United States and southern Canada. Owing to their highly variable wing pigmentation, they have become a model system for studying sexual selection and interspecific behavioral interference. Here, we sequence and assemble the genome of a female smoky rubyspot. Of the primary assembly (i.e. the principle pseudohaplotype), 98.8% is made up of 12 chromosomal pseudomolecules (2N = 22A + X). There are 75 scaffolds in total, an N50 of 120 Mb, a contig-N50 of 0.64 Mb, and a high arthropod BUSCO score [C: 97.6% (S: 97.3%, D: 0.3%), F: 0.8%, M: 1.6%]. We then compare our assembly to that of the blue-tailed damselfly genome (Ischnura elegans), the most complete damselfly assembly to date, and a recently published assembly for an American rubyspot damselfly (Hetaerina americana). Collectively, these resources make Hetaerina a genome-enabled genus for further studies of the ecological and evolutionary forces shaping biological diversity.

The Yuma myotis bat (Myotis yumanensis) is a small vespertilionid bat and one of 52 species of new world Myotis bats in the subgenus Pizonyx. While M. yumanensis populations currently appear relatively stable, it is one of 12 bat species known or suspected to be susceptible to white-nose syndrome, the fungal disease causing declines in bat populations across North America. Only two of these 12 species have genome resources available, which limits the ability of resource managers to use genomic techniques to track the responses of bat populations to white-nose syndrome generally. Here we present the first de novo genome assembly for Yuma myotis, generated as a part of the California Conservation Genomics Project. The M. yumanensis genome was generated using a combination of PacBio HiFi long reads and Omni-C chromatin-proximity sequencing technology. This high-quality genome is one of the most complete bat assemblies available, with a contig N50 of 28.03 Mb, scaffold N50 of 99.14 Mb, and BUSCO completeness score of 93.7%. The Yuma myotis genome provides a high-quality resource that will aid in comparative genomic and evolutionary studies, as well as inform conservation management related to white-nose syndrome.

Across internally fertilising species, males transfer ejaculate proteins that trigger wide-ranging changes in female behaviour and physiology. Much theory has been developed to explore the drivers of ejaculate protein evolution. The accelerating availability of high-quality genomes now allows us to test how these proteins are evolving at fine taxonomic scales. Here, we use genomes from 264 species to chart the evolutionary history of Sex Peptide (SP), a potent regulator of female post-mating responses in Drosophila melanogaster. We infer that SP first evolved in the Drosophilinae subfamily and has since followed markedly different evolutionary trajectories in different lineages. Outside of the Sophophora-Lordiphosa, SP exists largely as a single-copy gene with independent losses in several lineages. Within the Sophophora-Lordiphosa, the SP gene family has repeatedly and independently expanded. Up to seven copies, collectively displaying extensive sequence variation, are present in some species. Despite these changes, SP expression remains restricted to the male reproductive tract. Alongside, we document considerable interspecific variation in the presence and morphology of seminal microcarriers that, despite the critical role SP plays in microcarrier assembly in D. melanogaster, appears to be independent of changes in the presence/absence or sequence of SP. We end by providing evidence that SP's evolution is decoupled from that of its receptor, Sex Peptide Receptor, in which we detect no evidence of correlated diversifying selection. Collectively, our work describes the divergent evolutionary trajectories that a novel gene has taken following its origin and finds a surprisingly weak coevolutionary signal between a supposedly sexually antagonistic protein and its receptor.

Desertification is a major threat to biodiversity in arid areas of the world, partly because many organisms in these regions already exist at or near the limits of their movement and physiology. Here, we used molecular data to investigate patterns of persistence and dispersal in an ecologically and economically important carpenter bee (Xylocopa grisescens Lepeletier) found throughout the semiarid Caatinga region of Brazil. We used a genome-wide approach (double digest restriction-site associated DNA, ddRAD) to gather genetic data from bees sampled from eight sites within a semiarid region subject to desertification in Northeastern Brazil. Across all populations, we observed a consistent heterozygosity and effective population size deficit along with low genetic differentiation. We did not find strong evidence of dispersal limitations caused by desertification in this study system despite data collection from sites up to 300 km distant. Thus, our data suggest that human-mediated changes in the Caatinga, such as habitat loss, have impacted the population genetic patterns of X. grisescens. However, these impacts have also been softened by the species' biological characteristics, such as its relatively high capacity for movement. This study provides insights into how habitat changes might impact the long-term survival of large solitary bees.

Vocal rhythm plays a fundamental role in sexual selection and species recognition in birds, but little is known of its genetic basis due to the confounding effect of vocal learning in model systems. Uncovering its genetic basis could facilitate identifying genes potentially important in speciation. Here we investigate the genomic underpinnings of rhythm in vocal non-learning Pogoniulus tinkerbirds using 135 individual whole genomes distributed across a southern African hybrid zone. We find rhythm speed is associated with two genes that are also known to affect human speech, Neurexin-1 and Coenzyme Q8A. Models leveraging ancestry reveal these candidate loci also impact rhythmic stability, a trait linked with motor performance which is an indicator of quality. Character displacement in rhythmic stability suggests possible reinforcement against hybridization, supported by evidence of asymmetric assortative mating in the species producing faster, more stable rhythms. Because rhythm is omnipresent in animal communication, candidate genes identified here may shape vocal rhythm across birds and other vertebrates.

BackgroundThe Atribacterota are widely distributed in the subsurface biosphere. Recently, the first Atribacterota isolate was described and the number of Atribacterota genome sequences retrieved from environmental samples has increased significantly; however, their diversity, physiology, ecology, and evolution remain poorly understood.ResultsWe report the isolation of the second member of Atribacterota, Thermatribacter velox gen. nov., sp. nov., within a new family Thermatribacteraceae fam. nov., and the short-term laboratory cultivation of a member of the JS1 lineage, Phoenicimicrobium oleiphilum HX-OS.bin.34TS, both from a terrestrial oil reservoir. Physiological and metatranscriptomics analyses showed that Thermatribacter velox B11T and Phoenicimicrobium oleiphilum HX-OS.bin.34TS ferment sugars and n-alkanes, respectively, producing H2, CO2, and acetate as common products. Comparative genomics showed that all members of the Atribacterota lack a complete Wood-Ljungdahl Pathway (WLP), but that the Reductive Glycine Pathway (RGP) is widespread, indicating that the RGP, rather than WLP, is a central hub in Atribacterota metabolism. Ancestral character state reconstructions and phylogenetic analyses showed that key genes encoding the RGP (fdhA, fhs, folD, glyA, gcvT, gcvPAB, pdhD) and other central functions were gained independently in the two classes, Atribacteria (OP9) and Phoenicimicrobiia (JS1), after which they were inherited vertically; these genes included fumarate-adding enzymes (faeA; Phoenicimicrobiia only), the CODH/ACS complex (acsABCDE), and diverse hydrogenases (NiFe group 3b, 4b and FeFe group A3, C). Finally, we present genome-resolved community metabolic models showing the central roles of Atribacteria (OP9) and Phoenicimicrobiia (JS1) in acetate- and hydrocarbon-rich environments.ConclusionOur findings expand the knowledge of the diversity, physiology, ecology, and evolution of the phylum Atribacterota. This study is a starting point for promoting more incisive studies of their syntrophic biology and may guide the rational design of strategies to cultivate them in the laboratory. Video Abstract.

Long-read sequencing is driving rapid progress in genome assembly across all major groups of life, including species of the family Drosophilidae, a longtime model system for genetics, genomics, and evolution. We previously developed a cost-effective hybrid Oxford Nanopore (ONT) long-read and Illumina short-read sequencing approach and used it to assemble 101 drosophilid genomes from laboratory cultures, greatly increasing the number of genome assemblies for this taxonomic group. The next major challenge is to address the laboratory culture bias in taxon sampling by sequencing genomes of species that cannot easily be reared in the lab. Here, we build upon our previous methods to perform amplification-free ONT sequencing of single wild flies obtained either directly from the field or from ethanol-preserved specimens in museum collections, greatly improving the representation of lesser studied drosophilid taxa in whole-genome data. Using Illumina Novaseq X Plus and ONT P2 sequencers with R10.4.1 chemistry, we set a new benchmark for inexpensive hybrid genome assembly at US $150 per genome while assembling genomes from as little as 35 ng of genomic DNA from a single fly. We present 183 new genome assemblies for 179 species as a resource for drosophilid systematics, phylogenetics, and comparative genomics. Of these genomes, 62 are from pooled lab strains and 121 from single adult flies. Despite the sample limitations of working with small insects, most single-fly diploid assemblies are comparable in contiguity (>1 Mb contig N50), completeness (>98% complete dipteran BUSCOs), and accuracy (>QV40 genome-wide with ONT R10.4.1) to assemblies from inbred lines. We present a well-resolved multi-locus phylogeny for 360 drosophilid and 4 outgroup species encompassing all publicly available (as of August 2023) genomes for this group. Finally, we present a Progressive Cactus whole-genome, reference-free alignment built from a subset of 298 suitably high-quality drosophilid genomes. The new assemblies and alignment, along with updated laboratory protocols and computational pipelines, are released as an open resource and as a tool for studying evolution at the scale of an entire insect family.

The modification of fertile stamens into sterile staminodes has occurred independently many times in the flowering plant lineage. In the genus Aquilegia (columbine) and its closest relatives, the two stamen whorls closest to the carpels have been converted to staminodes. In Aquilegia, the only genetic analyses of staminode development have been reverse genetic approaches revealing that B-class floral identity genes are involved. A. jonesii, the only species of columbine where staminodes have reverted to fertile stamens, allows us to explore the genetic architecture of staminode development using a forward genetic approach. We performed QTL analysis using an outcrossed F2 population between A. jonesii and a horticultural variety that makes fully developed staminodes, A. coerulea 'Origami'. Our results reveal a polygenic basis for staminode loss where the two staminode whorls are under some level of independent control. We also discovered that staminode loss in A. jonesii is not complete, in which staminode-like traits sometimes occur in the inner fertile stamens, potentially representing a fading boundary of gene expression. The QTLs identified in this study provide a map to guide future reverse genetic and functional studies examining the genetic basis and evolutionary significance of this trait.

Abstract: Aim: Understanding the impacts of past and future climate change on genetic diversity and structure is a current major research gap. We ask whether past range shifts explain the observed genetic diversity of surfgrass species and if future climate change projections anticipate genetic diversity losses. Our study aims to identify regions of long‐term climate suitability with higher and unique seagrass genetic diversity and predict future impacts of climate change on them. Location: Northeast Pacific. Time Period: Analyses considered a timeframe from the Last Glacial Maximum (LGM; 20 kybp) until one Representative Concentration Pathway (RCP) scenario of future climate changes (RCP 8.5; 2100). Major Taxa Studied: Two seagrass species belonging to the genus Phyllospadix. Methods: We estimated population genetic diversity and structure using 11 polymorphic microsatellite markers. We predicted the distribution of the species for the present, LGM, and near future (RCP 8.5, no climate mitigation) using Species Distribution Models (SDMs). Results: SDMs revealed southward range shifts during the LGM and potential poleward expansions in the future. Genetic diversity of Phyllospadix torreyi decreases from north to south, but in Phyllospadix scouleri the trend is variable. Phyllospadix scouleri displays signals of genome admixture at the southernmost and northernmost edges of its distribution. Main Conclusions: The genetic patterns observed in the present reveal the influence of climate‐driven range shifts in the past and suggest further consequences of climate change in the future, with potential loss of unique gene pools. This study also shows that investigating climate links to present genetic information at multiple timescales can establish a historical context for analyses of the future evolutionary history of populations.

Amphiboreal taxa are often composed of vicariant phylogroups and species complexes whose divergence and phylogeographic affinities reflect a shared history of chronic isolation and episodic trans-Arctic dispersal. Ecological filters and shifting selective pressures may also promote selective sweeps, niche shifts and ecological speciation during colonization, but these are seldom considered at biogeographical scales. Here we integrate genetic data and Ecologic Niche Models (ENMs) to investigate the historical biogeography and cohesion of the polymorphic rockweed Fucus distichus throughout its immense amphiboreal range, focusing on trans-Arctic asymmetries, glacial/interglacial dynamics, and integrity of sympatric eco-morphotypes. Populations were sampled throughout the Pacific and the Atlantic, from southern rear-edges to the high-Arctic. They were genotyped for seven microsatellites and an mtDNA spacer, and genetic diversity and structure were assessed from global to local scales. ENMs were used to compare niche divergence and magnitude of post-glacial range shifts in Pacific versus Atlantic sub-ranges. Haplotypic and genotypic data revealed distinct and seemingly isolated Pacific vs Arctic/Atlantic gene-pools, with finer-scale regional sub-structuring pervasive in the Pacific. MtDNA diversity was highly structured and overwhelmingly concentrated in the Pacific. Regionally, Alaska showed the highest intra-population diversity but the lowest levels of endemism. Some sympatric/parapatric ecotypes exhibited distinct genotypic/haplotypic compositions. Strikingly, niche models revealed higher Pacific tolerance to maximum temperatures and predicted a much more consolidated presence in the NE Atlantic. Glacial and modern ranges overlapped extensively in the Pacific, whereas the modern Atlantic range was largely glaciated or emerged during the Last Glacial Maximum. Higher genetic and ecogeographic diversity supports a primary Pacific diversification and secondary Atlantic colonization, also likely reflecting the much larger and more stable climatic refugia in the Pacific. The relic distribution and reduced ecological/morphological plasticity in the NE Atlantic are hypothesized to reflect functional trans-Arctic bottlenecks, recent colonization or competition with congeners. Within the Pacific, Alaska showed signatures of a post-glacial melting pot of eastern and southern populations. Genetic/ecotypic variation was generally not sufficiently discontinuous or consistent to justify recognizing multiple taxonomic entities, but support a separate species in the eastern Pacific, at the southern rear-edge. We predict that layered patterns of phylogeographic structure, incipient speciation and niche differences might be common among widespread low-dispersal amphiboreal taxa.

Despite rapid evolution across eutherian mammals, the X-linked MIR-506 family miRNAs are located in a region flanked by two highly conserved protein-coding genes (SLITRK2 and FMR1) on the X chromosome. Intriguingly, these miRNAs are predominantly expressed in the testis, suggesting a potential role in spermatogenesis and male fertility. Here, we report that the X-linked MIR-506 family miRNAs were derived from the MER91C DNA transposons. Selective inactivation of individual miRNAs or clusters caused no discernible defects, but simultaneous ablation of five clusters containing 19 members of the MIR-506 family led to reduced male fertility in mice. Despite normal sperm counts, motility, and morphology, the KO sperm were less competitive than wild-type sperm when subjected to a polyandrous mating scheme. Transcriptomic and bioinformatic analyses revealed that these X-linked MIR-506 family miRNAs, in addition to targeting a set of conserved genes, have more targets that are critical for spermatogenesis and embryonic development during evolution. Our data suggest that the MIR-506 family miRNAs function to enhance sperm competitiveness and reproductive fitness of the male by finetuning gene expression during spermatogenesis.

BackgroundBiofilms in sulfide-rich springs present intricate microbial communities that play pivotal roles in biogeochemical cycling. We studied chemoautotrophically based biofilms that host diverse CPR bacteria and grow in sulfide-rich springs to investigate microbial controls on biogeochemical cycling.ResultsSulfide springs biofilms were investigated using bulk geochemical analysis, genome-resolved metagenomics, and scanning transmission X-ray microscopy (STXM) at room temperature and 87 K. Chemolithotrophic sulfur-oxidizing bacteria, including Thiothrix and Beggiatoa, dominate the biofilms, which also contain CPR Gracilibacteria, Absconditabacteria, Saccharibacteria, Peregrinibacteria, Berkelbacteria, Microgenomates, and Parcubacteria. STXM imaging revealed ultra-small cells near the surfaces of filamentous bacteria that may be CPR bacterial episymbionts. STXM and NEXAFS spectroscopy at carbon K and sulfur L2,3 edges show that filamentous bacteria contain protein-encapsulated spherical elemental sulfur granules, indicating that they are sulfur oxidizers, likely Thiothrix. Berkelbacteria and Moranbacteria in the same biofilm sample are predicted to have a novel electron bifurcating group 3b [NiFe]-hydrogenase, putatively a sulfhydrogenase, potentially linked to sulfur metabolism via redox cofactors. This complex could potentially contribute to symbioses, for example, with sulfur-oxidizing bacteria such as Thiothrix that is based on cryptic sulfur cycling. One Doudnabacteria genome encodes adjacent sulfur dioxygenase and rhodanese genes that may convert thiosulfate to sulfite. We find similar conserved genomic architecture associated with CPR bacteria from other sulfur-rich subsurface ecosystems.ConclusionsOur combined metagenomic, geochemical, spectromicroscopic, and structural bioinformatics analyses of biofilms growing in sulfide-rich springs revealed consortia that contain CPR bacteria and sulfur-oxidizing Proteobacteria, including Thiothrix, and bacteria from a new family within Beggiatoales. We infer roles for CPR bacteria in sulfur and hydrogen cycling. Video Abstract.

In this study, a Gram-stain-positive, non-motile, oxidase- and catalase-negative, rod-shaped, bacterial strain (SG_E_30_P1T) that formed light yellow colonies was isolated from a groundwater sample of Sztaravoda spring, Hungary. Based on 16S rRNA phylogenetic and phylogenomic analyses, the strain was found to form a distinct linage within the family Microbacteriaceae. Its closest relatives in terms of near full-length 16S rRNA gene sequences are Salinibacterium hongtaonis MH299814 (97.72 % sequence similarity) and Leifsonia psychrotolerans GQ406810 (97.57 %). The novel strain grows optimally at 20-28 °C, at neutral pH and in the presence of NaCl (1-2 w/v%). Strain SG_E_30_P1T contains MK-7 and B-type peptidoglycan with diaminobutyrate as the diagnostic amino acid. The major cellular fatty acids are anteiso-C15 : 0, iso-C16 : 0 and iso-C14 : 0, and the polar lipid profile is composed of diphosphatidylglycerol and phosphatidylglycerol, as well as an unidentified aminoglycolipid, aminophospholipid and some unidentified phospholipids. The assembled draft genome is a contig with a total length of 2 897 968 bp and a DNA G+C content of 65.5 mol%. Amino acid identity values with it closest relatives with sequenced genomes of

The order Sordariales is taxonomically diverse, and harbours many species with different lifestyles and large economic importance. Despite its importance, a robust genome-scale phylogeny, and associated comparative genomic analysis of the order is lacking. In this study, we examined whole-genome data from 99 Sordariales, including 52 newly sequenced genomes, and seven outgroup taxa. We inferred a comprehensive phylogeny that resolved several contentious relationships amongst families in the order, and cleared-up intrafamily relationships within the Podosporaceae. Extensive comparative genomics showed that genomes from the three largest families in the dataset (Chaetomiaceae, Podosporaceae and Sordariaceae) differ greatly in GC content, genome size, gene number, repeat percentage, evolutionary rate, and genome content affected by repeat-induced point mutations (RIP). All genomic traits showed phylogenetic signal, and ancestral state reconstruction revealed that the variation of the properties stems primarily from within-family evolution. Together, the results provide a thorough framework for understanding genome evolution in this important group of fungi.

Explorations in the past 20 years in the Plio-Pleistocene Zanda Basin (3,800–4,500 m above sea level) along the northern slopes of the Himalaya Mountains have substantially enriched our understanding of the paleoenvironments of the Tibetan Plateau and associated biologic evolution. Many elements of the mammalian fauna recovered are either new to science or shed new light about their special adaptations in this high elevation basin. Here we describe a new species of twisted-horned antelope, Gazellospira tsaparangensis, with a heteronymous spiral. Its small size and primitive morphology, such as relatively short horncore with less twisting, thin frontal bones, a lack of frontal and horncore sinuses, small size of supraorbital foramina, and lack of an anterior keel, helps to place it at the base of genus Gazellospira, substantially more stem-ward than the type species G. torticornis from the Plio-Pleistocene of Europe and western Asia. With an estimated age of 3.62 Ma, this also places G. tsaparangensis as one of the early occurrences in Eurasia, although some fragmentary records in Turkey may be slightly earlier. Considering this early appearance and primitive morphology, G. tsaparangensis once again may be a case of the ‘out-of-Tibet’ model of megafauna origin, with earlier progenitors adapted to cold environments in high Tibet before expanding their range to the rest of Eurasia.

Marine algae are central to global carbon fixation, and their productivity is dictated largely by resource availability. Reduced nutrient availability is predicted for vast oceanic regions as an outcome of climate change; however, there is much to learn regarding response mechanisms of the tiny picoplankton that thrive in these environments, especially eukaryotic phytoplankton. Here, we investigate responses of the picoeukaryote Micromonas commoda, a green alga found throughout subtropical and tropical oceans. Under shifting phosphate availability scenarios, transcriptomic analyses revealed altered expression of transfer RNA modification enzymes and biased codon usage of transcripts more abundant during phosphate-limiting versus phosphate-replete conditions, consistent with the role of transfer RNA modifications in regulating codon recognition. To associate the observed shift in the expression of the transfer RNA modification enzyme complement with the transfer RNAs encoded by M. commoda, we also determined the transfer RNA repertoire of this alga revealing potential targets of the modification enzymes. Codon usage bias was particularly pronounced in transcripts encoding proteins with direct roles in managing phosphate limitation and photosystem-associated proteins that have ill-characterized putative functions in "light stress." The observed codon usage bias corresponds to a proposed stress response mechanism in which the interplay between stress-induced changes in transfer RNA modifications and skewed codon usage in certain essential response genes drives preferential translation of the encoded proteins. Collectively, we expose a potential underlying mechanism for achieving growth under enhanced nutrient limitation that extends beyond the catalog of up- or downregulated protein-encoding genes to the cell biological controls that underpin acclimation to changing environmental conditions.

Life-history variation is the raw material of adaptation, and understanding its genetic and environmental underpinnings is key to designing effective conservation strategies. We used large-scale genetic pedigree reconstruction of anadromous steelhead trout (Oncorhynchus mykiss) from the Russian River, CA, USA, to elucidate sex-specific patterns of life-history traits and their heritability. SNP data from adults returning from sea over a 14-year period were used to identify 13,474 parent-offspring trios. These pedigrees were used to determine age structure, size distributions and family sizes for these fish, as well as to estimate the heritability of two key life-history traits, spawn date and age at maturity (first reproduction). Spawn date was highly heritable (h2  = 0.73) and had a cross-sex genetic correlation near unity. We provide the first estimate of heritability for age at maturity in ocean-going fish from this species and found it to be highly heritable (h2 from 0.29 to 0.62, depending on sex and method), with a much lower genetic correlation across sexes. We also evaluated genotypes at a migration-associated inversion polymorphism and found sex-specific correlations with age at maturity. The significant heritability of these two key reproductive traits in these imperiled fish, and their patterns of inheritance in the two sexes, is consistent with predictions of both natural and sexually antagonistic selection (sexes experience opposing selection pressures). This emphasizes the importance of anthropogenic factors, including hatchery practices and ecosystem modifications, in shaping the fitness of this species, thus providing important guidance for management and conservation efforts.

Animal pigment patterns are excellent models to elucidate mechanisms of biological organization. Although theoretical simulations, such as Turing reaction-diffusion systems, recapitulate many animal patterns, they are insufficient to account for those showing a high degree of spatial organization and reproducibility. Here, we study the coat of the African striped mouse (Rhabdomys pumilio) to uncover how periodic stripes form. Combining transcriptomics, mathematical modelling and mouse transgenics, we show that the Wnt modulator Sfrp2 regulates the distribution of hair follicles and establishes an embryonic prepattern that foreshadows pigment stripes. Moreover, by developing in vivo gene editing in striped mice, we find that Sfrp2 knockout is sufficient to alter the stripe pattern. Strikingly, mutants exhibited changes in pigmentation, revealing that Sfrp2 also regulates hair colour. Lastly, through evolutionary analyses, we find that striped mice have evolved lineage-specific changes in regulatory elements surrounding Sfrp2, many of which may be implicated in modulating the expression of this gene. Altogether, our results show that a single factor controls coat pattern formation by acting both as an orienting signalling mechanism and a modulator of pigmentation. More broadly, our work provides insights into how spatial patterns are established in developing embryos and the mechanisms by which phenotypic novelty originates.

Migration is driven by a combination of environmental and genetic factors, but many questions remain about those drivers. Potential interactions between genetic and environmental variants associated with different migratory phenotypes are rarely the focus of study. We pair low coverage whole genome resequencing with a de novo genome assembly to examine population structure, inbreeding, and the environmental factors associated with genetic differentiation between migratory and resident breeding phenotypes in a species of conservation concern, the western burrowing owl (Athene cunicularia hypugaea). Our analyses reveal a dichotomy in gene flow depending on whether the population is resident or migratory, with the former being genetically structured and the latter exhibiting no signs of structure. Among resident populations, we observed significantly higher genetic differentiation, significant isolation-by-distance, and significantly elevated inbreeding. Among migratory breeding groups, on the other hand, we observed lower genetic differentiation, no isolation-by-distance, and substantially lower inbreeding. Using genotype-environment association analysis, we find significant evidence for relationships between migratory phenotypes (i.e., migrant versus resident) and environmental variation associated with cold temperatures during the winter and barren, open habitats. In the regions of the genome most differentiated between migrants and residents, we find significant enrichment for genes associated with the metabolism of fats. This may be linked to the increased pressure on migrants to process and store fats more efficiently in preparation for and during migration. Our results provide a significant contribution toward understanding the evolution of migratory behavior and vital insight into ongoing conservation and management efforts for the western burrowing owl.

We announce the assembly of the first de novo reference genome for the California Scrub-Jay (Aphelocoma californica). The genus Aphelocoma comprises four currently recognized species including many locally adapted populations across Mesoamerica and North America. Intensive study of Aphelocoma has revealed novel insights into the evolutionary mechanisms driving diversification in natural systems. Additional insights into the evolutionary history of this group will require continued development of high-quality, publicly available genomic resources. We extracted high molecular weight genomic DNA from a female California Scrub-Jay from northern California and generated PacBio HiFi long-read data and Omni-C chromatin conformation capture data. We used these data to generate a de novo partially phased diploid genome assembly, consisting of two pseudo-haplotypes, and scaffolded them using inferred physical proximity information from the Omni-C data. The more complete pseudo-haplotype assembly (arbitrarily designated "Haplotype 1") is 1.35 Gb in total length, highly contiguous (contig N50 = 11.53 Mb), and highly complete (BUSCO completeness score = 97%), with comparable scaffold sizes to chromosome-level avian reference genomes (scaffold N50 = 66.14 Mb). Our California Scrub-Jay assembly is highly syntenic with the New Caledonian Crow reference genome despite ~10 million years of divergence, highlighting the temporal stability of the avian genome. This high-quality reference genome represents a leap forward in publicly available genomic resources for Aphelocoma, and the family Corvidae more broadly. Future work using Aphelocoma as a model for understanding the evolutionary forces generating and maintaining biodiversity across phylogenetic scales can now benefit from a highly contiguous, in-group reference genome.

Rattlesnakes play important roles in their ecosystems by regulating prey populations, are involved in complex coevolutionary dynamics with their prey, and exhibit a variety of unusual adaptations, including maternal care, heat-sensing pit organs, hinged fangs, and medically-significant venoms. The western rattlesnake (Crotalus oreganus) is one of the widest ranging rattlesnake species, with a distribution from British Columbia, where it is listed as threatened, to Baja California and east across the Great Basin to western Wyoming, Colorado and New Mexico. Here, we report a new reference genome assembly for one of six currently recognized subspecies, C. oreganus helleri, as part of the California Conservation Genomics Project (CCGP). Consistent with the reference genomic sequencing strategy of the CCGP, we used Pacific Biosciences HiFi long reads and Hi-C chromatin-proximity sequencing technology to produce a de novo assembled genome. The assembly comprises a total of 698 scaffolds spanning 1,564,812,557 base pairs, has a contig N50 of 64.7 Mb, a scaffold N50 of 110.8 Mb, and BUSCO complete score of 90.5%. This reference genome will be valuable for studies on the genomic basis of venom evolution and variation within Crotalus, in resolving the taxonomy of C. oreganus and its relatives, and for the conservation and management of rattlesnakes in general.

Snakes in the family Colubridae include more than 2,000 currently recognized species, and comprise roughly 75% of the global snake species diversity on Earth. For such a spectacular radiation, colubrid snakes remain poorly understood ecologically and genetically. Two subfamilies, Colubrinae (788 species) and Dipsadinae (833 species), comprise the bulk of colubrid species richness. Dipsadines are a speciose and diverse group of snakes that largely inhabit Central and South America, with a handful of small-body-size genera that have invaded North America. Among them, the ring-necked snake, Diadophis punctatus, has an incredibly broad distribution with 14 subspecies. Given its continental distribution and high degree of variation in coloration, diet, feeding ecology, and behavior, the ring-necked snake is an excellent species for the study of genetic diversity and trait evolution. Within California, six subspecies form a continuously distributed "ring species" around the Central Valley, while a seventh, the regal ring-necked snake, Diadophis punctatus regalis is a disjunct outlier and Species of Special Concern in the state. Here, we report a new reference genome assembly for the San Diego ring-necked snake, D. p. similis, as part of the California Conservation Genomics Project. This assembly comprises a total of 444 scaffolds spanning 1,783 Mb and has a contig N50 of 8.0 Mb, scaffold N50 of 83 Mb, and BUSCO completeness score of 94.5%. This reference genome will be a valuable resource for studies of the taxonomy, conservation, and evolution of the ring-necked snake across its broad, continental distribution.

Acarospora socialis, the bright cobblestone lichen, is commonly found in southwestern North America. This charismatic yellow lichen is a species of key ecological significance as it is often a pioneer species in new environments. Despite their ecological importance virtually no research has been conducted on the genomics of A. socialis. To address this, we used long-read sequencing to generate the first high-quality draft genome of A. socialis. Lichen thallus tissue was collected from Pinkham Canyon in Joshua Tree National Park, California and deposited in the UC Riverside herbarium under accession #295874. The de novo assembly of the mycobiont partner of the lichen was generated from Pacific Biosciences HiFi long reads and Dovetail Omni-C chromatin capture data. After removing algal and bacterial contigs, the fungal genome was approximately 31.2 Mb consisting of 38 scaffolds with contig and scaffold N50 of 2.4 Mb. The BUSCO completeness score of the assembled genome was 97.5% using the Ascomycota gene set. Information on the genome of A. socialis is important for California conservation purposes given that this lichen is threatened in some places locally by wildfires due to climate change. This reference genome will be used for understanding the genetic diversity, population genomics, and comparative genomics of A. socialis species. Genomic resources for this species will support population and landscape genomics investigations, exploring the use of A. socialis as a bioindicator species for climate change, and in studies of adaptation by comparing populations that occur across aridity gradients in California.

The California Pipevine Swallowtail Butterfly, Battus philenor hirsuta, and its host plant, the California Pipevine or Dutchman's Pipe, Aristolochia californica Torr., are an important California endemic species pair. While this species pair is an ideal system to study co-evolution, genomic resources for both are lacking. Here, we report a new, chromosome-level assembly of B. philenor hirsuta as part of the California Conservation Genomics Project (CCGP). Following the sequencing and assembly strategy of the CCGP, we used Pacific Biosciences HiFi long reads and Hi-C chromatin proximity sequencing technology to produce a de novo assembled genome. Our genome assembly, the first for any species in the genus, contains 109 scaffolds spanning 443 mega base (Mb) pairs, with a contig N50 of 14.6 Mb, a scaffold N50 of 15.2 Mb, and BUSCO complete score of 98.9%. In combination with the forthcoming A. californica reference genome, the B. philenor hirsuta genome will be a powerful tool for documenting landscape genomic diversity and plant-insect co-evolution in a rapidly changing California landscape.

Diaphorina citri is the hemipteran pest and vector of a devastating bacterial pathogen of citrus worldwide. In addition to the two core bacterial endosymbionts of D. citri, Candidatus Carsonella ruddii and Candidatus Profftella armatura, the genome of a novel endosymbiont and as of yet undescribed microbe was discovered in a Hawaiian D. citri population through deep sequencing of multiple D. citri populations. Found to be closely related to the genus Asaia in the family Acetobacteraceae by 16S rRNA gene sequence analysis, it forms a sister clade along with other insect-associated 16S rRNA gene sequences from uncultured bacterium found associated with Aedes koreicus and Sogatella furcifera. Multilocus sequence analysis confirmed the phylogenetic placement sister to the Asaia clade. Despite the culturable Asaia clade being the closest phylogenetic neighbour, attempts to culture this newly identified bacterial endosymbiont were unsuccessful. On the basis of these distinct genetic differences, the novel endosymbiont is proposed to be classified into a candidate genus and species 'Candidatus Kirkpatrickella diaphorinae'. The full genome was deposited in GenBank (accession number CP107052; prokaryotic 16S rRNA OP600170).

Phylogenetics has been foundational to SARS-CoV-2 research and public health policy, assisting in genomic surveillance, contact tracing, and assessing emergence and spread of new variants. However, phylogenetic analyses of SARS-CoV-2 have often relied on tools designed for de novo phylogenetic inference, in which all data are collected before any analysis is performed and the phylogeny is inferred once from scratch. SARS-CoV-2 data sets do not fit this mold. There are currently over 14 million sequenced SARS-CoV-2 genomes in online databases, with tens of thousands of new genomes added every day. Continuous data collection, combined with the public health relevance of SARS-CoV-2, invites an "online" approach to phylogenetics, in which new samples are added to existing phylogenetic trees every day. The extremely dense sampling of SARS-CoV-2 genomes also invites a comparison between likelihood and parsimony approaches to phylogenetic inference. Maximum likelihood (ML) and pseudo-ML methods may be more accurate when there are multiple changes at a single site on a single branch, but this accuracy comes at a large computational cost, and the dense sampling of SARS-CoV-2 genomes means that these instances will be extremely rare because each internal branch is expected to be extremely short. Therefore, it may be that approaches based on maximum parsimony (MP) are sufficiently accurate for reconstructing phylogenies of SARS-CoV-2, and their simplicity means that they can be applied to much larger data sets. Here, we evaluate the performance of de novo and online phylogenetic approaches, as well as ML, pseudo-ML, and MP frameworks for inferring large and dense SARS-CoV-2 phylogenies. Overall, we find that online phylogenetics produces similar phylogenetic trees to de novo analyses for SARS-CoV-2, and that MP optimization with UShER and matOptimize produces equivalent SARS-CoV-2 phylogenies to some of the most popular ML and pseudo-ML inference tools. MP optimization with UShER and matOptimize is thousands of times faster than presently available implementations of ML and online phylogenetics is faster than de novo inference. Our results therefore suggest that parsimony-based methods like UShER and matOptimize represent an accurate and more practical alternative to established ML implementations for large SARS-CoV-2 phylogenies and could be successfully applied to other similar data sets with particularly dense sampling and short branch lengths.

Eusocial Hymenoptera are often characterized by having facultatively or obligately sterile worker castes. However, findings across an increasing number of species are that some workers are non-natal-they have 'drifted' away from where they were born and raised. Moreover, drifters are often indistinguishable from natal workers in the work and benefits provided to joined groups. This seems an evolutionary paradox of providing benefits to potentially unrelated individuals over close kin. Rather than being mistakes, drifting is proposed to be adaptive if joiners either gain inclusive fitness by preferentially moving to other kin groups or through generalized reciprocity in which exchanging workers across groups raises group-level genetic diversity and creates social heterosis. It is unclear, however, if reciprocity is unlikely because of a susceptibility to cheating. In resolving this question, a series of evolutionary simulations show: (1) Reciprocity can persist under a range of genetic assumptions and scenarios of cheating, (2) cheating almost always evolves, but can be expressed in a variety of ways that are not always predictable, (3) the inclusive fitness hypothesis is equally or more susceptible to cheating. Moreover, existing data in Hymenoptera (although not extensive) are more consistent with generalized reciprocity. This supports a hypothesis that drifting, as a phenomenon, may more often reflect maximization of group and parental fitness rather than fitness gains for the individual drifters.