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New insights into the phylogenetics and population structure of the prairie falcon (Falco mexicanus)

Background Management requires a robust understanding of between- and within-species genetic variability, however such data are still lacking in many species. For example, although multiple population genetics studies of the peregrine falcon ( Falco peregrinus ) have been conducted, no similar studies have been done of the closely-related prairie falcon ( F. mexicanus ) and it is unclear how much genetic variation and population structure exists across the species’ range. Furthermore, the phylogenetic relationship of F. mexicanus relative to other falcon species is contested. We utilized a genomics approach (i.e., genome sequencing and assembly followed by single nucleotide polymorphism genotyping) to rapidly address these gaps in knowledge. Results We sequenced the genome of a single female prairie falcon and generated a 1.17 Gb (gigabases) draft genome assembly. We generated maximum likelihood phylogenetic trees using complete mitochondrial genomes as well as nuclear protein-coding genes. This process provided evidence that F. mexicanus is an outgroup to the clade that includes the peregrine falcon and members of the subgenus Hierofalco. We annotated > 16,000 genes and almost 600,000 high-quality single nucleotide polymorphisms (SNPs) in the nuclear genome, providing the raw material for a SNP assay design featuring > 140 gene-associated markers and a molecular-sexing marker. We subsequently genotyped ~ 100 individuals from California (including the San Francisco East Bay Area, Pinnacles National Park and the Mojave Desert) and Idaho (Snake River Birds of Prey National Conservation Area). We tested for population structure and found evidence that individuals sampled in California and Idaho represent a single panmictic population. Conclusions Our study illustrates how genomic resources can rapidly shed light on genetic variability in understudied species and resolve phylogenetic relationships. Furthermore, we found evidence of a single, randomly mating population of prairie falcons across our sampling locations. Prairie falcons are highly mobile and relatively rare long-distance dispersal events may promote gene flow throughout the range. As such, California’s prairie falcons might be managed as a single population, indicating that management actions undertaken to benefit the species at the local level have the potential to influence the species as a whole.

BMC Genomics

Perspectives: Gene Expression in Fisheries Management

Functional genes and gene expression have been connected to physiological traits linked to effective production and broodstock selection in aquaculture, selective implications of commercial fish harvest, and adaptive changes reflected in non-commercial fish populations subject to human disturbance and climate change. Gene mapping using single nucleotide polymorphisms (SNPs) to identify functional genes, gene expression (analogue microarrays and real-time PCR), and digital sequencing technologies looking at RNA transcripts present new concepts and opportunities in support of effective and sustainable fisheries. Genomic tools have been rapidly growing in aquaculture research addressing aspects of fish health, toxicology, and early development. Genomic technologies linking effects in functional genes involved in growth, maturation and life history development have been tied to selection resulting from harvest practices. Incorporating new and ever-increasing knowledge of fish genomes is opening a different perspective on local adaptation that will prove invaluable in wild fish conservation and management. Conservation of fish stocks is rapidly incorporating research on critical adaptive responses directed at the effects of human disturbance and climate change through gene expression studies. Genomic studies of fish populations can be generally grouped into three broad categories: 1) evolutionary genomics and biodiversity; 2) adaptive physiological responses to a changing environment; and 3) adaptive behavioral genomics and life history diversity. We review current genomic research in fisheries focusing on those that use microarrays to explore differences in gene expression among phenotypes and within or across populations, information that is critically important to the conservation of fish and their relationship to humans.

Current Zoology

Yosemite toad (Anaxyrus canorus) transcriptome reveals interplay between speciation genes and adaptive introgression

Genomes are heterogeneous during the early stages of speciation, with small ‘islands’ of DNA appearing to reflect strong adaptive differences, surrounded by vast seas of relative homogeneity. As species diverge, secondary contact zones between them can act as an interface and selectively filter through advantageous alleles of hybrid origin. Such introgression is another important adaptive process, one that allows beneficial mosaics of recombinant DNA (‘rivers’) to flow from one species into another. Although genomic islands of divergence appear to be associated with reproductive isolation, and genomic rivers form by adaptive introgression, it is unknown whether islands and rivers tend to be the same or different loci. We examined three replicate secondary contact zones for the Yosemite toad ( Anaxyrus canorus ) using two genomic data sets and a morphometric data set to answer the questions: (1) How predictably different are islands and rivers, both in terms of genomic location and gene function? (2) Are the adaptive genetic trait loci underlying tadpole growth and development reliably islands, rivers or neither? We found that island and river loci have significant overlap within a contact zone, suggesting that some loci are first islands, and later are predictably converted into rivers. However, gene ontology enrichment analysis showed strong overlap in gene function unique to all island loci, suggesting predictability in overall gene pathways for islands. Genome-wide association study outliers for tadpole development included LPIN3, a lipid metabolism gene potentially involved in climate change adaptation, that is island-like for all three contact zones, but also appears to be introgressing (as a river) across one zone. Taken together, our results suggest that adaptive divergence and introgression may be more complementary forces than currently appreciated.

Molecular Ecology

Concordant signal of genetic variation across marker densities in the desert annual Chylismia brevipes is linked with timing of winter precipitation

Climate change coupled with large-scale surface disturbances necessitate active restoration strategies to promote resilient and genetically diverse native plant communities. However, scarcity of native plant materials hinders restoration efforts, leading practitioners to choose from potentially viable but nonlocal seed sources. Genome scans for genetic variation linked with selective environmental gradients have become a useful tool in such efforts, allowing rapid delineation of seed transfer zones along with predictions of genomic vulnerability to climate change. When properly applied, genome scans can reduce the risk of maladaptation due to mismatches between seed source and planting site. However, results are rarely replicated among complimentary data sources. Here, we compared RAD-seq datasets with 819 and 2699 SNPs (in 625 and 356 individuals, respectively) from the Mojave Desert winter annual Chylismia brevipes . Overall, we found that the datasets consistently characterized both neutral population structure and genetic–environmental associations. Ancestry analyses indicated consistent spatial genetic structuring into four regional populations. We also detected a marked signal of isolation by resistance (IBR), wherein spatial genetic structure was better explained by habitat resistance than by geographic distance. Potentially adaptive loci identified from genome scans were associated with the same environmental gradients—fall precipitation, winter minimum temperature, and precipitation timing—regardless of dataset. Paired with our finding that habitat resistance best explained genetic divergence, our results suggest that isolation of populations within environmentally similar habitats—and subsequent local adaption along gradients parallel to these habitats—drive genome-wide divergence in this species. Moreover, strong genetic associations with winter precipitation timing, along with forecasted shifts in precipitation regime due to midcentury climate change, could impact future population dynamics, habitat distribution, and genetic connectivity for C. brevipes populations within the Mojave Desert.

Arizona, California, Nevada

Exchange of carbapenem-resistant Escherichia coli Sequence Type 38 intercontinentally and among wild bird, human, and environmental niches

Carbapenem-resistant Enterobacteriaceae (CRE) are a global threat to human health and are increasingly being isolated from nonclinical settings. OXA-48-producing Escherichia coli sequence type 38 (ST38) is the most frequently reported CRE type in wild birds and has been detected in gulls or storks in North America, Europe, Asia, and Africa. The epidemiology and evolution of CRE in wildlife and human niches, however, remains unclear. We compared wild bird origin E. coli ST38 genome sequences generated by our research group and publicly available genomic data derived from other hosts and environments to (i) understand the frequency of intercontinental dispersal of E. coli ST38 clones isolated from wild birds, (ii) more thoroughly measure the genomic relatedness of carbapenem-resistant isolates from gulls sampled in Turkey and Alaska, USA, using long-read whole-genome sequencing and assess the spatial dissemination of this clone among different hosts, and (iii) determine whether ST38 isolates from humans, environmental water, and wild birds have different core or accessory genomes (e.g., antimicrobial resistance genes, virulence genes, plasmids) which might elucidate bacterial or gene exchange among niches. Our results suggest that E. coli ST38 strains, including those resistant to carbapenems, are exchanged between humans and wild birds, rather than separately maintained populations within each niche. Furthermore, despite close genetic similarity among OXA-48-producing E. coli ST38 clones from gulls in Alaska and Turkey, intercontinental dispersal of ST38 clones among wild birds is uncommon. Interventions to mitigate the dissemination of antimicrobial resistance throughout the environment (e.g., as exemplified by the acquisition of carbapenem resistance by birds) may be warranted.

Applied and Environmental Microbiology

Mixotrophic iron-oxidizing Thiomonas isolates from an acid mine drainage-affected creek

Natural attenuation of heavy metals occurs via coupled microbial iron cycling and metal precipitation in creeks impacted by acid mine drainage (AMD). Here, we describe the isolation, characterization, and genomic sequencing of two iron-oxidizing bacteria (FeOB) species: Thiomonas ferrovorans FB-6 and Thiomonas metallidurans FB-Cd, isolated from slightly acidic (pH 6.3), Fe-rich, AMD-impacted creek sediments. These strains precipitated amorphous iron oxides, lepidocrocite, goethite, and magnetite or maghemite and grew at a pH optimum of 5.5. While Thiomonas spp. are known as mixotrophic sulfur oxidizers and As oxidizers, the FB strains oxidized Fe, which suggests they can efficiently remove Fe and other metals via coprecipitation. Previous evidence for Thiomonas sp. Fe oxidation is largely ambiguous, possibly because of difficulty demonstrating Fe oxidation in heterotrophic/mixotrophic organisms. Therefore, we also conducted a genomic analysis to identify genetic mechanisms of Fe oxidation, other metal transformations, and additional adaptations, comparing the two FB strain genomes with 12 other Thiomonas genomes. The FB strains fall within a relatively novel group of Thiomonas strains that includes another strain (b6) with solid evidence of Fe oxidation. Most Thiomonas isolates, including the FB strains, have the putative iron oxidation gene cyc2 , but only the two FB strains possess the putative Fe oxidase genes mtoAB . The two FB strain genomes contain the highest numbers of strain-specific gene clusters, greatly increasing the known Thiomonas genetic potential. Our results revealed that the FB strains are two distinct novel species of Thiomonas with the genetic potential for bioremediation of AMD via iron oxidation.

Thuringia

Identification of a novel Adélie penguin circovirus at Cape Crozier (Ross Island, Antarctica)

Understanding the causes of disease in Antarctic wildlife is crucial as many of these species are already threatened by environmental changes brought about by climate change. In recent years, Antarctic penguins have been showing signs of an unknown pathology: a feather disorder characterised by missing feathers resulting in exposed skin. During the 2018-19 austral summer breeding season at Cape Crozier colony on Ross Island, Antarctica, for the first time we observed an Adlie penguin chick missing down over most of its body. A guano sample was collected from the nest of the featherless chick and, using high throughput sequencing we identified a novel circovirus. Using abutting primers, we amplified the full genome, which we cloned, and Sanger sequenced to determine the complete genome of the circovirus. The Adlie penguin guano-associated circovirus genome shares <67% genome-wide nucleotide identity to other circoviruses, representing a new species of circovirus, therefore we named it penguin circovirus (PenCV). Using the same primer pair, we screened 25 previously collected cloacal swabs taken at Cape Crozier from known-age adult Adlie penguins during the 2014-15 season displaying no clinical signs of feather-loss disorder. Three of the 25 samples (12%) were positive for a PenCV, whose genome shared >99% pairwise identity to the one identified in 2018-19. This is the first report of a circovirus associated with a penguin species and could be a possible etiological agent of the feather-loss disorder in Antarctic penguins.

Viruses

Novel avian papillomaviruses identified in a south polar skua sampled on Ross Island, Antarctica

Papillomaviruses are small circular DNA viruses that infect epithelial cells of their hosts. Avian papillomaviruses are poorly sampled/documented compared to those infecting humans. We used a viral metagenomic approach to identify viruses from the oral swab taken from a deceased south polar skua ( Stercorarius maccormicki ) found at Cape Royds, Ross Island, Antarctica in late 2024. We identified three papillomaviruses and determined their complete genomes, Stercorarius maccormicki papillomavirus (SmacPV) 1-3. SmacPV1 is the most divergent of the three SmacPVs, sharing 62% genome-wide pairwise identity to SmacPV2 and SmacPV3 and <63.5% to other avian papillomaviruses. The genomes of SmacPV2 and SmacPV3 represent two new papillomavirus types sharing 82.4% genome-wide pairwise identity with each other and <72% to other papillomaviruses. SmacPV2 and SmacPV3 phylogenetically cluster with sequences of the Rissa tridactyla papillomavirus 1 from black-legged kittiwake ( Rissa tridactyla ), Larus smithsonianus papillomavirus 1 from American herring gull ( Larus smithsonianus ) and Fratercula arctica papillomavirus 1 from Atlantic puffin ( Fratercula arctica ), and they collectively represent a new papillomavirus species. These are the first papillomaviruses to be identified in Stercorarius spp . and add to the handful of known papillomaviruses in identified avian species. We also expand the known host range of papillomaviruses in Antarctic animals, which previously included Adélie penguins ( Pygoscelis adeliae ), Weddell seals ( Leptonychotes weddellii ), Antarctic fur seals ( Arctocephalus gazella ), leopard seals ( Hydrurga leptonyx ) and emerald notothen ( Trematomus bernacchii ).

Archives of Virology

Female persistence during toxicant treatment predicts survival probability of offspring in invasive brown treesnakes (Boiga irregularis)

Assessing the long-term efficacy of control methods is a critical component of invasive species management. For example, if traits related to control have significant heritability or are influenced by maternal effects, control methods may lose efficacy over time. The potential for these effects can be evaluated via parent/offspring survival analysis, which concomitantly recasts adaptive management as an evolutionary force for invasive species. However, difficulties can arise when the life history of an invasive is cryptic, precluding direct observations of familial relationships. Genomic pedigree reconstruction can facilitate such analyses by assigning offspring to parents in invasive species for which mating and reproduction are difficult to study. Here, we use genomic pedigree reconstruction to quantify parental longevity and probability of offspring survival for brown treesnakes ( Boiga irregularis ) on Guam in a landscape treated with toxic baits simulating application via an aerial delivery system (ADS). To do so, we used 398 single nucleotide polymorphisms (SNPs) to update an existing multi-generational genomic pedigree for a geographically-closed population of brown treesnakes. This facilitated assignment of parents to juveniles born during three consecutive years of toxic bait application under a simulated aerial treatment program (N = 72). We found that the offspring of dams that persisted until the end of the study displayed greater survival probability (cox proportional hazard model, P < 0.001), yet there was no such effect for sires. This sex-specific relationship between parental longevity and offspring survival indicates that heritability of traits contributing to resistance to ADS is unlikely, but it supports a role of maternal effects that could undermine ADS. Our study identifies potential risks associated with control efforts and also highlights the utility of parent-offspring survival analyses informed by genomic pedigree reconstruction as a tool for adaptive management.

Global Ecology and Conservation

Toxicogenomics in regulatory ecotoxicology

Recently, we have witnessed an explosion of different genomic approaches that, through a combination of advanced biological, instrumental, and bioinformatic techniques, can yield a previously unparalleled amount of data concerning the molecular and biochemical status of organisms. Fueled partially by large, well-publicized efforts such as the Human Genome Project, genomic research has become a rapidly growing topical area in multiple biological disciplines. Since 1999, when the term “toxicogenomics” was coined to describe the application of genomics to toxicology (1), a rapid increase in publications on the topic has occurred (Figure 1). The potential utility of toxicogenomics in toxicological research and regulatory activities has been the subject of scientific discussions and, as with any new technology, has evoked a wide range of opinion (2–6).

Environmental Science & Technology

Historical DNA reveals climate adaptation in an endangered songbird

To cope with climate change, species may shift their distributions or adapt in situ to changing environmental conditions. However, clear examples of genetic changes via adaptation are limited. We explore evolutionary responses to climate change in the endangered southwestern willow flycatcher ( Empidonax traillii extimus ) through whole-genome comparisons between historical specimens, collected from 1888 to 1909 near San Diego, California, United States, and contemporary individuals from across the breeding range. Genomic analyses revealed that introgression into San Diego increased adaptive potential over time and shifted genome-wide population structure towards that of neighbouring populations. In contrast, loci linked to climate (dew point temperature and precipitation) shifted away from neighbouring populations and in a direction consistent with adaptation to climate change in southern California. This research highlights the role of admixture in facilitating adaptive shifts through its impact on genome-wide genetic variation and represents one of the few studies to document climate adaptation in a wild population.

Nature Climate Change

Frequent transitions in mating-type locus chromosomal organization in Malassezia and early steps in sexual reproduction

Fungi in the basidiomycete genus Malassezia are the most prevalent eukaryotic microbes resident on the skin of human and other warm-blooded animals and have been implicated in skin diseases and systemic disorders. Analysis of Malassezia genomes revealed that key adaptations to the skin microenvironment have a direct genomic basis, and the identification of mating/meiotic genes suggests a capacity to reproduce sexually, even though no sexual cycle has yet been observed. In contrast to other bipolar or tetrapolar basidiomycetes that have either two linked mating-type-determining ( MAT ) loci or two MAT loci on separate chromosomes, in Malassezia species studied thus far the two MAT loci are arranged in a pseudobipolar configuration (linked on the same chromosome but capable of recombining). By generating additional chromosome-level genome assemblies, and an improved Malassezia phylogeny, we infer that the pseudobipolar arrangement was the ancestral state of this group and revealed six independent transitions to tetrapolarity, seemingly driven by centromere fission or translocations in centromere-flanking regions. Additionally, in an approach to uncover a sexual cycle, Malassezia furfur strains were engineered to express different MAT alleles in the same cell. The resulting strains produce hyphae reminiscent of early steps in sexual development and display upregulation of genes associated with sexual development as well as others encoding lipases and a protease potentially relevant for pathogenesis of the fungus. Our study reveals a previously unseen genomic relocation of mating-type loci in fungi and provides insight toward the identification of a sexual cycle in Malassezia , with possible implications for pathogenicity.

PNAS

Identical sequence types of Yersinia ruckeri associated with lethal disease in wild-caught invasive Blue Catfish and cultured hybrid catfish (Channel Catfish ♀ × Blue Catfish ♂) from disparate aquatic ecosystems

Objective The Blue Catfish Ictalurus furcatus is commonly raised in warmwater aquaculture in the United States to produce Channel Catfish I. punctatus × Blue Catfish hybrids. It is also a prominent aquatic invasive species of concern in the mid-Atlantic region of the United States. Here, Yersina ruckeri was isolated from moribund Blue Catfish and hybrid catfish from disparate regions of the USA. The goal of the research here was to compare these Y. ruckeri strains to each other and other known strains for which adequate sequence data was available. In addition, we sought to determine if the strain from Blue Catfish was pathogenic to Rainbow Trout Oncorhynchus mykiss . Methods Moribund hybrid catfish from culture ponds in Mississippi were processed for diagnostic evaluation in March 2016. In April 2022, a moribund Blue Catfish specimen was collected from a tributary of the Nanticoke River in Maryland. Bacterial isolates were identified and characterized using biochemical tests, antimicrobial sensitivity screening, serotyping, and complete or partial genome sequencing. Disease pathology was described via histology. The isolate from Blue Catfish was used in challenge experiments to determine if it was pathogenic to Rainbow Trout. Multilocus sequencing typing was conducted using the PubMLST database. Results Biochemical testing was consistent with Y. ruckeri . A draft genome of the Y. ruckeri isolate was assembled based on Oxford Nanopore Technology sequencing and identified a single genomic replicon (3,791,418 bp) consistent in size to other Y. ruckeri genomes and a pLT plasmid (60, 933 bp). The challenge study demonstrated no significant virulence of this isolate for Rainbow Trout ( Y. ruckeri ). This isolate was most similar to other strains isolated from ictalurids. Notably, the gyrase B gene from this isolate was identical to that of archived strains isolated from moribund Mississippi hybrid catfish aquaculture during 2016 and these isolates share identical PubMLST sequence type profiles. Similarly, they shared a pLT plasmid that differed by only 6 bp. This plasmid has never been reported from trout isolates and appears to be unique to ictalurids. Conclusions Analyses here provide preliminary genetic evidence that geographically distant (Maryland and Mississippi, USA) isolates of Y. ruckeri from ictalurids are genetically similar to each other and Y. ruckeri (strain SC09) that infects ictalurids in China. This strain is not a biothreat to Rainbow Trout at typical culture temperatures.

Journal of Aquatic Animal Health

Incomplete bioinformatic filtering and inadequate age and growth analysis lead to an incorrect inference of harvested-induced changes

Understanding the evolutionary impacts of harvest on fish populations is important for informing fisheries management and conservation and has become a growing research topic over the last decade. However, the dynamics of fish populations are highly complex, and phenotypes can be influenced by many biotic and abiotic factors. Therefore, it is vital to collect robust data and explore multiple alternative hypotheses before concluding that fish populations are influenced by harvest. In their recently published manuscript, Bowles et al, Evolutionary Applications, 13(6):1128 conducted age/growth and genomic analysis of walleye ( Sander vitreus ) populations sampled 13–15 years (1–2.5 generations) apart and hypothesized that observed phenotypic and genomic changes in this time period were likely due to harvest. Specifically, Bowles et al. (2020) documented differential declines in size-at-age in three exploited walleye populations compared to a separate, but presumably less-exploited, reference population. Additionally, they documented population genetic differentiation in one population pair, homogenization in another, and outlier loci putatively under selection across time points. Based on their phenotypic and genetic results, they hypothesized that selective harvest had led to fisheries-induced evolution (referred to as nascent changes) in the exploited populations in as little as 1–2.5 generations. We re-analyzed their data and found that (a) sizes declined across both exploited and reference populations during the time period studied and (b) observed genomic differentiation in their study was the result of inadequate data filtering, including retaining individuals with high amounts of missing data and retaining potentially undersplit and oversplit loci that created false signals of differentiation between time points. This re-analysis did not provide evidence for phenotypic or genetic changes attributable to harvest in any of the study populations, contrasting the hypotheses presented by Bowles et al. (2020). Our comment highlights the potential pitfalls associated with conducting age/growth analyses with low sample sizes and inadequately filtering genomic datasets.

Quebec

A genetic signature of the evolution of loss of flight in the Galapagos cormorant

INTRODUCTION Changes in the size and proportion of limbs and other structures have played a key role in the evolution of species. One common class of limb modification is recurrent wing reduction and loss of flight in birds. Indeed, Darwin used the occurrence of flightless birds as an argument in favor of his theory of natural selection. Loss of flight has evolved repeatedly and is found among 26 families of birds in 17 different orders. Despite the frequency of these modifications, we have a limited understanding of their underpinnings at the genetic and molecular levels. RATIONALE To better understand the evolution of changes in limb size, we studied a classic case of recent loss of flight in the Galapagos cormorant ( Phalacrocorax harrisi ). Cormorants are large water birds that live in coastal areas or near lakes, and P. harrisi is the only flightless cormorant among approximately 40 extant species. The entire population is distributed along the coastlines of Isabela and Fernandina islands in the Galapagos archipelago. P. harrisi has a pair of short wings, which are smaller than those of any other cormorant. The extreme reduction of the wings and pectoral skeleton observed in P. harrisi is an attractive model for studying the evolution of loss of flight because it occurred very recently; phylogenetic evidence suggests that P. harrisi diverged from its flighted relatives within the past 2 million years. We developed a comparative and predictive genomics approach that uses the genome sequences of P. harrisi and its flighted relatives to find candidate genetic variants that likely contributed to the evolution of loss of flight. RESULTS We sequenced and de novo assembled the whole genomes of P. harrisi and three closely related flighted cormorant species. We identified thousands of coding variants exclusive to P. harrisi and classified them according to their probability of altering protein function based on conservation. Variants most likely to alter protein function were significantly enriched in genes mutated in human skeletal ciliopathies, including Ofd1 , Evc , Wdr34 , and Ift122 . We carried out experiments in Caenorhabditis elegans to confirm that a missense variant present in the Galapagos cormorant IFT122 protein is sufficient to affect ciliary function. The primary cilium is essential for Hedgehog (Hh) signaling in vertebrates, and individuals affected by ciliopathies have small limbs and ribcages, mirroring the phenotype of P. harrisi . We also identified a 4–amino acid deletion in the regulatory domain of Cux1 , a highly conserved transcription factor that has been experimentally shown to regulate limb growth in chicken. The four missing amino acids are perfectly conserved in all birds and mammals sequenced to date. We tested the consequences of this deletion in a chondrogenic cell line and showed that it impairs the ability of CUX1 to transcriptionally up-regulate cilia-related genes (some of which contain function-altering variants in P. harrisi ) and to promote chondrogenic differentiation. Finally, we show that positive selection may have played a role in the fixation of the variants associated with loss of flight in P. harrisi . CONCLUSION Our results indicate that the combined effect of variants in genes necessary for the correct transcriptional regulation and function of the primary cilium likely contributed to the evolution of highly reduced wings and other skeletal adaptations associated with loss of flight in P. harrisi . Our approach may be generally useful for identification of variants underlying evolutionary novelty from genomes of closely related species.

Science

Detection of diazotrophy in the acetylene-fermenting anaerobe Pelobacter sp. strain SFB93

Acetylene (C2H2) is a trace constituent of the present Earth's oxidizing atmosphere, reflecting a mixture of terrestrial and marine emissions from anthropogenic, biomass-burning, and unidentified biogenic sources. Fermentation of acetylene was serendipitously discovered during C2H2 block assays of N2O reductase, and Pelobacter acetylenicus was shown to grow on C2H2 via acetylene hydratase (AH). AH is a W-containing, catabolic, low-redox-potential enzyme that, unlike nitrogenase (N2ase), is specific for acetylene. Acetylene fermentation is a rare metabolic process that is well characterized only in P. acetylenicus DSM3246 and DSM3247 and Pelobacter sp. strain SFB93. To better understand the genetic controls for AH activity, we sequenced the genomes of the three acetylene-fermenting Pelobacter strains. Genome assembly and annotation produced three novel genomes containing gene sequences for AH, with two copies being present in SFB93. In addition, gene sequences for all five compulsory genes for iron-molybdenum N2ase were also present in the three genomes, indicating the cooccurrence of two acetylene transformation pathways. Nitrogen fixation growth assays showed that DSM3426 could ferment acetylene in the absence of ammonium, but no ethylene was produced. However, SFB93 degraded acetylene and, in the absence of ammonium, produced ethylene, indicating an active N2ase. Diazotrophic growth was observed under N2 but not in experimental controls incubated under argon. SFB93 exhibits acetylene fermentation and nitrogen fixation, the only known biochemical mechanisms for acetylene transformation. Our results indicate complex interactions between N2ase and AH and suggest novel evolutionary pathways for these relic enzymes from early Earth to modern days.

Applied and Environmental Microbiology

Development and application of a qPCR-based genotyping assay for Ophidiomyces ophidiicola to investigate the epidemiology of ophidiomycosis

Ophidiomycosis (snake fungal disease) is an infectious disease caused by the fungus Ophidiomyces ophidiicola to which all snake species appear to be susceptible. Significant variation has been observed in clinical presentation, progression of disease, and response to treatment, which may be due to genetic variation in the causative agent. Recent phylogenetic analysis based on whole-genome sequencing identified that O . ophidiicola strains from the United States formed a clade distinct from European strains, and that multiple clonal lineages of the clade are present in the United States. The purpose of this study was to design a qPCR-based genotyping assay for O . ophidiicola , then apply that assay to swab-extracted DNA samples to investigate whether the multiple O . ophidiicola clades and clonal lineages in the United States have specific geographic, taxonomic, or temporal predilections. To this end, six full genome sequences of O . ophidiicola representing different clades and clonal lineages were aligned to identify genomic areas shared between subsets of the isolates. Eleven hydrolysis-based Taqman primer-probe sets were designed to amplify selected gene segments and produce unique amplification patterns for each isolate, each with a limit of detection of 10 or fewer copies of the target sequence and an amplification efficiency of 90–110%. The qPCR-based approach was validated using samples from strains known to belong to specific clades and applied to swab-extracted O . ophidiicola DNA samples from multiple snake species, states, and years. When compared to full-genome sequencing, the qPCR-based genotyping assay assigned 75% of samples to the same major clade (Cohen’s kappa = 0.360, 95% Confidence Interval = 0.154–0.567) with 67–77% sensitivity and 88–100% specificity, depending on clade/clonal lineage. Swab-extracted O . ophidiicola DNA samples from across the United States were assigned to six different clonal lineages, including four of the six established lineages and two newly defined groups, which likely represent recombinant strains of O . ophidiicola . Using multinomial logistic regression modeling to predict clade based on snake taxonomic group, state of origin, and year of collection, state was the most significant predictor of clonal lineage. Furthermore, clonal lineage was not associated with disease severity in the most intensely sampled species, the Lake Erie watersnake ( Nerodia sipedon insularum ). Overall, this assay represents a rapid, cost-effective genotyping method for O . ophidiicola that can be used to better understand the epidemiology of ophidiomycosis.

PLoS ONE

A phylogenomic supertree of birds

It has long been appreciated that analyses of genomic data (e.g., whole genome sequencing or sequence capture) have the potential to reveal the tree of life, but it remains challenging to move from sequence data to a clear understanding of evolutionary history, in part due to the computational challenges of phylogenetic estimation using genome-scale data. Supertree methods solve that challenge because they facilitate a divide-and-conquer approach for large-scale phylogeny inference by integrating smaller subtrees in a computationally-efficient manner. Here, we combined information from sequence capture and whole-genome phylogenies using supertree methods. However, available phylogenomic trees had limited overlap so we used taxon-rich (but not phylogenomic) megaphylogenies to weave them together. This allowed us to construct a phylogenomic supertree, with support values, that included 707 bird species (~7% of avian species diversity). We estimated branch lengths using mitochondrial sequence data and we used this to estimate divergence times. Our time-calibrated supertree supports radiation of all three major avian clades (Palaeognathae, Galloanseres, and Neoaves) near the Cretaceous-Paleogene (K-Pg) boundary. The approach we used will permit the continued addition of taxa to this supertree as new phylogenomic data are published, and it could be applied to other taxa as well.

Diversity