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Microbial rhodopsins are widely distributed in aquatic environments and may significantly contribute to phototrophy and energy budgets in global oceans. However, the study of freshwater rhodopsins has been largely limited. Here, we explored the diversity, ecological distribution, and expression of opsin genes that encode the apoproteins of Type I rhodopsins in humic and clearwater lakes with contrasting physicochemical and optical characteristics. Using metagenomes and metagenome-assembled genomes, we recovered opsin genes from a wide range of taxa, mostly predicted to encode green light-absorbing proton pumps. Viral opsin and novel bacterial opsin clades were recovered. Opsin genes occurred more frequently in taxa from clearwater than from humic water, and opsins in some taxa have non-typical ion-pumping motifs that might be associated with physicochemical conditions of these two freshwater types. Analyses of the surface layer of 33 freshwater systems revealed an inverse correlation between opsin gene abundance and lake dissolved organic carbon (DOC). In humic water with high terrestrial DOC and light-absorbing humic substances, opsin gene abundance was low and dramatically declined within the first few meters, whereas the abundance remained relatively high along the bulk water column in clearwater lakes with low DOC, suggesting opsin gene distribution is influenced by lake optical properties and DOC. Gene expression analysis confirmed the significance of rhodopsin-based phototrophy in clearwater lakes and revealed different diel expressional patterns among major phyla. Overall, our analyses revealed freshwater opsin diversity, distribution and expression patterns, and suggested the significance of rhodopsin-based phototrophy in freshwater energy budgets, especially in clearwater lakes.

Cyanobacteria and cyanotoxin producing cyanobacterial blooms are a trending focus of current research. Many studies focus on bloom events in lentic environments such as lakes or ponds. Comparatively few studies have explored lotic environments and fewer still have examined the cyanobacterial communities and potential cyanotoxin producers during ambient, non-bloom conditions. Here we used a metagenomics-based approach to profile non-bloom microbial communities and cyanobacteria in 12 major U.S. rivers at multiple time points during the summer months of 2019. Our data show that U.S. rivers possess microbial communities that are taxonomically rich, yet largely consistent across geographic location and time. Within these communities, cyanobacteria often comprise significant portions and frequently include multiple species with known cyanotoxin producing strains. We further characterized these potential cyanotoxin producing taxa by deep sequencing amplicons of the microcystin E (mcyE) gene. We found that rivers containing the highest levels of potential cyanotoxin producing cyanobacteria consistently possess taxa with the genetic potential for cyanotoxin production and that, among these taxa, the predominant genus of origin for the mcyE gene is Microcystis. Combined, these data provide a unique perspective on cyanobacteria and potential cyanotoxin producing taxa that exist in large rivers across the U.S. and can be used to better understand the ambient conditions that may precede bloom events in lotic freshwater ecosystems.

The acquisition of novel traits is central to organismal evolution, yet the molecular mechanisms underlying this process are elusive. The beetle forewings (elytra) are evolutionarily modified to serve as a protective shield, providing a unique opportunity to study these mechanisms. In the past, the orthologs of genes within the wing gene network from Drosophila studies served as the starting point when studying the evolution of elytra (candidate genes). Although effective, candidate gene lists are finite and only explore genes conserved across species. To go beyond candidate genes, we used RNA sequencing and explored the wing transcriptomes of two Coleopteran species, the red flour beetle (Tribolium castaneum) and the Japanese stag beetle (Dorcus hopei). Our analysis revealed sets of genes enriched in Tribolium elytra (57 genes) and genes unique to the hindwings, which possess more "typical" insect wing morphologies (29 genes). Over a third of the hindwing-enriched genes were "candidate genes" whose functions were previously analyzed in Tribolium, demonstrating the robustness of our sequencing. Although the overlap was limited, transcriptomic comparison between the beetle species found a common set of genes, including key wing genes, enriched in either elytra or hindwings. Our RNA interference analysis for elytron-enriched genes in Tribolium uncovered novel genes with roles in forming various aspects of morphology that are unique to elytra, such as pigmentation, hardening, sensory development, and vein formation. Our analyses deepen our understanding of how gene network evolution facilitated the emergence of the elytron, a unique structure critical to the evolutionary success of beetles.

Lignin is an abundant renewable source of aromatics and precursors for the production of other organic chemicals. However, lignin is a heterogeneous polymer, so the mixture of aromatics released during its depolymerization can make its conversion to chemicals challenging. Microbes are a potential solution to this challenge, as some can catabolize multiple aromatic substrates into one product. Novosphingobium aromaticivorans has this ability, and its use as a bacterial chassis for lignin valorization could be improved by the ability to predict product yields based on thermodynamic and metabolic inputs. In this work, we built a genome-scale metabolic model of N. aromaticivorans, iNovo479, to guide the engineering of strains for aromatic conversion into products. iNovo479 predicted product yields from single or multiple aromatics, and the impact of combinations of aromatic and non-aromatic substrates on product yields. We show that enzyme reactions from other organisms can be added to iNovo479 to predict the feasibility and profitability of producing additional products by engineered strains. Thus, we conclude that iNovo479 can help guide the design of bacteria to convert lignin aromatics into valuable chemicals.

Lignin is a potential source of valuable chemicals, but its chemical depolymerization results in a heterogeneous mixture of aromatics and other products. Microbes could valorize depolymerized lignin by converting multiple substrates into one or a small number of products. In this study, we describe the ability of Novosphingobium aromaticivorans to metabolize 1-(4-hydroxy-3-methoxyphenyl)propane-1,2-dione (G-diketone), an aromatic Hibbert diketone which is produced during formic acid-catalyzed lignin depolymerization. By assaying genome-wide transcript levels from N. aromaticivorans during growth on G-diketone and other chemically-related aromatics, we hypothesized that the Lig dehydrogenases, previously characterized as oxidizing β-O-4 linkages in aromatic dimers, were involved in G-diketone metabolism by N. aromaticivorans. Using purified N. aromaticivorans Lig dehydrogenases, we found that LigL, LigN, and LigD each reduced the Cα ketone of G-diketone in vitro but with different substrate specificities and rates. Furthermore, LigL, but not LigN or LigD, also reduced the Cα ketone of 2-hydroxy-1-(4-hydroxy-3-methoxyphenyl)propan-1-one (GP-1) in vitro, a derivative of G-diketone with the Cβ ketone reduced, when GP-1 was provided as a substrate. The newly identified activity of these Lig dehydrogenases expands the potential range of substrates utilized by N. aromaticivorans beyond what has been previously recognized. This is beneficial both for metabolizing a wide range of natural and non-native depolymerized lignin substrates and for engineering microbes and enzymes that are active with a broader range of aromatic compounds. Importance Lignin is a major plant polymer composed of aromatic units that have value as chemicals. However, the structure and composition of lignin has made it difficult to use this polymer as a renewable source of industrial chemicals. Bacteria like Novosphingobium aromaticivorans have the potential to make chemicals from lignin not only because of their natural ability to metabolize a variety of aromatics but also because there are established protocols to engineer N. aromaticivorans strains to funnel lignin-derived aromatics into valuable products. In this work, we report a newly discovered activity of previously characterized dehydrogenase enzymes with a chemically-modified byproduct of lignin depolymerization. We propose that the activity of N. aromaticivorans enzymes with both native lignin aromatics and those produced by chemical depolymerization will expand opportunities for producing industrial chemicals from the heterogenous components of this abundant plant polymer.

Lignin is a potential source of valuable chemicals, but its chemical depolymerization results in a heterogeneous mixture of aromatics and other products. Microbes could valorize depolymerized lignin by converting multiple substrates into one or a small number of products. In this study, we describe the ability of Novosphingobium aromaticivorans to metabolize 1-(4-hydroxy-3-methoxyphenyl)propane-1,2-dione (G-diketone), an aromatic Hibbert diketone that is produced during formic acid-catalyzed lignin depolymerization. By assaying genome-wide transcript levels from N. aromaticivorans during growth on G-diketone and other chemically-related aromatics, we hypothesized that the Lig dehydrogenases, previously characterized as oxidizing β-O-4 linkages in aromatic dimers, were involved in G-diketone metabolism by N. aromaticivorans. Using purified N. aromaticivorans Lig dehydrogenases, we found that LigL, LigN, and LigD each reduced the Cα ketone of G-diketone in vitro but with different substrate specificities and rates. Furthermore, LigL, but not LigN or LigD, also reduced the Cα ketone of 2-hydroxy-1-(4-hydroxy-3-methoxyphenyl)propan-1-one (GP-1) in vitro, a derivative of G-diketone with the Cβ ketone reduced, when GP-1 was provided as a substrate. The newly identified activity of these Lig dehydrogenases expands the potential range of substrates utilized by N. aromaticivorans beyond what has been previously recognized. This is beneficial both for metabolizing a wide range of natural and non-native depolymerized lignin substrates and for engineering microbes and enzymes that are active with a broader range of aromatic compounds. IMPORTANCE Lignin is a major plant polymer composed of aromatic units that have value as chemicals. However, the structure and composition of lignin have made it difficult to use this polymer as a renewable source of industrial chemicals. Bacteria like Novosphingobium aromaticivorans have the potential to make chemicals from lignin not only because of their natural ability to metabolize a variety of aromatics but also because there are established protocols to engineer N. aromaticivorans strains to funnel lignin-derived aromatics into valuable products. In this work, we report a newly discovered activity of previously characterized dehydrogenase enzymes with a chemically modified by-product of lignin depolymerization. We propose that the activity of N. aromaticivorans enzymes with both native lignin aromatics and those produced by chemical depolymerization will expand opportunities for producing industrial chemicals from the heterogenous components of this abundant plant polymer.

Developmental processes transduce diverse influences during phenotype formation, thereby biasing and structuring amount and type of phenotypic variation available for evolutionary processes to act on. The causes, extent, and consequences of this bias are subject to significant debate. Here we explore the role of developmental bias in contributing to organisms' ability to innovate, to adapt to novel or stressful conditions, and to generate well integrated, resilient phenotypes in the face of perturbations. We focus our inquiry on one taxon, the horned dung beetle genus Onthophagus, and review the role developmental bias might play across several levels of biological organization: (a) gene regulatory networks that pattern specific body regions; (b) plastic developmental mechanisms that coordinate body wide responses to changing environments and; (c) developmental symbioses and niche construction that enable organisms to build teams and to actively modify their own selective environments. We posit that across all these levels developmental bias shapes the way living systems innovate, adapt, and withstand stress, in ways that can alternately limit, bias, or facilitate developmental evolution. We conclude that the structuring contribution of developmental bias in evolution deserves further study to better understand why and how developmental evolution unfolds the way it does.

Background Blackbird (Icteridae) damage to ripening sunflower (Helianthus annuus L.) has been a persistent economic issue in the USA for the last five decades. To quantify losses, we surveyed blackbird damage from 2001 to 2013 (excluding 2004) to physiologically mature sunflower in eight states: North Dakota, South Dakota, Texas, Nebraska, Minnesota, Colorado, Kansas, and Vermont. Results We pooled data gathered during the most recent 5 years (2009 to 2013) of the survey and found losses averaged $US2.5 million and $US11.3 million for confectionery and oilseed hybrids, respectively. Three states, North Dakota, South Dakota, and Nebraska, had sufficient acreage and bird damage to warrant economic analyses using a regional economic model. The average annual total (direct plus indirect) economic impact of bird damage to sunflower production for North Dakota, South Dakota, and Nebraska was $US18.7 million, $US7.3 million, and $US2.6 million, respectively. Conclusion This study provides a better understanding of the broader economic implications of bird damage to sunflower by estimating the macroeconomic impacts of lost sunflower production in the region. Additionally, the findings of this study may be used to inform decisions regarding bird management policy by providing policymakers with information to determine the downstream value of bird management and the possible benefits of improved methods that further mitigate losses. © 2019 Society of Chemical Industry.

Typha is an iconic wetland plant found worldwide. Hybridization and anthropogenic disturbances have resulted in large increases in Typha abundance in wetland ecosystems throughout North America at a cost to native floral and faunal biodiversity. As demonstrated by three regional case studies, Typha is capable of rapidly colonizing habitats and forming monodominant vegetation stands due to traits such as robust size, rapid growth rate, and rhizomatic expansion. Increased nutrient inputs into wetlands and altered hydrologic regimes are among the principal anthropogenic drivers of Typha invasion. Typha is associated with a wide range of negative ecological impacts to wetland and agricultural systems, but also is linked with a variety of ecosystem services such as bioremediation and provisioning of biomass, as well as an assortment of traditional cultural uses. Numerous physical, chemical, and hydrologic control methods are used to manage invasive Typha, but results are inconsistent and multiple methods and repeated treatments often are required. While this review focuses on invasive Typha in North America, the literature cited comes from research on Typha and other invasive species from around the world. As such, many of the underlying concepts in this review are relevant to invasive species in other wetland ecosystems worldwide.

SummaryViruses influence the fate of nutrients and human health by killing microorganisms and altering metabolic processes. Organosulfur metabolism and biologically-derived hydrogen sulfide play dynamic roles in manifestation of diseases, infrastructure degradation, and essential biological processes. While microbial organosulfur metabolism is well-studied, the role of viruses in organosulfur metabolism is unknown. Here we report the discovery of 39 gene families involved in organosulfur metabolism encoded by 3,749 viruses from diverse ecosystems, including human microbiomes. The viruses infect organisms from all three domains of life. Six gene families encode for enzymes that degrade organosulfur compounds into sulfide, while others manipulate organosulfur compounds and may influence sulfide production. We show that viral metabolic genes encode key enzymatic domains, are translated into protein, are maintained after recombination, and that sulfide provides a fitness advantage to viruses. Our results reveal viruses as drivers of organosulfur metabolism with important implications for human and environmental health.

Here, we report the genome-wide identification of transcription start sites (TSSs) from two Alphaproteobacteria grown under conditions that result in significant changes in gene expression. TSSs that were identified as present in one condition or both will be an important resource for future studies of these, and possibly other, Alphaproteobacteria.

Descent with modification is the foundational framework of all of evolution. Yet evolutionary novelties are defined as lacking affinities to structures that already existed in the ancestral state, i.e. to somehow emerge in the absence of homology. We posit that reconciling both perspectives necessitates the existence of a type of innovation gradient that allows descent with modification to seed the initiation of a novel trait, which once in existence can then diversify into its variant forms. Recent work on diverse, textbook examples of morphological novelties illustrate the value of the innovation gradient concept. Innovations as profound and diverse as insect wings, beetle horns, and treehopper helmets derive from homologous source tissues instructed in their development by homologous gene regulatory networks. Yet rather than rendering these traits no longer novel, we posit that discoveries such as these call for a reassessment of the usefulness of defining evolutionary novelty as necessitating the absence of homology. Instead, we need to redirect our attention to how ancestral homologies scaffold and bias the innovation gradient to facilitate hotspots of innovation in some places, and deep conservation elsewhere.

Aquatic N-fixation is generally associated with the growth and mass development of Cyanobacteria in nitrogen-deprived photic zones. However, sequenced genomes and environmental surveys suggest active aquatic N-fixation also by many non-cyanobacterial groups. Here, we revealed the seasonal variation and genomic diversity of potential N-fixers in a humic bog lake using metagenomic data and nif gene clusters analysis. Groups with diazotrophic operons were functionally divergent and included Cholorobi, Geobacter, Desulfobacterales, Methylococcales, and Acidobacteria. In addition to nifH (a gene that encodes the dinitrogenase reductase component of the molybdenum nitrogenase), we also identified sequences corresponding to vanadium and iron-only nitrogenase genes. Within the Chlorobi population, the nitrogenase (nifH) cluster was included in a well-structured retrotransposon. Furthermore, the presence of light-harvesting photosynthesis genes implies that anoxygenic photosynthesis may fuel nitrogen fixation under the prevailing low-irradiance conditions. The presence of rnf genes (related to the expression of H+/Na+-translocating ferredoxin: NAD+ oxidoreductase) in Methylococcales and Desulfobacterales suggests that other energy-generating processes may drive the costly N-fixation in the absence of photosynthesis. The highly reducing environment of the anoxic bottom layer of Trout Bog Lake may thus also provide a suitable niche for active N-fixers and primary producers. While future studies on the activity of these potential N-fixers are needed to clarify their role in freshwater nitrogen cycling, the metagenomic data presented here enabled an initial characterization of previously overlooked diazotrophs in freshwater biomes.

Background How novel traits integrate within ancient trait complexes without compromising ancestral functions is a foundational challenge in evo-devo. The insect head represents an ancient body region patterned by a deeply conserved developmental genetic network, yet at the same time constitutes a hot spot for morphological innovation. However, the mechanisms that facilitate the repeated emergence, integration, and diversification of morphological novelties within this body region are virtually unknown. Using horned Onthophagus beetles, we investigated the mechanisms that instruct the development of the dorsal adult head and the formation and integration of head horns, one of the most elaborate classes of secondary sexual weapons in the animal kingdom. Results Using region-specific RNAseq and gene knockdowns, we (i) show that the head is compartmentalized along multiple axes, (ii) identify striking parallels between morphological and transcriptional complexity across regions, yet (iii) fail to identify a horn-forming gene module. Instead, (iv) our results support that sex-biased regulation of a shared transcriptional repertoire underpins the formation of horned and hornless heads. Furthermore, (v) we show that embryonic head patterning genes frequently maintain expression within the dorsal head well into late post-embryonic development, thereby possibly facilitating the repurposing of such genes within novel developmental contexts. Lastly, (vi) we identify novel functions for several genes including three embryonic head patterning genes in the integration of both posterior and anterior head horns. Conclusions Our results illuminate how the adult insect head is patterned and suggest mechanisms capable of integrating novel traits within ancient trait complexes in a sex- and species-specific manner. More generally, our work underscores how significant morphological innovation in developmental evolution need not require the recruitment of new genes, pathways, or gene networks but instead may be scaffolded by pre-existing developmental machinery.

Context-dependent trait exaggeration is a major contributor to phenotypic diversity. However, the genetic modifiers instructing development across multiple contexts remain largely unknown. We use the arthropod tibia, a hotspot for segmental differentiation, as a paradigm to assess the developmental mechanisms underlying the context-dependent structural exaggeration of size and shape through nutritional plasticity, sexual dimorphism and segmental differentiation. Using an RNAseq approach in the sexually dimorphic and male-polyphenic dung beetleDigitonthophagus gazella, we find that only a small portion (3.7%) of all transcripts covary positively in expression level with trait size across contexts. However, RNAi-mediated knockdown of the conserved sex-determination genedoublesexsuggests that it functions as a context-dependent master mediator of trait exaggeration inD. gazellaas well as the closely related dung beetleOnthophagus taurus. Taken together, our findings suggest (i) that the gene networks associated with trait exaggeration are highly dependent on the precise developmental context, (ii) thatdoublesexdifferentially shapes morphological exaggeration depending on developmental contexts and (iii) that this context-specificity ofdsx-mediated trait exaggeration may diversify rapidly. This mechanism may contribute to the resolution of conflict arising from environment-dependent antagonistic selection among sexes and divergent developmental contexts in a wide range of animals.

Frugivorous birds impose significant costs on tree fruit growers through direct consumption of fruit and grower efforts to manage birds. We documented factors that influenced tree fruit bird damage from 2012 through 2014 with a coordinated field study in Michigan, New York, and Washington. For sweet cherries, percent bird damage was higher in 2012 compared to 2013 and 2014, in Michigan and New York compared to Washington, and in blocks with more edges adjacent to non-sweet cherry land-cover types. These patterns appeared to be associated with fruit abundance patterns; 2012 was a particularly low-yield year for tree fruits in Michigan and New York and percent bird damage was high. In addition, percent bird damage to sweet and tart cherries in Michigan was higher in landscapes with low to moderate forest cover compared to higher forest cover landscapes. 'Honeycrisp' apple blocks under utility wires were marginally more likely to have greater bird damage compared to blocks without wires. We recommend growers prepare bird management plans that consider the spatial distribution of fruit and non-fruit areas of the farm. Growers should generally expect to invest more in bird management in low-yield years, in blocks isolated from other blocks of the same crop, and in blocks where trees can provide entry to the crop for frugivorous birds.

Some fruit-eating bird species commonly consume cultivated fruit. Species-specific variation in diet preferences could result in varying use of orchards and impacts on the fruit-producing industry. However, species-specific studies of avian orchard use are lacking, particularly throughout the fruit-growing season. Our objectives were to quantify the frequency of daily bird visits to orchards and the amount of time birds spent visiting orchards each day over the fruit-ripening season. Birds are well-documented consumers of cultivated sweet cherries (Prunus avium), which are relatively high in sugar and low in proteins and lipids. American Robins (Turdus migratorius) and Cedar Waxwings (Bombycilla cedrorum) are common fruit-consumers in sweet cherry orchards. Robins often consume larger proportions of invertebrates and prefer lipid-rich fruits, while waxwings choose sugary fruits. Given these species-specific diet differences, we hypothesized waxwings would spend a greater proportion of days and mo...

Microdiversity can lead to different ecotypes within the same species. These are assumed to provide stability in time and space to those species. However, the role of microdiversity in the stability of whole microbial communities remains underexplored. Understanding the drivers of microbial community stability is necessary to predict community response to future disturbances. Here, we analyzed 16S rRNA gene amplicons from eight different temperate bog lakes at the 97% OTU and amplicon sequence variant (ASV) levels and found ecotypes within the same OTU with different distribution patterns in space and time. We observed that these ecotypes are adapted to different values of environmental factors such as water temperature and oxygen concentration. Our results showed that the existence of several ASVs within a OTU favored its persistence across changing environmental conditions. We propose that microdiversity aids the stability of microbial communities in the face of fluctuations in environmental factors., This work was funded by grant CTM2016-80095-C2-1-R from the Spanish Ministerio de Economía, Industria y Competitividad. NG-G was funded by a student scholarship from the Severo Ochoa Program at CNB (SEV-2013-0347-17-2).

Intra-species diversity comprises different ecotypes within the same species. These are assumed to provide stability in time and space to those species. However, the role that microdiversity plays in the stability of whole microbial communities remains underexplored. Understanding the drivers of microbial community stability is necessary to predict community response to future disturbances. Here, we analyzed 16S rRNA gene amplicons from eight different temperate bog lakes at OTU-97% and amplicon sequence variant (ASV) levels, and we found ecotypes within the same species with different distribution patterns in space and time. We observed that these ecotypes are adapted to different values of environmental factors such as water temperature and oxygen concentration. Our results showed that the existence of several ASVs within a species favored its persistence across changing environmental conditions. We propose that microdiversity aids the stability of microbial communities in the face of fluctuations in environmental factors.

Where do horns come from? One of the most pronounced examples of a sexually selected trait is the prothoracic horns of scarab beetles, which, in the most extreme cases, can be nearly half as long as the length of the beetle. It is fairly easy to understand how selection might have shaped these horns, but understanding how development shaped them from a hornless ancestor is a much more complex proposition. Hu et al. show that these horns are generated from wing homologs and argue that many other insect traits may have followed similar transcriptional paths (see the Perspective by Nijhout). Science , this issue p. 1004 ; see also p. 946

Humic lakes and ponds receive large amounts of terrestrial carbon and are important components of the global carbon cycle, yet how their redox cycling influences the carbon budget is not fully understood. Here we compared metagenomes obtained from a humic bog and a clear-water eutrophic lake and found a much larger number of genes that might be involved in extracellular electron transfer (EET) for iron redox reactions and humic substance (HS) reduction in the bog than in the clear-water lake, consistent with the much higher iron and HS levels in the bog., Humic lakes and ponds receive large amounts of terrestrial carbon and are important components of the global carbon cycle, yet how their redox cycling influences the carbon budget is not fully understood. Here we compared metagenomes obtained from a humic bog and a clear-water eutrophic lake and found a much larger number of genes that might be involved in extracellular electron transfer (EET) for iron redox reactions and humic substance (HS) reduction in the bog than in the clear-water lake, consistent with the much higher iron and HS levels in the bog. These genes were particularly rich in the bog’s anoxic hypolimnion and were found in diverse bacterial lineages, some of which are relatives of known iron oxidizers or iron-HS reducers. We hypothesize that HS may be a previously overlooked electron acceptor and that EET-enabled redox cycling may be important in pelagic respiration and greenhouse gas budget in humic-rich freshwater lakes.

Metabolic processes at the microbial scale influence ecosystem functions because microbes are responsible for much of the carbon and nutrient cycling in freshwater. One approach to predict the metabolic capabilities of microbial communities is to search for functional marker genes in metagenomes. However, this approach does not provide context about co-occurrence with other metabolic traits within an organism or detailed taxonomy about those organisms. Here, we combine a functional marker gene analysis with metabolic pathway prediction of microbial population genomes (MAGs) assembled from metagenomic time series in eutrophic Lake Mendota and humic Trout Bog to identify how carbon and nutrient cycles are connected in freshwater. We found that phototrophy, carbon fixation, and nitrogen fixation pathways co-occurred in Cyanobacteria MAGs in Lake Mendota and in Chlorobiales MAGs in Trout Bog. Cyanobacteria MAGs also had strong temporal correlations to functional marker genes for nitrogen fixation in several years. Genes encoding steps in the nitrogen and sulfur cycles varied in abundance and taxonomy by lake, potentially reflecting the availability and composition of inorganic nutrients in these systems. We were also able to identify which populations contained the greatest density and diversity of genes encoding glycoside hydrolases. Populations with many glycoside hydrolases also encoded pathways for sugar degradation. By using both MAGs and marker genes, we were better able to link functions to specific taxonomic groups in our metagenomic time series, enabling a more detailed understanding of freshwater microbial carbon and nutrient cycling.

Yellow-headed Blackbirds (Xanthocephalus xanthocephalus) in central North Dakota undergo prebasic molt or prejuvenile molt during late summer. Nestling Yellow-headed Blackbirds initiate a complete prejuvenile molt, grow their primary and secondary regimes in about 40 days, completing molt after they leave the nest by the first week in August. Remiges are not replaced during the subsequent preformative molt, being retained until the second prebasic molt. Nonlinear (logistic) regression of primary remex growth during definitive prebasic molts of Yellow-headed Blackbirds indicated 38 days were required to complete the linear phase of growth (between 10% and 90% of total primary length). Males added 19.5 mm/d and females added 15.7 mm/d to the total length of all primaries during this linear growth phase; an average of 4–5 mm per primary remex per day. Definitive prebasic molting of primary remiges in males and females was initiated in late June, after nesting and brood rearing were completed. Molts ...

Bird-livestock interactions have been implicated as potential sources for bacteria within concentrated animal feeding operations (CAFO). European starlings (Sturnus vulgaris) in particular are known to contaminate cattle feed and water with Salmonella enterica through their fecal waste. We propose that fecal waste is not the only mechanisms through which starlings introduce S. enterica to CAFO. The goal of this study was to assess if starlings can mechanically move S. enterica. We define mechanical movement as the transportation of media containing S. enterica, on the exterior of starlings within CAFO. We collected 100 starlings and obtained external wash and gastrointestinal tract (GI) samples. We also collected 100 samples from animal pens. Within each pen we collected one cattle fecal, feed, and water trough sample. Isolates from all S. enterica positive samples were subjected to antimicrobial susceptibility testing. All sample types, including 17% of external starling wash samples, contained S. enterica. All sample types had at least one antimicrobial resistant (AMR) isolate and starling GI samples harbored multidrug resistant S. enterica. The serotypes isolated from the starling external wash samples were all found in the farm environment and 11.8% (2/17) of isolates from positive starling external wash samples were resistant to at least one class of antibiotics. This study provides evidence of a potential mechanism of wildlife introduced microbial contamination in CAFO. Mechanical movement of microbiological hazards, by starlings, should be considered a potential source of bacteria that is of concern to veterinary, environmental and public health.

Knowledge of factors influencing animal abundance is important to wildlife biologists developing management plans. This is especially true for economically important species such as blackbirds (Icteridae), which cause more than $100 million in crop damages annually in the United States. Using data from the North American Breeding Bird Survey, the National Land Cover Dataset, and the National Climatic Data Center, we modeled effects of regional environmental variables on relative abundance of 3 blackbird species (red-winged blackbird, Agelaius phoeniceus; yellow-headed blackbird, Xanthocephalus xanthocephalus; common grackle, Quiscalus quiscula) in the Prairie Pothole Region of the central United States. We evaluated landscape covariates at 3 logarithmically related spatial scales (1,000ha, 10,000ha, and 100,000ha) and modeled weather variables at the 100,000-ha scale. We constructed models a priori using information from published habitat associations. We fit models with WinBUGS using Markov chain Monte Carlo techniques. Both landscape and weather variables contributed strongly to predicting blackbird relative abundance (95% credibility interval did not overlap 0). Variables with the strongest associations with blackbird relative abundance were the percentage of wetland area and precipitation amount from the year before bird surveys were conducted. The influence of spatial scale appeared small—models with the same variables expressed at different scales were often in the best model subset. This large-scale study elucidated regional effects of weather and landscape variables, suggesting that management strategies aimed at reducing damages caused by these species should consider the broader landscape, including weather effects, because such factors may outweigh the influence of localized conditions or site-specific management actions. The regional species distributional models we developed for blackbirds provide a tool for understanding these broader landscape effects and guiding wildlife management practices to areas that are optimally beneficial. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

The origin of insect wings is still a highly debated mystery in biology, despite the importance of this evolutionary innovation. There are currently two prominent, but contrasting wing origin hypotheses (the tergal origin hypothesis and the pleural origin hypothesis). Through studies in the Tribolium beetle, we have previously obtained functional evidence supporting a third hypothesis, the dual origin hypothesis. Although this hypothesis can potentially unify the two competing hypotheses, it requires further testing from various fields. Here, we investigated the genetic regulation of the tissues serially homologous to wings in the abdomen, outside of the appendage-bearing segments, in Tribolium. We found that the formation of ectopic wings in the abdomen upon homeotic transformation relies not only on the previously identified abdominal wing serial homolog (gin-trap), but also on a secondary tissue in the pleural location. Using an enhancer trap line of nubbin (a wing lineage marker), we were able to visualize both of these two tissues (of tergal and pleural nature) contributing to form a complete wing. These results support the idea that the presence of two distinct sets of wing serial homologs per segment represents an ancestral state of the wing serial homologs, and can therefore further support a dual evolutionary origin of insect wings. Our analyses also uncovered detailed Hox regulation of abdominal wing serial homologs, which can be used as a foundation to elucidate the molecular mechanisms that have facilitated the evolution of bona fide insect wings, as well as the diversification of other wing serial homologs.

Although microbes mediate much of the biogeochemical cycling in freshwater, the categories of carbon and nutrients currently used in models of freshwater biogeochemical cycling are too broad to be relevant on a microbial scale. One way to improve these models is to incorporate microbial data. Here, we analyze both genes and genomes from three metagenomic time series and propose specific roles for microbial taxa in freshwater biogeochemical cycles. Our metagenomic time series span multiple years and originate from a eutrophic lake (Lake Mendota) and a humic lake (Trout Bog Lake) with contrasting water chemistry. Our analysis highlights the role of polyamines in the nitrogen cycle, the diversity of diazotrophs between lake types, the balance of assimilatory vs. dissimilatory sulfate reduction in freshwater, the various associations between types of phototrophy and carbon fixation, and the density and diversity of glycoside hydrolases in freshwater microbes. We also investigated aspects of central metabolism such as hydrogen metabolism, oxidative phosphorylation, methylotrophy, and sugar degradation. Finally, by analyzing the dynamics over time in nitrogen fixation genes and Cyanobacteria genomes, we show that the potential for nitrogen fixation is linked to specific populations in Lake Mendota. This work represents an important step towards incorporating microbial data into ecosystem models and provides a better understanding of how microbes may participate in freshwater biogeochemical cycling.

European starlings (Sturnus vulgaris, Figure 1)are an invasive species in the United States. The first recorded release of the birds was in 1890 in New York City’s Central Park. Because starlings easily adapt to a variety of habitats, nest sites and food sources, the birds spread quickly across the country. Today, there are about 150 million starlings in North America. Conflicts between people and starlings occur mostly in agricultural settings. Starlings damage apples, blueberries, cherries, figs, grapes, peaches, and strawberries. Starlings gather at concentrated animal feeding operations (CAFOs) during late fall and winter. Starlings also cause human health problem, airplane hazards, and nuisance problems. European starlings are not protected by the Migratory Bird Treaty Act (MBTA).

Bacteria play a key role in freshwater biogeochemical cycling, but long-term trends in freshwater bacterial community composition and dynamics are not yet well characterized. We used a multiyear time series of 16S rRNA gene amplicon sequencing data from eight bog lakes to census the freshwater bacterial community and observe annual and seasonal trends in abundance. The sites that we studied encompassed a range of water column mixing frequencies, which we hypothesized would be associated with trends in alpha and beta diversity. Each lake and layer contained a distinct bacterial community, with distinct levels of richness and indicator taxa that likely reflected the environmental conditions of each lake type sampled, including

Lakes are excellent systems for investigating bacterial community dynamics because they have clear boundaries and strong environmental gradients. The results of our research demonstrate that bacterial community composition varies by year, a finding which likely applies to other ecosystems and has implications for study design and interpretation. Understanding the drivers and controls of bacterial communities on long time scales would improve both our knowledge of fundamental properties of bacterial communities and our ability to predict community states. In this specific ecosystem, bog lakes play a disproportionately large role in global carbon cycling, and the information presented here may ultimately help refine carbon budgets for these lakes. Finally, all data and code in this study are publicly available. We hope that this will serve as a resource for anyone seeking to answer their own microbial ecology questions using a multiyear time series., Bacteria play a key role in freshwater biogeochemical cycling, but long-term trends in freshwater bacterial community composition and dynamics are not yet well characterized. We used a multiyear time series of 16S rRNA gene amplicon sequencing data from eight bog lakes to census the freshwater bacterial community and observe annual and seasonal trends in abundance. The sites that we studied encompassed a range of water column mixing frequencies, which we hypothesized would be associated with trends in alpha and beta diversity. Each lake and layer contained a distinct bacterial community, with distinct levels of richness and indicator taxa that likely reflected the environmental conditions of each lake type sampled, including Actinobacteria in polymictic lakes (i.e., lakes with multiple mixing events per year), Methylophilales in dimictic lakes (lakes with two mixing events per year, usually in spring and fall), and “Candidatus Omnitrophica” in meromictic lakes (lakes with no recorded mixing events). The community present during each year at each site was also surprisingly unique. Despite unexpected interannual variability in community composition, we detected a core community of taxa found in all lakes and layers, including Actinobacteria tribe acI-B2 and Betaprotobacteria lineage PnecC. Although trends in abundance did not repeat annually, each freshwater lineage within the communities had a consistent lifestyle, defined by persistence, abundance, and variability. The results of our analysis emphasize the importance of long-term multisite observations, as analyzing only a single year of data or one lake would not have allowed us to describe the dynamics and composition of these freshwater bacterial communities to the extent presented here. IMPORTANCE Lakes are excellent systems for investigating bacterial community dynamics because they have clear boundaries and strong environmental gradients. The results of our research demonstrate that bacterial community composition varies by year, a finding which likely applies to other ecosystems and has implications for study design and interpretation. Understanding the drivers and controls of bacterial communities on long time scales would improve both our knowledge of fundamental properties of bacterial communities and our ability to predict community states. In this specific ecosystem, bog lakes play a disproportionately large role in global carbon cycling, and the information presented here may ultimately help refine carbon budgets for these lakes. Finally, all data and code in this study are publicly available. We hope that this will serve as a resource for anyone seeking to answer their own microbial ecology questions using a multiyear time series.

The varied diet of European starlings (Sturnus vulgaris L.) can present challenges when working with starlings in experimental testing and holding situations and should be taken into account when testing repellents applied to food. Our purpose was to evaluate an anthraquinone-based repellent (Arkion Life Sciences, New Castle, DE, USA; active ingredient 50% 9,10-anthraquinone; hereafter anthraquinone) and SucraShield™ (Natural Forces, Davidson, NC, USA; active ingredient 40% sucrose octanoate esters) repellent for non-lethal protection of specialty crops (i.e., fruit, sweet corn) and grains. Our objectives were to evaluate (1) laboratory efficacy of anthraquinone applied topically to blueberries and sweet corn, (2) laboratory efficacy of anthraquinone applied to two pellet matrices, and (3) laboratory efficacy of SucraShield™ as a chemical repellent for European starlings. We found that anthraquinone was not an effective repellent for blueberries or sweet corn, although consumption of each matrix varied potentially due to sucrose content. Anthraquinone was an effective repellent on CU Bird Carrier pellets with 6275 ppm needed to achieve 80% repellency, whereas up to 35,000 ppm anthraquinone was not effective when the anthraquinone was not topically applied. SucraShield was not an effective repellent for starlings and in fact increased consumption of CU Bird Carrier as concentration increased.

Non-lethal alternatives are needed to manage the damage caused by wild birds to oilseed sunflower crops (Helianthus annuus Linnaeus). We evaluated field residues and experimental applications of an anthraquinone-based repellent (active ingredient 50% 9,10-anthraquinone) to minimize red-winged blackbird (Agelaius phoeniceus Linnaeus) depredation of oilseed sunflower. Chemical residues from experimental applications of the anthraquinone-based repellent (4.7 l/ha and 9.4 l/ha; low, high) in a ripening oilseed sunflower field were 481 ppm and 978 ppm anthraquinone at the beginning of blackbird damage, and 385 ppm and 952 ppm anthraquinone at the end of blackbird damage, respectively. Prior to harvest, we observed 402 ppm and 462 ppm anthraquinone in the oil, and 27 ppm and 165 ppm anthraquinone in the pomace from crushed sunflower achenes previously sprayed with the low and high applications, respectively. For the purpose of developing application strategies useful for avian repellents, we subsequently investigated blackbird feeding response to oilseed sunflower treated with the anthraquinone-based repellent and either a registered insecticide or a registered fungicide popularly used for ripening sunflower. We observed a positive concentration–response relationship among blackbirds exposed to anthraquinone and the insecticide (a.i. 8.4% esfenvalerate), or anthraquinone and the fungicide (a.i. 23.6% pyraclostrobin). Blackbirds reliably discriminated between untreated sunflower and that treated with 1810 ppm anthraquinone and 0.1% of the insecticide or 1700 ppm anthraquinone and 0.14% of the fungicide during our preference experiments. Given that ripening achenes are inverted from conventional pesticide applications throughout much of the period associated with blackbird depredation, we also evaluated blackbird repellency of the anthraquinone-based repellent applied to involucral bracts (i.e., the back of sunflower heads) of oilseed sunflower. Blackbirds did not discriminate between untreated involucral bracts and those treated with foliar applications comparable to 4.7 l/ha or 9.4 l/ha; blackbirds consumed more achenes from untreated sunflower heads than from those treated with 18.7 l/ha of the anthraquinone-based repellent. Supplemental repellent efficacy studies should investigate blackbird response to anthraquinone-based repellents (e.g., ≥4.7 l/ha) within 10–100 ha sunflower fields and include independent field replicates with predicted bird damage, repellent application strategies developed for protection of ripening crops, pre- and at-harvest repellent residues, and bird damage and crop yield measurements.