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At least 109 records · Page 6

Lunar outpost agriculture

The development of a CELSS for a lunar outpost is discussed. It is estimated that a lunar outpost life support system with a crew of four that produces food would break even in terms of mass and cost to deliver the system to the lunar surface after 2.5 years when compared to the cost of resupply from earth. A brief review is made of research on life support systems and NASA projects for evaluating CELSS components. The use of on-site materials for propellants, construction materials, and agriculture is evaluated, and the use of microbes for waste decomposition and stabilization of ecological balance is touched upon. Areas for further investigation include the behavior of organisms in microgravity, genetic alteration, gas exchange capabilities of organisms, integration of biological and physicochemical components, and automation. The development stages leading to lunar deployment are outlined.

Hossner, Lloyd R.↗

CRISPR/Cas9-based gene activation and base editing in Populus

The genus Populus has long been used for environmental, agroforestry and industrial applications worldwide. Today Populus is also recognized as a desirable crop for biofuel production and a model tree for physiological and ecological research. As such, various modern biotechnologies, including CRISPR/Cas9-based techniques, have been actively applied to Populus for genetic and genomic improvements for traits such as increased growth rate and tailored lignin composition. However, CRISPR/Cas9 has been primarily used as the active Cas9 form to create knockouts in the hybrid poplar clone “717-1B4” (P. tremula x P. alba clone INRA 717-1B4). Alternative CRISPR/Cas9-based technologies, e.g. those involving modified Cas9 for gene activation and base editing, have not been evaluated in most Populus species for their efficacy. Here we employed a deactivated Cas9 (dCas9)-based CRISPR activation (CRISPRa) technique to fine-tune the expression of two target genes, TPX2 and LecRLK-G which play important roles in plant growth and defense response, in hybrid poplar clone “717-1B4” and poplar clone “WV94” (P. deltoides “WV94”), respectively. We observed that CRISPRa resulted in 1.2-fold to 7.0-fold increase in target gene expression through transient expression in protoplasts and Agrobacterium-mediated stable transformation, demonstrating the effectiveness of dCas9-based CRISPRa system in Populus. In addition, we applied Cas9 nickase (nCas9)-based cytosine base editor (CBE) to precisely introduce premature stop codons via C-to-T conversion, with an efficiency of 13%–14%, in the target gene PLATZ which encodes a transcription factor involved in plant fungal pathogen response in hybrid poplar clone “717-1B4”. Overall, we showcase the successful application of CRISPR/Cas-based technologies in gene expression regulation and precise gene engineering in two Populus species, facilitating the adoption of emerging genome editing tools in woody species.

59 BASIC BIOLOGICAL SCIENCES↗

The first two chromosome‐scale genome assemblies of American hazelnut enable comparative genomic analysis of the genus Corylus

Summary The native, perennial shrub American hazelnut ( Corylus americana ) is cultivated in the Midwestern United States for its significant ecological benefits, as well as its high‐value nut crop. Implementation of modern breeding methods and quantitative genetic analyses of C. americana requires high‐quality reference genomes, a resource that is currently lacking. We therefore developed the first chromosome‐scale assemblies for this species using the accessions ‘Rush’ and ‘Winkler’. Genomes were assembled using HiFi PacBio reads and Arima Hi‐C data, and Oxford Nanopore reads and a high‐density genetic map were used to perform error correction. N50 scores are 31.9 Mb and 35.3 Mb, with 90.2% and 97.1% of the total genome assembled into the 11 pseudomolecules, for ‘Rush’ and ‘Winkler’, respectively. Gene prediction was performed using custom RNAseq libraries and protein homology data. ‘Rush’ has a BUSCO score of 99.0 for its assembly and 99.0 for its annotation, while ‘Winkler’ had corresponding scores of 96.9 and 96.5, indicating high‐quality assemblies. These two independent assemblies enable unbiased assessment of structural variation within C. americana , as well as patterns of syntenic relationships across the Corylus genus. Furthermore, we identified high‐density SNP marker sets from genotyping‐by‐sequencing data using 1343 C. americana , C. avellana and C. americana × C. avellana hybrids, in order to assess population structure in natural and breeding populations. Finally, the transcriptomes of these assemblies, as well as several other recently published Corylus genomes, were utilized to perform phylogenetic analysis of sporophytic self‐incompatibility (SSI) in hazelnut, providing evidence of unique molecular pathways governing self‐incompatibility in Corylus .

54 ENVIRONMENTAL SCIENCES↗

ENIGMA: Ecosystems and Networks Integrated with Genes and Molecular Assemblies. Final Report

ENIGMA research is performed at a very large scale involving collaborations between multiple research groups to address two major driving concepts: For any microbe, it is possible to discover efficiently the genetic determinants of adaptation to life in dynamic environments in contact with other living members of its ecology and reveal hidden functional phenomena that are only possible by using a systems biology approach; and for any environment, it is possible to predict the temporal changes in geochemistry and accompanying system behavior, for some length of time, given biogeochemical and physical inputs. These driving concepts also map directly to grand challenges the ENIGMA team has outlined: “Define the levels of biological organization most relevant to scaling from single cells to ecosystems and global cycles; capture how that organization varies in time and space; and identify critical interactions that dictate rates of carbon, nutrient, and energy transformation in different environments.”

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Viromics approaches for the study of viral diversity and ecology in microbiomes

Viruses are found across all ecosystems and infect every type of organism on Earth. Traditional culture-based methods have proven insufficient to explore this viral diversity at scale, driving the development of viromics, the sequence-based analysis of uncultivated viruses. Viromics approaches have been particularly useful for studying viruses of microorganisms, which can act as crucial regulators of microbiomes across ecosystems. They have already revealed the broad geographic distribution of viral communities and are progressively uncovering the expansive genetic and functional diversity of the global virome. Moving forward, large-scale viral ecogenomics studies combined with new experimental and computational approaches to identify virus activity and host interactions will enable a more complete characterization of global viral diversity and its effects.

Ecology↗

Cross-family and phage-specific gene requirements for Klebsiella infection revealed by scalable RB-TnSeq genetic screens.

Bacteriophages are being cataloged at an accelerating pace and are recognized as key players in nutrient and energy cycling across ecosystems. Yet the bacterial genetic determinants that govern phage-host specificity and infection success remain poorly understood, particularly in clinically and ecologically important genera such as Klebsiella where prior receptor characterization has been almost entirely limited to capsulated strains. Here we used a randomly barcoded, genome-wide, loss-of-function transposon mutant library (RB-TnSeq) of Klebsiella sp. M5al, a naturally acapsular, nitrogen-fixing rhizobacterium, to generate the first systematic, cross-family map of phage receptor gene dependencies in Klebsiella. Challenging the library against 25 double-stranded DNA phages spanning five families in 213 parallel assays, we identified 42 bacterial genes associated with phage infection, of which 15 had no prior association with phage infection in any bacterial system. Disruption of surface receptor biosynthesis genes conferred cross-resistance across multiple phage families, while intracellular gene disruptions had predominantly phage-specific effects. Clonal validation of eight genes confirmed LPS outer core biosynthesis genes as primary receptor determinants alongside additional host factors spanning outer membrane transport, cofactor biosynthesis, and two-component signaling. Comparative analysis across all 25 phages revealed that phage genus rather than family is the stronger predictor of host gene dependency profiles, a finding with direct implications for the functional annotation of uncharacterized phage isolates and rational phage cocktail design. Together, these findings provide a community resource for linking phage genomic diversity to functional host interaction space in this ecologically and clinically important genus.

Gittrich, Marissa R↗

A natural view of microbial biodiversity within hot spring cyanobacterial mat communities

This review summarizes a decade of research in which we have used molecular methods, in conjunction with more traditional approaches, to study hot spring cyanobacterial mats as models for understanding principles of microbial community ecology. Molecular methods reveal that the composition of these communities is grossly oversimplified by microscopic and cultivation methods. For example, none of 31 unique 16S rRNA sequences detected in the Octopus Spring mat, Yellowstone National Park, matches that of any prokaryote previously cultivated from geothermal systems; 11 are contributed by genetically diverse cyanobacteria, even though a single cyanobacterial species was suspected based on morphologic and culture analysis. By studying the basis for the incongruity between culture and molecular samplings of community composition, we are beginning to cultivate isolates whose 16S rRNA sequences are readily detected. By placing the genetic diversity detected in context with the well-defined natural environmental gradients typical of hot spring mat systems, the relationship between gene and species diversity is clarified and ecological patterns of species occurrence emerge. By combining these ecological patterns with the evolutionary patterns inherently revealed by phylogenetic analysis of gene sequence data, we find that it may be possible to understand microbial biodiversity within these systems by using principles similar to those developed by evolutionary ecologists to understand biodiversity of larger species. We hope that such an approach guides microbial ecologists to a more realistic and predictive understanding of microbial species occurrence and responsiveness in both natural and disturbed habitats.

Review↗

Editorial overview: Unlocking the secrets of nongenetic plasticity, one cell at a time

Cellular noise, the non-genetic variability observed among isogenic cells, arises from factors such as growth conditions, aging, and stochastic gene expression, influencing cell stress-response, metabolism, morphology, and size. Here, such plasticity, while critical for adaptation, often goes unnoticed with traditional population-averaging biotechnologies that inevitably mask cell-specific variations and prompting the question, "What else might we be missing". However, recent breakthroughs in optical imaging, microfluidics, and omics, are beginning to uncover the complexity of cellular plasticity. This special is-sue highlights some of these breakthroughs, with key contributions including innovations in multimodal chemical imaging, label-free microscopy, spatial and temporal omics, and droplet-based microfluidics. Collectively, these cutting-edge tools provide unprecedent-ed insights into non-genetic cell-to-cell variability, enhancing our understanding of cellu-lar plasticity and its implications for health, energy, and ecology.

59 BASIC BIOLOGICAL SCIENCES↗

Growth instabilities shape morphology and genetic diversity of microbial colonies

Abstract Cellular populations assume an incredible variety of shapes ranging from circular molds to irregular tumors. While we understand many of the mechanisms responsible for these spatial patterns, little is known about how the shape of a population influences its ecology and evolution. Here, we investigate this relationship in the context of microbial colonies grown on hard agar plates. This a well-studied system that exhibits a transition from smooth circular disks to more irregular and rugged shapes as either the nutrient concentration or cellular motility is decreased. Starting from a mechanistic model of colony growth, we identify two dimensionless quantities that determine how morphology and genetic diversity of the population depend on the model parameters. Our simulations further reveal that population dynamics cannot be accurately described by the commonly-used surface growth models. Instead, one has to explicitly account for the emergent growth instabilities and demographic fluctuations. Overall, our work links together environmental conditions, colony morphology, and evolution. This link is essential for a rational design of concrete, biophysical perturbations to steer evolution in the desired direction.

Golden, Alexander↗

Metabolic patterns in Septoria canker resistant and susceptible Populus trichocarpa genotypes 24 hours post-inoculation

Sphaerulina musiva is an economically and ecologically important fungal pathogen that causes Septoria stem canker and leaf spot disease of Populus species. Here, to bridge the gap between genetic markers and structural barriers previously found to be linked to Septoria (Sphaerulina musiva) canker disease resistance in poplar, we employed hydrophilic interaction liquid chromatography (HILIC) and tandem mass spectrometry (MS/MS) to identify and quantify metabolites involved with signaling and cell wall re-modelling. Fluctuations in signaling molecules, organic acids, amino acids, sterols, phenolics, and saccharides in resistant and susceptible Populus trichocarpa inoculated with Sphaerulina musiva, were observed. The patterns of 222 metabolites in the resistant host implicates systemic acquired resistance (SAR), cell wall apposition, and lignin deposition as modes of resistance to this hemibiotrophic pathogen. This pattern is consistent with the expected response to the biotrophic phase of S. musiva colonization during the first 24 hours post-inoculation. The fungal pathogen metabolized key regulatory signals of SAR, other phenolics, and precursors of lignin biosynthesis which were depleted in the susceptible host. This is the first study to characterize metabolites associated with the response to initial colonization by S. musiva between resistant and susceptible hosts.

59 BASIC BIOLOGICAL SCIENCES↗

What are mycorrhizal traits?

Traits are inherent properties of organisms, but how are they defined for organismal networks such as mycorrhizal symbioses? Mycorrhizal symbioses are complex and diverse belowground symbioses between plants and fungi that have proved challenging to fit into a unified and coherent trait framework. Here we propose an inclusive mycorrhizal trait framework that classifies traits as morphological, physiological, and phenological features that have functional implications for the symbiosis. We further classify mycorrhizal traits by location – plant, fungus, or the symbiosis – which highlights new questions in trait-based mycorrhizal ecology designed to charge and challenge the scientific community. This new framework is an opportunity for researchers to interrogate their data to identify novel insights and gaps in our understanding of mycorrhizal symbioses.

59 BASIC BIOLOGICAL SCIENCES↗

The silicon citizen naturalist

Smartphone-wielding citizen scientists and an AI called FLORIST are transforming ecology at the continental scale. Here, in this issue of Cell, when Tibbs-Cortes et al. pair the crowdsourced data with controlled genetics, they discover how switchgrass times its flowering to outwit both frost and heat, depending on latitude.

Hudson, Matthew E. [University of Illinois at Urba↗

Synthetic communities as a model for determining interactions between a biofertilizer chassis organism and native microbial consortia

Biofertilizers are critical for sustainable agriculture because they can replace ecologically disruptive chemical fertilizers while improving the trajectory of soil and plant health. However, for improving deployment, the persistence of biofertilizers within native soil consortia must be elucidated and enhanced. In this study we characterized a high-throughput, modular, and automation-friendly in vitro approach to screen for biofertilizer persistence within soil-derived consortia after co-cultivation with stable synthetic soil microbial communities (SynComs) obtained through a top-down cultivation process. Here, we profiled ~1200 SynComs isolated from various soil sources and cultivated in divergent media types, and we detected significant phylogenetic diversity (e.g. Shannon index >4) and richness (observed richness >400) across these communities. We observed high reproducibility in SynCom community structure from common soil and media types, which provided a testbed for assessing biofertilizer persistence within representative native consortia. Furthermore, we demonstrated that the screening method described herein can be coupled with microbial engineering to efficiently identify soil-derived SynComs in which an engineered biofertilizer organism (i.e. Bacillus subtilis) persists. Accordingly, we discovered that B. subtilis persisted in ~10% of SynComs that generally followed the diversity–invasion principle. Additionally, our approach enabled analysis of the ecological impact of B. subtilis inoculation on SynCom structure and profile alterations in community diversity and richness associated with the presence of a genetically modified model bacterium. Ultimately, this work has established a modular pipeline that could be integrated into a variety of microbiology/microbiome-relevant workflows or related applications that would benefit from assessment of the persistence of a specific organism of interest and its interaction with native consortia.

biofertilizers↗

Enabling Experimental Evolution: Multi-Parameter Sensor System Integration into a Culture/Stressor Biofluidics System

Experimental evolution (EE) exposes microbial communities to ecological stressors, simulating dynamics up to near-extinction events. Combined with comparative sequencing and other molecular tools, such data can inform the genetic and other biological mechanisms underlying extremophile adaptation, and other observed effects. Automating this type of experiment using biofluidics can mitigate many traditional obstacles, including delays in assay results and environment adjustment and the need for many replicates. A first-generation device for automating EE procedures, the Automated Adaptive Directed Evolution Chamber (AADEC), was developed at NASA Ames. UV-C radiation was the stressor, an LED-photodiode array measured optical density, magnetic agitation and peristaltic pump systems ensured nutrient availability, and Arduino microcontrollers provided control. Escherichia coli in LB kanamycin media was used for testing and performance verification. A manual laboratory procedure with timed exposure to UV-C was performed to typify tolerance acquisition. Approximately a 106 factor increase in survival ratio was recorded over multiple iterations. Currently, a second-generation device is being developed integrating more real-time sensors: redox potential (ORP), indicating available/consumed metabolic energy; dissolved oxygen (DO), indicating aerobic/anaerobic growth; pH, indicating metabolic products; and electrical conductivity (EC), another indicator of metabolic products. The EC sensor system was constructed and calibrated in-house and matched commercial sensors in the required range. A Raspberry Pi computer automated the electrical system, allowing real-time data acquisition. The fluidics card was made of CNC-milled polycarbonate for biocompatibility. Each sensor parameter can also be used as a selection pressure alone or in combination with others to create extreme microbial environments. As a proof of concept, this work demonstrated sensor operation in one pair of growth-sensor chambers. It can be expanded to a multi-chamber system to enable inter-culture comparisons and multi-population studies. The prior Arduino system will be ported to the RPi system. Future stressors to be added include thermal, reactive oxygen species, and varying nutrient availability.

Govinda Raj, Chinmayee↗

Genetic time traveling: sequencing old herbarium specimens, including the oldest herbarium specimen sequenced from kingdom Fungi, reveals the population structure of an agriculturally significant rust

Summary Sequencing herbarium specimens can be instrumental in answering ecological, evolutionary, and taxonomic inquiries. We developed a protocol for sequencing herbarium specimens of rust fungi (Pucciniales) and proceeded to sequence specimens ranging from 4 to 211 yr old from five different genera. We then obtained sequences from an economically important biological control agent, Puccinia suaveolens , to highlight the potential of sequencing herbarium specimens in an ecological sense and to evaluate the following hypotheses: (1) The population structure of a plant pathogen changes over time, and (2) introduced pathogens are more diverse in their native range. Our efforts resulted in sequences from 87 herbarium specimens that revealed a high level of diversity with a population structure that exhibited spatial–temporal patterns. The specimens sequenced from Europe showed more diversity than the ones from North America, uncovering an invasion pattern likely related to its European native host in North America. Additionally, to the best of our knowledge, the specimen from France collected in c. 1811 is the oldest herbarium specimen sequenced from kingdom Fungi. In conclusion, sequencing old herbarium specimens is an important tool that can be extrapolated to better understand plant–microbe evolution and to evaluate old type specimens to solidify the taxonomy of plant pathogenic fungi.

Plant Sciences↗

Effects of Genetic and Physiological Divergence on the Evolution of a Sulfate-Reducing Bacterium under Conditions of Elevated Temperature

ABSTRACT Adaptation via natural selection is an important driver of evolution, and repeatable adaptations of replicate populations, under conditions of a constant environment, have been extensively reported. However, isolated groups of populations in nature tend to harbor both genetic and physiological divergence due to multiple selective pressures that they have encountered. How this divergence affects adaptation of these populations to a new common environment remains unclear. To determine the impact of prior genetic and physiological divergence in shaping adaptive evolution to accommodate a new common environment, an experimental evolution study with the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH) was conducted. Two groups of replicate populations with genetic and physiological divergence, derived from a previous evolution study, were propagated in an elevated-temperature environment for 1,000 generations. Ancestor populations without prior experimental evolution were also propagated in the same environment as a control. After 1,000 generations, all the populations had increased growth rates and all but one had greater fitness in the new environment than the ancestor population. Moreover, improvements in growth rate were moderately affected by the divergence in the starting populations, while changes in fitness were not significantly affected. The mutations acquired at the gene level in each group of populations were quite different, indicating that the observed phenotypic changes were achieved by evolutionary responses that differed between the groups. Overall, our work demonstrated that the initial differences in fitness between the starting populations were eliminated by adaptation and that phenotypic convergence was achieved by acquisition of mutations in different genes. IMPORTANCE Improving our understanding of how previous adaptation influences evolution has been a long-standing goal in evolutionary biology. Natural selection tends to drive populations to find similar adaptive solutions for the same selective conditions. However, variations in historical environments can lead to both physiological and genetic divergence that can make evolution unpredictable. Here, we assessed the influence of divergence on the evolution of a model sulfate-reducing bacterium, Desulfovibrio vulgaris Hildenborough, in response to elevated temperature and found a significant effect at the genetic but not the phenotypic level. Understanding how these influences drive evolution will allow us to better predict how bacteria will adapt to various ecological constraints.

59 BASIC BIOLOGICAL SCIENCES↗