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At least 19 records

MicroFisher: Fungal taxonomic classification for metatranscriptomic and metagenomic data using multiple short hypervariable markers

AbstractProfiling the taxonomic and functional composition of microbes using metagenomic (MG) and metatranscriptomic (MT) sequencing is advancing our understanding of microbial functions. However, the sensitivity and accuracy of microbial classification using genome– or core protein-based approaches, especially the classification of eukaryotic organisms, is limited by the availability of genomes and the resolution of sequence databases. To address this, we propose the MicroFisher, a novel approach that applies multiple hypervariable marker genes to profile fungal communities from MGs and MTs. This approach utilizes the hypervariable regions of ITS and large subunit (LSU) rRNA genes for fungal identification with high sensitivity and resolution. Simultaneously, we propose a computational pipeline (MicroFisher) to optimize and integrate the results from classifications using multiple hypervariable markers. To test the performance of our method, we applied MicroFisher to the synthetic community profiling and found high performance in fungal prediction and abundance estimation. In addition, we also used MGs from forest soil and MTs of root eukaryotic microbes to test our method and the results showed that MicroFisher provided more accurate profiling of environmental microbiomes compared to other classification tools. Overall, MicroFisher serves as a novel pipeline for classification of fungal communities from MGs and MTs.

Wang, Haihua↗

Molecular Transformation and Metabolic Insights of Microbial Electrolysis Treatment and Valorization of Post-hydrothermal Liquefaction Wastewater

Hydrothermal liquefaction (HTL) presents a promising approach for the conversion of wet waste into biocrude and biofuels. However, the post-hydrothermal liquefaction wastewater (PHW) poses significant challenges for treatment and valorization due to its high concentration and complex nature. In this study, we investigated the conversion pathways of major organic contaminants within the microbial electrolytic treatment of PHW from food waste HTL. To achieve this, we employed high-performance liquid chromatography (HPLC) and 2D nuclear magnetic resonance (NMR). Our findings demonstrate volatile fatty acids (VFAs) and monohydric and polyhydric alcohols were effectively transformed through the synergistic metabolism of fermentative and electroactive bacteria, which led to over 70% COD removal of the recalcitrant compounds and a record high H2 production rate (1.62 L L -1 d -1 ). We also employed the liquid-state 15N NMR on wastewater samples for the first time and revealed that the nitrogen-containing heteroaromatics were persistent to microbial electrochemical treatment. By integrating the chemical profiles with bioanode community profiles, we constructed a metabolic network that provides insights for enhancing treatment efficiency and facilitating resource recovery.

08 HYDROGEN↗

Carbon dioxide (CO2) flux, amplicon sequencing and liquid chromatography–mass spectrometry (LC-MS) data for pyrogenic organic matter (PYOM) amended soil incubations in lab, Madison, WI, 2021-22

This dataset comprises soil-pyrogenic organic matter (PyOM) incubation experiments conducted in a laboratory setting. The focus was on measuring carbon dioxide (CO2) flux emitted in the headspace of incubation jars over a one-month period. The aim was to estimate carbon mineralization from two different carbon fractions in PyOM, as well as carbon mineralization from soil organic carbon (SOC). Bacterial and fungal community profiles were tracked at various time points during the incubation to observe changes in the overall community and specific responders to PyOM. LC-MS analysis of soil samples was also performed to monitor changes in the soil's chemical composition.The provided dataset includes processed CO2 flux measurements obtained from the Picarro-multiplexer measurement setup for each sample incubation jar. This data is available in the "CO2Flux.csv" file. Additionally, the dataset includes CO2 flux data partitioned to estimate mean carbon mineralization from the two PyOM carbon fractions, bulk PyOM, and SOC. The cumulative carbon mineralization data is provided in "cml_respired.csv," while the rate of carbon mineralization data is in "rate_respired.csv." The file "cml_respired_lastcyc.csv" contains cumulative carbon mineralization data for the PyOM carbon fractions, bulk PyOM, and SOC throughout the entire incubation period.For the LC-MS analysis, the raw data can be found in the "20220721_NZ_PeakHeights_FinalAnalysis.csv" file, while sample metadata is provided in "20220721_NZpilot_Metadata.csv". Processed LC-MS data is available in the file "20220721_NZ_FINAL_msdat_sub3.csv". Lastly, the dataset includes the relative abundance data for bacterial genera that exhibited a significant positive response to the addition of PyOM produced at 350 degrees Celsius, which can be found in "resp_genera_350.csv".

54 ENVIRONMENTAL SCIENCES↗

Impact of intense sanitization procedures on bacterial communities recovered from floor drains in pork processing plants

Pork processing plants in the United States (US) cease operations for 24–48 h every six or twelve months to perform intense sanitization (IS) using fogging, foaming, and further antimicrobial treatments to disrupt natural biofilms that may harbor pathogens and spoilage organisms. The impact such treatments have on short-term changes in environmental microorganisms is not well understood, nor is the rate at which bacterial communities return. Swab samples were collected from floor drains to provide representative environmental microorganisms at two US pork processing plants before, during, and after an IS procedure. Samples were collected from four coolers where finished carcasses were chilled and from four locations near cutting tables. Each sample was characterized by total mesophile count (TMC), total psychrophile count (TPC), and other indicator bacteria; their biofilm-forming ability, tolerance of the formed biofilm to a quaternary ammonium compound (300 ppm, QAC), and ability to protect co-inoculated Salmonella enterica. In addition, bacterial community composition was determined using shotgun metagenomic sequencing. IS procedures disrupted bacteria present but to different extents depending on the plant and the area of the plant. IS reduced TPC and TMC, by up to 1.5 Log 10 CFU only to return to pre-IS levels within 2–3 days. The impact of IS on microorganisms in coolers was varied, with reductions of 2–4 Log 10 , and required 2 to 4 weeks to return to pre-IS levels. The results near fabrication lines were mixed, with little to no significant changes at one plant, while at the other, two processing lines showed 4 to 6 Log 10 reductions. Resistance to QAC and the protection of Salmonella by the biofilms varied between plants and between areas of the plants as well. Community profiling of bacteria at the genus level showed that IS reduced species diversity and the disruption led to new community compositions that in some cases did not return to the pre-IS state even after 15 to 16 weeks. The results found here reveal the impact of using IS to disrupt the presence of pathogen or spoilage microorganisms in US pork processing facilities may not have the intended effect.

59 BASIC BIOLOGICAL SCIENCES↗

Inclusive Shared Solar: The State Policy Landscape and Select Community Solar Project Profiles

This report provides an overview of the LMI community solar landscape to help State Energy Offices, LIHEAP offices, and other key stakeholders understand and take advantage of opportunities to expand the reach and impact of community solar programs. It discusses common LMI community solar policy and program considerations and profiles four projects in three different states to show how state policy decisions can impact community solar program design and implementation.

14 SOLAR ENERGY↗

Community-Centric Microgrid Feasibility Analysis Framework to Accelerate Resilience of Island Communities

Marine energy offers a reliable energy solution for island and coastal communities, which often lack traditional local generation, to support their transition to energy independence and reduce reliance on externally imported fuels. Successful deployment of new technologies in these isolated locations requires community acceptance and approval from the outset, as these communities typically lack the financial and technical resources to operate and maintain new systems. This report presents a community-centric microgrid planning framework for remote coastal and island communities. Community engagement is integrated as the first step in the planning process, incorporating community profiles and visions into energy development scenarios. A case study was conducted in St. George, Pribilof Islands, Alaska, which relies entirely on diesel yet has significant wind and wave energy potential. Community engagement revealed a unique history and current economic status, with an interest in adopting advanced energy technologies despite past failures. Various microgrid configurations were optimized, considering different technologies to meet current and future energy needs while balancing cost and energy resilience. Wave energy converters (WECs) were a key component, integrated with other energy sources using the Xendee optimization tool. The Marine Energy Microgrid Toolkit, developed as part of this work, uses commercial power system analysis tools to optimize and analyze microgrid scenarios. The developed framework and toolkit can be applied to island and coastal communities to enhance resilience and support microgrid deployments. Future enhancements will include incorporating new marine resources, developing dynamic models, and automating the integration of Xendee and PowerFactory simulations.

02 PETROLEUM↗

Community-level physiological profiling performed with an oxygen-sensitive fluorophore in a microtiter plate

Community-level physiological profiling based upon fluorometric detection of oxygen consumption was performed on hydroponic rhizosphere and salt marsh litter samples by using substrate levels as low as 50 ppm with incubation times between 5 and 24 h. The rate and extent of response were increased in samples acclimated to specific substrates and were reduced by limiting nitrogen availability in the wells.

NASA Discipline Life Support Systems↗

Geobacter sp. Strain IAE Dihaloeliminates 1,1,2-Trichloroethane and 1,2-Dichloroethane

We report that chlorinated ethanes, including 1,2-dichloroethane (1,2-DCA) and 1,1,2-trichloroethane (1,1,2-TCA), are widespread groundwater contaminants. Enrichment cultures XR DCA and XR TCA derived from river sediment dihaloeliminated 1,2-DCA to ethene and 1,1,2-TCA to vinyl chloride (VC), respectively. The XR TCA culture subsequently converted VC to ethene via hydrogenolysis. Microbial community profiling demonstrated the enrichment of Geobacter 16S rRNA gene sequences in both the XR DCA and XR TCA cultures, and Dehalococcoides mccartyi (Dhc) sequences were only detected in the ethene-producing XR TCA culture. The presence of a novel Geobacter population, designated as Geobacter sp. strain IAE, was identified by the 16S rRNA gene-targeted polymerase chain reaction and Sanger sequencing. Time-resolved population dynamics attributed the dihaloelimination activity to strain IAE, which attained the growth yields of 0.93 ± 0.06 x 10 7 and 1.18 ± 0.14 x 10 7 cells per μmol Cl -1 released with 1,2-DCA and 1,1,2-TCA as electron acceptors, respectively. In contrast, Dhc growth only occurred during VC-to-ethene hydrogenolysis. Our findings discover a Geobacter sp. strain capable of respiring multiple chlorinated ethanes and demonstrate the involvement of a broader diversity of organohalide-respiring bacteria in the detoxification of 1,2-DCA and 1,1,2-TCA.

1,1,2-trichloroethane↗

Linking Spatiotemporal Biological Data to Predict Harmful Algal Blooms

Cyanobacterial Harmful Algal Blooms (cHABs) have significant impacts on an affected region’s economy, ecology, and human health. The blooms can release toxins that kill fish and poison water for people and animals. The global adverse effects of cHABs are exacerbated by the consequences of climate change and increased pollution. Though the phenomena are well documented, scientists’ efforts to mitigate the damage are hampered by insufficient predictive models and incomplete granular knowledge of cHAB community structure. With a goal of leveraging bioinformatics and machine learning tools to better understand and predict cHABs, we are first exploring water sample data sets. Using nearly four thousand samples from the National Center for Biotechnology Information Sequence Read Archive (NCBI-SRA) across 16 years with latitude and longitude embedded in the metadata, we mapped the location of the samples onto a Lake Erie shape file. We combined information about location, date, and community taxa in the NCBI samples to discover factors that determine cHAB features. The data are separated into three distinct zones, with the majority pooled at the southwest end of the lake and occurring in 2017. The samples are rich in biological data; our next steps are to carry out whole genome sequence analysis and use the community profiles as part of our predictive machine learning model.

59 BASIC BIOLOGICAL SCIENCES↗

Final Report for CSP Tower Public Opinion and Education Project

As part of the CSP Plant Optimization Study for the California Power Market (DE-EE0009809) the project wanted to understand the public’s opinion of the technology and and explore the types of community engagement that is needed to support such development. The initial objectives for the public perception activities were twofold. The primary objective was to gather public opinion and feedback from the communities living near the two operating solar power tower plants, Ivanpah and Crescent Dunes, and to distill lessons learned from the community engagement conducted before, during, and after the plants were developed to inform future development. This included outreach to nearby airports. The other objective was to gauge public opinion about large-scale solar, specifically CSP towers, to start educating the public on the benefits and to begin building relationships with communities of interest, initially targeting the Kingman, Arizona area. The objectives shifted after the first exploratory trip to Ivanpah and Kingman, however, as it became apparent that it would be challenging to gather public opinion from the community surrounding Ivanpah that could be useful for other community profiles, and the company wasn’t ready to address the concerns in Kingman. Another area was chosen, therefore, to represent those where future development is possible. The recently published Lawrence Berkeley National Lab Perceptions of Large-Scale Solar Project Neighbors Study exemplified public perception polling based on social science and served as the foundation for the survey questions taken to the field. The intent was to ensure that people knew their input was valued and that the time they spent was valuable for the participant as well. It has been noted in the literature that in-person interaction has greater benefits than activities online or via mail, as well as limits the expense.

14 SOLAR ENERGY↗

Response of soybean rhizosphere communities to human hygiene water addition as determined by community level physiological profiling (CLPP) and terminal restriction fragment length polymorphism (TRFLP) analysis

In this report, we describe an experiment conducted at Kennedy Space Center in the biomass production chamber (BPC) using soybean plants for purification and processing of human hygiene water. Specifically, we tested whether it was possible to detect changes in the root-associated bacterial assemblage of the plants and ultimately to identify the specific microorganism(s) which differed when plants were exposed to hygiene water and other hydroponic media. Plants were grown in hydroponics media corresponding to four different treatments: control (Hoagland's solution), artificial gray water (Hoagland's+surfactant), filtered gray water collected from human subjects on site, and unfiltered gray water. Differences in rhizosphere microbial populations in all experimental treatments were observed when compared to the control treatment using both community level physiological profiles (BIOLOG) and molecular fingerprinting of 16S rRNA genes by terminal restriction fragment length polymorphism analysis (TRFLP). Furthermore, screening of a clonal library of 16S rRNA genes by TRFLP yielded nearly full length SSU genes associated with the various treatments. Most 16S rRNA genes were affiliated with the Klebsiella, Pseudomonas, Variovorax, Burkholderia, Bordetella and Isosphaera groups. This molecular approach demonstrated the ability to rapidly detect and identify microorganisms unique to experimental treatments and provides a means to fingerprint microbial communities in the biosystems being developed at NASA for optimizing advanced life support operations.

NASA Discipline Life Support Systems↗

Disruption of the endogenous indole glucosinolate pathway impacts the Arabidopsis thaliana root exudation profile and rhizobacterial community

Root exudates are composed of primary and secondary metabolites known to modulate the rhizosphere microbiota. Glucosinolates are defense compounds present in the Brassicaceae family capable of deterring pathogens, herbivores and biotic stressors in the phyllosphere. In addition, traces of glucosinolates and their hydrolyzed byproducts have been found in the soil, suggesting that these secondary metabolites could play a role in the modulation and establishment of the rhizosphere microbial community associated with this family. Here, we used Arabidopsis thaliana mutant lines, including the cyp79B2cyp79B3 double mutant line with a disruption in the indole glucosinolate pathway and atr1D, which overexpresses ATR1 and increases glucosinolate production. These lines were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and 16S rRNA amplicon sequencing to evaluate how genetic modifications to the indole glucosinolate pathway affects the root exudate profile of Arabidopsis thaliana, and, in turn, impacts the rhizosphere microbial community. Metabolic analysis of root exudates from the wild-type Columbia (Col-0), along with the mutant lines, confirmed that alterations to the indole glucosinolate biosynthetic pathway result in shifts in the root exudate profile of the plant. We observed changes in the relative abundance of exuded metabolites. Moreover, 16S rRNA amplicon sequencing results provided evidence that the rhizobacterial communities associated with the plant lines used were directly impacted in diversity and community composition. Here, this work provides further information on the involvement of secondary metabolites and their role in modulating the rhizobacterial community. Root metabolites dictate the presence of different bacterial species, including plant growth-promoting rhizobacteria (PGPR). Our results suggest that genetic alterations in the indole glucosinolate pathway cause disruptions beyond the endogenous levels of the plant, significantly changing the abundance and presence of different metabolites in the root exudates of the plants as well as the microbial rhizosphere community.

59 BASIC BIOLOGICAL SCIENCES↗

Distinct Depth-Discrete Profiles of Microbial Communities and Geochemical Insights in the Subsurface Critical Zone

Microbial assembly and metabolic potential in the subsurface critical zone (SCZ) are substantially impacted by subsurface geochemistry and hydrogeology, selecting for microbes distinct from those in surficial soils. In this study, we integrated metagenomics and geochemistry to elucidate how microbial composition and metabolic potential are shaped and impacted by vertical variations in geochemistry and hydrogeology in terrestrial subsurface sediment. A sediment core from an uncontaminated, pristine well at Oak Ridge Field Research Center in Oak Ridge, Tennessee, including the shallow subsurface, vadose zone, capillary fringe, and saturated zone, was used in this study. Our results showed that subsurface microbes were highly localized and that communities were rarely interconnected. Microbial community composition as well as metabolic potential in carbon and nitrogen cycling varied even over short vertical distances. Further analyses indicated a strong depth-related covariation of community composition with a subset of 12 environmental variables. An analysis of dissolved organic carbon (DOC) quality via ultrahigh resolution mass spectrometry suggested that the SCZ was generally a low-carbon environment, with the relative portion of labile DOC decreasing and that of recalcitrant DOC increasing along the depth, selecting microbes from copiotrophs to oligotrophs and also impacting the microbial metabolic potential in the carbon cycle. Our study demonstrates that sediment geochemistry and hydrogeology are vital in the selection of distinct microbial populations and metabolism in the SCZ.

59 BASIC BIOLOGICAL SCIENCES↗

Relative effectiveness of kinetic analysis vs single point readings for classifying environmental samples based on community-level physiological profiles (CLPP)

The relative effectiveness of average-well-color-development-normalized single-point absorbance readings (AWCD) vs the kinetic parameters mu(m), lambda, A, and integral (AREA) of the modified Gompertz equation fit to the color development curve resulting from reduction of a redox sensitive dye from microbial respiration of 95 separate sole carbon sources in microplate wells was compared for a dilution series of rhizosphere samples from hydroponically grown wheat and potato ranging in inoculum densities of 1 x 10(4)-4 x 10(6) cells ml-1. Patterns generated with each parameter were analyzed using principal component analysis (PCA) and discriminant function analysis (DFA) to test relative resolving power. Samples of equivalent cell density (undiluted samples) were correctly classified by rhizosphere type for all parameters based on DFA analysis of the first five PC scores. Analysis of undiluted and 1:4 diluted samples resulted in misclassification of at least two of the wheat samples for all parameters except the AWCD normalized (0.50 abs. units) data, and analysis of undiluted, 1:4, and 1:16 diluted samples resulted in misclassification for all parameter types. Ordination of samples along the first principal component (PC) was correlated to inoculum density in analyses performed on all of the kinetic parameters, but no such influence was seen for AWCD-derived results. The carbon sources responsible for classification differed among the variable types with the exception of AREA and A, which were strongly correlated. These results indicate that the use of kinetic parameters for pattern analysis in CLPP may provide some additional information, but only if the influence of inoculum density is carefully considered. c2001 Elsevier Science Ltd. All rights reserved.

NASA Center KSC↗

Composition and physiological profiling of sprout-associated microbial communities

The native microfloras of various types of sprouts (alfalfa, clover, sunflower, mung bean, and broccoli sprouts) were examined to assess the relative effects of sprout type and inoculum factors (i.e., sprout-growing facility, seed lot, and inoculation with sprout-derived inocula) on the microbial community structure of sprouts. Sprouts were sonicated for 7 min or hand shaken with glass beads for 2 min to recover native microfloras from the surface, and the resulting suspensions were diluted and plated. The culturable fraction was characterized by the density (log CFU/g), richness (e.g., number of types of bacteria), and diversity (e.g., microbial richness and evenness) of colonies on tryptic soy agar plates incubated for 48 h at 30 degrees C. The relative similarity between sprout-associated microbial communities was assessed with the use of community-level physiological profiles (CLPPs) based on patterns of utilization of 95 separate carbon sources. Aerobic plate counts of 7.96 +/- 0.91 log CFU/g of sprout tissue (fresh weight) were observed, with no statistically significant differences in microbial cell density, richness, or diversity due to sprout type, sprout-growing facility, or seed lot. CLPP analyses revealed that the microbial communities associated with alfalfa and clover sprouts are more similar than those associated with the other sprout types tested. Variability among sprout types was more extensive than any differences between microbial communities associated with alfalfa and clover sprouts from different sprout-growing facilities and seed lots. These results indicate that the subsequent testing of biocontrol agents should focus on similar organisms for alfalfa and clover, but alternative types may be most suitable for the other sprout types tested. The inoculation of alfalfa sprouts with communities derived from various sprout types had a significant, source-independent effect on microbial community structure, indicating that the process of inoculation alters the dynamics of community development regardless of the types of organisms involved.

NASA Center KSC↗

Microgrid Hardening Design Toolkit User Guide: Software v0.29

Microgrid Hardening Design Toolkit helps microgrid stakeholders assess community-specific natural hazard risks to microgrid components and provides mitigation strategies to reduce those risks. Using census tract location inputs and the Federal Emergency Management Agency (FEMA) Risk Index dataset, the tool provides a ranked list of the highest risk hazards for a selected community and hazard-specific mitigation options for microgrid components. It also provides risk twin communities with similar hazard risk profiles and includes a radar-chart to visually compare risk profiles across communities. The toolkit is expected to help users quickly understand community-specific hazard risks and make decisions for microgrid hardening.

24 POWER TRANSMISSION AND DISTRIBUTION↗

Ectomycorrhizal fungi alter soil food webs and the functional potential of bacterial communities

Most of Earth’s trees rely on critical soil nutrients that ectomycorrhizal fungi (EcMF) liberate and provide, and all of Earth’s land plants associate with bacteria that help them survive in nature. Yet, our understanding of how the presence of EcMF modifies soil bacterial communities, soil food webs, and root chemistry requires direct experimental evidence to comprehend the effects that EcMF may generate in the belowground plant microbiome. To this end, we grew Pinus muricata plants in soils that were either inoculated with EcMF and native forest bacterial communities or only native bacterial communities. We then profiled the soil bacterial communities, applied metabolomics and lipidomics, and linked omics data sets to understand how the presence of EcMF modifies belowground biogeochemistry, bacterial community structure, and their functional potential. We found that the presence of EcMF (i) enriches soil bacteria linked to enhanced plant growth in nature, (ii) alters the quantity and composition of lipid and non-lipid soil metabolites, and (iii) modifies plant root chemistry toward pathogen suppression, enzymatic conservation, and reactive oxygen species scavenging. Using this multi-omic approach, we therefore show that this widespread fungal symbiosis may be a common factor for structuring soil food webs.

59 BASIC BIOLOGICAL SCIENCES↗

Potential for functional divergence in ectomycorrhizal fungal communities across a precipitation gradient

Abstract Functional traits influence the assembly of microbial communities, but identifying these traits in the environment has remained challenging. We studied ectomycorrhizal fungal (EMF) communities inhabiting Populus trichocarpa roots distributed across a precipitation gradient in the Pacific Northwest, USA. We profiled these communities using taxonomic (meta-barcoding) and functional (metagenomic) approaches. We hypothesized that genes involved in fungal drought-stress tolerance and fungal mediated plant water uptake would be most abundant in drier soils. We were unable to detect support for this hypothesis; instead, the abundance of genes involved in melanin synthesis, hydrophobins, aquaporins, trehalose-synthases, and other gene families exhibited no significant shifts across the gradient. Finally, we studied variation in sequence homology for certain genes, finding that fungal communities in dry soils are composed of distinct aquaporin and hydrophobin gene sequences. Altogether, our results suggest that while EMF communities exhibit significant compositional shifts across this gradient, coupled functional turnover, at least as inferred using community metagenomics is limited. Accordingly, the consequences of these distinct EMF communities on plant water uptake remain critically unknown, and future studies targeting the expression of genes involved in drought stress tolerance are required.

59 BASIC BIOLOGICAL SCIENCES↗