Rob Knight - US grants

DESCRIPTION (provided by applicant): The microbes that inhabit human bodies outnumber the human cells by an order of magnitude, and impact many aspects of health and disease including obesity, vaginosis, and Crohn's disease. Understanding this endogenous microbiota is emerging as a key extension of efforts to understand the human genome and the role of genetic variation on health and disease. The Human Microbiome Project (HMP) will characterize microbial communities in a large number of individual healthy humans using metagenomic sequencing. Consequently, new methods for interpreting sequence data to understand microbial community composition and dynamics are urgently needed. This project unites disciplines ranging from ecology to evolutionary biology to applied mathematics, to develop new methods for understanding which body habitats are more or less similar in terms of their microbial communities, by evaluating measures of microbial diversity and change, and creating needed new metrics of community composition. This will enable understanding of how clinically relevant parameters such as age, sex, or the pH of specific body habitats affect these communities, and of how the dynamics of change in microbial communities within an individual, in transmission between individuals, and in transmission between humans and the environment. This project is directly responsive to the Roadmap RFA for Development of New tools for Computational Analysis of Human Microbiome Project Data. The specific aims of this proposal are: Aim 1. Develop, characterize, and apply enriched descriptors of microbial community diversity. Aim 2. Develop methods for describing how human microbial communities vary over time and space. Aim 3. Develop new methods for tracing the flow of organisms among different communities. Some key aspects of the proposed work are: the development of new statistical methods for estimating microbial diversity within a body habitat; development of enriched methods for describing microbial community diversity; exhaustive validation of methods for comparing microbial communities through large-scale simulations and by using the largest available data sets that characterize microbial communities empirically; and the development of new methods for tracing the sources of the microbes that inhabit the human body using both marker genes and whole-metagenome data. Key outcomes include the ability to help determine the extent to which there is a core human microbiome, and how best to sample human microbial diversity. All methods developed will be made available under open source licenses and will be deposited with the HMP Data Analysis and Coordination Center (DACC). The investigators intend to work closely with other researchers involved in the HMP in order to ensure rapid progress. PUBLIC HEALTH RELEVANCE: Microbial communities associated with the human body play critical roles in human health and disease. This project will provide methods that help establish the nature and variability of microbial communities in healthy human individuals. Using these individuals as a baseline, this work will will pave the way for studies of a wide range of medical conditions that these communities affect by looking for abnormal communities associated with specific disease states, allowing the development of new diagnostics and therapeutic approaches.

All species of plants and animals harbor microbes (bacteria and fungi) that live symbiotically in and on them. How those microbes are related to the health of their host organisms is largely undetermined and, in particular, there is a limited understanding of how symbiotic microbes influence a host's ability to cope with disease-causing pathogens. The proposed research will use amphibians as a model system and a fungal skin pathogen that is linked to the decline of amphibian populations around the world. Different amphibian species are either highly susceptible or tolerant to disease caused by the fungal pathogen, and there is reasonable evidence to suggest that naturally occurring skin microbes may play an important role. This research will use both field observations from wild amphibian populations and a series of specific experiments that will be conducted in the laboratory. In the field, amphibians will be sampled for microbes to compare differences among species in pond habitats. In the lab, controlled experiments will examine how microbes on the skin respond to the pathogen over time and also how they respond to probiotic treatments (beneficial bacteria) in an effort to understand the ecology of the microorganisms that live on the skin. Novel high-throughput DNA sequencing techniques and innovative computer-based analysis tools will be used to examine the identity and composition of different bacterial and fungal groups in each sample.

Broader Impacts: This research aims to advance our knowledge about the role of symbiotic skin microbes and how probiotic treatments can be developed for use in conservation of amphibians as well as a broad range of human and wildlife disease systems. The project will advance the research program of an early career scientist (PI) and support the training of a graduate student, a postdoctoral researcher, and undergraduates from under-represented backgrounds. All DNA sequence data and analytic tools produced during this project will be made available for use by other researchers. Boreal toads, an endangered amphibian species in Colorado, will serve as a focal species in this project and the PIs will build collaborations with the Boreal Toad Recovery Program in Colorado.

SUMMARY Core B will provide database infrastructure and coordinate data deposition to public resources such as INSDC (the International Nucleotide Sequence Database Collaboration), as well as coordinate the data for the project overall to facilitate comparison with other datasets and deployment of advanced algorithms. It will build on the Knight lab's extensive experience with meta-analysis, sequence databases, and data visualization to provide these methods to Project 1, Project 2, and Project 3, and will work closely with Core A to mirror metabolomics data and integrate metabolomics datasets with the rest of the multi-omic data to be collected. Core B has three Aims. Aim 1-organize the data and metadata collected in Project 1 from mice and Project 2 from humans, curate these datasets, and ensure that analyses are reproducible in an automated fashion using virtual machines. Aim 2-deposit the data and metadata in standards-compliant form to INSDC, the Gene Expression Omnibus, and other resources (e.g., metabolomics repositories) as they emerge. Aim 3-Through analyses of existing microbiome datasets, provide best-practices recommendations to investigators in Project 1 and Project 2 to optimize experimental design. Core B will build on an extensive multi-omics data repository funded by multiple sources that is able to accommodate the types of data to be collected in the project overall, including links between human subjects with defined family relationships (e.g. dizygotic twins), humanized gnotobiotic mice colonized with strains derived from these human subjects, timeseries study designs in both humans and mice, combinations of data at multiple levels including 16S rRNA gene sequencing, RNA-Seq, and metabolomics, and other advanced features of this complex project. A key component of our approach is to enable investigators in the laboratory collecting the datasets to perform their own first-pass analyses while at the same time making the data available more broadly within the project for additional advanced analyses, such as those being developed in Project 3, to be applied, and also making the data available to the public in a relatively user-friendly form to supplement public deposition in permanent government-backed sequence data repositories.

SUMMARY Project 3 will develop new computational tools for understanding how gut microbial communities change their membership, gene content, gene expression, and metabolic activities in obesity and during diet interventions, in both humans and 'humanized' gnotobiotic mouse models. Its overarching goals are to: (i) understand the levels at which multi-omics data should be collected and analyzed in order to maximize our understanding of complex multifactorial pathophysiological conditions such as obesity and its associated metabolic abnormalities; (ii) develop improved genome assembly techniques and predictions about culture conditions and syntrophic interactions to improve the utility of personalized bacterial culture collections and data derived from them; and (iii) understand how best to characterize the diversity of gut microbial communities, and the functional profiles of these communities, observed in the human population and use them for patient stratification. Project 3 has three Aims: (1) develop new tools for relating multi-omics data across analysis levels and relating information from mouse models, specifically gnotobiotic humanized mice characterized in Project 1 and Core A, to information about the discordant twins (phenotyped in Project 2 and Core A) from which those animal models were derived and to the human population at large; (2) to develop improved methods for assembly of complete bacterial genomes as a reference for shotgun metagenomic and meta- transcriptomic data, including meta-transcriptome data collected from gnotobiotic mice colonized with bacterial culture collections generated from the fecal microbiota of co-twins in discordant twin pairs where the complete bacterial genomes are known, thus eliminating a major computational bottleneck and providing more relevant types of assemblies for downstream annotation and interpretation tasks; and (3) to provide a broader understanding of the major patterns of variation in human gut microbial communities and their genes, transcripts and metabolites in individuals with and without obesity and obesity associated abnormalities, and to test whether these major patterns can be used for stratification of human subjects in terms of their response to specific dietary or other interventions. Project 3 and Core B will work closely together to make new analysis tools and datasets available to the scientific community.

The evolutionary relationships among species of animals, plants, fungi and microbes is a large, complex, branching network that biologists call the Tree of Life. Obtaining the complete Tree of Life for all living things is a grand challenge in science, on the same intellectual scale as investigating the nature of matter or the origin of the universe: it is fundamental to understanding our world, central to human sustainability, and critical as a framework for future discovery. Scientists have made major progress toward this challenge by combining biology with computer science and successfully generating large genealogies of thousands and even millions of species. These large phylogenetic trees are now available to researchers in many branches of biology -- from genomics to ecology -- and have great promise for applied science in medicine, agriculture, industry, and climate-change mitigation. It is critical that scientists develop a future vision for this area of scientific exploration and make clear the central role of phylogenetics in the strategic integration of biodiversity science across US academic, corporate and government institutions. To achieve this vision, this project has developed a three-year program of catalysis meetings and workshops to bring together and enlarge the Tree of Life community. By uniting a wide range of biologists, computational experts, and representatives from corporate entities, foundations, and government agencies, the meeting series will generate a mechanism for scientific input into the strategic vision of biodiversity science.

The workshops series will impact many areas of research involving biodiversity and evolution by addressing three over-arching goals: (1) identifying challenges and progress in generating, storing and visualizing the genealogy of life, (2) integrating large data layers with the tree of life, and (3) developing a clear plan and compelling vision for the future of phylogenetic biology. The workshops and catalysis meetings will include students and early-career scientists, as well as educators and members of key corporate, government and non-profit organizations, with the aim to create new, lasting partnerships that bridge across interest groups. Tangible products of the meetings will include educational materials, publications that highlight broader uses of Tree of Life as well as advances in software that will be open source and available to a wide audience. At completion of the workshop series the participants will produce a white paper discussing ideas and challenges for long-term sustainability of biodiversity data.

Single-cellular organisms (microbes) represent a vast component of the diversity of life on Earth and perform an amazing array of biological functions. They rarely live or act alone and instead exist in complex communities composed of many interacting species that make up the microbiome. This award supports the development of the next generation of Quantitative Insights Into Microbial Ecology (QIIME, pronounced "chime"), a free and open source software platform for analyzing microbiomes based on DNA sequencing data. Microbiome science is in a transformation from being descriptive and technically challenging, to becoming hypothesis-driven, actionable, and technically straight-forward, in part enabled by QIIME. We now know that the traditional approach for studying microbial communities, which relied on culturing microbes in the lab, is insufficient because we don't know the conditions required for the growth of most microbes. Recent advances link microbiomes to functional processes via 'culture independent' techniques, such as sequencing fragments of microbial genomes, and then using those fragments as 'molecular fingerprints' to profile the microbiome. The bottleneck in microbiome analysis is not DNA sequencing, but in interpreting the large quantities of sequence data generated. QIIME 2 will advance knowledge of microbiomes by helping users derive insight through interactive exploratory analysis capabilities, understand the underlying methods, and report their results in ways accessible to end users from outside of the field, including physicians, engineers and policymakers who urgently need access to conclusions drawn from studies of complex microbial ecosystems. Societal benefits range from global to personal (from understanding cycling of biologically essential nutrients, such as carbon and nitrogen in the environment to curing disease, including obesity and cancer). QIIME has been cited over 4,000 times and has active user and developer communities. Educational workshops on QIIME are taught approximately monthly in the USA and around the world.

At its core, QIIME 2 will provide a stable application programming interface (API) relying on existing community standards for documentation, coding style, and testing. It will have a novel 'documentation-driven' graphical user interface that will make QIIME accessible to users without requiring advanced computational skills. At the same time, it will help users improve their computational skills through exposure to the underlying bioinformatics methods. QIIME 2 will have fully integrated provenance tracking, which will simplify reporting and the reproducibility of bioinformatics workflows. A first-class plugin system will decentralize development by allowing outside developers to add new methods to the QIIME 2 platform. The API will also support improved integration of QIIME as a component of other widely used systems, such as Illumina BaseSpace® and Qiita, and an automatically generated command line interface will be provided for power users. QIIME 2 will have a completely redeveloped parallel framework, which will support deployment on diverse high-performance computing resources, from locally owned and operated computer clusters to commercially available cloud computing platforms. All stages of QIIME 2 development will be driven by user community input through the QIIME Forum (currently over 2500 active users) and our public GitHub repository. Further details on this project are on the QIIME website (www.qiime.org).

The project will characterize the functional, taxonomic, biotic and abiotic drivers of soil ecosystems in the Trans Antarctic Mountains (one of the most remote and harsh terrestrial landscapes on the planet). The work will utilize new high-throughput DNA and RNA sequencing technologies to identify members of the microbial communities and determine if the microbial community structures are independent of local environmental heterogeneities. In addition the project will determine if microbial diversity and function are correlated with time since the last glacial maximum (LGM). The expected results will greatly contribute to our knowledge regarding rates of microbial succession and help define the some of the limits to life and life-maintaining processes on Earth.

The project will analyze genomes and RNA derived from these genomes to describe the relationships between biodiversity and ecosystem functioning from soils above and below LGM elevations and to correlate these with the environmental drivers associated with their development during the last ~18,000 years. The team will identify the taxonomic diversity and the functional genetic composition within a broad suite of soil biota and examine their patterns of assembly and distribution within the framework of their geological legacies. The project will mentor participants from undergraduate students to postdoctoral researchers and prepare them to effectively engage in research to meet their career aspirations. The project will contribute to ongoing public education efforts through relationships with K-12 teachers and administrators- to include University-Public School partnerships. Less formal activities include public lecture series and weblogs aimed at providing information on Antarctic polar desert ecosystems to the general public. Targeted classrooms near each PI's institution will participate in online, real-time discussions about current topics in Antarctic ecosystems research.

The Hispanic/Latino population is the fasting growing segment of the US population. Diabetes disproportionately affects this group. National US 2007-09 data found that >20 yr old Hispanics (11.8%) have a 66% higher rate of diabetes compared to non-Hispanic whites (7.1%). In the population-based Hispanic Community Health Study (HCHS)/Study of Latinos (SOL), diabetes had a baseline prevalence of approximately 17%. Very recent data implicates the gut microbiome (GMB) as a key determinant of diabetes. Since different ancestral populations harbor different diabetes-associated sets of GMBs, it is necessary to study Hispanic/Latino populations with high rates of diabetes to determine the relationship between the GMB and diabetes. Understanding the relationship of the GMB to diabetes is anticipated to lead to a new era of prevention and treatment options, especially since therapeutic interventions are available that target the GMB. Nevertheless, there are major gaps in understanding the epidemiology of the GMB in the population and its role in the development of diabetes. The proposed study will leverage the HCHS/SOL study that will re-examine the participants in 2014-2017. This study has a major focus on diabetes including a fasting 2h glucose tolerance test (GTT), a standardized and universally accepted metric of glucose metabolism, in addition to specific other laboratory and clinical measurements. This proposed ancillary study will test the hypothesis that specific patterns of the gut microbiome will be significantly associated with pre-diabetes and diabetes, building upon recent advances in understanding the importance of the GMB in human health and metabolic diseases. This project will collect and determine the genetic composition of the fecal microbiome from 2,000 cohort members. The proposed study has developed a unique multidisciplinary team to address the following specific aims: (1) to investigate epidemiological factors affecting the gut microbiome in the sample of Hispanic/Latino individuals of diverse background who have normal indices of carbohydrate metabolism. We will test the association of geographic/ancestral background (e.g., Mexican, Puerto Rican), US birth status, gender, age, BMI, shared household and relatedness, and other variables with the GBM composition; (2) to utilize a cross-sectional design to evaluate the association of the gut microbiome (GMB) with the presence of disorders of carbohydrate metabolism including diabetes and prediabetes; and (3) to examine the longitudinal association of the GMB with risk of developing diabetes. We will use the active follow-up in the entire cohort to identify individuals who develop diabetes and estimate the relative risk of disease associated with different microbiomes. We hypothesize that the microbiomes found to be cross-sectionally associated with diabetes in Aim 2 will be predictive of the development of diabetes among initially pre-diabetic and normoglycemic individuals. 1

Summary/Abstract Our prior work in HIV cohort studies provides insights into the viral, inflammation, immune activation and antiretroviral therapy related risk factors for HIV-related CVD risk. Yet, an understanding of its pathophysiology remains incomplete. Emerging evidence suggests that gut microbiota (GMB) altered during HIV infection correlates with increased immune activation and disrupted metabolite profiles, but the role of GMB in HIV- related CVD is unknown. Our preliminary data show that in HIV infection, progression of atherosclerosis is associated with higher circulating sCD14, a marker of monocyte activation, and increased tryptophan catabolism. This preliminary work presents two promising candidates linking GMB and CVD risk in HIV infection which we propose to study using integrated ?Omics? approaches. In addition to these hypothesis- driven study aims, we will also generate novel hypotheses linking GMB, host immune activation and metabolomics profiles associated with CVD risk. We will leverage the Women?s Interagency HIV Study (WIHS) and Multicenter AIDS Cohort Study (MACS) >20 year follow-up for biospecimens, atherosclerosis and other CVD measures, and HIV parameters. Our longitudinal semi-annual measures allow us to subset individuals according to long-term HIV treatment, disease progression markers (CD4+ T-cell count, viral load), and comorbidity, with inclusion of matched HIV-uninfected participants. In this project, We will extend our established collaborations with leading labs to gather multi-dimensional data among 400 women and men (~65% of whom are HIV+), including stool GMB metagenomics, serum and cellular inflammation and immunologic markers (sCD14, monocyte transcriptomics), metabolomics, and measures (carotid artery ultrasound imaging over 4 year follow-up). Findings from this intensively studied group will then be extended to a larger sample of 746 women and men with metabolomics and longitudinal atherosclerosis data. In this project, we will have the opportunity to identify immune activation and metabolites underlying the role of GMB in CVD, which may be specific to HIV+ individuals, or accentuated in the setting of HIV.

ABSTRACT ? Overall Behavioral, emotional and cognitive disorders have been historically studied as diseases of the central nervous system (CNS). However, emerging data suggests a gut-brain connection in a variety of diseases that affect the brain. Our own data and others? suggests a gut-brain connection in Alzheimer?s disease (AD), a progressive neurodegenerative disorder that is the leading cause of dementia. There are currently no therapies to prevent or slow AD progression, causing a huge socioeconomic burden and highlighting our incomplete knowledge. Given an emerging role for gut microbiome and hypotheses implicating viral and bacterial contributions to AD pathogenesis, defining the bidirectional biochemical communication between the brain and the gut will improve understanding of neurodegenerative and psychiatric diseases. Indeed, it is crucial to study the brain not in isolation, but in the context of peripheral influences including diet, lifestyle, and microbiome. In this proposal we build on large initiatives and infrastructures co-established by our multi-disciplinary team including: The American Gut Project, The AD Metabolomics Consortium, Alzheimer?s Disease Research Centers (ADRCs), National Centralized Repository for Alzheimer's Disease and Related Dementias (NCRAD), The National Alzheimer?s Coordinating Center (NACC) and centers of excellence in informatics and systems biology. We aim to define the role of gut microbiome in AD pathogenesis and biochemical axis of communication between gut and brain. Aim 1: Examine the association between the gut microbiome and AD phenotypes. Aim 2: Define the biochemical axis of communication between the gut microbiome and the brain and identify metabolites that contribute to AD endophenotypes. Aim 3: Examine mechanistic links between the activity of the gut microbiome and AD pathogenesis, and identify new approaches for AD prevention that target the gut-brain axis. These aims will be enabled three projects supported by an Omics and Technology Core, a Computational and Systems Biology Core, and an Administrative Core that provides data and biorepository infrastructure. Project 1 will define changes in gut microbiome and metabolome across the AD trajectory; Project 2 leverages three existing clinical trials of controlled diets to examine dietary effects on gut microbiome, metabolome, cognition and brain imaging; Project 3 examines mechanism by defining gut- brain communication and microbiome-based interventions in animal models of AD. In this U19 we will create an unprecedented, high-quality dataset and resources specifically for the AD research community, and make these available under open science, FAIR (findable, accessible, interoperable, reusable) data principles. With our cross-disciplinary team of experts in clinical AD, gut microbiome research, imaging, metabolomics, informatics, deep learning and systems biology, this effort will yield novel biomarkers for AD progression and prognosis, and insight into mechanisms opening the door to development of transformative options for AD.

Summary Complex interactions between numerous specialized host cells in gut, liver and other locations, polymorphisms of multiple susceptibility genes and their associated signaling pathways and functions, and a myriad of microbial and other environmental factors are emerging as central to the pathogenesis of many of the most important digestive diseases, as exemplified by inflammatory bowel disease (IBD) and non-alcoholic fatty liver disease (NAFLD). It is becomingly increasingly evident that more comprehensive views of the players and interactions are needed to unravel many of the most perplexing digestive diseases. Consequently, tools to characterize and define multiple host and microbial cells and their gene expression responses are increasingly important and essential for digestive diseases research. Most advanced among these tools are technologies based on high- throughput sequencing, as it is now routinely possible to obtain hundreds of millions of base pairs of sequence at modest costs. Next-generation sequencing technologies have revolutionized basic and translational research in medicine over the last several years, and are in increasing demand in digestive diseases research. Consequently, we have designed the Microbiomics and Functional Genomics Core to provide advanced microbiomics and genomics services to Center members, and offer consultation and training in genomics technologies to enhance the ability of members to implement these technologies in their research. Specifically, the Core will: 1) Conduct sequencing-based assays for microbiomics and functional genomics; 2) Perform bioinformatics analyses of sequencing-based assays; 3) Consult and advise in studies involving microbiomics and functional genomics; and 4) Provide validated experimental protocols and training. The Core leverages several established institutional shared resources, including the Center for Microbiome Innovation at the University of California, San Diego (UCSD), the Genomics Center of the Institute for Genomic Medicine at UCSD, and the UCSD Center for Computational Biology and Bioinformatics. This unique combination and integration of outstanding existing resources will ensure that Center members have access to cutting-edge technologies for studies involving microbiomics and functional genomics. The Core will be directed by internationally recognized experts in microbiomics (Dr. Rob Knight) and functional genomics (Dr. Christopher Glass), and other Core personnel are highly experienced in microbiome science, molecular genetics, bioinformatics and computational biology. The Core has established rigorous procedures for quality control and data validation. Together, this infrastructure and expertise will enable the Microbiomics and Functional Genomics Core to provide a range of high quality and cost-effective microbiomics and functional genomics services that greatly enhance the ability of Center members to advance digestive diseases research.

Project Summary There is a clear imperative to develop potent, cost effective therapeutics to confront the challenge cancer poses to society. Here we address this need by developing synthetically engineered cells effective against a broad range of cancer types with a special emphasis on colorectal cancer (CRC). This cancer type is the second most common cause of cancer death in the US, with more than 50,000 Americans dying every year. Recent research demonstrates the power of genetic engineering to make significant advances towards more efficacious cancer therapy. The introduction of genetically engineered cells, such as chimeric antigen receptor T (CAR T) cells, has shown great promise for treating many types of B cell malignancies, but unfortunately targeting CAR T cells to solid tumors remains challenging. In this project we will use the tools of synthetic biology to make new engineered therapies based on bacterial rather than mammalian cells. Certain bacterial species have demonstrated a useful ability to ?home in? and selectively colonize solid tumors without infecting healthy tissue. This tumor targeting property will be exploited in the proposed work to deliver safe, effective therapies directly to the locations where they are needed most: the solid core of tumors. Previously we developed a bacterial therapeutic and tested it in an animal model of metastatic disease. In contrast to other approaches utilizing bacterial cells, this ?lysis strain? does not require specialized genetic modifications for the secretion of encoded cargo, it simply releases it into the environment when the cells burst. Initially we will genetically modify the lysis strain to produce a wide range of therapeutics for testing, including toxins (from bacteria, animals and plants), enzymes, antibiotics, and apoptotic peptides. Next we will analyze the tumor microbiome from human samples since we hypothesize that the native bacterial population's composition will provide a unique signature (analogous to a fingerprint) that can be used to divide tumors into distinct subtypes. We expect to use these fingerprints to identify other species with superior suitability for therapeutic delivery in treating CRC. Once identified we will develop two in vitro assays for testing the candidate strains. We will use microfluidic technology to create a high throughput co-culturing system for bacteria and a cancer cell line. In parallel, we will develop a co-culturing system for bacteria and organoids that are generated from the same human tumor samples which had been previously used for strain identification and fingerprinting. Lastly we will test the most promising therapies in an animal model of colorectal cancer to determine efficacy in a pre- clinical model.

Project Summary Antimicrobial resistance is an increasing problem, and current drug pipelines are not keeping pace with the rise of antimicrobial resistance. An alternative strategy is to boost host immunity. An often overlooked side-effect of the vitamin and mineral supplementation projects of the 1940s is that these supplements greatly reduced infectious disease burden. Recent work has shown that further gains may be possible, especially in adding phytochemicals back to highly processed diets typically consumed in the United States. However, we lack a fundamental understanding of how these components are processed by the microbiome, and how diet-derived molecules, microbiome and host immune system work together to resist infectious disease. A key barrier preventing us from making these discoveries is that each individual assay (microbiome, host gene expression metabolome, dietary compounds) is expensive and highly multivariate. Three key insights that enable the current project are the miniaturization of DNA and RNA sequencing assays on advanced nanoliter- scale liquid handling robots, greatly reducing the cost, the combination of untargeted and targeted mass spectrometry on the same samples in high throughput to enable discovery of a much greater chemical space, and the ability to use explicitly spatial maps on multiple scales to integrate the dataset throughout the body and enable both visual analytics and deep learning approaches based on spatial data. These breakthroughs will provide a fundamentally new understanding of how dietary metabolites promote disease resistance, and will allow us to develop a new infrastructure to integrate results from many investigators in different laboratories studying various aspects of these systems. Additionally, the results will allow us to choose biomaterials and biomarkers in human subjects that provide maximum information about internal nutritional and immune status. Results will be tested against the NHANES and American Gut cohorts. The results of this project will therefore be: 3D maps of mouse models showing how the microbiome, diet, and host gene expression produce immunity; an infrastructure for creating and sharing these maps; and a preliminary test of whether the results extend to large human populations.

The current COVID-19 pandemic prompts an urgent response including improved bioinformatics tools to help the community analyze relevant datasets from infected patients and their environments. This project will update the popular QIIME pipeline for microbiome analysis, which is very widely used by people studying bacteria and archaea in the microbiome, optimizing it for researchers studying viruses. This will bring features such as data provenance tracking and reproducibility of analysis workflows to the viral research community. Such features enhance the reliability of bioinformatics work in a rapidly paced of research environment such as emergency response to a pandemic, where processing errors are more likely.

The intellectual merit of this work will be to move studies of viral communities from a nonphylogenetic to a phylogenetic basis, accelerate time-to-result, and to make QIIME 2 far more useful to the increasing number of researchers moving from bacterial community analysis to viral community analysis in response to the COVID-19 pandemic. Specific enhancements to be implemented are to build a reference database of viral sequences from diverse genome, metagenome, and metatranscriptome sources; to enhance storage and compute to resolve limitations posed by large-scale datasets generated for SARS-CoV-2; to extend computational pipelines to accommodate the recombination and lack of recognizable common phylogenetic tree roots characteristic of viruses; and to support genome assembly from reads recruiting to viral databases. Results will be disseminated as new QIIME 2 plugins, for broad distribution to the community, including the development of new educational materials and new workshop modules.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.