Integrated annotation prioritizes metabolites with bioactivity in inflammatory bowel disease.

Microbial biochemistry is central to the pathophysiology of inflammatory bowel diseases (IBD). Improved knowledge of microbial metabolites and their immunomodulatory roles is thus necessary for diagnosis and management. Here, we systematically analyzed the chemical, ecological, and epidemiological properties of ~82k metabolic features in 546 Integrative Human Microbiome Project (iHMP/HMP2) metabolomes, using a newly developed methodology for bioactive compound prioritization from microbial communities. This suggested >1000 metabolic features as potentially bioactive in IBD and associated ~43% of prevalent, unannotated features with at least one well-characterized metabolite, thereby providing initial information for further characterization of a significant portion of the fecal metabolome. Prioritized features included known IBD-linked chemical families such as bile acids and short-chain fatty acids, and less-explored bilirubin, polyamine, and vitamin derivatives, and other microbial products. One of these, nicotinamide riboside, reduced colitis scores in DSS-treated mice. The method, MACARRoN, is generalizable with the potential to improve microbial community characterization and provide therapeutic candidates.

International Society for Extracellular Vesicles Workshop. QuantitatEVs: multiscale analyses, from bulk to single extracellular vesicle.

The 'QuantitatEVs: multiscale analyses, from bulk to single vesicle' workshop aimed to discuss quantitative strategies and harmonized wet and computational approaches toward the comprehensive analysis of extracellular vesicles (EVs) from bulk to single vesicle analyses with a special focus on emerging technologies. The workshop covered the key issues in the quantitative analysis of different EV-associated molecular components and EV biophysical features, which are considered the core of EV-associated biomarker discovery and validation for their clinical translation. The in-person-only workshop was held in Trento, Italy, from January 31st to February 2nd, 2023, and continued in Milan on February 3rd with "Next Generation EVs", a satellite event dedicated to early career researchers (ECR). This report summarizes the main topics and outcomes of the workshop.

Longitudinal gut microbiome changes in immune checkpoint blockade-treated advanced melanoma.

Multiple clinical trials targeting the gut microbiome are being conducted to optimize treatment outcomes for immune checkpoint blockade (ICB). To improve the success of these interventions, understanding gut microbiome changes during ICB is urgently needed. Here through longitudinal microbiome profiling of 175 patients treated with ICB for advanced melanoma, we show that several microbial species-level genome bins (SGBs) and pathways exhibit distinct patterns from baseline in patients achieving progression-free survival (PFS) of 12 months or longer (PFS ≥12) versus patients with PFS shorter than 12 months (PFS <12). Out of 99 SGBs that could discriminate between these two groups, 20 were differentially abundant only at baseline, while 42 were differentially abundant only after treatment initiation. We identify five and four SGBs that had consistently higher abundances in patients with PFS ≥12 and <12 months, respectively. Constructing a log ratio of these SGBs, we find an association with overall survival. Finally, we find different microbial dynamics in different clinical contexts including the type of ICB regimen, development of immune-related adverse events and concomitant medication use. Insights into the longitudinal dynamics of the gut microbiome in association with host factors and treatment regimens will be critical for guiding rational microbiome-targeted therapies aimed at enhancing ICB efficacy.

Machine learning for microbiologists.

Machine learning is increasingly important in microbiology where it is used for tasks such as predicting antibiotic resistance and associating human microbiome features with complex host diseases. The applications in microbiology are quickly expanding and the machine learning tools frequently used in basic and clinical research range from classification and regression to clustering and dimensionality reduction. In this Review, we examine the main machine learning concepts, tasks and applications that are relevant for experimental and clinical microbiologists. We provide the minimal toolbox for a microbiologist to be able to understand, interpret and use machine learning in their experimental and translational activities.

BugSigDB captures patterns of differential abundance across a broad range of host-associated microbial signatures.

The literature of human and other host-associated microbiome studies is expanding rapidly, but systematic comparisons among published results of host-associated microbiome signatures of differential abundance remain difficult. We present BugSigDB, a community-editable database of manually curated microbial signatures from published differential abundance studies accompanied by information on study geography, health outcomes, host body site and experimental, epidemiological and statistical methods using controlled vocabulary. The initial release of the database contains >2,500 manually curated signatures from >600 published studies on three host species, enabling high-throughput analysis of signature similarity, taxon enrichment, co-occurrence and coexclusion and consensus signatures. These data allow assessment of microbiome differential abundance within and across experimental conditions, environments or body sites. Database-wide analysis reveals experimental conditions with the highest level of consistency in signatures reported by independent studies and identifies commonalities among disease-associated signatures, including frequent introgression of oral pathobionts into the gut.

Curated single cell multimodal landmark datasets for R/Bioconductor.

BACKGROUND: The majority of high-throughput single-cell molecular profiling methods quantify RNA expression; however, recent multimodal profiling methods add simultaneous measurement of genomic, proteomic, epigenetic, and/or spatial information on the same cells. The development of new statistical and computational methods in Bioconductor for such data will be facilitated by easy availability of landmark datasets using standard data classes. RESULTS: We collected, processed, and packaged publicly available landmark datasets from important single-cell multimodal protocols, including CITE-Seq, ECCITE-Seq, SCoPE2, scNMT, 10X Multiome, seqFISH, and G&T. We integrate data modalities via the MultiAssayExperiment Bioconductor class, document and re-distribute datasets as the SingleCellMultiModal package in Bioconductor's Cloud-based ExperimentHub. The result is single-command actualization of landmark datasets from seven single-cell multimodal data generation technologies, without need for further data processing or wrangling in order to analyze and develop methods within Bioconductor's ecosystem of hundreds of packages for single-cell and multimodal data. CONCLUSIONS: We provide two examples of integrative analyses that are greatly simplified by SingleCellMultiModal. The package will facilitate development of bioinformatic and statistical methods in Bioconductor to meet the challenges of integrating molecular layers and analyzing phenotypic outputs including cell differentiation, activity, and disease.

Evaluation of software impact designed for biomedical research: Are we measuring what's meaningful?

Software is vital for the advancement of biology and medicine. Through analysis of usage and impact metrics of software, developers can help determine user and community engagement. These metrics can be used to justify additional funding, encourage additional use, and identify unanticipated use cases. Such analyses can help define improvement areas and assist with managing project resources. However, there are challenges associated with assessing usage and impact, many of which vary widely depending on the type of software being evaluated. These challenges involve issues of distorted, exaggerated, understated, or misleading metrics, as well as ethical and security concerns. More attention to the nuances, challenges, and considerations involved in capturing impact across the diverse spectrum of biological software is needed. Furthermore, some tools may be especially beneficial to a small audience, yet may not have comparatively compelling metrics of high usage. Although some principles are generally applicable, there is not a single perfect metric or approach to effectively evaluate a software tool's impact, as this depends on aspects unique to each tool, how it is used, and how one wishes to evaluate engagement. We propose more broadly applicable guidelines (such as infrastructure that supports the usage of software and the collection of metrics about usage), as well as strategies for various types of software and resources. We also highlight outstanding issues in the field regarding how communities measure or evaluate software impact. To gain a deeper understanding of the issues hindering software evaluations, as well as to determine what appears to be helpful, we performed a survey of participants involved with scientific software projects for the Informatics Technology for Cancer Research (ITCR) program funded by the National Cancer Institute (NCI). We also investigated software among this scientific community and others to assess how often infrastructure that supports such evaluations is implemented and how this impacts rates of papers describing usage of the software. We find that although developers recognize the utility of analyzing data related to the impact or usage of their software, they struggle to find the time or funding to support such analyses. We also find that infrastructure such as social media presence, more in-depth documentation, the presence of software health metrics, and clear information on how to contact developers seem to be associated with increased usage rates. Our findings can help scientific software developers make the most out of the evaluations of their software so that they can more fully benefit from such assessments.

RaggedExperiment: the missing link between genomic ranges and matrices in Bioconductor.

SUMMARY: The RaggedExperiment R / Bioconductor package provides lossless representation of disparate genomic ranges across multiple specimens or cells, in conjunction with efficient and flexible calculations of rectangular-shaped summaries for downstream analysis. Applications include statistical analysis of somatic mutations, copy number, methylation, and open chromatin data. RaggedExperiment is compatible with multimodal data analysis as a component of MultiAssayExperiment data objects, and simplifies data representation and transformation for software developers and analysts. MOTIVATION AND RESULTS: Measurement of copy number, mutation, single nucleotide polymorphism, and other genomic attributes that may be stored as VCF files produce "ragged" genomic ranges data: i.e. across different genomic coordinates in each sample. Ragged data are not rectangular or matrix-like, presenting informatics challenges for downstream statistical analyses. We present the RaggedExperiment R/Bioconductor data structure for lossless representation of ragged genomic data, with associated reshaping tools for flexible and efficient calculation of tabular representations to support a wide range of downstream statistical analyses. We demonstrate its applicability to copy number and somatic mutation data across 33 TCGA cancer datasets.

Severe Acute Respiratory Syndrome Coronavirus 2 Incidence and Risk Factors in a National, Community-Based Prospective Cohort of US Adults.

BACKGROUND: Prospective cohort studies of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) incidence complement case-based surveillance and cross-sectional seroprevalence surveys. METHODS: We estimated the incidence of SARS-CoV-2 infection in a national cohort of 6738 US adults, enrolled in March-August 2020. Using Poisson models, we examined the association of social distancing and a composite epidemiologic risk score with seroconversion. The risk score was created using least absolute shrinkage selection operator (LASSO) regression to identify factors predictive of seroconversion. The selected factors were household crowding, confirmed case in household, indoor dining, gathering with groups of ≥10, and no masking in gyms or salons. RESULTS: Among 4510 individuals with ≥1 serologic test, 323 (7.3% [95% confidence interval (CI), 6.5%-8.1%]) seroconverted by January 2021. Among 3422 participants seronegative in May-September 2020 and retested from November 2020 to January 2021, 161 seroconverted over 1646 person-years of follow-up (9.8 per 100 person-years [95% CI, 8.3-11.5]). The seroincidence rate was lower among women compared with men (incidence rate ratio [IRR], 0.69 [95% CI, .50-.94]) and higher among Hispanic (2.09 [1.41-3.05]) than white non-Hispanic participants. In adjusted models, participants who reported social distancing with people they did not know (IRR for always vs never social distancing, 0.42 [95% CI, .20-1.0]) and with people they knew (IRR for always vs never, 0.64 [.39-1.06]; IRR for sometimes vs never, 0.60 [.38-.96]) had lower seroconversion risk. Seroconversion risk increased with epidemiologic risk score (IRR for medium vs low score, 1.68 [95% CI, 1.03-2.81]; IRR for high vs low score, 3.49 [2.26-5.58]). Only 29% of those who seroconverted reported isolating, and only 19% were asked about contacts. CONCLUSIONS: Modifiable risk factors and poor reach of public health strategies drove SARS-CoV-2 transmission across the United States.

Household factors and the risk of severe COVID-like illness early in the U.S. pandemic.

OBJECTIVE: To investigate the role of children in the home and household crowding as risk factors for severe COVID-19 disease. METHODS: We used interview data from 6,831 U.S. adults screened for the Communities, Households and SARS/CoV-2 Epidemiology (CHASING) COVID Cohort Study in April 2020. RESULTS: In logistic regression models, the adjusted odds ratio [aOR] of hospitalization due to COVID-19 for having (versus not having) children in the home was 10.5 (95% CI:5.7-19.1) among study participants living in multi-unit dwellings and 2.2 (95% CI:1.2-6.5) among those living in single unit dwellings. Among participants living in multi-unit dwellings, the aOR for COVID-19 hospitalization among participants with more than 4 persons in their household (versus 1 person) was 2.5 (95% CI:1.0-6.1), and 0.8 (95% CI:0.15-4.1) among those living in single unit dwellings. CONCLUSION: Early in the US SARS-CoV-2 pandemic, certain household exposures likely increased the risk of both SARS-CoV-2 acquisition and the risk of severe COVID-19 disease.

High-sensitivity pattern discovery in large, paired multiomic datasets.

MOTIVATION: Modern biological screens yield enormous numbers of measurements, and identifying and interpreting statistically significant associations among features are essential. In experiments featuring multiple high-dimensional datasets collected from the same set of samples, it is useful to identify groups of associated features between the datasets in a way that provides high statistical power and false discovery rate (FDR) control. RESULTS: Here, we present a novel hierarchical framework, HAllA (Hierarchical All-against-All association testing), for structured association discovery between paired high-dimensional datasets. HAllA efficiently integrates hierarchical hypothesis testing with FDR correction to reveal significant linear and non-linear block-wise relationships among continuous and/or categorical data. We optimized and evaluated HAllA using heterogeneous synthetic datasets of known association structure, where HAllA outperformed all-against-all and other block-testing approaches across a range of common similarity measures. We then applied HAllA to a series of real-world multiomics datasets, revealing new associations between gene expression and host immune activity, the microbiome and host transcriptome, metabolomic profiling and human health phenotypes. AVAILABILITY AND IMPLEMENTATION: An open-source implementation of HAllA is freely available at http://huttenhower.sph.harvard.edu/halla along with documentation, demo datasets and a user group. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

GenomicSuperSignature facilitates interpretation of RNA-seq experiments through robust, efficient comparison to public databases.

Millions of transcriptomic profiles have been deposited in public archives, yet remain underused for the interpretation of new experiments. We present a method for interpreting new transcriptomic datasets through instant comparison to public datasets without high-performance computing requirements. We apply Principal Component Analysis on 536 studies comprising 44,890 human RNA sequencing profiles and aggregate sufficiently similar loading vectors to form Replicable Axes of Variation (RAV). RAVs are annotated with metadata of originating studies and by gene set enrichment analysis. Functionality to associate new datasets with RAVs, extract interpretable annotations, and provide intuitive visualization are implemented as the GenomicSuperSignature R/Bioconductor package. We demonstrate the efficient and coherent database search, robustness to batch effects and heterogeneous training data, and transfer learning capacity of our method using TCGA and rare diseases datasets. GenomicSuperSignature aids in analyzing new gene expression data in the context of existing databases using minimal computing resources.

Refining colorectal cancer classification and clinical stratification through a single-cell atlas.

BACKGROUND: Colorectal cancer (CRC) consensus molecular subtypes (CMS) have different immunological, stromal cell, and clinicopathological characteristics. Single-cell characterization of CMS subtype tumor microenvironments is required to elucidate mechanisms of tumor and stroma cell contributions to pathogenesis which may advance subtype-specific therapeutic development. We interrogate racially diverse human CRC samples and analyze multiple independent external cohorts for a total of 487,829 single cells enabling high-resolution depiction of the cellular diversity and heterogeneity within the tumor and microenvironmental cells. RESULTS: Tumor cells recapitulate individual CMS subgroups yet exhibit significant intratumoral CMS heterogeneity. Both CMS1 microsatellite instability (MSI-H) CRCs and microsatellite stable (MSS) CRC demonstrate similar pathway activations at the tumor epithelial level. However, CD8+ cytotoxic T cell phenotype infiltration in MSI-H CRCs may explain why these tumors respond to immune checkpoint inhibitors. Cellular transcriptomic profiles in CRC exist in a tumor immune stromal continuum in contrast to discrete subtypes proposed by studies utilizing bulk transcriptomics. We note a dichotomy in tumor microenvironments across CMS subgroups exists by which patients with high cancer-associated fibroblasts (CAFs) and C1Q+TAM content exhibit poor outcomes, providing a higher level of personalization and precision than would distinct subtypes. Additionally, we discover CAF subtypes known to be associated with immunotherapy resistance. CONCLUSIONS: Distinct CAFs and C1Q+ TAMs are sufficient to explain CMS predictive ability and a simpler signature based on these cellular phenotypes could stratify CRC patient prognosis with greater precision. Therapeutically targeting specific CAF subtypes and C1Q + TAMs may promote immunotherapy responses in CRC patients.

Cross-cohort gut microbiome associations with immune checkpoint inhibitor response in advanced melanoma.

The composition of the gut microbiome has been associated with clinical responses to immune checkpoint inhibitor (ICI) treatment, but there is limited consensus on the specific microbiome characteristics linked to the clinical benefits of ICIs. We performed shotgun metagenomic sequencing of stool samples collected before ICI initiation from five observational cohorts recruiting ICI-naive patients with advanced cutaneous melanoma (n = 165). Integrating the dataset with 147 metagenomic samples from previously published studies, we found that the gut microbiome has a relevant, but cohort-dependent, association with the response to ICIs. A machine learning analysis confirmed the link between the microbiome and overall response rates (ORRs) and progression-free survival (PFS) with ICIs but also revealed limited reproducibility of microbiome-based signatures across cohorts. Accordingly, a panel of species, including Bifidobacterium pseudocatenulatum, Roseburia spp. and Akkermansia muciniphila, associated with responders was identified, but no single species could be regarded as a fully consistent biomarker across studies. Overall, the role of the human gut microbiome in ICI response appears more complex than previously thought, extending beyond differing microbial species simply present or absent in responders and nonresponders. Future studies should adopt larger sample sizes and take into account the complex interplay of clinical factors with the gut microbiome over the treatment course.

Inverting the model of genomics data sharing with the NHGRI Genomic Data Science Analysis, Visualization, and Informatics Lab-space.

The NHGRI Genomic Data Science Analysis, Visualization, and Informatics Lab-space (AnVIL; https://anvilproject.org) was developed to address a widespread community need for a unified computing environment for genomics data storage, management, and analysis. In this perspective, we present AnVIL, describe its ecosystem and interoperability with other platforms, and highlight how this platform and associated initiatives contribute to improved genomic data sharing efforts. The AnVIL is a federated cloud platform designed to manage and store genomics and related data, enable population-scale analysis, and facilitate collaboration through the sharing of data, code, and analysis results. By inverting the traditional model of data sharing, the AnVIL eliminates the need for data movement while also adding security measures for active threat detection and monitoring and provides scalable, shared computing resources for any researcher. We describe the core data management and analysis components of the AnVIL, which currently consists of Terra, Gen3, Galaxy, RStudio/Bioconductor, Dockstore, and Jupyter, and describe several flagship genomics datasets available within the AnVIL. We continue to extend and innovate the AnVIL ecosystem by implementing new capabilities, including mechanisms for interoperability and responsible data sharing, while streamlining access management. The AnVIL opens many new opportunities for analysis, collaboration, and data sharing that are needed to drive research and to make discoveries through the joint analysis of hundreds of thousands to millions of genomes along with associated clinical and molecular data types.

Patterns of SARS-CoV-2 Testing Preferences in a National Cohort in the United States: Latent Class Analysis of a Discrete Choice Experiment.

BACKGROUND: Inadequate screening and diagnostic testing in the United States throughout the first several months of the COVID-19 pandemic led to undetected cases transmitting disease in the community and an underestimation of cases. Though testing supply has increased, maintaining testing uptake remains a public health priority in the efforts to control community transmission considering the availability of vaccinations and threats from variants. OBJECTIVE: This study aimed to identify patterns of preferences for SARS-CoV-2 screening and diagnostic testing prior to widespread vaccine availability and uptake. METHODS: We conducted a discrete choice experiment (DCE) among participants in the national, prospective CHASING COVID (Communities, Households, and SARS-CoV-2 Epidemiology) Cohort Study from July 30 to September 8, 2020. The DCE elicited preferences for SARS-CoV-2 test type, specimen type, testing venue, and result turnaround time. We used latent class multinomial logit to identify distinct patterns of preferences related to testing as measured by attribute-level part-worth utilities and conducted a simulation based on the utility estimates to predict testing uptake if additional testing scenarios were offered. RESULTS: Of the 5098 invited cohort participants, 4793 (94.0%) completed the DCE. Five distinct patterns of SARS-CoV-2 testing emerged. Noninvasive home testers (n=920, 19.2% of participants) were most influenced by specimen type and favored less invasive specimen collection methods, with saliva being most preferred; this group was the least likely to opt out of testing. Fast-track testers (n=1235, 25.8%) were most influenced by result turnaround time and favored immediate and same-day turnaround time. Among dual testers (n=889, 18.5%), test type was the most important attribute, and preference was given to both antibody and viral tests. Noninvasive dual testers (n=1578, 32.9%) were most strongly influenced by specimen type and test type, preferring saliva and cheek swab specimens and both antibody and viral tests. Among hesitant home testers (n=171, 3.6%), the venue was the most important attribute; notably, this group was the most likely to opt out of testing. In addition to variability in preferences for testing features, heterogeneity was observed in the distribution of certain demographic characteristics (age, race/ethnicity, education, and employment), history of SARS-CoV-2 testing, COVID-19 diagnosis, and concern about the pandemic. Simulation models predicted that testing uptake would increase from 81.6% (with a status quo scenario of polymerase chain reaction by nasal swab in a provider's office and a turnaround time of several days) to 98.1% by offering additional scenarios using less invasive specimens, both viral and antibody tests from a single specimen, faster turnaround time, and at-home testing. CONCLUSIONS: We identified substantial differences in preferences for SARS-CoV-2 testing and found that offering additional testing options would likely increase testing uptake in line with public health goals. Additional studies may be warranted to understand if preferences for testing have changed since the availability and widespread uptake of vaccines.

Multivariable association discovery in population-scale meta-omics studies.

It is challenging to associate features such as human health outcomes, diet, environmental conditions, or other metadata to microbial community measurements, due in part to their quantitative properties. Microbiome multi-omics are typically noisy, sparse (zero-inflated), high-dimensional, extremely non-normal, and often in the form of count or compositional measurements. Here we introduce an optimized combination of novel and established methodology to assess multivariable association of microbial community features with complex metadata in population-scale observational studies. Our approach, MaAsLin 2 (Microbiome Multivariable Associations with Linear Models), uses generalized linear and mixed models to accommodate a wide variety of modern epidemiological studies, including cross-sectional and longitudinal designs, as well as a variety of data types (e.g., counts and relative abundances) with or without covariates and repeated measurements. To construct this method, we conducted a large-scale evaluation of a broad range of scenarios under which straightforward identification of meta-omics associations can be challenging. These simulation studies reveal that MaAsLin 2's linear model preserves statistical power in the presence of repeated measures and multiple covariates, while accounting for the nuances of meta-omics features and controlling false discovery. We also applied MaAsLin 2 to a microbial multi-omics dataset from the Integrative Human Microbiome (HMP2) project which, in addition to reproducing established results, revealed a unique, integrated landscape of inflammatory bowel diseases (IBD) across multiple time points and omics profiles.

Cohort profile: a national, community-based prospective cohort study of SARS-CoV-2 pandemic outcomes in the USA-the CHASING COVID Cohort study.

PURPOSE: The Communities, Households and SARS-CoV-2 Epidemiology (CHASING) COVID Cohort Study is a community-based prospective cohort study launched during the upswing of the USA COVID-19 epidemic. The objectives of the cohort study are to: (1) estimate and evaluate determinants of the incidence of SARS-CoV-2 infection, disease and deaths; (2) assess the impact of the pandemic on psychosocial and economic outcomes and (3) assess the uptake of pandemic mitigation strategies. PARTICIPANTS: We began enrolling participants from 28 March 2020 using internet-based strategies. Adults≥18 years residing anywhere in the USA or US territories were eligible. 6740 people are enrolled in the cohort, including participants from all 50 US states, the District of Columbia, Puerto Rico and Guam. Participants are contacted regularly to complete study assessments, including interviews and dried blood spot specimen collection for serologic testing. FINDINGS TO DATE: Participants are geographically and sociodemographically diverse and include essential workers (19%). 84.2% remain engaged in cohort follow-up activities after enrolment. Data have been used to assess SARS-CoV-2 cumulative incidence, seroincidence and related risk factors at different phases of the US pandemic; the role of household crowding and the presence of children in the household as potential risk factors for severe COVID-19 early in the US pandemic; to describe the prevalence of anxiety symptoms and its relationship to COVID-19 outcomes and other potential stressors; to identify preferences for SARS-CoV-2 diagnostic testing when community transmission is on the rise via a discrete choice experiment and to assess vaccine hesitancy over time and its relationship to vaccine uptake. FUTURE PLANS: The CHASING COVID Cohort Study has outlined a research agenda that involves ongoing monitoring of the incidence and determinants of SARS-CoV-2 outcomes, mental health outcomes and economic outcomes. Additional priorities include assessing the incidence, prevalence and correlates of long-haul COVID-19.

SARS-CoV-2 incidence and risk factors in a national, community-based prospective cohort of U.S. adults.

BACKGROUND: Epidemiologic risk factors for incident SARS-CoV-2 infection as determined via prospective cohort studies greatly augment and complement information from case-based surveillance and cross-sectional seroprevalence surveys. METHODS: We estimated the incidence of SARS-CoV-2 infection and risk factors in a well-characterized, national prospective cohort of 6,738 U.S. adults, enrolled March-August 2020, a subset of whom (n=4,510) underwent repeat serologic testing between May 2020 and January 2021. We examined the crude associations of sociodemographic factors, epidemiologic risk factors, and county-level community transmission with the incidence of seroconversion. In multivariable Poisson models we examined the association of social distancing and a composite score of several epidemiologic risk factors with the rate of seroconversion. FINDINGS: Among the 4,510 individuals with at least one serologic test, 323 (7.3%, 95% confidence interval [CI] 6.5%-8.1%) seroconverted by January 2021. Among 3,422 participants seronegative in May-September 2020 and tested during November 2020-January 2021, we observed 161 seroconversions over 1,646 person-years of follow-up (incidence rate of 9.8 per 100 person-years [95%CI 8.3-11.5]). In adjusted models, participants who reported always or sometimes social distancing with people they knew (IRRalways vs. never 0.43, 95%CI 0.21-1.0; IRRsometimes vs. never 0.47, 95%CI 0.22-1.2) and people they did not know (IRRalways vs. never 0.64, 95%CI 0.39-1.1; IRRsometimes vs. never 0.60, 95%CI 0.38-0.97) had lower rates of seroconversion. The rate of seroconversion increased across tertiles of the composite score of epidemiologic risk (IRRmedium vs. low 1.5, 95%CI 0.92-2.4; IRRhigh vs. low 3.0, 95%CI 2.0-4.6). Among the 161 observed seroconversions, 28% reported no symptoms of COVID-like illness (i.e., were asymptomatic), and 27% reported a positive SARS-CoV-2 diagnostic test. Ultimately, only 29% reported isolating and 19% were asked about contacts. INTERPRETATION: Modifiable epidemiologic risk factors and poor reach of public health strategies drove SARS-CoV-2 transmission across the U.S during May 2020-January 2021. FUNDING: U.S. National Institutes of Allergy and Infectious Diseases (NIAID).