regionalpcs : improved discovery of DNA methylation associations with complex traits.

We have developed the regional principal components (rPCs) method, a novel approach for summarizing gene-level methylation. rPCs address the challenge of deciphering complex epigenetic mechanisms in diseases like Alzheimer's disease (AD). In contrast to traditional averaging, rPCs leverage principal components analysis to capture complex methylation patterns across gene regions. Our method demonstrated a 54% improvement in sensitivity over averaging in simulations, offering a robust framework for identifying subtle epigenetic variations. Applying rPCs to the AD brain methylation data in ROSMAP, combined with cell type deconvolution, we uncovered 838 differentially methylated genes associated with neuritic plaque burden-significantly outperforming conventional methods. Integrating methylation quantitative trait loci (meQTL) with genome-wide association studies (GWAS) identified 17 genes with potential causal roles in AD, including MS4A4A and PI-CALM . Our approach is available in the Bioconductor package regionalpcs , opening avenues for research and facilitating a deeper understanding of the epigenetic landscape in complex diseases.

Intraoperative Evaluation of Breast Tissues During Breast Cancer Operations Using the MasSpec Pen.

Importance: Surgery with complete tumor resection remains the main treatment option for patients with breast cancer. Yet, current technologies are limited in providing accurate assessment of breast tissue in vivo, warranting development of new technologies for surgical guidance. Objective: To evaluate the performance of the MasSpec Pen for accurate intraoperative assessment of breast tissues and surgical margins based on metabolic and lipid information. Design, Setting, and Participants: In this diagnostic study conducted between February 23, 2017, and August 19, 2021, the mass spectrometry-based device was used to analyze healthy breast and invasive ductal carcinoma (IDC) banked tissue samples from adult patients undergoing breast surgery for ductal carcinomas or nonmalignant conditions. Fresh-frozen tissue samples and touch imprints were analyzed in a laboratory. Intraoperative in vivo and ex vivo breast tissue analyses were performed by surgical staff in operating rooms (ORs) within 2 different hospitals at the Texas Medical Center. Molecular data were used to build statistical classifiers. Main Outcomes and Measures: Prediction results of tissue analyses from classification models were compared with gross assessment, frozen section analysis, and/or final postoperative pathology to assess accuracy. Results: All data acquired from the 143 banked tissue samples, including 79 healthy breast and 64 IDC tissues, were included in the statistical analysis. Data presented rich molecular profiles of healthy and IDC banked tissue samples, with significant changes in relative abundances observed for several metabolic species. Statistical classifiers yielded accuracies of 95.6%, 95.5%, and 90.6% for training, validation, and independent test sets, respectively. A total of 25 participants enrolled in the clinical, intraoperative study; all were female, and the median age was 58 years (IQR, 44-66 years). Intraoperative testing of the technology was successfully performed by surgical staff during 25 breast operations. Of 273 intraoperative analyses performed during 25 surgical cases, 147 analyses from 22 cases were subjected to statistical classification. Testing of the classifiers on 147 intraoperative mass spectra yielded 95.9% agreement with postoperative pathology results. Conclusions and Relevance: The findings of this diagnostic study suggest that the mass spectrometry-based system could be clinically valuable to surgeons and patients by enabling fast molecular-based intraoperative assessment of in vivo and ex vivo breast tissue samples and surgical margins.

Public health factors help explain cross country heterogeneity in excess death during the COVID19 pandemic.

The COVID-19 pandemic has taken a devastating toll around the world. Since January 2020, the World Health Organization estimates 14.9 million excess deaths have occurred globally. Despite this grim number quantifying the deadly impact, the underlying factors contributing to COVID-19 deaths at the population level remain unclear. Prior studies indicate that demographic factors like proportion of population older than 65 and population health explain the cross-country difference in COVID-19 deaths. However, there has not been a comprehensive analysis including variables describing government policies and COVID-19 vaccination rate. Furthermore, prior studies focus on COVID-19 death rather than excess death to assess the impact of the pandemic. Through a robust statistical modeling framework, we analyze 80 countries and show that actionable public health efforts beyond just the factors intrinsic to each country are important for explaining the cross-country heterogeneity in excess death.

Confidence intervals for the Cox model test error from cross-validation.

Cross-validation (CV) is one of the most widely used techniques in statistical learning for estimating the test error of a model, but its behavior is not yet fully understood. It has been shown that standard confidence intervals for test error using estimates from CV may have coverage below nominal levels. This phenomenon occurs because each sample is used in both the training and testing procedures during CV and as a result, the CV estimates of the errors become correlated. Without accounting for this correlation, the estimate of the variance is smaller than it should be. One way to mitigate this issue is by estimating the mean squared error of the prediction error instead using nested CV. This approach has been shown to achieve superior coverage compared to intervals derived from standard CV. In this work, we generalize the nested CV idea to the Cox proportional hazards model and explore various choices of test error for this setting.

Improved Digital Therapy for Developmental Pediatrics Using Domain-Specific Artificial Intelligence: Machine Learning Study.

BACKGROUND: Automated emotion classification could aid those who struggle to recognize emotions, including children with developmental behavioral conditions such as autism. However, most computer vision emotion recognition models are trained on adult emotion and therefore underperform when applied to child faces. OBJECTIVE: We designed a strategy to gamify the collection and labeling of child emotion-enriched images to boost the performance of automatic child emotion recognition models to a level closer to what will be needed for digital health care approaches. METHODS: We leveraged our prototype therapeutic smartphone game, GuessWhat, which was designed in large part for children with developmental and behavioral conditions, to gamify the secure collection of video data of children expressing a variety of emotions prompted by the game. Independently, we created a secure web interface to gamify the human labeling effort, called HollywoodSquares, tailored for use by any qualified labeler. We gathered and labeled 2155 videos, 39,968 emotion frames, and 106,001 labels on all images. With this drastically expanded pediatric emotion-centric database (>30 times larger than existing public pediatric emotion data sets), we trained a convolutional neural network (CNN) computer vision classifier of happy, sad, surprised, fearful, angry, disgust, and neutral expressions evoked by children. RESULTS: The classifier achieved a 66.9% balanced accuracy and 67.4% F1-score on the entirety of the Child Affective Facial Expression (CAFE) as well as a 79.1% balanced accuracy and 78% F1-score on CAFE Subset A, a subset containing at least 60% human agreement on emotions labels. This performance is at least 10% higher than all previously developed classifiers evaluated against CAFE, the best of which reached a 56% balanced accuracy even when combining "anger" and "disgust" into a single class. CONCLUSIONS: This work validates that mobile games designed for pediatric therapies can generate high volumes of domain-relevant data sets to train state-of-the-art classifiers to perform tasks helpful to precision health efforts.

Identification of Social Engagement Indicators Associated With Autism Spectrum Disorder Using a Game-Based Mobile App: Comparative Study of Gaze Fixation and Visual Scanning Methods.

BACKGROUND: Autism spectrum disorder (ASD) is a widespread neurodevelopmental condition with a range of potential causes and symptoms. Standard diagnostic mechanisms for ASD, which involve lengthy parent questionnaires and clinical observation, often result in long waiting times for results. Recent advances in computer vision and mobile technology hold potential for speeding up the diagnostic process by enabling computational analysis of behavioral and social impairments from home videos. Such techniques can improve objectivity and contribute quantitatively to the diagnostic process. OBJECTIVE: In this work, we evaluate whether home videos collected from a game-based mobile app can be used to provide diagnostic insights into ASD. To the best of our knowledge, this is the first study attempting to identify potential social indicators of ASD from mobile phone videos without the use of eye-tracking hardware, manual annotations, and structured scenarios or clinical environments. METHODS: Here, we used a mobile health app to collect over 11 hours of video footage depicting 95 children engaged in gameplay in a natural home environment. We used automated data set annotations to analyze two social indicators that have previously been shown to differ between children with ASD and their neurotypical (NT) peers: (1) gaze fixation patterns, which represent regions of an individual's visual focus and (2) visual scanning methods, which refer to the ways in which individuals scan their surrounding environment. We compared the gaze fixation and visual scanning methods used by children during a 90-second gameplay video to identify statistically significant differences between the 2 cohorts; we then trained a long short-term memory (LSTM) neural network to determine if gaze indicators could be predictive of ASD. RESULTS: Our results show that gaze fixation patterns differ between the 2 cohorts; specifically, we could identify 1 statistically significant region of fixation (P<.001). In addition, we also demonstrate that there are unique visual scanning patterns that exist for individuals with ASD when compared to NT children (P<.001). A deep learning model trained on coarse gaze fixation annotations demonstrates mild predictive power in identifying ASD. CONCLUSIONS: Ultimately, our study demonstrates that heterogeneous video data sets collected from mobile devices hold potential for quantifying visual patterns and providing insights into ASD. We show the importance of automated labeling techniques in generating large-scale data sets while simultaneously preserving the privacy of participants, and we demonstrate that specific social engagement indicators associated with ASD can be identified and characterized using such data.

A maximum flow-based network approach for identification of stable noncoding biomarkers associated with the multigenic neurological condition, autism.

BACKGROUND: Machine learning approaches for predicting disease risk from high-dimensional whole genome sequence (WGS) data often result in unstable models that can be difficult to interpret, limiting the identification of putative sets of biomarkers. Here, we design and validate a graph-based methodology based on maximum flow, which leverages the presence of linkage disequilibrium (LD) to identify stable sets of variants associated with complex multigenic disorders. RESULTS: We apply our method to a previously published logistic regression model trained to identify variants in simple repeat sequences associated with autism spectrum disorder (ASD); this L1-regularized model exhibits high predictive accuracy yet demonstrates great variability in the features selected from over 230,000 possible variants. In order to improve model stability, we extract the variants assigned non-zero weights in each of 5 cross-validation folds and then assemble the five sets of features into a flow network subject to LD constraints. The maximum flow formulation allowed us to identify 55 variants, which we show to be more stable than the features identified by the original classifier. CONCLUSION: Our method allows for the creation of machine learning models that can identify predictive variants. Our results help pave the way towards biomarker-based diagnosis methods for complex genetic disorders.

Estimating sequencing error rates using families.

BACKGROUND: As next-generation sequencing technologies make their way into the clinic, knowledge of their error rates is essential if they are to be used to guide patient care. However, sequencing platforms and variant-calling pipelines are continuously evolving, making it difficult to accurately quantify error rates for the particular combination of assay and software parameters used on each sample. Family data provide a unique opportunity for estimating sequencing error rates since it allows us to observe a fraction of sequencing errors as Mendelian errors in the family, which we can then use to produce genome-wide error estimates for each sample. RESULTS: We introduce a method that uses Mendelian errors in sequencing data to make highly granular per-sample estimates of precision and recall for any set of variant calls, regardless of sequencing platform or calling methodology. We validate the accuracy of our estimates using monozygotic twins, and we use a set of monozygotic quadruplets to show that our predictions closely match the consensus method. We demonstrate our method's versatility by estimating sequencing error rates for whole genome sequencing, whole exome sequencing, and microarray datasets, and we highlight its sensitivity by quantifying performance increases between different versions of the GATK variant-calling pipeline. We then use our method to demonstrate that: 1) Sequencing error rates between samples in the same dataset can vary by over an order of magnitude. 2) Variant calling performance decreases substantially in low-complexity regions of the genome. 3) Variant calling performance in whole exome sequencing data decreases with distance from the nearest target region. 4) Variant calls from lymphoblastoid cell lines can be as accurate as those from whole blood. 5) Whole-genome sequencing can attain microarray-level precision and recall at disease-associated SNV sites. CONCLUSION: Genotype datasets from families are powerful resources that can be used to make fine-grained estimates of sequencing error for any sequencing platform and variant-calling methodology.

Indels in SARS-CoV-2 occur at template-switching hotspots.

The evolutionary dynamics of SARS-CoV-2 have been carefully monitored since the COVID-19 pandemic began in December 2019. However, analysis has focused primarily on single nucleotide polymorphisms and largely ignored the role of insertions and deletions (indels) as well as recombination in SARS-CoV-2 evolution. Using sequences from the GISAID database, we catalogue over 100 insertions and deletions in the SARS-CoV-2 consensus sequences. We hypothesize that these indels are artifacts of recombination events between SARS-CoV-2 replicates whereby RNA-dependent RNA polymerase (RdRp) re-associates with a homologous template at a different loci ("imperfect homologous recombination"). We provide several independent pieces of evidence that suggest this. (1) The indels from the GISAID consensus sequences are clustered at specific regions of the genome. (2) These regions are also enriched for 5' and 3' breakpoints in the transcription regulatory site (TRS) independent transcriptome, presumably sites of RNA-dependent RNA polymerase (RdRp) template-switching. (3) Within raw reads, these indel hotspots have cases of both high intra-host heterogeneity and intra-host homogeneity, suggesting that these indels are both consequences of de novo recombination events within a host and artifacts of previous recombination. We briefly analyze the indels in the context of RNA secondary structure, noting that indels preferentially occur in "arms" and loop structures of the predicted folded RNA, suggesting that secondary structure may be a mechanism for TRS-independent template-switching in SARS-CoV-2 or other coronaviruses. These insights into the relationship between structural variation and recombination in SARS-CoV-2 can improve our reconstructions of the SARS-CoV-2 evolutionary history as well as our understanding of the process of RdRp template-switching in RNA viruses.

Selection of trustworthy crowd workers for telemedical diagnosis of pediatric autism spectrum disorder.

Crowd-powered telemedicine has the potential to revolutionize healthcare, especially during times that require remote access to care. However, sharing private health data with strangers from around the world is not compatible with data privacy standards, requiring a stringent filtration process to recruit reliable and trustworthy workers who can go through the proper training and security steps. The key challenge, then, is to identify capable, trustworthy, and reliable workers through high-fidelity evaluation tasks without exposing any sensitive patient data during the evaluation process. We contribute a set of experimentally validated metrics for assessing the trustworthiness and reliability of crowd workers tasked with providing behavioral feature tags to unstructured videos of children with autism and matched neurotypical controls. The workers are blinded to diagnosis and blinded to the goal of using the features to diagnose autism. These behavioral labels are fed as input to a previously validated binary logistic regression classifier for detecting autism cases using categorical feature vectors. While the metrics do not incorporate any ground truth labels of child diagnosis, linear regression using the 3 correlative metrics as input can predict the mean probability of the correct class of each worker with a mean average error of 7.51% for performance on the same set of videos and 10.93% for performance on a distinct balanced video set with different children. These results indicate that crowd workers can be recruited for performance based largely on behavioral metrics on a crowdsourced task, enabling an affordable way to filter crowd workforces into a trustworthy and reliable diagnostic workforce.

Game theoretic centrality: a novel approach to prioritize disease candidate genes by combining biological networks with the Shapley value.

BACKGROUND: Complex human health conditions with etiological heterogeneity like Autism Spectrum Disorder (ASD) often pose a challenge for traditional genome-wide association study approaches in defining a clear genotype to phenotype model. Coalitional game theory (CGT) is an exciting method that can consider the combinatorial effect of groups of variants working in concert to produce a phenotype. CGT has been applied to associate likely-gene-disrupting variants encoded from whole genome sequence data to ASD; however, this previous approach cannot take into account for prior biological knowledge. Here we extend CGT to incorporate a priori knowledge from biological networks through a game theoretic centrality measure based on Shapley value to rank genes by their relevance-the individual gene's synergistic influence in a gene-to-gene interaction network. Game theoretic centrality extends the notion of Shapley value to the evaluation of a gene's contribution to the overall connectivity of its corresponding node in a biological network. RESULTS: We implemented and applied game theoretic centrality to rank genes on whole genomes from 756 multiplex autism families. Top ranking genes with the highest game theoretic centrality in both the weighted and unweighted approaches were enriched for pathways previously associated with autism, including pathways of the immune system. Four of the selected genes HLA-A, HLA-B, HLA-G, and HLA-DRB1-have also been implicated in ASD and further support the link between ASD and the human leukocyte antigen complex. CONCLUSIONS: Game theoretic centrality can prioritize influential, disease-associated genes within biological networks, and assist in the decoding of polygenic associations to complex disorders like autism.

Precision Telemedicine through Crowdsourced Machine Learning: Testing Variability of Crowd Workers for Video-Based Autism Feature Recognition.

Mobilized telemedicine is becoming a key, and even necessary, facet of both precision health and precision medicine. In this study, we evaluate the capability and potential of a crowd of virtual workers-defined as vetted members of popular crowdsourcing platforms-to aid in the task of diagnosing autism. We evaluate workers when crowdsourcing the task of providing categorical ordinal behavioral ratings to unstructured public YouTube videos of children with autism and neurotypical controls. To evaluate emerging patterns that are consistent across independent crowds, we target workers from distinct geographic loci on two crowdsourcing platforms: an international group of workers on Amazon Mechanical Turk (MTurk) (N = 15) and Microworkers from Bangladesh (N = 56), Kenya (N = 23), and the Philippines (N = 25). We feed worker responses as input to a validated diagnostic machine learning classifier trained on clinician-filled electronic health records. We find that regardless of crowd platform or targeted country, workers vary in the average confidence of the correct diagnosis predicted by the classifier. The best worker responses produce a mean probability of the correct class above 80% and over one standard deviation above 50%, accuracy and variability on par with experts according to prior studies. There is a weak correlation between mean time spent on task and mean performance (r = 0.358, p = 0.005). These results demonstrate that while the crowd can produce accurate diagnoses, there are intrinsic differences in crowdworker ability to rate behavioral features. We propose a novel strategy for recruitment of crowdsourced workers to ensure high quality diagnostic evaluations of autism, and potentially many other pediatric behavioral health conditions. Our approach represents a viable step in the direction of crowd-based approaches for more scalable and affordable precision medicine.

Coalitional Game Theory Facilitates Identification of Non-Coding Variants Associated With Autism.

Studies on autism spectrum disorder (ASD) have amassed substantial evidence for the role of genetics in the disease's phenotypic manifestation. A large number of coding and non-coding variants with low penetrance likely act in a combinatorial manner to explain the variable forms of ASD. However, many of these combined interactions, both additive and epistatic, remain undefined. Coalitional game theory (CGT) is an approach that seeks to identify players (individual genetic variants or genes) who tend to improve the performance-association to a disease phenotype of interest-of any coalition (subset of co-occurring genetic variants) they join. This method has been previously applied to boost biologically informative signal from gene expression data and exome sequencing data but remains to be explored in the context of cooperativity among non-coding genomic regions. We describe our extension of previous work, highlighting non-coding chromosomal regions relevant to ASD using CGT on alteration data of 4595 fully sequenced genomes from 756 multiplex families. Genomes were encoded into binary matrices for three types of non-coding regions previously implicated in ASD and separated into ASD (case) and unaffected (control) samples. A player metric, the Shapley value, enabled determination of individual variant contributions in both sets of cohorts. A total of 30 non-coding positions were found to have significantly elevated player scores and likely represent significant contributors to the genetic coordination underlying ASD. Cross-study analyses revealed that a subset of mutated non-coding regions (all of which are in human accelerated regions (HARs)) and related genes are involved in biological pathways or behavioral outcomes known to be affected in autism, suggesting the importance of single nucleotide polymorphisms (SNPs) within HARs in ASD. These findings support the use of CGT in identifying hidden yet influential non-coding players from large-scale genomic data, to better understand the precise underpinnings of complex neurodevelopmental disorders such as autism.

Coalitional game theory as a promising approach to identify candidate autism genes.

Despite mounting evidence for the strong role of genetics in the phenotypic manifestation of Autism Spectrum Disorder (ASD), the specific genes responsible for the variable forms of ASD remain undefined. ASD may be best explained by a combinatorial genetic model with varying epistatic interactions across many small effect mutations. Coalitional or cooperative game theory is a technique that studies the combined effects of groups of players, known as coalitions, seeking to identify players who tend to improve the performance--the relationship to a specific disease phenotype--of any coalition they join. This method has been previously shown to boost biologically informative signal in gene expression data but to-date has not been applied to the search for cooperative mutations among putative ASD genes. We describe our approach to highlight genes relevant to ASD using coalitional game theory on alteration data of 1,965 fully sequenced genomes from 756 multiplex families. Alterations were encoded into binary matrices for ASD (case) and unaffected (control) samples, indicating likely gene-disrupting, inherited mutations in altered genes. To determine individual gene contributions given an ASD phenotype, a "player" metric, referred to as the Shapley value, was calculated for each gene in the case and control cohorts. Sixty seven genes were found to have significantly elevated player scores and likely represent significant contributors to the genetic coordination underlying ASD. Using network and cross-study analysis, we found that these genes are involved in biological pathways known to be affected in the autism cases and that a subset directly interact with several genes known to have strong associations to autism. These findings suggest that coalitional game theory can be applied to large-scale genomic data to identify hidden yet influential players in complex polygenic disorders such as autism.