CBEA: Competitive balances for taxonomic enrichment analysis

Research in human associated microbiomes often involves the analysis of taxonomic count tables generated via high-throughput sequencing. It is difficult to apply statistical tools as the data is high-dimensional, sparse, and compositional. An approachable way to alleviate high-dimensionality and sparsity is to aggregate variables into pre-defined sets. Set-based analysis is ubiquitous in the genomics literature, and has demonstrable impact in improving interpretability and power of downstream analysis. Unfortunately, there is a lack of sophisticated set-based analysis methods specific to microbiome taxonomic data, where current practice often employs abundance summation as a technique for aggregation. This approach prevents comparison across sets of different sizes, does not preserve inter-sample distances, and amplifies protocol bias. Here, we attempt to fill this gap with a new single sample taxon enrichment method that uses a novel log-ratio formulation based on the competitive null hypothesis commonly used in the enrichment analysis literature. Our approach, titled competitive balances for taxonomic enrichment analysis (CBEA), generates sample-specific enrichment scores as the scaled log ratio of the subcomposition defined by taxa within a set and the subcomposition defined by its complement. We provide sample-level significance testing by estimating an empirical null distribution of our test statistic with valid p-values. Herein we demonstrate using both real data applications and simulations that CBEA controls for type I error even under high sparsity and high inter-taxa correlation scenarios. Additionally, it provides informative scores that can be inputs to downstream analyses such as prediction tasks. Author summary The study of human associated microbiomes relies on genomic surveys via high-throughput sequencing. However, microbiome taxonomic data is sparse and high dimensional which prevents the application of standard statistical techniques. One approach to address this problem is to perform analyses at the level of taxon sets. Set-based analysis has a long history in the genomics literature, with demonstrable impact in improving both power and interpretability. Unfortunately, there is limited interest in developing new set-based tools tailored for microbiome taxonomic data given its unique features compared to other ‘omics data types. We developed a new tool to generate taxon set enrichment scores at the sample level through a novel log-ratio formulation based on the competitive null hypothesis. Our scores can be used for statistical inference at both the sample and population levels, as well as inputs to other downstream analyses such as prediction models. We demonstrate the performance of our method against competing approaches across both real data analyses and simulation studies.