ABSTRACT
Comprehensive and accurate comparisons of transcriptomic distributions of cells from samples taken from two different biological states, such as healthy versus diseased individuals, are an emerging challenge in single-cell RNA sequencing (scRNA-seq) analysis. Current methods for detecting differentially abundant (DA) subpopulations between samples rely heavily on initial clustering of all cells in both samples. Often, this clustering step is inadequate since the DA subpopulations may not align with a clear cluster structure, and important differences between the two biological states can be missed. Here, we introduce DA-seq, a targeted approach for identifying DA subpopulations not restricted to clusters. DA-seq is a multiscale method that quantifies a local DA measure for each cell, which is computed from its k nearest neighboring cells across a range of k values. Based on this measure, DA-seq delineates contiguous significant DA subpopulations in the transcriptomic space. We apply DA-seq to several scRNA-seq datasets and highlight its improved ability to detect differences between distinct phenotypes in severe versus mildly ill COVID-19 patients, melanomas subjected to immune checkpoint therapy comparing responders to nonresponders, embryonic development at two time points, and young versus aging brain tissue. DA-seq enabled us to detect differences between these phenotypes. Importantly, we find that DA-seq not only recovers the DA cell types as discovered in the original studies but also reveals additional DA subpopulations that were not described before. Analysis of these subpopulations yields biological insights that would otherwise be undetected using conventional computational approaches.