DEsingle is an R package for differential expression (DE) analysis of single-cell RNA-seq (scRNA-seq) data. It defines and detects 3 types of differentially expressed genes between two groups of single cells, with regard to different expression status (DEs), differential expression abundance (DEa), and general differential expression (DEg). DEsingle employs Zero-Inflated Negative Binomial model to estimate the proportion of real and dropout zeros and to define and detect the 3 types of DE genes. Results showed that DEsingle outperforms existing methods for scRNA-seq DE analysis, and can reveal different types of DE genes that are enriched in different biological functions.