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Abstract Rapid expansion of the probiotics industry demands fast, sensitive, comprehensive, and low‐cost strategies for quality assessment. Here, we introduce a culture‐free, one‐cell‐resolution, phenome‐genome‐combined strategy called Single‐Cell Identification, Viability and Vitality tests, and Source‐tracking (SCIVVS). For each cell directly extracted from the product, the fingerprint region of D2O‐probed single‐cell Raman spectrum (SCRS) enables species‐level identification with 93% accuracy, based on a reference SCRS database from 21 statutory probiotic species, whereas the C–D band accurately quantifies viability, metabolic vitality plus their intercellular heterogeneity. For source‐tracking, single‐cell Raman‐activated Cell Sorting and Sequencing can proceed, producing indexed, precisely one‐cell‐based genome assemblies that can reach ~99.40% genome‐wide coverage. Finally, we validated an integrated SCIVVS workflow with automated SCRS acquisition where the whole process except sequencing takes just 5 h. As it is >20‐fold faster, >10‐time cheaper, vitality‐revealing, heterogeneity‐resolving, and automation‐prone, SCIVVS is a new technological and data framework for quality assessment of live‐cell products.

Abstract Massive microbiome sequencing data has been generated, which elucidates associations between microbes and their environmental phenotypes such as host health or ecosystem status. Outstanding bioinformatic tools are the basis to decipher the biological information hidden under microbiome data. However, most approaches placed difficulties on the accessibility to nonprofessional users. On the other side, the computing throughput has become a significant bottleneck of many analytical pipelines in processing large‐scale datasets. In this study, we introduce Parallel‐Meta Suite (PMS), an interactive software package for fast and comprehensive microbiome data analysis, visualization, and interpretation. It covers a wide array of functions for data preprocessing, statistics, visualization by state‐of‐the‐art algorithms in a user‐friendly graphical interface, which is accessible to diverse users. To meet the rapidly increasing computational demands, the entire procedure of PMS has been optimized by a parallel computing scheme, enabling the rapid processing of thousands of samples. PMS is compatible with multiple platforms, and an installer has been integrated for full‐automatic installation.