Reference genomes are essential for metagenomic analyses and functional characterization of the human gut microbiota. We present the Culturable Genome Reference (CGR), a collection of 1,520 nonredundant, high-quality draft genomes generated from >6,000 bacteria cultivated from fecal samples of healthy humans. Of the 1,520 genomes, which were chosen to cover all major bacterial phyla and genera in the human gut, 264 are not represented in existing reference genome catalogs. We show that this increase in the number of reference bacterial genomes improves the rate of mapping metagenomic sequencing reads from 50% to >70%, enabling higher-resolution descriptions of the human gut microbiome. We use the CGR genomes to annotate functions of 338 bacterial species, showing the utility of this resource for functional studies. We also carry out a pan-genome analysis of 38 important human gut species, which reveals the diversity and specificity of functional enrichment between their core and dispensable genomes.

Butyrate-producing bacteria can biosynthesize butyrate and alleviate inflammatory diseases. However, few studies have reported that the genus Collinsella has the ability to produce butyric acid. Here, our study depicts a Collinsella strain, which is a rod-shaped obligate anaerobe that is able to produce butyric acid. This microorganism was isolated from a human gut, and the optimal growth conditions were found to be 37 °C on PYG medium with pH 6.5. The 16S rRNA gene sequence demonstrated that this microorganism shared 99.93% similarity with C. aerofaciens ATCC 25986^T, which was higher than the threshold (98.65%) for differentiating two species. Digital DNA–DNA hybridization and average nucleotide identity values also supported that this microorganism belonged to the species C. aerofaciens. Distinct phenotypic characteristics between TF06-26 and the type strain of C. aerofaciens, such as the fermentation of D-lactose, D-fructose and D-maltose, positive growth under pH 5 and 0.2% (w/v) cholate, suggested this strain was a novel subspecies. Comparative genome analysis revealed that butyric acid kinase and phosphate butyryltransferase enzymes were coded exclusively by this strain, indicating a specific butyric acid-producing function of this C. aerofaciens subspecies within the genus Collinsella. Thus, Collinsella aerofaciens subsp. shenzhenensis subsp. nov. was proposed, with set strain TF06-26^T (=CGMCC 1.5216^T = DSM 105138^T) as the type strain.

Exploiting a pure culture strategy to investigate the composition of the human gut microbiota, two novel anaerobes, designated strains AF52-21^T and CM04-06^T, were isolated from faeces of two healthy Chinese donors and characterized using a polyphasic approach. The two strains were observed to be gram-negative, non-motile, and rod-shaped. Both strains grew optimally at 37 °C and pH 7.0. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the two strains clustered with species of the genus Faecalibacterium and were most closely related to Faecalibacterium prausnitzii ATCC 27768^T with sequence similarity of 97.18% and 96.87%, respectively. The two isolates shared a 16S rRNA gene sequence identity of 98.69%. Draft genome sequencing was performed for strains AF52-21^T and CM04-06^T, generating genome sizes of 2.85 Mbp and 3.01 Mbp. The calculated average nucleotide identity values between the genomes of the strains AF52-21^T and CM04-06^T compared to Faecalibacterium prausnitzii ATCC 27768^T were 83.20% and 82.54%, respectively, and 90.09% when comparing AF52-21^T and CM04-06^T. Both values were below the previously proposed species threshold (95–96%), supporting their recognition as novel species in the genus Faecalibacterium. The genomic DNA G + C contents of strains AF52-21^T and CM04-06^T calculated from genome sequences were 57.77 mol% and 57.51 mol%, respectively. Based on the phenotypic, chemotaxonomic and phylogenetic characteristics, we conclude that both strains represent two new Faecalibacterium species, for which the names Faecalibacterium butyricigenerans sp. nov. (type strain AF52-21^T = CGMCC 1.5206^T = DSM 103434^T) and Faecalibacterium longum sp. nov. (type strain CM04-06^T = CGMCC 1.5208^T = DSM 103432^T) are proposed.

The Lachnospiraceae family holds promise as a source of next‐generation probiotics, yet a comprehensive delineation of its diversity is lacking, hampering the identification of suitable strains for future applications. To address this knowledge gap, we conducted an in‐depth genomic and functional analysis of 1868 high‐quality genomes, combining data from public databases with our new isolates. This data set represented 387 colonization‐selective species‐level clusters, of which eight genera represented multilineage clusters. Pan‐genome analysis, single‐nucleotide polymorphism (SNP) identification, and probiotic functional predictions revealed that species taxonomy, habitats, and geography together shape the functional diversity of Lachnospiraceae. Moreover, analyses of associations with atherosclerotic cardiovascular disease (ACVD) and inflammatory bowel disease (IBD) indicated that several strains of potentially novel Lachnospiraceae species possess the capacity to reduce the abundance of opportunistic pathogens, thereby imparting potential health benefits. Our findings shed light on the untapped potential of novel species enabling knowledge‐based selection of strains for the development of next‐generation probiotics holding promise for improving human health and disease management.

Culture-independent metagenomic studies have revolutionized our understanding of the gut microbiota. However, the lack of full genomes from cultured species is still a limitation for in-depth studies of the gut microbiota. Here we present a substantially expanded version of our Cultivated Genome Reference (CGR), termed CGR2, providing 3324 high-quality draft genomes from isolates selected from a large-scale cultivation of bacterial isolates from fecal samples of healthy Chinese individuals. The CGR2 classifies 527 species (179 previously unidentified species) from 8 phyla, and uncovers a genomic and functional diversity of Collinsella aerofaciens. The CGR2 genomes match 126 metagenome-assembled genomes without cultured representatives in the Unified Human Gastrointestinal Genome (UHGG) collection and harbor 3767 unidentified secondary metabolite biosynthetic gene clusters, providing a source of natural compounds with pharmaceutical potentials. We uncover accurate phage–bacterium linkages providing information on the evolutionary characteristics of interaction between bacteriophages and bacteria at the strain level.

The incidence of young-onset colorectal cancer (yCRC) has been increasing in recent decades, but little is known about the gut microbiome of these patients. Most studies have focused on old-onset CRC (oCRC), and it remains unclear whether CRC signatures derived from old patients are valid in young patients. To address this, we assembled the largest yCRC gut metagenomes to date from two independent cohorts and found that the CRC microbiome had limited association with age across adulthood. Differential analysis revealed that well-known CRC-associated taxa, such as Clostridium symbiosum, Peptostreptococcus stomatis, Parvimonas micra and Hungatella hathewayi were significantly enriched (false discovery rate <0.05) in both old- and young-onset patients. Similar strain-level patterns of Fusobacterium nucleatum, Bacteroides fragilis and Escherichia coli were observed for oCRC and yCRC. Almost all oCRC-associated metagenomic pathways had directionally concordant changes in young patients. Importantly, CRC-associated virulence factors (fadA, bft) were enriched in both oCRC and yCRC compared to their respective controls. Moreover, the microbiome-based classification model had similar predication accuracy for CRC status in old- and young-onset patients, underscoring the consistency of microbial signatures across different age groups.