Advances in the isolation, cultivation, and identification of gut microbes

Abstract

The gut microbiome is closely associated with human health and the development of diseases. Isolating, characterizing, and identifying gut microbes are crucial for research on the gut microbiome and essential for advancing our understanding and utilization of it. Although culture-independent approaches have been developed, a pure culture is required for in-depth analysis of disease mechanisms and the development of biotherapy strategies. Currently, microbiome research faces the challenge of expanding the existing database of culturable gut microbiota and rapidly isolating target microorganisms. This review examines the advancements in gut microbe isolation and cultivation techniques, such as culturomics, droplet microfluidics, phenotypic and genomics selection, and membrane diffusion. Furthermore, we evaluate the progress made in technology for identifying gut microbes considering both non-targeted and targeted strategies. The focus of future research in gut microbial culturomics is expected to be on high-throughput, automation, and integration. Advancements in this field may facilitate strain-level investigation into the mechanisms underlying diseases related to gut microbiota.

Keywords: Culturomics; Droplet microfluidics; Gut microbes; Microbial identification; Strain-level investigation.

Fig. 1

Advances in the isolation, cultivation, and identification of gut microbes. The isolation and cultivation techniques of gut microbes are depicted on the left. Traditional methods involve plate-based techniques and limiting dilution in liquid media. The renaissance of culturomics focuses on refining and improving culture media and conditions to enhance species diversity through streamlined procedures. Droplet microfluidics in the cultivation of gut microbes is characterized by high-throughput, automation, single-cell, and miniaturization, which are compatible with anaerobic workstations. Phenotypic and genomics selection mostly rely on FACS. In situ cultivation and co-culture techniques are based on membrane diffusion. The identification technique used for gut microbes is depicted on the right. Non-targeted identification methods include classical phenotyping, 16S rRNA gene sequencing, and mass-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Targeted identification mainly employs the principle of nucleic acid amplification or hybridization techniques