Raman‐activated sorting of antibiotic‐resistant bacteria in human gut microbiota

Abstract

Summary The antibiotic‐resistant bacteria (ARB) and antibiotic‐resistant genes (ARGs) in human gut microbiota have significant impact on human health. While high throughput metagenomic sequencing reveals genotypes of microbial communities, the functionality, phenotype and heterogeneity of human gut microbiota are still elusive. In this study, we applied Raman microscopy and deuterium isotope probing (Raman–DIP) to detect metabolic active ARB (MA‐ARB) in situ at the single‐cell level in human gut microbiota from two healthy adults. We analysed the relative abundances of MA‐ARB under different concentrations of amoxicillin, cephalexin, tetracycline, florfenicol and vancomycin. To establish the link between phenotypes and genotypes of the MA‐ARB, Raman‐activated cell sorting (RACS) was used to sort MA‐ARB from human gut microbiota, and mini‐metagenomic DNA of the sorted bacteria was amplified, sequenced and analysed. The sorted MA‐ARB and their associated ARGs were identified. Our results suggest a strong relation between ARB in human gut microbiota and personal medical history. This study demonstrates that the toolkit of Raman–DIP, RACS and DNA sequencing can be useful to unravel both phenotypes and genotypes of ARB in human gut microbiota at the single‐cell level.

A single-cell Raman-based platform to identify developmental stages of human pluripotent stem cell-derived neurons

“…The SCRS of undifferentiated iPSCs and their neural lineage descendants showed distinct characteristics, and different cell phenotypes were visualized as clear clusters using t-SNE multivariate analysis. These results, along with noninvasive Raman-activated cell sorting techniques (45), such as the recent development of a high-throughput Raman flow cytometer (17,46), demonstrate the potential of transforming our platform into a noninvasive and label-free cell sorting platform to facilitate clinical uses of stem cell-related interventions and therapies. While biomolecular profiles of Raman spectra are complex, it can be challenging and time consuming to collect, process, and analyze a huge number of data manually.…”

One-Cell Metabolic Phenotyping and Sequencing of Soil Microbiome by Raman-Activated Gravity-Driven Encapsulation (RAGE)

“…For example, SCRS can detect not just D 2 O intake (and phenotypes that are correlated with D 2 O intake such as antibiotic resistance) ( 17 ) but assimilatory activity of carbon or nitrogen sources labeled by stable isotopes of C or N ( 19 , 61 ). Similarly, the ability to mine both the cellular carotenoids (in both abundance and structure) and the underlying genotypes encoding biosynthetic pathways at the individual cell level should allow mining for not just carotenoids but lipids, polysaccharides, protein, and even antibiotics ( 24 , 25 , 62 , 63 ). Therefore, it is possible that RAGE-Seq would become a universal and highly versatile tool for precisely probing “who is doing what” and for mining cells or metabolites of interest, from soil and other complex natural ecosystems.…”

Isolation and Culture of Single Microbial Cells by Laser Ejection Sorting Technology

“…In the LIFT isolating process, we are able to observe the isolation of a single cell under the microscope, and LIFT can also be combined with other optical techniques such as fluorescence imaging ( 33 ) and Raman spectroscopy ( 25 – 27 , 34 , 35 ). LIFT has been applied for isolation and cultivation of microbial cells as well because of its ability to isolate the bacterial cells without destroying the microenvironment (for example, ejecting the bacteria and the surrounding soil together, but not focusing on single-cell ejection).…”