Enriched Aptamer Libraries in Fluorescence-Based Assays for Rikenella microfusus-Specific Gut Microbiome Analyses

Area of Science:

• Microbiology
• Biotechnology
• Molecular Biology

Background:

• Intestinal flora imbalance is linked to various diseases, including intestinal, immune, and metabolic disorders.
• Rikenella microfusus is a key intestinal probiotic with potential as a disease biomarker.
• Current Rikenella detection methods are slow, complex, and costly, hindering clinical applications.

Purpose of the Study:

• To develop a rapid, low-cost method for detecting Rikenella microfusus.
• To evolve a specific DNA aptamer library for Rikenella microfusus labeling.
• To construct aptamer-based biosensors for microbiome monitoring.

Main Methods:

• In vitro whole Cell-SELEX (Systematic Evolution of Ligands by EXponential Enrichment) was used to enrich DNA aptamers targeting Rikenella microfusus.
• Counterselection against five other anaerobic gut bacteria was performed to ensure specificity.
• The enriched aptamer library (R.m-R13) was characterized for affinity (K_d = 9.597 nM).

Main Results:

• An aptamer library (R.m-R13) with high specificity and strong affinity for Rikenella microfusus was successfully evolved.
• Rikenella microfusus was effectively distinguished from control bacteria in complex mixtures.
• Detection was demonstrated using fluorescence microscopy and fluorometric suspension assays, even in human stool samples.

Conclusions:

• The evolved aptamer library offers a promising tool for rapid and reliable monitoring of Rikenella microfusus.
• This work paves the way for aptamer-based biosensing applications in microbiome analysis.
• The findings support the potential of Rikenella microfusus as a biomarker and therapeutic target.