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Updated: Jul 19, 2025
Murine Fecal Isolation and Microbiota Transplantation
Published on: May 26, 2023
A Comparison of Methods of Gut Microbiota Transplantation for Preclinical Studies
Jonas Mingaila1, Alessandro Atzeni1, Aurelijus Burokas1
1Department of Biological Models, Institute of Biochemistry, Life Sciences Center, Vilnius University, Sauletekio Ave. 7, LT-10257 Vilnius, Lithuania.
Abstract:
The experimental details reported in preclinical fecal microbiota transplantation (FMT) protocols are highly inconsistent, variable, and/or incomplete. We therefore evaluated FMT from a human donor to antibiotic-induced microbial-depleted mice by exploring the effects of six techniques based on antibiotic (AB) or antibiotic + antimycotic (AB + T) gut decontamination, different administration routes, and different dosing intervals on the gut microbial population, assessed using 16S and 18S sequencing. In addition, we explored the effectiveness of FMT in terms of inflammation, physiological, and behavioral outcomes. Our results showed that intrarectal FMT at low dosing intervals better preserved the donor's gut bacterial community at genus level. Furthermore, we showed a lower abundance of several genera of fungi in animals treated with AB + T. In addition, we observed that AB + T gut decontamination followed by per os FMT, once every 3 days, affected behavioral parameters when compared to other FMT techniques. Accordingly, the same FMT groups that showed an association with some of the behavioral tests were also related to specific gut fungal genera, suggesting a possible mediation. Our findings may be useful for optimizing the practice of FMT and also in terms of donor microbiota preservation. This information may help to improve the reproducibility and reliability of FMT studies.
Insights
Optimizing fecal microbiota transplantation (FMT) protocols is crucial for research reproducibility. Intrarectal FMT with specific decontamination methods and dosing intervals best preserves donor gut microbes and influences host outcomes.
Area of Science:
- Microbiology
- Gastroenterology
- Immunology
Background:
- Preclinical fecal microbiota transplantation (FMT) protocols exhibit significant inconsistencies and variability.
- Standardizing FMT procedures is essential for reliable and reproducible research outcomes.
Purpose of the Study:
- To evaluate the impact of different gut decontamination methods, administration routes, and dosing intervals on FMT efficacy in mice.
- To assess the influence of FMT on gut microbial populations, inflammation, and host physiology and behavior.
Main Methods:
- Fecal microbiota transplantation (FMT) was performed on mice with antibiotic-induced gut microbial depletion.
- Six techniques involving antibiotic (AB) or antibiotic + antimycotic (AB + T) decontamination, varied administration routes, and dosing intervals were tested.
- Gut microbial communities were analyzed using 16S and 18S sequencing; host outcomes were assessed for inflammation, physiology, and behavior.
Main Results:
- Intrarectal FMT administered at low dosing intervals demonstrated superior preservation of the donor's gut bacterial community at the genus level.
- Antibiotic + antimycotic (AB + T) decontamination led to a lower abundance of specific fungal genera.
- A specific FMT protocol (AB + T decontamination followed by per os FMT every 3 days) significantly affected host behavioral parameters and showed associations with specific gut fungal genera.
Conclusions:
- The findings provide insights into optimizing FMT protocols for improved donor microbiota preservation and experimental reproducibility.
- Specific decontamination and administration strategies can influence both microbial engraftment and host-microbe interactions, including behavioral outcomes.
- This study offers a foundation for refining FMT methodologies in preclinical research to enhance reliability and validity.
