Poster Presentation: Comparative Analysis of Intervertebral Disc and Gut Microbiomes in Germ Free and Specific-Pathogen-Free Mice using full length 16S rRNA Sequencing
Description
52nd Annual Meeting of the International Society for the Study of the Lumbar Spine (ISSLS)
Cape Town, South Africa
Cape Town, South Africa
Abstract
Introduction: Recent studies and evidence suggest that the microbiome may play a key role in inducing and promoting intervertebral disc (IVD) degeneration (IDD). Bacteria that infiltrate the disc space could contribute to IDD and the generation of low back pain. Several studies have identified Cutibacterium acnes (previously known as Propionibacterium acnes) in pathological IVD tissue. This study aims to determine whether mice represent a suitable model for studying IVD and microbial colonization from IVD and gut samples of specific pathogen free (SPF) (Mice that are free from particular known pathogens, but still have a normal microbiota) and germ-free (GF) (Mice that are completely free of all microorganisms).
Methods: DNA was extracted using the QIAamp UCP Pathogen Mini Kit from IVD and gut tissue samples of female C57BL6J mice (n= 3) kept either under SPF or GF conditions. DNA concentration was quantified using the Qubit system. After extracting the genomic DNA from the samples, a small amount of a known bacterial mixture (called ZymoBIOMICS™ Spike-in Control I) was added before running 16S ribosomal RNA (rRNA) amplification. DNA sequencing was performed using nanopore technology on a MinION platform, followed by base calling with Dorado and filtering with Nanofilt (1400–1600 bp, q-score > 9).
Results: Bacterial DNA was not detected in any IVD sample from either SPF or GF mice (Fig.1), suggesting no microbial load, which aligns with the findings in gut samples of germ-free mice, confirming their sterile status. In contrast, bacterial DNA was identified in the gut tissue of control/SPF mice in only two of three samples. Evaluation of the ZymoBIOMICS™ Microbial Community Standard yielded a strong correlation between expected and measured relative abundances. (Fig.2a, 2b, and 2c)
Discussion: The study revealed that SPF mice are possibly unsuitable as a model for studying a native IVD microbiome. Moreover, it also confirmed that GF-standards were maintained as in the intestine, no bacteria were detected. The future studies will include the analysis of faecal samples from the mice to gain a more understanding of the relationship between gut and IVD microbiomes.
Acknowledgments:
This work was supported by the SNSF Weave Grant (#320030E_224175) and the DFG (#437213841).
Methods: DNA was extracted using the QIAamp UCP Pathogen Mini Kit from IVD and gut tissue samples of female C57BL6J mice (n= 3) kept either under SPF or GF conditions. DNA concentration was quantified using the Qubit system. After extracting the genomic DNA from the samples, a small amount of a known bacterial mixture (called ZymoBIOMICS™ Spike-in Control I) was added before running 16S ribosomal RNA (rRNA) amplification. DNA sequencing was performed using nanopore technology on a MinION platform, followed by base calling with Dorado and filtering with Nanofilt (1400–1600 bp, q-score > 9).
Results: Bacterial DNA was not detected in any IVD sample from either SPF or GF mice (Fig.1), suggesting no microbial load, which aligns with the findings in gut samples of germ-free mice, confirming their sterile status. In contrast, bacterial DNA was identified in the gut tissue of control/SPF mice in only two of three samples. Evaluation of the ZymoBIOMICS™ Microbial Community Standard yielded a strong correlation between expected and measured relative abundances. (Fig.2a, 2b, and 2c)
Discussion: The study revealed that SPF mice are possibly unsuitable as a model for studying a native IVD microbiome. Moreover, it also confirmed that GF-standards were maintained as in the intestine, no bacteria were detected. The future studies will include the analysis of faecal samples from the mice to gain a more understanding of the relationship between gut and IVD microbiomes.
Acknowledgments:
This work was supported by the SNSF Weave Grant (#320030E_224175) and the DFG (#437213841).
Date Issued
2026
Publication Type
Conference Item
Subjects
Microbiome
•
low back pain
•
intervertebral disc
•
degeneration
•
gut-disc axis
Language(s)
en
Author(s)
Additional Credits
Related Collection(s)
Access(Rights)
open.access