FACTS AND ARTIFACTS ABOUT METAGENOMIC ANALYSIS IN HUMAN ADIPOSE TISSUE

Objectives: The existence of a microbiota in the stroma-vascular fraction of human adipose tissue has been recently suggested. The present study was aimed to characterize viable cultivable bacteria and non-viable bacteria in human adipose tissue. Methodology: The presence of bacteria was assessed through standard microbiological cultural techniques and 16S rRNA gene sequencing (Illumina technology) on DNA and RNA from a) whole abdominal subcutaneous (SAT) and visceral (VAT) adipose tissue of 14 obese and 5 normal-weight subjects, b) mature adipocytes isolated from these tissues after collagenase digestion and/or mechanical separation. Transmission electron microscopy (TEM) was used to identify bacteria at intracellular level. Results: Microbiological cultures were negative in all samples. Contaminating bacterial DNA was amplified and sequenced in enzymatically isolated adipocytes. No amplification of the 16S rRNA gene was obtained from DNA and RNA of whole SAT and VAT and mature adipocytes mechanically isolated. Ultra-structure analyses by TEM did not show presence of bacteria within adipocytes. Conclusion: Our results do not confirm the existence of human adipose tissue microbiota. Further investigations are needed to clarify whether the discrepancy between our and previous results is due to methodological differences.