Adipose tissue microbiota in humans: an open issue.
A specific 'adipose tissue' microbiota has been recently identified in mice and hypothesized in humans. The purpose of this study was to verify the presence of microbiota of human whole adipose tissue and isolated adipocytes by combining culture-dependent and independent methods.



Standard microbiological cultural techniques and 16S ribosomal RNA (16S rRNA) gene sequencing (Illumina technology) on DNA and RNA were employed to study (a) whole abdominal subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) from 14 obese and five normal-weight subjects and (b) mature adipocytes isolated from SAT and VAT after collagenase digestion or mechanical separation. To optimize the 16S rRNA gene detection, we used different DNA extraction methods (lysis with proteinase K, proteinase K+lysozyme and microbeads) and amplification procedures (semi-quantitative standard PCR and real-time quantitative PCR).



Microbiological cultures were negative in all analyzed samples. In enzymatically isolated adipocytes, 90% of the sequenced bacterial DNA belonged to Clostridium histolyticum, the bacterium from which the collagenase enzyme was isolated. Bacterial 16S rRNA gene was not detected from DNA and RNA of whole SAT and VAT, as well as of mechanically isolated mature adipocytes, even after blocking with a specific primer the nonspecific amplification of human mitochondrial 12S rRNA.



Our results do not support the presence of a human adipose tissue microbiota. In addition, they emphasized the technical problems encountered when applying metagenomic studies to human tissues with very low or absent bacterial load.
IRCCS Istituto Auxologico Italiano, Diabetes Research Laboratory, Milan, Italy.|IRCCS Istituto Auxologico Italiano, Laboratory of Molecular Biology, Milan, Italy.
A
A
Zulian
IRCCS Istituto Auxologico Italiano, Diabetes Research Laboratory, Milan, Italy.|IRCCS Istituto Auxologico Italiano, Laboratory of Molecular Biology, Milan, Italy.
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Cancello
IRCCS Istituto Auxologico Italiano, Microbiology Laboratory, Milan, Italy.
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E
Cesana
National Research Council, Institute for Biomedical Technologies, Segrate, Milan, Italy.
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E
Rizzi
National Research Council, Institute for Biomedical Technologies, Segrate, Milan, Italy.
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Consolandi
National Research Council, Institute for Biomedical Technologies, Segrate, Milan, Italy.
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M
Severgnini
INCO and Department of General Surgery, Istituto Clinico Sant'Ambrogio, Milan, Italy.
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V
Panizzo
IRCCS Istituto Auxologico Italiano, Laboratory of Molecular Biology, Milan, Italy.
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Di Blasio
Department of Pathophysiology and Transplantation University of Milan, INCO and Department of General Surgery, Istituto Clinico Sant'Ambrogio, Milan, Italy.
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G
Micheletto
IRCCS Istituto Auxologico Italiano, Diabetes Research Laboratory, Milan, Italy.
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C
Invitti
D000328:Adult; D015992:Body Mass Index; D005260:Female; D000069196:Gastrointestinal Microbiome / Q000502:physiology; D005786:Gene Expression Regulation; D006801:Humans; D007249:Inflammation / Q000382:microbiology* / Q000473:pathology; D007413:Intestinal Mucosa / Q000382:microbiology* / Q000473:pathology; D050152:Intra-Abdominal Fat / Q000382:microbiology* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009765:Obesity / Q000382:microbiology* / Q000473:pathology; D012333:RNA, Messenger / Q000378:metabolism; D012336:RNA, Ribosomal, 16S; D060888:Real-Time Polymerase Chain Reaction
D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S