Detection of Tissue-resident Bacteria in Bladder Biopsies by 16S rRNA Fluorescence In Situ Hybridization

Michael L. Neugent, Jashkaran Gadhvi, Kelli L. Palmer, Philippe E. Zimmern, Nicole J. De Nisco

Research output: Contribution to journalArticle

Abstract

Visualization of the interaction of bacteria with host mucosal surfaces and tissues can provide valuable insight into mechanisms of pathogenesis. While visualization of bacterial pathogens in animal models of infection can rely on bacterial strains engineered to express fluorescent proteins such as GFP, visualization of bacteria within the mucosa of biopsies or tissue obtained from human patients requires an unbiased method. Here, we describe an efficient method for the detection of tissue-associated bacteria in human biopsy sections. This method utilizes fluorescent in situ hybridization (FISH) with a fluorescently labeled universal oligonucleotide probe for 16S rRNA to label tissue-associated bacteria within bladder biopsy sections acquired from patients suffering from recurrent urinary tract infection. Through use of a universal 16S rRNA probe, bacteria can be detected without prior knowledge of species, genera, or biochemical characteristics, such as lipopolysaccharide (LPS), that would be required for detection by immunofluorescence experiments. We describe a complete protocol for 16S rRNA FISH from biopsy fixation to imaging by confocal microscopy. This protocol can be adapted for use in almost any type of tissue and represents a powerful tool for the unbiased visualization of clinically-relevant bacterial-host interactions in patient tissue. Furthermore, using species or genera-specific probes, this protocol can be adapted for the detection of specific bacterial pathogens within patient tissue.

Original languageEnglish (US)
JournalJournal of visualized experiments : JoVE
Issue number152
DOIs
StatePublished - Oct 18 2019

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Biopsy
Fluorescence In Situ Hybridization
Bacteria
Urinary Bladder
Fluorescence
Tissue
Visualization
Pathogens
Mucous Membrane
Oligonucleotide Probes
Confocal microscopy
Oligonucleotides
Confocal Microscopy
Urinary Tract Infections
Fluorescent Antibody Technique
Lipopolysaccharides
Animal Models
Labels
Animals
Proteins

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

Detection of Tissue-resident Bacteria in Bladder Biopsies by 16S rRNA Fluorescence In Situ Hybridization. / Neugent, Michael L.; Gadhvi, Jashkaran; Palmer, Kelli L.; Zimmern, Philippe E.; De Nisco, Nicole J.

In: Journal of visualized experiments : JoVE, No. 152, 18.10.2019.

Research output: Contribution to journalArticle

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