title: Hybridization-based detection of Helicobacter pylori at human body temperature using advanced locked nucleic acid (LNA) probes.
abstract: The understanding of the human microbiome and its influence upon human life has long been a subject of study. Hence, methods that allow the direct detection and visualization of microorganisms and microbial consortia (e.g. biofilms) within the human body would be invaluable. In here, we assessed the possibility of developing a variant of fluorescence in situ hybridization (FISH), named fluorescence in vivo hybridization (FIVH), for the detection of Helicobacter pylori. Using oligonucleotide variations comprising locked nucleic acids (LNA) and 2'-O-methyl RNAs (2'OMe) with two types of backbone linkages (phosphate or phosphorothioate), we were able to successfully identify two probes that hybridize at 37 °C with high specificity and sensitivity for H. pylori, both in pure cultures and in gastric biopsies. Furthermore, the use of this type of probes implied that toxic compounds typically used in FISH were either found to be unnecessary or could be replaced by a non-toxic substitute. We show here for the first time that the use of advanced LNA probes in FIVH conditions provides an accurate, simple and fast method for H. pylori detection and location, which could be used in the future for potential in vivo applications either for this microorganism or for others.
authors.affiliation: LEPABE, Laboratory for Process Engineering, Environment, Biotechnology and Energy, Department of Chemical Engineering, Faculty of Engineering, University of Porto, Porto, Portugal ; IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal ; Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense M, Denmark ; ICBAS, Institute of Biomedical Sciences Abel Salazar, University of Porto, Porto, Portugal.
authors.forename: Sílvia
authors.initials: S
authors.lastname: Fontenete
authors.forename: Nuno
authors.initials: N
authors.lastname: Guimarães
authors.forename: Marina
authors.initials: M
authors.lastname: Leite
authors.forename: Céu
authors.initials: C
authors.lastname: Figueiredo
authors.forename: Jesper
authors.initials: J
authors.lastname: Wengel
authors.forename: Nuno
authors.initials: N
authors.lastname: Filipe Azevedo
mesh_terms: D005434:Flow Cytometry; D016481:Helicobacter Infections / Q000175:diagnosis; D016480:Helicobacter pylori / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008856:Microscopy, Fluorescence; D009693:Nucleic Acid Hybridization; D015341:Nucleic Acid Probes; D013270:Stomach / Q000382:microbiology
chemical_list: D015341:Nucleic Acid Probes