PMID- 26848853 OWN - NLM STAT- MEDLINE DCOM- 20160719 LR - 20231111 IS - 1932-6203 (Electronic) IS - 1932-6203 (Linking) VI - 11 IP - 2 DP - 2016 TI - Fluorescence In Vivo Hybridization (FIVH) for Detection of Helicobacter pylori Infection in a C57BL/6 Mouse Model. PG - e0148353 LID - 10.1371/journal.pone.0148353 [doi] LID - e0148353 AB - INTRODUCTION: In this study, we applied fluorescence in vivo hybridization (FIVH) using locked nucleic acid (LNA) probes targeting the bacterial rRNA gene for in vivo detection of H. pylori infecting the C57BL/6 mouse model. A previously designed Cy3_HP_LNA/2OMe_PS probe, complementary to a sequence of the H. pylori 16S rRNA gene, was used. First, the potential cytotoxicity and genotoxicity of the probe was assessed by commercial assays. Further, the performance of the probe for detecting H. pylori at different pH conditions was tested in vitro, using fluorescence in situ hybridization (FISH). Finally, the efficiency of FIVH to detect H. pylori SS1 strain in C57BL/6 infected mice was evaluated ex vivo in mucus samples, in cryosections and paraffin-embedded sections by epifluorescence and confocal microscopy. RESULTS: H. pylori SS1 strain infecting C57BL/6 mice was successfully detected by the Cy3_HP_LNA/2OMe_PS probe in the mucus, attached to gastric epithelial cells and colonizing the gastric pits. The specificity of the probe for H. pylori was confirmed by microscopy. CONCLUSIONS: In the future this methodology can be used in combination with a confocal laser endomicroscope for in vivo diagnosis of H. pylori infection using fluorescent LNA probes, which would be helpful to obtain an immediate diagnosis. Our results proved for the first time that FIVH method is applicable inside the body of a higher-order animal. FAU - Fontenete, Silvia AU - Fontenete S AD - LEPABE, Laboratory for Process Engineering, Environment, Biotechnology and Energy, Faculty of Engineering, University of Porto, Porto, Portugal. AD - i3S, Instituto de Investigacao e Inovacao em Saude, Universidade do Porto, Porto, Portugal. AD - IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal. AD - Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense M, Denmark. AD - ICBAS, Institute of Biomedical Sciences Abel Salazar, University of Porto, Porto, Portugal. FAU - Leite, Marina AU - Leite M AD - i3S, Instituto de Investigacao e Inovacao em Saude, Universidade do Porto, Porto, Portugal. AD - IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal. FAU - Cappoen, Davie AU - Cappoen D AD - Laboratory of Microbiology, Parasitology and Hygiene (LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Antwerp, Belgium. FAU - Santos, Rita AU - Santos R AD - LEPABE, Laboratory for Process Engineering, Environment, Biotechnology and Energy, Faculty of Engineering, University of Porto, Porto, Portugal. AD - i3S, Instituto de Investigacao e Inovacao em Saude, Universidade do Porto, Porto, Portugal. AD - IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal. AD - Laboratory of General Biochemistry and Physical Pharmacy, Ghent University, Gent, Belgium. FAU - Ginneken, Chris Van AU - Ginneken CV AD - Laboratory of Applied Veterinary Morphology, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Antwerp, Belgium. FAU - Figueiredo, Ceu AU - Figueiredo C AD - i3S, Instituto de Investigacao e Inovacao em Saude, Universidade do Porto, Porto, Portugal. AD - IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal. AD - FMUP, Faculty of Medicine of the University of Porto, University, Porto, Portugal. FAU - Wengel, Jesper AU - Wengel J AD - Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense M, Denmark. FAU - Cos, Paul AU - Cos P AD - Laboratory of Microbiology, Parasitology and Hygiene (LMPH), Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Antwerp, Belgium. FAU - Azevedo, Nuno Filipe AU - Azevedo NF AD - LEPABE, Laboratory for Process Engineering, Environment, Biotechnology and Energy, Faculty of Engineering, University of Porto, Porto, Portugal. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20160205 PL - United States TA - PLoS One JT - PloS one JID - 101285081 RN - 0 (Oligonucleotides) RN - 0 (RNA, Bacterial) RN - 0 (locked nucleic acid) SB - IM MH - Animals MH - Cell Line MH - Disease Models, Animal MH - Female MH - Gastric Mucosa/microbiology MH - Helicobacter Infections/*diagnosis MH - Helicobacter pylori/*genetics/*physiology MH - Humans MH - Mice MH - Mice, Inbred C57BL MH - Oligonucleotides/metabolism MH - RNA, Bacterial/genetics/metabolism MH - Staining and Labeling/*methods PMC - PMC4743915 COIS- Competing Interests: The authors have read the journal's policy and the authors of this manuscript have the following competing interests: Two of the authors of this manuscript have commercial interests in this area. Jesper Wengel is cofounder of RiboTask ApS which offer LNA/2'-OMe-RNA probes for RNA targeting. Nuno Filipe Azevedo is cofounder of Biomode SA which develops molecular methods for the rapid detection of microorganisms. This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials. EDAT- 2016/02/06 06:00 MHDA- 2016/07/20 06:00 PMCR- 2016/02/05 CRDT- 2016/02/06 06:00 PHST- 2015/12/02 00:00 [received] PHST- 2016/01/18 00:00 [accepted] PHST- 2016/02/06 06:00 [entrez] PHST- 2016/02/06 06:00 [pubmed] PHST- 2016/07/20 06:00 [medline] PHST- 2016/02/05 00:00 [pmc-release] AID - PONE-D-15-52300 [pii] AID - 10.1371/journal.pone.0148353 [doi] PST - epublish SO - PLoS One. 2016 Feb 5;11(2):e0148353. doi: 10.1371/journal.pone.0148353. eCollection 2016.