Novel strategy to detect and locate periodontal pathogens: The PNA-FISH technique

• Type: Article in International Scientific Journal
• Date: 2016

Authors

Luzia Mendes (Author) FMDUP; Rui Rocha (Author) FEUP; Andreia Sofia Azevedo (Author) FEUP; Catarina Ferreira (Author) Other; Mariana Henriques (Author) Other; Miguel Gonçalves Pinto (Author) FMDUP; Nuno Filipe Azevedo (Author) FEUP

Journal

Title: Microbiological Research

Vol. 192

Pages: 185-191

ISSN: 0944-5013

Publisher: Elsevier

Indexing

ISI Web of Knowledge - 3 Citations

ISI Web of Science

Scopus - 11 Citations

Pubmed / Medline

Scientific classification

CORDIS: Health sciences

FOS: Medical and Health sciences

Associated Projects

DNA mimics - Desenvolvimento e aplicação de mímicos de DNA para a rápida identificação de microrganismos patogénicos

LEPABE - Laboratório de Engenharia de Processos, Ambiente, Biotecnologia e Energia

Other information

Authenticus ID: P-00K-NWE

DOI: 10.1016/j.micres.2016.07.002.

Abstract (EN): Purpose: We aim to develop peptic nucleic acid (PNA) probes for the identification and localization of Aggregatibacter actinomycetemcomintans and Porphyromonas gingivalis in sub-gingival plaque and gingival biopsies by Fluorescence in situ Hybridization (FISH). Methods: A PNA probe was designed for each microorganism. The PNA-FISH method was optimized to allow simultaneous hybridization of both microorganisms with their probe (PNA-FISH multiplex). After being tested on representative strains of P. gingivalis and A. actinomycetemcomitans,the PNA-FISH method was then adapted to detect microorganisms in the subgingival plaque and gingival samples, collected from patients with severe periodontitis. Results: The best hybridization conditions were found to be 59 ◦C for 150 min for both probes (PgPNA1007 and AaPNA235). The in silico sensitivity and specificity was both 100% for PgPNA1007 probe and 100% and 99.9% for AaPNA235 probe, respectively. Results on clinical samples showed that the PNA-FISH method was able to detect and discriminate target bacteria in the mixed microbial population of the subgingival plaque and within periodontal tissues. Conclusion: This investigation presents a new highly accurate method for P. gingivalis and A. actinomycetemcomitans detection and co-location in clinical samples, in just few hours. With this technique we were able to observe spatial distribution of these species within polymicrobial communities in the periodontal pockets and, for the first time with the FISH method, in the organized gingival tissue.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 7