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Author(s): Diana Vilas Boas
Carina Almeida
Sanna Sillankorva
Ana Nicolau
Joana Azeredo
Nuno F. Azevedo
Title: Discrimination of bacteriophage infected cells using locked nucleic acid fluorescent in situ hybridization (LNA-FISH)
Issue Date: 2016
Abstract: Bacteriophage-host interaction studies in biofilm structures are still challenging due to the technicallimitations of traditional methods. The aim of this study was to provide a direct fluorescence in situhybridization (FISH) method based on locked nucleic acid (LNA) probes, which targets the phagereplication phase, allowing the study of population dynamics during infection. Bacteriophagesspecific for two biofilm-forming bacteria, Pseudomonas aeruginosa and Acinetobacter, were selected.Four LNA probes were designed and optimized for phage-specific detection and for bacterialcounterstaining. To validate the method, LNA-FISH counts were compared with the traditionalplaque forming unit (PFU) technique. To visualize the progression of phage infection within a biofilm,colony-biofilms were formed and infected with bacteriophages. A good correlation (r = 0.707) wasobserved between LNA-FISH and PFU techniques. In biofilm structures, LNA-FISH provided a gooddiscrimination of the infected cells and also allowed the assessment of the spatial distribution ofinfected and non-infected populations.
Related Information: info:eu-repo/grantAgreement/FCT - Fundação para a Ciência e Tecnologia/Projectos de Investigação Clínica/PIC/IC/82815/2007/Desenvolvimento e aplicação de mímicos de DNA para a rápida identificação de microrganismos patogénicos/DNA mimics
info:eu-repo/grantAgreement/FCT - Fundação para a Ciência e Tecnologia/Projetos Estratégicos/UID/EQU/00511/2013 - POCI-01-0145-FEDER-006939/Laboratório de Engenharia de Processos, Ambiente, Biotecnologia e Energia/LEPABE
Document Type: Artigo em Revista Científica Internacional
Rights: restrictedAccess
Appears in Collections:FEUP - Artigo em Revista Científica Internacional

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