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TitleA phage-based assay for the rapid diagnosis of bloodstream infections
Author(s)Costa, Susana P.
Dias, Nicolina Marques
Melo, Luís Daniel Rodrigues
Santos, Sílvio Roberto Branco
Azeredo, Joana
Carvalho, Carla Alexandra Oliveira Cunha Medeiros
Issue date4-Aug-2019
CitationCosta, Susana P.; Dias, Nicolina M.; Melo, Luís D. R.; Santos, Sílvio B.; Azeredo, Joana; Carvalho, Carla M., A phage-based assay for the rapid diagnosis of bloodstream infections. 23rd Biennial Evergreen International Phage Meeting. Olympia, USA, Aug 4-9, 2019.
Abstract(s)Bloodstream infections (BSIs) are considered a major cause of death worldwide. Staphylococcus aureus is one of the most prevalent pathogens in these infections (Tong et al., 2015) and has been classified as high priority bacteria due to the increasing of multidrug resistant strains like methicillin-resistant S. aureus (MRSA) (World Health Organization, 2017). Thus, specific, sensitive, rapid and cost detection of this pathogen is of great importance. Some bacteriophage proteins present high potential as biorecognition elements for the detection of pathogens. Examples include the cell wall binding domains (CBDs) of phage endolysins that recognize and bind to the bacterial cell wall (Loessner, 2005). In this study, we described a novel assay for detection of S. aureus in blood samples, which combines the advantages of CBDs as specific probes with the accuracy and high throughput of flow cytometry techniques. For this purpose, a CBD from S. aureus phage LM12 endolysin (Melo et al., 2018) was fused with a Green Fluorescent Protein (GFP), expressed by heterologous recombination and their functional analysis was performed by microscopy analysis. In the flow cytometry assays, the GFP_CBD (GFP-AMI_SH3) was tested against Staphylococcus strains and other Gram-positive and Gram-negative bacteria, and proved to be specific to Staphylococcus spp. Also, the optimized assay allowed the rapid detection of S. aureus loads of approximately 1 CFU per 10 mL of blood after an enrichment step. Overall the developed assay allowed a fast, specific and sensitive detection of Staphylococcus in blood and thus can be a promising approach for diagnosis of BSIs. Moreover, we are exploiting its potential as a multiplex tool by using another phage protein from an Enterococcus phage coupled with a red fluorescent protein, in order to allow the detection of both pathogens simultaneously in blood.
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Appears in Collections:CEB - Resumos em Livros de Atas / Abstracts in Proceedings

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