A bacteriophage detection tool for viability assessment of Salmonella cells.
Salmonellosis, one of the most common food and water-borne diseases, has a major global health and economic impact. Salmonella cells present high infection rates, persistence over inauspicious conditions and the potential to preserve virulence in dormant states when cells are viable but non-culturable (VBNC). These facts are challenging for current detection methods. Culture methods lack the capacity to detect VBNC cells, while biomolecular methods (e.g. DNA- or protein-based) hardly distinguish between dead innocuous cells and their viable lethal counterparts. This work presents and validates a novel bacteriophage (phage)-based microbial detection tool to detect and assess Salmonella viability. Salmonella Enteritidis cells in a VBNC physiological state were evaluated by cell culture, flow-cytometry and epifluorescence microscopy, and further assayed with a biosensor platform. Free PVP-SE1 phages in solution showed the ability to recognize VBNC cells, with no lysis induction, in contrast to the minor recognition of heat-killed cells. This ability was confirmed for immobilized phages on gold surfaces, where the phage detection signal follows the same trend of the concentration of viable plus VBNC cells in the sample. The phage probe was then tested in a magnetoresistive biosensor platform allowing the quantitative detection and discrimination of viable and VBNC cells from dead cells, with high sensitivity. Signals arising from 3 to 4 cells per sensor were recorded. In comparison to a polyclonal antibody that does not distinguish viable from dead cells, the phage selectivity in cell recognition minimizes false-negative and false-positive results often associated with most detection methods.
IBB-Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, University of Minho Campus de Gualtar, 4700-057 Braga, Portugal; International Iberian Nanotechnology Laboratory, Av. Mestre José Veiga, 4715-330 Braga, Portugal. Electronic address: elisabete.fernandes@deb.uminho.pt.
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E
Fernandes
V C
VC
Martins
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C
Nóbrega
C M
CM
Carvalho
F A
FA
Cardoso
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Cardoso
J
J
Dias
D
D
Deng
L D
LD
Kluskens
P P
PP
Freitas
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Azeredo
D001435:Bacteriophages / Q000302:isolation & purification*; D015374:Biosensing Techniques / Q000379:methods*; D002470:Cell Survival; D006801:Humans; D012475:Salmonella / Q000821:virology; D012480:Salmonella Infections / Q000175:diagnosis / Q000628:therapy
Bacteriophage; Cell viability; Magnetic nanoparticles; Magnetoresistive biochip; Salmonella; Viable but non-culturable bacteria