Phagomagnetic immunoassay for the rapid detection of Salmonella

Abstract

This work explores the use of the phage P22 in a phagomagnetic immunoassay for the rapid detection of Salmonella. The covalent attachment of wild-type phages was performed on two different magnetic carriers: carboxyl-activated magnetic nanoparticles (300 nm) and tosyl-activated magnetic microparticles (2.8 μm). The bacteria were captured and preconcentrated by the phage-modified magnetic particles, followed by the detection using specific anti-Salmonella antibodies conjugated to horseradish peroxidase as an optical reporter. Outstanding selectivity and sensitivity was obtained with this approach, achieving detection limits of 19 CFU mL(-1) in 2.5 h without any pre-enrichment, in milk samples. Moreover, if the samples were pre-enriched for 6 h, the method was able to detect as low as 1.4 CFU in 25 mL of milk. Therefore, the proposed strategy based on the combined use of phagomagnetic separation with immunological labeling is promising as a rapid and simple method for food safety.

Modified Bacteriophage S16 Long Tail Fiber Proteins for Rapid and Specific Immobilization and Detection of Salmonella Cells

“…Interestingly, by combining phage P22-based magnetic separation with anti-Salmonella antibodies, Laube et al reported a detection limit of 19 CFU · ml Ϫ1 Salmonella, representing one of the most sensitive phage-based detection assays to date (34). However, it was also noted that the phage P22 particles used caused cell lysis (34).…”

“…Interestingly, by combining phage P22-based magnetic separation with anti-Salmonella antibodies, Laube et al reported a detection limit of 19 CFU · ml Ϫ1 Salmonella, representing one of the most sensitive phage-based detection assays to date (34). However, it was also noted that the phage P22 particles used caused cell lysis (34). While phage-induced lysis can be combined with IMS to detect E. coli (62) and Salmonella (63) based on phage-encoded biomarker amplification, lysis may also be problematic if downstream identification requires the recovery of viable cells-an issue not associated with LTFs and other phage-encoded proteins with no inherent lytic activity.…”

“…The limitations of phages as affinity molecules include their relatively large size in comparison to that of antibodies, the retention of enzymatic activity by their binding proteins, and their basal lytic activity for target cells, which releases cellular components that could hinder downstream detection (34). A simplified alternative to applying whole phages for bacterial detection is to utilize phage-encoded host interaction proteins as affinity molecules.…”

Magnetic Bead‐Based Immunoassay Coupled with Tyramide Signal Amplification for Detection of Salmonella in Foods

“…Similar sensitivity decreases in testing food samples have been observed in other studies. Laube et al used phagoomagnetic immunoassay to detect S. Typhimurium in LB broth and milk, and obtained detection limits of 10 and 19 CFU/mL without any pre-enrichment step, respectively, which indicates the sensitivity of the assay decreased by nearly twofold in food testing (Laube et al, 2014). Similarly, Amaro et al detected S. Typhimurium in various food samples, including chicken broth, low fat milk and freshly squeezed orange juice, using scano-magneto immunoassay based on a nano-composite (Amaro et al, 2012).…”

Current and Emerging Innovations for Detection of Food-Borne Salmonella

“…Bacteriophages have also been explored as a means to capture Salmonella cells. Phages may ofer some advantages over antibodies given their inherent speciicity for host cells, their ease of production in bacteria versus animals or eukaryotic cell culture, and their relative stability in harsh conditions such as pH and temperature extremes [70].…”