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Capture, Concentration, and Detection of Salmonella in Foods Using Magnetic Ionic Liquids and Recombinase Polymerase Amplification.

Stephanie A HiceKevin D ClarkJared L AndersonByron F Brehm-Stecher
Published in: Analytical chemistry (2018)
We previously investigated the extraction and concentration of bacteria from model systems using magnetic ionic liquid (MIL) solvents while retaining their viability. Here, we combine MIL-based sample preparation with isothermal amplification and detection of Salmonella-specific DNA using recombinase polymerase amplification (RPA). After initial developmental work with Serratia marcescens in water, Salmonella Typhimurium ATCC 14028 was inoculated in water, 2% milk, almond milk, or liquid egg samples and extracted using one of two MILs, including trihexyl(tetradecyl)phosphonium cobalt(II) hexafluoroacetylacetonate ([P66614+][Co(hfacac)3-]) and trihexyl(tetradecyl)phosphonium nickel(II) hexafluoroacetylacetonate ([P66614+][Ni(hfacac)3-]). Viable cells were recovered from the MIL extraction phase after the addition of modified LB broth, followed by a 20 min isothermal RPA assay. Amplification was carried out using supersaturated sodium acetate heat packs and results compared to those using a conventional laboratory thermocycler set to a single temperature. Results were visualized using either gel electrophoresis or nucleic acid lateral flow immunoassay (NALFIA). The combined MIL-RPA approach enabled detection of Salmonella at levels as low as 103 CFU mL-1. MIL-based sample preparation required less than 5 min to capture and concentrate sufficient cells for detection using RPA, which (including NALFIA or gel-based analysis) required approximately 30-45 min. Our results suggest the utility of MILs for the rapid extraction and concentration of pathogenic microorganisms in food samples, providing a means for physical enrichment that is compatible with downstream analysis using RPA.
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