Human heat-shock protein 60 receptor-coated paramagnetic beads show improved capture of Listeria monocytogenes in the presence of other Listeria in food

Abstract

Aims: To investigate the suitability of human Hsp60, a receptor for Listeria adhesion protein (LAP), on paramagnetic beads (PMB) to capture Listeria monocytogenes from food in the presence of other Listeria to facilitate rapid and specific detection of this pathogen. Methods and Results: Commercially available streptavidin‐coated PMBs were linked with biotinylated Hsp60 (PMB‐Hsp60), and the bacterial capture efficiency from pure culture and meat samples was determined. Capture rate was also compared with the monoclonal antibody (MAb)‐C11E9‐coated beads (PMB‐C11E9) and the commercial Dynabeads anti‐Listeria. Captured cells were detected and quantified by plating on selective medium, quantitative real‐time PCR (qPCR) and a light‐scattering sensor. Overall, all ligand‐coated beads had similar capture efficiency (varied from 1·8 to 9·2%) for L. monocytogenes under the conditions employed, and the minimum cell number required to achieve such capture was 103 CFU ml−1. PMB‐Hsp60 had significantly greater capture efficiency for pathogenic Listeria (P < 0·0001) than the nonpathogenic Listeria. In contrast, PMB‐C11E9 and Dynabeads anti‐Listeria had similar capture efficiency for both. The efficacy of all PMBs to capture L. monocytogenes in the presence of Listeria innocua from food matrices was compared. Although Dynabeads anti‐Listeria had the overall best capture efficiency, PMB‐Hsp60 was able to selectively capture L. monocytogenes even in the presence of 10–100‐fold more L. innocua cells from enriched meat samples. Conclusions: Data show that the human cell receptor, Hsp60, is suitable for the capture of pathogenic Listeria on PMB in the presence of other Listeria in food. Significance and Impact of the Study: As pathogen interaction with host cells is highly specific, host cell receptors could be used as alternate capture molecules on PMB to aid in specific detection of pathogens.

Highly specific fiber optic immunosensor coupled with immunomagnetic separation for detection of low levels of Listeria monocytogenes and L. ivanovii

“…IMS using PMBs is used to separate and concentrate target pathogens from food samples before detection by plating, immunoassay, PCR, or biosensor methods [31,37,39,42,45,51]. Antibodies [14] or alternative molecules [19,51,52] are used as capture molecules for IMS, and improvements in reagents and assay platform development are essential to enhance assay performance.…”

“…Furthermore, the binding of bacteriophage to host cells is an irreversible process, which may lead to higher capture efficiency than with antibody-coated PMBs. Koo et al [19] used Hsp60-coated M-280, which showed a capture efficiency for L. monocytogenes of 1.8%–9.2%.…”

“…PMBs were recovered using MPC-S, washed 3 times using 1 mL of PBST, and resuspended in 200 μL of PBS. Finally, PMBs were subjected to vigorous vortexing to release the captured bacteria and 100 μL of each suspension was surface-plated onto BHI or MOX agar plates for enumeration [19]. In some experiments, Dynabeads Anti- Listeria (Invitrogen) were used in parallel as a control.…”

“…An automated light-scattering sensor, BARDOT (BActerial Rapid Detection using Optical light-scattering Technology; Advanced Bioimaging Systems, LLC, West Lafayette, IN) was used to identify colonies of Listeria captured by IMS (described above) on BHI or MOX agar plates [19,61]. This system collects scatter images of bacterial colonies (diameter, 1.3 ± 0.2 mm) through a diode laser (635 nm), and the bacteria were identified by comparing scatter images with library-stored images [61].…”

“…However, when an enrichment step is used, the non-pathogenic species may overgrow and outcompete L. monocytogenes [17-19], leading to false-negative results. L. innocua is the most frequently found bacteria in Listeria -contaminated foods [17,20], thus presenting a challenge for the specific capture and detection of pathogenic Listeria [21].…”