Evaluation of microplate immunocapture method for detection of Vibrio cholerae, Salmonella Typhi and Shigella flexneri from food

Abstract

BackgroundImproved methods with better separation and concentration ability for detection of foodborne pathogens are in constant need. The aim of this study was to evaluate microplate immunocapture (IC) method for detection of Salmonella Typhi, Shigella flexneri and Vibrio cholerae from food samples to provide a better alternative to conventional culture based methods.ResultsThe IC method was optimized for incubation time, bacterial concentration, and capture efficiency. 6 h incubation and log 6 CFU/ml cell concentration provided optimal results. The method was shown to be highly specific for the pathogens concerned. Capture efficiency (CE) was around 100% of the target pathogens, whereas CE was either zero or very low for non-target pathogens. The IC method also showed better pathogen detection ability at different concentrations of cells from artificially contaminated food samples in comparison with culture based methods. Performance parameter of the method was also comparable (Detection limit- 25 CFU/25 g; sensitivity 100%; specificity-96.8%; Accuracy-96.7%), even better than culture based methods (Detection limit- 125 CFU/25 g; sensitivity 95.9%; specificity-97%; Accuracy-96.2%).ConclusionThe IC method poses to be the potential to be used as a method of choice for detection of foodborne pathogens in routine laboratory practice after proper validation.

The Role of Background Bacteria in Preventing the Isolation and Detection of Shigella and STEC in Foods and Novel MuSIC ddPCR Method to Improve STEC Detection in the Presence of Background Bacteria

“…Recent efforts to improve detection of Shigella in foods have been limited. There have been a few studies looking to improve the molecular methods for detection and characterization of Shigella such as microplate immunocapture, targeted sequencing and DNA microarray assays (Fakruddin, Hossain, & Ahmed, 2017;Ferrario et al, 2017;Y. Li, 2016).…”

Title: Microplate Immunocapture Coupled with LAMP-BART and Selective Plating for the Rapid Detection of Salmonella Infantis in Dry Dog Food and Treats

“…monocytogenes at levels of 10 0 , 10 2 , and 10 4 CFU/25g at rates of 88.9%, 94.4%, and 100%, respectively, but microplate IC with selective plating yielded 0% recovery at 10 0 CFU/25g and 44.4% at 10 2 CFU/25g. Fakruddin et al (2017) found that coupling microplate IC with PCR allowed for detection of Salmonella Typhi in 62.7% of food samples inoculated with concentrations of 10 1 -10 5 CFU/25 g, as compared to 56% detection for samples that underwent traditional enrichment plus PCR. Neither study used a pre-enrichment step prior to conducting microplate IC and selective plating…”

“…A potential means for shortening the enrichment period for the detection of low concentrations of Salmonella is with microplate immunocapture (IC). Microplate IC utilizes an antibody-coated microtiter plate to concentrate bacterial cells for greater detection efficiency (Arbault et al 2014ab;Fakruddin et al 2017;Rogers et al 2018). Although microplate IC is not as widely used as immunomagnetic separation, it is considerably less expensive because it does not require production of antibody-coated beads.…”

“…Although microplate IC is not as widely used as immunomagnetic separation, it is considerably less expensive because it does not require production of antibody-coated beads. Microplate IC has been used in previous studies to concentrate bacterial cells for detection with PCR and/or selective plating (Rogers et al 2017;Fakruddin et al 2017). For example, Fakruddin et al (2017) found that coupling microplate IC with PCR allowed for detection of Salmonella Typhi in 62.7% of food samples inoculated with concentrations of 10 1 -10 5 CFU/25 g, as compared to 56% detection for samples that underwent traditional enrichment plus PCR.…”

“…Microplate IC has been used in previous studies to concentrate bacterial cells for detection with PCR and/or selective plating (Rogers et al 2017;Fakruddin et al 2017). For example, Fakruddin et al (2017) found that coupling microplate IC with PCR allowed for detection of Salmonella Typhi in 62.7% of food samples inoculated with concentrations of 10 1 -10 5 CFU/25 g, as compared to 56% detection for samples that underwent traditional enrichment plus PCR. Rogers et al (2018) found that microplate IC coupled with PCR could detect L. monocytogenes at levels of 10 0 , 10 2 , and 10 4 CFU/25g at rates of 88.9%, 94.4%, and 100% respectively, but microplate IC with selective plating yielded 0% recovery at 10 0 CFU/25g and 44.4% at 10 2 CFU/25g.…”