Biofunctionalized Magnetic Nanoparticles with Multiplex Touchdown PCR for Simultaneous and Rapid Detection/Identification of Campylobacter jejuni and Campylobacter coli. (12th October 2022)

• Record Type: Journal Article
• Authors: Wenbap, Pattarapong; Rattanarojpong, Triwit; Khunrae, Pongsak; Luangtongkum, Taradon; Erickson, Larry E.; Hansen, Ryan R.; Tuitemwong, Pravate
• Other Names: Parvinzadeh Gashti Mazeyar Academic Editor.
• Is Part Of: Journal of nanomaterials. Volume 2022(2022)
• Publication Date: 2022-10-12
• Subjects: Nanostructured materials -- Periodicals; Nanotechnology -- Periodicals; Nanomatériaux; Nanostructured materials; Nanotechnology; Nanostructures; Nanotechnology; Periodicals; Fulltext; Internet Resources; Periodicals; 620.115
• DOI: 10.1155/2022/5104187
• Languages: English
• ISSNs: 1687-4110

Abstract

Abstract : The simple, accurate, and rapid detection of foodborne pathogens is essential for public health. Development of an immunomagnetic separation (IMS) multiplex touchdown PCR (IMS–multiplex TD–PCR) assay for simultaneous detection and distinguishing of C. jejuni and C. coli is reported herein. Polyclonal antibody (pAb) against multiepitope antigen (MEA) was conjugated to ferromagnetic nanoparticles (FMNs) to produce anti-MEA FMNs. Optimal anti-MEA FMNs loading yielded 26.7 μ g of immunoglobulin G (IgG) molecules per mg of FMNs with an average size of 72 ± 9 nm, corresponding to an 83% rate of pAb conjugation. Anti-MEA FMNs (20 μ g) for IMS captured culturable C. jejuni cells at 3.54 × 10 2 colony-forming unit (CFU)/mL in pure culture, while higher amounts (40 and 60 μ g) reduced the recovery. The scanning electron microscope (SEM) analysis revealed the attachment of anti-MEA FMNs to target bacteria, forming aggregated cells and magnetic nanoparticles in ellipse-like shapes. The subsequent multiplex TD–PCR assay simultaneously detected and distinguished C. jejuni and C. coli at 10 4 CFU/mL in mixed culture and at 10 3 CFU/mL for each individual species. Furthermore, the limit of detection (LOD) of the IMS–multiplex TD–PCR assay was 10 4 CFU/g in spiked chicken breast samples. Specificity was 100% for both C. jejuni and C. coli as none of the amplicons were detected in control samples where Campylobacter was absent. This assay is able to detect and distinguish C. jejuni and C. coli simultaneously and is simple, accurate, and rapid with a time to result of 4 h without an enrichment step, making it a promising approach for rapid and culture-free detection of Campylobacter in chicken products.