title: Rapid pathogen detection with bacterial-assembled magnetic mesoporous silica.
abstract: We report rapid and accurate pathogen detection by coupling with high efficiency magnetic separation of pathogen by Ni(2+)-heterogeneous magnetic mesoporous silica (Ni-HMMS) and real time-polymerase chain reaction (RT-PCR) technique. Ni-HMMS was developed with a significant incorporation of Fe particles within the silica mesopores by programmed thermal hydrogen reaction and functionalized with Ni(2+) ion on the surface by the wet impregnation process. High abundant Ni(2+) ions on the Ni-HMMS surface were able to assemble with cell wall component protein NikA (nickel-binding membrane protein), which contains several pathogenic bacteria including Escherichia coli O157:H7. NikA protein expression experiment showed the outstanding separation rate of the nikA gene-overexpressed E. coli (pSY-Nik) when comparing with wild-type E. coli (44.5 ± 13%) or not over-expressed E. coli (pSY-Nik) (53.2 ± 2.7%). Moreover, Ni-HMMS showed lower obstacle effect by large reaction volume (10 mL) than spherical core/shell-type silica magnetic nanoparticles functionalized with Ni(2+) (ca. 40 nm-diameters). Finally, the Ni-HMMS was successfully assessed to separate pathogenic E. coli O157:H7 and applied to direct and rapid RT-PCR to quantitative detection at ultralow concentration (1 Log10 cfu mL(-1)) in the real samples (milk and Staphylococcus aureus culture broth) without bacterial amplification and DNA extraction step.
authors.affiliation: Korea Institute of Ceramic Engineering & Technology, Seoul 153-801, Korea.
authors.forename: Soo Youn
authors.initials: SY
authors.lastname: Lee
authors.forename: Jiho
authors.initials: J
authors.lastname: Lee
authors.forename: Hye Sun
authors.initials: HS
authors.lastname: Lee
authors.forename: Jeong Ho
authors.initials: JH
authors.lastname: Chang
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000302:isolation & purification*; D000818:Animals; D015374:Biosensing Techniques / Q000379:methods*; D019453:Escherichia coli O157 / Q000302:isolation & purification* / Q000472:pathogenicity; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000302:isolation & purification*; D005516:Food Microbiology; D060328:Magnetic Phenomena; D008460:Meat / Q000382:microbiology; D008892:Milk / Q000382:microbiology; D009532:Nickel / Q000737:chemistry; D012822:Silicon Dioxide / Q000737:chemistry; D013211:Staphylococcus aureus / Q000302:isolation & purification / Q000472:pathogenicity
chemical_list: D018528:ATP-Binding Cassette Transporters; D029968:Escherichia coli Proteins; C089849:nikA protein, E coli; D012822:Silicon Dioxide; D009532:Nickel
keywords: Hydrogen reduction; Magnetic mesoporous silica; Nickel-binding protein; Pathogen detection; Real-time polymerase chain reaction (RT-PCR); Ultralow concentration