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Immunomagnetic nanoparticle based quantitative PCR for rapid detection of Salmonella

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Abstract

We have developed a rapid and sensitive method for immunomagnetic separation (IMS) of Salmonella along with their real time detection via PCR. Silica-coated magnetic nanoparticles were functionalized with carboxy groups to which anti-Salmonella antibody raised against heat-inactivated whole cells of Salmonella were covalently attached. The immuno-captured target cells were detected in beverages like milk and lemon juice by multiplex PCR and real time PCR with a detection limit of 104 cfu.mL−1 and 103 cfu.mL−1, respectively. We demonstrate that IMS can be used for selective concentration of target bacteria from beverages for subsequent use in PCR detection. PCR also enables differentiation of Salmonella typhi and Salmonella paratyphi A using a set of four specific primers. In addition, IMS—PCR can be used as a screening tool in the food and beverage industry for the detection of Salmonella within 3–4 h which compares favorably to the time of several days that is needed in case of conventional detection based on culture and biochemical methods.

The method uses silica coated magnetic nanoparticles immobilized with anti-Salmonella antibody for immunomagnetic separation of Salmonella from beverages followed by detection by multiplex PCR (mPCR) and real time PCR (qPCR). This methodology contributes to rapid screening and accurate detection of Salmonella contaminations in beverages.

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References

  1. Valdés MG, González ACV, Calzón JAG, Díaz-García ME (2009) Analytical nanotechnology for food analysis. Microchim Acta 166: 1–19

    Article  Google Scholar 

  2. María Isabel Pividori MI, Salvador Alegret S (2010) Micro and nanoparticles in biosensing systems for food safety and environmental monitoring. An example of converging technologies. Microchim Acta 170:227–242

    Article  Google Scholar 

  3. Lu AH, Salabas EL, Ferdi S (2007) Magnetic nanoparticles: synthesis, protection, functionalization, and application. Angew Chem Int Ed 46:1222–1244

    Article  CAS  Google Scholar 

  4. Ashtari P, He X, Wang K, Gong P (2005) An efficient method for recovery of target ssDNA based on amino-modified silica-coated magnetic nanoparticles. Talanta 67:548–554

    Article  CAS  Google Scholar 

  5. Fu A, Hu W, Xu L, Wilson RJ, Yu H, Osterfeld SJ, Gambhir SS, Wang SX (2009) Protein-functionalized synthetic antiferromagnetic nanoparticles for biomolecule detection and magnetic manipulation. Angew Chem Int Ed Engl 48:1620–1624

    Article  CAS  Google Scholar 

  6. Chou TC, Hsu W, Wang CH, Chen YJ, Fang JM (2011) Rapid and specific influenza virus detection by functionalized magnetic nanoparticles and mass spectrometry. J Nanobiotechnology 9:52

    Article  CAS  Google Scholar 

  7. Huang C, Neoh KG, Wang L, Kang ET, Shuter B (2010) Magnetic nanoparticles for magnetic resonance imaging: modulation of macrophage uptake by controlled PEGylation of the surface coating. J Mater Chem 20:8512–8520

    Article  CAS  Google Scholar 

  8. Fuchigami T, Kitamoto Y, Namiki Y (2012) Size-tunable drug-delivery capsules composed of a magnetic nanoshell. Biomatter 2:313–320

    Article  Google Scholar 

  9. Li Z, He L, He N, Deng Y, Shi Z, Wang H, Li S, Liu H, Wang Z, Wang D (2011) Polymerase chain reaction coupling with magnetic nanoparticles-based biotin-avidin system for amplification of chemiluminescent detection signals of nucleic acid. J Nanosci Nanotechnol 11:1074–1078

    Article  CAS  Google Scholar 

  10. Tang Y, Zou J, Ma C, Ali Z, Li Z, Li X, Ma N, Mou X, Deng Y, Zhang L, Li K, Lu G, Yang H, He N (2013) Highly sensitive and rapid detection of Pseudomonas aeruginosa based on magnetic enrichment and magnetic separation. Theranostics 3:85–92

    Article  CAS  Google Scholar 

  11. Pappert G, Rieger M, Niessner R, Seidel M (2010) Immunomagnetic nanoparticle-based sandwich chemiluminescence-ELISA for the enrichment and quantification of E. coli. Microchim Acta 168:1–8

    Article  CAS  Google Scholar 

  12. Iqbal Z, Lai EPC, Avis TJ (2012) Development of polymer-modified magnetic nanoparticles and quantum dots for Escherichia coli binding test. Microchim Acta 176:193–200

    Article  CAS  Google Scholar 

  13. Roda A, Mirasoli M, Roda B, Bonvicini F, Colliva C, Reschiglian P (2012) Recent developments in rapid multiplexed bioanalytical methods for foodborne pathogenic bacteria detection. Microchim Acta 178:7–28

    Article  CAS  Google Scholar 

  14. Benoit PW, Donahue DW (2003) Methods for rapid separation and concentration of bacteria in food that bypass time-consuming cultural enrichment. J Food Prot 66:1935–1948

    Google Scholar 

  15. Liu G, Yu X, Xue F, Chen W, Ye Y, Yang X, Lian Y, Yan Y, Zong K (2012) Screening and preliminary application of a DNA aptamer for rapid detection of Salmonella O8. Microchim Acta 178:237–244

    Article  CAS  Google Scholar 

  16. Wang H, Zhang C, Xing D (2011) Simultaneous detection of Salmonella enterica, Escherichia coli O157:H7, and Listeria monocytogenes using oscillatory-flow multiplex PCR. Microchim Acta 173:503–512

    Article  CAS  Google Scholar 

  17. Salomonsson AC, Aspa’n A, Johansson S, Heino A, Ha¨ggblom P (2005) Salmonella detection by polymerase chain reaction after pre-enrichment of feed samples. J Rapid Meth Automat Microbiol 13:96–110

    Article  CAS  Google Scholar 

  18. Huang YF, Wang YF, Yan XP (2010) Amine-functionalized magnetic nanoparticles for rapid capture and removal of bacterial pathogens. Environ Sci Technol 44:7908–7913

    Article  CAS  Google Scholar 

  19. Lian W, Litherland SA, Badrane H, Tan W, Wu D, Baker HV, Gulig PA, Lim DV, Jin S (2004) Ultrasensitive detection of biomolecules with fluorescent dye-doped nanoparticles. J Anal Biochemistry 334:135–144

    Article  CAS  Google Scholar 

  20. Padmavathy B, Patel A, Vinoth Kumar R, Jaffar Ali BM (2012) Superparamagnetic nanoparticles based immunomagnetic separation - multiplex PCR assay for detection of Salmonella. Sci Adv Mat 4:1–7

    Article  Google Scholar 

  21. Marathe SA, Chowdhury R, Bhattacharya R, Nagarajan AG, Chakravorthy D (2012) Direct detection of Salmonella without pre-enrichment in milk, ice-cream and fruit juices by PCR against hilA gene. Food Control 23:559–563

    Article  CAS  Google Scholar 

  22. Rieger M, Schaumann GE, Mouvenchery YK, Niessner R, Seidel M, Baumann T (2012) Development of antibody-labelled superparamagnetic nanoparticles for the visualisation of benzo[a]pyrene in porous media with magnetic resonance imaging. Anal Bioanal Chem 403:2529–2540

    Article  CAS  Google Scholar 

  23. Feifel SC, Lisdat F (2011) Silica nanoparticles for the layer-by-layer assembly of fully electro-active cytochrome c multilayers. J Nanobiotechnology 9:59

    Article  CAS  Google Scholar 

  24. Wang S, Wen S, Shen M, Guo R, Cao X, Wang J, Shi X (2011) Aminopropyltriethoxysilane-mediated surface functionalization of hydroxyapatite nanoparticles: synthesis, characterization, and in vitro toxicity assay. Int J Nanomedicine 6:3449–3459

    CAS  Google Scholar 

  25. Spanova A, Rittich B, Karpiskova R, Cechova L, Skapova D (2000) PCR identification of Salmonella cells in food and stool samples after immunomagnetic separation. Bioseparation 9:379–384

    Article  CAS  Google Scholar 

  26. Beuchat LR, Mann DA (2008) Survival and growth of acid-adapted and unadapted Salmonella in and on raw tomatoes as affected by variety, stage of ripeness, and storage temperature. J Food Prot 71:1572–1579

    Google Scholar 

  27. Bai Y, Song M, Cui Y, Shi C, Wang D, Paoli GC, Shi X (2013) A rapid method for the detection of foodborne pathogens by extraction of a trace amount of DNA from raw milk based on amino-modified silica-coated magnetic nanoparticles and polymerase chain reaction. Anal Chim Acta. doi:10.1016/j.aca.2013.05.043

    Google Scholar 

  28. Yang Y, Xu F, Xu H, Aguilar ZP, Niu R, Yuan Y, Sun J, You X, Lai W, Xiong Y, Wan C, Wei H (2013) Magnetic nano-beads based separation combined with propidium monoazide treatment and multiplex PCR assay for simultaneous detection of viable Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes in food products. Food Microbiol 34:418–424

    Article  CAS  Google Scholar 

  29. Wang L, Li Y, Mustaphai A (2007) Rapid and simultaneous quantitation of Escherichia coli 0157:H7, Salmonella, and Shigella in ground beef by multiplex real-time PCR and immunomagnetic separation. J Food Prot 70:1366–1372

    CAS  Google Scholar 

  30. Mercanoglu TB, Ben U, Aytac SA (2009) Rapid detection of Salmonella in milk by combined immunomagnetic separation-polymerase chain reaction assay. J Dairy Sci 92:2382–2388

    Article  Google Scholar 

  31. Moreira AN, Conceição FR, Conceição Rde C, Ramos RJ, Carvalhal JB, Dellagostin OA, Aleixo JA (2008) Detection of Salmonella typhimurium in raw meats using in-house prepared monoclonal antibody coated magnetic beads and PCR assay of the fimA gene. J Immunoassay Immunochem 29:58–69

    CAS  Google Scholar 

  32. Notzon A, Helmuth R, Bauer J (2006) Evaluation of an immunomagnetic separation-real-time PCR assay for the rapid detection of Salmonella in meat. J Food Prot 69:2896–2901

    CAS  Google Scholar 

  33. Mercanoglu B, Griffiths MW (2005) Combination of immunomagnetic separation with real-time PCR for rapid detection of Salmonella in milk, ground beef, and alfalfa sprouts. J Food Prot 68:557–561

    CAS  Google Scholar 

  34. Hagren V, von Lode P, Syrjälä A, Korpimäki T, Tuomola M, Kauko O, Nurmi J (2008) An 8-hour system for Salmonella detection with immunomagnetic separation and homogeneous time-resolved fluorescence PCR. Int J Food Microbiol 125:158–161

    Article  CAS  Google Scholar 

  35. Li A, Zhang H, Zhang X, Wang Q, Tian J, Li Y, Li J (2010) Rapid separation and immunoassay for low levels of Salmonella in foods using magnetosome-antibody complex and real-time fluorescence quantitative PCR. J Sep Sci 33:3437–3443

    Article  CAS  Google Scholar 

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Acknowledgment

We acknowledge Central Instrumentation Facility, Pondicherry University for SEM imaging. We thank Dr. Vaidehi, SMF Hospital for providing the clinical isolates. BP thanks Council of Scientific and Industrial Research (CSIR) for the award of Senior Research Fellowship. This work was partially supported by KBC Research Foundation.

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Authors and Affiliations

  1. AU-KBC Research Centre, M.I.T Campus of Anna University, Chennai, 600044, India

    Padmavathy Bakthavathsalam, Vinoth Kumar Rajendran, Uttara Saran & Suvro Chatterjee

  2. Centre for Green Energy Technology, Pondicherry University, Puducherry, 605014, India

    Baquir Mohammed Jaffar Ali

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  1. Padmavathy Bakthavathsalam
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  2. Vinoth Kumar Rajendran
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  3. Uttara Saran
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  4. Suvro Chatterjee
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  5. Baquir Mohammed Jaffar Ali
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Corresponding author

Correspondence to Baquir Mohammed Jaffar Ali.

Additional information

Padmavathy Bakthavathsalam and Vinoth Kumar Rajendran contributed equally to this work.

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Bakthavathsalam, P., Rajendran, V.K., Saran, U. et al. Immunomagnetic nanoparticle based quantitative PCR for rapid detection of Salmonella . Microchim Acta 180, 1241–1248 (2013). https://doi.org/10.1007/s00604-013-1052-1

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  • DOI: https://doi.org/10.1007/s00604-013-1052-1

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