Bacterial pathogens pose an increasing food safety and bioterrorism concern. Current DNA detection methods utilizing sensitive nanotechnology and biosensors have shown excellent detection, but require expensive and time-consuming polymerase chain reaction (PCR) to amplify DNA targets; thus, a faster, more economical method is still essential. In this proof-of-concept study, we investigated the ability of a gold nanoparticle-DNA (AuNP-DNA) biosensor to detect non-PCR amplified genomic Salmonella enterica serovar Enteritidis (S. enteritidis) DNA, from pure or mixed bacterial culture and spiked liquid matrices. Non-PCR amplified DNA was hybridized into sandwich-like structures (magnetic nanoparticles/DNA/AuNPs) and analyzed through detection of gold voltammetric peaks using differential pulse voltammetry. Our preliminary data indicate that non-PCR amplified genomic DNA can be detected at a concentration as low as 100 ng/mL from bacterial cultures and spiked liquid matrices, similar to reported PCR amplified detection levels. These findings also suggest that AuNP-DNA biosensors are a first step towards a viable detection method of bacterial pathogens, in particular, for resource-limited settings, such as field-based or economically limited conditions. Future efforts will focus on further optimization of the DNA extraction method and AuNP-biosensors, to increase sensitivity at lower DNA target concentrations from food matrices comparable to PCR amplified DNA detection strategies.
USA Food and Drug Administration. HACCP Principles. Avaialble online: http://www.fda.gov/Food/FoodSafety/HazardAnalysisCriticalControlPointsHACCP/HACCPPrinciplesApplicationGuidelines/default.htm (accessed on 31 May 2012).
[3]
Robinson-Dunn, B. The microbiology laboratory's role in response to bioterroism. Arch. Pathol. Lab. Med. 2002, 126, 291–294. 11860302
[4]
Tsoka, S.; Gill, A.; Brookman, J.L.; Hoare, M. Rapid monitoring of virus-like particles using an optical biosensor: A feasibility study. J. Biotechnol. 1988, 63, 147–153.
[5]
Ding, C.; Zhang, Q.; Lin, J.-M.; Zhang, S.S. Electrochemical detection of DNA hybridization based on bio-bar code method. Biosens. Bioelectron. 2009, 24, 3140–3143, doi:10.1016/j.bios.2009.03.015. 19362810
Pal, S.P.; Ying, W.; Alocilja, E.C.; Downes, F.P. Sensitivity and specificity performance of a direct-charge transfer biosensor for detecting Bacillus cereus in selected food matrices. Biosys. Eng. J. 2008, 99, 461–468, doi:10.1016/j.biosystemseng.2007.11.015.
[9]
Pal, S.; Alocilja, E.C. Electrically active polyaniline coated magnetic (EAPM) nanoparticle as novel transducer in biosensor for dection of Bacillus anthracis spores in food samples. Biosens. Bioelectron. 2009, 24, 1437–1444, doi:10.1016/j.bios.2008.08.020. 18823768
[10]
Pal, S.; Alocilja, E.C. Electrically active magnetic nanoparticles as novel concentrator and electrochemical redox transducer in Bacillus anthracis DNA detection. Biosens. Bioelectron. 2010, 26, 1624–1630, doi:10.1016/j.bios.2010.08.035. 20864333
[11]
Poonam, P.; Deo, N. Current correlation functions for chemical sensors based on DNA decorated carbon nanotube. Sens. Actuators B Chem. 2008, 135, 327–335, doi:10.1016/j.snb.2008.09.003.
Wang, J.; Zhang, S.; Zhang, Y. Fabrication of chronocoulometric DNA sensor based on gold nanoparticles/poly (L-lysine) modified glassy carbon electrode. Anal. Biochem. 2010, 396, 304–309, doi:10.1016/j.ab.2009.10.004. 19818728
[14]
Zhang, D.; Alocilja, E.C. Characterization of nano-porus silicon-based DNA biosensor for the detection of Salmonella Enteritidis. IEEE Sens. J. 2008, 8, 775–780, doi:10.1109/JSEN.2008.923037.
[15]
Zhang, D.; Carr, D.J.; Alocilja, E.C. Fluorescent bio-barcode DNA assay for the detection of Salmonella enterica serovar Enteritidis. Biosens. Bioelectron. 2009, 24, 1377–1381, doi:10.1016/j.bios.2008.07.081. 18835708
[16]
Zhang, D.; Huarng, M.C.; Alocilja, E.C. A multiplex nanoparticle-based bio-barcode DNA senosr for the simultaneous detection of multiple pathogens. Biosens. Bioelectron. 2010, 26, 1736–1742, doi:10.1016/j.bios.2010.08.012. 20810267
[17]
Saha, K.; Agasti, S.S.; Kim, C.; Li, X.; Rotello, V. Gold nanoparticles in chemical and biolgical sensing. Chem. Rev. 2012, 112, 2739–2779, doi:10.1021/cr2001178. 22295941
[18]
Doria, G.; Conde, J.; Veigas, B.; Giestas, L.; Almeida, C.; Assuncao, M.; Rosa, J.; Baptista, P.V. Nobel metal nanoparticles for biosensing applications. Sensors 2012, 12, 1657–1687, doi:10.3390/s120201657. 22438731
[19]
Drummond, T.G.; Hill, M.G.; Barton, J.K. Electrochemical DNA sensors. Nat. Biotechnol. 2003, 21, 1192–1199, doi:10.1038/nbt873. 14520405
Weng, J.; Zhang, J.; Li, H.; Sun, L.; Lin, C.; Zhang, Q. Label-free DNA sensor by boron-dooped diamond electrode using an AC impedimetric approach. Anal. Chem. 2008, 80, 7075–7083, doi:10.1021/ac800610z. 18707136
[22]
Goluch, E.D.; Nam, J.M.; Georganopoulou, D.G.; Chiesl, T.N.; Shaikh, K.A.; Ryu, K.S.; Barron, A.E.; Mirkin, C.A.; Liu, C. A bio-barcode assay for on-chip attomolar sensitivity protein detection. Lab Chip 2006, 6, 1293–1299, doi:10.1039/b606294f. 17102842
[23]
Nam, J.M.; Thaxton, C.S.; Mirkin, C.A. Nanoparticle-based bio-bar codes for the ultrasenstive detection of proteins. Science 2003, 301, 1884–1886, doi:10.1126/science.1088755. 14512622
[24]
Nam, J.M.; Stoeva, S.I.; Mirkin, C.A. Bio-bar-code-based DNA detection with PCR-like sensitivity. J. Am. Chem. Soc. 2004, 126, 5932–5933, doi:10.1021/ja049384+. 15137735
[25]
Salmonella Surveillance: Annual Summary, 2005; US Department of Health and Human Services CDC: Atlanta, GA, USA, 2007.
[26]
Hill, H.D.; Mirkin, C.A. The bio-barcode assay for the detection of protein and nuclei acid targets using DTT-induced ligand exchange. Nat. Protoc. 2006, 1, 324–336, doi:10.1038/nprot.2006.51. 17406253
[27]
Wang, S.J.; Yeh, D.B. Designing of polymerase chain reaction primers for the dection of Salmonella enteritidis in foods and faecal samples. Lett. Appl. Microbiol. 2005, 34, 422–427.
[28]
Saito, H; Miura, Ki. Preparation of transforming deoxyribonucleic acid by phenol treatment. Biochim. Biophys. Acta 1963, 17, 619–629.
[29]
Ambjornsson, T.; Lomholt, M.A.; Metzler, R. Directed motion emerging from two coupled random processes: Translocation of a chain through a membrane nanopore driven by binding proteins. J. Phys. Condens. Matt. 2005, 17, S1841–S1869, doi:10.1088/0953-8984/17/20/013.
[30]
Hochmeister, M.N.; Budowle, B.; Rudin, O.; Gehrig, C.; Borer, U.; Thali, M.; Dirnhofer, R. Evaluation of prostate-specific antigen (PSA) membrane test assays for the forensic identification of seminal fluid. J. Forensic Sci. 1999, 44, 1057–1060. 10486959
[31]
Lu, C.H.; Kalmar, B.; Malaspina, A.; Greensmith, L.; Petzold, A. A method to solubilise protein aggregates for immunoassay quantification which overcomes the neurofilament “hoo” effect. J. Neurosci. Meth. 2011, 195, 143–150, doi:10.1016/j.jneumeth.2010.11.026.
[32]
Mackay, I.M.; Arden, K.E.; Nitsche, A. Real-time PCR in virology. Nucl. Acids Res. 2002, 30, 1292–1305, doi:10.1093/nar/30.6.1292. 11884626
[33]
Stocher, M.; Leb, V.; Berg, J. Aconvenient approach to the generation of multiple internal control DNA for a panel of real-time PCR assays. J. Virol. Meth. 2003, 108, 1–8, doi:10.1016/S0166-0934(02)00266-5.