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Detection of non-PCR amplified S. enteritidis genomic DNA from food matrices using a gold-nanoparticle DNA biosensor: a proof-of-concept study.

Vetrone SA, Huarng MC, Alocilja EC - Sensors (Basel) (2012)

Bottom Line: Bacterial pathogens pose an increasing food safety and bioterrorism concern.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.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Whittier College, 13406 E. Philadelphia St., Whittier, CA 90608, USA. svetrone@whittier.edu

ABSTRACT
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.

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Related in: MedlinePlus

Non-PCR amplified S. enteritidis genomic DNAt can be detected in acidic liquid food matrices using the AuNP-DNA biosensor. Graphs represent the average differential current peak values vs. DNAt concentration attained from genomic DNAt extracted from PBC and MBC spiked orange juice. H2O, blank control; NS-DNA, non-specific PCR amplified B. anthracis DNAt (0.1 ng/μL) negative control; HCl, 1M hydrogen chloride. Graphs represent the average value of duplicate samples for each condition. Error bars represent the standard deviation of the mean.
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f5-sensors-12-10487: Non-PCR amplified S. enteritidis genomic DNAt can be detected in acidic liquid food matrices using the AuNP-DNA biosensor. Graphs represent the average differential current peak values vs. DNAt concentration attained from genomic DNAt extracted from PBC and MBC spiked orange juice. H2O, blank control; NS-DNA, non-specific PCR amplified B. anthracis DNAt (0.1 ng/μL) negative control; HCl, 1M hydrogen chloride. Graphs represent the average value of duplicate samples for each condition. Error bars represent the standard deviation of the mean.

Mentions: As the AuNP-DNA biosensor exhibited a trend towards the detection of non-PCR amplified genomic DNAt from PBC and MBC samples, we next determined if it could also detect non-PCR amplified genomic DNAt extracted from basic and acidic liquid food matrices spiked/contaminated with S. enteritidis PBC or MBC. We first spiked 9 mL samples of 2% milk (a basic food matrix, Figure 4A) and 100% orange juice (an acidic food matrix, Figure 5) with 1.0 mL of fresh, PBC of S. enteritidis at three bacterial concentrations: 1.04 cfu/mL (orange juice only), 1.05 cfu/mL and 1.06 cfu/mL. The spiked matrices were allowed to incubate at room temperature for 15 min before aliquoting a 1.0 mL sample of each for TRIzol® extraction of genomic DNAt. The DNAt samples were then hybridized with AuNPs and MNPs, and detected on the AuNP-DNA biosensor using DPV. The results display that the AuNP-DNA biosensor was also able to detect S. enteritidis genomic DNAt from PBC spiked basic and acidic food matrices at all levels of contamination (Figures 4A and 5).


Detection of non-PCR amplified S. enteritidis genomic DNA from food matrices using a gold-nanoparticle DNA biosensor: a proof-of-concept study.

Vetrone SA, Huarng MC, Alocilja EC - Sensors (Basel) (2012)

Non-PCR amplified S. enteritidis genomic DNAt can be detected in acidic liquid food matrices using the AuNP-DNA biosensor. Graphs represent the average differential current peak values vs. DNAt concentration attained from genomic DNAt extracted from PBC and MBC spiked orange juice. H2O, blank control; NS-DNA, non-specific PCR amplified B. anthracis DNAt (0.1 ng/μL) negative control; HCl, 1M hydrogen chloride. Graphs represent the average value of duplicate samples for each condition. Error bars represent the standard deviation of the mean.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3472839&req=5

f5-sensors-12-10487: Non-PCR amplified S. enteritidis genomic DNAt can be detected in acidic liquid food matrices using the AuNP-DNA biosensor. Graphs represent the average differential current peak values vs. DNAt concentration attained from genomic DNAt extracted from PBC and MBC spiked orange juice. H2O, blank control; NS-DNA, non-specific PCR amplified B. anthracis DNAt (0.1 ng/μL) negative control; HCl, 1M hydrogen chloride. Graphs represent the average value of duplicate samples for each condition. Error bars represent the standard deviation of the mean.
Mentions: As the AuNP-DNA biosensor exhibited a trend towards the detection of non-PCR amplified genomic DNAt from PBC and MBC samples, we next determined if it could also detect non-PCR amplified genomic DNAt extracted from basic and acidic liquid food matrices spiked/contaminated with S. enteritidis PBC or MBC. We first spiked 9 mL samples of 2% milk (a basic food matrix, Figure 4A) and 100% orange juice (an acidic food matrix, Figure 5) with 1.0 mL of fresh, PBC of S. enteritidis at three bacterial concentrations: 1.04 cfu/mL (orange juice only), 1.05 cfu/mL and 1.06 cfu/mL. The spiked matrices were allowed to incubate at room temperature for 15 min before aliquoting a 1.0 mL sample of each for TRIzol® extraction of genomic DNAt. The DNAt samples were then hybridized with AuNPs and MNPs, and detected on the AuNP-DNA biosensor using DPV. The results display that the AuNP-DNA biosensor was also able to detect S. enteritidis genomic DNAt from PBC spiked basic and acidic food matrices at all levels of contamination (Figures 4A and 5).

Bottom Line: Bacterial pathogens pose an increasing food safety and bioterrorism concern.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.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Whittier College, 13406 E. Philadelphia St., Whittier, CA 90608, USA. svetrone@whittier.edu

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus