A nanoparticle-based method for culture-free bacterial DNA enrichment from whole blood

Point-of-care (POC) diagnostics are one of the quick and sensitive detection approaches used in current clinical applications, but always face a performance tradeoff between time-to-result and assay sensitivity. One critical setting where these limitations are evident is the detection of sepsis, whe...

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Published inBiosensors & bioelectronics Vol. 99; pp. 150 - 155
Main Authors Hassan, Marwa M., Ranzoni, Andrea, Cooper, Matthew A.
Format Journal Article
LanguageEnglish
Published England Elsevier B.V 15.01.2018
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Abstract Point-of-care (POC) diagnostics are one of the quick and sensitive detection approaches used in current clinical applications, but always face a performance tradeoff between time-to-result and assay sensitivity. One critical setting where these limitations are evident is the detection of sepsis, where 6–10mL of whole blood may contain as little as one bacterial colony forming unit (cfu). The large sample volume, complex nature of the sample and low analyte concentration necessitates signal enhancement using culture-based or molecular amplification techniques. In the time-critical diagnosis of sepsis, waiting for up to 24h to produce sufficient DNA for analysis is not possible. As a consequence, there is a need for integrated sample preparation methods that could enable shorter detection times, whilst maintaining high analytical performance. We report the development of a culture-free bacterial enrichment method to concentrate bacteria from whole blood in less than 3h. The method relies on triple-enrichment steps to magnetically concentrate bacterial cells and their DNA with a 500-fold reduction in sample volume (from 10 to 0.02mL). Using this sample preparation method, sensitive qPCR detection of the extracted S. aureus bacterial DNA was achieved with a detection limit of 5±0.58cfu/mL within a total elapsed time of 4h; much faster than conventional culture-based approaches. The method could be fully automated for integration into clinical practice for point-of-care or molecular detection of bacterial DNA from whole blood. [Display omitted]
AbstractList Point-of-care (POC) diagnostics are one of the quick and sensitive detection approaches used in current clinical applications, but always face a performance tradeoff between time-to-result and assay sensitivity. One critical setting where these limitations are evident is the detection of sepsis, where 6–10mL of whole blood may contain as little as one bacterial colony forming unit (cfu). The large sample volume, complex nature of the sample and low analyte concentration necessitates signal enhancement using culture-based or molecular amplification techniques. In the time-critical diagnosis of sepsis, waiting for up to 24h to produce sufficient DNA for analysis is not possible. As a consequence, there is a need for integrated sample preparation methods that could enable shorter detection times, whilst maintaining high analytical performance. We report the development of a culture-free bacterial enrichment method to concentrate bacteria from whole blood in less than 3h. The method relies on triple-enrichment steps to magnetically concentrate bacterial cells and their DNA with a 500-fold reduction in sample volume (from 10 to 0.02mL). Using this sample preparation method, sensitive qPCR detection of the extracted S. aureus bacterial DNA was achieved with a detection limit of 5±0.58cfu/mL within a total elapsed time of 4h; much faster than conventional culture-based approaches. The method could be fully automated for integration into clinical practice for point-of-care or molecular detection of bacterial DNA from whole blood. [Display omitted]
Point-of-care (POC) diagnostics are one of the quick and sensitive detection approaches used in current clinical applications, but always face a performance tradeoff between time-to-result and assay sensitivity. One critical setting where these limitations are evident is the detection of sepsis, where 6-10mL of whole blood may contain as little as one bacterial colony forming unit (cfu). The large sample volume, complex nature of the sample and low analyte concentration necessitates signal enhancement using culture-based or molecular amplification techniques. In the time-critical diagnosis of sepsis, waiting for up to 24h to produce sufficient DNA for analysis is not possible. As a consequence, there is a need for integrated sample preparation methods that could enable shorter detection times, whilst maintaining high analytical performance. We report the development of a culture-free bacterial enrichment method to concentrate bacteria from whole blood in less than 3h. The method relies on triple-enrichment steps to magnetically concentrate bacterial cells and their DNA with a 500-fold reduction in sample volume (from 10 to 0.02mL). Using this sample preparation method, sensitive qPCR detection of the extracted S. aureus bacterial DNA was achieved with a detection limit of 5±0.58cfu/mL within a total elapsed time of 4h; much faster than conventional culture-based approaches. The method could be fully automated for integration into clinical practice for point-of-care or molecular detection of bacterial DNA from whole blood.
Author Cooper, Matthew A.
Hassan, Marwa M.
Ranzoni, Andrea
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Keywords qPCR
POCT
Blood
Bacterial sepsis detection
Magnetic nanoparticles
Language English
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Snippet Point-of-care (POC) diagnostics are one of the quick and sensitive detection approaches used in current clinical applications, but always face a performance...
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SubjectTerms Bacteria
Bacterial sepsis detection
Biosensing Techniques
Blood
DNA
DNA, Bacterial - genetics
DNA, Bacterial - isolation & purification
Humans
Limit of Detection
Magnetic nanoparticles
Nanoparticles
POCT
Point-of-Care Systems
qPCR
Sensitivity and Specificity
Sepsis - blood
Sepsis - microbiology
Staphylococcus aureus - growth & development
Staphylococcus aureus - isolation & purification
Staphylococcus aureus - pathogenicity
Title A nanoparticle-based method for culture-free bacterial DNA enrichment from whole blood
URI https://dx.doi.org/10.1016/j.bios.2017.07.057
https://www.ncbi.nlm.nih.gov/pubmed/28753457
https://search.proquest.com/docview/1924604377
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