Publication
Development of a novel phagomagnetic-assisted isothermal DNA amplification system for endpoint electrochemical detection of Listeria monocytogenes
| dc.contributor.author | Maciel, Cláudia | |
| dc.contributor.author | Silva, Nádia F. D. | |
| dc.contributor.author | Teixeira, Paula | |
| dc.contributor.author | Magalhães, Júlia M. C. S. | |
| dc.date.accessioned | 2023-05-03T17:18:34Z | |
| dc.date.available | 2023-05-03T17:18:34Z | |
| dc.date.issued | 2023-04-07 | |
| dc.description.abstract | The hitherto implemented Listeria monocytogenes detection techniques are cumbersome or require expensive non-portable instrumentation, hindering their transposition into on-time surveillance systems. The current work proposes a novel integrated system resorting to loop-mediated isothermal amplification (LAMP), assisted by a bacteriophage P100–magnetic platform, coupled to an endpoint electrochemical technique, towards L. monocytogenes expeditious detection. Molybdophosphate-based optimization of the bacterial phagomagnetic separation protocol allowed the determination of the optimal parameters for its execution (pH 7, 25 °C, 32 µg of magnetic particles; 60.6% of specific capture efficiency). The novel LAMP method targeting prfA was highly specific, accomplishing 100% inclusivity (for 61 L. monocytogenes strains) and 100% exclusivity (towards 42 non-target Gram-positive and Gram-negative bacteria). As a proof-of-concept, the developed scheme was successfully validated in pasteurized milk spiked with L. monocytogenes. The phagomagnetic-based approach succeeded in the selective bacterial capture and ensuing lysis, triggering Listeria DNA leakage, which was efficiently LAMP amplified. Methylene blue-based electrochemical detection of LAMP amplicons was accomplished in 20 min with remarkable analytical sensitivity (1 CFU mL−1). Hence, the combined system presented an outstanding performance and robustness, providing a 2.5 h-swift, portable, cost-efficient detection scheme for decentralized on-field application. | pt_PT |
| dc.description.version | info:eu-repo/semantics/publishedVersion | pt_PT |
| dc.identifier.doi | 10.3390/bios13040464 | pt_PT |
| dc.identifier.eid | 85153678617 | |
| dc.identifier.issn | 2079-6374 | |
| dc.identifier.uri | http://hdl.handle.net/10400.14/41020 | |
| dc.identifier.wos | 000977509500001 | |
| dc.language.iso | eng | pt_PT |
| dc.peerreviewed | yes | pt_PT |
| dc.rights.uri | http://creativecommons.org/licenses/by/4.0/ | pt_PT |
| dc.subject | Bacteriophage P100 | pt_PT |
| dc.subject | Electrochemical detection | pt_PT |
| dc.subject | Listeria monocytogenes | pt_PT |
| dc.subject | Loop-mediated isothermal DNA amplification | pt_PT |
| dc.subject | Magnetic capture | pt_PT |
| dc.subject | Milk analysis | pt_PT |
| dc.subject | prfA | pt_PT |
| dc.title | Development of a novel phagomagnetic-assisted isothermal DNA amplification system for endpoint electrochemical detection of Listeria monocytogenes | pt_PT |
| dc.type | journal article | |
| dspace.entity.type | Publication | |
| oaire.citation.issue | 4 | pt_PT |
| oaire.citation.title | Biosensors | pt_PT |
| oaire.citation.volume | 13 | pt_PT |
| rcaap.rights | openAccess | pt_PT |
| rcaap.type | article | pt_PT |