Development of DNA-based nano-biosensors for food and environmental applications

Date

2014

Abstract

The Philippine still ranks high on death and morbidity cases due to diarrhea and bacterial infection caused by foodborne and environmental pathogens such as Salmonella typhimurium, S. enteriditis. Enterohemorrhagic Escherichia coli (EHEC) including E. coli 0157:H7, Staphylococcus aureus, Listeria monocytogenes, Vibrio cholera, among others. Several factors contribute to this situation such as poor sanitation specially in rural and urban-poor communities and in small or medium-scale food manufacturing units; increasing number of high risk, elderly, immune compromised and neonatal babies; increased consumption of ready-to-eat and raw foods; and highly integrated world food supply resulting to growing food importation. The common microbiological, immunological and molecular tools such as polymerase chain reaction (PCR) have their advantages and disadvantages. Although microbiological detection is accurate, it often relies in time-consuming growth in culture media, followed by isolation, biochemical identification, and sometimes serology, and need special reagents and facilities. The immunological detection systems are specific but their sensitivity is low. PCR-based detection method is sensitive, but is complex, expensive, time consuming, and labor-intensive. Almost all assays used to detect specific pathogens require some growth in an enrichment medium before analysis. So a rapid, specific and sensitive system for identification of microbial pathogens is vital within the overall context of Hazard Analysis Critical Control Points (HACCP) analysis, point-of-care (POC) diagnosis of foodborne illness, and environmental monitoring. A field operable devices for rapid microbial detection such as DNA-based nanobiosensor would be in a much better position for use in our country. The DNA-based detection device being developed in this project is a 'proof concept' of a novel design of a bio-barcode DNA amplification and nanoparticle-based electrochemical disgnosis of E. coli, E. coli 0157:H7 and Listeria monocytogenes. The detection system does not need polymerase chain reaction (PCR) amplification since the signal was amplified by the nanoparticle tracers. The oxidation potential is unique for each nanoparticle tracer and the peak current is related to its concentration. It is composed of a handheld potentiostat powered by a pocket PC and a 2-cm sup 2 electrochemical cell. Functionalization of the nanoparticles was done following the method described by Pal and Alocilja (2010). The florescence intensity of the functionalized supernatant each of that of E. coli 0157:H7 (malB) and L. monocytogenes (lis O) was lower than the pure oligo probes with PANI. This indicates good conjugation of the oligo probes to the polyaniline modified magnetic nanoparticles (PANI). The DNA probes functionalized in nanoparticles were found to be sensitive and stable. The developed probe was attached to the electrochemical sensing apparatus. It was shown that as target concentration increases, the current response of the sensor also increase. The DNA-based nanobiosensor device was constructed and initially validated in the laboratory.

Document Type

Article

Pages /Collation

69 leaves

En – AGROVOC descriptors

FOODS; HYGIENE; DNA; BIOSENSORS; PCR; ENVIRONMENT; MONITORING; SALMONELLA TYPHIMURIUM; STAPHYLOCOCCUS AUREUS; LISTERIA MONOCYTOGENES; VIBRIO CHOLERAE; PATHOGENS

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