Optimization of PCR Detection of Listeria Spp. and Listeria Monocytogenes in Vegetable Salad

Ng, Soo Ching (2016) Optimization of PCR Detection of Listeria Spp. and Listeria Monocytogenes in Vegetable Salad. Final Year Project (Bachelor), Tunku Abdul Rahman University College.

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Abstract

Listeria monocytogenes is a foodborne pathogen with the capacity to cause severe illness called listeriosis with high fatality rates compared with other foodborne pathogens. Over the years, minimally processed ready-to-eat (RTE) vegetables and fruits, the high risk foods to L. monocytogenes contamination, have become an established product in worldwide markets. A sensitive and rapid detection of Listeria spp. and L. monocytogenes is needed to improve as it is very important with regard to food safety. The main objective of this study is to optimize the PCR detection of Listeria spp. and L. monocytogenes in RTE vegetable salad. Three parameters were optimized to increase the efficiency of PCR detection which included the annealing temperature (50C to 60C), magnesium ion concentration (1.5 mM to 3.5 mM), and primer concentration (0.05 μM to 1.0 μM). Two pairs of primer were used to target the hlyA gene and 16S rRNA which produced amplicons at 702 bp and 938 bp for each targeted bacteria, L. monocytogenes and Listeria spp., respectively. From the results obtained we can conclude that the optimum annealing temperature was 53C. The optimum MgCl2 concentration and primer concentration for PCR concurrently detection of Listeria spp. and L. monocytogenes were 1.5 mM and 0.5 M, respectively Key words: PCR, optimization, RTE, vegetable salad, Listeria monocytogenes