Purpose: The objectives of this research are to develop a DNA microarray containing genomic DNA fragments of Listeria monocytogenes, to verify its diagnostic ability using a laboratory medium, and to apply the microarray to detect L. monocytogenes in milk.
Methods: Sixty genomic DNA fragments of L. monocytogenes digested using several pairs of restriction enzymes were randomly selected and affixed to a slide glass to fabricate the DNA microarray. The diagnostic ability of the microarray was verified using seven strains of L. monocytogenes, seven strains of other Listeria spp., and thirteen strains of foodborne pathogens in different genera. Finally, the diagnostic ability of the DNA microarray in milk was confirmed and the sample preparation procedure was established.
Results: DNA microarray was able to detect L. monocytogenes and distinguish it from other Listeria spp. and other foodborne pathogens in a laboratory medium. The detection limits of the microarray were approximately 8 log CFU/ml when other types of bacterial strains (B. cereus, S. Montevideo, Bacillus spp., P. aeruginosa) were present in high number in milk. For the enrichment at 37 °C, UVMB was better than TSB or BHIB in increasing the population of L. monocytogenes. When milk containing L. monocytogenes (2.2 log CFU/ml) and other five types of bacterial strains (> 2.0 log CFU/ml each) was enriched in UVMB at 37 °C for 24 h, the populations of L. monocytogenes increased to 7.7 log CFU/ml. When genomic DNAs extracted from these enriched suspensions were hybridized on the microarrays, L. monocytogenes were successfully detected.
Significance: The microarray containing non sequenced genomic DNA fragments was able to detect L. monocytogenes in milk without any interference of other microbes.