DNA microarray chip identifies, differentiates Listeria species.

We all know that L. monocytogenes is a leading cause of deaths attributed to foodborne bacterial pathogens. A lack of sufficient scientific information has led regulatory agencies to consider any strain of L. monocytogenes to be potentially pathogenic for humans, although evidence indicates that there are differences in the virulence potential among isolates of L. monocytogenes.

The long-term goal of scientists at the University of Georgia is to develop a DNA microarray chip that can differentiate and identify Listeria species, including L. monocytogenes, as well as subtype and assess the virulence potential of L. monocytogenes isolates. Their initial approach was to develop a DNA microarray chip for identifying Listeria species and for the partial serotyping and genotyping of L. monocytogenes.

In addition, the researchers evaluated the use of spacer molecules to reduce steric interference from the supporting matrix to the probes. A redundant and hierarchically structured set of oligonucleotide probes (17 mer to 37 mer) that targeted five genes (16S rRNA, iap, gltA, gltB and inlB) was designed, synthesized and spotted onto epoxy-derivatized glass slides. The target DNA was amplified from purified genomic DNA using an asymmetric polymerase chain reaction (PCR).

Results from the hybridization of the chip with target DNA from 18 L. monocytogenes strains (for which serotype and genotype data were available) revealed the array's ability to unambiguously serotype and genotype. The addition of 12-mer spacer molecules significantly increased the intensity of hybridization signals.

The DNA microarray chip will make it possible to rapidly and accurately discriminate among six Listeria species, as well as partially serotype and genotype L. monocytogenes isolates. Furthermore, with the addition of more probes that target virulence-associated genetic markers, the chip should be useful for the rapid and more accurate global assessment of the virulence potential of L. monocytogenes isolates. Such differentiation may offer an additional opportunity for the prioritization of treatments for Listeriae.

Further information. Michael Doyle, Center for Food Safety, University of Georgia, Griffin Campus, Melton Building, Griffin, GA 30223; phone: [phone]; fax: [phone]; email: [e-mail].