Submitted to: Veterinary Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/17/2002
Publication Date: 1/20/2002
Citation: Borucki, M.K., Krug, M.J., Muraoka, W.T., Call, D.R. 2002. Discrimination among listeria monocytogenes isolates using a mixed genome DNA microarray. Veterinary Microbiology. 92:351-362. Interpretive Summary: A Listeria monocytogenes DNA array was constructed and used for phylogenetic reconstruction, strain typing, and identification of genetic sequences that may be specific to clinical strains of L. monocytogenes. This work represents the first description of a mixed genome microarray and its application in comparative genomics. While this array is specific to Listeria, the methods we describe for developing and using the array are relatively simple to implement and should be universal for any bacteria.
Technical Abstract: Listeria monocytogenes can cause serious illness in both humans and domestic livestock. Therefore, epidemiological investigations require subtyping methods that are rapid, discriminatory, and reproducible. Furthermore, genetic markers are needed to differentiate environmental versus potential pathogenic strains of Listeria. To address both of these issues, we constructed a 585 spot microarray from a mixed genomic library. By hybridizing genomic DNA from 29 strains of Listeria to the array and comparing the number of shared probes between strains, we were able reconstruct previously described phylogenetic relationships between major serogroups and we showed consistent classification within PFGE groups. Cluster l consisted of 1/2a and 1/2c isolates, all of which shared four unique probe sequences. This cluster could be further subdivided into two sub-clusters, one of which (cluster 1b) included all 1/2a and 1/2c clinical isolates. Cluster ll consisted of clinically relevant 1/2b and 4b serotypes as well as 4e and 4c and all members shared a single unique probe sequence. Importantly, all members of clusters 1b and 11 shared six unique gene sequences suggesting a possible relationship between clinical isolates from these two phylogenetically divergent serogroups. Sequencing 15 probes revealed redundancy between only two of the probes and BLAST searches identified 13 putative matches with previously described bacterial genes. These results demonstrate that mixed genomic microarrays are relatively simple to construct and are very powerful tools for identifying genetic markers. These arrays may also prove to be very valuable for making phylogenetic, phylogeographic, and of bacteria.