Discovery of Natural Atypical Nonhemolytic Listeria seeligeri Isolates

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Abstract
We found seven Listeria isolates, initially identified as isolates with the Xyl + Rha − biotype of Listeria welshimeri by phenotypic tests, which exhibited discrepant genotypic properties in a well-validated Listeria species identification oligonucleotide microarray. The microarray gives results of these seven isolates being atypical hly -negative L. seeligeri isolates, not L. welshimeri isolates. The aberrant L. seeligeri isolates were d -xylose fermentation positive, l -rhamnose fermentation negative (Xyl + Rha − ), and nonhemolytic on blood agar and in the CAMP test with both Staphylococcus aureus (S − reaction) and Rhodococcus equi (R − reaction). All genes of the prfA cluster of L. seeligeri , located in the prs-ldh region, including the orfA2 , orfD , prfA , orfE , plcA , hly , orfK , mpl , actA , dplcB , plcB , orfH , orfX , orfI , orfP , orfB , and orfA genes, were checked by PCR and direct sequencing for evidence of their presence in the atypical isolates. The prs-prfA cluster- ldh region of the L. seeligeri isolates was approximately threefold shorter due to the loss of orfD , prfA , orfE , plcA , hly , orfK , mpl , actA , dplcB , plcB , orfH , orfX , and orfI . The genetic map order of the cluster genes of all the atypical L. seeligeri isolates was prs-orfA2-orfP-orfB-orfA-ldh , which was comparable to the similar region in L. welshimeri , with the exception of the presence of orfA2 . DNA sequencing and phylogenetic analysis of 17 housekeeping genes indicated an L. seeligeri genomic background in all seven of the atypical hly -negative L. seeligeri isolates. Thus, the novel biotype of Xyl + Rha − Hly − L. seeligeri strains can only be distinguished from Xyl + Rha − L. welshimeri strains genotypically, not phenotypically. In contrast, the Rha + Xyl + biotype of L. welshimeri would not present an identification issue.