- Top of page
- Materials and methods
- Bacterial strains, media and cultures
- RNA purification
- Reverse transcription and TaqMan assay
- Cluster analyses of gene expression data
- Definition of terminology
- Amplified fragment length polymorphism analyses
- Multilocus sequencing analyses
- Comparison of gene expression and genotype data
- Determination of the accuracy of the quantitative gene expression assay
- Effect of growth conditions on hlyA gene expression for L. monocytogenes type strains DSMZ 20600 and NTNC 7973
- Diversity of hlyA gene expression patterns
- Genotype clustering
- Comparison of gene expression and genotype
- Potential errors in the gene expression analyses
- Diverse gene expression patterns
- The lack of correlation between hlyA gene expression and the identified genotypes
- Potential for defining expression patterns for pathogenic L. monocytogenes
Aims: A major challenge for Listeria monocytogenes diagnostics is that this bacterium is ubiquitous in the environment, and that only a small fraction of the lineages are potential human pathogens. The aim of this work was to obtain a better subtyping of L. monocytogenes through utilization of combined analyses of genotype and the expression of the virulence determinant hlyA.
Methods and Results: We investigated the effect of growth temperature and medium on the hlyA expression. The gene expression levels were determined by real-time quantitative reverse transcription PCR. The expression pattern of hlyA was highly diverse among the different strains tested. The expression ranged from repression to a 1000-fold induction for growth at 42°C, as compared with 0°C. The expression patterns were compared with the corresponding genotypes. There were surprisingly low correlations between the expression patterns and the genotype clusterings.This is exemplified for the virulent type strain NTNC 7973 and non-virulent type strain DSMZ 20600. These strains are genetically nearly identical, while the hlyA gene expression patterns are very different.
Conclusions: The hlyA gene expression was highly diverse even within genetically clustered subgroups of L. monocytogenes. Consequently, the gene expression patterns can be used to further differentiate the strains within these genetic subgroups.
Significance and Impact of the Study: A major limitation in the control of L. monocytogenes is that the current tools for subtyping are not accurate enough in determining the potential virulent strains. The impact of this study is that we have developed a subtyping approach that actually targets a virulence property.