Try phylogenetic typing of L. monocytogenes.

L. monocytogenes can cause listeriosis with high fatality rates--up to 25% in large outbreaks. Subtyping isolated strains of the bacteria may help track transmission sources within food processing plants.

Scientists at Oklahoma State University examined the discriminatory power of several subtyping methods, including multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE) and ribotyping, for various virulence-related genes. They wanted to type strains of L. monocytogenes obtained from processing plants, retail raw meat and animal production facilities. The researchers performed MLST on 45 food isolates of L. monocytogenes based on genetic loci in several genes: the hemolysin (hlyA), internalin (inlA), positive regulatory factor (prfA), and actin polymerization (actA1, actA2) genes.

PCR products of 500 bp to 590 bp were amplified for each isolate, sequenced in both directions, and manually joined to form an artificial composite gene. These were compared by multiple sequence alignment and clustal analysis. Forty-five isolates were identified and differentiated into two main groups based on the degree of divergence between the strains.

To compare the discriminatory power of these approaches, the data obtained from MLST will be compared with PFGE and ribotyping for selected strains. The 13 strains selected from MLST groupings were run with PFGE using the restriction enzymes AscI and ApaI. Dendrographic groupings of the PFGE fingerprints have also been examined using clustal analysis.

The researchers indicate that DNA-based methods can define bacterial subtypes using PCR amplification and sequence analysis, or restriction digestion of bacterial DNA to generate DNA fragment banding patterns. Typing pathogenic bacteria from environmental sources involved in food processing operations may help identify strains that are persistent and which may have harborage sites within the processing facility. The scientists believe strongly that MLST, or other sequence-based typing techniques, will be the method of choice in the future.

Further information. Peter M. Muriana, 109 FAPC, Department of Animal Science, 101 Animal Science Building, Stillwater, OK 74078; phone: [phone]; fax: [phone]; email: [e-mail].