Multi-locus variable-number tandem repeat analysis for outbreak studies of Salmonella enterica serotype Enteritidis

Salmonella enterica subsp. enterica serotype Enteritidis is known as an important and pathogenic clonal group which continues to cause worldwide sporadic cases and outbreaks in humans.

Here a new multiple-locus variable-number tandem repeat analysis (MLVA) method is reported for high-resolution subtyping of Salmonella Enteritidis. Emphasis was given on the most predominant phage types PT4 and PT8.

The method comprises of a multiplex PCR specifically amplifying repeated sequences from nine different loci followed by an automatic fragment size analysis using a multicolor capillary electrophoresis instrument. A total of 240 human, animal, food and environmental isolates of S.

Enteritidis including 23 definite phage types were used for the development and validation. Furthermore, the MLVA types of several isolates from two recent outbreaks were compared to the phage types to determine the concordance between both methods and their in vivo stability.

The in vitro stability of the two MLVA types specifically for PT4 and PT8 strains were determined by multiple freeze-thaw cycles.

Results: Seventy-nine different MLVA types have been identified in the 240 S. Enteritidis strains tested.

The Simpsonas diversity index for the MLVA method was 0.919 and Nei diversity values for the nine VNTR loci ranged from 0.07 to 0.65. Twenty-four MLVA types have been assigned to 62 PT4 strains tested and 21 types to 81 PT8 strains tested.

All outbreak isolates revealed an indistinguishable outbreak specific MLVA type. The in vitro stability experiments showed no changes of the MLVA type compared to the original isolate.

Conclusion: This MLVA method is useful to discriminate S.

Enteritidis strains even within a single phage type. It is easy in use, fast, and cheap compared to other high-resolution molecular methods and therefore an important tool for surveillance and outbreak studies for S.

Enteritidis.

Author: Burkhard Malorny, Ernst Junker and Reiner Helmuth
Credits/Source: BMC Microbiology 2008, 8:84

Published on: 2008-05-30

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