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. 2010 Oct 23:10:269.
doi: 10.1186/1471-2180-10-269.

Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system

Sascha Al Dahouk  1 , Holger C Scholz, Herbert Tomaso, Peter Bahn, Cornelia Göllner, Wolfram Karges, Bernd Appel, Andreas Hensel, Heinrich Neubauer, Karsten Nöckler
Affiliations

Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system

Sascha Al Dahouk et al. BMC Microbiol. .

Abstract

Background: A commercial biotyping system (Taxa Profile™, Merlin Diagnostika) testing the metabolization of various substrates by bacteria was used to determine if a set of phenotypic features will allow the identification of members of the genus Brucella and their differentiation into species and biovars.

Results: A total of 191 different amines, amides, amino acids, other organic acids and heterocyclic and aromatic substrates (Taxa Profile™ A), 191 different mono-, di-, tri- and polysaccharides and sugar derivates (Taxa Profile™ C) and 95 amino peptidase- and protease-reactions, 76 glycosidase-, phosphatase- and other esterase-reactions, and 17 classic reactions (Taxa Profile™ E) were tested with the 23 reference strains representing the currently known species and biovars of Brucella and a collection of 60 field isolates. Based on specific and stable reactions a 96-well "Brucella identification and typing" plate (Micronaut™) was designed and re-tested in 113 Brucella isolates and a couple of closely related bacteria.Brucella species and biovars revealed characteristic metabolic profiles and each strain showed an individual pattern. Due to their typical metabolic profiles a differentiation of Brucella isolates to the species level could be achieved. The separation of B. canis from B. suis bv 3, however, failed. At the biovar level, B. abortus bv 4, 5, 7 and B. suis bv 1-5 could be discriminated with a specificity of 100%. B. melitensis isolates clustered in a very homogenous group and could not be resolved according to their assigned biovars.

Conclusions: The comprehensive testing of metabolic activity allows cluster analysis within the genus Brucella. The biotyping system developed for the identification of Brucella and differentiation of its species and biovars may replace or at least complement time-consuming tube testing especially in case of atypical strains. An easy to handle identification software facilitates the applicability of the Micronaut™ system for microbiology laboratories.

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Figures

Figure 1
Figure 1
Cluster analysis of Brucella reference strains based on biochemical reactions. Cluster analysis of the 23 Brucella reference strains based on 570 (A) and 93 (B) biochemical reactions tested with the Taxa Profile™ system (plate A, C, and E) and the newly developed Brucella specific Micronaut™ microtiter plate, respectively. Hierarchical cluster analysis was performed by the Ward's linkage algorithm using the binary coded data based on the empirically set cut-off.
Figure 2
Figure 2
The Brucella specific Micronaut™ microtiter plate. Design of the newly developed Brucella specific Micronaut™ microtiter plate including 93 selected substances.
Figure 3
Figure 3
Cluster analysis of Brucella field isolates based on biochemical reactions. Cluster analysis of 113 Brucella strains including the reference strains and two isolates of a potentially new species that originated from Austrian foxes based on 93 biochemical reactions tested with the newly developed Brucella specific Micronaut™ microtiter plate. Hierarchical cluster analysis was performed by the Ward's linkage algorithm using the binary coded data based on the empirically set cut-off.
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References

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