Comparison of phenotypic and genotypic methods for detection of diphtheria toxin among isolates of pathogenic corynebacteria

Abstract

We have compared molecular, immunochemical, and cytotoxic assays for the detection of diphtheria toxin from 55 isolates of Corynebacterium diphtheriae and Corynebacterium ulcerans originally isolated in five different countries. The suitabilities and accuracies of these assays for the laboratory diagnosis of diphtheria were compared and evaluated against the "gold standard" in vivo methods. The in vivo and Vero cell cytotoxicity assays were accurate in their abilities to detect diphtheria toxin but were time-consuming; however, the cytotoxicity assay is a suitable in vitro alternative to the in vivo virulence test. There was complete concordance between all the phenotypic methods. Genotypic tests based upon PCR were rapid; however, PCR must be used with caution because some isolates of C. diphtheriae possessed toxin genes but failed to express a biologically active toxin. Therefore, phenotypic confirmation of toxigenicity for the microbiological diagnosis of diphtheria is recommended.

FIG. 1

Immunoblot detection of DT from clinical isolates of C. diphtheriae. Lane 1, low-molecular-mass marker; lanes 2 to 5, whole-cell protein lysates of non-toxin-producing strains of C. diphtheriae; lanes 6 to 10, strong toxin producers; lane 11, very weak toxin producer. The toxin protein (58 kDa) was detected with a monoclonal antibody specific for the catalytic domain.