The plate virulence test for diphtheria.

A method has recently been described (Elek, 1948) for the detection in vitro of toxin-producing organisms including C. diphtheriae. The present paper has two purposes. The first is the examination of a larger series of strains of C. diphtheriae with a view to establishing the reliability of the procedure, for if the method is to be of any practical value for clinical purposes it must demonstrate all toxin-producing strains within a reasonably short time. The second purpose is the standardization of the ingredients used so as to yield the best results and to ensure that the conditions of the test may be accurately reproducible in different laboratories. Petrie and Steabben (1943) first suggested the use of nutrient media as a matrix for toxinantitoxin reactions. They evolved a method for the detection of toxicogenic clostridia and briefly reported some experiments with diphtheria bacilli. Ouchterlony (1948) showed that when diphtheria antitoxin was incorporated in a serum ditch plate and a toxin-producing strain streaked across, several lines were produced, one of which was caused by the toxin. For the practical test of his in vitro method he incorporated various concentrations of antitoxin in serum agar plates, as did Petrie and Steabben (1943), and each strain was tested on a series of plates with falling antitoxin concentrations. The appearance of a halo around the inoculum was regarded as positive. Two series of strains were examined by this method and by the subcutaneous guinea-pig test. In the first series of 237 strains there was a discrepancy amounting to 8.5%. Of the guinea-pig positive strains 5.1% failed to give a halo, and 3.4% of avirulent strains gave false positives. In the second series of 308 strains the discrepancy was 6.2%, and included 4.2% false positives. Elek (1948) criticized Petrie