IV. INVESTIGATION OF C. DIPHTHEHZAE
145
degrees of consistency; they tend to break into fragments (gravis, atypical
corynebacteria), or show a soft buttery consistency. They are easily
homogenised (mitis, intermedius, C . ulcerans).
( c ) Enzymic characters. Carbohydrate fermentation: glucose, sucrose,
maltose and laevulose are used for species differentiation. Dextrine,
starch and glycogen are used to define C . diphtheriae biotypes (gravis,
mitis, intermedius) (see above). Sugar fermentations are performed in test
tubes on Hiss medium with Andrade’s indicator.
A micromethod, Corynetest, using carbohydrate impregnated discs,
was recently recommended (Ionescu-Stoian et al., 1977).
Urease test is perfwmed on Blake-Christensen’s medium; this test is
negative for C . diphtheriae and the variant with dwarf colonies and positive
for C . ulcerans; 75-90y0 of diphtheroids display positive reaction.
Cystinase test is performed in stab cultures in tubes of 10/100 mm on
Pisu’s modified medium. The test is positive in 24 h (starting from the
fifth hour of growth at 37°C) for C . diphtheriae and C . ulcerans. A positive
reaction consists of a black to brown halo along the streaked culture.
Diphtheroids, atypical corynebacteria and the variant with dwarf
colonies are always negative or produce a blackening of the medium
surface only.
(d) Slide agglutination. For the rapid confirmation of C . diphtheriae a
polyvalent antidiphtherial serum for slide agglutination is recommended.
Positive agglutination is specific for C . diphtheriae (87%). Spontaneously
agglutinable or inagglutinable strains can also occur (13%). Therefore it
should be stressed that a negative slide agglutination is not always contrary
to a C . diphtheriae diagnosis.
Variants of C . diphtheriae or C . ulcerans with dwarf colonies may also
agglutinate, but inconstantly. No other species of Corynebacterium agglutinate with antidiphtherial polyvalent serum.
Slide agglutination can be performed also with the isolated colonies
from Tinsdale medium as a supplementary test for a rapid identification
of C . diphtheriae.
(e) Toxigenicity. Toxigenicity tests present different degrees of sensitivity :
the in vitro Elek-Ouchterlony-Frobisher test is a qualitative test showing a
high sensitivity (98%); tissue cultures and rabbit skin i.d. test are quantitative tests. They can detect 1/500 and 1/1000-1/2000 DLM respectively.
Guinea pig tests detect at least 1 DLM.
(i) In vitro tests. Elek-Ouchterlony-Frobisher test. The most commonly
used test, economical, reproducible and easy to perform, is the ElekOuchterlony-Frobisher immunodiffusion test (EOF). It is based upon
6
145
degrees of consistency; they tend to break into fragments (gravis, atypical
corynebacteria), or show a soft buttery consistency. They are easily
homogenised (mitis, intermedius, C . ulcerans).
( c ) Enzymic characters. Carbohydrate fermentation: glucose, sucrose,
maltose and laevulose are used for species differentiation. Dextrine,
starch and glycogen are used to define C . diphtheriae biotypes (gravis,
mitis, intermedius) (see above). Sugar fermentations are performed in test
tubes on Hiss medium with Andrade’s indicator.
A micromethod, Corynetest, using carbohydrate impregnated discs,
was recently recommended (Ionescu-Stoian et al., 1977).
Urease test is perfwmed on Blake-Christensen’s medium; this test is
negative for C . diphtheriae and the variant with dwarf colonies and positive
for C . ulcerans; 75-90y0 of diphtheroids display positive reaction.
Cystinase test is performed in stab cultures in tubes of 10/100 mm on
Pisu’s modified medium. The test is positive in 24 h (starting from the
fifth hour of growth at 37°C) for C . diphtheriae and C . ulcerans. A positive
reaction consists of a black to brown halo along the streaked culture.
Diphtheroids, atypical corynebacteria and the variant with dwarf
colonies are always negative or produce a blackening of the medium
surface only.
(d) Slide agglutination. For the rapid confirmation of C . diphtheriae a
polyvalent antidiphtherial serum for slide agglutination is recommended.
Positive agglutination is specific for C . diphtheriae (87%). Spontaneously
agglutinable or inagglutinable strains can also occur (13%). Therefore it
should be stressed that a negative slide agglutination is not always contrary
to a C . diphtheriae diagnosis.
Variants of C . diphtheriae or C . ulcerans with dwarf colonies may also
agglutinate, but inconstantly. No other species of Corynebacterium agglutinate with antidiphtherial polyvalent serum.
Slide agglutination can be performed also with the isolated colonies
from Tinsdale medium as a supplementary test for a rapid identification
of C . diphtheriae.
(e) Toxigenicity. Toxigenicity tests present different degrees of sensitivity :
the in vitro Elek-Ouchterlony-Frobisher test is a qualitative test showing a
high sensitivity (98%); tissue cultures and rabbit skin i.d. test are quantitative tests. They can detect 1/500 and 1/1000-1/2000 DLM respectively.
Guinea pig tests detect at least 1 DLM.
(i) In vitro tests. Elek-Ouchterlony-Frobisher test. The most commonly
used test, economical, reproducible and easy to perform, is the ElekOuchterlony-Frobisher immunodiffusion test (EOF). It is based upon
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