Diphtheria (Corynebacterium diphtheriae)

Corynebacteria are Gram-positive, aerobic, nonmotile, rod-shaped bacteria
related to the Actinomycetes. They do not form spores or branch as do the
actinomycetes, but they have the characteristic of forming irregular shaped,
club-shaped or V-shaped arrangements in normal growth. They undergo snapping
movements just after cell division which brings them into characteristic
arrangements resembling Chinese letters.

The genus Corynebacterium consists of a diverse group of bacteria including
animal and plant pathogens, as well as saprophytes. Some corynebacteria are part
of the normal flora of humans, finding a suitable niche in virtually every
anatomic site. The best known and most widely studied species is Corynebacterium
diphtheriae, the causal agent of the disease diphtheria.

History and Background

No bacterial disease of humans has been as successfully studied as diphtheria.
The etiology, mode of transmission, pathogenic mechanism and molecular basis of
exotoxin structure, function, and action have been clearly established.
Consequently, highly effective methods of treatment and prevention of diphtheria
have been developed.

The study of Corynebacterium diphtheriae traces closely the development of
medical microbiology, immunology and molecular biology. Many contributions to
these fields, as well as to our understanding of host-bacterial interactions,
have been made studying diphtheria and the diphtheria toxin.

Hippocrates provided the first clinical description of diphtheria in the 4th
century B.C. There are also references to the disease in ancient Syria and Egypt.

In the 17th century, murderous epidemics of diphtheria swept Europe; in Spain
"El garatillo" (the strangler"), in Italy and Sicily, "the gullet disease".

In the 18th century, the disease reached the American colonies and reached
epidemic proportions in 1735. Often, whole families died of the disease in a few

The bacterium that caused diphtheria was first described by Klebs in 1883, and
was cultivated by Loeffler in 1884, who applied Koch\'s postulates and properly
identified Corynebacterium diphtheriae as the agent of the disease.

In 1884, Loeffler concluded that C. diphtheriae produced a soluble toxin, and
thereby provided the first description of a bacterial exotoxin.

In 1888, Roux and Yersin demonstrated the presence of the toxin in the cell-free
culture fluid of C. diphtheriae which, when injected into suitable lab animals,
caused the systemic manifestation of diphtheria.

Two years later, von Behring and Kitasato succeeded in immunizing guinea pigs
with a heat-attenuated form of the toxin and demonstrated that the sera of
immunized animals contained an antitoxin capable of protecting other susceptible
animals against the disease. This modified toxin was suitable for immunizing
animals to obtain antitoxin but was found to cause severe local reactions in
humans and could not be used as a vaccine.

In 1909, Theobald Smith, in the U.S., demonstrated that diphtheria toxin
neutralized by antitoxin (forming a Toxin-Anti-Toxin complex, TAT) remained
immunogenic and eliminated local reactions seen in the modified toxin. For some
years, beginning about 1910, TAT was used for active immunization against
diphtheria. TAT had two undesirable characteristics as a vaccine. First, the
toxin used was highly toxic, and the quantity injected could result in a fatal
toxemia unless the toxin was fully neutralized by antitoxin. Second, the
antitoxin mixture was horse serum, the components of which tended to be
allergenic and to sensitize individuals to the serum.

In 1913, Schick designed a skin test as a means of determining susceptibility or
immunity to diphtheria in humans. Diphtheria toxin will cause an inflammatory
reaction when very small amounts are injected intracutaneously. The Schick Test
involves injecting a very small dose of the toxin under the skin of the forearm
and evaluating the injection site after 48 hours. A positive test (inflammatory
reaction) indicates susceptibility (nonimmunity). A negative test (no reaction)
indicates immunity (antibody neutralizes toxin).

In 1929, Ramon demonstrated the conversion of diphtheria toxin to its nontoxic,
but antigenic, equivalent (toxoid) by using formaldehyde. He provided humanity
with one of the safest and surest vaccines of all time-the diphtheria toxoid.

In 1951, Freeman made the remarkable discovery that pathogenic (toxigenic)
strains of C. diphtheriae are lysogenic, (i.e., are infected by a temperate B
phage), while non lysogenized strains are avirulent. Subsequently, it was shown
that the gene for toxin production is located on the DNA of the B phage.

In the early 1960s, Pappenheimer and his group at Harvard conducted experiments
on the mechanism of a action of the diphtheria toxin. They studied the effects
of the toxin in HeLa cell cultures and in cell-free systems, and concluded that
the toxin inhibited protein synthesis by blocking the transfer of amino acids
from tRNA to the growing polypeptide chain on the ribosome. They found that this
action of the toxin could be neutralized by prior treatment with diphtheria

Subsequently, the exact mechanism of action of the toxin was shown, and the
toxin has become