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rare to see any gonococci outside the pus-cells. A cell may be so
filled with them as to lose all its characteristic structure and appear
only as a clump of cocci. The relation of the bacteria to the pus-
corpuscles is regarded by some as evidence of its activity, by others
as an illustration of the protective action of the phagocytes.

The gonococci are stained well with methyl-blue. They cfo not
adapt themselves to the Gram method, as the iodide of potassium
deprives them of their color. Neisser recommends the following


method : A cover-glass having been prepared with a layer of the
fluid to be examined in the usual way, it is treated for a few
minutes with a concentrated alcoholic solution of eosin, the
action of which is reinforced by heat. The excess of eosin
being removed by blotting-paper, a concentrated alcoholic solu-
tion of methyl-blue is next applied for fifteen seconds, and then
washed off with distilled water. The cocci are now seen colored
blue, while the protoplasm of the leucocytes is stained a delicate
pink and their nuclei blue (Fig. 12).

The gonococci do not grow on any of the ordinary culture-
media, such as gelatin or agar or potatoes. Even on the media on
which they do develop they are so
frequently mixed up with other
forms that the latter grow rapidly
and present appearances which
make it difficult to distinguish
them from the genuine gonococci.

Bum m has succeeded, however,
in making them grow on human
blood-serum, but this growth is
accomplished with considerable
difficulty. The materials used
must, in the first place, be as free
as possible from other organisms,
otherwise the latter will outgrow
the coccus. The gonorrheal pus,
containing the organism in large

numbers, must be placed on the surface of the blood-serum in
drops of considerable size. Scratch- or stab-cultures are of no
value. The test-tube must be placed in an oven at a tempera-
ture of from 33 to 37 C. The growth forms a delicate film
with well-defined, irregular borders. It appears like a layer
of varnish upon the top of the serum. When somewhat thicker
it has a grayish-white or a slightly brownish tinge. The growth
is slow and scant in amount. At the end of two or three days the
cocci begin to die off, and the culture must therefore be often trans-
planted if it is desired to preserve the organisms. As nothing
further will be said about this disease, it may be well to study here
the action of the gonococci in the human epithelium.

For some time it was thought that sufficient proof had not been
afforded of the specific character of the gonococcus. There is no
one sinsfle characteristic which distinguishes this organism from all

V. *

FIG. 12. Gonococci.


others, but the combination of peculiarities which have just been
mentioned is such as is not found in other forms of bacteria.
These peculiarities are the diplococcus or " breakfast-roll " shape,
the characteristic arrangement of the organism in the pus-cells,
the bleaching caused by Gram's solution of iodide of potassium,
and the difficulty of cultivation on ordinary media.

Proof positive has been afforded, however, by several experi-
menters of its contagious character. Buinm transplanted the
twentieth generation of a gonococcus culture to the urethra of a
bedridden paralytic, and produced a typical gonorrhoea. This
experiment has recently been made upon the healthy urethrse of
medical students. Bumm also examined the different stages of the
gonorrhoeal inflammation in the conjunctiva of new-born infants.
Twenty-six fragments were taken from the conjunctiva! mem-
brane at periods of the disease varying from thirty-six hours to
thirty-two days. He found that the cocci, once having entered
the conjunctival sac, reproduce themselves rapidly in the secre-
tions, next invade the epithelial layer, and finally force their way
down to the papillary layer. On the second day an enormous
immigration of leucocytes takes place into the invaded layer of
epithelium and the surrounding cells, pushing the epithelial cells,
so as to lift them from their bed. On the papillary layer thus
exposed there forms an exudation of a fibre-cellular character in
which are clumps and rows of growing cocci. The bacterial
growth does not invade the deeper tissue ; it does not go beyond
the most superficial of the sub-epithelial layers. A regeneration
X)f the epithelium soon covers over the denuded spots, and the
cocci, after growing for some time longer on the surface, gradually
disappear. It is only on certain types of mucous membrane that
these organisms will grow namely, those which possess a cylin-
der epithelium or one closely allied to it. These are the mem-
branes of the male and female urethra, the uterus, Bartholin's
glands, and the conjunctiva. The more deep-seated secondary
inflammations, such as involve the prostate, the epididymis, the
testicles, the uterus, and the tubes, are frequently due to the pres-
ence of some of the pyogenic bacteria ; but suppurative inflamma-
tion of both tubes and ovaries has been found to be due largely to
the presence of the gonococcus. The aureus has been found as a
frequent companion of the gonococcus in the urethral discharge
and in the pus from gonorrhoeal buboes. The metastatic inflamma-
tion of joints and the endocarditis which occur as sequelae of
gonorrhoea have been supposed to be due tv the presence of


pyogenic cocci, but Councilman and others have shown that the
gonococcus may be the sole organism concerned in the inflamma-
tory process.

Why the gonococcus should grow only in the superficial layers
of certain mucous membranes, and nowhere else in the body,
has not yet been satisfactorily explained. The most plausible
theory seems to be that inasmuch as the gonococci possess a very
marked preference for oxygen, they find a better culture-soil in the
epithelium than in the subjacent connective tissue. Bumm has
shown that injections of pure gonorrhoeal discharge or of pure
cultures of the gonococcus into the subcutaneous tissue do not
produce suppuration. That this loss of activity is not explained
by the action of phagocytes is shown by the fact that the organ-
isms, when the tissues are examined later, are not taken up by
the cells of the part, but are nearly all to be found outside the

After the gonococci have existed for a certain length of time in
the epithelium of the part, they disappear spontaneously in a
certain number of cases. This disappearance is accounted for by
the casting off of the cylinder epithelium during the inflammatory
processes, and its replacement by a pavement epithelium which
resists the efforts of the cocci to penetrate it. In this way proper
nutriment gradually fails them and they die out. It is generally
accepted that cure is effected in this way rather than by the phago-
cytes, for such leucocytes as are invaded by the gonococci are
destroyed by the latter during the active growth and multiplication
of the organisms which take place in the protoplasm of those cells.

Streptococcus erysipelatis in all respects so closely resembles
the streptococcus pyogenes that the majority of bacteriologists are
unable to detect any constant differences between them either by
the microscope or by culture. The description of the organism
coincides with that already given to the streptococcus : therefore it
is needless to repeat it here. Rosenbach undertakes to recognize
certain distinguishing marks between the two. He thinks the
cocci and the chains of the erysipelas coccus are larger than those
of the pyogenic coccus. His delineations of the culture show a
growth of the erysipelas coccus more transparent and more
irregular and nodular in outline than is seen in the cultures of the
other organisms. The brownish tint of the culture is also want-
ing. The weight of evidence at the present time is, however, in
favor of the identity of the two organisms. The question is dis-
cussed more at length in the chapter on Erysipelas.


Bacillus Tctani. The first observations on the nature and origin
of this organism were made in 1884 by Nicolaier, who found a
bacillus in garden soil, and who succeeded in producing tetanus
in mice, guinea-pigs, and rabbits by injecting the soil into them
subcutaneously. The same organisms were found in the diseased
animals, but there was great difficulty in obtaining a pure culture
of the bacilli, thus giving conclusive evidence of its power to
produce the disease. This culture was finally accomplished in
1889 by Kitasato, who planted on a suitable culture-soil a frag-
ment of tissue from the neighborhood of a wound in a man
dead of the disease. He found that the spores of this bacillus
germinated before those of the other forms of bacilli mixed with
it. As soon as these spores had formed he subjected the culture
to a temperature of 80 C., which killed off all bacteria; con-
sequently, the spores of the tetanus bacillus alone, remained, and
a pure culture of this organism was obtained as soon as the bacilli
had developed from them. The spores are found in garden soil,
in masonry, in decomposing liquids, and in manure.

The tetanus bacillus is a large slender rod with somewhat
rounded ends. It resembles the bacillus of mouse septicaemia, but

v \


FIG. 13. Bacillus Tetani.

is longer: in fact, it sometimes grows into long chains which show
very imperfectly the lines of division. The spore-formation takes
place at the end of the bacillus, and, as it enlarges the cell
considerably, gives it a "pin" or "drumstick" shape (Fig. 13).
The spore germinates at a temperature of 37.5 C. in thirty hours;
in the temperature of a house, in about a week. It is motile, and
belongs to the strictly anaerobic organisms, rapidly dying when
exposed to the air. It is readily colored by* methyl-blue and



fuchsin, and is brought out very perfectly by the Gram method.
It can be cultivated in gelatin mixed with grape-sugar, which aids
in its rapid development. The upper portions of the gelatin
remain sterile, but in the lower portions of the puncture there is
an active bacterial growth which sends out innumerable little pro-
longations, giving to the culture the appearance of an inverted fir
tree. After the first week the gelatin begins to liquefy and to
obscure the peculiar features of the growth, until, finally, the
gelatin is changed into a whitish-gray, tenacious, shining mass.
To obtain cultures of the tetanus bacillus from cases of trau-
matic tetanus in man or from experiment animals the following
method may be employed, which is a modification by Frothingham
of Kitasato's method :

Inoculate tubes of decidedly alkaline bouillon with pus from the wound or
point of inoculation. If there is no pus, small fragments of tissue are snipped
from the region of the wound and used for this purpose. The tubes should
now be placed in an atmosphere of hydrogen at a temperature of from 37 to
39 C. At the end of forty eight hours a microscopic examination may be
made, and if the tetanus bacilli are found, the 'tubes are to be heated for
three-quarters of an hour to one hour in a water-bath previously heated to
80 C. From these heated tubes fresh alkaline bouillon may be inoculated.

FIG. 14. Hydrogen Jar for Anaerobic Cultures. The stop-cock on the right allows the
air to escape from the jar, while the hydrogen is passed in from the left.

These fresh cultures may be allowed to develop under hydrogen at a temper-
ature of 37 C. for forty eight hours (Fig 14). Pure cultures should be obtained
in this way, the purity of the culture being verified by microscopic examina-
tion and growth on solid culture-media, and the virulence being determined
by inoculation experiments.

Brieger has obtained from cultures a toxine which he named
"tetanin," and in addition " tetanotoxin " and " spasmotoxin,"
all of which, when injected into animals, produce convulsive
movements and, finally, paralysis. Inasmuch as the same group


of symptoms were obtained by the toxines as were obtained by the
bacilli, and as the latter are hard to find in the blood and internal
organs of individuals affected with tetanus, it has been thought
probable that the symptoms of the disease are largely produced by
these chemical substances.

Although Rosenbach and Shakspeare have stated that the
bacilli are to be seen in the central nervous system, subsequent
observers have not been able to find them, and it is probable, there-
fore, that the convulsions are produced by the tetanin elaborated
by these organisms.

Bacillus Tuberculosis. Although experiments were made as
early as 1865 by Villemin to prove the inoculability of tubercle,
and as Cohnheim in the following decade decided that tuberculosis
was a specific infectious disease, it was not until 1882 that Bauni-
garten and Koch simultaneously discovered the organism which
causes the disease. Baumgarten should receive credit for first
having seen the bacillus with the microscope, but it remained for
Koch to cultivate it successfully and by inoculation to prove beyond
question its right to be considered the cause and only cause of

The tubercle bacilli are small and thin rods about 2 to 4/>< in
length ; that is, about one-quarter to three-quarters the length of
the diameter of a red blood-corpuscle. The ends of the rods are
generally slightly rounded, and are usually slightly bent near the
middle or are more or less curved. In artificial cultures the rods
are a little smaller than when growing in the tissues. The longest
rods are usually seen in phthisical sputa. They are generally
single, occasionally being found in pairs arranged like a V, and
sometimes several are strung together. They do not possess the
power of motion. Whether spore-formation takes place is unde-
termined, although Baumgarten thinks it highly probable that
it does occur, as a cheesy material, in which it is impossible to
demonstrate the bacilli by any method of staining, when inocu-
lated into animals produces the disease. Free spores have never
been seen, nor have the bacilli been observed in the act of spore-
formation. In the fresh state none of those bright, glistening
spots are seen which are characteristic of spores. When colored,
the bacilli exhibit, placed in regular order, bright spots which are
very suggestive of spores. The expectorations can be kept months,
and even years, in a dried state without destroying the vitality of
the bacilli. The acids of the stomach and the products of decom-
position have no effect upon them. Pure culttfres of bacilli have


been mixed with the food of animals, and have thus been passed
through the digestive tract without any effect upon their vitality.
This durability seems to be due to the unusually tough cell-wall
which the bacillus possesses. The organism is a facultative
anaerobic ; that is, it may grow without oxygen, although it pre-
fers to grow with oxygen.

This is one of the few bacteria which have a pathognomonic
stain. Though taking the ordinary watery and alcoholic aniline
stains with difficulty, yet when properly stained it does not give
up its coloring material even in the presence of mineral acids a
property which the bacillus of leprosy alone holds in common with

The following is a convenient method (Ziehl) of examining the
sputa :

The sputum selected is spread out upon a glass with a dark background to
enable one to detect the various details, such as the fragments of the diseased
lung, the secretions of the upper air-passages, and the saliva. The bacilli
are usually found in the lung-fragments, which are small, tough, yellow
clumps floating in the saliva. One of these clumps is removed by the steril-
ized platinum needle and placed upon a cover ; a second cover-glass is then
placed upon the first, and the specimen is gently pressed between the two so
as to form a thin layer, whereupon the glasses are separated by a sliding
motion and are allowed to dry in the air. To complete this process the glass
to be stained is rapidly passed three times through a flame. A few drops of
carbolic fuchsin l are allowed to trickle over the glass, and it is held over the
flame until the coloring fluid partially evaporates. More staining fluid is
now added, and the heating repeated until a satisfactory coloring is obtained,
or the coloring fluid containing the specimen can be placed in a watch-glass
and heated for a few moments over a water-bath. The specimen is then
washed with distilled water. To decolorize the surrounding cells and other
forms of bacteria a strong decolorizing agent must be used, as, for example,
a 5 to 10 per cent, solution of sulphuric acid. The glass is moved up and
down in this solution until the deep-red color becomes a yellowish brown.
Next place the glass in 70 per cent, alcohol to wash out the dissolved fuchsin.
Wash with distilled water and color again with ordinary watery solution of
methyl-blue. Wash, finally, with distilled water and examine, wet or dry,
the specimen, and mount it permanently in Canada balsam.

The specimen is examined to the best advantage when wet, as
the bacilli are not so much shrivelled as when mounted in Canada
balsam. The tubercle bacilli will be found colored red, and any
other bacteria which happen to be present, and which have been
deprived of their red color by the acid, are stained blue, so that
the different kinds can thus readily be distinguished from one
another (Figs. 15, 16). The method may be simplified by dipping

1 Fuchsin lo parts in 100 parts of a saturated aqueous solution of carbolic acid.


the red-colored specimens into a solution in which the acid and


FIG. 15. Tuberculous Sputum.

the methyl-blue are both present: water, 50 parts, alcohol 30
parts, nitric acid 20 parts, and methyl-blue to saturation. This

FIG. 1 6. Tuberculous Urine.

simplifies the process somewhat. Sections are colored very much
in the same way :

Place the section for half an hour in a dish of carbolic fuchsin ; allow it
to float for one minute in a 5 per cent, solution of sulphuric acid ; wash in 60
per cent, alcohol. Next stain with methyl-blue for two or three minutes.
Wash in water and weak alcohol, dehydrate in absolute Alcohol, and, having
cleared it in oil of cedar, mount in Canada balsam.


Although for clinical work the short methods may be used in the
hands of experts, still it must be remembered that the Ziehl
solution stains a number of spores, which, unless recognized,
may prove a source of error. It is not generally known that
under certain conditions for example, age the bacilli may not
be stained by the quick methods. When stained for twenty-
four hours according to the now nearly-forgotten Koch-Ehrlich
method the bacilli are well defined. For this reason the Koch-
Ehrlich method is given. It should however, be remembered that
this method shows crystalline forms which may be mistaken for

Place the section in aniline-water fuchsin for twenty-four hours ; decolor-
ize in a 25 per cent, solution of nitric acid ; wash in 60 per cent, alcohol ;
place in watery methyl-blue for a few moments ; wash and mount.

Under the microscope is seen the miliary tubercle consisting of
leucocytes and epithelial cells, and a giant-cell in or near the
centre of the growth. The bacilli are found lying in small
numbers between the leucocytes and in the giant-cell. The nuclei
of the giant-cell appear to be arranged in a radiating manner at its
periphery, as do also the bacilli. This arrangement is due to the
fact that the centre of the cell has undergone degeneration and its
contents at this part have disappeared. This appearance is quite
characteristic of the tubercular giant-cell, and distinguishes it
from the giant-cell of sarcoma (Fig. 76). The degenerative pro-
cess is seen also in the other cells at the centre of the tubercle,
while new cells and bacilli are seen on the borders. In this way
the growing tubercle undergoes a cheesy degeneration at its
centre. If the disease at this stage is on the surface of the skin or
a membrane, ulceration will occur.

The growth of the organisms is exceedingly slow, and takes
place at the temperature of the human body, and very slight
deviations from this point are likely to arrest their development.

Koch devised expressly for this organism the hardened blood-
serum. Nocard and Roux have suggested a combination of agar
with glycerin, upon which it grows even better, as the bacillus
seems to have a predilection for glycerin, and this being also
much more easily sterilized. When cultivated in the test-tube on
agar thus prepared, a well-developed growth is procured at the end
of fourteen days, while on blood-serum from one to two weeks
more must pass before the culture reaches the same point. It then
appears as thick crusts of a dull grayish-white color, which crusts



are very dry and brittle and are made up of minute scale-like
masses (Fig. 17). If the growth meets a drop of condensed moist-
ure, it will form a thin film over the latter, without
in the least disturbing the clearness of the fluid.

To obtain materials with which to make a series
of pure cultures tuberculous sputa may be injected
into a guinea-pig. When tuberculosis is established,
the animal should be killed, and a fragment of tu-
bercle taken, with due precautions, from the lung
and placed upon the culture-soil. After develop-
ment takes place the fragment of lung can be seen
under a low power, and surrounding it are seen
S-shaped, wavy, and scroll-like masses of bacilli.

The tubercle bacillus is not found growing out-
side the living tissues of man and animals, the
necessary conditions of nutrition and temperature
not existing elsewhere. They must be regarded,
therefore, as true parasitic organisms. Although
they are unable to grow around us, their great power
1 of resistance permits of their being preserved for a
I long time in a dried state mixed with dust, and of
taking on an active growth whenever an opportunity
occurs for them to become grafted again upon the
living tissues.

This inoculation may take place on the skin fol-
lowing slight blows or bruises or cuts. The hands
of attendants on the sick may be cut with a glass
containing sputa. Anatomical tubercle is an exam-
ple of this form of contagion. The disease known
as "lupus" is but a variety of tuberculosis of the
skin. As has been seen, the bacillus is extremely
resistant to the action of the digestive fluids, and
animals experimentally fed with this organism have
succumbed to a general tuberculosis. Whether in-
oculation can take place through the uninjured mu-
cous membrane has not been demonstrated, but it is
probable that if bacteria can penetrate the uninjured skin, they can
also work their way through a normal mucous membrane. As a
rule, the mesenteric glands are found first affected, and afterward
the mucous membrane a sequence which is at least suggestive that
the membrane was previously in a healthy condition. Later, the
spleen and liver are found infected. A very practical deduction from

FIG. 17. Bacillus
of Tuberculosis
on glycerin -


these experiments is the necessity for the supervision of food, par-
ticularly the milk of tuberculous cows. It is now well known that
the organisms are found in the milk. H. C. Ernst has shown that
six drops of infected milk injected subcutaneously into a guinea-
pig will produce a general tuberculosis even though there be no
manifestations of disease in the udder.

The question of an infection through the respiratory tract
appears to be a disputed one. According to Baumgarten, experi-
mental work seems to point against such mode of entrance into the
system. Frankel, who writes with the authority of Koch behind
him, takes the opposite view, and believes that breathing infected
air is the most frequent mode of acquiring the disease. Experi-
ment shows that the disease appears first at the point of infection,
and therefore is at first local. The frequency of the disease in the
lungs surely points strongly to the respiratory tract as the route
through which infection takes place. Inasmuch as it has been
proved that bacilli can float in the air when dry, it is probable that
they are in this way conveyed from one individual to another.

Online LibraryJohn Collins WarrenSurgical pathology and therapeutics → online text (page 5 of 84)