It is necessary to transfer the bacteria to fresh gelatine about every six weeks, lest the products of growth and decay given off by the organisms destroy them.

CHAPTER XII.

CULTIVATION OF ANAEROBIC BACTERIA.

FIG. 38.

SPECIAL methods are necessary for the culture of the anærobic variety of bacteria in order to procure a space devoid of oxygen. Liborius's High Cultures. The tube is filled about full with gelatine, which is then steamed in a water bath and allowed to cool to 40° C., when it is inoculated by means of a long platinum rod with small loop, the movement being a rotary vertical one, and the rod going to the bottom of the tube.

The gelatine is next quickly solidified under ice; very little air is present. The anærobic germs will grow from the bottom upward, and any ærobins present will develop first on top, this method being one of isolation.

From the anaerobic germs grown in the lower part, a stab culture is made into another tube containing gelatine, the material being obtained by breaking test-tube with the culture.

Hesse's Method. A stab culture having been made with anaerobic germs, gelatine in a semi-solid condition is poured into the tube until it is full, thus displacing the air. (Fig. 39.)

Liborius's

method.

Esmarch's Method. Having inoculated a tube with the microbe, the gelatine is rolled out on the walls of the tube, a roll culture," and the rest of the interior filled with gelatine, the tube being held in ice water. The colonies develop upon the sides of the tube and can be examined microscopically.

Gases like Hydrogen to replace the Oxygen. Several arrangements for passing a stream of hydrogen through the culture:

Fränkel puts in the test tube, a rubber cork containing two glass tubes, one reaching to the bottom and connected with a hydrogen apparatus, the other very short, both bent at right angles. When the hydrogen has passed through ten to thirty minutes, the short tube is annealed and then the one in connection with the hydrogen bottle, and the gelatine rolled out upon the walls of the tube. (Fig. 40.) Hüppe uses eggs as described in Chapter IX.

Use of Ærobic Bacteria to remove the Oxygen. Roux inoculates an agar tube through a needle thrust after which semisolid gelatine is poured in on top. When the gelatine has solidified, the surface is inoculated with a small quantity of bacillus subtilis or some other ærobic germ. The subtilis does not allow the oxygen to pass by, appropriating it to itself.

Buchner's Method. The test tube containing the culture is placed within a larger tube, the lower part of which contains an alkaline solution of pyrogallic acid. The tube is then closed with a rubber stopper. (Fig. 41.)

Botkin's Method. Petri dishes, uncovered, are placed on a rack under a large bell-jar, into which hydrogen gas is con

ducted. Alkaline pyrogallic acid is placed in the upper and lower dishes to absorb what oxygen remains.

CHAPTER XIII.

INFECTION.

The Properties of Blood Serum. So much has been written in recent years concerning blood serum that it is thought best to dilate somewhat upon it.

Fresh blood serum prevents the growth of many forms of bacteria. This property is gradually lost, and finally the serum serves as good culture media. These general bactericidal properties are lost if the blood is dialysed, if it is exposed to sunlight, or if it is heated for half an hour at 55° C. Its strength is variable in different specimens taken from the same animal and in different animals. Alexins is the term applied to the albuminoid bactericidal substances separated by Buchner and Hankin. Hankin believes these alexins to be formed by leucocytes. Special germicidal properties are given to the blood when an animal is rendered immune or is subjected to the action of toxins. Of the two forms of specific properties, anti· bacterial and anti-toxic, the former is present in all forms of immune serum, the latter only when toxins have been used to produce immunity.

Anti-toxic substances are more resistant than alexins or antibacterial substances. They can be dried and subjected to a heat of 100° C. without destruction. Anti-toxic substances form in 3 or 4 days and retain power for 3 or 4 weeks.

Specific anti-bodies of Pfeiffer are found in the blood serum of animals immunized against cholera and typhoid. These bodies are claimed to be quantitatively specific-i. e., while pure serum will produce the Pfeiffer phenomenon known as agglutination, if typhoid serum is used with typhoid cultures a very weak dilution of the serum will be effective, and the same can be said of cholera serum with the cholera cultures. This is utilized in the Widal test. (See Bacillus of Typhoid.)

As certain yeasts are specific for certain fermentations, Pfeiffer considers these anti-bodies specific.

How Bacteria Cause Disease. Many theories have been put forward to explain the action of bacteria in causing dis

ease, but only a few of the more important ones can be taken up here.

What are the Conditions Necessary to Produce Infection? First. As to the Infective Agent. The organism must have the power to produce disease. It must, in other words, be pathogenic. A non-pathogenic bacterium under certain conditions may cause disease, but this is not an infectious disease; it is rather a toxemia, and is due to the absorption of poisons generated outside of the body. It must be parasitic-have the power of growing within the body of an animal.

Essentially an infectious disease is a toxemia, because it depends upon poisons or toxins produced in the body. Parasitic or infectious bacteria cause disease by growing in the animal organism and generating products therein which are toxic. Saprophytic bacteria grow outside of the animal organism in dead matter, decaying particles, etc., and they may give rise to products which also are toxic to the animal economy.

Second. The toxins or poisons elaborated must be present in sufficient amount. Undoubtedly each animal organism is a law unto itself in regard to the amount of poison it will tolerate before disease is actually produced. The period of incubation can be explained on the supposition that the germ requires so much time to elaborate the amount of toxin necessary. This time period varies with different organisms, some carrying the toxin with them at the time of entry.

Third. The animal infected must be susceptible. Susceptibility varies in different species of animals, in different members of the same species, in the same individual at different times, and in the same individual to the different forms of disease germs. Susceptibility may be natural to the race, it may be acquired, it may be inherited. Mice are naturally susceptible to anthrax. Acquired susceptibility occurs upon exposure to conditions which lower vitality, as hunger, cold, advanced age, and surgical shock. Inherited susceptibility is a less important factor now than formerly. Many diseases were at one time considered inherited which now are known to be acquired during the lifetime of an individual. Still, certain physical characteristics, such as narrow chest, mouth-breathing, etc.-clearly inheritable