Fig. 72.—Cholera vibrios, short forms. (MacNeal from Kolle and Schurmann after Zettrow.)
It is Gram-negative and stains best with a dilute (1-10) carbol fuchsin. There is a single terminal flagellum, which endows the organism with great motility, which may best be termed scintillating. It has been estimated that its motility is five times greater than that of the typhoid bacillus. It grows best on media with an alkaline reaction (—0.4%) and it is this tolerance for media of high alkalinity that permits the separation of the cholera spirillum from the ordinary faecal bacteria by the use of Dieudonne’s alkaline blood agar or similar media. This is equal parts of defibrinated ox blood and N/1 NaOH, 3 parts of which are added to 7 parts of nutrient agar. It thus has 15% of normal sodium hydrate, instead of the 1% acid reaction of the usual media. Unfortunately, other spirilla tolerate this high alkalinity.
The cholera organism is strongly aerobic and grows quickly and luxuriantly in the upper part of a tube of Dunham’s peptone solution, this property enabling one to separate it from other organisms of faeces by taking up loopfuls from the surface layer to plate out on agar of about 0 or -0.3% reaction. When grown in peptone solution the cholera spirillum produces a nitroso body so that one obtains an indol reaction (cholera red) by simply adding 5 or 6 drops of concentrated H2SO4.
When this test is employed it is necessary to determine whether the peptone used is suitable for the reaction. As a matter of fact this test is now rather discredited. Blood serum is digested. Recently much discussion has arisen as to the value of the haemolytic power possessed by noncholera vibrios on blood agar plates.
It is true that the digestive action which true cholera has on the red cells of the medium may give the appearance of a zone of haemolysis. Therefore, for the demonstration of this haemolytic action of noncholera spirilla, fluid blood media should be used. The El Tor spirillum, isolated from Egyptian pilgrims without symptoms of cholera, is haemolytic, but gives the immunity reactions of the true cholera vibrios which are not haemolytic.
Gelatine is liquefied and the stab shows an air bubble liquefaction at the summit of the stab. On gelatine plates a powdered glass center with an encircling zone of liquefaction was formerly considered characteristic of cholera, but at the present time gelatine cultures have been almost abandoned in practical work.
As a rule animals cannot be infected by feeding them cholera material unless the acidity of the gastric juice be neutralized and intestinal peristalsis checked by opium (procedure of Koch). Injected intraperitoneally, the cholera vibrio produces a fatal peritonitis. Recently monkeys have been infected after purgation with sulphate of soda and administration of bicarbonate of soda. They died in from one to forty-eight hours with symptoms of cholera.
There have been instances where cholera has been caused in laboratory workers by the accidental ingestion of cholera cultures, thus Orgel was infected from sucking up peritoneal fluid in doing Pfeiffer tests for bacteriolysis and died.
Emmerich and Pettenkofer swallowed cholera cultures, the former experiencing a severe attack of cholera and the latter a diarrhoea in which cholera spirilla were present. On the other hand similar experiments have resulted negatively but this is what should be expected from the epidemiological facts as to carriers.
The virulence of the cholera vibrio can be exalted by passage through guinea pigs—successive culturing of the peritoneal exudate of intraperitoneally infected animals alternating with culture media growth inoculations. Such a fixed virus, the virulence of which cannot be exalted, is the material used by Haffkine in his cholera vaccine. The toxicity of cholera is supposed to be due to an endotoxin which is set free when the vibrios undergo disintegration when lying between the basement membrane and epithelial lining of Lieberkühn’s glands. Others think the vibrios may enter the blood stream, there to be immediately disintegrated with toxin production. The usual idea, however, is that the cholera spirilla never invade the blood stream—they are confined to the alimentary canal. Macfadyen obtained the endotoxin by grinding the frozen spirilla. This toxin was destroyed by a temperature of 60°C.