SPORES 243 



fresh blood preparations. If the blood or the juice from the spleen 

 is obtained a considerable time after death, when putrefactive changes 

 and the admission of air has set in, longer chains and spores may be 

 found. 



Staining Properties. The anthrax bacillus stains well with the 

 ordinary watery anilin stains, and is Gram positive. When fixed and 

 stained in the usual manner the bacilli show certain features not seen 

 in the fresh, unstained specimen. The ends of the bacilli appear 

 sharper and are not so regularly cylindrical as before, but slightly 

 swollen, and show a little excavation of the outer surface, so that a 

 small lenticular empty space is formed between two individual bacilli 

 of a chain. A chain of anthrax bacilli after it is stained looks very 

 much like a stick of bamboo. In the stained blood or splenic juice 

 preparation the anthrax bacillus shows a gelatinous capsule, which, 

 however, is not present in bacilli obtained from pure cultures on the 

 ordinary artificial media. Johne has devised a special method for 

 clearly demonstrating the anthrax bacillus capsule in blood prepa- 

 rations. His method is as follows: 



1. Prepare a blood smear on a cover-glass, allow it to become air 

 dry, and fix by drawing carefully and rapidly through a flame. Do 

 not overheat or the capsules will be burned. 



2. Apply to the fixed cover-glass 2 per cent, watery gentian-violet 

 solution and heat slightly for one-quarter to one-half minute over a 

 flame. 



3. Wash rapidly in water. 



4. Apply for six to ten seconds a 1 to 2 per cent, solution of acetic 

 acid. 



5. Wash in water and mount in water (not in Canada balsam) on 

 a slide and examine while moist. 



The capsules may also be stained by any of the other methods for 

 exhibiting this plasmatic envelope. 



Spores. Spores can easily be demonstrated in cultures raised on 

 artificial culture media in the presence of oxygen. As far as is known, 

 the organism never forms spores in the absence of free oxygen, and 

 they are never seen in the living blood. They are formed in artificial 

 media at blood temperature after eighteen hours, at 18 C., after 

 fifty hours; at 14 C. spores are no longer formed. The spore of the 

 anthrax bacillus is found in the middle of the rod, but since the proto- 

 plasm of the latter soon degenerates and perishes after sporulation, 

 it may appear as if this were not the case. Every bacillus forms a 

 single spore, which is not perfectly spherical but somewhat egg-shaped 

 or elliptical. These spores possess a very tough membrane; some 

 observers even claim that the anthrax spore has two membranes. 

 The following method of staining the spores is recommended as the 

 most trustworthy: 



1. Take several platinum loopfuls of material from an anthrax 

 growth on a solid medium and rub it up well with a 0.85 per cent, 

 salt solution. 



