276 



SCIENCE 



[Vol. IV., No. 85. 



a cloven hoof indicates to the observer the 

 forms of the teeth, of all the big bones, thighs, 

 shoulders, and of the trunk of the body, of the 

 animal which left the mark." x 



W. P. Blake. 



Mill Rock, New Haven. 



THE VIRULENCE OF CULTIVATED 

 ANTHRAX VIRUS. 



Experimental studies on the artificial attenuation of the 

 infectious properties of the bacillus of A nthrax by 

 means of cultivation. By Dr. R. Koch, Dr. 

 Gaffky, and Dr. Loeffler. 



Mittheilungen aus dem Kaiserlichen gesundheits-amte. 

 Zweiter band. Berlin, 1884. (Extract from the 

 publication of the imperial board of health of 

 Germany.) 



Pasteur's announcement that the parasites 

 of malignant Anthrax were capable of changing 

 their characteristics when cultivated under cer- 

 tain conditions, and that when thus modified 

 they could be used for protective inoculation, 

 aroused the greatest interest among investiga- 

 tors. Such a statement could not be accepted 

 without confirmation at the hands of other 

 observers ; and none were better fitted for 

 this task than the Roj^al health commission of 

 Germany, at the head of which stands Dr. 

 Robert Koch. 



The experiments, which were instituted un- 

 der his direction, have been carried on for two 

 years, and have shown, that, although the ba- 

 cilli could be rendered harmless, their protec- 

 tive power was not so great as was expected. 



The original communication of the French 

 savant was not exact in the details by which 

 the experiments were to be carried out, and 

 Koch had to employ much time in preliminary 

 studies. This cannot, however, be considered 

 as lost, since many valuable facts have been 

 obtained by it. 



According to Pasteur, if the cultivations were 

 kept at a temperature constantly maintained 

 between 42° and 43° C, the virulence gradu- 

 ally decreased until the ninth day, when it was 

 entirely lost. By removing a specimen on any 

 day, and allowing it to germinate at a tempera- 

 ture of 37° C, its activity at that stage could be 

 perpetuated, and thus any degree of virulence 

 that was required could be preserved. Two 

 such cultures of different strength were used 

 for protective inoculation, the weaker of which 

 was called the premier, and the stronger the 

 deuxi^me vaccin. 



Koch commenced his investigations in the 

 following way. A mouse was injected with 



1 Proc. am. assoc. ad. sc, xiv. 146. 



blood containing spores of the bacillus, which 

 had been kept five years, and was known to 

 be of great virulence. 



The animal was killed at the end of twenty- 

 four hours, and a minute quantity of the spleen 

 was taken on the point of a platinum needle 

 which had been sterilized by heat, and sown in 

 a glass bulb containing twent}^ cubic centi- 

 metres of chicken-bouillon neutralized with 

 sodic carbonate. The bulb was then sealed, 

 and placed at once in a constant-temperature 

 apparatus, which was kept between 42° and 

 43° C. 



Samples were taken daily, and tested upon 

 animals ; but, contrary to the promised result, 

 the growth was found to be as deadly for small 

 animals on the ninth as on the first da} 7 . Fur- 

 ther cultivation proved, however, that, in a 

 period varying from eighteen to twenty-nine 

 da} T s, the infectious property was entirely lost, 

 although the external appearance of the bacil- 

 lus was unaltered. Thus far, Pasteur's asser- 

 tion was substantiated, except in regard to the 

 length of time. A portion of this was taken, 

 and allowed to grow at an ordinary tempera- 

 ture for two years ; and during this time there 

 has been no evidence of a return of virulence, 

 nor has the form changed. These bacilli are 

 as immovable as the active ones ; their ends 

 appear sharply truncated ; they form long fila- 

 ments, in which are developed oval glancing 

 spores. Vaccination with this entirety inac- 

 tive form did not give immunity against inocu- 

 lation with the virulent one. 



Those of a slight degree of force were next 

 tried. At the end of twent}^-four days a cul- 

 ture was obtained which would kill mice, but 

 not guinea-pigs or other small animals, but 

 still did not render them safe. This particular 

 form Koch speaks of as ' mouse anthrax.' 



It was thought that perhaps this represented 

 the second and the inactive form, the first vac- 

 cination of Pasteur. Accordingly, a sheep 

 was tried, but it succumbed to the malignant 

 form as quickly as ever. It was next proposed 

 to use three or more preventive inoculations ; 

 and, accordingly, cultures from the fifteenth day 

 were taken as the first, from the eleventh as the 

 second, from the ninth as the third, and from 

 the fifth as the fourth, and these were followed 

 by the virulent form. In this manner seven 

 sheep, seven rabbits, and eleven guinea-pigs 

 were tried. At the end, all the rabbits and 

 guinea-pigs, and five of the sheep, had died. 



In order to determine whether there might 

 not be some other difference, specimens of the 

 vaccinating-material, as furnished by Pasteur 

 through his agent, were purchased, and proved 



