THE CULTIVATON OF SPIROCHAETA NOVYI* WITHOUT THE USE 
OF TISSUE FROM ANIMAL ORGANS. 
CHARLES A, BEHRENS. 
(Purdue University, Lafayette, Indiana. ) 
In 1909 Schereschewsky' by deeply imbedding a piece of human tissue 
containing Spirochaeta Pallida in gelatinized horse serum, first demon- 
strated that the treponema might be cultivated in vitro. He was not able 
to obtain a pure culture of the organism, for bacteria grew along with the 
pallidum, nor did he succeed in reproducing syphilitic lesions by inoculating 
animals. 
In 1910, Muhlen*® obtained the first generation of the spirochaete by 
utilizing the above mentioned method. By the use of horse serum agar, 
he further succeeded in obtaining a culture devoid of bacteria. Muhlen’s 
pure cultures also were evidently non-pathogenic. 
During the same year, Bruchner and Galasesco® succeeded in cultivating 
“young impure cultures” by using Schereschewsky’s medium. But upon 
inoculating rabbits with the material which also contained the original 
spirochaetal tissue, syphilitic lesions were produced. They were, however, 
unable to obtain a second generation of these organisms. 
In 1911, Hoffman* succeeded in getting pure cultures of the spirochaete 
by the utilization of Schereschewsky’s and Miihlen’s methods. Although 
his cultures were morphologically typical, he, like the above mentioned 
experimenters, was not able to demonstrate their pathogenic properties. 
Also at this time, Sowade® succeeded in procuring impure virulent cul- 
tures by using the gelatinized horse serum or gelatinized ascitie fluid. This 
inocuable material also still contained the original pallidum tissue. 
In 1911, Noguchi® was able to cultivate pure virulent cultures in vitro 
of this organism. He accomplished this by placing a small piece of fresh 
sterile rabbit kidney or testicle tissue into each tube containing about 
sixteen cubic centimeters of serum water (one part of serum and three 
parts of distilled water) which was previously fractionally sterilized. A 
layer of sterile paraffin oil was added to each sterile tube containing this 
cultural medium and placed under strict anaerobiosis and incubated at 
35-37° Centigrade. Under these conditions the spirochaetes which were 
morphologically typical and virulent were obtained and cultivated for 
many generations. 
In the following year, Noguchi’ cultivated for the first time the follow- 
ing relapsing-fever spirochaetes: Spirochaeta Duttoni, Spirochaeta Kochi, 
Spirochaeta Obermeieri and Spirochaeta Novyi. 
In 1912, Treponema macrodentium and microdentium’, Treponema Re- 
fringens’, Treponema mucosum”™, Spirochaeta Phagedenis™, and Spirochaeta 
Gallinarum”™ were likewise successfully cultivated. 
The media used in their cultivation was of a similar nature to that used 
in the pallidum cultural work. It was not essential in some instances to 
*The author is greatly indebted to Dr. F. G. Novy, Ann Arbor, Michigan, thru 
whose kindness this strain of Spirochaete was obtained. 
