406 CLEGG. 



method, a change in the morphology and a multiplication of the leprosy 

 bacilli, while smears made from the control tubes stained in a similar 

 manner showed no evidence of a multiplication of the bacilli inoculated 

 and no noticeable changes in morphology. Transplants of the mixed 

 cultures of amoeba, cholera and leprosy bacilli were made to fresh agar 

 tubes and incubated at the same temperature. Smears from these tubes 

 made three days later and stained as before showed a great increase in 

 niunber of the acid-fast bacilli and also a change in their morphology 

 from the long, slender bacilli found in smears from leprous lesions to 

 short plump rods and occasionally coccus forms. This deviation in mor- 

 phology is not constant, however, on continued cultivation ; some cultures 

 have resumed a more typical morphology, as is shown in the illustrations. 

 (Plate I, fig. 1.) Transplants of the mixed cultures were then made 

 once a week for three months. After having been on the artificial 

 medium for tbis length of time, the cultures were placed at a temperature 

 of 60° C. for thirty minutes. This temperature was sufficient to kill 

 the amoebBe and the cholera vibrios, but not the acid-fast l^acilli. Trans- 

 plants were made from the heated cultures to fresh tubes containing 

 amoebse and cholera vibrios and also to tubes containing plain, sterile 

 agar. After three days' incubation, smears from the tubes containing 

 the amoebse and cholera vibrios showed a development of the acid-fast 

 bacilli, and at the end of six days small, brownish colonies with regular 

 margins appeared on the surface of the tubes containing plain agar only. 

 Smear preparations made from these colonies and stained by the Ziehl- 

 Neelson method showed microscopically an acid-fast bacillus, the cultural 

 characteristics of which will be described in another part of this paper. 

 Case G. — This was a well-marked case of tubercular leprosy. The autopsy was 

 performed two hours after death. A small nodule was present in the apex of 

 the left lung which might have been of tubercular origin; there were no evidences 

 of gross tubercular lesions in the other organs. The spleen was removed in the 

 same manner as described in Case F. Smear preparations from the spleen stained 

 by the Ziehl-Neelson method showed very few leprosy bacilli. Cultures were 

 made on media containing amoebae and cholera vibrios and on plain sterile agar. 

 Because of the small niunber of leprosy bacilli present in the spleen, a large nodule 

 situated in the thigh and containing large numbers of the organism was also 

 selected for use in making the inoculations and was excised. In making the 

 cultures the surface of the nodule was first burned with a hot spatula and an 

 opening made with a sterilized scalpel. 



Ten tubes containing amoebae and cholera vibrios were inoculated with 

 material taken from the center of the nodule, as were also ten tubes 

 containing plain agar. All these cultures, together with several tubes 

 containing amoebae and cholera vibrios, but not inoculated with leprosy 

 materia], were placed in the incubator at a temperature of 37° C. At 

 the end of six days smear preparations from two of the tubes containing 

 amoebse, cliolera vibrios and material from tbe nodule showed a change 

 in morphology and an evident multiplication of the leprosy bacilli. 



