LABORATORY AND GREENHOUSE STUDIES. 129 



solution of ammonium oxalate. This work was done in the usual 

 way and left until April 5, at which time (after 47 days) the material 

 was collected and i)latings from the diseased tissue were made bf»th 

 by the writer and by Miss Lucia McCulloch. The appearance of the 

 inoculations was as follows: 



Bacillus coli. — The outermost point of the inoculation was merely a trifle browned 

 and water soaked and not at all extensive. The next inner leaf and the one inclosing 

 the central leaf had uppermost an inoculation hole which was browned and water 

 soaked, but only 8 millimeters in extent. On the other side of this same leafstalk was 

 a soft-rotted white area about 5 centimeters long. The innermost leaf, which was 

 still folded, showed the result of the inoculation extending over a distance of 9 centi- 

 meters. The diseased part at the lower end was only slightly browned and dry, the 

 middle was soft rotted and water soaked, and the upper part was considerably black- 

 ened. The rot was a typical soft rot, although it had not reduced the tissues to a 

 watery fluid. 



Ammonium oxalate. — In the outer tissues this inoculation had no characteristic 

 effect. In the inner tissues the leaf was somewhat blackened and dry. No soft rot 

 was in evidence. The action seems to have been a poisonous one rather than one 

 having any effect in dissolving the tissues. 



The isolation of the organism from the diseased material as carried 

 out by Miss McCulloch is described in the following paragraphs : 



Young coconut leaf, brown to black with rot at base. Bacteria only moderately 

 abundant as seen by the microscope. Some mycelium found. 



Plates poured with ordinary beef agar showed in 20 hours numerous round, white 

 colonies up to 2 millimeters in diameter. Transfers were made to agar and to litmus 

 milk. 



In 48 hours the agar colonies which had been white were cream color, opaque, and 

 not quite round. Transfers were made from the agar tubes to fermentation tubes 

 containing 1 per cent peptone water plus 1 per cent dextrose plus neutral red and to 

 tubes containing nitrate bouillon. 



In three days the fermentation tubes contained gas to the amount of 2.5 to 3 centi- 

 meters and the closed arms were canary yellow. Five out of the six tubes showed 

 this reaction. The nitrate-bouillon cultures were then tested for the reduction of 

 nitrates to nitrites and the same five out of the six tubes responded to the test. 



Transfers were made from the five fermentation tubes which produced gas to slant 

 tubes of Dolt's synthetic medium. In two days the medium became reddened and 

 the cultures showed a good pink, wet-shining growth. 



Transfers were made from these slant agar tubes to litmus milk and to agar con- 

 taining dextrose and neutral red. 



The litmus-milk cultures made directly from the plates, for the most part, reddened 

 and coagulated. The litmus-milk cultures, from the slant-agar Dolt's medium, 

 likewise reddened and coagulated. 



The agar tubes containing dextrose and neutral red after inoculation showed good 

 growth and a subsequent bleaching of the color, but no change to canary yellow. 

 As the reaction is an inconstant one, however, it can not be considered e\'idence 

 against the identification of Bacillus coli. 



Thus, in Miss McCuUoch's isolations of the organisms she obtained 

 bacteria which coagulated milk, reddened litmus, grew well on Dolt's 

 litmus-lactose-glycerin agar, produced gas, and caused a change to 



6389°— Bui. 228—12 9 



