BY R. GEEIG-SMITH. 55 



extract to seventeen oi water, thus briugmg the original approximately 5 % of 

 tannin down to about | %. The solution was divided into three sets, each set 

 containing a control and two other portions, one with 0.2 % and one with 0.4 % 

 of calcium lactate. The sets were seeded with phases Al, A2 and B2. Phase Al 

 did not become ropy. Phase B2 developed ropmess in the control only, while 

 phase A2 showed ropiness in all tests . In these, the ropiness did not appear until 

 the third day at 28° and it had disappeared by the sixth, giving place to cob- 

 webby growths consisting of bacteria emmeshed in coagulated slime. The ex- 

 periment showed that ropiness may develop in dilutions of old extract of wattle- 

 bark and that it soon disappears. It also seemed to sliow that calcium lactate, a 

 substance probably occurring in old tan liquors, has little or no influence in as- 

 sisting the ropy fermentation. 



During the investigation the bacteria were tested to see if they retained the 

 power of making infusions of wattle bark ropy. Their- physiological activities 

 were being tested in sj'nthetic solutions, and in these the characteristic ropiness 

 was being produced, but it -vras considered advisable to prove that this also hap- 

 ]iened in bark infusions. Thirty gram portions of raw bark were put into sterile 

 4-oiince bottles and 50 e.c. portions of sterile water were added. The water 

 just covered the bark. The liquids were seeded with the bacteria and incubated 

 at 28°. Upon the first occasion of this routine testing, phase Al produced the 

 characteristic ropiness in a day and phases B2 and A2 in two days. 



Twelve days later the test was repeated. Phases B2 and A2a developed the 

 ropiness in two days, a duplicate race of B2 in three days. By the fourth day, 

 phase A2 had developed ropiness. Phase Al, which had given a positive result 

 twelve days before, was negative. 



Other tests made from time to time showed, like the above, a certain varia- 

 bility in the activity of the phases. Tliis was to be expected, for a stock culture 

 could not be kept on account of the alteration . of one phase into another. The 

 bacteria were carried over from colony to colony, that is, plates were smeared 

 every few days and from these, colonies were picked out and seeded into bouillon. 

 It was only by proceeding in this way that the phases Al and A2 could be main- 

 tained in a pure state. 



When infected bark is covered with water and allowed to stand, the bacteria 

 gr-ow and produce the ropy substance, while the solution increases in strength. 

 The bacteria apparently grow in clumps of slime, that is to. say, they form a 

 coherent sHrne and remain imbedded in this slimey environment. This is demon- 

 strated when the bacteria are gTown in saccharine nutrient solutions containing 

 chalk ; the blobs of cohesive slime can be seen upon rotating the flask, and they are 

 meapable of being broken up by the rotation of the flask. Once the blob of 

 slime around the bacteria is admitted, it becomes a matter of question as to the 

 diffusive speed of the tannins and non-tannins through the slimes, just as it is 

 a question about the diffusive speed of the non-tannins and tannins from the bark. 



If in making an extract, the non-tannins, which we will presume are chiefly 

 bacterial nutrients are the first to diffuse, or preponderate in the initial diffusion, 

 the bacteria will grow and, in doing so, form a protective slime envelope which 

 may be protective until the tannins become sufficiently concentrated to coagulate 

 it. Such a coagulation occurs experimentally in dilute infusions of bark, but it 

 has not been observed to occur in cases where the bark has been covered with 

 water, and the infusion allowed to remain in contact with the bark. 



In an endeavour to throw some light upon this matter, portions of raw bark 

 were treated with water in the ratio of three of bark to five of water, and after 



