104 BACTERIAL DISEASES OF PLANTS 



in advance of nitration it is desirable that such heating should 

 not be carried to the boiling point. We extract potato juice 

 from thin slices of the tubers by heating for an hour in double 

 their weight of water at a temperature not higher than 60C., 

 in order that we may be able to filter the fluid without difficulty. 

 Fluids extracted from vegetables by grinding and pressure are 

 usually not very well adapted in their concentrated form to 

 the growth of bacteria and must, therefore, be diluted with a 

 considerable volume of water before they are steam sterilized 

 (1 :4, 1:10 may be taken as standard dilutions, but I also try 

 them full strength). Such juices may be sterilized cold, if de- 

 sired, by passing them through a Chamberland filter free from 

 cracks. 



Animal Substances. Cow's milk is one of the easiest and 

 most satisfactory of animal substances for use, provided it can 

 be obtained in a fresh condition and is properly sterilized. I 

 use it both with and without cream, especially without. For 

 separating the cream we use a centrifuge. Small quantities 

 may, however, be prepared nearly cream-free by filtration 

 through coarse filter paper but this is a slow process. Over- 

 heating changes the nature of milk somewhat and should be 

 avoided. We generally use it in 10-cc. portions in sterile test 

 tubes, cotton-plugged, which portions are steamed on 4 consecu- 

 tive days, each time for a short period only. Often three times 

 heating is enough, but occasionally the fourth heating is re- 

 quired to render the milk free from resistant bacteria, and there- 

 fore we have adopted the four heatings, as a routine practice. 

 These steamings may be for 15 minutes on the first day, 10 min- 

 utes on the second and third days, and 5 minutes on the fourth 

 day. This destroys all aerobes, but some anaerobes may re- 

 main. Milk should not only be used in the manner described 

 but should also be used in the form of litmus milk, only enough 

 sensitive blue litmus being added to render the fluid a lilac or 

 lavender blue. If the curd is precipitated in the steaming, then 

 something is wrong either with the litmus or with the milk. 

 Usually it means that such milk is old and full of organisms and 

 their by-products. Methylene blue may also be added to milk 

 (6 cc. of 1 per cent, methylene blue in distilled water to each 

 200 cc. of milk) for study of the reducing activities of organ- 



