132 A MANUAL OF BACTERIOLOGY 



the seed. Purely physical sterilization is impossible, because the 

 bacteria are even more resistant than the seed. Chemical agents 

 have been found to give the best results. The ordinary antiseptics, 

 such as alcohol, ether, chloroform, and toluol, have proved unsuitable 

 for this purpose. The substances commonly employed for sterilizing 

 seed are corrosive sublimate, silver nitrate, bichromate of potash, 

 copper sulphate, bromine, and many of the mineral acids. The dis- 

 infecting power of these substances depends largely upon four factors 

 — strength of solution, time of action, temperature, and pressure. 



The ordinary procedure of soaking the seeds in mercuric chloride 

 solution does not always give satisfactory results, because of the per- 

 sistance with which occasional air bubbles remain on or inside the 

 seeds, and thus prevent complete sterilization. 



Very good results have been obtained in sterilizing the seeds of 

 corn, wheat, and peas by a modification of the method first described 

 by Hutchinson and Miller,'' which consists in sterilizing the seeds in 

 a mercuric chloride solution in vacuum. In the first place, well-dried 

 seeds of even size are necessary. These should be thoroughly washed 

 and cleaned in flowing water and perhaps 60 per cent alcohol before 

 beginning sterilization. Transfer the seeds to a sterile bottle and, 

 after adding mercuric chloride, remove the air bubbles with a vacuum 

 pump. The apparatus for this purpose consists of the following 

 (Fig. 43) : 



Five stout-walled glass flasks, wide at the mouth (capacity about 

 500-800 cc), carrying steam-proof rubber stoppers with two holes in 

 each for glass tubes, which are connected by rubber tubing, as seen 

 in the diagram. Flask B is connected to flask A on one side, and on 

 the other, by means of a three-way tube, to two glass bottles, C and 

 D. ' These, in turn, are connected to flask E. Fill flask C with a 

 0.25 per cent solution of mercuric chloride, D with distilled water, 

 and E, which is smaller than the other flasks, about half full of 

 50 per cent sulphuric acid. This bottle has a drawnout tube passing 

 down through the cork almost to the bottom and serves to purify 

 the air. 



After connecting the whole apparatus, place screw clamps on- the 

 rubber tubing between all flasks, in order to prevent the liquid 

 from flowing from one bottle into another; then sterilize in the 

 autoclave at 125° C. for half an hour, or three times by fractional 

 steam. Cool to 40° C. and connect flask ^ to a vacuum pump ; place 



1 Journal of Agricultural Science 3 : 2, 185. 1908. 



