GALL ORGANISM SCARCE IN TISSUES. 193 



bacteria — often none, or less than 1 per loop (see p. 168)." If he 

 had mashed his cube in the first tube of bouillon, allowed the con- 

 tents of the crushed cells to diffuse for an hour, and then inoculated 

 directly from this first tube, rather copiously, e. g., with several 

 3 mm. loops of the fluid, he would probably have had colonies of the 



a As the result of poured plates made in 1910 by Lucia McCulloch, using a sound old hop gall received 

 from the Pacific coast, it would seem that there were less than 500 living bacteria per cubic inch of the mate- 

 rial used. The right organism was plated out and tumors obtained with it on sugar beet and daisy, but 

 two of the three colonies selected were noninfectious. 



Plates of +15 nutrient agar, poured by Miss Brown in the fall of 1910 from tumors on sugar beet, gave the 

 following results: 



(1) First set of Arlington (Va.) plates. Two c. c. of a rather old and tough tumor were mashed in 10 

 c. c. of bouillon. Eleven plates were poured, all from the original tube, inoculating as follows: 



3 plates each Ave 3 mm. loops. 

 3 plates each four 3 mm. loops. 

 2 plates each three 3 mm. loops. 

 2 plates each two 3 mm. loops. 



1 plate one 3 mm. loop. 



Five favorable colonies appeared on this set of plates. 



(2) Second set of Arlington (Va.) plates made from another tumor— material good. Three c. c. were 

 mashed in 10 c. c. of bouillon. Eight plates were poured. The first six were from the original tube, the 

 other two from the first dilution. The inoculation was heavy, viz: 



2 plates with five 3 mm. loops. 

 2 plates with four 3 mm. loops. 



2 plates with three 3 mm. loops. 

 1 plate with three 3 mm. loops. 



1 plate with two 3 mm. loops. 



Fifteen favorable colonies appeared on this set of plates. 



(3) First set of Blissfield (Mich.) plates. 



Of this tumor 3.4 c. c. were mashed in 10 c. c. of bouillon. Eight tubes were poured, the first six from 

 the original tube, the other two from the first dilution, inoculating as follows: 



3 plates each with three 3 mm. loops. 



2 plates each with two 3 mm. loops. 

 1 plate with one 3 mm. loops. 



1 plate with two 3 mm. loops. 

 1 plate with one 3 mm. loop. 

 Five colonies resembling gall colonies came up on this set of plates. 



(4) Second set [of Blissfield plates (same tumor, next day), using 0.5 c c, which was mashed in 10 c. c. 

 bouillon. Eight plates were poured, all from the original tube, inoculating as follows: 



4 plates each with four 3 mm. loops. 

 1 plate with three 3 mm. loops. 



1 plate with two 3 mm. loops. 



2 plates each with one 3 mm. loop. 

 No gall colonies appeared on this set of plates. 



(5) First set of Fairfield (Wash.) plates. A smooth tumor 3.5 to 4 cm. in diameter was selected and about 

 one-half of it (possibly 10 c. c.) was mashed in 10 c. c. of bouillon for the plates. All of the eight plates were 

 poured from the original tube, inoculating as follows: 



5 plates each with five 3 mm. loops. 

 2 plates each with four 3 mm. loops. 



1 plate with two 3 mm. loops. 

 No gall colonies appeared. 



(6) Second set of Fairfield plates (same tumor). About one cubic centimeter was mashed in 10 c. c. of 

 bouillon. Eight plates were poured, all being inoculated copiously from the original tube, viz: 



4 plates each with five 3 mm. loops. 



2 plates each with three 3 mm. loops. 

 2 plates each with two 3 mm. loops. 



Four colonies looking very much like the gall-forming organism grew on these plates. 



(7) Plates were also poured in December from a gall on another Arlington boot which had been trans- 

 planted to the hothouse for six weeks. These yielded only one colony resembling Bacterium tumefaciens, 

 and this gave no positive result when inoculated into sugar beets. 



Of thes§ 30 colonies, as already stated, only 5 have proved infectious and all of them are possessed only of 

 feeble virulence. 

 (For a quantitative study made by the senior writer In November, 1910, see under "Sugar beet." p. 81.) 



78026°— Bull. 213—11 13 



