ROOT-NODULES OF LEGUMINOSAE. 119 



They are then rinsed in water and placed in the iodine solution. After washing this off they were 

 allowed to remain one or two minutes in the orange G. They were then washed in absolute alcohol 

 as long as gentian violet came off abundantly or needed to be removed. They were cleared in clove 

 oil, this being preferred to xylol. He states that the bacteria were distinctly differentially stained in 

 the tissues. 



In the Bur clover the proportion of root-hairs attacked naturally by the nodule organism was 

 estimated to be about one in a thousand. He, however, was able to confirm Miss Dawson's statement 

 respecting the abundance of infected root-hairs when the roots of legumes were subjected to special 

 methods of infection, e. g., placed on damp filter paper under a tumbler, and watered with water 

 containing the bacteria in suspension, nearly every hair on a root in some fields of the microscope was 

 found to be enlarged and twisted at the end, and showed the beginning of an infection thread. A 

 striking characteristic of the infection of the root-hairs is the curvature of the hair about the infected 

 portion (fig. 21). It is believed that the organism dissolves or softens the walls of the root-hair and 

 thus enters. A wound is not necessary. In fact, wounded root-hairs lose the power of curvature 

 displayed so strikingly in the ones showing the infections. The curvature is unquestionably due to 

 the presence of the bacteria. 



"The bending is the evident response to irritation." * * * " Since the majority of infected 

 root-hairs show the bending at or near the tip, * we may infer that the bacteria enter unin- 



jured hairs which are able by growth-curvatures to respond to mechanical or chemical stimuli." 



* * * "Having entered the root-hair by softening or dissolving a small portion of the cell-wall, 

 and moving or growing through this, the tubercle bacteria multiply rapidly, forming a thread-like 

 zoogloea from the infection spot along the hair into the epidermal cell of which the hair is a branch. 

 From the epidermis the infecting zoogloeae grows fairly straight into the underlying cortical paren- 

 chyma." * * * "The direction of the infection thread -which is solid, and is incorrectly termed 

 infection ' tube ' is too regular not to encourage one to suppose that the course of the growing strand 

 of bacteria is determined by attraction exerted by the host-cells upon the bacteria. This then is 

 chemotropic growth of the strand or, if the bacteria are motile in the cells, chemotactic movement of 

 the bacteria. The course of the thread is toward the conducting tissues of the host." * * * "The 

 growth does not extend into the central cylinder and the conducting tissues, so far as I have seen. 

 Instead, in the layer of cells just outside the endodermis of the root, division takes place in the cell 

 into which the infection thread has penetrated and in the cells adjacent to it. The daughter-cells 

 grow, repeated divisions and growth follow, and there arises a conical mass of cells which are some- 

 what larger, and which contain more protoplasm than the adjacent cortical parenchyma cells. This 

 conical mass is the young tubercle. At first all of its cells are merismatic, but later the divisions 

 become more and more limited to the cells near the rounded apex of the blunt cone. Thus a regular 

 cambium is differentiated in the tubercle. This cambium * * * lies near the tip of the tubercle, 

 and forms a bowl-shaped or shallow thimble-shaped layer." 



"The growing tubercle pushes out the overlying cortical parenchyma and epidermis, forming an 

 increasing swelling on the side of the root. Cortical parenchyma and epidermis, at least for a time, 

 nearly keep pace with the growth of the tubercle. Thus, although the cortical cells are compressed 

 somewhat, the epidermis is not ruptured, and the tubercle does not burst out of the side of the root as 

 a lateral root does." 



Morphologically, in origin the nodules are indistinguishable from the nascent lateral roots. In 

 subsequent growth they are more and more dissimilar. 



"Morphologically, then, the root-tubercles are lateral roots." * * "The bacteria in the 

 infection thread, which grows through the root-hair and the cortical parenchyma cells of the root 

 to the pericambium layer, multiply, but they multiply most rapidly in the infected cells farthest from 

 the surface of the root. New threads form, which grow out into and infect the cells of the mass of 

 new cells composing the embryo-tubercle. Thus a majority of the cells in the young tubercle contain 

 bacteria." 



"Though infected cells do divide, they probably divide less often than the uninfected cells." 



* * * "The infection of the daughter-cells composing the embryo-tubercle is accomplished by 

 branching infection threads growing in fairly straight lines radiating from the base of the tubercle. 

 In this way the cells near the base of the growing tubercle are most infected, those near the tip least." 



The meristem continues to form new cells between itself and the cells containing bacteria and 

 infection threads. When, however, he imprisoned the nodules in plaster of Paris, the meristematic 

 cells also were infected. 



The infected cells became enlarged and in their definitive condition are said to be from half as 

 large again to twice as large as the uninfected cells. There was less difference in size of normal and 

 infected cells in nodules imprisoned in plaster casts. 



The infected cells are thin-walled and contain only one large vacuole. This is not traversed by 



