22 
BULLETIN 780, U. S. DEPARTMENT OF AGRICULTURE. 
constrictions are less marked, and the transparency is diminished. 
In late stages of the disease, however, the stomach approaches the 
normal in size and the constrictions are again well marked. The 
organ is then white and opaque and the tissues are friable and easily 
crushed. When crushed the mass presents a milky appearance. 
Upon microscopic examination Nosema apis is found in very large 
numbers in the crushed tissues. The presence of the parasite is 
almost invariably recognized by its spore form. The presence of 
Nosema-infected bees in a colony is the one constant colony symptom 
of the disease. 
METHODS EMPLOYED IN EXPERIMENTAL STUDIES. 
As Nosema apis has not been grown in the laboratory by artificial 
methods, in carrying out these investigations it has been necessary 
to inoculate a large number of 
colonies of bees. The use of a 
few bees in cages was found to 
be inadequate for experimental 
purposes. A 4 to 6 frame nu- 
cleus in a 10-frame hive body 
(fig. 5) serves well the purposes 
of an experimental colony. 
The experimental apiary (PI. 
IV) , consisting usually of about 
50 colonies, was the same one 
that was used in the sacbrood 
studies. During the bee sea- 
son the colonies were inocu- 
lated and kept in the apiary 
in the open under conditions 
similar to those occurring in 
nature. Precautions similar 
to those observed in the sac- 
brood studies were followed in 
the present studies. During the winter colonies to be inoculated 
were removed to and kept in the laboratory. The top of the hive 
body was screened and the bees given free opportunity for flight 
through a hole in the window. 
The manner of obtaining the parasite Nosema apis from diseased 
bees for use in the inoculations is described under " Diagnosis' ' 
(p. 48). The stomachs of from 5 to 10 infected bees are amply suffi- 
cient for each inoculation. After their removal from the bees they 
are crushed, suspended in sirup, and fed to a colony free or practically 
free from Nosema infection. The methods throughout are similar 
Fig. 5.— Experimental hive, having four Hoffman 
frames, a division board, Petri dishes as feeders, the 
entrance nearly closed with wire cloth, and the open- 
ing on the side of the hive body occupied by the 
frames. (Original.) 
