The Sterilization of American Foulbrood Combs 23 



remain alive to start the disease, as experience in cultivating Bacil- 

 lus larvae from diseased material on artificial culture media has 

 demonstrated that a certain quantity of inoculum is necessary to 

 start growth, whereas growth will not begin when there are only a 

 few spores. These are among the problems awaiting solution. 



Another important fact brought out by the various tests of solu- 

 tions containing 20 per cent of formalin is that in the case of all 

 combs that were treated for 48 hours no cultures made from scales 

 taken from open cells or from cells the cappings of which had been 

 removed gave any growth of Bacillus larvae. For this length of 

 treatment complete sterilization was accomplished by the water- 

 formalin solution as well as by the alcohol-formalin solution. In 

 the case of the 24-hour tests a few of the scales from open cells of 

 combs treated with alcohol-formalin solution failed to be completely 

 sterilized, 6 of the 220 such scales cultured giving growths of 

 B. larvae. Apparently a 24-hour treatment is somewhat below the 

 minimum time in which complete sterilization of the scales in open 

 cells may be expected. On the other hand, none of the cultures from 

 220 scales from open cells of comb treated 24 hours in water-forma- 

 lin solution showed any growth of B. larvae. These results indicate 

 that the water-formalin solution is more efficient as a germicide for 

 American foulbrood than is the alcohol-formalin solution, provided 

 the solutions come in actual contact with the scales for at least 24 

 hours. 



All cappings, those over brood cells as well as honey cappings, 

 shoutd be carefully removed to insure sterilization of combs infected 

 with American foulbrood. Even when this is done, apparently a 

 48-hour treatment is still necessary when an alcohol-formalin solu- 

 tion is used. From the fact that all cultures were negative which 

 were made from scales in open cells of combs treated 24 hours in 

 a water solution containing 20 per cent of formalin, a treatment of 

 24 hours in such a solution appears to be sufficient. When, after 

 combs have been soaked in the disinfectant solution, the excess liquid 

 has been removed in an extractor, and the combs are allowed to dry 

 without further treatment, there is still some disinfectant left which 

 continues to act while they are drying, until it is entirely evaporated. 

 If circumstances, such as the necessity for drying treated combs at 

 a comparatively low temperature, require the washing of the treated 

 combs in water in order to prevent the formation of an undesirable 

 residue caused by the retarded evaporation, the combs should be 

 treated 48 hours, whether in water-formalin or in alcohol-formalin 

 solution; an additional 12 hours in the case of alcohol-formalin 

 solution would give a greater margin of safety. If this is not done, 

 the discontinuance of the germicidal action may permit a few scales 

 to emerge with spores still capable of causing disease, while a period 

 of treatment shorter than 48 hours before washing in water would 

 be entirely insufficient for complete germicidal action. 



The uncapping of all brood cells removes the necessity of using 

 a solution capable of penetrating the wax. With the cappings re- 

 moved, the dissolving action, or penetration of the wax of the comb, 

 seems of questionable value. It is noteworthy that not one case has 

 definitely been recorded of disease resulting from the use in healthy 

 colonies of tons of comb foundation that for years has been made 

 from wax rendered from combs once containing American foulbrood. 



