12 Department Circular 2S4, U. S. Dept. of Agriculture 



of the wax, as well as with the fatty residue in the diseased remains 

 of the brood, thereby nullifying the action of the iodine as a germi- 

 cide. 



The use of denatured alcohol in various dilutions containing 5 

 per cent concentrated hydrochloric acid, and a small quantity of 

 formalin to prevent deleterious action of the acid (10) on the 

 supporting wires of the frame, gave unsatisfactory results with im- 

 mersions of 24 and of 43 hours. Each of the sealed cells tested gave 

 a good growth of Bacillus larvae: in two cases growth was obtained 

 from open cells, although some spores apparently were killed. This 

 type of solution was not tested further. 



Varying quantities of acetic acid added to a 20 per cent solution 

 of formalin in water contributed nothing to the germicidal or pene- 

 trating action of the solution so far as sealed cells were concerned. 

 as several such cells in each series tested gave each a good growth of 

 Bacillus larvae. These results are very similar to those obtained 

 with plain water-formalin solution, which will be considered later. 



Several other miscellaneous solutions were tried whose composition 

 is indicated in the table, all containing 20 per cent of formalin, but 

 varying in the composition of the carrier. All proved unsatisfactory 

 when used with sealed cells, and will not be discussed further. 



SOLUTIONS WITH DILUTED ALCOHOL 



A few dilutions of denatured alcohol, as well as dilutions of 

 alcohol-formalin solution, were made, varying the alcohol content 

 from about 30 per cent to over 60 per cent, as indicated in Table 4, 

 the 20 per cent formalin, however, being kept constant in the mix- 

 tures with denatured alcohol. Tests were made of combs treated 

 for 24 hours and for 48 hours. In every case no growth was ob- 

 tained from open cells, and, as would be expected, there were fewer 

 positive cultures from sealed cells in combs treated 48 hours than 

 from those treated 24 hours, although several sealed cells showed no 

 growth, even in the 24-hour series. In each of the 43-hour series 

 there was at least one sealed cell from which was obtained a growth 

 of Bacillus larvae, but, with the few observations made, no signifi- 

 cant differences could be found between the few dilutions tested of 

 alcohol or alcohol-formalin solution. Further work is necessary to 

 demonstrate whether results comparable with those of Kronig and 

 Paul can be obtained. 



For purposes of comparison, as a preliminary test, three series 

 of six samples each from diseased combs were treated with a solution 

 containing no alcohol, composed of formalin 20 parts and water 

 80 parts (Table 9). No significant difference could be seen between 

 these results and those obtained with the various alcoholic solutions. 

 No open cells treated with this solution gave cultures showing 

 growth, but 8 sealed cells, 1 or more in each series, gave cultures 

 showing growth of Bacillus larvae, the growths in 3 of the 8 cultures 

 being noted as " few spores germinated." 



