10 CIRCULAR 3 92, U. S. DEPARTMENT OF AGRICULTURE 



To run the test, place the sample in the vial, and add 20 drops of 

 warm water (about ±4 of a vial) and shake gently; then add 10 drops 

 of the powdered milk suspension and again shake gently. After this 

 do not further distrnb the vial. If less than an entire scale is avail- 

 able, add 20 drops of water as before, but reduce the number of drops 

 of milk proportionately. Until one is familiar with the test it is 

 advisable to have a check vial, with only water and milk in suspension 

 for comparison. There may or may not be a fine curd after 5 minutes, 

 but this is not a significant part of the test. The test is positive (i. e.. 

 American f oulbrood is present ) if the milky suspension clears, usually 

 within 15 minutes, leaving a transparent, pale-yellow liquid. Some- 

 times the clearing is so rapid that a test is definitely positive after 5 

 minutes. The difference between a positive test and a negative or 

 check test is very striking if the vials are held to the light. With non- 

 American foulbrood scales the liquid may sometimes become dis- 

 colored, but the suspension remains cloudy during the 15 minutes of 

 the test, and the test is considered negative. 



In case only one or a few tests are to be run. add 20 drops of warm 

 water as before and shake : then add 5 drops of fresh skim milk, shake, 

 and read as before. Pasteurized milk should not be used, since it 

 does not give consistent results. If less than an entire scale is avail- 

 able, reduce the amount of milk proportionately. For example, if 

 half a scale is tested, add 2 to 3 drops of milk. 



Care should be exercised in disposing of the material in the vial after 

 the test is run. The vials should be washed clean, and boiled 20 

 minutes in water before they are used again. 



It has been noted that scales treated with formaldehyde fumes 

 give a negative reaction. Similarly, the test is retarded when scales 

 are exposed to paradichlorobenzene vapors. On the other hand, sulfa 

 drugs had no effect. In any doubtful cases the Division of Bee Culture 

 of the Bureau of Entomology and Plant Quarantine will make a 

 laboratory analysis of the material. 



European Foulbrood 

 cause 



European foulbrood is an infectious bacterial disease of the brood 

 of honeybees. The bacteria grow within the stomach of infected 

 worker, queen, and drone larvae and cause their death, but pupae are 

 rarely attacked. Adult bees are not affected by this disease. 



The earliest studies on European foulbrood seemed to indicate that 

 it was caused by a rod-shaped bacterium. Bacillus alvei, which is 

 commonly found in decayed brood. However, pure cultures of this 

 organism did not produce European foulbrood. Later it was observed 

 that lancet-shaped bacteria, different in shape and size from the rods 

 and spores of B. alvei found in decayed brood, are usually present in 

 large numbers in sick and recently dead larvae. This lancet-shaped 

 bacterium, which was given the name Bacillus pluton, is now com- 

 monly considered to be the cause of European foulbrood. It has been 

 found recently, however, that the rod-shaped B. alvei and Bacterium 

 t urydice are capable of changing their form to a lancet-shaped bac- 

 terium resembling B. pluton or other forms of bacteria found in larvae 



