DISEASES OF BEES 13 



almost always lanceolate-shaped with definitely pointed ends. 

 When " S. apis " is demonstrable microscopically in affected larvae 

 it can always be cultured readily on a variety of substrates. 



" S. apis " is not, as has hitherto been assumed, a single species, 

 and already three different varieties have been isolated. Two of 

 these appear to be identical with 5. liquefaciens and S. glycerinaceus, 

 which are well-known lactic acid bacteria. Though it may prove to 

 be more convenient to speak of these species collectively as " S. 

 apis " when they are foimd in larvae affected with European foul 

 brood, the term is strictly incorrect, and for purposes of classifica- 

 tion the species name should be dropped. The source of these species 

 of lactic acid bacteria is uncertain, but it is likely that the bees 

 come in contact with them when collecting water. Such species 

 are commonly found in cattle faeces. The fact that " S. apis " 

 does not exist as a distinct species makes it increasingly difficult to 

 accept the theory that European foul brood is caused by a pleo- 

 morphic organism (Lochhead, Bumside), especially in view of other 

 evidence given in this paper. 



The intestine of a single larva recently infected with European 

 foul brood, and containing B. pluton in apparently pure cultxire, 

 will soon initiate disease in a healthy nucleus when it is emulsified 

 in water and the resulting suspension is sprayed over developing 

 brood. However, as in the case of American foul brood, a certain 

 " mass inoculum " of B. pluton cells is necessary before disease can 

 be produced in a healthy colony. In one experiment a fairly uniform 

 suspension of B. pluton cells was prepared by macerating the guts 

 of two recently infected larvae, in which this organism appeared 

 in apparently pure culture, in 25 cc. of dilute phosphate buffer 

 (pH. 7.2). When portions of this suspension were examined bac- 

 teriologically, no coccus shaped cells grew on the medium used, the 

 suspension being practically " sterile " as regards bacteria capable 

 of multiplying on this medium, only a few colonies of small rod- 

 shaped bacteria appearing (probably Bacterium eurydice). The 

 approximate number of B. pluton cells in this suspension was 

 estimated using a Thoma haemocjrtometer slide. Three different 

 healthy nuclei were inoculated on June 25 by feeding them 1, 5 

 and 10 cc. amounts of this suspension in 30 cc. of syrup. Within 

 18 days the two nuclei receiving the larger doses (approximately 

 980 and 490 million cells of B. pluton) showed European foul 

 brood, while the nucleus receiving the smallest dose (approximately 

 98 million cells) showed no disease up to the beginning of September. 



All attempts to initiate European foul brood in healthy nuclei, 

 by spraying the developing brood with suspensions containing large 

 numbers of the organisms occurring as secondary invaders in this 

 disease (" S. apis," Bacterium eurydice, B. alvei or B. para-alvei), 

 have met with failure. However, it has been found that the course 

 of the disease can be profoundly modified by spraying cultures of 

 certain of these organisms over the developing brood of nuclei 



