Apr. 12, 1924 
Development of American Foulbrood 
135 
that of larvae at the age when European foulbrood makes its attack, comprises 
nearly 45 per cent of the dried substance, or nearly 14 per cent of the fresh 
substance. From these facts it may be assumed that, because of the great 
amount of food given the larva at this age, there must be present, in the larval 
intestine, at all times during the active feeding period, considerable amounts 
of this food rich in sugar unassimilated, up to and even after active feeding 
ceases. A number of larval and prepupal intestines were dissected from 
healthy larvse and tested roughly with Benedict’s qualitative solution (34) 
for the presence of reducing sugars. The results indicated the presence of 
relatively large amounts of reducing sugar in the intestines of larvae just prior 
to sealing. Little or no reducing sugar could be demonstrated in the intestines 
of sealed larvae or prepupae. 
It may therefore be assumed that certain of the organisms associated with the 
early stages of European foulbrood are able to grow in the presence of a high 
sugar concentration. Experiments were devised in 
which a medium containing 10 per cent dextrose was 
used. It was found that while a few types of organ¬ 
isms, such as one resembling Streptococcus apis (28), 
could be grown in varying numbers, an organism sim¬ 
ilar to that described by Maassen (29), resembling the 
larger forms of Bacillus pluton, called Bacillus lanceolatus, 
could be isolated and grown from over 50 per cent 
of the samples cultured. As described by the writer 
(47), “This organism w^as found-to grow best on a 
10 per cent dextrose yeast extract agar with a reac¬ 
tion slightly acid. It is differentiated from Bacillus 
pluton and Streptococcus apis in being gram-negative, 
and does not grow readily if at all in media with¬ 
out sugar.” From these studies it was suggested 
that possibly this comparatively high sugar content of 
the unassimilated food in the larval intestine may 
have an influence on the germination of the spores and growth of Bacillus 
larvae and that a change may occur when the sugar content is sufficiently re¬ 
duced by assimilation in the larval intestine after it has been capped and when 
the intestines have been emptied by the opening of the ventriculus into the 
end gut. Therefore, with these preliminary observations as a basis, experi- 
nental work on this subject was begun during the spring of 1922. 
GROWTH OF BACILLUS LARVAE IN CULTURES IN RELATION TO 
VARIATION IN SUGAR CONCENTRATION 
The first step in the substantiation of this theoretical assumption is to de¬ 
termine whether there is a correlation between germination of the spores of 
Bacillus larvae and vigor of vegetative growth and variations in concentration of 
reducing sugars in culture media. Ordinary culture media are unsuitable for 
the growth and isolation of Bacillus larvae; in fact, one of the confirmatory testa 
for this organism in laboratory diagnosis of American foulbrood (81) is the 
absence of growth on plain beef infusion agar plates, since the spores will not 
germinate thereon. There are rarely any secondary invaders associated with 
Bacillus larvae in the decayed material, and these plates practically never show 
growth. 
GROWTH REQUIREMENTS OF BACILLUS LARVAE 
Various special culture media have been devised which answer more or less 
satisfactorily the requirements for the ordinary growth of the organism. The 
Fig. 5—Healthy coiled larva 
at age of maximum intestinal 
sugar content and approxi¬ 
mately the age when the ma¬ 
jority die from European foul¬ 
brood. (White ( 56 )) 
