P. larvae Project Pilot

The overall goal of this experiment is to determine LD50 for P. larvae at the first and second instar in honey bee larva. This will likely require around 100 bee larva to accomplish. We have a number of other possible directions to head with this project, but the LD50s will be the start. Right now, I am focusing on culturing, creating freezer stocks, and isolating spores from P. larvae. As only the spores are invective to the bees, that is what I'll need.

Paenibacillus larvae arrived in the mail today from Microbiologics. ATCC number 9545

Struck out onto Chocolate Agar and MYPGP Agar and incubated at 37C 5% CO2. P. larvae grows very well in bee hemolymph, but is otherwise difficult to grow in media. Growth on MYPGP seemed to result in higher sporulation, per literature review.

Agar information can be located at the Coloss Honey Bee Research Association (Link).
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 MYPGP agar (per litre):

• 10 g Mueller-Hinton broth (Oxoid CM0405)
• 15 g yeast extract
• 3 g K₂HPO₄
• 1 g Na-pyruvate
• 20 g agar
•  Autoclave at 121°C / 15 min.
•  Add 20 ml 10 % glucose (autoclaved separately).

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Incubation of the P. larvae plates will likely take 6-7 days before spore collection. 

//EWW