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# 2.2.4.2.2. Spore suspension concentration

Dilution of the purified spore suspension to the desired concentration can be performed using the formula: C1V1 = C2V2 where: C1 = initial concentration; V1 = initial volume; C2 = final concentration; V2 = final volume.

For example, if you want to feed 10,000 Nosema spores in 10 µl 50 % (weight / volume) sucrose solution, you must, as determined by proportions, create a final spore concentration of 1,000,000 spores / ml of 50 % (w/v) sucrose solution. To create 10 ml of this final spore suspension in 50 % sucrose solution when your initial spore suspension is:

Example 1. 10,000,000 spores / ml water, then 1 ml of initial spore suspension must be added to 9 ml 55.6 % (w / v) sucrose solution. See below for specific details:

Calculating initial volume of spore-water suspension needed

1. C1V1 = C2V2
2. V1 = C2V2/C1
3. V1 = 1,000,000 spores per ml X 10 ml / 10,000,000 spores per ml
4. V1 = 1 ml

Calculating initial concentration of spore-free sucrose solution needed*

1. C1V1 = C2V2
2. C1 = C2V2 / V1
3. C1 = 50 g sucrose per 100 ml water X 10 ml final sucrose solution / 9 initial sucrose solution
4. 55.56 g sucrose per 100 ml water

*An initial concentration of spore-free sucrose solution of 50 % is generally adequate when a low volume of spore suspension is added to create a final spore suspension in approximately 50 % sucrose solution.

Example 2. 12,456,000 spores / ml water, then 0.8 ml of initial spore suspension must be added to 9.2 ml 54.3 % (w / v) sucrose solution.

Control suspensions to infected control bees should be created using uninfected individuals from the same colonies the spores were sourced from.