Direct Count Method [164]
This method is used when the numbers of nematodes are less than
50.
Materials Required
l
Nematode suspension.
l
Petri dish.
l
Micropipette.
l
Dissecting microscope.
Procedure
1. Transfer the nematode suspension (n < 50) into a petri dish
using a micropipette.
2. Sum the nematodes in the suspension independently under a
dissecting microscope (magnification 40 for infective juveniles; 10–20 for adults).
Dilution Method [164]
This method is used when the number of nematodes is more than
50.
Materials Required
l
Nematode suspension.
l
Micropipette.
l
Beaker.
l
Nematode counting slide (eelworm counter).
l
Dissecting microscope.
Procedure
1. Prepare a dilution of about 100 IJs/ml to facilitate counting.
2. Mix the nematode initial suspension in a beaker to get a homogeneous mixture.
3. Take 1 ml of nematode initial suspension and add to X ml of
water, where X is projected to produce another suspension of
around 100 IJs/ml, i.e., 1: X + 1 dilution.
4. Take samples from a systematically mixed, diluted suspension
culture.
5. Place 1 ml suspension on eelworm counter (Fig. 10).
6. Count the IJs under a dissecting microscope.
7. Repeat the counting thrice or more by taking fresh samples
every time.
8. Determine the concentration of nematodes in the original
suspension, using the following formula:
S ¼
N Â 1 X þ 1
ð
Þ
M
N ¼ average number of nematodes per counted sample.
M ¼ number of ml per counted sample.
66
Laboratory Techniques for Entomopathogenic Nematodes