Temperatures may need adjustment when incubation and storage of nematode strains/species have specific adaptation to either
cold or warm surroundings. The nematodes are then collected into
tissue culture flasks with distilled water and placed in storage.
3.7.1 Materials Required
l
Nematode infective juveniles.
l
Tissue culture flask.
l
BOD incubator.
l
Distilled water.
l
Triton X-100.
3.7.2 Procedure
1. Take suspensions of nematode species.
2. Further dilute the suspension with approximately 1500–3000
IJs/ml concentration of distilled water.
3. Take a clean tissue culture flask (250 ml) and pour the
suspension.
4. Gently pour a drop of triton X-100 to avoid the IJs on adhering
to the flask surface.
5. Close with a lid and label it.
6. Nematode in distilled water suspension must be at not more
than 1 cm in depth to ensure sufficient oxygen availability.
7. Store the culture flask in BOD incubator at 10–15
C until
further use (Fig. 11).
Note: Steinernematids can be stored up to a maximum of 6–9
months at 4–15
C.
Heterorhabditids can be saved for 3–4 months at 4–15
C.
4 Morphological Characterization
Accurate documentation/analysis of these taxa necessitates the
consideration of suitable taxonomic methods. However, recognizing a species appropriate to either the genus Steinernema or Heterorhabditis is rather multifaceted, involving morphometry, DNA
analysis, and mating studies. An understanding of the identification
and characterization of the nematode species is important for their
utilization as biocontrol agents.
Morphological characteristics for species identification require
substantial skills to ensure accuracy. The morphology of nematodes
varies between the species. The morphometry is based on the
length and width of the body or of specific anatomical features of
the nematode. Nematodes are examined either live or heat killed.
68
Laboratory Techniques for Entomopathogenic Nematodes
cold or warm surroundings. The nematodes are then collected into
tissue culture flasks with distilled water and placed in storage.
3.7.1 Materials Required
l
Nematode infective juveniles.
l
Tissue culture flask.
l
BOD incubator.
l
Distilled water.
l
Triton X-100.
3.7.2 Procedure
1. Take suspensions of nematode species.
2. Further dilute the suspension with approximately 1500–3000
IJs/ml concentration of distilled water.
3. Take a clean tissue culture flask (250 ml) and pour the
suspension.
4. Gently pour a drop of triton X-100 to avoid the IJs on adhering
to the flask surface.
5. Close with a lid and label it.
6. Nematode in distilled water suspension must be at not more
than 1 cm in depth to ensure sufficient oxygen availability.
7. Store the culture flask in BOD incubator at 10–15
C until
further use (Fig. 11).
Note: Steinernematids can be stored up to a maximum of 6–9
months at 4–15
C.
Heterorhabditids can be saved for 3–4 months at 4–15
C.
4 Morphological Characterization
Accurate documentation/analysis of these taxa necessitates the
consideration of suitable taxonomic methods. However, recognizing a species appropriate to either the genus Steinernema or Heterorhabditis is rather multifaceted, involving morphometry, DNA
analysis, and mating studies. An understanding of the identification
and characterization of the nematode species is important for their
utilization as biocontrol agents.
Morphological characteristics for species identification require
substantial skills to ensure accuracy. The morphology of nematodes
varies between the species. The morphometry is based on the
length and width of the body or of specific anatomical features of
the nematode. Nematodes are examined either live or heat killed.
68
Laboratory Techniques for Entomopathogenic Nematodes