Freezing and Storing Ditylenchus dipsaci in Liquid Nitrogen.

Area of Science:

• Nematology
• Cryobiology
• Plant Pathology

Background:

• Ditylenchus dipsaci, a significant plant-parasitic nematode, requires effective long-term storage methods for research and management.
• Previous methods for cryogenic storage of nematodes often necessitated the use of the quiescent stage, limiting applicability.

Purpose of the Study:

• To evaluate the viability of Ditylenchus dipsaci larvae after long-term cryopreservation.
• To determine if active larvae, rather than quiescent ones, can be successfully stored at cryogenic temperatures.

Main Methods:

• Larvae of Ditylenchus dipsaci were treated with a 7.5% solution of dimethyl sulphoxide.
• Treated larvae were cooled to -25°C and stored for 18 months at -150°C.
• Viability was assessed upon thawing, with survivors tested for their ability to penetrate and develop in alfalfa seedlings.

Main Results:

• A significant proportion of Ditylenchus dipsaci larvae remained viable after 18 months of storage at -150°C.
• Surviving larvae successfully penetrated and developed within alfalfa seedling stems, indicating normal biological function.
• Direct freezing of active larvae proved effective, negating the need for the quiescent stage.

Conclusions:

• Cryopreservation using dimethyl sulphoxide and low temperatures is a viable method for the long-term storage of Ditylenchus dipsaci.
• This technique simplifies nematode preservation and can potentially be adapted for other plant-parasitic nematode species.