A technique has been elaborated that enabled the plerocercoid larvae of Schistocephalus solidus to be removed from the body cavity of Gasterosteus aculeatus without bacterial contamination. Larvae were cultured in plugged test-tubes under completely aseptic conditions in a variety of balanced salines, glucose salines and nutrient peptone broth.
The most successful results were obtained with peptone broth at room temperatures (16-19° C) in which plerocercoids remained active and showed normal behaviour for periods up to 300 days. In ¾ strength Locke's solution, which was found by experiment to be approximately isotonic with Schistocephalus (δ = -0.44 ± 0.02° C), the mean period of normal behaviour was 114 days. In the remaining saline and saline-glucose media, the mean viability and period of normal behaviour was considerably less.
In the plerocercoid, histological examination revealed that the genitalia are in an immature condition. During cultivation at room temperatures, the genitalia remained in this undifferentiated condition and showed no signs of undergoing spermatogenesis, oogenesis or vitellogenesis.
Plerocercoids were induced to develop into sexually mature adults by raising the temperature of cultivation in peptone broth to 40° C. (i.e. the body temperature of the final host in the natural life cycle). Oviposition took place after 48-60 hr. at this temperature, and histological examination revealed that spermatogenesis, oogenesis, vitellogenesis and shell formation had taken place in a normal manner. The viability of artificially matured Schistocephalus was 4-6 days in vitro--a period equivalent to the viability of the adult in vivo.
The eversion of the cirris was observed in each proglottid after 40 hr. cultivation at 40° C. During the sexual process the cirris everted and invaginated at the rate of about once per second. Cross-fertilization between segments of the same worm or with segments of another worm was not observed. Except for one specimen in ¾ strength Locke's solution which underwent spermatogenesis and partial vitellogenesis, larvae cultured in salines or glucose salines at 40° C. died within 1-3 days without further development.
Attempts to hatch out the eggs produced by the cultivation of larvae in peptone broth at 40° C. proved unsuccessful. Histological examination revealed that spermatozoa had not been taken into the vagina. It was concluded that the eggs were not fertilized owing to the failure of normal copulation to take place.