A preliminary study of the prospect of inducing immunity in fascioliasis by means of infections with X-irradiated metacercarial cysts and subsequent challenge with normal cysts of Fasciola hepatica L.

Extract

In three series of infections 32 mice were given metacercarial cysts of Fasciola hepatica X-irradiated at 3 Kr.: 8 mice received 20 cysts each and 10 mice 40 cysts each. In serial kills 9–24 days after infection 127 young flukes were recovered, 10 + 49 entire and 39 + 29 in serial sections of infected liver. These flukes were stunted and inhibited in their development. During a period of 8–22 days the length and breadth of the flukes increased from 0·53 mm. x 0·27 mm. to 1·12 mm. x 0·37 mm., whereas the mean length and breadth of normal flukes 22 days old is 3·4 mm. x 1·9 mm. The intestines of the flukes have about 10–14 lateral diverticula or sacculations but not normal branches during this period. In the sections of 12– to 29–day infections 16 dead flukes were observed in various stages of disintegration but flukes deemed alive at the time of fixation were noted in all stages from 8 days onwards, although some of them were injured by inflammatory processes set up in the liver. Injury is caused by massive infiltrations into the inner substance of the cuticle of band-type neutrophil leucocytes, which raise the epicuticle in extensive blister-like formations, but the basal portion of the cuticle (lamina vitrea) remains more or less intact. The cuticular sheets ultimately fragment, apparently as a result of chemical action, and leucocytes which had infiltrated into the cuticle rejoin the aggregate of similar cells inside the burrow. How the leucocytes effect an entry into the cuticle is not clear, but it is more probably by chemical than by physical means. When the epicuticle has been destroyed the fluke is in a dying condition. Few leucocytes penetrate into the parenchyma, suggesting that the lamina vitrea is relatively resistant to these cells and that death is due to chemical action following widespread but superficial damage to the cuticle.

The remaining 14 mice which received 40 X-irradiated cysts each were challenged 22 days later with 10 or 5 normal cysts. This procedure was followed by serial kills at intervals during the period 8–53 days after infection. Three almost mature flukes were recovered at 32 days and one mature fluke at 39 days. The mature fluke was of unusually small size (6·4 mm. x 4·3 mm.), but it had shed many viable eggs into the bile duct and gall bladder. The numbers of flukes of the challenge infections did not differ significantly from those of normal controls: 62 flukes in groups of 1–8 per mouse provided means of 4·4 flukes per mouse and 55·4 flukes per 100 cysts. In similar experimental infections of mice (which, however, received 2 doses of cysts X-irradiated at 4 Kr.) Hughes (1963) recovered 39 flukes in groups of 1–7 per mouse, providing means of 3·5 flukes per mouse and 35·3 flukes per 100 cysts. In his control experiments, which also serve as the writer's controls, 10 mice received 10 normal cysts each and in 22– to 29–day infections 34 flukes were recovered in groups of 3–5 per mouse, providing means of 4·25 flukes per mouse and 42·5 flukes per 100 cysts. There is therefore no evidence of immunity in terms of either delayed maturation or reduced fluke burdens. At all stages of infection flukes of the challenge infection bear comparison with flukes of single normal infections, except for an apparent diminution of growth (which requires confirmation) about 3–4 weeks after infection. This accords with generally accepted opinion that any small degree of immunity which is acquired by the hosts of liver flukes is almost undemonstrable. Further experiments with reduced periods between infections with X-irradiated cysts and a challenge infection may be profitable and are in progress but it is suggested that in such experiments nutritional effects may arise as a result of modification of the hepatic tissue on which young flukes feed by leucocytic infiltrations of the inflammatory reaction which is normally directed towards the repair of damage caused by the flukes.

My former pupil Denys H. Hughes, of the Parasitology Group, Allen and Hanburys (Ware), kindly set up the experimental mice to my specifications and I am deeply grateful to him for this kindness and consideration and for the respect he has shown for my wishes. Mice and cysts—X-irradiated as required— were supplied by Allen and Hanburys, for which my thanks are also proffered.

My personal technician, Mr John Wells, has attended to my technical needs constantly. In addition to making numerous series of excellent sections and whole mounts, he undertook the task of measuring several dimensions of more than 100 flukes. His patience and perseverance in the face of continual requests for more and more, good and still better materials has been commendable and I am glad to express my gratitude to him. I am grateful also to Mr R. D. Reed of this Department, who prepared the photomicrographs for Pls. 1–4 of this paper, sometimes when under the pressure of other duties.