Immunity to Fasciola hepatica in rats: the effect of different levels of primary exposure on superinfection.

Abstract

Seventy-six per cent fewer flukes developed from a challenge infection in rats with an existing infection of one F. hepatica than in rats not previously exposed. Rats first exposed to 10 metacercariae were able to resist completely a challenge exposure. Existing fluke burdens from the primary exposure were not significantly affected by challenge. The results corroborate and extend an earlier study on immunity to superinfection with F. hepatica in rats, and the ability of one existing fluke to cause a marked reduction from challenge is taken to indicate that nonspecific liver inflammation and damage existing at the time of challenge may not play a necessary role in immunity to F. hepatica in rats. Hayes et al. (1972) reported that rats are highly immune to superinfection with Fasciola hepatica. Previous reports had indicated that exposure of rats to irradiated metacercariae, lymphoid cells from infected donors, or multiple stimulating infections (Thorpe and Broome, 1962; Corba et al., 1971; Riiter cited in Geyer, 1971) caused reduced fluke recoveries from challenge in rats. Much work remains to be performed, however, to characterize the rat's ability to respond against F. hepatica, and to determine the mechanism involved in immunity. The study reported herein was designed to determine whether minimal infection with F. hepatica (one metacercaria/rat) would affect a challenge infection, to compare this situation with rats exposed to a higher level of infection (10 metacercariae/rat), and to evaluate the effect of challenge on an existing infection. MATERIALS AND METHODS The experiment consisted of rats that were exposed to 1 or 10 metacercariae (cysts) of F. hepatica per rat followed by reexposure to 10 cysts per rat 49 days later and the control group necessary to show expected fluke burdens from each exposure. For this purpose, 100 male outbred albino rats (Charles River, CD) 27 days old and with a mean weight of 85 g at the time of first exposure (experiment day 0) were divided into 6 groups. Group 1 consisted of 30 rats exposed to 1 cyst per rat on day 0 and then to 10 cysts on experiment day 49, Group 2 of 30 rats exposed to 1 cyst per rat on day 0 only, Group 3 of 10 rats exposed to 10 cysts per rat on day 49 only, Group 4 of 10 rats exposed to 10 cysts per rat on day 0 only, and Group 5 of 10 rats Received for publication 26 March 1973. exposed to 10 cysts per rat on day 0 and again on experiment day 49. A host control consisting of 10 rats not exposed at either time was also kept and found to be negative for liver lesions at necropsy on experiment day 76 (Group 6). Metacercariae were collected from the walls of glass aquaria containing Lymnaea tomentosa artificially infected by exposure to miracidia of F. hepatica hatched from eggs recovered from the bile of experimentally infected sheep. The cysts were individually selected on the basis of typical internal morphology (Hayes, 1970) and given to rats orally. All cysts used in this experiment were from snails infected on the same date, shed 6 to 7 weeks after snail exposure, pooled, and stored at 4 C till needed. Six of the 100 rats died prior to scheduled termination, one in Group 1, two in Group 2, and three in Group 4. None of these deaths could definitely be ascribed to fascioliasis, and data from these animals are not included in the results. One in Group 4 that died 1 day prior to termination of the experiment was necropsied, and had 8 flukes in the bile duct and none in the liver parenchyma. Necropsy was performed on rats exposed on experiment day 0 only (Groups 2 and 4) and on the host control (Group 6) on experiment day 76. Rats exposed on experiment day 49 (Groups 1, 3, and 5) were necropsied on experiment day 77 (28 days after the second infection). The liver was examined in situ and any flukes in the extrahepatic portion of the biliary system were collected (bile duct flukes). The liver was then re oved, placed in saline, manually crushed, teased apart, and the flukes recovered. Following storage for 18 hr or longer in saline at 4 C, the flukes were measured (mean length followed by range in mm given in results). Flukes from each infection of the doubly exposed rats were distinguished by virtue of a 7-week difference in age between them and were thus disparate in size, degree of maturation, and site of location in the liver.