Fasciola hepatica (liver fluke), is a trematode parasite, particularly of
ruminants, that is found throughout the world. Sheep show no resistance to the
parasite, cattle may develop partial resistance and rats show complete resistance,
which is immune-mediated, to challenge infections. In rats, resistance occurs early
during infection, at the level of the gut and/or peritoneum. Little is known regarding
the natural immune responses in cattle that occur during this period.
This study was designed to investigate the effect that pre-exposure of cattle to
1-2 and 5-6 day old flukes had upon subsequent challenge infections, when
compared to naive and chronically infected animals. Infection parameters, together
with peripheral and local cellular and antibody responses to various F. hepatica
protein preparations were investigated.
A primary exposure to 5-6 day old flukes, terminated by triclabendazole
treatment, was found to reduce the degree of liver damage and eosinophilia
experienced after a challenge infection. Lower levels of the enzymes gammaglutamyl transferase and glutamate dehydrogenase were detected in sera, compared
to those in naive animals (P < 0.05). Eosinophilia was also reduced (P < 0.01), as
was egg output in the faeces during the early patent period (P < 0.05). Calves that
were pre-exposed to 1-2 day old flukes showed no significant differences in the
levels of these parameters, when compared to their naive counterparts.
Sera taken prior to the secondary challenge from pre-exposed animals and
those receiving an unterminated primary infection recognised a variety of proteins in
Western blots of whole adult fluke somatic antigen (WFA) and excretory-secretory
(ES) preparations. The IgGl antibody response to protein bands of 96-82, 76-68 and
60-52 kDa predominated. After secondary challenge the response of the chronically
infected animals to these protein species was reduced and extremely strong
recognition of bands in the region 30-28 kDa was observed. Pre-exposed animals
maintained the response to the higher weight bands, showing a similar, but initially
stronger recognition pattern, to that of naive challenged animals. The lower weight
protein bands were not detected in these groups until much later.
The IgGl/IgG2 isotype antibody response to purified cathepsin and haemcontaining high molecular weight fractions were also examined by ELISA. A
monophasic, IgGl response was seen to the cathepsin fraction, which occurred late
during the infection process and was not seen prior to secondary challenge in any of
the four groups. A mixed IgGl/IgG2 antibody response to the haem fraction was
seen within 14 days of primary infection. After secondary challenge, this response
was boosted in the pre-exposed, but not the chronically infected, animals. No
significant difference was noted in antibody titre to either protein fraction between
pre-exposed or nai've animals.
Peripheral blood mononuclear cells responded to stimulation with WFA and
ES preparations, giving a strong proliferative response as early as 7 days post¬
infection. This suggests the presence of common antigens between adult and very
early fluke stages. Proliferation was decreased after secondary challenge, particularly
in the chronically infected group.
In order to examine the local immune response in the 5-6 day pre-exposed
group, mesenteric and hepatic lymph nodes were removed 10 days after secondary
challenge. Substantial proliferation to WFA was seen with hepatic, but not
mesenteric derived lymph node cells, suggesting that the gut response to this antigen
preparation was not important. Pre-exposed animals showed a lower mean level of
IFN-y and IL2 cytokines in supernatant fluid. Both groups produced similar levels of
B cell stimulating factor. IgG antibody present in hepatic lymph node cell culture
supernatant fluid from pre-exposed animals recognised proteins on WFA and ES
Western blots of sizes 190-120, 96-82, 72-68 and 60-57.5 kDa. No such response
was seen with supernatant fluid from naive challenged animals.
These results suggest that pre-exposure to flukes of up to 5-6 days old
provides a degree of protection against secondary challenge infections in cattle.
Peripheral and local antibody is mainly directed against a series of proteins ranging
from 96-57.5 kDa in size and to particular bands in metacercarial preparations.
Although the main antibody response was IgGl dominated, the cellular response
showed a mixed cytokine pattern, suggesting an unrestricted T helper cell response.
Further investigation into the nature of immunoreactive bands identified, together
with examination of IgE antibody responses, will aid understanding of the naturally
occurring immune response during early infection.