Abstract
The central aim of this study is to investigate the mechanisms of action
of plant secondary metabolites (PSM), their direct effect on gastrointestinal
nematodes (GIN) and their indirect effect on the local immune response of
parasitised sheep. The first approach was through an in vivo study using a
tannin-rich crop, sainfoin (Onobrichys viciifolia), for feeding growing lambs
trickle infected with Trichostrongylus colubriformis. Thirty-two lambs, 16
weeks of age, were allocated to 1 of 4 treatment groups (n=8) that were
offered either grass (G) or sainfoin (S) hay from day -14, while concurrently
either infected (+), or not (-) with 12 000 L3 T. colubriformis larvae per week,
from day 0, for 6 weeks. Liveweight gains were affected by diet (P=0.002) and
reduced by infection (P<0.005). Faecal egg count was reduced in the S+ group
compared with the G+ group from days 35 to 42 (P=0.001); however, total egg
output, worm burdens at day 42 and worm fecundity were similar between
diets (P>0.05). Feeding sainfoin appeared to enhance immune cell
development with tissue eosinophils, mast cells and Pan T cells present in
greater concentrations in S+ than in G+ animals. However, further studies are
required to determine if the enhanced immune cell development is a
consequence of a greater nutrient supply or a direct influence of sainfoin
metabolites on local inflammatory responses to the gastrointestinal nematode
T. colubriformis.
The second approach was to investigate in vitro the potential anthelmintic
effect of PSM different from tannins, namely plant lectins.
Phytohaemagglutinin E3L (PHA-E3L), Wheat germ agglutinin (WGA) and
Concanavalin A (Con A or Jack bean lectin) were used with the larval
feeding inhibition assay (LFIA), where the direct effect of the lectins at
different concentration on the feeding of first stage larvae (LI) was
investigated. The results showed that these plant lectins had a profound
effect on the feeding of the larvae, especially PHA E3L (P=0.0006) at low
concentrations, while Con A and WGA were effective at higher
concentrations and T. colubriformis appeared to be the most susceptible
(P=0.07) of the three nematode species tested.
In a third approach the intriguing results achieved with the LFIA employing
plant lectins were confirmed through an in vivo experiment. Twenty-four
lambs were allocated to one of four groups (n=6) in a 2x2 factorial design
with treatments being either infection (-P: no infection vs. +P: mixed infection
with Teladorsagia circumcincta and T. colubriformis) or PHA administration (-L:
no PHA vs. +L: oral dose of 80 mg PHA/animal/d), for 6 weeks. Animals
received a dose of 167 mg/animal/d of 40.24% w/w semi-purified PHA lectin
(equivalent to 80 mg of pure PHA) administered orally five times per week.
Compared with their non-dosed counterparts, PHA significantly reduced the
faecal egg counts (epg) between days 25 and 36 post infection (P=0.033) and
had a
tendency to reduce the ability of larvae to penetrate abomasal tissue
(P=0.063), as evaluated by an in vitro direct challenge assay (IVDC) used to
investigate the effect of lectin treatment on the ability of the larvae to
penetrate the mucus layer and migrate to the gastric pits in the abomasal
tissue. PHA did not affect cell populations of the intestine. In abomasal tissue
of parasitised animals, PHA induced an increase in the number of
eosinophils (P<0.001) and PAS-positive cells (P=0.034). No changes in
mucosal mast cells were observed in any of the animals. These results
indicate that the oral dosing of PHA may affect local mucosal immune
responses during GIN infection, although further studies are required to
define both the direct and indirect effects of PHA in vivo.
A clear direct effect of lectins in vitro on the feeding of first stage larvae was
found in the laboratory and clear evidence of an enhanced immune response
of the host was shown through both in vivo studies. However, there is still
more studies to be done to find the mechanisms of action of PSM on the GIN
of sheep. The use of PSM or nutraceutical plants, seen from the perspective of
being able to influence the immune response of the animals,
'immunoceuticals' is an important target for future studies. If we are able in
the future not only to have a direct effect on the worms but also to induce
changes in the immune response of the animals through their food,
alternative parasite control methods could have an important role to play in
commercial farm conditions.
