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Animal
(2009), 3:12, pp 1644–1661 &The Animal Consortium 2009
doi:10.1017/S1751731109004236
animal
Natural alternatives to in-feed antibiotics in pig production: can
immunomodulators play a role?
M. Gallois
1
, H. J. Rothko
¨tter
2
, M. Bailey
3
, C. R. Stokes
3
and I. P. Oswald
1
-
1
INRA, UR66, Laboratoire de Pharmacologie-Toxicologie, Centre de Recherche de Toulouse, BP3 31931, Toulouse cedex 9, France;
2
Institut fu
¨r Anatomie,
Medizinische Fakultaet, Otto-von-Guericke-Universitaet Magdeburg, Leipziger Strasse 44, 39120 Magdeburg, Germany;
3
Division of Veterinary Pathology Infection
& Immunity, University of Bristol, Langford House, Langford BS40 5DU, UK
(Received 9 June 2008; Accepted 2 February 2009; First published online 13 March 2009)
As a result of the European ban of in-feed growth-promoting antibiotics, new strategies are being developed to increase
the resistance to disease in farm animals. In pig production, this is of particular importance during the weaning transition
when piglets are subjected to major stressful events, making them highly sensitive to digestive disorders. At this time, the
development of both innate and adaptive immunity at the mucosal surface is critical in preventing the potential harmful effects
of intestinal pathogenic agents. Strategies aiming at stimulating natural host defences through the use of substances able
to modulate immune functions have gained increasing interest in animal research, and different bioactive components
a
priori
sharing those properties have been the subject of
in vivo
nutritional investigations in pig. Among these, yeast derivates
(b-glucans and mannans) are able to interact with immune cells, particularly phagocytic cells. However, studies where they
have been fed to pigs have shown inconsistent results, suggesting that their ability to target the sensitive immune cells
through the oral route is questionable. The plant extracts, which would benefit from a positive image in the public opinion,
have also been tested. However, due to a lack of data on the bioactive components of particular plants and the large diversity
of species, it has proved difficult to prepare extracts of equivalent potency and thus, the literature on their influence on pig
immunity remains inconclusive. In considering piglet immunity and health benefits, the most promising results to date have
been obtained with spray-dried animal plasma, whose positive effects would be provided by specific antibodies and non-
specific competition of some plasma components with bacteria for intestinal receptors. The major positive effect of spray-dried
animal plasma is in reducing the infiltration of gut-associated lymphoid tissue by immune cells, which is likely to be the result
of a decreased colonisation by potentially harmful bacteria. This review also highlights the limitations of some of the published
in vivo
studies on the immunomodulatory activity of certain feed additives. Among those, the lack of standardisation of extracts
and the heterogeneity of piglet-rearing conditions (e.g. exposure to pathogens) are likely the most limiting.
Keywords: pig, immunity, disease sensitivity, feed additive, immunomodulators
Introduction
Adding sub-therapeutic doses of antibiotics (AB) to feed has
been widely used in the pig industry to enhance production
efficiency (Cromwell, 2002). Concerns about potential risks
for human health due to the use and misuse of AB in animal
feeds (Dewey
et al
., 1997) have led to their ban as growth-
promoters throughout Europe since 1 January 2006 (Reg-
ulation (EC) No. 1831/2003). Risks to human health include
the possibility of AB residues in meat, unapparent carriage
of anti-microbial drug-resistant bacteria, and exchange of
plasmids from AB-resistant bacteria of swine to human
pathogens making them resistant to AB (Dewey
et al
., 1997;
Anadon and Martinez-Larranaga, 1999; Pugh, 2002). In-feed
AB were used not only to improve growth but also to control
enteric infections during critical periods such as weaning.
Initial experience of an in-feed AB ban in Sweden and
Denmark indicated that there was a reduced performance and
increased morbidity in nursery pigs, which emphasised the
therapeutic use of AB in farms (Stein, 2002). This underlines
the need to develop alternative strategies.
In pigs, the post-weaning period is characterised by an
immediate, but transient, drop in feed intake, resulting in
alterations in gut architecture and function, making piglets
highly sensitive to digestive diseases (Pluske
et al
., 1997;
Lalle
`s
et al
., 2004). To help piglets to cope with this transition,
-
E-mail: Isabelle.Oswald@toulouse.inra.fr
1644
various nutritional approaches have been proposed (Lalle
`s
et al
., 2007), including supplementing the diet with sub-
stances that increase appetence or have anti-microbial and/
or immunostimulating properties. Amongst the alternatives
to in-feed AB, strategies aiming at boosting natural host
defences (e.g. immunomodulators), as opposed to those
that act directly on the microflora, are attracting a greater
level of attention. Indeed, the correct functional develop-
ment of the gastro-intestinal tract is of crucial importance in
controlling potential pathogens during the neonatal and
post-weaning period. The development of both innate and
adaptive immunity at the mucosal intestinal surface is cri-
tical in determining the outcome of the large exposure to
new antigens at these times. Paradoxically, the gut is
required to be sufficiently permeable to allow the absorp-
tion of nutrients, whilst at the same time it must prevent
the harmful effects associated with the replication and
absorption of potential pathogens (or their products). If for
the adult the main aim is the maintenance of gut homo-
eostasis, for the young animal it is to establish a favourable
steady state. Weaning affects the ontogeny of immune
functions, largely as a consequence of the withdrawal of
milk, which has important implications for passively mod-
ulating immune responses through both suppressive and
enhancing pathways. The aim of immunomodulating sub-
stances is to help piglets develop ‘appropriate’ active
responses from both innate and acquired immunity.
Immunomodulating substances used as in-feed additives
have to fulfil a variety of properties. From a technical point
of view, they have to be resistant to the processes used in
feed manufacturing. From a regulatory viewpoint, they have
to be safe for farm animals and must not affect food safety.
Those products should also share a positive image towards
the public, who are increasingly sensitive to ethical con-
siderations and whose opinion influences legislation (Flor-
kowski
et al
., 1998). Thus, products from natural sources
will probably be easily accepted by the public. Finally, these
alternatives must be effective in their purpose, namely to
act as growth-promoters and provide health benefits to
piglets, which will be the main focus of the present review.
Following a brief description of the development of the
piglet mucosal immune system, this paper will focus on
groups of in-feed alternatives that have been suggested to
show immunomodulatory properties and that have been the
purpose of
in vivo
investigations in pigs. These include
yeast derivates, plant extracts and animal by-products.
Piglet immunity: implications for the oral application
of immunomodulators
The immune response to oral delivery of antigens is orche-
strated by a well-developed local intestinal mucosal immune
system (Figure 1). On the one hand it must respond
with vigour to potential pathogens, whilst on the other
inappropriate responses to dietary antigens and commensal
bacteria may lead to harmful allergic or damaging inflam-
matory conditions. In adult animals, these potentially harmful
responses are downregulated by a process known as oral
tolerance, but at birth the piglet’s mucosal immune system is
poorly developed and it has been hypothesised that this may
lead to inappropriate responses to food and microbial anti-
gens in the post-weaning period. The kinetics of develop-
ment of the gut immune system, which will be described in
the following (summarised in Table 1), may therefore play
a critical role in determining the outcome of the response
to feed antigens.
The development of the gut mucosal immune system
Organised lymphoid tissues
. The porcine small intestinal
wall contains Peyer’s patches (PP) in the jejunum and ileum
(Figure 1). Whereas jejunal PP are several centimetres in
length and located at the anti-mesenteric site of the gut,
the ileal PP is a continuous band of up to 2 m in length
(Rothko
¨tter and Pabst, 1989). The postnatal development of
these two types of PP is different. For a short period of time,
the ileal PP resembles the characteristics of the ileal PP in
lambs with tall, elongated follicles and a small dome area
(Barman
et al
., 1997). Jejunal PPs have rounded follicles
and a larger dome area. Microfold cells (M cells) are present
in the follicle-associated epithelium of both types of PP
(Gebert
et al
., 1994), and play a specialised role in sampling
antigen from the gut lumen. Their microfolded apical cell
membrane enables endocytotic antigen uptake, and with
lymphocytes located in pockets of their basolateral cell
membrane, a close interaction of antigens and lymphocytes
is possible (Gebert
et al
., 1996). Similarly, cellular processes
of dendritic cells (DC) are also in close to the basolateral
membrane of M cells (Bimczok
et al
., 2006). Following
Table 1
Stages in the development of the gut mucosal immune system of the neonatal piglet
Postnatal period Development of the gut mucosal immune system
Stage 1 The newborn pig >Rudimentary Peyer’s patches
>Small numbers of mucosal antigen-presenting cells and T-cells
Stage 2 1 day to 2 weeks >Non-specific expansion of Peyers patches and B-cells
>Appearance of some conventional, activated, CD4
1
T-cells
>Influx of MHCII
1
cells
Stage 3 2 to 4 weeks >Appearance of mature CD4
1
T-cells
>Expansion of B-cell repertoire
Stage 4 4 to 6 weeks >Appearance of CD8
1
T-cells
In-feed modulators of piglet immunity
1645
antigen uptake and presentation in the PP, studies from
various species have demonstrated that PP B-cells migrate
via the mesenteric lymph prior to homing back to the
lamina propria crypts where they differentiate into immu-
noglobulin (Ig)-producing plasma cells (Bienenstock
et al
.,
1983). The rapid migration of IgA
1
immunoblasts from
intestinal lymph into the intestinal mucosa has also been
demonstrated in lymph duct cannulation experiments in
pigs (Rothko
¨tter
et al
., 1999b).
Lamina propria lymphocytes
. The lamina propria of adult
pig is well supplied with leucocytes (Figure 1), and in
contrast to many other species the immunological organi-
sation of the lamina propria in the pig intestine shows a
high level of organisation (Bailey
et al
., 1996; Haverson
et
al
., 1999). Within the villous lamina propria the tissue deep
to the capillary plexus contains predominately CD4
1
T-cells
whilst CD8
1
cells occur luminally and in the epithelium.
Antigen-presenting cells are present in large numbers in the
lamina propria of many species including the pig (Haverson
et al
., 2000). The lamina propria around the intestinal
crypts contains cells staining for Igs (predominantly IgA,
presumably plasma cells), with few T-cells or DC, but with
myeloid cells with the characteristics of macrophages and
granulocytes (Rothko
¨tter
et al
., 1991).
At birth, only small numbers of leucocytes are found in the
lamina propria and in pigs it becomes populated according
to a clearly staged time course. DC that are strongly major
histocompatibility complex class II (MHC II
1
) and co-express
CD45 and CD16 appear within the first week (Haverson
et al
.,
2000). In contrast, T-cells appear more slowly and undergo a
phased pattern of appearance. An unusual cell type, char-
acterised by the expression of CD2, but lacking CD4 and CD8,
has been shown to co-express CD3 and can therefore be
classified as CD4
2
CD8
2
T-cells (Rothko
¨tter
et al
., 1991;
Haverson
et al
., 1999). Together with a second T-cell
population, characterised as CD2
1
CD3
1
CD4
2
CD8a/a
1
,
they form the dominant T-cells migrating into the jejunal
tissue during the first week to 10 days of life. Interestingly,
while conventional CD4
1
and CD8a/b
1
T-cells in this site
in adult animals express low levels of CD45RC, consistent
with advanced memory status, a significant proportion of
these unusual CD2
1
CD3
1
CD4
2
CD8a/a
1
T-cells express
moderate to high levels of CD45RC, suggesting that they may
be less antigen-experienced (Bailey
et al
., 1998; Haverson
et al
., 1999). During the second and third weeks of life,
increasing numbers of CD4
1
T-cells appear and CD8
1
cytotoxic T-cells only appear in significant numbers after the
third week of life (Rothko
¨tter
et al
., 1991; Haverson
et al
.,
1999). Similarly, IgA
1
plasma cells have been reported
to appear in significant numbers as late as 3 to 6 weeks of
life. The final architecture containing large numbers of DC
and CD4
1
T-cells of resting, advanced memory phenotype,
transcribing interleukin-4 (IL-4) but being unable to secrete
IL-2 and responding to further activation by apoptosis is
not achieved until the pig is approximately 6 weeks old
(Bailey
et al
., 2005).
Intraepithelial lymphocytes
. The intraepithelial lymphocytes
(IEL) are a large population of cells with up to 2 310
7
cells
present per gram of porcine jejunal mucosa (Figure 1). They
are mostly CD8
1
T-cells with a significant proportion expres-
sing the CD8aa homo-dimer (Rothko
¨tter
et al
., 1999a). In the
postnatal period, the frequency of IEL increases with age and
this can be influenced by the luminal contents. For example,
their number is markedly reduced with total parenteral
nutrition (Kansagra
et al
., 2003).
The induction of immune responses in the postnatal period
In order to generate an immune response, the ingested
antigens must pass the epithelial barrier by either transcellular
(vesicular transport) or paracellular routes. The uptake of
B cell
follicle
M cell
B cells
Follicle-associated
epithelium
Lamina propria
Plasmocytes
DC
T cells
Enterocyte
IEL
Villus
Dome
DC
Crypt
T cells
DC
Interfollicular
areas
B cell
follicle
M cell
B cells
Follicle-associated
epithelium
Lamina propria
Plasmocytes
DC
T cells
Enterocyte
IEL
Villus
Dome
DC
Crypt
T cells
DC
Interfollicular
areas
Figure 1 Schematic representation of the distribution of ‘immune cells’ in the small intestinal tract. Abbreviations: IEL 5intraepithelial lymphocyte;
DC 5dendritic cell.
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1646
antigens occur via either fluid-phase or receptor-mediated
transport, followed by intracellular processing or simply
exocytosis.
Enterocytes are non-phagocytic cells that, in health, pre-
vent the passage of macromolecules through the epithelium
(Shah and Walker, 2002; Oswald, 2006). Additionally, access
to the enterocyte surface is restricted by local secretions like
IgA and mucins, closely packed villi, and a thick glycocalyx.
However, the epithelial barrier cannot completely prevent
antigen uptake (Macdonald and Monteleone, 2005). Although
the paracellular antigen entry by diffusion from the lumen is
avoided by tight junctions at the apical poles, antigens can
cross through breaks in tight junctions (Neutra
et al
., 2001).
Additionally, the tightness of tight junctions can be affected
by host mediators and soluble environmental factors (Bouhet
et al
., 2004). For example, in cases of immaturity and diseases
like viral gastroenteritis, bacterial infection and inflammatory
bowel disease, the well-integrated function of the intestinal
epithelium is affected, and antigens of every kind can enter. In
health, only penetrating pathogens (Niedergang and Kweon,
2005) or invading bacteria (Cossart and Sansonetti, 2004)
find a way into the epithelial cell.
DC are located throughout the intestinal immune system
and as sentinel cells they perform important immuno-
surveillance functions (Stockwin
et al
., 2000). They are
found in the villous lamina propria, in the sub-epithelial
dome and the interfollicular regions of the PP (Figure 1), as
well as in the mesenteric lymph nodes (MLN). Although it is
feasible that DC residing in the MLN take up free antigens
or pathogens derived from afferent lymph, antigen uptake
is primarily performed by the immature peripheral DC
located in the villous lamina propria and the sub-epithelial
dome of PP. Interestingly, even non-replicating particulate
antigens can generate specific immune responses, although
they cannot normally cross the intestinal barrier. One
important mechanism responsible for this may be antigen
uptake by lamina propria DC that extend cytoplasmic pro-
cesses into the gut lumen between the enterocytes (Niess
and Reinecker, 2005). These cytoplasmic processes also
exist in the pig, but they are relatively rare (Bimczok
et al
.,
2006). After antigen uptake, the cells alter their chemokine
receptor pattern, which allows them to migrate to T-cell
regions of organised lymphoid tissue (i.e. interfollicular
regions of the PP or MLN), where antigen presentation takes
place (Fleeton
et al
., 2004). Cannulation studies of pseudo-
afferent intestinal lymph in rats and in pigs have shown
that DC are constantly migrating from the gut wall to the
MLN, even in the absence of overt antigenic stimulation
(Macpherson and Uhr, 2004). This would suggest that these
cells play an important role in the maintenance of tolerance.
Indeed, a significant population of these cells contained par-
ticles of apoptotic cells (Huang
et al
., 2000). Phenotypic
analyses of migrating DC in porcine pseudo-afferent intestinal
lymph suggest that migration routes are mainly from the
villous lamina propria to MLN, and from the sub-epithelial
domes to the interfollicular regions of the PP (Bimczok
et al
.,
2005).
The postnatal period is critical in terms of the development
of an effective mucosal immune system that is capable of the
adaptation to commensal bacteria, tolerance towards nutri-
tional components and defence against pathogens in the
gut lumen (Pie
´
et al
., 2004 and 2007; Stokes
et al
., 2004;
Rothko
¨tter
et al
., 2005). Weaning is a period during which
there is considerable change in the magnitude and diversity of
exposure to environmental antigens derived from food and
potentially pathogenic organisms. Under ‘natural conditions’
weaning is a gradual process, and in piglets it is not complete
until 10 to 12 weeks of age. This is however not normal
husbandry practice and piglets are more commonly weaned
abruptly at 3 to 5 weeks. As a consequence, neonatal pigs
experience a transient immune hypersensitivity to dietary
antigens when weaned at 3 weeks of age, often associated
with clinical symptoms, e.g. diarrhoea, weight loss and
enteritis (Bailey
et al
., 1993; Stokes
et al
., 2004). In contrast, if
piglets are allowed to suckle their dams, until they voluntarily
wean themselves at approximately 12 weeks, they develop
oral tolerance to the dietary antigens without clinical symp-
toms (Bailey
et al
., 1994). The initial and characteristic lesions
of post-weaning diarrhoea are a crypt hyperplasia and a
severe villous atrophy in the small intestine, which is asso-
ciated with a fall in brush border enzymes (disaccharidases,
sucrase and lactase) and malabsorption. These changes occur
within 3 to 4 days of weaning and a partial recovery occurs
after 7 to 10 days. They are partly related to a transient
hypersensitivity reaction to antigens in the post-weaning diet
as comparable effects are observed in the complete absence
of bacteria. However, the weaning effects are most commonly
associated with an increase in the proliferation of gut bacteria
and the hypersecretory effects of bacterial enterotoxins (Sahin
et al
., 1991).
Natural alternatives to in-feed antibiotics
In the following, the capacity of selected plant extracts and
other natural substances to improve the immune status of
the pig is reviewed. For each type or class of substances,
effort has been made to synthesise in tables major elements
concerning experimental designs, and immune parameters
which have been studied by differentiating the local
intestinal immune response and the systemic immunity.
Indeed, in most of the experiments, the peripheral blood
immune cells are more easily studied than mucosal immune
cells. However, only two percent of all lymphocytes are in
the peripheral blood (Blum and Pabst, 2007) and their half
life in the blood is about 30 min. Thus, the observed reac-
tions reflect the responses of cells that may have their
origin in the PP, in the MLN or in the tonsils – all com-
partments of the mucosal immune system. Alternatively, the
peripheral blood lymphocytes may be spleen-derived or
come from peripheral lymph nodes. Thus, immune reactions
after an in-feed application of natural immunomodulators
cannot directly be linked to a defined site of origin. So far, it
is often difficult to explain whether their reaction pattern
depends on mucosal or systemic immune responses.
In-feed modulators of piglet immunity
1647
Yeast derivates
Interest in the impact of polysaccharides on immunity has
increased rapidly over the past decade and this is reflected in
the large number of reviews dealing with this topic (Bohn and
BeMiller, 1995; Williams, 1997; Tzianabos, 2000; Brown and
Gordon, 2003). A variety of polysaccharides from different
natural sources have the ability to modulate the immune
system. Among those, b-D-glucans and the carbohydrate
portion of mannoproteins, the a-D-mannans, have been
recognised to be responsible for modulating the immune
responses in mammals through specific interactions with dif-
ferent immunocompetent cells (Kogan and Kocher, 2007). A
direct interaction of b-D-glucans with macrophages and poly-
morphonuclear cells confers to these products their immuno-
modulatory properties (Tzianabos, 2000; Brown and Gordon,
2003). Mannans alter the immune response by specific links
with mannose receptors, present on many cells of the immune
system (Lee, 1988), such as macrophages (Tzianabos, 2000).
Glucans and mannans are already proposed as potential
immunomodulatory agents for prophylaxis and therapy of
infections in human (Masihi, 2000) and in farm animals
including pigs (Sohn
et al
., 2000; Kogan and Kocher, 2007).
Numerous commercial preparations derived from yeast cell
walls, particularly rich in glucans and mannans, are avail-
able as in-feed supplements for pigs (Table 3). However, the
composition and the purity of these products often remain
unknown. This could partly explain the high variation of
incorporation levels (from a factor 1 to 500 for glucans, and
a factor 1 to 30 for mannans) of these commercial pre-
parations in feed (Table 2). Moreover, the literature dealing
with diet incorporation of yeast cell wall extracts often
refers to only one of those two compounds, but yeast
derivates usually contain both polysaccharides.
Glucans
.b-glucans extracted from the cell wall of
Sac-
charomyces cerevisiae
is the most common source used for
animal in-feed complements. It is a b-1,3-glucan with long
b-1,6-glucan branches, whose structure is different from
the b-glucans extracted from bacteria (linear b-1,3-glucan),
cereals (b-1,3/1,4-glucan) and fungi (short b-1,6-glucan
branched b-1,3-glucan). Their different chemical structures
would be expected to be reflected in their different bio-
activities (Bohn and BeMiller, 1995). Furthermore, depending
on the extraction procedure, soluble and insoluble fractions
that have different activities can be present together or
separately (Tzianabos, 2000). Thus, the form under which
glucans are included in diet will probably influence their
biological properties.
Literature dealing with effects of glucans on local
intestinal immune response in pigs is very scarce (Table 2).
In finishing pigs, the ileal contents of IgM, IgA, or CD4
1
and CD8
1
T-cells were not modified by glucans supple-
mentation (Sauerwein
et al
., 2007). A higher incorporation
dose of b-glucans (2.5%) led to increased intestinal tumour
necrosis factor-a(TNF) and IL-1bmRNA but also in
intestinal IL1-receptor antagonist (IL-1Ra) mRNA in pigs
challenged 4 h earlier with lipopolysaccharide (LPS) (Eicher
et al
., 2006). These changes were not associated with
alterations in blood TNF-awithin hours following the
inflammatory challenge. Biological significance of these
results is difficult to assess as those cytokines are impli-
cated in both pro-inflammatory (TNF-aand IL-1b) and anti-
inflammatory (IL-1Ra) processes.
Much interest has been paid to the effects of glucans on
systemic immune responses, particularly innate immunity.
b-glucans have been shown to have anti-inflammatory prop-
erties.
In vitro
,IL-6andTNF-aproduction by lymphocytes
isolated from peripheral blood of weaned piglets supple-
mented with b-glucan and stimulated with LPS was
decreased, whereas IL-10 production was enhanced (Li
et al
.,
2005). Those cytokine patterns were confirmed
in vivo
in
piglets fed diets supplemented with glucans, and challenged
intraperitoneally with LPS (Li
et al
., 2005 and 2006). Taken
together, these
in vitro
and
in vivo
analyses show that dietary
b-glucans prevent the elevation in pro-inflammatory cytokines
whilst enhancing the production of anti-inflammatory cyto-
kines in response to an inflammatory challenge. Consistently,
glucans have been shown to modulate the acute phase
response, whose regulation is known to be orchestrated by
pro-inflammatory cytokines like IL-1, IL-6 or TNF (Baumann
and Gauldie, 1994; Niewold, 2007). In piglets, blood hap-
toglobin concentrations increase during the 2 weeks following
an early weaning (18 days), then remain stable (Dritz
et al
.,
1995). A supplementation with 0.025% or 0.05% of b-glucans
partly suppresses this increase (Dritz
et al
., 1995). These
results were not confirmed in piglets weaned at 28 days (Hiss
and Sauerwein, 2003; Sauerwein
et al
., 2007). However, it
should be noted that in these later experiments, haptoglobin
concentration increased only during 1 week post-weaning,
which could have limited the impact of glucans on this
response.
As a known target for glucans, the function of neutrophils
and macrophages has been widely investigated (Brown and
Gordon, 2003). Dietary b-glucans had no consistent effects
on the ability of peripheral blood neutrophils to generate
reactive oxygen intermediates, neutrophil-mediated antibody-
dependent cellular cytotoxicity and on neutrophil phagocytic
activity (Dritz
et al
., 1995; Sauerwein
et al
., 2007). Similarly,
lung macrophage production of superoxide anion and bac-
tericidal activity, as well as the expression of CD14, were not
influenced by the inclusion of b-glucans in the diet (Dritz
et al
., 1995). Thus, the predicted modulation of neutrophil
and macrophage functions by glucans was not confirmed in
in vivo
experiments, which may partly be explained by the
limiting ability of glucans to be delivered to those cells after
an oral supplementation. Whereas the ability of lymphocytes
to proliferate is also insensitive to a feed supplementation
with glucans (Hiss and Sauerwein, 2003), the production of
the different classes of Igs is influencedinadose-dependent
manner with lower and higher doses favouring IgA and IgG
responses, respectively (Sauerwein
et al
., 2007). This could be
partly correlated to the different pattern of lymphocyte sub-
classes (helper and cytotoxic T-cells) observed in response
to a supplementation with b-glucans (Hahn
et al
., 2006).
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1648
Table 2
Immune parameters measured in
in vivo
experiments where pigs are dietary supplied with b-glucans
Glucans
Feed content
(%) Weaning age
Time
supplementation PW Challenge or vaccination
Intestinal immune
response Systemic immune response Reference
A 0.05 25–26 d 4 wk None Not measured
Blood:
Ig titre Decuypere
et al
. (1998)
0.4 21 d 4 wk None Not measured
Blood:
leukocyte subset Kim
et al
. (2000)
0.1 21 d 4 wk
a
None Not measured
Blood:
neutrophil function Dritz
et al
. (1995)
0.1 14 d 4 wk None Not measured None Dritz
et al
. (1995)
0.025–0.05 18 d 6 wk
Streptococcus suis
(6.5 310
8
colony forming unit
(CFU) i.v.) 4 wk PW
Not measured
Blood:
neutrophil function,
haptoglobin
Lung
: macrophage function
Dritz
et al
. (1995)
B 0.015–0.03 28 d 4 wk PRRS virus, vaccination 1 d PW
c
Not measured
Blood:
Ig titre, LC proliferation,
haptoglobin
Hiss and Sauerwein (2003)
0.03 27 d 4 wk None Not measured
Blood:
neutrophil phagocytic
activity, serum IgG and IgA,
haptoglobin
Sauerwein
et al
. (2007)
0.3 30 d 4 wk None Not measured
Blood:
neutrophil phagocytic
activity, serum IgG and IgA,
haptoglobin
Sauerwein
et al
. (2007)
0.03 Fattening pigs 2 wk before slaughter None
Ileum
: IgM, IgA,
CD4
1
, CD8
1
, T-cells
Not measured Sauerwein
et al
. (2007)
C 0.02–0.04 6.1 kg (? d) 8 wk Atrophic rhinitis vaccination,
i.m. 9 d PW
d
Not measured
Blood:
Ig titre, lymphocyte
subset
Hahn
et al
. (2006)
D 2.5 14 d 2 wk
b
LPS (150 mg/kg BW i.v.)
14 d PW
Ileum
: mRNA for
TNF-a, IL-1b, IL-1Ra
Blood:
leukocyte count, TNF-a
Liver, lung, spleen
: mRNA for
TNF-a, IL-1b, IL-1Ra
Eicher
et al
. (2006)
E 0.005 28 d 5 wk None Not measured
Blood:
LC proliferation Li
et al
. (2006)
0.005 28 d 31 d LPS (25 mg/kg BW i.p.) 31 d PW Not measured
Blood:
TNF-a, IL-6, IL-10 Li
et al
. (2006)
0.005 28 d 31 d Ovalbumin (5 mg/kg BW)
14 d PW; LPS (25 mg/kg BW
i.p.) 31 d PW
Not measured
Blood:
anti-ovalbumin Ig,
TNF-a, IL-6, IL-10
Li
et al
. (2005)
PW 5post-weaning; d 5day; wk 5week; PRRS 5porcine reproductive and respiratory syndrome; Ig 5immunoglobulin; LC5lymphocyte; LPS 5lipopolysaccharide; TNF-a5tumour necrosis factor-a;IL5interleukin;
IL-1Ra 5interleukin1-receptor antagonist.
A: Macrogard-S
TM
(Biotec-Mackzymal A/S, Tromso, Norway; or Provesta Corp., Bartlesville, OK, USA; or Jeil Vet. Chem. Co., Seoul, Korea); B: b1,3/1,6-glucans (Antaferm MG, Dr Eckel GmbH, Niederzissen, Germany) –
insoluble fraction containing 25% of b-D-glucans, and 10% of mannans; C: Glucagen (Enbiotec Company, Seoul, Korea); D: Energy Plus (Natural Chem Industries, Ltd, Houston, TX, USA); E: From baker’s yeast, manual
manufacturing – should contain 86% of b-glucans.
a
Starting 1 wk post-weaning;
b
Treatment also administered in suckling piglets by gavage;
c
PRRSV (Ingelvac
R
PRRS MLV, Boehringer, Ingelheim, Germany);
d
Pfizer Co., Seoul, Korea.
In-feed modulators of piglet immunity
1649
However, those variations are inconstantly reported, and
appear to be related to the period of supplementation (Kim
et al
., 2000; Hahn
et al
., 2006).
The effect of dietary supplementation with b-glucans on
the response to systemic immunisation has produced con-
trasting results. For example, whereas b-glucan-supplemented
piglets vaccinated with atrophic rhinitis vaccine produced a
transiently lower antibody response (Hahn
et al
., 2006), pigs
injected with ovalbumin and receiving b-glucans at a dose
of 0.005% mounted a transiently higher antibody response
(Li
et al
., 2005). b-glucans did not enhance the efficiency of
a vaccination with porcine reproductive and respiratory
syndrome (PRRS) virus (Hiss and Sauerwein, 2003), or different
EnteroToxinogenic
Escherichia coli
(ETEC) antigens (Decuypere
et al
., 1998).
The effect of b-glucans on the passive transfer of
immunity has also been studied by immunisation of sows
with strains of
E. coli
implicated in neonatal diarrhoea
(Decuypere
et al
., 1998). Whereas the titre of antibodies
specific to K88ab and K99 was not enhanced in colostrum
of glucan-fed sows, it was elevated in their milk as com-
pared to control sows. The authors noted that the health
score (mainly due to neonatal diarrhoea) of piglets whose
mother was submitted to a b-glucan regimen was severely
impaired during 14 days
post partum
, but that the aetio-
logical agent was not one of the ETEC strains against which
sows were immunised.
In conclusion, the effects of glucans on immunity are not
predictable and their ability to act as growth-promoters is also
not reliable (Table 2). It should be highlighted that many of
the studies have been performed in ‘clean’ environments,
where morbidity rates are low, and that beneficial effects on
health are much more difficult to detect in such environments.
Mannans
. The potential protective activities of mannans may
include their ability to ‘adsorb’ enteric pathogens and to
modulate immune functions (Sohn
et al
., 2000). To our
knowledge, only two publications have investigated the effects
of dietary mannan supplementation on local intestinal immune
response in swine (Table 3). Even if mannans do not have any
effect on the number of macrophages in intestinal lamina
propria, their function seems to be enhanced. Indeed, the
phagocytosis of sheep red blood cells by macrophages
isolated from jejunal lamina propria was increased by
inclusion of 0.3% mannans in the basal diet (Davis
et al
.,
2004a). The recruitment of lymphocytes into the small
intestinal lamina propria is reduced in piglets fed mannans
(Lizardo
et al
., 2008) and their profile is also influenced.
Indeed, the ratio of CD3
1
CD4
1
/CD3
1
CD8
1
T-cells was
lowered 21 days post-weaning, indicating a greater pro-
portion of CD8
1
T-cells in mannan-supplemented pigs
(Davis
et al
., 2004a). After birth, infiltration of lamina
propria with CD8
1
T-cells is usually delayed (4 weeks after
birth) as compared to infiltration with CD4
1
cells, which
can begin as early as 2 weeks of age (Bailey
et al
., 2001).
This observation could be an indication that mannan sup-
plementation would enhance the establishment of a mature
Table 3
Immune parameters measured in
in vivo
experiments where pigs are dietary supplied with mannans
Mannans
Feed content
(%) Weaning age
Time supplementation
PW Challenge or vaccination Intestinal immune response Systemic immune response Reference
A 0.2 18 d 5 wk None Not measured
Blood:
lymphocyte proliferation Davis
et al
. (2002)
0.1 21 d 4 wk None Not measured
Blood:
leukocyte subset Kim
et al
. (2000)
0.3 19 d 4 wk None
Jejunum
: IEL, leukocyte subset
and macrophage phagocytosis
in
lamina propria
Blood:
leukocyte subset, a-1-
acylglycoprotein, lymphocyte proliferation,
monocyte phagocytosis
Davis
et al
. (2004a)
0.3 then 0.2 19 d 10 d then 28d None Not measured
Blood:
lymphocyte proliferation Davis
et al
. (2004b)
0.2–0.3 19 d 38 d None Not measured
Blood:
lymphocyte proliferation Davis
et al
. (2004b)
0.3 19 d 35 d None Not measured
Blood:
lymphocyte proliferation Davis
et al
. (2004b)
B 3 22 d 4 wk None Not measured
Blood:
IgA, IgG, IgM White
et al
. (2002)
3 11 d 39d
Escherichia coli
K88 (9.5 310
8
CFU
p.o.) 29 d P W
Not measured
Blood:
IgA, IgG, IgM White
et al
. (2002)
C 0.15 6.8 kg (? d) 4 wk
Salmonella enterica
(1.3 310
9
CFU
p.o.) 14 d P W
Not measured
Blood:
haptoglobin, IL-6 Burkey
et al
. (2004)
0.1 28 d 5 wk None
Jejunum–ileum
: IEL
Blood:
leukocyte subset Lizardo
et al
. (2008)
PW 5post-weaning; d 5day; wk 5week; IEL 5intraepithelial lymphocytes; Ig 5immunoglobulin; CFU5colony forming unit; IL 5interleukin.
A: Bio-Mos
R
(Alltech Inc., Nicholasville, KY, USA); B: From brewer’s yeast (Brewtech, International Ingredient Corp., St Louis, MO, USA) – mannan content was found to be 5.2%; C: SAF-Mannan
R
(Lesaffre Feed
Additives, Marquette-Lez-Lille, France).
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1650
T-cell repertoire within the gastro-intestinal tract. However,
further work is still needed to confirm this hypothesis.
Systemic immune response to dietary mannans has been
studied in more detail. For example under normal breeding
conditions, serum a-1-acid glycoprotein concentration was not
altered by inclusion of dietary mannans (Davis
et al
., 2004a),
whereas piglets fed with 0.3% of phosphorylated mannans
exhibited a decreased blood neutrophils : lymphocytes ratio,
suggesting that mannan supplementation could alleviate the
inflammatory response associated with the stress of weaning
(Davis
et al
., 2004a). The increased blood lymphocyte popu-
lation among leucocytes in mannan-fed animals could be
linked more to B than to T-cells. Indeed, 3% of brewer’s yeast
(which corresponds to a final level of 0.16% of mannan oli-
gosaccharide) only tended to increase the piglet serum level of
IgG when used alone, but substantially increased this level
when fed with the acidifier, citric acid (White
et al
., 2002).
Conversely, blood proportions of CD4
1
or CD8
1
lymphocytes
are insensitive to mannan supplementation (Kim
et al
., 2000).
The proliferative response of peripheral blood lymphocytes
was generally found not to be affected by the inclusion of
mannans in the diet of nursery piglets (Davis
et al
., 2002,
2004a and 2004b), but when continuously incorporated at a
dose rate of 0.3% the ability of lymphocytes to proliferate
after a stimulation with a pokeweed mitogen (non-specific
proliferation of B- and T-cells) or with phytohemaglutinin
(proliferation of primarily T-cells) was reduced (Davis
et al
.,
2004b). The authors suggested that this altered immune
function may explain the beneficial effects of mannans for
piglet growth. However, there are no data concerning mor-
bidity or mortality to assess the health effects of such an
immune depression.
In piglets challenged with
Salmonella enterica
serotype
Typhimurium, serum haptoglobin concentrations were
increased in 0.15% mannan-fed piglets 6 and 13 days post-
infection as compared to piglets fed the basal diet either
alone or enriched with AB (Burkey
et al
., 2004). Serum IL-6
was not altered by mannan supplementation, but this could
be explained by the absence of a response on this cytokine
following challenge with this pathogen (Burkey
et al
.,
2004). Contrary to carbadox, mannans failed to reduce the
length of the period of hyperthermia observed after infec-
tion with
S. enterica
and did not promote growth (Burkey
et al
., 2004). Total levels of serum IgA, IgM and IgG were
not modified by dietary mannan, in piglets challenged or
not with an enterotoxigenic strain of
E. coli
K88 (White
et al
., 2002).
As for glucans, the influence of mannans on immunity is
not always reliable, as well as their effects on piglet per-
formances. In piglets challenged with enteric pathogens
(
E. coli
K88,
S. enterica
), health benefits of dietary mannans
are not consistent.
Plant extracts
Empirical evidence suggests that plant extracts may offer
benefits in boosting the immune system; thus, preventing
disease in production animals (Wenk, 2003) and plant-
derived products have gained increasing interest as possible
feed additives for non-ruminant species (Windisch
et al
.,
2008). Plants, and where they have been identified their
bioactive components, are very diverse and their potential
to enhance pig health and immunity has only been scarcely
evaluated
in vivo
(see Table 4). Moreover, most studies
have used a mixture of compounds, which does not allow
the investigation of the immune properties of a specific
bioactive component.
Herbaceous plants
. Herbs and spice extracts have been
used extensively in different parts of the World to treat a
variety of human diseases (Stein and Kil, 2006). Gastro-
intestinal disorders have been treated with a number of
different plant extracts including garlic, peppermint, cha-
momile or aloe (Amagase
et al
., 2001; Akerreta
et al
., 2007;
Rodriguez-Fragoso
et al
., 2008). Some herbs like
Echinacea
,
liquorice, cat’s claw and garlic are claimed as immuno-
enhancers (Craig, 1999). The essential oil of the plant is
often the biologically active component, but other extracts
can also display biological activities.
The
Labiatae
constitutes a large family of herbs: basil, dill,
fennel, marjoram, mint, rosemary, oregano, sage and thyme
(Craig, 1999). Mixtures of essential oils based on thymol and
carvacrol whose major sources are thyme and oregano,
respectively (Burt, 2004), seem promising due to their anti-
microbial (Kim
et al
., 1995; Cosentino
et al
., 1999; Baydar
et al
., 2004) and potential immunomodulatory properties
(Woollard
et al
., 2007). A plant extract containing 6% of
carvacrol and 0.14% of thymol, incorporated at 0.05% to
0.15% in a pig diet, had no effect on the plasma levels of the
acute phase proteins, haptoglobin and C-reactive protein
(Muhl and Liebert, 2007) and the inclusion of a commercial
plant product composed of oregano oil mixed with anis and
citrus oils did not improve the health status of piglets
(Kommera
et al
., 2006). In contrast, an extract of
Origanum
vulgare
, enriched with both thymol and carvacrol in similar
proportions, was reported to protect low-weight growing-
finishing pigs from disease (Walter and Bilkei, 2004). This
health benefit was associated with an increased proportion
of CD4
1
,CD8
1
and double-positive T-cells in peripheral
blood and MLN (Walter and Bilkei, 2004). Thymol used alone
enhances total IgA and IgM serum levels and exhibits some
local anti-inflammatory properties, as indicated by a reduction
in TNF-amRNA in the stomach (Trevisi
et al
., 2007).
In vitro
,
cinnamaldehyde, the main component of cinnamon essence,
also has anti-microbial (Burt, 2004) and immunomodulatory
(Koh
et al
., 1998) properties. A plant extract containing 5% of
carvacrol (
Origanum
spp.), 3% of cinnamaldehyde (
Cinna-
monum
spp.) and 2% of capsicum oleoresin (
Capsicum
annum
), included in the feed at a 0.03% level, led to a
decreased number of jejunal IEL, and an increased number of
lymphocytes in the colonic lamina propria (Manzanilla
et al
.,
2006). Conversely, mononuclear cell subsets from ileal PP
were not affected by this plant extract combination (Nofrarias
et al
., 2006) and only the percentage of B lymphocytes was
reduced in lymph nodes of piglets (Nofrarias
et al
., 2006).
In-feed modulators of piglet immunity
1651
Table 4
Immune parameters measured in
in vivo
experiments where pigs are dietary supplied with plant extracts
Source
Suspected bioactive
component(s) Feed content (%) Weaning age
Time
supplementation
PW Challenge or vaccination
Intestinal immune
response Systemic immune response Reference
Herbaceous plants
Bahzen Chinese herbal
medicine
a
Flavonoids and
polyphenols
1 28 d 8 wk LPS (100 mg/kg BW i.m.),
ovalbumin (1 mg/pig
i.m.), 5 wk PW
Not measured
Blood
: IL-6, TNF-a, leukocyte
count, neutrophil function,
IgG, anti-ovalbumin Ig
Lien
et al
. (2007)
Unknown Thymol 1 24 d 25 d
S. enterica
ser. Typhi. (1.5 3
10
9
CFU p.o.) 5 d PW
Stomach
: mRNA TNF-a
Blood
: IgM, IgA
Saliva
: IgA
Trevisi
et al
. (2007)
Phytogenic additive
b
Carvacrol, thymol 0.05–0.1–0.15 ? 5 wk None Not measured
Blood
: C-reactive protein,
haptoglobin
Muhl and Liebert
(2007)
Origanum
spp. Carvacrol,
cinnamaldehyde,
capsicum oleoresin
0.03 18–22 d 3 wk None
Jejunum
,
ileum
,
colon
:
IEL,
lamina propria
lymphocytes
Not measured Manzanilla
et al
. (2006)
0.03 18–22 d 3 wk None
Ileocolic lymph nodes
,
ileal PP
: mononuclear cell
phenotype
Blood
: leukocyte subset Nofrarias
et al
. (2006)
Cinnamonum
spp.
Capsicum annum
c
Oregano feed additive
d
Carvacrol 0.3 60 kg Until slaughter None Not measured
Blood
,
MLN
: leukocyte subset Walter and Bilkei
(2004)
Thymol (? d)
Astragalus
membranaceus
e
b-glucans 0.05–0.1 28 d 4 wk LPS (200 mg/kg BW i.m.)
7 and 21 d PW
Not measured
Blood
: lymphocyte
proliferation, IL-2
Mao
et al
. (2005)
Astragalus
membranaceus
f
b-glucans 0.01–0.1 26–30 d 3 wk Ovalbumin (1 mg/kg BW)
14 d PW
Not measured
Blood
: leukocyte count,
lymphocyte subset and
proliferation, anti-ovalbumin
Ig,IL-2,IL-4,IL-10,IFN-g,IgG
Yuan
et al
. (2006)
Echinacea purpurea
g
Chicory acid, alkamids 1.8 (cobs) 5.8 kg 6 wk None Not measured
Blood
: cell count, lymphocyte
proliferation
Maass
et al
. (2005)
1.5 (cobs) 4–6 ml/d
(juice)
Finishing pigs
(32 kg)
9wk
p
Erysipelothrix
rhusiopathiae
wks
1 and 5
r
Not measured
Blood
: cell count, specific Ig Maass
et al
. (2005)
Sugar cane extract
Saccharum
officinarum
spp.
h
? 0.5–1–1.5–2 g/kg BW 35 d 4 wk
q
None Not measured
Blood
: PRRS Ig, leukocyte
count, NK cell cytotoxicity,
monocyte and neutrophil
phagocytosis
Lo
et al
. (2005)
Soy daidzein
i
Daidzein 0.02–0.04–0.08 11d 6 wk PRRS virus (2 310
4.3
virus oronasally) 17 d PW
Not measured
Blood
: PRRS Ig and virus,
IFN-g,AGP
Greiner
et al
. (2001b)
Soy genistein
j
Genistein 0.02–0.04–0.08 10 d 5 wk PRRS virus (2 310
4.3
virus oronasally) 9 d PW
Not measured
Blood
: PRRS Ab and virus,
IFN-g,AGP
Greiner
et al
. (2001a)
Ligneous plants
Quillaja saponaria
k
Saponin fraction 0.0125–0.025–0.05 24 d 4 wk
Salmonella enterica
ser.
Typhi. (10.5 310
9
CFU
p.o.) 14 d PW
Not measured
Blood
: haptoglobin, AGP, IgG,
IgM, phagocytic function
Turner
et al
. (2002b)
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1652
Chinese pharmacopoeia describes the use of numerous
herbal formulations to cure a wide variety of diseases. Among
those, Bahzen is a medicine composed of eight different herbs
(
Atractylodes ovata
,
Codonopsis pilosula
,
Poria cocos
,
Gly-
cyrrihiza uralensis
,
Angelica sinensis
,
Ligusticum chuanxiong
,
Paeonia albiflora
and
Rehmannia glutinosa
) whose effects
have been tested
in vivo
in pigs (Lien
et al
., 2007). Bahzen
increased white blood cell counts, without leukocytosis, and
enhanced IL-6 and TNF-ain the serum of pigs challenged
with LPS, compared to negative and positive (AB) control
animals (Lien
et al
., 2007). Serum IgG levels and blood
neutrophil activity in Bahzen and AB groups were increased.
However, Bahzen did not modulate acquired immunity, as
measured by specific antibody responses directed towards
different antigens (sheep red blood cells, ovalbumin) (Lien
et al
., 2007). Piglets fed with Bahzen also grew faster than
control piglets (Lien
et al
., 2007). The authors speculated that
Bahzen-fed piglets had an increased ability to respond to an
inflammatory stress, and that this may be beneficial for piglet
health. Equally, it is possible to argue that an increased
inflammatory response could be detrimental to piglet health.
An abundance of plants of the
Echinacea
family in
pasture is used as an indicator of ‘good health’. The main
bioactive components of
Echinacea purpurea
are chicory
acid and alkamids. When included as juice or cobs in the
post-weaning diet or that of finishing pigs, growth perfor-
mances are not improved, but feed efficiency tends to be
increased (Maass
et al
., 2005). Blood parameters, including
cell count and lymphocyte proliferation, were not modified
by dietary treatment but the health status of piglets was
good throughout the trial. Moreover, the response to
immunisation of piglets with a vaccine against
Erysipelo-
thrix rhusiopathiae
was enhanced by the inclusion of
E.
purpurea
into the diet of finishing pigs (Maass
et al
., 2005).
This could be an indication for the health-promoting prop-
erties of
Echinacea
, but further studies would be required to
confirm this.
Whilst the bulk of b-glucans used in feed industry is
derived from yeast cell wall (see previous section), b-glucans
from the Chinese herb
Astragalus membranaceus
have also
been tested (Mao
et al
., 2005; Yuan
et al
., 2006). Dietary
A.
membranaceus
increased the white blood cell count, mainly
through the contribution of CD4
1
lymphocytes (Yuan
et al
.,
2006). The proliferation of T-cells isolated from peripheral
blood in weanling pigs was also increased in a dose-depen-
dent manner in b-glucans-fed piglets (Mao
et al
., 2005).
Concomitantly, b-glucans from
Astragalus
increased blood
concentration in IL-2 (Mao
et al
., 2005; Yuan
et al
., 2006) and
interferon-g(IFN-g)(Yuan
et al
., 2006), but IL-4 and IL-10
concentrations remained unchanged (Yuan
et al
., 2006). This
cytokine profile suggests a Th1 bias, which is in accordance
with the enhancement of cellular immunity conferred by glu-
cans. Plant b-glucans do not influence humoral immunity, as
their inclusion in diet did not alter specific antibody titres
following immunisation with ovalbumin (Yuan
et al
., 2006).
A 0.05% level of glucans extracted from
A. membranaceus
counteracted the increased plasma concentrations of
Table 4 Continued
Source
Suspected bioactive
component(s) Feed content (%) Weaning age
Time
supplementation
PW Challenge or vaccination
Intestinal immune
response Systemic immune response Reference
Quillaja saponaria
l
Saponin Curcumin 0.03 29 d 3 wk None Not measured
Blood
: IgG, IgA, AGP, IFN-gIlsley
et al
. (2005)
Curcuma longa
m
0.02
Seaweed extract
Ascophyllum nodosum
n
Fucan polysaccharide? 0.5–1–2 24 d 4 wk
S. enterica
ser. Typhi.
(6 310
9
CFU p.o.)
14 d PW
Not measured
Blood
: haptoglobin, AGP,
IgG, IgM
Spleen
: IL-10
Turner
et al
. (2002a)
PW 5post-weaning; d 5day; wk 5week; IL 5interleukin; TNF-a5tumour necrosis factor-a;Ig5immunoglobulin; CFU 5colony forming unit; IEL 5intraepithelial lymphocytes; PP 5Peyer’s patches; MLN 5mesenteric
lymph nodes; LPS 5lipopolysaccharide; PRRS 5porcine reproductive and respiratory syndrome; NK 5natural killer; IFN-g5interferon-g;Ab5antibody; AGP 5alpha-1-acid glycoprotein.
a
Equal amount of powder extracted from eight herbal medicines purchased from a drug market in Taiwan;
b
Commercial phytogenic additive containing 53% inulin, 8% essential oil mix, 3% tannins – analysis revealed a
6% carvacrol and 0.14% thymol content of this additive;
c
Plant extract combination (Pancosma, S.A., Geneva, Switzerland) containing 5% carvacrol (
Origanum
spp.), 3% cinnamaldehyde (
Cinnamonum
spp.) and 2%
capsicum oleoresin (
Capsicum annum
);
d
Oregpig
R
, Oregpig GmbH, Pecs, Hungary;
e
HongSheng Herbal Drug Store, Beijing, China, manual manufacturing;
f
Beijing, People’s Republic of China, manual manufacturing;
g
Echinacea
pressed juice from aerial part of the plant, commercial product;
h
Shin Mitsui Sugar Co., Ltd, Japan;
i
CSI Metabolics, Newport Beach, CA, USA;
j
Wiley Organics, Coshocton, OH, USA;
k
Desert King International,
Chula Vista, CA, USA;
l
Acros Organics, Geel, Belgium;
m
Axiss France SAS, Bellegarde-sur-Valserine, France;
n
No information;
o
Except the 1st wk: 0.075%;
p
Supplementation only during wk 1–3 and 7–9;
q
Three
consecutive d/wk;
r
Porcilis Ery ad us. Vet., Intervet, Unterschleißheim, Germany.
In-feed modulators of piglet immunity
1653
IL-1band prostaglandin E2 induced by an LPS challenge.
Surprisingly, this effect was less pronounced when included
at a dose of 0.1% in the diet (Mao
et al
., 2005). Thus,
b-glucans from
A. membranaceus
exhibited anti-inflamma-
tory properties (decrease in IL-1bsynthesis) and promoted
T-lymphocyteproliferation(whichmaybepartlylinkedtothe
enhanced serum IL-2), but had no positive effects on humoral
immunity. These immune modulations may be beneficial for
the piglets to fight against infections, but the health status of
piglets was not stated in these studies.
Sugar cane extracts display a wide range of biological
effects, including immunostimulating and growth-promot-
ing properties as shown in chicken (El-Abasy
et al
., 2002).
In pigs, sugar cane extract prepared by chromatographic
separation on an ion exchange column could enhance
innate immunity, through increased NK (natural killer) cell
cytotoxicity and phagocytic function of monocytes and
neutrophils (Lo
et al
., 2005). As morbidity and mortality
rates remained very low in this experiment, it is not possible
to speculate whether those immune modulations induced
by sugar cane extract would improve the resistance of
piglets to infection. Moreover, the potential bioactive
component(s) from this particular extract of sugar cane
have not been identified.
Genistein and daidzein, two isoflavones found in soy-
bean products, have also been suggested to act as immuno-
modulators when given orally. After oronasal inoculation
of piglets with PRRS virus, dietary daidzein failed to
decrease serum titres of virus (Greiner
et al
., 2001b),
whereas genistein minimised the viremia from day 4 to day
24 post-inoculation, as well as the serum concentration of
IFN-g(Greiner
et al
., 2001a). Serum a-1-acid glycoprotein
concentration was not modulated by daidzein (Greiner
et al
., 2001b), but was increased during periods of high
viremia by genistein (Greiner
et al
., 2001a). This enhanced
a-1-acid glycoprotein response in genistein-fed piglets
supports the hypothesis of a greater and more effective
immune response, which could explain the lower viremia
(Greiner
et al
., 2001a). Accordingly, lower serum IFN-g
concentration in genistein-fed animals is in agreement with
the greater virus elimination and a quicker return of IFN-g
to basal levels (Greiner
et al
., 2001a). Despite the different
effects on viremia, both isoflavones were efficient in pro-
moting growth in piglets challenged with PRRS virus, which
suggests that their mechanisms of action would differ. It
would be of interest in future to test their ability to enhance
immune responses and health during a challenge with an
intestinal pathogen.
Ligneous plants
. An extract of the South American tree
Quillaja saponaria
is widely used as a vaccine adjuvant
(Kensil
et al
., 2004), and whose active ingredient appears to
be the saponin fraction (Milgate and Roberts, 1995). As
saponins are known to enhance immunity via the intestinal
route, their use in animal nutrition has been the subject of
investigation (Francis
et al
., 2002). Dietary treatment with
crude soap bark of
Q. saponaria
, ranging from 125 to
500 mg/kg, did not counteract the negative effects on feed
intake and growth induced transiently by a challenge with
Salmonella enterica
serovar Typhimurium (Turner
et al
.,
2002b).
Q. saponaria
also failed to modulate the rise in
serum haptoglobin, a-1-acid glycoprotein and IgM con-
centrations induced by this challenge 7 and 14 days post-
infection (Turner
et al
., 2002b). The phagocytic function of
peripheral white blood cells tended to be depressed in
challenged pigs fed with high doses of
Q. saponaria
, but
not with low doses (Turner
et al
., 2002b). It has been
suggested that these ‘weak immune modulations’ may be
due to the low purity of the extract used (Ilsley
et al
., 2005),
which could reach a maximal level of 10% of saponins.
Moreover, tannins, which could be as high as 15% in a
crude extract of
Q. saponaria
, might have induced a detri-
mental response, as they are known to have anti-nutritional
properties (Singh
et al
., 2003). Thus, Ilsley
et al
. (2005)
incorporated a saponin extract from
Q. saponaria
in the diet,
alone or in combination with curcumin, which has been
shown to modulate lymphocyte-mediated immune functions
in mice (Churchill
et al
., 2000). The combination of both
products did not result in adverse or synergistic effects (Ilsley
et al
., 2005). Whereas piglet immune responses were not
influenced by curcumin, the feed intake and serum IgA, IgG
and C-reactive protein concentrations were transiently
increased in saponin-fed piglets. The subsequent negative
impact of saponin on feed utilisation, assessed by a decreased
feed :weight gain ratio, could result from increased dietary
requirements to mount an immune response (Ilsley
et al
.,
2005). However, as the health status of piglets remained very
good throughout the experiment, it is difficult to predict
whether such a response may have been beneficial or detri-
mental to piglets during the time of an infection.
Seaweed extracts
. Seaweed extracts are also known to
have immunomodulatory properties (Yoshizawa
et al
.,
1993). To date, we are only aware of one study that pre-
sents data on influence of a seaweed extract on the
immune parameters in pig production. This extract was
derived from
Ascophyllum nodosum
, and different levels of
incorporation ranging from 0% to 2% were tested in piglets
orally challenged with 6 310
9
CFU (colony forming units)
of
Salmonella enterica
serovar Typhimurium (Turner
et al
.,
2002a). Increasing levels of
A. nodosum
tended to linearly
enhance the feed intake, but decreased the feed efficiency
during the 4 weeks following the weaning of piglets.
The challenge with
Salmonella
had only moderate effects
on piglets, and dietary
A. nodosum
had no influence on
immune responses irrespective of whether the piglets were
infected or not.
In vitro
, a dose of 10 mg/ml of
A. nodosum
(greatly exceeding the highest level of feed incorporation
level of 2%) was able to activate porcine alveolar macro-
phages to secrete prostaglandin E2. However, this dose did
not alter the secretion of IL-10 by splenocytes (Turner
et al
.,
2002a). Taken together, the
in vitro
and
in vivo
data suggest
that
A. nodosum
extract probably has only little direct effect
on gut immune system in pig.
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1654
Animal by-products
Spray-dried animal plasma
. The biological properties of
spray-dried plasma (SDP), a by-product of commercial
slaughtering facilities, have been investigated in pig nutri-
tion (Coffey and Cromwell, 2001). Spray-dried animal
plasmas are characterised by a rich protein content, whose
digestibility and amino acid composition are similar to those
of the proteins in sow’s milk (van Dijk
et al
., 2001), and
whose Igs content can represent 24% to 25% (Niewold
et
al
., 2007). Plasma from both bovine and porcine origins
seem efficient in improving performances of piglets, and the
IgG fraction would appear to be the main component
responsible for the growth-promoting properties for these
products (Pierce
et al
., 2005). The Igs may prevent viruses
and bacteria from interacting with the gut wall, resulting in
an improvement of gut function (Coffey and Cromwell,
2001). One major problem is the stability of Igs during the
manufacturing process of SDP. The most critical step would
be spray-drying, which can lead to a 30% loss of Igs activity
(Niewold
et al
., 2007). Further processing to pellets would
not lead to appreciable losses. It is important to remember
that the use of non-sterilised products of animal origin as
feed ingredients for animals poses a potential health risk,
through the spread of certain infectious diseases. Despite
spray-drying being shown to eliminate viable pseudorabies
and PRRS viruses (Polo
et al
., 2005), the use of SDP origi-
nating from slaughter animals that have been subjected to
veterinary inspection is required. Indeed, it should be kept
in mind that the use of non-sterilised products of animal
origin as feed ingredients for animals remains a health risk
practice, which is strictly regulated in Europe (Regulation
(EC) No. 1774/2002 currently under revision). Moreover, as
recently illustrated by the use of animal flour in animal feed
and the bovine spongiform encephalopathy crisis, the public
opinion is concerned by the inclusion of animal by-products
in livestock feed. The meat consumers’ confidence could, in
the future, represent a limiting factor to the use of SDP and
other animal by-products in animal nutrition.
The effects of SDP on local intestinal immune responses
have been widely studied in pigs (Table 5). Results are
concordant and reveal that SDP prevents the infiltration of
gut-associated lymphoid tissue by immune cells, including
macrophages and lymphocytes (Jiang
et al
., 2000; Nofrarias
et al
., 2006 and 2007). This decreased infiltration is likely
to be a reflection of a lower antigenic challenge (Nofrarias
et al
., 2007). Additionally, the jejunal expression of the pro-
inflammatory cytokines IL-8 and TNF-awas decreased in
6% SDP-fed piglets 15 days after a challenge with ETEC
K88, irrespective of their status for K88 receptor (Bosi
et al
.,
2004). This decreased inflammatory response was asso-
ciated with a lower histopathological score in SDP-fed
piglets, thus suggesting that SDP would be efficient in
helping piglets fight against infections.
Concerning systemic immune responses, a 6% or 10%
supplementation of the diet with porcine plasma did not
modulate the increase in blood white blood cell count that
normally occurs during the 2 weeks following weaning
(Jiang
et al
., 2000; Nofrarias
et al
., 2006 and 2007). SDP did
not influence serum IFN-gor TNF-aunder basal conditions
(Touchette
et al
., 2002), but when stimulated with an
intraperitoneal LPS injection, SDP-fed piglets showed exa-
cerbated increases in serum levels of IFN-gand TNF-a
associated with severe intestinal damage, suggesting that
SDP-fed piglets would be more susceptible to certain
immunological challenges (Touchette
et al
., 2002). Similarly,
in piglets challenged intravenously with LPS, the increase in
serum IL-6 and IL-1bwas exacerbated in SDP-fed piglets,
but there was no evidence of an effect on mRNA cytokine
levels in the liver, thymus or spleen (Frank
et al
., 2003).
Conversely, SDP supplementation led to a decrease in C-
reactive protein concentration but not in haptoglobin (Frank
et al
., 2003). In those models, the extremely high dose
of LPS (75–150 mg/kg of BW) as well as the route of
administration (intraperitoneally or intravenously) preclude
from concluding on the plasma effects on inflammatory
responses when piglets are exposed to pathogens via the
enteral route.
The growth-promoting properties of SDP that are usually
reported (Jiang
et al
., 2000; Bosi
et al
., 2004; Nofrarias
et al
., 2006 and 2007; Niewold
et al
., 2007) are more
commonly observed with SDP from porcine than bovine
origin (Hansen
et al
., 1993; van Dijk
et al
., 2001). Moreover,
their anti-microbial/immunomodulatory properties would be
more likely to be beneficial when piglet growth may be
compromised, such as in conventional on-farm nursery
setting as compared to ‘cleaner’ off-site nursery (Coffey and
Cromwell, 1995), or in pigs weaned early
v.
a late weaning
(Torrallardona
et al
., 2002). This could be related to the
health-promoting properties of SDP observed in challenged
piglets. Indeed, in piglets challenged via the oral route with
an ETEC strain, inclusion of porcine SDP at an 8% level in
diet can reduce post-weaning diarrhoea and increase
growth of piglets that are positive for F4 receptor (Niewold
et al
., 2007). This effect was enhanced in piglets fed a diet
containing an SDP obtained from pigs previously immunised
with a vaccine against neonatal
E. coli
, where a con-
comitant decreased ETEC excretion was observed. These
results confirm that protection conferred by SDP would
be mainly due to specific antibodies, but also highlight
the implication of less-specific components. Indeed, the
decreased intestinal inflammatory response observed in
SDP-fed pigs challenged with ETEC K88 may be explained
by a lower colonisation or binding of
E. coli
to intestinal
receptors as assessed by their lower serum IgA titre and
histopathological score (Bosi
et al
., 2004). For instance, the
glycan moieties of glycoproteins would compete with nat-
ural intestinal receptors, thus reducing bacterial adhesion to
the gut (Nollet
et al
., 1999). Indeed, the protective effect of
SDP may also be non-specific as the use of a plasma
powder depleted of antibodies directed towards adhesion
factors of the challenge strain of
E. coli
(O141:K85ab
expressing the F18ac fimbriae) was efficient in protecting
piglets against oedema disease (Nollet
et al
., 1999). In piglets
orally challenged with different strains of pathogenic
E. coli
In-feed modulators of piglet immunity
1655
Table 5
Immune parameters measured in
in vivo
experiments where pigs are dietary supplied with animal by-products
Feed content
Weaning
age
Time
supplementation
PW Challenge or vaccination Intestinal immune response Systemic immune response Reference
Spray-dried animal plasma
7%
a
17 d 12 d LPS (75 mg/kg BW i.v.) 12 d PW Not measured
Blood
: TNF-a, IL-1b, IL-6, C-reactive protein,
haptoglobin
Frank
et al
. (2003)
Thymus
,
spleen
,
liver
: mRNA TNF-a, IL-1b, IL-6
6%
a
18–22 d 3 wk None
Ileocolic lymph nodes
,
ileal PP
:
mononuclear cell phenotype
Blood
: leukocyte count and phenotype Nofrarias
et al
. (2006)
6%
a
18–22 d 3 wk None
Jejunum
,
ileum
,
colon
: IEL, intravillous
lamina propria
cell density
Blood
: leukocyte subset Nofrarias
et al
. (2007)
Ileocolic lymph nodes
,
ileal PP
: leukocyte
cell subset
7%
b
14 d 1 wk LPS (150 mg/kg BW i.p.) 7 d PW Not measured
Blood
: IFN-g, TNF-aTouchette
et al
. (2002)
Thymus
,
spleen
,
liver
: mRNA TNF-a, IL-1b, IL-6
6%
c
21 d 2 wk ETEC K88 (10
10
bacteria p.o.) 4 d
PW
Jejunum
: inflammatory cell infiltration,
mRNA IL-8, TNF-a, IFN-g
Blood
: IgA anti-K88 Bosi
et al
. (2004)
10%
d
14 d 2 wk None
Jejunum
: intravillous
lamina propria
cell
density
Blood
: leukocyte count Jiang
et al
. (2000)
8%
e
21 d 2 wk
i
Rotavirus strain RV277 (2 310
6
p.o.) 5 d PW 1ETEC O149K91
(5 310
9
CFU p.o.) 7 d PW
Not measured
Blood
: anti-thermolabile toxin Ig, anti-F4
receptor Ig
Niewold
et al
. (2007)
7%
f
17 d 11 d ETEC K88 (5.5 310
8
bacteria p.o.)
12 d PW
Not measured
Blood
: IL-6 Yi
et al
. (2005)
Bovine colostrum
1 g or 5 g/d
g
21 d 3 wk None
MLN
,
jejunum
,
ileal PP
: mononuclear cell
subset and proliferation, mRNA IL-2, IL-
12, IFN-g, IL-4, IL-10
Ileum
: total Ig, anti-colostrum Ig
Blood
: leukocyte count, mononuclear cell subset
and proliferation, IgG, IgA, anti-bovine colostrum Ig
Spleen
: mononuclear cell subset and proliferation,
mRNA IL-2, IL-12, IFN-g,IL-4,IL-10
Boudry
et al
. (2007)
Lactoferrin
0.1%
h
28 d 30 d None Not measured
Blood
: lymphocyte proliferation, IL-1a, IL-2, IgG,
IgM, IgA, C3, C4
Shan
et al
. (2007)
Spleen
: lymphocyte proliferation
CFU 5colony forming unit; PW 5post-weaning; d 5day; TNF-a5tumour necrosis factor-a;IL5interleukin; wk 5week; PP 5Peyer’s patches; LPS 5lipopolysaccharide; PP 5Peyer’s patches; IEL 5intraepithelial
lymphocytes; LPS 5lipopolysaccharide; IFN-g5interferon-g;Ig5immunoglobulin; ETEC 5EnteroToxinogenic
Escherichia coli
; MLN 5mesenteric lymph nodes.
a
Appetein (SDPP), APC-Europe, Barcelona, Spain;
b
MP722, Merrick’s, Union Center, WI, origin unknown;
c
APC-Europe, Barcelona, Spain;
d
Porcine plasma protein concentrate from unknown origin, one group restricted to
control group feed intake in this study;
e
porcine SDP produced from non-immunised and immunised (
Porcilis coli
, Intervet International BV, Boxmeer, The Netherlands) pigs;
f
Unknown origin;
g
By daily gavage, spray-dried
bovine colostrum (CER, Marloie, Belgium);
h
Institute of Feed Science (Zhejiang University, Hangzhou, China);
i
Supplemented diet from d-3 post-weaning.
Gallois, Rothko
¨tter, Bailey, Stokes and Oswald
1656
(van Dijk
et al
., 2002; Bosi
et al
., 2004; Yi
et al
., 2005;
Torrallardona
et al
., 2007), SDP generally prevents growth
retardation and clinical signs, a phenomenon inconstantly
associated with decreased circulation or excretion of patho-
genic strains. This supports the theory of a direct competition
of SDP with intestinal receptors for pathogenic
E. coli
,more
than a direct anti-microbial effect (Bosi
et al
., 2004). Both
theories concerning mechanisms of action of SDP are likely,
and may act synergistically to decrease disease susceptibility
of piglets. Glycoproteins, but probably other components, may
prevent attachment of pathogens to intestine in a non-specific
way, which could still be reduced by specific antibodies
linking to pathogens. The elimination of undesired pathogens
from the gastro-intestinal tract would be facilitated.
Other animal by-products
. Bovine colostrum has a critical
role in postnatal health as an immune booster. Apart from
the passive transfer of antibody, it provides growth- and
anti-microbial factors (IGF, transforming growth factor,
epidermal growth factor, lactoferrin, lysozyme, etc.), which
will be beneficial to the neonate (Pakkanen and Aalto,
1997). In piglets, bovine colostrum has been shown to
enhance mucosa restoration by stimulating migration of
epithelial cells along the crypt–villous axis in the intestine,
and by decreasing apical cell apoptosis (Huguet
et al
.,
2007). Recent studies would suggest that bovine colostrum
has immunomodulatory properties that are directly related
to a specific region of the porcine gut-associated lymphoid
tissue. Depending upon the region studied, bovine colos-
trum could lead to Th1 (IL-2, IFN-gand IL-12) or Th2 (IL-4
and IL-10) cytokine profiles (Boudry
et al
., 2007). In ileal PP,
the increase in Th2 mRNA expression (IL-4 and IL-10) was
associated with an impaired Th1 profile (IFN-g), suggesting
that bovine colostrum supplementation may cause a more
marked Th2 immune response (Boudry
et al
., 2007). In
contrast, a Th1 profile was more pronounced in the jeju-
num. This bipolarity activities associated with bovine
colostrum would be of importance in the context of expo-
sure to a wide range of antigens (Boudry
et al
., 2007).
Bovine colostrum significantly decreased the total number
of mononuclear cells in ileal PP, but their proliferative
responses were increased (Boudry
et al
., 2007). Phenotyp-
ing of the ileal PP cells showed a very significant dose-
related decrease in CD21
1
cell count, concomitant with an
increase in the CD3
1
cell population (involving mainly the
CD4
1
subpopulation) 21 days post-weaning. Perhaps not
surprisingly, the piglets mounted a strong local antibody
response to the bovine colostral Igs by 21 days post-
weaning (Boudry
et al
., 2007). The activities associated
with fed bovine colostrum were mainly associated with the
intestinal tract, and systemic immune responses were not
altered. To our knowledge, its health benefits that could
result from its orchestration of immune responses have not
yet been studied and could be the subject of further
investigations.
Lactoferrin is a member of the transferrin family of iron-
binding glycoproteins and is ubiquitous in animal secretions
(colostrum, milk, tears). It displays a wide range of phy-
siological functions including protection against microbial
infections, regulation of non-specific immune response and
modulation of the inflammatory response (see Levay and
Viljoen (1995) for review). Both specific and innate systemic
immune responses seem dependent on lactoferrin. In pigs,
it has been shown that a 15-day lactoferrin treatment
increased the concentration of IL-2 in blood as well as the
C4 component of complement, at levels much higher than
those reached by positive controls receiving AB (Shan
et al
.,
2007). Conversely, lactoferrin did not seem to have any
impact on IL-1aor C3 blood concentrations. Lactoferrin
enhanced the proliferation of peripheral blood and spleen
lymphocytes after stimulation with phytohemagglutinin and
concanavalin A (for spleen lymphocytes only). Serum con-
centrations of IgG, IgA and IgM were also increased in
lactoferrin-supplemented piglets. To our knowledge, its
impact on local intestinal immunity as well as its potential
to enhance specific immune responses has not yet been
investigated in pigs. The use of lactoferrin is promising, as it
seems efficient in preventing diarrhoea in piglets (Shan
et
al
., 2007; Wang
et al
., 2007).
Concluding remarks
Literature dealing with natural alternatives to in-feed AB
and their impact on pig immunity is still scarce. In our
opinion, two main reasons are responsible for this lack of
knowledge. First, the total ban of in-feed AB is recent and
published works on that topic may arise in the following
years. Secondly, in-feed alternatives that can act directly on
immunity, and not through the control of the microflora,
have only recently gained interest. Indeed, the main
mechanism proposed for the action of AB as growth-
enhancers was only linked to their ability to control flora. It
was suggested that AB improve the efficiency of animal
growth via the inhibition of intestinal microflora, leading to
a reduction in the maintenance costs of the gastro-intest-
inal system including immune responses (Gaskins
et al
.,
2002). However, new data are emerging and suggest that
AB may directly interact with host cells (mainly inflamma-
tory ones), and decrease their production and excretion of
catabolic mediators (Niewold, 2007). Niewold (2007)
argues that the changes in microflora would be more likely
the consequence of the altered condition of the intestinal
wall and that this may explain the highly reproducible
effects of AB, as opposed to the inconstant results obtained
with alternatives aiming at controlling the microflora.
However, data obtained with different classes of alternative
substances able to directly interact with natural defences of
the host are not so conclusive.
As highlighted in this review, substances whose immuno-
modulatory properties often issue from
in vitro
experiments
are not so potent when tested
in vivo
as feed additives. The
heterogeneity of experiments aiming at studying the effects
of feed additives on pig immunity can partly explain the
discrepancies on their efficacy: variable composition of
In-feed modulators of piglet immunity
1657
additives, different time for supplementation, diversity of
experimental designs or measured parameters, etc. Indeed,
from the feed processing to the intestinal absorption, the
different compounds will be submitted to a myriad of
events that can reduce their activity before they reach the
cells of the gut-associated immune system (feed storage,
interactions with other nutrients, digestive processes,
absorption kinetics, etc.). Moreover, the level of the feed
additive in the final diet often remains unknown, and in
some cases, the composition of the additive itself is not
revealed. This is particularly true for yeast extracts that are
mainly commercial products, but also for plant extracts
whose active principle(s) content is highly dependent on
environmental growth conditions, harvesting time and state
of maturity or storage management (Wenk, 2003). Inde-
pendent of these basic information that should be men-
tioned in every scientific article, some experimental designs
do not seem to be fully adapted to the study of the
immunomodulatory potential of in-feed additives. Indeed,
this review highlights that effects of in-feed supplements on
systemic immunity are quite well documented, whereas
local intestinal immune responses have only received little
attention. It can be understood from both a practical and an
ethical point of view as the study of systemic immunity via
the blood does not require the slaughter of animals, and
offers the possibility of kinetic studies. However, mucosal
responses can occur independently of systemic immunity
(Cunningham-Rundles and Lin, 1998; Hannant, 2002). Thus,
the study of systemic immune responses may not reflect
immune functions and dysfunctions occurring in the gut.
In spite of all these experimental design considerations,
in the context of testing immunomodulatory compounds,
one main problem is to define what ‘desired’ immune
functions should be targeted. If there is evidence that
immunosuppressive effects are expected to suppress
potentially damaging reactions (chronic inflammation for
instance), promotion of the active immunity is conversely
required for development and education of the immune
system for both short-term and long-term health (Kelly and
Coutts, 2000). This is particularly true for mucosal surfaces
where a homeostatic balance has to be reached to both
tolerate antigens from commensal bacteria or feed, and
eliminate invading pathogens (Sansonetti, 2004). The defi-
nition of a ‘desired’ immune function is thus highly com-
plex, and in the context of animal productions, it could be
defined as the one that offers both the best growth and the
best health status to animals. This requires, however, that
piglets are not kept under very clean conditions but are
exposed to immune/infectious challenges to assess the
impact of in-feed immunomodulators simultaneously on
immunity and on performances and health.
In conclusion, the effects of in-feed additives on the
development of immune responses in gut-associated lymphoid
tissues should be more largely documented in the future, as
their effects are mainly expected in this compartment. Con-
comitantly, pharmacokinetic studies are necessary to know the
fate of these compounds in the organism, in order to precise
their site of action and biological effects. Finally, relationships
between immunomodulations induced by in-feed supplements
and health status of animals have to be clearly stated, as the
final aim of using such compounds in animal nutrition is to
promote piglet performance and health.
Acknowledgments
The authors thank the European Union for its financial support
for research in gut health and nutrition in piglets (Feed for
Pig Health project no. FP6-FOOD-1-506144). ‘The authors are
solely responsible for the work described in this article, and
their opinions are not necessarily those of the European
Union’.
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