Immunity to schistosomes in
man and snail
Source: http://www.parasitologie.nl/index.php?id=152
Written by: Marjon Kamp (3517330)
External supervisor: Dr. Jaap van Hellemond, Department of Medical Microbiology and
Infectious Diseases, Erasmus University Medical Center, Rotterdam
Examiner: Prof.dr. Lodewijk Tielens, Dept. Biochemistry & Cell Biology, Fac. Veterinary
Medicine Utrecht University
Time span: 01-11-2013/8-12-2013
1
Layman summary
Schistosomiasis or Bilharziasis is a parasitic disease found in tropical areas, caused by fluke
worms of the genus Schistosoma (S.). This parasite can infect humans and other mammals,
and freshwater snails are the intermediate host. In humans infections can cause acute
schistosomiasis, characterized by fever, cough, diarrhoea and joint pain. Chronic infections
mostly occur in endemic areas, where the schistosomes can stay in the body for several
years. These infections might result in fibrosis in different organs, including the urinary tract,
intestines, liver and spleen, with a possible fatal outcome. Infections in snails result in an
increased mortality, and a lower reproduction and thus it seems that Schistosoma infections
in snails are more severe than infections in humans because of the high mortality.
In this report the differences and similarities between snail and human immune
responses against schistosomiasis have been described. The first similarity is the
encapsulation of the parasite. In humans a capsule or granuloma is made around the
schistosome consisting of several different cell-types. In snails this capsule exists of a
multilayer of hemocytes, which are a type of blood cell. Thus it seems that containment of
the parasite is a very important part of the immune response. The killing mechanism is also
similar, in that both humans and snails need superoxides and reactive oxygen species to kill
the schistosomes. The last similarity is between human antibodies and snail molecules called
FREPs. Both antibodies and FREPs have immunoglobulin domains which are highly variable
within one individual, and both play a role in neutralizing and precipitating proteins from
schistosomes.
The biggest difference between the human and snail immune response is that snails
lack an adaptive immune response, and that infection in snails is more lethal. However the
rate of resistance in snails is higher than in humans. In the discussion several theories have
been proposed as to why snails have a higher resistance rate. The first theory that might be
considered is that recognition of the parasite is faster in snails than in humans. Snails have
their highly variable FREPs immediately available for recognition and aggregation of
schistosome molecules. However in humans, it takes at least a week to get an antibody
response. A fast response is important, for soon upon infection schistosomes cover
themselves in host- and host-like proteins in order to escape immune recognition. If
schistosomes can be detected before this transition, the chance of killing is much higher. The
second theory involves the different evolution rates. Snails have a much shorter life cycle,
and thus can adapt faster compared to humans. Besides this snails also have more to gain by
getting resistance since infections are more lethal compared to human infections.
2
Introduction
Schistosomiasis or Bilharziasis is a parasitic disease found in tropical areas, caused by fluke
worms of the genus Schistosoma (S.). Approximately 200 million people are infected
worldwide and endemic areas are mostly found in developing countries (Gryseels, 2006).
Human infections occur in surface water contaminated with cercariae. Upon entering the
body by penetrating the skin, the cercariae transform into schistosomulae and travel via the
lungs to the liver (Fig.1). Here they migrate through the tissue and enter the blood vessels of
the portal veins. Here the schistosomulae mature into adult worms, mate and then migrate
towards either the mesenteric or perivesicular venules. In this place the adult worm pairs
secrete hundreds to thousands eggs a day, which produce proteolytic enzymes in order to
reach the stool or urine. Upon entering the water the eggs hatch and release miracidia,
which then search and infect the intermediate host, freshwater snails. Inside the snail the
miracidia transform into sporocysts which multiply asexually and eventually mature into
bifurcated cercariae. An infected snail can produce up to thousands of cercariae a day, and
these cercariae are once again ready to infect humans (Gryseels, 2006).
There are different Schistosoma species, of which S. mansoni, S. haematobium and S.
japonicum are most important for human infections. S. mansoni, known to cause hepatic
and intestinal schistosomiasis, is found in Africa, South-America and Arabia, and is
transmitted by Biomphalaria freshwater snails. The species S. haematobium is found in
Africa and Arabia, transmitted by Bulinus snails and can cause urinary schistosomiasis,
whereas S. japonicum is restricted to China, the Phillippines and Indonesia. It is transmitted
by the Oncomelania snail and causes intestinal and hepatosplenic schistosomiasis. Besides
these three species there are other Schistosoma species known, however these have either a
very local spread, or use other animals as their main host (Gryseels, 2006).
The acute form of schistosomiasis, also known as Katayama fever, is characterized by
fever, cough, diarrhoea, anorexia and arthralgia (Caldas, 2008). The acute form occurs a few
weeks after primary infection and is mostly found in visitors of endemic areas, whereas the
endemic population mostly suffers from chronic schistosomiasis. Theories as to why the
endemic population does not suffer from the acute form are the early exposure of children
to the pathogen, and in-utero desensitisation. Katayama fever is a result from a
hypersensitive reaction to the schistosomes, but most patients recover spontaneously after
2-10 weeks (Gryseels, 2006; Caldas, 2008).
Chronic schistosomiasis is a problem in endemic areas and can result in different
pathological outcomes, also depending on the Schistosoma species. Initially there is a high
eosinophilic and granulamatory response towards the secreted eggs. At a later time this can
result in fibrosis in different organs, including the urinary tract, intestines, liver and spleen.
Many infected patients are asymptomatic, however, symptoms that may occur are:
haematuria, abdominal pain, diarrhoea and enlargement of the liver and spleen. In severe
cases schistosomiasis may lead to calcification of the urinary tract and severe fibrosis of the
liver, eventually leading to death (Gryseels, 2006).
Currently, treatment programs are set up, using the drug praziquantel, which
effectively paralyzes adult worms and damages their outer tegument layer. The treatment is
effective and has few side-effects, but after treatment the population is still susceptible to
re-infection. In addition, drug resistance in schistosomes should be monitored closely to
ensure that treatment remains effective, especially since cases of resistance have already
been observed, (Gryseels, 2006, Bayne, 2009).
3
Adult schistosomes can survive for decades in the human host, and give chronic
morbidity in infected humans. The schistosomes survive by complex immune evasion
mechanisms and interactions and modulations of the host immune system. Some people,
especially adults, can develop resistance against schistosomes, and also in snails resistance
has been observed. This paper will discuss the interaction between schistosomes and the
immune system, and what factors contribute to resistance in men and snail.
Figure 1: The life cycle of Schistosoma
After F. Scélo and P. Brennan (2007), The epidemiology of bladder and kidney cancer. Nature Clinical Practice
Urology. 4:205-217
1. Immune response in acute schistomiasis
Acute schistosomiasis occurs in the first few weeks after infection, and is mostly found in
naïve adults traveling to endemic areas. It is characterized as a hypersensitive reaction
towards the schistosomes. Symptoms include; coughing, fever, fatigue, myalgia, malaise and
eosinophilia (Gryseels, 2006). Larval antigens have been found in the skin only 3 hours after
penetration (Beschin, 2013).
The immune response to acute schistosomiasis involves several different immune
cells. Neutrophils in the skin detect the migrating schistosomes and their antigens, and
within 24 hours macrophages and dendritic cells (DC’s) also get involved in recognition and
induction of immune responses. The antigens are transported towards the lymph nodes by
macrophages and DC’s within 40 hours, to activate an adaptive immune response (Beschin,
2013). The DC’s activated by the antigens of immature worms, induce a T-helper1 (Th1)
response (Pearce, 2012).
4
As a response to the invading schistosomes, eosinophils, basophils and mast cells are
activated and recruited towards the site of infection. They excrete reactive oxygen- and
nitrogen intermediates (ROI/RNI) and lytic enzymes to kill the parasites, as well as employing
antibody dependent cell-mediated cytotoxicity (ADCC) (Beschin, 2013).
In acute schistosomiasis patients, high concentrations of IgG, IgE and IgM are
detectable in serum, against antigens present in cercariae and adult worms. IgM antibodies
against cercarial antigens were found in mice one week after infection. An important
schistosomal antigen is a protein expressed on the surface of miracidia and schistosomula,
with similar carbohydrate epitopes as the protein keyhole limpet hemocyanin (KLH). KLH is a
protein from the keyhole limpet that is known to induce strong antibody responses, and this
KLH-like antigen from schistosomes also induces a strong IgG and IgM response,
characteristic for acute infection (Rabello, 1995). Antibodies play an important role in killing
of the schistosomes through ADCC, primarily by eosinophils (Beschin, 2013) and there are
indications that acquired immunity is mediated by IgE against larvae and adult worms
(Gryseels, 2006).
Elevated levels of IFN-γ and TNF-α have been found in patients with acute
schistosomiasis, as well as the pro-inflammatory cytokines IL-1, IL-2 and IL-6 (Caldas, 2008;
Pearce, 2012). Both INF-γ and TNF-α are indicators of a Th1 response, but IL-6 is associated
with Th2 responses.
Alltogether, the immune response in acute schistosomiasis is characterized by a
strong pro-inflammatory reaction against the schistosomes. It results in a Th1 response,
inducing antibody production and eosinophilia. The eosinophils are able to kill the parasites
by excreting destructive molecules and by ADCC.
2. Chronic disease and immune response
As the infection progresses, the worms migrate to the liver where they mature. Upon mating
and migration to the urinary or mesenteric veins they start excreting eggs. Egg secretion
marks the transition from acute to chronic schistosomiasis, as the immune reaction gets
modulated by egg antigens. Part of the produced eggs gets stuck in the liver, intestines or
perivesicular structures, where they keep secreting their proteolytic and immunogenic
enzymes. The eggs and the secreted enzymes induce a strong granulomatous reaction,
resulting in an accumulation of granulomas in the tissue. As the disease progresses, the
granulomas may result in fibrosis in different organs, eventually resulting in lethal
complications like liver fibrosis and bladder calcification (Gryseels, 2006).
2.1. Cellular response
As a response to egg production and release of egg antigens, a switch occurs from a Th1towards a Th2-response (Fig.2). Simultaneously the acute pathology proceeds to chronic
schistosomiasis (Fairfax, 2012). Certain carbohydrates on egg antigens are responsible for
this shift, and in particular lacto-N-fucopentaose III is a strong Th2 adjuvant (Pearce, 2002).
Recently the role of omega-1 in the Th2 shift was established. Omega-1 is a RNase secreted
by S. mansoni eggs, and is found to be responsible for priming DC’s to a Th2 reaction.
Omega-1 is taken up via the Mannose Receptor (MR) on DC’s, and degrades intracellular
messenger- and ribosomal RNA, leading to an altered, Th2-driven, protein expression
(Everts, 2012; Everts, 2009).
The Th2 cells promote formation of granuloma by orchestrating different cell types,
thereby protecting the organs from damage by the proteolytic enzymes. The granulomas
5
give immediate protection of the tissue, however, in the long run the granulomas can lead to
fibrosis, thus indicating that a Th2 response has beneficial effects in the short term, but
detrimental effects on the long term (Fairfax, 2012).
An important cell type in chronic schistosomiasis is the macrophage. The cytokines IL4 and IL-13, which are produced by Th2 cells, induce ‘’alternative activation’’ (AA)
macrophages. These macrophages play an important role in wound repair and in supressing
inflammation. They achieve this by producing arginase-1, which is an enzyme that converts
L-arginine into L-ornithine and urea. Hereby it is able to compete with Th2 cells for arginine,
and is able to limit the Th2 response (Fairfax, 2012). The AA-macrophages are able to inhibit
inflammation, but thereby promote fibrosis. They produce metalloproteinases (MMP’s),
which are necessary for degradation of extracellular matrix (ECM) and tissue remodelling.
However these MMP’s also induce fibrosis of the tissue. The balance between Th2 and AAmacrophages is an important factor to determine the outcome of the disease, however both
are necessary to prevent disease (Fairfax, 2012).
Th2 cells will also activate B-cells, which are responsible for antibody production
against the different egg antigens. The antibodies are able to neutralize some of the
immunogenic proteolytic enzymes within the granuloma and reduce inflammation. A lack of
B-cells leads to portal hypertension and worsening of the disease (Fairfax, 2012).
Next to the Th2 response, there are also regulatory T-cells (Tregs) induced, following
egg antigen presentation. Treg induction requires the presence of Toll like receptor 2 (TLR2).
Absence of TLR2 will lead to a prolonged Th1 response and more immunopathology. Tregs
have an immune-modulating role, and reduce the granuloma size (Fairfax, 2012). In the
granuloma many eosinophils have been found, and they are known to secrete IL-4 and IL-13,
thereby inducing AA-macrophages. However, the main function of eosinophils in granuloma
is still unknown (Fairfax, 2012).
Altogether, the cellular immune response in the chronic infection is dominated by
reaction towards eggs, forming granuloma around them. The immune reactions are
supressed by different cell types to reduce immune-response related tissue damage. Tissue
remodelling is an important part, which is mostly regulated by AA-macrophages, but which
can lead to fibrosis.
2.2. Humoral responses
In chronic schistosomiasis patients there is higher concentration of antibodies against eggs
than to adult worms. These antibodies help with neutralizing the excreted proteins from the
eggs in the granuloma (Fairfax, 2012). It has been indicated that IgG4 antibodies against egg
antigens might be a risk factor for fibrosis, since higher concentrations are found in fibrotic
patients. However, it has not yet been determined if IgG4 is causative of fibrosis, or is an
effect (Caldas, 2008).
2.3. Cytokine reaction
In chronic schistosomiasis patients the cytokine pattern changes from a Th1-, to a Th2pattern. The IFN-γ production is decreased, and more anti-inflammatory cytokines are
induced, including IL-10, IL-4 and IL-13 (Caldas, 2008).
6
Figure 2: Development of the immune response in schistosome infections
After E. J. Pearce and A.S. MacDonald (2002), The immunobiology of schistosomiasis. Nature reviews Immunology,
2(7), 499–511
The switch towards a Th2-response results in lower IFN-γ production. IFN-γ seems to
play a role in inhibiting fibrosis by regulating fibroblasts and collagen production. This
corresponds with the very low concentrations of IFN-γ found in patients with severe fibrosis
(Caldas, 2008).
IL-10 is a well-known anti-inflammatory cytokine, which decreases the amount of
inflammation. This cytokine is important in keeping the tissue damage in control. IL-10 is
responsible for modulating Th2 responses and inhibits the proliferation of peripheral blood
mononuclear cells (Caldas, 2008). It is also involved in inducing the anti-inflammatory AAmacrophages (Fairfax, 2012).
IL-4, as produced by Th2 cells, has an anti-inflammatory effect as well and is an
important activator of AA-macrophages. Absence of IL-4 in mice leads to profound tissue
injury in the liver and intestines, and is lethal. This indicates that IL-4 is essential in
controlling inflammatory reactions in response to schistosomiasis (Fairfax, 2012).
The concentration of IL-13 is correlated to the amount of fibrosis (Caldas, 2008). In
accordance with this, IL-13 is responsible for activation of AA-macrophages which, as
described above, are involved in tissue remodelling and promote fibrosis (Beschin, 2013).
To conclude, it is clear that the immune reaction in chronic patients is much more
subdued. Many of the cells and cytokines involved in the chronic reaction are involved with
suppression of inflammation and promoting tissue repair and remodelling. This
inflammatory inhibition is essential to prevent disease in the short term, however in the
long-term it may lead to substantial fibrosis which can lead to organ failure and death of the
host.
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3. Resistance and aging
3.1. Age-related resistance versus acquired immunity
Adults are more resistant to schistosomiasis
infections than children. In endemic areas this
resistance is partly due to acquired immunity upon
years of exposure. However, in recently endemic
sites a lower infection rate in adults compared to
children was observed as well, which was not
correlated to amount of exposure (Kabatereine,
1999). Children aged between 10 and 15 years old
are most susceptible (Fig.3), and show high
individual variety in susceptibility to re-infection.
This variety is especially clear in endemic areas
with high parasitic burdens (Gryseels, 1994). This
indicates that besides acquired immunity, there Figure 3: Age distribution of infection
are general factors that play a role in increased in recent immigrants to an endemic
resistance in adults. Besides this there are area . The peak of infection is found in
individual differences in susceptibility, which is 10-15 year-old children.
most clearly visible in children (Gryseels, 1994, Gryseels, 1994
Gryseels 2006).
3.2. Immune mechanisms that mediate resistance
Acquired immunity is mostly mediated by IgE directed against larvae and adult worms,
triggering ADCC by eosinophils (Gryseels, 2006). Another study showed that individuals from
endemic areas with high resistance had a high concentration of IgG against a 37kDa antigen
derived from larvae (Dessein, 1988). Yet another study describes the role of both IgE and
IgG4 against the tegumental antigen smTal1, as a factor of resistance. It seems that the main
theme in resistance lies in antibody production, however, this does not explain the variation
of resistance throughout aging. Neither does it explain why vaccine development, using
several dominant antigens, has not yet resulted in a successful vaccine.
3.3. Role of hormones
One explanation for the variation in resistance throughout aging might be variation of
hormone levels. The levels of testosterone precursor dehydroepiandrosterone (DHEA) and
its anologue DHEA sulphate (DHEAS) rise in male teenagers (15-19 years), reaching a peak at
20-29 year-olds. Simultaneously the infection intensity drops and the lowest infection
intensity coincides with the DHEA-peak. Treatment of female mice with DHEAS resulted in
significant decrease of worm burden, and oviposition in S. mansoni was affected both in vivo
and in vitro by DHEAS (Morales-Montor, 2007). However this reason would only account for
a decrease in susceptibility in the male population, whereas the correlation between age
and susceptibility is found in both males and females.
3.4. Role of Cytokines
Another point of interest in age-related resistance is the role of cytokines. An IL-10 response
occurs early in life, and peaks in 11-12 year-old children. As described earlier, IL-10 has an
immune-modulatory role, and is associated with chronic schistosomiasis. However, a direct
8
link between high serum levels of IL-10 and an increased susceptibility has not yet been
found. A possible explanation is that IL-10 production increases early in infection, and
influences infection intensity but not susceptibility. Cytokine levels of IL-4 and IL-5 were
highest in age groups where infection intensity and prevalence were the lowest, indicating a
protective role. It was also suggested that IL-10 is able to inhibit IL-5 production, and thereby
hinders the protective role of IL-5 (Mutapi, 2007).
3.5. Reduced resistance in elderly
A point not often addressed, is the reduced resistance in elderly. This reduced resistance
might be due to a decrease in the T-cell repertoire as an effect of thymic involution.
However, there are elderly individuals that maintain resistance over the years. This
resistance is achieved by compensating the T-cell loss with an enhanced innate immune
system, including more Natural Killer cells (NK-cells) producing IFN-γ and higher expression
of the pattern recognition receptors called TLR’s, in particular TLR-1 (Comin, 2008).
Whereas an enhanced innate immune system works protective, it seems that
increasing immune regulation inhibits resistance in elderly. In susceptible elderly, high levels
of IL-10 secretion were found, while IFN-γ production decreased. This in combination with
an increase of Tregs in susceptible elderly, suggests that immune-modulation is
unfavourable for development of resistance (Comin, 2008)
4. Snail Immunity
The infection cyclus in snails starts with infection by miracidia, which hatch from the eggs
excreted in human urine of faeces. The miracidia enter the snail by secreting the contents of
their penetration glands and by muscular squirming. Upon infection the miracidia shed their
ciliated surface ‘plates’ and discharge lipids and cytoplasm, which forms a tegument layer,
transforming the miracidium into a sporocyst. The sporocyst stays in the body cavity for
several weeks, exposed to the immune system of the snail, while the germinal cells are
developing into a second generation of daughter sporocysts (Bayne, 2009). These newly
formed daughter sporocysts migrate to the hepatopancreas where cercariae are produced
(Gryseels, 2006). Approximately one month after infection shedding starts, lasting several
weeks. Thousands of cercariae are then shed into the water, ready to infect humans (Bayne,
2009). Infection of snails results in higher mortality rates of the snail and causes partial
castration (Webster, 2004).
The specificity of miracidia for their intermediate hosts is quite remarkable. The S.
mansoni species can only infect Biomphalaria snails, whereas S. haematobium is specific for
Bulinus snails and S. japonicum specific for Oncomelania snails. This specificity is less
pronounced in their definitive hosts, as all these schistosome species are able to infect
humans, and often several other mammals. This high specificity is an indication that the
schistosomes and snail species have co-evolved and are highly adapted (Webster, 2004).
4.1. Immune system of the snail
The immune system of a snail is less advanced when compared to mammals, and has no
adaptive immune response. The different components of the immune system in a snail will
be described shortly, to get a better understanding of the infection process.
Snails have several different cells that play a role in immunity. The first one is the
antigen trapping endothelial cell, lining blood vessels. This cell is stationary, preventing
dissemination of microbes, and presents microbes to phagocytic cells. Reticulum cells and
9
pore cells are also immobile, and play a role in recognizing and internalizing non-self
material and proteins, respectively. The most important immune cells however, are the
hemocytes (van der Knaap, 1990).
Hemocytes are mobile cells, which can phagocytose microbes, and produce oxygen
radicals and hydrogen peroxide, used for killing. They are somewhat similar in their role to
human neutrophils. The mobility of these cells enables them to carry out a surveillance
function in different tissues. They detect non-self material, and subsequently induce an
immune reaction. This immune reaction can involve phagocytosis, cytotoxic killing, or
production of lectins.
Lectins are carbohydrate-binding proteins which are found as receptors on
hemocytes, and soluble in the plasma. On the hemocytes the lectins function as receptors
for non-self pattern recognition. Upon an infection they can also be secreted as soluble
factors, and here they play a role in agglutination of microbes and opsonisation for
phagocytosis by hemocytes. Lectins play a very important and central role in almost all
immune reactions in invertebrates (van der Knaap, 1990).
4.2. Immediate response
Upon entering snails, the trematodes release a range of molecules now known as their
excretory-secretory products (ESP). These ESP molecules include superoxide dismutases,
cysteine proteases and polymorphic mucins. It has been suggested that these polymorphic
mucins are recognized by plasma factors and hemocytes.
The immediate response is directed towards these invading miracidia. This reaction is
fast and eliminates the miracidia, preventing infection (Carton, 2005). The outcome depends
on the fact if the miracidium manages to transform into a mother-sporocyst, or is eliminated
before (van der Knaap, 1990).
4.3. Late response
A later response is directed against the sporocysts themselves, mediated by hemocytes.
Especially the granulomous hemocytes (granulocytes) are important in the control and killing
of sporocysts. The immune reaction can be divided into three steps: recognition, capsule
formation and killing (van der Knaap, 1990).
Recognition is regulated by the lectin receptors on the hemocytes. The lectins are
pattern recognition receptors, which activate the hemocytes upon encountering non-self
material (van der Knaap, 1990). The fibrinogen-related proteins (FREPs) also seem to play a
role in the recognition. FREPs contain one or two immunoglobulin-like (Ig) domains, which
are highly diverse between individual snails, and contain a fibrinogen domain as well. Plasma
levels of FREPs rise following trematode infections, precipitating glycan-bearing molecules
that are released from the schistosomes. However, the presence FREPs in plasma is not
predictive of snail resistance, and FREPs have not yet been identified as membrane receptors
(Bayne, 2009). After the initial recognition, hemocytes are activated and migrate toward the
sporocysts. They accumulate and form a multi-layered capsule around the sporocysts.
The last step in the immune response is killing of the sporocysts. Killing is achieved
through oxygen-dependent mechanisms, phagocytosis and cytotoxicity (Carton, 2005;
Bayne, 2009). The sporocysts are too big for proper phagocytosis, causing hemocytes to
cover part of the sporocyst and release their phagosomal and lysosomal components in this
cleft (frustrated phagocytosis). The importance of ROS in killing was demonstrated by adding
10
ROS-scavenging molecules or inhibitors of the respiratory burst pathway, which led to a
compromised killing capacity of hemocytes (Bayne, 2009). IL-1 like molecules seem essential
in priming superoxide production (Carton, 2005). Peroxidases, superoxide and hydrogen
peroxide were all found in the capsules of hemocytes, indicating a similar killing mechanism
as ADCC in humans. It seems that plasma factors play an important role in killing, since
addition of plasma from resistant snails resulted in killing by hemocytes from susceptible
snails (van der Knaap, 1990)
5.
Snail resistance
Snails have a high degree of resistance to schistosomes compared to humans, and there is
very stringent host snail specificity. A miracidium is often able to only infect some individuals
of the same snail strain, because of the high variety in susceptibility even within strains. The
susceptibility of a snail is both determined by the genotype of the miracidium as well as the
snail (Bayne, 2009). Several mechanisms involved in snail resistance will be described below.
5.1. Resistance difference between young and old snails
In snails there is a great variation in resistance to schistosome infections among and within
populations. Some resistance traits seem to be associated with aging, as susceptible juvenile
snails can gain resistance upon maturation. However there are also juvenile snails that are
already resistant. Most research in the field of resistance has been performed with the B.
glabrata and S. mansoni model. Different factors that play a role in the snail resistance will
be discussed below (Carton, 2005).
5.2. Genetic factors
According to a breeding study, resistance appeared to be genetically determined by a single
gene, with a dominant profile for the resistance allele (Knight, 1999). The gene found in this
study seems to correlate with the R-gene, which is expressed in both young and adult snails.
However, upon further breeding studies it became clear that the resistance profile was very
specific for a certain S. mansoni strain, since resistance was lost when the snails were
challenged with another S. mansoni strain. Upon challenge with the new S. mansoni strain
the resistance in snails was no longer divided following mendelian expectations, indicating
that other genes may play a role (Carton, 2005).
It is implied that susceptibility in juvenile snails is partly dependent on multiple
genes, called the juvenile susceptibility gene complex. The Mr gene, which is only expressed
in adult snails, is responsible for the suppression of the juvenile susceptibility gene complex
in adult worms, rendering them resistant and allowing R-gene expression (Carton, 2005).
It seems that several genes play a role in the resistance in snails, and that each gene
has several alleles. This may also explain the high variation in susceptibility in snails.
5.3. Expression patterns
Next to genetic differences, there were also differences found between expression patterns
in resistant and susceptible snails. One example is the down regulation of a gene
homologous to P450 in the ovotestes of resistant snails (Carton, 2005). In humans the P450
superfamily is responsible oxygenation of many substrates in several metabolic processes,
including the oxygenation and degradation of drugs.
11
Another example of differential expression is that cDNA of the Heat Shock Protein 70
(HSP70) gene was only found in infected resistant snails, suggesting a protective role
(Carton, 2005).
In a large screen in hemocytes, using differential display reverse-transcription PCR
(DDRT-PCR), many genes were identified to be differentially expressed between resistant
and susceptible snails. The most intriguing of which, show homology to adhesion molecules,
neutrophil defensins, glycosidases and peroxidases (Schneider, 2001).
5.4. Recognition
Recognition of the parasite is of course a very important factor in determining the outcome
of infection. However, the recognition is dependent on both snail and schistosome factors.
As described earlier, miracidia secrete ESP molecules, including polymorphic mucins. These
mucins are highly diverse between individual miracidia, which is partly regulated by
epigenetic changes. This high variety helps the miracidia in evading detection (Roger, 2008).
It seems that the snail counteracts against these polymorphic mucins, with the family of
FREP proteins. These FREP proteins exhibit a high variety as well, and play a role in
neutralizing and precipitating ESP molecules (Bayne, 2009)
5.5. Killing
If the differences in immune reactions between resistant and susceptible snails are
compared, it becomes clear that resistant snails are more capable of killing schistosomes.
Resistant snails have a higher phagocytic and cytotoxic capacity compared to susceptible
snails. In resistant snails, a higher concentration of hemocytes produces superoxides, used in
killing of schistosomes (Carton, 2005).
The induction of cytotoxic functions seems to be dependent on plasma factors, since
co-incubation of plasma from resistant snails and hemocytes from susceptible snails often
induces cytotoxicity from hemocytes in vitro. Plasma alone, from resistant snails does not
possess killing capacities, indicating that plasma factors play a role in inducing the
cytotoxicity (van der Knaap, 1990).
The production of H2O2 was found to be decreased in susceptible snails. Related to
this, the activity of superoxide dismutase (SOD), which accelerates the generation of H2O2,
was approximately half compared to SOD activity in resistant snails. Three alleles have been
found of the associated gene (SOD1) in B. glabrata, one of which is significantly associated
with snail resistance. It has been suggested that other genes involved in the H2O2 production
could be potential resistance genes (Bayne, 2009).
Lastly, IL-1 like molecules might be involved in priming superoxide-release in resistant
snails. In conclusion, the immune reaction towards the parasite is more intense in resistant
snails, leading to a more efficient killing (Carton, 2005).
5.6. Costs and trade-off
Knowing that schistosome infections in snails result in significantly increased mortality, and
in partial castration, it seems obvious that resistance is highly beneficial. However, there are
certain costs associated with resistance (Webster, 2004). It seems that resistant snails have a
loss of fertility and have a delay in becoming reproductively mature. The ratio of
resistant/susceptible snails seems to drop after several generations without the presence of
schistosomes, since it is no longer beneficial (Carton, 2005).
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7.
Discussion
7.1. Snail and human resistance
Snails are quite adept at clearing schistosome infections, which is partly because of the fast
immune response, often initiated by the recognition of secreted ESP of the miracidia. These
ESP include highly variable mucins, however snails seem to have adapted to this variety by
using their own variable FREPs (Bayne,2009). Though FREPs have mostly been found in
plasma, we might hypothesize that they play a role in the initial recognition of infection, and
subsequent activation of the hemocytes.
It seems that many factors play a role in determining the outcome of the infection in
snails, and this is both dependent on phenotype of snail and schistosome. Factors that play a
role in schistosome-snail compatibility include: Mucin-FREP interactions, snail maturity,
production of hemocyte activating plasma factors, and killing capacity of hemocytes. All
these factors contribute to clearance of the schistosomes, but infection dose and evasion
mechanisms of the schistosome may counteract the snail immune response. The balance
between all these factors determines successful infection or clearance. However, it seems
that snails have become well adapted and have taken the overhand, since most snails are
resistant.
A short overview will be given of the resemblance and differences between the
immune response in humans and snails. At the end some ideas will be offered why snails
have developed a better resistance mechanism than humans.
7.2. Similarities in immune response
One remarkable finding, is that both in man and snail resistance increases through aging. In
snails this effect is mostly attributable to the juvenile susceptibility gene complex (Carton,
2005). However in humans it seems that the increase of resistance through aging is multifactorial. Although in humans this increase can partially be explained by acquired immunity,
there are still other more obscure factors contributing. The resistance might partially be
explained by changing hormone levels throughout aging, of which DHEAS is a good example.
DHEAS is a hormone which expression rises during puberty in boys. DHEAS can decrease
parasitic burden in mice (Morales-Montor, 2007). Another factor that might play a role is
differential cytokine levels throughout aging, where it seems that IL-4 and IL-5 have a
protective function (Mutapi, 2007).
Another similarity is the encapsulation of the schistosomes. Where in humans the
granuloma exist of several cell-types, in snails they exist of a multilayer of hemocytes. But in
both man and snail it seems important to contain the schistosomes. One of the reasons why
encapsulation may be a similar theme is because of the size of schistosomes. The parasites
are too big for phagocytosis, thus resulting in encapsulation by several immune cells.
Both in the immune response of man and snail, the killing mechanisms share a close
resemblance. Though the cells responsible might be different, neutrophils and eosinophils
and hemocytes respectively, the mechanism of killing is very similar. In both hosts
superoxides and ROS are used in killing schistosomes. This seems to be a consistent and thus
effective mechanism for killing.
The last similarity that will be brought to attention is the similarity between human
antibodies and snail FREPs. Both antibodies and FREPs have immunoglobulin domains which
are highly variable within one individual, and both play a role in neutralizing and
precipitating proteins from schistosomes.
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7.3. Differences in Immune response
While there are many similarities between the human and snail immune response, there are
also some major differences. The first and most obvious is that snails don’t have an adaptive
immune response. In humans antibody production seems to play an important role in
developing resistance, while in snails the adaptive immune response is completely lacking.
However, as mentioned before, it seems that snails have adapted to this by producing a high
variety of antibody-like molecules called FREPs.
Another big difference is the consequence of infection in man and snail. In humans
infection is often chronic, resulting in some symptoms which in the long term become lethal.
However the disease does not influence reproduction and the overall lethality is low. In
snails the impact of infection is much bigger, causing partial castration and a significant
increase in mortality.
7.4. Why are snails more effective in clearing schistosomes?
Upon reviewing the immune responses of both snails and humans it became clear how the
responses are similar and how they differ. However, a point that has not yet been discussed
in great detail is why snails are better at clearing schistosome infections. Here we pose
several theories that may contribute to this phenomenon.
The first theory that might be considered is that recognition of the parasite is faster
in snails than in humans. Snails have their highly variable FREPs immediately available for
recognition and aggregation of schistosome molecules. However in humans, it takes at least
a week to get a specific antibody response, which carry out similar functions as the FREPs in
snails. This fast response is important, for soon upon infection schistosomes cover
themselves in host- and host-like proteins in order to escape immune recognition. If
schistosomes can be detected before this transition, the chance of killing is much higher.
The second theory involves the different evolution rates of snails, schistosomes and
humans. The life cycle of the snail is quite short compared to schistosomes, and very short
compared to humans. This means that in a set amount of time, snails will have produced the
most generations, and therefore have had the most chance to adapt and pass on beneficial
genes. Therefore snails are able to adapt faster than both schistosomes and humans, and
this might explain why they are so capable of dealing with schistosome infections.
The last hypothesis involves another aspect of evolution, namely the selection
pressure. Schistosome infections in snails greatly reduce the reproduction rate, both by
partial castration and by higher mortality rates. These factors contribute notably to the
reduction of susceptible snails in endemic areas. However, in humans the infection often has
no effect on reproductive abilities whatsoever. Therefore there is barely any selection
pressure in humans, whereas the selection in snails is very strict.
14
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