SCHISTOSOMIASIS VACCINE TECHNICAL FIELD

[0001] The invention relates to prevention and/or treatment of schistosomiasis. In particular, the present invention provides a vaccine that comprises one or more schistosome immunogenic proteins.

BACKGROUND

[0002] The carcinogenic blood fluke, Schistosoma haematobium, infects more than 100 million people throughout Africa and is the most prevalent of the human schistosomes, causing more than half of all infections (1). S. haematobium adult flukes migrate to the vasculature of the organs of the pelvis. Severe morbidity results from host immune responses to eggs in tissues and includes periportal fibrosis, portal hypertension, and hepato-splenic disease (2). Formerly known as urinary schistosomiasis, S. haematobium infection was recently renamed "urogenital schistosomiasis" in recognition that the disease affects both the urinary and genital tracts of women and men. Female S. haematobium lay between 20 and 200 eggs daily (3), which penetrate the vessel wall and move towards the lumen of the bladder. Some of the eggs become sequestered in the tissue of the pelvic organs such as the urinary bladder, ureters, cervix, vagina, prostate gland, and seminal vesicles, where they cause chronic inflammation, pelvic pain, bleeding, and an altered cervical epithelium in women (4). S. haematobium is unique among the schistosomes in its recognition as a group I carcinogen by the International Agency for Research on Cancer because of its robust association with urothelial carcinoma (5). S. haematobium infection also increases susceptibility to infection with HIV-1, progression to disease, and results in a higher likelihood of transmitting infection to others (6).

[0003] Praziquantel (PZQ) is widely used to treat human schistosome infections and has two main effects on schistosomes - paralysis and tegument damage (7). An added benefit of PZQ treatment is that it mediates destruction of flukes thereby exposing antigens on the worm surface to the host immune system. This release of surface antigens induces and/or enhances parasite-specific immune responses (8), resulting in immune-mediated killing of the parasite. Early studies reported modifications in T cell proliferative responses (9), whereas recent studies noted modifications in the levels and types of antibody (10-13) and cytokine responses (14-16) following PZQ treatment. The immune response triggered by PZQ treatment is thought to last for more than one year (14, 17-19) and confer at least some level of resistance to reinfection. This phenomenon is referred to as "drug-induced resistance" (DIR) (20). The mechanisms behind DIR differ significantly from those of putative natural resistance (PR, resistant individuals who have not received PZQ therapy) and can be related to the origin (developmental stage) and concentration of the released antigen, as well as the type of antigen-presenting cells (APCs) involved. PZQ treatment introduces a large amount of adult fluke antigen directly into the bloodstream as a result of many worms dying at once (21), whereas naturally acquired resistance in the absence of PZQ treatment (PR) is stimulated by the introduction smaller quantities of adult antigen due to a more gradual worm death. The process of PR is additionally stimulated by the release of antigens from naturally dying larval schistosomes (schistosomula) primarily through the skin and pulmonary vasculature, thus inducing different APCs and resulting in different interactions between the antigens and the immune system (22). This additional stimulus does not appear to factor significantly in DIR due to the ineffectiveness of PZQ against schistosomula (7, 8). Whatever the mechanism, it is important that an antigen threshold is reached in order to sufficiently stimulate anti- schistosome immunity (23, 24).

[0004] Studies with car washers in schistosome-infected waters of Lake Victoria in Kenya showed that a subset of the men developed resistance to re-infection after PZQ therapy while others remained susceptible despite treatment (25, 26). It was found that IgE production to soluble worm antigen preparation (SWAP) paralleled the development of resistance, and did not occur in those who remained susceptible to reinfection (25). Additionally, immuno-proteomic studies have used S. haematobium SWAP to identify a number of antigens that are released by PZQ treatment and/or are the target of DIR immune responses (27, 28). However, despite the power of these proteomic studies in identifying individual parasite proteins, the utilization of SWAP (where worms are homogenized and solubilized under native conditions in the absence of detergents that will solubilize the cell membranes) does not result in full representation of the S. haematobium proteome. Indeed, numerous abundantly expressed proteins with multiple membrane spanning domains that are released from the tegument with detergents (29, 30) are accessible to chemical labeling on the surface of live worms (30), are recognized by sera from PR individuals, and are lead vaccine antigens against schistosomiasis (31-33). [0005] A third mechanism of resistance to schistosomiasis is seen in the rhesus macaque (Macacca mulatto). It is unique among animal models of schistosomiasis in that, once an infection reaches patency, worm death starts to occur from week 10 (34) and egg output diminishes over time until the infection is eliminated (35, 36). This phenomenon only occurs above a threshold worm burden (35, 36), presumably as sufficient immune stimulus is required for this process to occur (23, 24). This self- cure mechanism is thought to be antibody-mediated because of a strong inverse association between the rapidity and intensity of the IgG response and the number and morphology of surviving worms (34). Two-dimensional immunoblotting of worm extracts showed the immune response to be directed at gut digestive enzymes, tegument surface hydrolases and anti-oxidant enzymes (34).

SUMMARY

[0006] To overcome the deficiencies of the approaches listed above, a novel immunomics approach was undertaken to identify antigens which are the targets of humoral immune responses in (1) DIR human subjects from an S. haematobium- endemic area in Africa, and (2) rhesus macaques that had undergone self-cure after experimental S. japonicum infection.

[0007] It is therefore an object of this invention to provide one or more Schistosoma haematobium immunogens that, preferably, are useful in a safe and efficacious vaccine against various Schistosoma species, including but not limited to S. mansoni, S. japonicum and S. haematobium.

[0008] A broad form of the invention provides one or a plurality of isolated, immunogenic proteins obtained, or obtainable from, a schistosome such as S. haematobium, or a variant or fragment thereof, for use in preventing, treating or immunizing against a schistosome infection.

[0009] An aspect of the invention provides an immunogenic composition comprising one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof; one or a plurality of isolated nucleic acids encoding SEQ ID NOS: 1-3, or a fragment or variant thereof; or an antibody or antibody fragment which binds one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof.

[0010] In an embodiment, the immunogenic composition is capable of eliciting a protective immune response upon administration to a mammal. [0011] In a particular embodiment, the immunogenic composition is a vaccine.

[0012] Another aspect of the invention provides a method of eliciting an immune response in a mammal, said method including the step of administering an immunogenic composition to the mammal to thereby elicit an immune response to a schistosome in the mammal, said immunogenic composition comprising one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof; one or a plurality of isolated nucleic acids encoding SEQ ID NOS: 1-3, or a fragment or variant thereof; or an antibody or antibody fragment which binds one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof.

[0013] In an embodiment, the immunogenic composition elicits a protective immune response after administration to the mammal.

[0014] Yet another aspect of the invention provides a method of immunizing a mammal, said method including the step of administering an immunogenic composition to the mammal to thereby immunize the mammal against infection by a schistosome, said immunogenic composition comprising one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof; one or a plurality of isolated nucleic acids encoding SEQ ID NOS: 1-3, or a fragment or variant thereof; or an antibody or antibody fragment which binds one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof.

[0015] Still yet another aspect of the invention provides a method of preventing or treating a schistosome infection in a mammal, said method including the step of administering an immunogenic composition to the mammal to thereby prevent or treat a Schistosome infection in the mammal, said immunogenic composition comprising one or a plurality of isolated proteins one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof; one or a plurality of isolated nucleic acids encoding SEQ ID NOS: 1- 3, or a fragment or variant thereof; or an antibody or antibody fragment which binds one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof.

[0016] In some embodiments of the aforementioned aspects, SEQ ID NOS: 1-3, or fragments or variants thereof, are in the form of a single protein, such as a chimeric protein. In a particular embodiment, the chimeric protein comprises an amino acid sequence set forth in SEQ ID NO:4 or SEQ ID NO:5.

[0017] A further aspect of the invention provides an isolated, immunogenic chimeric protein comprising at least two of the respective amino acid sequences set forth in SEQ ID NOS: 1-3, or at least two fragments or variants of SEQ ID NOS: 1-3. In particular embodiments, the isolated, immunogenic chimeric protein comprises an amino acid sequence set forth in SEQ ID NO: 4 or SEQ ID NO: 5.

[0018] This further aspect of the invention also provides an isolated nucleic acid encoding the isolated, immunogenic chimeric protein, a genetic construct comprising the isolated nucleic acid and/or a host cell comprising the genetic construct.

[0019] This further aspect of the invention also provides an antibody which binds and/or is raised against the isolated chimeric protein.

[0020] Throughout this specification, unless the context requires otherwise, the words "comprise", "comprises" and "comprising" will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

[0021] As used in this specification the indefinite articles "a" and "an" may refer to one entity or a plurality of entities (e.g. proteins) and are not to be read or understood as being limited to a single entity.

BRIEF DESCRIPTION OF THE FIGURES

[0022] Figure 1. Characterization of study cohort and sub-cohort used for the study described herein. *Treatment efficacy was assessed by urinalysis 6 weeks after praziquantel therapy - all subjects were egg-negative (no eggs found in any of 3 urine samples, each collected on a separate day). ^{^} Subjects remained in the endemic study area and had regular water contact for the study duration.

[0023] Figure 2. Antibody responses to arrayed antigens differ in Schistosoma haematobium-infected humans before and after praziquantel treatment. (A) IgGl . (B) IgE. Average adjusted signal intensity values depicting the antibody response to each reactive antigen are shown for the drug-induced resistant (DIR) cohort before and after praziquantel treatment. The dashed and solid lines are the respective cut-offs for IgGl (8239) and IgE (1861) reactivity, calculated as one standard deviation of the mean of the no-DNA control spots on the array. Statistical analysis was performed using student's t test. *p<0.05, **p<0.01, ***p<0.001. [0024] Figure 3. IgGl antibody profiles to arrayed antigens differ between Schistosoma haematobium-infected humans who do and do not acquire drug-induced resistance after praziquantel treatment. Average adjusted signal intensity values depicting IgGl antibody responses to each reactive antigen are shown for the drug- induced resistant (DIR) and chronically infected (CI) cohorts after praziquantel treatment. Boxed antigens indicate homologues of known vaccine candidates. The dashed line is the cut-off for IgGl reactivity (8239), calculated as one standard deviation of the mean of the no-DNA control spots on the array. Statistical analysis was performed using student's t test. *p<0.05, **p<0.01, ***p<0.001.

[0025] Figure 4. Some arrayed antigens that induce IgGi responses in Schistosoma haematobium-infected humans who acquire drug-induced resistance after praziquantel treatment are not the targets of IgE. Average adjusted signal intensity values depicting IgGl and IgE antibody responses to each IgGi antigen reactive to post-treatment sera from drug-induced resistant (DIR) humans. The dashed and solid lines are the respective cut-offs for IgGl (8239) and IgE (1861) reactivity, calculated as one standard deviation of the mean of the no-DNA control spots on the array. Schistosoma japonicum SEA is included for comparative purposes.

[0026] Figure 5. Gene transcription in the adult and egg stages of S. haematobium for all arrayed proteins inducing significantly different and reactive IgG responses to DIR post-treatment sera. Data was assembled from publicly available RNA-seq databases (Young et al, 2012.). These data were filtered for quality (PHRED score of >30) using Trimmomatic [8] and aligned to the open reading frames of the published gene set [7] using Bowtie (v2.1.0) [9] Normalised levels of gene transcription were calculated using the software package RSEM (v 1.2.11) [10] and reported as the numbers of transcripts per million reads sequenced (TPMs). The TPM value of each gene was log ₂ -transformed and subjected to heat map visualisation using R.

[0027] Figure 6. IgG antibody profiles to arrayed antigens differ in Schistosoma japonicum-infected, self-curing rhesus macaques during the course of infection from exposure to perfusion. Average adjusted signal intensity values depicting IgG antibody responses to each significantly reactive antigen are shown at baseline (0 weeks), 12 weeks post-infection and elimination (20 weeks post-infection). The dashed line is the cut-off for IgG reactivity (3210), calculated as one standard deviation of the mean of the no-DNA control spots on the array. Statistical analysis was performed using student's i test. *p<0.05, **p<0.01, ***p<0.001. [0028] Figure 7. Different disease models of shistosomiasis resistance show common IgG responses to some arrayed antigens. Venn Diagram depicting common IgG reactive proteins between Schistosoma haematobium- infected humans from an endemic area in Africa who acquire drug-induced resistance after praziquantel treatment (DIRs), Schistosoma japonicum-infected self-curing rhesus macaques and Schistosoma mansoni- infected humans from an endemic area of Brazil who are naturally resistant (PRs). *data from Gaze et al, 2014; ^% IgGi response to AY812195 is significantly different between DIRs before and after praziquantel treatment but not between DIRs and CIs post-treatment; AY814977 and Smp_124240 are the respective S. japonicum and S. mansoni orthologues of SNaKip. ^{^} we believe the sequence represented by AY815690 ("myosin-7 [S. japonicum]") has been incorrectly annotated due to its high degree of homology with other parasite orthologues of ribosome-binding protein 1 and lack of blastP hits with any form of myosin.

DETAILED DESCRIPTION

[0029] The present invention is directed to an immunogenic composition, such as a vaccine, that is capable of preventing or treating schistosomiasis. The immunogenic composition suitably comprises one or a plurality of immunogenic proteins that respectively comprise amino acid sequences set forth in one or a plurality of SEQ ID NOS: 1-3. These proteins are the targets of humoral immune responses in "drug- induced resistance" human subjects from an S. haematobin-endemic area in Africa and also in rhesus macaques that had undergone self-cure after experimental S. japonicum infection. Other immunogenic proteins may be chimeras comprising respective amino acid sequences of two or more of SEQ ID NOS: 1-3, such as the embodiments set forth in SEQ ID NOS:4 and 5. It is therefore proposed that the isolated, immunogenic proteins disclosed herein and/or their encoding nucleic acids, may be useful in a safe and efficacious vaccine against various Schistosoma species, including but not limited to S. mansoni, S. japonicum and S. haematobium.

[0030] An aspect of the invention therefore provides an immunogenic composition comprising one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof; one or a plurality of isolated nucleic acids encoding SEQ ID NOS: 1-3, or a fragment or variant thereof; or an antibody which binds one or a plurality of isolated proteins comprising respective amino acid sequences set forth in SEQ ID NOS: 1-3, or a fragment or variant thereof. [0031] A further aspect of the invention provides an isolated, immunogenic chimeric protein comprising at least two of the respective amino acid sequences set forth in SEQ ID NOS: 1-3, or at least two fragments or variants of SEQ ID NOS: 1-3. In particular embodiments, the isolated, immunogenic chimeric protein comprises an amino acid sequence set forth in SEQ ID NO:4 or SEQ ID NO:5, or a chimeric fragment or variant thereof; or an immunogenic composition comprising said isolated immunogenic chimeric protein..

[0032] As used herein, terms such as "schistosome" "schistosomal" and "schistosomiasis" refer to trematode blood flukes of the genus "Schistosoma" and/or diseases or conditions associated with, or caused by, trematodes of this genus. Particular Schistosoma species, include but are not limited to S. mansoni, S. japonicum and S. haematobium. Infective schistosomulae migrate through several tissues and stages to their residence in the veins. Adult worms in humans reside in the mesenteric venules in various locations, which at times seem to be specific for each species. For instance, S. japonicum is more frequently found in the superior mesenteric veins draining the small intestine, and S. mansoni occurs more often in the superior mesenteric veins draining the large intestine. S. haematobium most often occurs in the venous plexus of bladder but it can also be found in the rectal venules. Eggs are deposited in the small venules of the portal and perivesical systems. The eggs progressively move toward the lumen of the intestine (S. mansoni and S. japonicum) and of the bladder and ureters (S. haematobium). Pathology of S. mansoni and S. japonicum schistosomiasis includes Katayama fever, hepatic perisinusoidal egg granulomas, Symmers' pipe stem periportal fibrosis, portal hypertension, and occasional embolic egg granulomas in brain or spinal cord. Pathology of S. haematobium schistosomiasis includes hematuria, scarring, calcification, squamous cell carcinoma, and occasional embolic egg granulomas in brain or spinal cord. Various mammals, such as dogs, cats, rodents, pigs, horse and goats, serve as reservoirs for S. japonicum.

[0033] For the purposes of this invention, by "isolated" is meant material that has been removed from its natural state or otherwise been subjected to human manipulation. Isolated material may be substantially or essentially free from components that normally accompany it in its natural state, or may be manipulated so as to be in an artificial state together with components that normally accompany it in its natural state. Isolated material includes material in native and recombinant form. The term "isolated" also encompasses terms such as "enriched", "purified" and/or "synthetic". The term "synthetic" includes recombinant synthetic and chemical synthetic.

[0034] As used herein a "protein" is an amino acid polymer that may comprise natural and/or non-natural amino acids, D- or L- amino acids and/or amino acid derivatives as are well known in the art. Typically, a peptide is a protein comprising no more than fifty (50) contiguous amino acids and a polypeptide is a protein that comprises more than fifty (50) contiguous amino acids.

[0035] A "chimeric protein" is a protein that comprises at least two different amino acid sequences that are not normally present in the same protein. The at least two different amino acid sequences may be contiguous or non-contiguous in the chimeric protein. In ceryain embodiments, the chimeric protein comprises respective amino acid sequences of two or three of SEQ ID NOS: 1-3, or of fragments of SEQ ID NOS: 1-3.. Particular embodiments of a chimeric protein comprise an amino acid sequence set forth in SEQ ID NO:4 or SEQ ID NO:5.

[0036] As used herein a protein "fragment" may be an epitope, sub-sequence, domain or other portion of a protein. Preferably, the fragment is an immunogenic fragment. Fragments may comprise no more than 6, 12, 15, 18, 20, 25, 30, 40, 50, 60, 78, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600 or 650 contiguous amino acids of a protein.

[0037] As used herein a protein "variant" may be a homolog, ortholog, allelic variant, polymorphic variant, mutant or artificially modified form of a protein disclosed herein. Artificial modification may be performed using recombinant DNA mutagenesis, chemical mutagenesis or by chemical synthesis, although without limitation thereto. In particular embodiments, the protein variant may comprise an amino acid sequence at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequence of any one of SEQ ID NOS: 1-5. Suitably, the variant is immunogenic. Amino acid sequence similarity and identity may be defined with reference to: GAP (Wisconsin Package, Accelerys, San Diego USA) which uses the Needleman and Wunsch algorithm; BLAST (which uses the method of Altschul et al., 1990, J. Mol. Biol. 215 405-410); or FAST A (which uses the method of Pearson & Lipman, 1988, PNAS USA 85 2444-2448), or the Smith- Waterman algorithm (Smith & Waterman, 1981, J. Mol Biol. 147 195-197). Particular versions of BLAST include the TBLASTN program and the psi-Blast algorithm. It is preferable to maximize the number of matches and minimize the number of gaps. Preferably, sequence comparisons are performed over substantially the entire length of the amino acid sequence of any one of SEQ ID NOS: 1-5.

[0038] As generally used herein, "immunogen" and "immunogenic" refer to an ability or property of a composition, protein, fragment, variant, encoding nucleic acid and/or antibody to elicit an immune response to a schistosome upon administration to a mammal.

[0039] By "elicits an immune response" is meant that upon administration to a mammal, an immunogenic protein, epitope or other fragment thereof, a nucleic acid encoding same, or an antibody or antibody fragment, stimulates, provokes, induces, potentiates or otherwise elicits an immune response to a schistosome parasite in the mammal. In a preferred form, the immune response includes an antibody response. A particularly preferred immune response generates specific antibodies at a titer of about >1 to about 1 x 10 ⁶ or greater. Preferably, the titer is from about 1 x 10 ⁴ or 1 x 10 ⁵ to about 1 x 10 ⁶ or more, such as measured by Enzyme Linked Immunosorbent Assay (ELISA) or greater than 1,000 antibody units as defined previously (Malkin et al, 2005a; 2005b). Preferably, the antibody response includes IgG (e.g IgGi) and/or IgE antibodies. In a particularly preferred form, the elicited immune response includes the generation of a T cell response such as during the acquisition of drug induced resistance (DIR) to agents such as Praziquantel (PZQ). The elicited immune response is not necessarily protective, but may reduce, alleviate or decrease one or more symptoms of schistosome infection in the mammal. For example, a decrease in schistosome burden of a least about 30% in a mammal may occur compared to a mammal that has not received the immunogenic composition. With respect to schistosome burden, the level of decrease in parasite egg production and/or worm burden could exceed 40%, as per standards set previously by the World Health Organization.

[0040] In a preferred embodiment, the immune response is a protective immune response. Accordingly, the immunogenic composition is preferably a vaccine. By "vaccine" is meant an immunogenic composition that elicits a protective immune response. By "vaccinate" or "immunize" is meant delivery of an immunogenic composition to a mammal to thereby elicit a protective immune response in the mammal. [0041] Another aspect of the invention therefore provides a method of immunizing a mammal, said method including the step of administering an immunogenic composition disclosed herein to the mammal to thereby immunize the mammal against infection by a schistosome.

[0042] Still yet another aspect of the invention provides a method of preventing or treating a schistosome infection in a mammal, said method including the step of administering an immunogenic composition disclosed herein to the mammal to thereby prevent or treat a schistosome infection in the mammal.

[0043] As used herein, "treating" (or "treat" or "treatment") refers to a therapeutic intervention that ameliorates a sign or symptom of a schistosome infection after it has begun to develop. The term "ameliorating," with reference to a schistosome infection, refers to any observable beneficial effect of the treatment. The beneficial effect can be determined using any methods or standards known to the ordinarily skilled artisan.

[0044] As used herein, "preventing" (or "prevent" or "prevention") refers to a course of action initiated prior to the onset of a symptom, aspect, or characteristic of schistosome infection so as to prevent or reduce the symptom, aspect, or characteristic. It is to be understood that such preventing need not be absolute to be beneficial to a mammalian subject. A "prophylactic" treatment is typically administered to a mammalian subject who is not infected with schistosomes, does not exhibit signs of schistosome infection or exhibits only early signs or symptoms consistent with a schistosome infection. Thus, a vaccine, vaccination or method of immunization is an example of a "prophylactic" composition or treatment.

[0045] In one particular embodiment, immunization, prevention or treatment of schistosomiasis may be facilitated by administration of immunogenic compositions comprising one or more antibodies or antibody fragments that bind and/or are raised against one or more isolated proteins, such as according to SEQ ID NOS: 1-5, or fragments or variants thereof. Such antibodies may confer or invoke passive immunity to schistosomes upon administration to a mammal. Preferred antibodies include IgG (e.g. IgGi) and IgE antibodies. The antibodies may be polyclonal or monoclonal as are well understood in the art and include synthetic antibody constructs such as diabodies and triabodies. Antibody fragments may include Fab, F(ab') ² and single chain fragments such as scFv fragments. It will also be appreciated that antibodies may be recombinant antibodies and include antibodies of non-human origin that have been "humanized" to optimize administration to humans. [0046] In some embodiments, isolated proteins such as comprising an amino acid sequence according to SEQ ID NOS: 1-5, fragments or variants thereof, encoding nucleic acids and/or antibodies thereto may be present in an immunogenic composition and/or administered in combination with a pharmaceutically- acceptable carrier, diluent or excipient.

[0047] Preferably, the pharmaceutically-acceptable carrier, diluent or excipient is suitable for administration to mammals, and more preferably, to humans.

[0048] By "pharmaceutically-acceptable carrier, diluent or excipient" is meant a solid or liquid filler, diluent, binder or encapsulating substance that may be safely used in systemic administration. Depending upon the particular route of administration, a variety of carriers, well known in the art may be used. These carriers may be selected from a group including sugars, starches, cellulose and its derivatives, malt, gelatine, talc, calcium sulfate, vegetable oils, synthetic oils, polyols such as glycerol, alginic acid, phosphate buffered solutions, dextrose, alcohols such as ethanol, zwiterrionic detergents emulsifiers, isotonic saline and salts such as mineral acid salts including hydrochlorides, bromides and sulfates, organic acids such as acetates, propionates and malonates, and pyrogen-free water.

[0049] A useful reference describing pharmaceutically acceptable carriers, diluents and excipients is Remington's Pharmaceutical Sciences (Mack Publishing Co. NJ USA, 1991).

[0050] Any safe route of administration may be employed for providing a subject with compositions comprising one or more isolated proteins such as according to SEQ ID NOS: 1-5, or fragment or variants thereof. For example, oral, rectal, parenteral, sublingual, buccal, intravenous, intra-articular, intra-muscular, intra-dermal, subcutaneous, inhalational, intra-nasal, intraocular, intraperitoneal, intracerebroventricular, transdermal, and other routes may be employed.

[0051] Dosage forms may include tablets, dispersions, suspensions, injections, solutions, syrups, troches, capsules, suppositories, aerosols, transdermal patches, and the like. These dosage forms may also include injecting or implanting controlled releasing devices designed specifically for this purpose or other forms of implants modified to act additionally in this fashion. Controlled release of one or more isolated proteins such as according to SEQ ID NOS: 1-5, or a fragment or variant thereof, may be effected by coating the same, for example, with hydrophobic polymers including acrylic resins, waxes, higher aliphatic alcohols, polylactic and polyglycolic acids, and certain cellulose derivatives such as hydroxypropylmethyl cellulose. In addition, the controlled release may be affected by using other polymer matrices, liposomes and/or microspheres.

[0052] The above compositions may be administered in a manner compatible with the dosage formulation, and in such amount as is pharmaceutically/therapeutically- effective. The dose administered to a subject, in the context of the present invention, should be sufficient to effect a beneficial response (e.g., prevention or treatment of schistosomiasis) in a subject over an appropriate period of time. The quantity of one or more isolated proteins comprising an amino acid sequence according to SEQ ID NOS: 1-5, or a fragment or variant thereof, to be administered may depend on the subject to be treated, inclusive of the age, sex, weight and general health condition thereof, factors that will depend on the judgement of a practitioner of ordinary skill in the art.

[0053] In some embodiments, isolated proteins comprising an amino acid sequence according to SEQ ID NOS: 1-5, fragments or variants thereof, encoding nucleic acids and/or antibodies or antibody fragments, may be present in an immunogenic composition and/or administered in combination with an immunostimulatory agent. An immunostimulatory agent may be an adjuvant, nucleic acid, bacterial toxin, cytokine or other immunoregulatory molecule that enhances the immunogenicity of the isolated proteins comprising an amino acid sequence according to SEQ ID NOS: 1-5, fragments or variants thereof, or encoding nucleic acids, upon administration to a mammalian subject.

[0054] Non-limiting examples of immunostimulatory agents include squalane and squalene (or other oils of plant or animal origin); block copolymers; detergents such as Tween®-80; Quil® A, mineral oils such as Drakeol or Marcol, vegetable oils such as peanut oil; Corynebacterium-derived adjuvants such as Corynebacterium parvum; Propionibacterium-derived adjuvants such as Propionibacterium acne; Mycobacterium bovis (Bacille Calmette and Guerin or BCG); Bordetella pertussis antigens; tetanus toxoid; diphtheria toxoid; surface active substances such as hexadecylamine, octadecylamine, octadecyl amino acid esters, lysolecithin, dimethyldioctadecylammonium bromide, N,N-dicoctadecy1-N', N'bis(2-hydroxyethy1- propanediamine), methoxyhexadecylglycerol, and pluronic polyols; polyamines such as pyran, dextransulfate, poly IC carbopol; peptides such as muramyl dipeptide and derivatives, dimethylglycine, tuftsin; oil emulsions; and mineral gels such as aluminium phosphate, aluminium hydroxide or alum; interleukins such as interleukin 2 and interleukin 12; monokines such as interleukin 1 ; tumour necrosis factor; interferons such as gamma interferon; immunostimulatory DNA such as CpG DNA, adjuvants and adjuvant combinations such as saponin-aluminium hydroxide or Qui1-A aluminium hydroxide; liposomes; ISCOM® and ISCOMATRIX® adjuvant; mycobacterial cell wall extract; synthetic glycopeptides such as muramyl dipeptides or other derivatives; Avridine; Synthetic lipid A and/or Lipid A derivatives; dextran sulfate; DEAE-Dextran alone or with aluminium phosphate; carboxypolymethylene such as Carbopol' EMA; acrylic copolymer emulsions such as Neocryl A640; water in oil emulsifiers such as Montanide ISA 720; poliovirus, vaccinia or animal poxvirus proteins; or mixtures thereof.

[0055] Examples of suitable adjuvants include but are not limited to an aluminum- based adjuvant, CpG and Synthetic Lipid A.

[0056] Other immunological agents administrable together with the isolated nucleic acids and/or nucleic acids disclosed herein may include carrier proteins such as thyroglobulin; albumins such as human serum albumin; toxins, toxoids or any mutant cross-reactive material (CRM) of the toxin from tetanus, diphtheria, pertussis, Pseudomonas, E. coli, Staphylococcus, and Streptococcus; polyamino acids such as poly(lysine: glutamic acid); influenza; Rotavirus VP6, Parvovirus VP1 and VP2; hepatitis B virus core protein; hepatitis B virus recombinant vaccine and the like.

[0057] As hereinbefore described, an isolated nucleic acid may encode one or a plurality of isolated protein comprising an amino acid sequence set forth in any one of SEQ ID NOS: 1-5, or a fragment thereof.

[0058] An aspect of the invention provides an isolated nucleic acid encoding an amino acid sequence set forth in SEQ ID NO: 4 or 5, or a chimeric fragment thereof.

[0059] As used herein a "nucleic acid" may be single- or double-stranded DNA inclusive of cDNA and genomic DNA or RNA inclusive of mRNA, tRNA and inhibitory RNA (e.g interfering RNA such as siRNA), although without limitation thereto.

[0060] In an embodiment, the isolated nucleic acid may be suitable for recombinant expression of an isolated protein disclosed herein. [0061] In an embodiment, the isolated nucleic acid may be administered to a mammalian subject to elicit an immune response to a schistosome in the mammalian subject.

[0062] Suitably, in either embodiment the isolated nucleic acid may be present in a genetic construct. A genetic construct may comprise the isolated nucleic acid operably linked or connected to one or more other nucleotide sequences. Such nucleotide sequences may include regulatory nucleotide sequences such as promoters, enhancers, polyadenylation sequences, splice sites, translation initiation or termination sequences, antibiotic resistances genes and selection marker genes although without limitation thereto. Promoters are typically selected according to a host cell for intended expression of the encoded isolated protein, such as yeast, bacterial, insect, plant or mammalian host cells. Fusion partner or epitope tag sequences may also be added, such as hexahistidine, MBP, GST, haemagglutinin, FLAG and/or c-myc sequences. The genetic construct is suitably manipulated, propagated and/or expressed in a host cell engineered or manipulated to comprise the genetic construct. Such host cells may include yeast, bacterial, insect, plant or mammalian host cells, although without limitation thereto.

[0063] In embodiments relating to expression of recombinant proteins, such methods are well known in the art and the skilled person may refer to standard protocols as for example described in Sambrook et al, MOLECULAR CLONING. A Laboratory Manual (Cold Spring Harbor Press, 1989), in particular Sections 16 and 17; CURRENT PROTOCOLS IN MOLECULAR BIOLOGY Eds. Ausubel et al, (John Wiley & Sons, Inc. 1995-1999), in particular Chapters 10 and 16; and CURRENT PROTOCOLS IN PROTEIN SCIENCE Eds. Coligan et al, (John Wiley & Sons, Inc. 1995-1999), in particular Chapters 1, 5 and 6.

[0064] In embodiments relating to delivery of isolated nucleic acids to a mammalian subject, genetic constructs may be, or comprise, viral vectors that comprise one or more regulatory nucleotide sequences from vaccinia, adenovirus, adenovirus- associated viruses (AAV), retroviruses, lentiviruses, herpes simplex virus or cytomegalovirus, although without limitation thereto. Immunization with DNA or "DNA vaccination" is well known in the art and a non-limiting overview is provided in Ferraro et al, 2011, Clin. Infect. Dis. 53 296.

[0065] It will be appreciated that the compositions and/or methods disclosed herein may be suitable for immunizing, preventing or treating one or more diseases or conditions caused by, or associated with, a schistosome infection. Diseases or conditions caused by, or associated with, S. mansoni and S. japonicum schistosomiasis include Katayama fever, hepatic perisinusoidal egg granulomas, Symmers' pipe stem periportal fibrosis, portal hypertension, and occasional embolic egg granulomas in brain or spinal cord. Diseases or conditions caused by, or associated with, S. haematobium schistosomiasis includes hematuria, scarring, calcification, squamous cell carcinoma, and occasional embolic egg granulomas in brain or spinal cord.

[0066] So that the invention may be readily understood and put into practical effect, the following non-limiting Examples are provided.

EXAMPLES

[0067] Given the extensive similarities in protein-coding gene sequences between the three major human schistosomes (86-92%) (43), as well as the extensive recognition of S. japonicum proteins on our array by sera from S. mansoni infected individuals (20), we reasoned that sera from S. haematobium infected individuals would strongly recognize many of the arrayed S. mansoni and S. japonicum proteins. Moreover, these cross-reactive antigens would potentially form the basis of a pan-schistosome vaccine that protects against all three human species. Leveraging existing protein arrays from our previous study, which contain antigens primarily from the antibody-accessible teguments of the adult fluke and the immunologically-vulnerable schistosomulum stage, we found that DIR individuals and self-curing rhesus macaques make robust antibody responses to a number of tegument-associated proteins, including novel and previously described schistosome vaccine candidates.

[0068] The study participants were residents of a S. haematobium endemic rural village in Murewa in the Mashonaland East Province of Zimbabwe (31°94'E; 17°67'S). The village was selected because health surveys regularly conducted in the region showed little or no infection with soi1-transmitted helminths (STH) and a low S. mansoni prevalence (<2%). Serum samples were provided from a cohort of S. haematobium-infected individuals (n=106) aged 5-14 years who had never been treated with PZQ prior to this study and were free from co-infection with other helminths, Plasmodium and HIV (14, 44). At the start of the study (baseline), subjects who were positive for S. haematobium eggs (at least one egg found in at least one of 3 urine samples, each collected on a separate day) following urinalysis were treated with PZQ by weight (40 mg/kg) and then assessed by urinalysis at 6 weeks to confirm clearance of the infection (no eggs found in any of 3 urine samples, each collected on a separate day). Individuals were followed for 18 months and maintained regular water contact throughout this period. Subjects were assessed for infectivity with S. haematobium at 6 months and at the end of the study. Individuals who were egg- positive at 18 months post treatment (n=32) were deemed CI and those who were egg- negative (n=74) were categorized as DIR (Figure 1). Serum samples were obtained from both 0- and 18-month timepoints.

[0069] For this study we selected a subset of subjects as follows: CI subjects that had the highest post-treatment egg burdens (eggs/10 ml 10-104; n=10) and DIR subjects that had some of the highest egg burdens at baseline (eggs/10 ml 44-743; n=10), reasoning that these individuals represented extremities of the DIR and CI spectrums and therefore would maximise the likelihood of identifying differences in antibody signatures between CIs and DIRs. Subject ages (in years) were as follows: CIs (5, 8, 8, 9, 10, 10, 11, 11, 12, 14), range = 5-14, mean = 9.8, median = 10; DIRs (6, 8, 8, 8, 8, 9, 9, 10, 11, 12) range = 6-12, mean = 8.9, median - 8.5.

[0070] The study used six captive-bred adult male rhesus macaques (Macaca rnulaita: mean age 9.67±0.82 years, mean weight 7.98+0.85 kg) from the Kunming Primate Research Center, CAS. Macaques were group-housed prior to the experiment but then singly after infection for faecal sampling. Cercariae of S. japonicum were shed from patent snails {Oncomelania hupensis) provided by the Jiangsu Institute of Parasitic Diseases (Wuxi, China), collected from the water surface using a bacteriological loop and placed on glass cover slips for infection. Rhesus macaques anaesthetized with ketamine hydrochloride (6 mg/kg body weight, Gutian Pharmaceutical Corporation, Fujian, China) were infected percutaneously with 600 cercariae via the shaved abdominal skin for 30 minutes. Blood was obtained by intravenous sampling prior to infection (week 0) and at 12 and 20 weeks after exposure. Elimination of infection was confirmed at week 20 by assessment of eggs per gram of feces using both the Percoll technique (45) and Kato-Katz method (46).

[0071] Protein microarrays were leveraged from a previous study by us (20) and contained both S. mansoni (n=45) and S. japonicum (n=172) proteins which were either (1) known or predicted to be localized to the tegument and/or (2) expressed in the schistosomulum (41), which is vulnerable to immune attack. Human IgGl and IgE responses to antigens were determined by probing arrays with sera as previously described (20). Macaque antibody responses were determined by probing of arrays with sera as described for human sera with the exception that a goat anti-monkey IgG- biotin (1 :500) (Sigma) secondary antibody was used.

[0072] Array data analysis was conducted using the "group average" method (20), where the mean signal intensity (SI) of the negative control (empty vector) spots for all sera were subtracted from the SI of each protein spot. The following reactivity cutoffs (calculated as one standard deviation above the negative control spots for all groups) were used: human IgGl - 8239; human IgE - 1861; macaque IgG - 3210. Statistical analyses (Student's t test) were conducted with Graphpad Prism 6 to determine significant differences between samples for a given reactive protein.

[0073] The transcription of genes in the adult and egg stages of S. haematobium was assessed for S. haematobium orthologues of all arrayed S. mansoni and S. japonicum proteins that were the targets of significantly different IgG responses between DIR and CI post-treatment sera using publicly available RNA-seq data (43). These data were filtered for quality (PHRED score of >30) using Trimmomatic (47) and aligned to the open reading frames of the published gene set (43) using Bowtie (v2.1.0) (48) Normalised levels of gene transcription were calculated using the software package RSEM (vl .2.11) (49) and reported as the numbers of transcripts per million reads sequenced (TPMs). The TPM value of each gene was log ₂ -transformed and subjected to heat map visualisation using R (v3.1.2; http://www.R-project.org), and utilising the heatmap.plus package.

[0074] To investigate the difference in antibody responses to arrayed antigens of the DIR cohort before and after PZQ treatment (therefore identifying antigens which are putatively exposed by drug therapy), sera from this group at baseline and 18 months after drug therapy were used to probe protein microarrays. IgGl responses were significantly higher in DIRs at 18 months post-treatment compared to baseline for all 24 reactive proteins. Antigens which were the target of the most significantly different (p<0.0001) responses pre- and post-drug treatment included AY810700 (glucose transporter), AY815303 (glutathione-S-transferase) and AY809911 (Ig domain- containing, sensory guidance protein) (Figure 2 A). In contrast, IgGl responses of the CI cohort to reactive proteins before and after PZQ treatment were not significantly different for any protein (data not shown). Additionally, IgE responses in the DIR group were significantly lower at 18 months post-PZQ treatment compared to baseline for the majority (78%) of the 18 reactive antigens (Figure 2B). Arrayed antigens that were the targets of IgE in post-treatment DIRs included AY814430 (calpain), AY812195 (extracellular superoxide dismutase [SOD]) and AY814497 (Na ⁺ /K ⁺ ATPase β subunit - SNaKlp).

[0075] In order to analyse changes in antibody signatures to arrayed antigens related to the acquisition of DIR (thereby identifying proteins which are potential inducers of a protective antibody response), arrays were interrogated with sera from post- treatment CIs and DIRs and probed for IgGl reactivity. IgGl responses were significantly elevated in DIRs compared to CIs at 18 months to 20 of the 24 (83%) reactive proteins. The 3 antigens that were targets of the most significantly different (pO.0001) IgGl responses were AY810792 (butylcholinesterase), AY812951 (mastin) and AY815196 (a homologue of human tetraspanin-33) (Figure 3). Homologues and/or family members of known schistosome vaccine candidates such as calpain (50) (AY814430), a 28 kilodalton glutathione- s-transferase (GST) - S/z28GST (51) (AY815303) and the tetraspanins (TSP)s Sm-TSP-1 and Sm-TSP-2 (33, 52) (AY815196) were also identified. Table 1 lists all of the antigens depicted in Figure 3 along with their S. haematobium orthologues as we reasoned that these were probably the native parasite antigens that our DIR and CI sera were targeting during the course of S. haematobium infection. Of the 20 antigens that were targets of significantly elevated IgGl responses in post-treatment DIRs compared to CIs, only 7 (35%) were targets of IgE responses that were deemed to be above the reactivity cutoff (Figure 4).

[0076] The transcription of genes in the adult and egg stages of S. haematobium was assessed for orthologues of all 20 arrayed S. mansoni and S. japonicum proteins that were the target of significantly different DIR IgG responses post-treatment using publicly available RNA-seq data. We did not find any significant difference in the level of transcription between life stages for a given protein. MS3_02176 (the gene encoding microsomal GST-3) was expressed most highly and relatively constitutively in all developmental stages examined (Figure 5).

[0077] To investigate IgG responses of rhesus macaques to arrayed proteins during the course of a self-curing infection, protein arrays were probed with sera taken at week 0 (primary infection), week 12 and week 20 (after parasite elimination). Antibody responses to all (8 proteins - Table 1) but one reactive protein (AY812195 - extracellular SOD) were significantly elevated between 0-12 weeks post-infection (p.L), with the 3 most robust and highly significant responses being aimed at proteins of unknown function (AY815838 and AY812161) and a MARVEL domain- containing lipid-raft associated protein (AY815056). The IgG reactivity of only one protein (AY812195 - extracellular SOD) was elevated at 20 weeks compared to 12 weeks p.i. (Figure 6 and Table 1).

[0078] We searched for reactive proteins common to DIR human subjects, S. japonicum-infected self-curing rhesus macaques (both described herein) and humans living in an S. mansoni-endemic area of Brazil who, unlike DIRs, have never been treated with PZQ but are putatively resistant to infection (20). Three reactive proteins were common targets of "protective" antibody responses in the DIR and macaque models: a MARVEL domain-containing lipid-raft associated protein; a glucose transporter (SGTP1); and an extracellular SOD (although the IgG response to this protein was not significantly elevated between DIRs and CIs after PZQ treatment). Two reactive antigens were commonly recognised by both DIRs and PRs: ribosome- binding protein 1 and the beta subunit of Na ⁺ /K ⁺ ATPase (SNaKip) (Figure 7).

[0079] The benefit of this approach is the identification of proteins that are cross- reactive between S. haematobium, S. japonicum and S. mansoni, a desirable feature of a vaccine antigen if it is to be protective against all medically important schistosome species.

[0080] The critical role that antibodies play in resistance to schistosomiasis has been well established in animal models by numerous passive transfer studies (eg: (53, 54), and there is evidence that some mechanisms of protective immunity in humans are antibody-mediated, both in individuals naturally resistant to schistosomiasis (20) and those who acquire resistance after PZQ therapy (25). Herein we describe the antibody reactivity profiling of a schistosome protein array with sera from S. haematobium- exposed DIR and CI individuals and rhesus macaques self-cured of a S. japonicum experimental infection (34) in an effort to identify schistosome antigens that might be the targets of resistant human and non-human primate hosts. We previously utilized this protein microarray to define the antibody signatures of individuals that are either naturally resistant to or chronically infected with S. mansoni in a schistosomiasis endemic area of Brazil (20). We restricted our antibody isotype analyses to IgGl and IgE. IgGl is one of the main drivers of the protective humoral response to schistosomiasis (23, 24), an observation supported by studies showing that key tegument vaccine antigens like Smp80 (calpain), Sm-TSP-2 and Sm29 are the targets of these responses in schistosome-resistant individuals (32, 33, 55). IgE is thought to be critical in resistance to schistosomiasis, including the DIR process (25, 56, 57), but caution is warranted in development of anti-helminth vaccines that drive IgE responses due to potential anaphylactic responses in individuals who are pre- sensitized from chronic helminth infection/exposure (58).

[0081] Significantly elevated IgGi responses were detected to 24 antigens in DIR subjects 18 months after therapy compared to pre-treatment responses. In stark contrast, we did not detect elevated IgGl responses to any proteins in CI subjects at 18 months post-treatment compared to pre-treatment levels. None of these antigens were recognised in a previous study by us in which pooled sera from S. haematobium- exposed individuals before and after PZQ treatment were used to probe 2D gels containing S. haematobium SWAP (27), likely because the majority of proteins on the array are membrane-associated tegument proteins and might not be well represented in SWAP due to the very mild solubilizing nature (Tris) of the preparation.

[0082] It is noteworthy that IgGi reactivity to a further 105 (48%) arrayed antigens was significantly higher in post- compared to pre-treatment DIRs but signal intensities were below the cut-off, so the proteins were deemed non-reactive. This decreased level of reactivity possibly reflects the heterogeneity of the antigen-antibody interaction, i.e. antibodies to S. haematobium proteins are reacting with a protein array containing S. mansoni and S. japonicum antigens. Indeed, significant differences in antibody recognition patterns were observed when using sera from S. haematobium-exposed people to probe crude antigen preparations from the closely related Schistosoma bovis, and vice versa (59). Moreover, sequence variation in the epitopes of Sh28GST, and its homologues from S. mansoni and S. bovis significantly altered the immune response generated by the host (60).

[0083] Twenty reactive arrayed antigens were the targets of significantly greater IgGi responses in DIRs compared to CIs post-treatment. A further 72 (33%) proteins were the target of significantly different IgGl recognition profiles between DIRs and CIs but were below the reactivity cut-off. We hypothesize that at least some of these IgGi- reactive proteins are major targets of protective immunity, engendering resistance to schistosomiasis through an antibody-mediated neutralization of the cognate antigen, the role of which is essential to the survival of the parasite within the host (eg: nutrient acquisition, immune evasion) such that disruption of its function results in worm impairment. Indeed, some of these antigens are protective in animal challenge models of schistosomiasis; for example, vaccination with and the Ca ²⁺ -activated protease, calpain (AY814430), induces 64% in baboons (50). Sh28GST (a homologue of the arrayed immunoreactive protein AY815303) is a multi-functional enzyme present in the tegument and sub-tegument of adult (61) and larval (62) schistosomes and the current focus of vaccine trials in humans (51). Its exact function is unknown (studies suggest it may aid in immune evasion by the parasite through its role in fatty acid metabolism and prostaglandin D2 synthesis (63)) but vaccine efficacy has been attributed to the induction of antibodies that neutralize enzyme activity (64). Other extracellular enzymes were prominent amongst the IgGl -reactive proteins, including proteases (calpain, mastin), esterases and superoxide dismutase, so it is tempting to speculate that antibodies to these enzymes neutralize key physiological processes (65, 66), and this now warrants further investigation. Members of the TSP family in schistosomes (Sm-TSP-l and Sm-TSP-2) are four-transmembrane domain proteins located within the tegument of larval and adult worms that have functions in membrane biogenesis (67). TSP-based vaccines have shown to be efficacious against schistosomiasis with Sm-TSP-l and Sm-TSP-2 (33, 52) and 5 23 (68, 69) conferring protection in animal challenge models.

[0084] Other significant IgGi responses were aimed at tegument-associated proteins that play fundamental roles in parasitism. Surface-associated acetylcholinesterase (AChE) (AY810792) has been implicated in the regulation of glucose scavenging from host blood (70) and anti-AChE antibodies facilitate complement-mediated killing of larval schistosomes (71). Genes encoding the glucose transporter SGTP1 (AY810700), Na ⁺ /K ⁺ ATPase subunit SNaKlp (AY814977) and ectonucleotide pyrophosphatase/phosphodiesterase Sm PP-5 (AY814261) have all been functionally silenced within schistosomes using RNAi (72-74), resulting in impairment of the worm's ability to establish infection in the host and highlighting their importance to parasite survival.

[0085] Significantly IgGi-reactive proteins whose therapeutic potential has not yet been examined include mastin (AY812951) and a MARVEL domain-containing lipid- raft associated protein (AY815056). Mastin is a trypsin-like serine protease and, in schistosomes, proteases of this class are known as cercarial elastases (CEs) for their role in skin degradation to facilitate penetration of the free-living cercaria into the definitive host (75). Mastin, however, differs in structural homology to CEs and has been assigned to a group of "non-CE" serine proteases (76). The five members of this group are yet to be functionally characterised in terms of their roles in parasitism, but mastin is unique in that it is highly upregulated in the intra-mammalian schistosomula and adult stages (60% and 150% relative to the constitutively-expressed smcoxl, respectively (76)) compared to the free-living stages of the parasite (77, 78), alluding to a fundamental parasitic function. MARVEL domains have a four-transmembrane helix architecture and proteins containing these motifs associate with membrane micro-domains and have been implicated in membrane biogenesis (79). In a pathogenesis context, the MARVEL domain-containing protein Nee 102 regulates actin organization and invasive growth of Candida albicans, with Nee 102 deletion mutants showing decreased virulence in mice (80). Antigens such as mastin and the MARVEL domain protein are attractive vaccine candidates for the reasons described herein as well as the successful use of proteases (81-83) and membrane structural proteins as anti-helminth vaccines (eg: (33, 69, 84, 85)).

[0086] A group of ribosome-associated proteins were also the targets of significantly higher IgGi responses in DIRs compared to CIs post-treatment and included ribosome-binding protein 1. Ribosome-associated proteins have received attention in the field of parasite immunology because of their classification as "patho-antigens" - conserved intracellular molecules capable of inducing an immunopathological response (86). Patho-antigens such as acidic ribosomal protein P0 conferred protection as vaccines against the intracellular parasites Leishmania major (86) and Plasmodium yoelii (87) in mouse challenge models of infection, and antibodies to P. falciparum P0 have been detected in individuals who are immune to malaria (88). The roles of these antigens, such as ribosome-binding protein 1, in the induction of anti- schistosome immunity is unclear, but it is possible that these intracellular molecules are stimulating host immune effectors through exo some-mediated pathways (recently identified in related helminths (89, 90)). It should also be noted that ribosome-binding protein 1 was one of the two antigens recognised by both S. mansoni-exposed PR subjects in Brazil and S. haematobium-exposed DIR subjects in Africa (Figure 6), possibly highlighting a common role in different mechanisms of schistosomiasis resistance.

[0087] IgE responses to arrayed antigens were, for the most part, significantly weaker in post-therapy DIRs compared to pre-treatment responses, which appears to be in contrast to the positive association between IgE levels and the process of acquiring DIR status (25, 56). This could be likely for 2 reasons: (1) these earlier studies on DIR employed soluble antigen preparations to detect IgE responses, whereas the majority of arrayed proteins are membrane-associated and therefore would not have been present in buffer-soluble parasite extracts or (2) the DIR cohort, being egg negative, do not receive the IgE-inducing stimulus of egg antigens (91). The latter explanation may be supported by the case of extracellular SOD (AY812195); the IgE response to this protein was significantly lower in egg-negative, post-treatment DIRs (Figure 2B) but significantly higher in egg-positive, post-treatment CIs (data not shown). Indeed, a recent study describing the prediction of IgE-binding antigens in S. mansoni-infected individuals reported no significant change in the IgE response to extracellular SOD before and 5 weeks after PZQ treatment (92), which lends support to the observation that the waning IgE response to some antigens in DIRs might be due to the reduced amount of IgE-inducing stimulus received by this cohort. Less than half of the antigens that were significantly reactive for DIR post-treatment IgGl compared to pre-treatment levels were reactive (above the cut-off) for IgE responses.

[0088] IgE poses somewhat of a conundrum for helminth vaccinologists due to its clear association with naturally acquired protection (22, 57), but the accompanying risk of vaccinating people with a recombinant protein that is the target of pre-existing IgE responses poses the risk of inducing atopy (58), or potentially anaphylaxis. Instead of excluding potentially protective IgGi -inducing antigens that are the targets of parasite-derived IgE in exposed individuals from further vaccine development, we propose that the molecules be assessed for allergenicity through the use of basophil activation studies, given that the induction of IgE and clinical manifestation of allergy are not mutually inclusive events (93). Another strategy aimed at minimising potential allergenicity of helminth proteins involves their fusion to Fey, thereby directing the chimeric protein to the negative signalling receptor FcγRIIb expressed on pro-allergic cells (94).

[0089] The IgGi response in S. japonicum-infected self-curing macaques to the majority of reactive antigens was significantly higher at 12 weeks p.i. (around the time that worm death starts to occur (34)) compared to week 0. Proteins that were the target of these antibodies included a protein with weak sequence homology to a bacterial hydrolase (AY815838), extracellular SOD (AY812195) and the previously discussed glucose transport and MARVEL domain-containing proteins. Extracellular SOD is thought to facilitate the parasite' s evasion of the immune response by neutralizing the effects of reactive oxygen and nitrogen species and has proven efficacious in murine vaccine trials (95). Moreover, both hydrolases and anti-oxidant enzymes were suggested to be the targets of IgG-mediated worm elimination in a previously established macaque self-cure model of schistosomiasis (34).

[0090] Given the cognate recognition of antigen by both B and helper T cells in the immune response, we hypothesise that the best antigens for a recombinant protein vaccine are those that elicit responses by both antibodies and T cells during the acquisition of DIR. The antigens described herein should now be subjected to further refinement by assessing their ability to drive T cell proliferation ex vivo. T cell profiling of B cell antigens has been conducted for the vaccinia virus (discovered using protein array profiling) where plasmids encoding arrayed proteins were expressed as inclusion bodies and screened for T cell reactivity in a high-throughput format (96).

[0091] Throughout this specification, the aim has been to describe the preferred embodiments of the invention without limiting the invention to any one embodiment or specific collection of features. Various changes and modifications may be made to the embodiments described and illustrated herein without departing from the broad spirit and scope of the invention.

[0092] All computer programs, algorithms, patent and scientific literature referred to herein is incorporated herein by reference in their entirety.

REFERENCES

1. van der Werf MJ, de Vlas S J, Brooker S, Looman CWN, Nagelkerke NJD, et al. (2003) Quantification of clinical morbidity associated with schistosome infection in sub-Saharan Africa. Acta Trop 86: 125-139.

2. King CH, Dickman K, Tisch DJ (2005) Reassessment of the cost of chronic helmintic infection: a meta-analysis of disability-related outcomes in endemic schistosomiasis. Lancet 365: 1561-1569.

3. King CH, Dangerfield-Cha M (2008) The unacknowledged impact of chronic schistosomiasis. Chronic Illn 4: 65-79.

4. Steinmann P, Keiser J, Bos R, Tanner M, Utzinger J (2006) Schistosomiasis and water resources development: systematic review, meta-analysis, and estimates of people at risk. Lancet Infect Dis 6: 411-425.

5. Gray DJ, McManus DP, Li Y, Williams GM, Bergquist R, Ross AG (2010) Schistosomiasis elimination: lessons from the past guide the future. Lancet Infect Dis 10: 733-736.

6. Fallon PG, Doenhoff MJ (1994) Drug-resistant schistosomiasis: resistance to praziquantel and oxamniquine induced in Schistosoma mansoni in mice is drug specific. Am J Trop Med Hyg 51 : 83-88.

7. Wang W, Wang L, Liang Y-S (2012) Susceptibility or resistance of praziquantel in human schistosomiasis: a review. Parasitol Res 11 1 : 1871-1877.

8. Fenwick A (2006) Waterborne infectious diseases—could they be consigned to history? Science 313 : 1077-1081.

9. Clements ACA, Bosque-Oliva E, Sacko M, Landoure A, Dembele R, et al. (2009) A comparative study of the spatial distribution of schistosomiasis in Mali in 1984- 1989 and 2004-2006. PLoS Negl Trop Dis 3 : e431.

10. Woolhouse ME (1998) Patterns in parasite epidemiology: the peak shift. Parasitol Today 14: 428-434.

11. Black CL, Mwinzi PNM, Muok EMO, Abudho B, Fitzsimmons CM, et al. (2010) Influence of exposure history on the immunology and development of resistance to human Schistosomiasis mansoni. PLoS Negl Trop Dis 4: e637. 12. Skelly PJ, Alan Wilson R (2006) Making sense of the schistosome surface. Adv Parasitol 63 : 185-284.

13. Nuttall PA, Trimnell AR, Kazimirova M, Labuda M (2006) Exposed and concealed antigens as vaccine targets for controlling ticks and tick-borne diseases. Parasite Immunol 28: 155-163.

14. Knox DP (2000) Development of vaccines against gastrointestinal nematodes. Parasitology 120 Suppl: S43-S61.

15. Johnson KS, Harrison GB, Lightowlers MW, O'Hoy KL, Cougle WG, et al. (1989) Vaccination against ovine cysticercosis using a defined recombinant antigen. Nature 338: 585-587.

16. Lightowlers MW, Lawrence SB, Gauci CG, Young J, Ralston MJ, et al. (1996) Vaccination against hydatidosis using a defined recombinant antigen. Parasite Immunol 18: 457-462.

17. Riveau G, Deplanque D, Remoue F, Schacht A-M, Vodougnon H, et al. (2012) Safety and immunogenicity of rSh28GST antigen in humans: phase 1 randomized clinical study of a vaccine candidate against urinary schistosomiasis. PLoS Negl Trop Dis 6: el704.

18. Tendler M, Simpson AJG (2008) The biotechnology- value chain: development of Sml4 as a schistosomiasis vaccine. Acta Trop 108: 263-266.

19. Hotez PJ, Bethony JM, Diemert DJ, Pearson M, Loukas A (2010) Developing vaccines to combat hookworm infection and intestinal schistosomiasis. Nature Rev Microbiol 8: 814-826.

20. McManus DP, Loukas A (2008) Current status of vaccines for schistosomiasis. Clin Microbiol Rev 21 : 225-242.

21. Loukas A, Gaze S, Mulvenna JP, Gasser RB, Brindley PJ, et al. (2011) Vaccinomics for the major blood feeding helminths of humans. OMICS 15: 567-577.

22. Martins VP, Pinheiro CS, Figueiredo BCP, Assis NRG, Morais SB, et al. (2012) Vaccination with enzymatically cleaved GPI-anchored proteins from Schistosoma mansoni induces protection against challenge infection. Clin Dev Immunol 2012: 962538.

23. Castro-Borges W, Dowle A, Curwen RS, Thomas-Oates J, Wilson RA (2011) Enzymatic shaving of the tegument surface of live schistosomes for proteomic analysis: a rational approach to select vaccine candidates. PLoS Negl Trop Dis 5: e993. 24. Cardoso FC, Macedo GC, Gava E, Kitten GT, Mati VL, et al. (2008) Schistosoma mansoni tegument protein Sm29 is able to induce a Th1-type of immune response and protection against parasite infection. PLoS Negl Trop Dis 2: e308.

25. Ahmad G, Zhang W, Torben W, Ahrorov A, Damian RT, et al. (2011) Preclinical prophylactic efficacy testing of Sm-p80-based vaccine in a nonhuman primate model of Schistosoma mansoni infection and immunoglobulin G and E responses to Sm-p80 in human serum samples from an area where schistosomiasis is endemic. J Infect Dis 204: 1437-1449.

26. Tran MH, Pearson MS, Bethony JM, Smyth DJ, Jones MK, et al. (2006) Tetraspanins on the surface of Schistosoma mansoni are protective antigens against schistosomiasis. Nature Med 12: 835-840.

27. Young ND, Jex AR, Li B, Liu S, Yang L, et al. (2012) Whole-genome sequence of Schistosoma haematobium. Nature Genet 44: 221-225.

28. Schistosoma japonicum Genome Sequencing and Functional Analysis Consortium (2009) The Schistosoma japonicum genome reveals features of host-parasite interplay. Nature 460: 345-351.

29. Berriman M, Haas BJ, LoVerde PT, Wilson RA, Dillon GP, et al. (2009) The genome of the blood fluke Schistosoma mansoni. Nature 460: 352-358.

30. Braschi S, Wilson RA (2006) Proteins exposed at the adult schistosome surface revealed by biotinylation. Mol Cell Proteomics 5: 347-356.

31. Mulvenna J, Moertel L, Jones MK, Nawaratna S, Lovas EM, et al. (2010) Exposed proteins of the Schistosoma japonicum tegument. Int J Parasitol 40: 543-554.

32. Gazzinelli A, Bethony J, Fraga LA, LoVerde PT, Correa-Oliveira R, Kloos H (2001) Exposure to Schistosoma mansoni infection in a rural area of Brazil. I: water contact. Trop Med Int Health 6: 126-135.

33. Bethony J, Williams JT, Kloos H, Blangero J, Alves-Fraga L, et al. (2001) Exposure to Schistosoma mansoni infection in a rural area in Brazil. II: household risk factors. Trop Med Int Health 6: 136-145.

34. Bethony J, Williams JT, Brooker S, Gazzinelli A, Gazzinelli MF, et al. (2004) Exposure to Schistosoma mansoni infection in a rural area in Brazil. Part III: household aggregation of water-contact behaviour. Trop Med Int Health 9: 381-389.

35. Viana IR, Sher A, Carvalho OS, Massara CL, Eloi-Santos SM, et al. (1994) Interferon-gamma production by peripheral blood mononuclear cells from residents of an area endemic for Schistosoma mansoni. Trans R Soc Trop Med Hyg 88: 466-470. 36. Caldas IR, Correa-Oliveira R, Colosimo E, Carvalho OS, Massara CL, et al. (2000) Susceptibility and resistance to Schistosoma mansoni reinfection: parallel cellular and isotypic immunologic assessment. Am J Trop Med Hyg 62: 57-64.

37. Bahia-Oliveira LM, Simpson AJ, Alves-Oliveira LF, Carvalho-Queiroz C, Silveira AM, et al. (1996) Evidence that cellular immune responses to soluble and membrane associated antigens are independently regulated during human schistosomiasis mansoni. Parasite Immunol 18: 53-63.

38. Driguez P, Doolan DL, Loukas A, Feigner PL, McManus DP (2010) Schistosomiasis vaccine discovery using immunomics. Parasit Vectors 3 : 4.

39. Diemert DJ, Pinto AG, Freire J, Jariwala A, Santiago H, et al. (2012) Generalized urticaria induced by the Na-ASP-2 hookworm vaccine: implications for the development of vaccines against helminths. J Allergy Clin Immunol 130: 169-76. e6.

40. Hartigan JA, Wong, MA (1979) Algorithm AS 136: a k-means clustering algorithm. J Royal Stat Soc Series C (Appl Stats) 28: 100-108.

41. Defays D (1977) An efficient algorithm for a complete link method. The Computer Journal 20: 364-366.

42. De Groot AS (2009) Exploring the immunome: A brave new world for human vaccine development. Hum Vaccin 5: 790-793.

43. Vigil A, Davies DH, Feigner PL (2010) Defining the humoral immune response to infectious agents using high-density protein microarrays. Future Microbiol 5: 241- 251.

44. Davies DH, Molina DM, Wrammert J, Miller J, Hirst S, et al. (2007) Proteome- wide analysis of the serological response to vaccinia and smallpox. Proteomics 7: 1678-1686.

45. Harrop R, Jennings N, Mountford AP, Coulson PS, Wilson RA (2000) Characterization, cloning and immunogenicity of antigens released by transforming cercariae of Schistosoma mansoni. Parasitology 121 : 385-394.

46. Moloney NA, Webbe G (1990) Antibody is responsible for the passive transfer of immunity to mice from rabbits, rats or mice vaccinated with attenuated Schistosoma japonicum cercariae. Parasitology 100: 235-239.

47. Cioli D, Blum K, Ruppel A (1978) Schistosoma mansoni: relationship between parasite age and time of spontaneous elimination from the rat. Exp Parasitol 45: 74- 80. 48. Cetre C, Pierrot C, Cocude C, Lafitte S, Capron A, et al (1999) Profiles of Thl and Th2 cytokines after primary and secondary infection by Schistosoma mansoni in the semipermissive rat host. Infect Immun 67: 2713-2719.

49. Barker RH, Srivastava BS, Suri P, Goldberg M, Knopf PM (1985) Immunoprecipitation analysis of radiolabelled protein antigens biosynthesized in vitro by S. mansoni. I. Identification of antigens uniquely recognized by protective antibodies. J Immunol 134: 1192-1201.

50. Sepulveda J, Tremblay JM, DeGnore JP, Skelly PJ, Shoemaker CB (2010) Schistosoma mansoni host-exposed surface antigens characterized by sera and recombinant antibodies from schistosomiasis-resistant rats. Int J Parasitol 40: 1407- 1417.

51. Doenhoff MJ, Sabah AA, Fletcher C, Webbe G, Bain J (1987) Evidence for an immune-dependent action of praziquantel on Schistosoma mansoni in mice. Trans R Soc Trop Med Hyg 81 : 947-951.

52. Brindley PJ, Sher A (1987) The chemotherapeutic effect of praziquantel against Schistosoma mansoni is dependent on host antibody response. J Immunol 139: 215- 220.

53. Walter K, Fulford AJC, McBeath R, Joseph S, Jones FM, et al. (2006) Increased human IgE induced by killing Schistosoma mansoni in vivo is associated with pretreatment Th2 cytokine responsiveness to worm antigens. J Immunol 177: 5490- 5498.

54. Joseph S, Jones FM, Walter K, Fulford AJ, Kimani G, et al. (2004) Increases in human T helper 2 cytokine responses to Schistosoma mansoni worm and worm- tegument antigens are induced by treatment with praziquantel. J Infect Dis 190: 835- 842.

55. Hagan P, Blumenthal UJ, Dunn D, Simpson AJ, Wilkins HA (1991) Human IgE, IgG4 and resistance to reinfection with Schistosoma haematobium. Nature 349: 243- 245.

56. Bahia-Oliveira LM, Gazzinelli G, Eloi-Santos SM, Cunha-Melo JR, Alves- Oliveira LF, et al. (1992) Differential cellular reactivity to adult worm antigens of patients with different clinical forms of schistosomiasis mansoni. Trans R Soc Trop Med Hyg 86: 57-61.

57. Correa-Oliveira R, Caldas IR, Gazzinelli G (2000) Natural versus drug-induced resistance in Schistosoma mansoni infection. Parasitol Today 16: 397-399. 58. Viana IR, Correa-Oliveira R, Carvalho ODS, Massara CL, Colosimo E, et al (1995) Comparison of antibody isotype responses to Schistosoma mansoni antigens by infected and putative resistant individuals living in an endemic area. Parasite Immunol 17: 297-304.

59. Bethony JM, Simon G, Diemert DJ, Parenti D, Desrosiers A, et al. (2008) Randomized, placebo-controlled, double-blind trial of the Na-ASP-2 hookworm vaccine in unexposed adults. Vaccine 26: 2408-2417.

60. Gobert GN, Moertel L, Brindley PJ, McManus DP (2009) Developmental gene expression profiles of the human pathogen Schistosoma japonicum. BMC Genomics 10: 128.

61. Gobert GN, Tran MH, Moertel L, Mulvenna J, Jones MK, et al. (2010) Transcriptional changes in Schistosoma mansoni during early schistosomula development and in the presence of erythrocytes. PLoS Negl Trop Dis 4: e600.

62. You H, McManus DP, Hu W, Smout MJ, Brindley PJ, Gobert GN (2013) Transcriptional responses of in vivo praziquantel exposure in schistosomes identifies a functional role for calcium signalling pathway member CamKII. PLoS Pathog 9: el003254.

63. van Balkom BWM, van Gestel RA, Brouwers JFHM, Krijgsveld J, Tielens AGM, et al. (2005) Mass spectrometric analysis of the Schistosoma mansoni tegumental sub- proteome. J Proteome Res 4: 958-966.

64. Taft AS, Yoshino TP (2011) Cloning and functional characterization of two calmodulin genes during larval development in the parasitic flatworm Schistosoma mansoni. J Parasitol 97: 72-81.

65. Katsumata T, Kohno S, Yamaguchi K, Hara K, Aoki Y (1989) Hatching of Schistosoma mansoni eggs is a Ca2+/calmodulin-dependent process. Parasitol Res 76: 90-91.

66. Geary TG, Divo AA, Jensen JB (1986) Effect of calmodulin inhibitors on viability and mitochondrial potential of Plasmodium falciparum in culture. Antimicrob Agents Chemother 30: 785-788.

67. Pearson MS, Pickering DA, McSorley HJ, Bethony JM, Tribolet L, et al. (2012) Enhanced protective efficacy of a chimeric form of the schistosomiasis vaccine antigen Sm-TSP-2. PLoS Negl Trop Dis 6: el 564. 68. Doolan DL, Mu Y, Unal B, Sundaresh S, Hirst S, et al. (2008) Profiling humoral immune responses to P. falciparum infection with protein microarrays. Proteomics 8: 4680-4694.

69. Crompton PD, Kayala MA, Traore B, Kayentao K, Ongoiba A, et al. (2010) A prospective analysis of the Ab response to Plasmodium falciparum before and after a malaria season by protein microarray. Proc Natl Acad Sci U S A 107: 6958-6963.

70. Barry AE, Trieu A, Fowkes FJI, Pablo J, Kalantari-Dehaghi M, et al. (2011) The stability and complexity of antibody responses to the major surface antigen of Plasmodium falciparum are associated with age in a malaria endemic area. Mol Cell Proteomics 10: Ml 11.008326.

71. Trieu A, Kayala MA, Burk C, Molina DM, Freilich DA, et al. (2011) Sterile protective immunity to malaria is associated with a panel of novel P. falciparum antigens. Mol Cell Proteomics 10: Ml 11.007948.

72. Mutapi F, Burchmore R, Mduluza T, Foucher A, Harcus Y, et al. (2005) Praziquantel treatment of individuals exposed to Schistosoma haematobium enhances serological recognition of defined parasite antigens. J Infect Dis 192: 1108-1118.

73. Marcilla A, Trelis M, Cortes A, Sotillo J, Cantalapiedra F, et al. (2012) Extracellular vesicles from parasitic helminths contain specific excretory/secretory proteins and are internalized in intestinal host cells. PLoS One 7: e45974.

74. Kariuki TM, Van Dam GJ, Deelder AM, Farah IO, Yole DS, et al. (2006) Previous or ongoing schistosome infections do not compromise the efficacy of the attenuated cercaria vaccine. Infect Immun 74: 3979-3986.

75. Yole DS, Pemberton R, Reid GD, Wilson RA (1996) Protective immunity to Schistosoma mansoni induced in the olive baboon Papio anubis by the irradiated cercaria vaccine. Parasitology 112: 37-46.

76. Rollinson D (2009) A wake up call for urinary schistosomiasis: reconciling research effort with public health importance. Parasitology 136: 1593-1610.

77. Davies DH, Liang X, Hernandez JE, Randall A, Hirst S, et al. (2005) Profiling the humoral immune response to infection by using proteome microarrays: high- throughput vaccine and diagnostic antigen discovery. Proc Natl Acad Sci U S A 102: 547-552.

78. Sundaresh S, Doolan DL, Hirst S, Mu Y, Unal B, et al. (2006) Identification of humoral immune responses in protein microarrays using DNA microarray data analysis techniques. Bioinformatics 22: 1760-1766. 79. Brock G, Pihur V, Datta S, Datta S (2008) clValid, an R package for cluster validation. J Stat Software 25(4).

80. RDC Team (2011) R: a language and environment for statistical computing, reference index version 2.13.0. Vienna: R Foundation for Statistical Computing.

81. Protasio AV, Tsai IJ, Babbage A, Nichol S, Hunt M, et al. (2012) A systematically improved high quality genome and transcriptome of the human blood fluke Schistosoma mansoni. PLoS Negl Trop Dis 6: el455.

SEQUENCE LISTING

Schistosoma haematobium AY810792 protein

DQFVIRTNYGAVSGKQKYIHGKNVFQFLGIPFAKPP IGDLRFRFPEPPDPWSNILDA TKPPNTCMQPLPESQEFLDATTRVWLNTTKMSEDCLYLNIWTRANNNYSHDIGDRKH MMLESSAIFGPKGGRPVMVWIHGGSLIRGSSS IEMYNGAYLAGKMNVVVVS IQYRLG PLGFLYLGNDEIPGNQGLMDQVAGLQWVRENIAYFGGNPQQITLFGHSGGVICVALH LI SP I SNHLFQQAILQSGSPLAWWAVESSHTALEKTRLLAQLSGCNAVTDSNGQYSK ELVKCLRSVESETLVINQWHMHLLRNGENSSRTNQLKKLYRNRASHHLLSTAGYYFD VPFKPVVSAPFLPEWPYEVLGSGKLNIRHRIMLGVNKDEGMYHLVQSLRMYFMSPGL WPQMPREFRHDIALMDPLDLLAFYIMDENFLQSVLLQATVFEYQIPSRALGTGSWTS LEVVKALNEVGGDYNIKCPVVEFADFYSRGPNAQVFLYSFEHRTTGLTWPEWTGVMQ GYEAEYIFGAPFNQAFIDHYYNFTLEEKRLSEEIMQFWTNFASTGSPNLNPGEFHTR NKELYWDRYETYSPQSVATRNDDSRKHMVFTLPRSYISKNLRRHYCMFWREQLPMLR ERILYNSACKSRQPTTQHEMTGNINKQS IESFGTLYPNI IKSSELNTTKRNQA (SEQ ID NO: l)

Schistosoma haematobium AY812951 protein

SEYDSNKEYIDGEALNRFI SDTDDDNDHHVHHNHRHHQQDRHHRRDIHKNVGRKTQL KLFPWRIHFNRPNTQEMYVFNTDRYSNWSDWKECLPMECIEIRYRKCLDNSWKTI SP NLIHTTRCI SKYYTEKRTCLNKTQCNEHTGEQI IKNLTNTCGIRKQDNKI IEKILGG KSAEPHSWPWAVRLSVKLPHRRPVTFCGGTLIAPQWILTAAHCVLVENKHIPVGKPV MLADHMKSTIYAHLGDHDRYKKEAAQVDHRVTVAILHPNYHRKLQTDGYDIALLRLS EPVKTSPEIDFACLPTKDLKLTPNSKCYAVGWGSNKGAKIPTFDNIHS ILESLFLPF PSLFTTPFRFGRSESTIWNIKKLEEEESSRELHEVELPIVSIEDCRKHYADISSKVH VCAGARNKDTCAGDSGGGLYCYLEETNRWHVVGVTSFGLARGCGLNPGVYTSTTSHM DWLSKQLATKIF (SEQ ID NO:2)

Schistosoma haematobium AY815196 protein

RESTVHRLEELIKTTFVIQYREIGFEDTTNFMDF IQKELNCCGPKSYLDWTANRYFS CDKSNI SPEACGVPYSCCRQMNDI SVNI INTSCGFGVQKLTTAEANRMVWTTGCVDA LISAIENN (SEQ ID NO:3) Chimeric protein comprising amino acid sequence from AY810792 and AY812951

DQFVIRTNYGAVSGKQKYIHGKNVFQFLGIPFAKPP IGDLRFRFPEPPDPWSNILDATKPPN TCMQPLPESQEFLDATTRVWLNTTKMSEDCLYLNIWTRANNNYSHDIGDRKHMMLESSAI FG PKGGRPVMVWIHGGSLIRGSSS IEMYNGAYLAGKMNWVVS IQYRLGPLGFLYLGNDEIPGN QGLMDQVAGLQWVRENIAYFGGNPQQITLFGHSGGVICVALHLISPISNHLFQQAILQSG SP LAWWAVESSHTALEKTRLLAQLSGCNAVTDSNGQYSKELVKCLRSVESETLVINQWHMHL LR NGENSSRTNQLKKLYRNRASHHLLSTAGYYFDVPFKPVVSAPFLPEWPYEVLGSGKLNIR HR IMLGVNKDEGMYHLVQSLRMYFMSPGLWPQMPREFRHDIALMDPLDLLAFYIMDENFLQS VL LQATVFEYQIPSRALGTGSWTSLEWKALNEVGGDYNIKCPWEFADFYSRGPNAQVFLYSF EHRTTGLTWPEWTGVMQGYEAEYIFGAPFNQAFIDHYYNFTLEEKRLSEEIMQFWTNFAS TG SPNLNPGEFHTRNKELYWDRYETYSPQSVATRNDDSRKHMVFTLPRSYI SKNLRRHYCMFWR EQLPMLRERILYNSACKSRQPTTQHEMTGNINKQSIESFGTLYPNI IKSSELNTTKRNQASE YDSNKEYIDGEALNRFISDTDDDNDHHVHHNHRHHQQDRHHRRDIHKNVGRKTQLKLFPW RI HFNRPNTQEMYVFNTDRYSNWSDWKECLPMECIE IRYRKCLDNSWKTISPNLIHTTRCI SKY YTEKRTCLNKTQCNEHTGEQI IKNLTNTCGIRKQDNKI IEKILGGKSAEPHSWPWAVRLSVK LPHRRPVTFCGGTLIAPQWILTAAHCVLVENKHIPVGKPVMLADHMKSTIYAHLGDHDRY KK EAAQVDHRVTVAILHPNYHRKLQTDGYDIALLRLSEPVKTSPEIDFACLPTKDLKLTPNS KC YAVGWGSNKGAKIPTFDNIHSILESLFLPFPSLFTTPFRFGRSESTIWNIKKLEEEESSR EL HEVELP IVSIEDCRKHYADI SSKVHVCAGARNKDTCAGDSGGGLYCYLEETNRWHVVGVTSF GLARGCGLNPGVYTSTTSHMDWLSKQLATKIF (SEQ ID NO: 4)

Chimeric protein comprising amino acid sequence from AY815196, AY812951 and AY810792

RESTVHRLEELIKTTFVIQYREIGFEDTTNFMDF IQKELNCCGPKSYLDWTANRYFSCDKSN I SPEACGVPYSCCRQMNDISVNI INTSCGFGVQKLTTAEANRMVWTTGCVDALISAIENNSE YDSNKEYIDGEALNRFISDTDDDNDHHVHHNHRHHQQDRHHRRDIHKNVGRKTQLKLFPW RI HFNRPNTQEMYVFNTDRYSNWSDWKECLPMECIE IRYRKCLDNSWKTISPNLIHTTRCI SKY YTEKRTCLNKTQCNEHTGEQI IKNLTNTCGIRKQDNKI IEKILGGKSAEPHSWPWAVRLSVK LPHRRPVTFCGGTLIAPQWILTAAHCVLVENKHIPVGKPVMLADHMKSTIYAHLGDHDRY KK EAAQVDHRVTVAILHPNYHRKLQTDGYDIALLRLSEPVKTSPEIDFACLPTKDLKLTPNS KC YAVGWGSNKGAKIPTFDNIHSILESLFLPFPSLFTTPFRFGRSESTIWNIKKLEEEESSR EL HEVELP IVSIEDCRKHYADI SSKVHVCAGARNKDTCAGDSGGGLYCYLEETNRWHVVGVTSF GLARGCGLNPGVYTSTTSHMDWLSKQLATKIFDQFVIRTNYGAVSGKQKYI HGKNVFQFLGI PFAKPP IGDLRFRFPEPPDPWSNILDATKPPNTCMQPLPESQEFLDATTRVWLNTTKMSEDC LYLNIWTRANNNYSHDIGDRKHMMLESSAIFGPKGGRPVMVWIHGGSLIRGSSSIEMYNG AY LAGKMNWWSIQYRLGPLGFLYLGNDEIPGNQGLMDQVAGLQWVRENIAYFGGNPQQITLF GHSGGVICVALHLI SP ISNHLFQQAILQSGSPLAWWAVESSHTALEKTRLLAQLSGCNAVTD SNGQYSKELVKCLRSVESETLVINQWHMHLLRNGENSSRTNQLKKLYRNRASHHLLSTAG YY FDVPFKPVVSAPFLPEWPYEVLGSGKLNIRHRIMLGVNKDEGMYHLVQSLRMYFMSPGLW PQ MPREFRHDIALMDPLDLLAFYIMDENFLQSVLLQATVFEYQIPSRALGTGSWTSLEWKAL N EVGGDYNIKCPVVEFADFYSRGPNAQVFLYSFEHRTTGLTWPEWTGVMQGYEAEYIFGAP FN QAFIDHYYNFTLEEKRLSEEIMQFWTNFASTGSPNLNPGEFHTRNKELYWDRYETYSPQS VA TRNDDSRKHMVFTLPRSYISKNLRRHYCMFWREQLPMLRERILYNSACKSRQPTTQHEMT GN INKQSIESFGTLYPNI IKSSELNTTKRNQA (SEQ ID NO:5)