Biomarkers And Immunogenic Compositions For Filarial Parasites

Abstract

Disclosed herein are immunogenic compositions for preventing or treating infection with filarial parasites and biomarkers for diagnosing infection with filarial parasites.

Description

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] The present application claims the benefit of U.S. Provisional Patent Application 62/317,243 filed Apr. 1, 2016, the entire contents of which is incorporated by reference herein.

BACKGROUND

[0003] Onchocerciasis (river blindness) is a neglected tropical disease, caused by infection with Onchocerca volvulus, that has been targeted for control and elimination through mass drug administration (MDA) of ivermectin that ultimately interrupts transmission. The ultimate success of MDA for onchocerciasis will largely depend on additional tools (macrofilaricidal drugs, vaccines, sensitive diagnostic biomarkers) that in turn rely on a comprehensive understanding of the biology of O. volvulus and the O. volvulus-human host interaction.

[0004] Because of the genetic similarity between O. volvulus and Dirofilaria immitis, the causative agent of heartworm in dogs, it is expected that the D. immitis orthologs of protective O. volvulus proteins will provide protection in dogs against infection with D. immitis as well. Vaccination of `at risk` dogs is an increasingly important activity as dogs are becoming resistant to ivermectin, the current prophylactic drug for canine heartworm.

SUMMARY

[0005] The present disclosure relates to immunogenic compositions for preventing or treating infection with filarial parasites and biomarkers for diagnosing infection with filarial parasites.

[0006] Thus, disclosed herein are immunogenic compositions for preventing or treating infection with a filarial parasite, wherein the filarial parasite is Onchocerca volvulus, and wherein the immunogenic composition comprises at least one filarial parasite protein having at least 85%, 90%, 95%, or 98% sequence identity to the full length mature protein of OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), or OVOC8227 (SEQ ID NO:30).

[0007] Also disclosed herein are immunogenic compositions for preventing infection with a filarial parasite, wherein the filarial parasite is Dirofilaria immitis, and wherein the immunogenic composition comprises at least one filarial parasite mature protein having at least 85%, 90%, 95%, or 98% sequence identity to the full length of OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), OVOC8227 (SEQ ID NO:30), OVOC9988 (SEQ ID NO:31), or OVOC4230 (SEQ ID NO:32), or an ortholog thereof. In some embodiments, the ortholog comprises a filarial parasite protein having at least 85%, 90%, 95%, or 98% sequence identity to the full length of one of SEQ ID NOs:33-49.

[0008] In some embodiments, an immunogenic composition further comprises an adjuvant. In certain embodiments, the immunogenic composition comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins. In some embodiments, the at least two filarial parasite proteins are present in the immunogenic composition in a mixture. In certain embodiments, the at least two filarial parasite proteins are present in the immunogenic composition as a fusion protein comprising the amino acid sequences of the at least two filarial parasite proteins. In some embodiments, the fusion protein optionally further comprises at least one linker sequence separating the at least two filarial parasite amino acid sequences.

[0009] Also disclosed herein are methods of preventing infection with, or transmission of, O. volvulus, the method comprising administering an immunogenic composition disclosed herein to a subject in need thereof, wherein the immunogenic composition prevents the infection or prevents transmission of the infection to another subject. In some embodiments, the immunogenic composition further includes an adjuvant.

[0010] In some embodiments, the immunogenic composition is administered to a subject at risk of O. volvulus infection, and the administration prevents infection with O. volvulus and/or prevents transmission of O. volvulus. In some embodiments, the subject is a human.

[0011] Also disclosed herein are methods of preventing an infection with D. immitis, the method comprising administering an immunogenic composition disclosed herein to a canine subject in need thereof, wherein the immunogenic composition prevents the infection.

[0012] In some embodiments, the immunogenic composition is administered to a subject at risk of D. immitis infection, and the administration prevents infection with D. immitis.

[0013] Also disclosed herein are methods of detecting infection with O. volvulus, comprising identifying in a specimen from a subject at least one filarial full length mature protein having at least 85%, 90%, 95%, or 98% sequence identity to OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC8422 (SEQ ID NO:14), OVOC6395 (SEQ ID NO:15), or OVOC10384 (SEQ ID NO:13) or an immunoreactive fragment thereof.

[0014] Also disclosed herein are methods of detecting infection with O. volvulus, comprising identifying in the blood of a subject, antibodies to at least one filarial protein having at least 85%, 90%, 95%, or 98% sequence identity to the full length mature protein of OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC10384 (SEQ ID NO:13), OVOC8422 (SEQ ID NO:14), OVOC9988 (SEQ ID NO:31), or OVOC6395 (SEQ ID NO:15), or an immunoreactive fragment thereof.

[0015] In certain embodiments, the immunoreactive fragment is OVOC10469_Pep2 (SEQ ID NO:51), OVOC3261_Pep1 (SEQ ID NO:52), OVOC3261_Pep3 (SEQ ID NO:53), OVOC10469_Pep1 (SEQ ID NO:54), OVOC10469_Pep3 (SEQ ID NO:55), OVOC3261_Pep2 (SEQ ID NO:56), OVOC5127_Pep1 (SEQ ID NO:57), OVOC5127_Pep2 (SEQ ID NO:58), OVOC5127_Pep4, (SEQ ID NO:59), OVOC5127_Pep5 (SEQ ID NO:60), or OVOC5127_PepX (SEQ ID NO:61).

[0016] In certain embodiments, the specimen comprises blood, a skin biopsy, or urine.

[0017] In some embodiments, the method comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins. In some embodiments, the at least two filarial parasite proteins comprise Ov16 and OVOC3261. In some embodiments, the at least four filarial parasite proteins comprise Ov16, OVOC3261, OVOC10469, and OVOC5127.

[0018] In some embodiments, the filarial protein or antibody to the filarial protein are detected by a method selected from the group consisting of ELISA, dipstick tests, lateral flow, microfluidic devices, luciferase immunoprecipitation systems, luminex, multiplex-formats, polymerase chain reaction, and microarrays.

BRIEF DESCRIPTION OF THE DRAWINGS

[0019] FIG. 1A-B depicts an overview of transcriptome and proteome of Onchocerca volvulus. FIG. 1A depicts the number of O. volvulus-specific genes (light gray) and proteins (dark bars) identified across the stages by transcriptomic and proteomic analyses. ND denotes samples that were not analyzed. FIG. 1B depicts the number of Wolbachia (wOv) proteins identified across the O. volvulus stages.

[0020] FIG. 2A-B depicts immunoreactivity of disclosed O. volvulus proteins. FIG. 2A depicts a scatter plot of representative proteins with significant IgG4 reactivity in infected individuals, plotted as normalized intensity. FIG. 2B depicts scatter plots of representative proteins with significant IgG1, IgG3 and IgE responses in putatively immune individuals, plotted as ratios to normal sera. The four columns of data for each protein are, from left to right: Putatively Immune; Normal; Infected individuals; Endemic Normal. P values are represented by * (p.ltoreq.0.05), **(p.ltoreq.0.01), *** (p.ltoreq.0.001), ****(p.ltoreq.0.001).

[0021] FIG. 3A-H depicts the sensitivity of the biomarkers for O. volvulus infection. FIG. 3A: Ov16; FIG. 3B: OVOC10469; FIG. 3C: OVOC3261, FIG. 3D: OVOC5127; FIG. 3E: Ov16; FIG. 3F: Ov16+OVOC10469; FIG. 3G: Ov16+OVOC10469+OVOC3261. FIG. 3H depicts the positivity (in black) for each protein based on ROC (receiver operating curves) values. The false negatives for Ov16 (gray, boxed) can be picked up using combination of the proteins (OVOC10469, OVOC3261 and OVOC5127). The white denotes samples not assayed for that protein.

[0022] FIG. 4A-C depicts the effect of immunization with a single vaccine antigen expressed by either Escherichia coli or Pichia pastoris on the development of protective immunity to O. volvulus larvae in mice. FIG. 4A: Ov-103; FIG. 4B: Ov-RAL-2; FIG. 4C: Ov-CPI-2M. Each dot represents larval recovery from an individual animal. Data presented are mean.+-.S.D. Asterisk represents statistical difference in larval recoveries; P.ltoreq.0.05.

[0023] FIG. 5A-B depicts the effect of immunization with fusion antigens on the development of protective immunity to O. volvulus larvae in mice. FIG. 5A depicts Ov-RAL-2/103 fusion protein expressed in E. coli and P. pastoris expressed protein. FIG. 5B depicts Ov-RAL-2/CPI-2M expressed in E. coli. Each dot represents larval recovery from an individual animal. Data presented are mean.+-.S.D. Asterisk represents statistical difference in larval recoveries; P.ltoreq.0.05.

[0024] FIG. 6 depicts the comparative effect of immunization with concurrent injections of O. volvulus Ov-103 (expressed in P. pastoris), Ov-RAL-2 (expressed in E. coli) and Ov-CPI-2M (expressed in E. coli) compared with immunization with the combined fusion antigen Ov-RAL-2/103/CPI-2M (expressed in E. coli). Each dot represents larval recovery from an individual animal. Data presented are mean.+-.S.D. Asterisk represents statistical difference in larval recoveries; P.ltoreq.0.05.

DETAILED DESCRIPTION

[0025] Onchocerciasis, or river blindness, caused by infection with Onchocerca volvulus, is a neglected tropical disease (NTD) that is associated with significant morbidity and disability in the 17 million people estimated to be infected. Infection leads to severe and disfiguring skin disease, lymphadenopathy and visual impairment (including blindness). Onchocerciasis was the first NTD targeted for control in 1974 by the World Health Organization (WHO) and is now one of the six NTDs targeted for elimination. Elimination efforts for O. volvulus are presently aimed by controlling transmission through ivermectin-based mass drug administration (MDA) programs, that have largely eliminated onchocerciasis in the Americas and that have made significant progress toward that goal in some regions of Africa. However, according to a new WHO evaluation, elimination would require an estimated 1.30 billion ivermectin treatments, lasting until 2045, and a recent report has suggested that onchocerciasis cannot solely be eliminated through MDA with ivermectin. Moreover, ivermectin is contraindicated in areas of marked co-endemicity with Loa loa, where the risk of severe adverse events is associated with high levels of circulating Loa loa microfilariae (mf). Furthermore, the potential for ivermectin resistance, the lack of macrofilaricidal activity by ivermectin, and the long timeline (>20 years) for transmission interruption has prompted research into the development of new tools (macrofilaricidal drugs, diagnostics, vaccines, etc.), the basis of which relies on a fundamental understanding of the parasite biology.

[0026] Humans are the only definitive host for O. volvulus. Because there are no existing small animal models for propagating the life cycle of O. volvulus, approaches that require sufficient amounts of stage-specific parasite material have been difficult, as the adult parasites must be obtained surgically from subcutaneous nodules and microfilariae from human skin. Moreover, the larval stages must be obtained from the infected blackflies--a process that to date requires feeding of newly hatched naive black flies on infected microfiladermic humans. Nevertheless, using parasite material from most of the life cycle stages, a comprehensive profile of the stage-specific transcriptomes and proteomes of O. volvulus has been developed. Systematic comparisons across the parasite stages and across related nematodes and `immunomics" has enabled the identification of novel vaccine and diagnostic candidates.

[0027] Systems biology aims at understanding biological processes by integrating various omic's data. Compared to transcriptomic data, attaining complete coverage at the protein level is fraught with technological limitations as well as the dynamic nature of any proteome. Although a difference in transcript (RNA) and protein recovery from the various stages is expected, normalization (RPKM and spectral abundance) provides provisional evidence for relative abundance of any particular gene/protein in a given stage. Using a combination of transcriptomic and proteomic analyses comprehensive stage-specific analyses of O. volvulus was undertaken. This dataset provides an in-depth resource for understanding and analyzing the biological pathways that are critical for the development of the various stages of the parasite in the vector and human hosts, host-O. volvulus interaction, and for the identification of novel biomarkers and targets for interventions.

[0028] Natural immunity against O. volvulus can be acquired in a few individuals within affected populations; these individuals are known as putatively immune and exhibit protective immune response against L3 larvae, suggesting that E/S products released by molting larvae and/or surface proteins of L3 larvae are an important source of protective antigens. The identification of proteins that are highly expressed by the mf and that are specifically recognized by sera from protected individuals who never developed a clinically relevant infection also suggests other suitable vaccine candidates. The identification of O. volvulus-unique proteins that are adult and/or mf stage-specific identified by infected individuals, provided additional novel biomarkers needed for better mapping the prevalence of infection and for post-control surveillance.

[0029] As used herein the term "transcriptome" refers to the full range of messenger RNA, or mRNA, molecules expressed by an organism at a certain time.

[0030] As used herein, the term "proteome" refers to the entire set of proteins expressed by a genome, cell, tissue, or organism at a certain time.

[0031] The life cycle of O. volvulus includes the following stages: nodular microfilariae (NodMF), skin microfilariae (SknMF), embryos (OvEMB), larva L1 (OvL1), larva L2 (OvL2), larva L3 (OvL3), molting L3s (L3 Day 1 and L3 Day 3), larva L4 (OvL4), adult male (OvAM), and adult female (OvAF).

[0032] Analyses of transcript levels or protein abundance for each of the stages identified 363 proteins that were found as core elements by having been present across all somatic stages. Functionally, proteins involved in metabolism, cytoskeletal processes and protein modification comprised more than 50% of these core genes. Proteins shared between OvEMB and OvAF are likely to play a role in embryogenesis. Similarly, proteins identified exclusively during the L3 to L4 transition highlight the machinery required during the developmental molt, and possibly adaptation to the human host environment. Based on C. elegans RNAi data, O. volvulus homologs of C. elegans that exhibit phenotypes of embryonic lethality (EMB), larval arrest (LVA), larval lethal (LVL), molting defective (MLT), or lethal (LET) were observed to be clustered not only in embryos, microfilariae (and thereby adult females), and L3 larval stages but also in adult males. This could either be due to C. elegans being primarily a hermaphroditic organism or to differences between gene families of parasitic and free-living nematodes.

[0033] Similarly, the O. volvulus genome encodes orthologues of the most critical genes essential for molting (based on C. elegans), orthologues that appear to be highly expressed during the in vitro molting process of the L3 larvae. However, it also highlights other proteins, some of which have already been shown to be essential for molting and/or other developmental processes of filarial parasites. For example, embryogenesis and molting in filarial parasites is dependent on the activity of cathepsin L-like cysteine proteases (CPLs).

[0034] Establishment of infection in humans depends on the successful molt from L3 to L4 larvae and subsequent development into adults. During molting, CPLs are stored in the glandular esophagus and their release during molting helps breakdown the old cuticle and drives synthesis of a new cuticle by processing the pro-proteins. Comparative analyses suggest an expansion of CPL-like enzymes in the O. volvulus genome. Significant transcriptional regulation of CPL and CPZ molecules was observed in L2 and L3 larvae compared to other stages. Inferring from Brugia malayi, a related filiarial parasite, these enzymes are probably needed for the L2 to L3 molt in the black fly. Interestingly, the GO gene categories of nucleotide binding (GO:0000166), molecular function (GO:0003674), and phosphoprotein phosphatase activity (GO:0004721), were the most represented categories of differentially expressed genes during L2/L3 and L3/L4 molting. Gene set enrichment analysis (GSEA) identified immunologically important classes of molecules as enriched in L3 larval stages, and a set of extracellular matrix-related genes distinct from the ones overexpressed in adult female worms. The collagens making up the cuticle are regulated by a number of factors, one of which is prolyl-4 hydroxylase, a family that is expanded in the O. volvulus genome, and that is expressed in a stage-specific manner.

[0035] In contrast to those gene families upregulated during development, nuclear hormone receptors (NHR), known to play an important role in other nematode developmental processes, are comparatively less expanded in O. volvulus but still appear to play a role in molting and embryogenesis, as seen in B. malayi. Indeed the O. volvulus ecdysone receptor (EcR, Accession No. OVOC9104) and NHR RXR (Accession No. OVOC2435) are upregulated during the L3 to L4 developmental molt. Furthermore GSEA indicate enrichment of OVOC351 and OVOC353 (other potential NHRs) in adult female worms (p-value <0.0001, FDR <1%). Similarly, the orthologues of the C. elegans NHRs-nhr-6 (OVOC8200), nhr-23 (OVOC464), nhr-25 (OVOC2839), nhr-41 (OVOC4741) and nhr-85 (OVOC827)--known to be involved in molting and metamorphosis, are present in the O. volvulus genome and detected as transcripts or proteins during the in vitro molting of L3 to L4. In addition, NHRs implicated in neural differentiation (OVOC635, OVOC3708) and sex determination (OVOC5276) were upregulated in the molting stages reflecting their probable role in molting, growth, and sex determination.

[0036] Protein OVOC2265 has a rather unique expression profile in the nodular microfilariae (mf) that corresponded with the proteome of embryonic stages. Among the embryo-enriched transcripts and proteins, OVOC11613 (immunodominant antigen or major antigen), and OVOC9384 (Oveg1) have been shown to be related to embryogenesis as well.

[0037] The O. volvulus sequences disclosed here correspond to the WS245 release of the genome by WormBase. The D. immitis sequences disclosed herein correspond to the WPBS1 release of the genome by WormBase. Subsequent genome releases by WormBase may have nucleotide or amino acid revisions.

[0038] I. Biomarkers

[0039] Thus provided herein are biomarkers for infection with a filarial parasite. In certain embodiments, the filarial parasite is O. volvulus.

[0040] In certain embodiments, if the filarial parasite is O. volvulus, the biomarker is a protein having at least 85%, at least 90%, at least 95%, or at least 98% sequence identity to the full length mature protein of OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC10384 (SEQ ID NO:13), OVOC8422 (SEQ ID NO:14), or OVOC6395 (SEQ ID NO:15).

[0041] In addition, the biomarker can also include proteins and peptides sharing a sequence identity or substantial sequence identity to the biomarker proteins provided herein.

[0042] As used herein, "sequence identity" or "identity" in the context of two protein or peptide sequences makes reference to a specified percentage of residues in the two sequences that are the same when aligned for maximum correspondence over a specified comparison window, as measured by sequence comparison algorithms or by visual inspection. When percentage of sequence identity is used in reference to proteins it is recognized that residue positions which are not identical often differ by conservative amino acid substitutions, where amino acid residues are substituted for other amino acid residues with similar chemical properties (e.g., charge or hydrophobicity) and therefore do not change the functional properties of the molecule. When sequences differ in conservative substitutions, the percent sequence identity may be adjusted upwards to correct for the conservative nature of the substitution. Sequences that differ by such conservative substitutions are said to have "sequence similarity" or "similarity." Means for making this adjustment are well known to those of skill in the art.

[0043] The term "substantial identity" in the context of a protein or peptide indicates that a protein or peptide comprises a sequence with at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, or 94%, or even 95%, 96%, 97%, 98%, or 99% sequence identity to the reference sequence over a specified comparison window. In certain embodiments, optimal alignment is conducted using the homology alignment algorithm of Needleman and Wunsch (Needleman and Wunsch, JMB, 48:443, 1970). An indication that two peptide sequences are substantially identical is that one peptide is immunologically reactive with antibodies raised against the second peptide. Thus, a peptide is substantially identical to a second peptide, for example, where the two peptides differ only by a conservative substitution. Thus, also provided herein are proteins and peptides that are substantially identical to the proteins and peptides presented herein.

[0044] In certain embodiments, the term "sequence identity" refers to identity across the entire amino acid sequence of one of SEQ ID NOs:1-66 but can include proteins or peptides which have additional amino acids at the C-terminus or N-terminus of the protein or peptide and which have at least 85%, at least 90%, at least 95%, or at least 98% sequence identity to the portion of the sequence which is the same length as the disclosed sequences.

[0045] Accordingly, some embodiments disclosed herein comprise a method of diagnosing an infection with a filarial parasite comprising: (a) providing a blood sample from at least one subject suspected of having a filarial parasite infection; and (b) contacting the sample with at least one protein selected from OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC8422 (SEQ ID NO:14), OVOC6395 (SEQ ID NO:15) or OVOC10384 (SEQ ID NO:13); wherein if the sample contains specific antibodies which bind to the at least one protein, the subject has an active filarial parasite infection. In certain embodiments, the method includes contacting the sample with at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

[0046] Additionally, some embodiments disclosed herein comprise a method of diagnosing an infection with a filarial parasite comprising: (a) providing a tissue or fluid sample from at least one subject suspected of having an filarial parasite infection; (b) providing a binding agent which binds to at least one filarial parasite-associated protein selected from OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC8422 (SEQ ID NO:14), OVOC6395 (SEQ ID NO:15) OVOC10384 (SEQ ID NO:13); and (c) detecting the proteins, individually and/or in combination, associated with the filarial parasite infection in the subject and contained in the sample; wherein if the sample contains at least one filarial parasite-associated protein, the subject has an active filarial parasite infection. In certain embodiments, the fluid sample is urine, blood, serum, plasma, or a skin biopsy. In some embodiments, the method includes detecting at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

[0047] Also disclosed herein are immunoreactive fragments of filarial parasite proteins which can be used in the methods disclosed herein. The immunoreactive fragments include, but are not limited to, OVOC10469_Pep2 (SEQ ID NO:51), OVOC3261_Pep1 (SEQ ID NO:52), OVOC3261_Pep3 (SEQ ID NO:53), OVOC10469_Pep1 (SEQ ID NO:54), OVOC10469_Pep3 (SEQ ID NO:55), OVOC3261_Pep2 (SEQ ID NO:56), OVOC5127_Pep1 (SEQ ID NO:57), OVOC5127_Pep2 (SEQ ID NO:58), OVOC5127_Pep4, (SEQ ID NO:59), OVOC5127_Pep5 (SEQ ID NO:60), and OVOC5127_PepX (SEQ ID NO:61).

[0048] Ideally, several methods, including ELISA, dipstick tests, lateral flow, microfluidic devices, luciferase immunoprecipitation systems (LIPS), and microarrays can be used to detect filarial parasite-associated biomarkers in patients with filarial parasite infections.

[0049] ELISA is a widely used method for the detection of specific antibodies and proteins in a biological sample. It involves the immobilization of an antibody (primary antibody), or an antigen, to a solid support, such as plastic microplates, and detecting binding of components of a patient sample to the immobilized antibody or antigen, followed by the addition of secondary antibody or antibodies, the latter usually being conjugated to a detectable moiety in order to facilitate measurement.

[0050] Hence, according to some embodiments, the immune affinity procedure may be an ELISA immunoassay selected from the group consisting of direct enzyme-linked immunosorbent assays, indirect enzyme-linked immunosorbent assays, direct sandwich enzyme-linked immunosorbent assays, indirect sandwich enzyme-linked immunosorbent assays, and competitive enzyme-linked immunosorbent assays.

[0051] In one embodiment, detection is effected through capture ELISA. Capture ELISA (also known as "sandwich" ELISA) is a sensitive assay to quantify picogram to microgram quantities of substances such as hormones, cell signaling chemicals, infectious disease antigens and cytokines. This type of ELISA is particularly sought after when the substance to be analyzed may be too dilute to bind to the microtiter plate (such as a protein in a cell culture supernatant) or does not bind well to plastics (such as a small organic molecule). Optimal dilutions for the capture antibody, samples, controls, and detecting antibodies as well as incubation times are determined empirically and may require extensive titration. Ideally, one would use an enzyme-labeled detection antibody. However, if the detection antibody is unlabeled, the secondary antibody should not cross-react with either the coating antibody or the sample. Optimally, the appropriate negative and positive controls should also be included.

[0052] Detection of the biomarkers, or of any fragment or derivative thereof, may be performed using antibodies specific to said biomarkers. These antibodies may be labeled directly or indirectly by a detectable moiety.

[0053] As used herein in the specification, the term "detectable moiety" refers to any element, molecule, or a portion thereof, the presence, absence or level of which may be monitored directly or indirectly. One example includes radioactive isotopes. Other examples include enzymes which can catalyze color or light emitting (luminescence) reactions, fluorophores, and gold or magnetic labels. The detection of the detectable moiety can be direct provided that the detectable moiety is itself detectable (i.e. can be directly visualized or measured), such as, for example, in the case of fluorophores. Alternatively, the detection of the detectable moiety can be indirect. In the latter case, a second moiety that reacts with the detectable moiety, itself being directly detectable is preferably employed. The detectable moiety may be inherent to the antibody. For example, the constant region of an antibody can serve as an indirect detectable moiety to which a secondary antibody having a direct detectable moiety can specifically bind.

[0054] Thus, secondary antibodies are particular suitable means for the detection of the anti-biomarker antibody. This secondary antibody may be itself conjugated to a detectable moiety. One of the ways in which an antibody can be detectably labeled is by linking the same to an enzyme. The enzyme, in turn, when exposed to an appropriate substrate, will react with the substrate in such a manner as to allow its detection, for example, by producing a chemical moiety, which can be detected, for example, by spectrophotometric, fluorometric, or by visual means. Enzymes which may be used to label the antibody include, but are not limited to, horseradish peroxidase, alkaline phosphatase, malate dehydrogenase, staphylococcal nuclease, .delta.-5-steroid isomerase, yeast alcohol dehydrogenase, .alpha.-glycerophosphate dehydrogenase, triose phosphate isomerase, asparaginase, glucose oxidase, beta-galactosidase, ribonuclease, urease, catalase, glucose-6-phosphate dehydrogenase, glucoamylase and acetylcholinesterase, or any other enzyme which can be conjugated to an antibody and its reaction with a substrate, measured or detected.

[0055] The detection may be accomplished by colorimetric methods, which employ a chromogenic substrate for the enzyme. Detection may also be accomplished by visual comparison of the extent of enzymatic reaction of a substrate in comparison with similarly prepared standards.

[0056] The solid support to which the first antibody is bound may be any water-insoluble, water-insuspensible, solid support. Examples of suitable solid support include, but are not limited to, large beads (e.g., of polystyrene), filter paper, slides, chips, test tubes, and microtiter plates. The first antibody may be bound to the solid support by covalent bonds or by adsorption. The advantage in using a solid support is that no centrifugation step is needed for the separation of the solid and liquid phase.

[0057] The solid support mentioned above may include polymers, such as polystyrene, agarose, SEPHAROSE.RTM., cellulose, glass beads and magnetizable particles of cellulose or other polymers. The solid-support can be in the form of large or small beads or particles, tubes, plates, slides, chips or other forms.

[0058] As a solid support, a test tube, the inner walls of a test tube or a microtiter plate are coated with a first antibody, e.g., antibodies specific to a peptide or protein disclosed herein, or of any fragment or derivative thereof.

[0059] In a further embodiment, dipstick assays may be used to detect filarial parasite biomarkers. Dipstick assays use the lateral flow format, wherein capture antibodies are striped or banded onto nitrocellulose membrane and a wicking pad draws the sample up through the dipstick, whereby the filarial parasite biomarkers interact with a filarial parasite biomarker antibody, or combination of antibodies. Other antibodies specific to filarial parasites, or other proteins of interest may be included. Subsequent analysis of enzyme activity and protein quantity may be done using standard methods known to a person skilled in the art, or as discussed above regarding ELISAs.

[0060] In another preferred embodiment, microfluidic devices, which may also be referred to as "lab-on-a-chip" systems, biomedical micro-electro-mechanical systems (bioMEMs), or multicomponent integrated systems, may be used for detecting filarial parasite biomarkers. Such systems miniaturize and compartmentalize processes that allow for detection of filarial parasite biomarkers, and other processes.

[0061] Array-based assays and bead-based assays may be used with microfluidic device. For example, a binding agent can be coupled to beads and the binding reaction between the beads and filarial parasite biomarker can be performed in a microfluidic device. Multiplexing, or detecting more than one filarial parasite biomarker at once, can also be performed using a microfluidic device. Different compartments can comprise different binding agents for different populations of filarial parasite biomarkers, where each population has a different bio-signature.

[0062] In another embodiment, microarrays are used to detect filarial parasite biomarkers. Microarrays are typically small, high throughput chips generally made of a solid support structure, typically glass slides, nitrocellulose, or microtiter plates. Generally, antibodies to specific biomarker are bound to the solid support; however, other molecules, such as, but not limited to other proteins, aptamers, DNA, RNA, sugars or lipids can be bound to the solid surface as well. Detection of the captured biomarker can also be accomplished as discussed above for ELISA detection.

[0063] In another further embodiment, recognition of filarial parasite specific biomarker is achieved through an immune affinity procedure such as Western blot, immuno-precipitation, FACS, biochip array, lateral flow, time resolved fluorometry, ECL procedures, luminex, LIPS, multiplex-immunoassay formats or any procedure based on immune recognition known to one of ordinary skill in the art.

[0064] II. Immunogenic Compositions

[0065] Also provided herein are immunogenic compositions for preventing infection with, or preventing transmission of, a filarial parasite. In certain embodiments, the filarial parasite is O. volvulus. In other embodiments, the filarial parasite is D. immitis. As used herein, "preventing transmission of" refers to the inability of an infected subject, who has been immunized with an immunogenic disclosed herein, to transmit infectious parasites to another subject via an intermediate vector.

[0066] In certain embodiments, the filarial parasite is O. volvulus, and the immunogenic composition comprises as least one protein having at least 85%, at least 90%, at least 95%, or at least 98% sequence identity to the full length mature protein of OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), OVOC8227 (SEQ ID NO:30), OVOC9988 (SEQ ID NO:31), OVOC4230 (SEQ ID NO:32), or an immunogenic fragment thereof, or a nucleic acid encoding the protein. In certain embodiments, the filarial parasite immunogenic composition for preventing infection with, or preventing transmission of, O. volvulus is not OVOC9988 (Ov-RAL-2), OVOC4230 (Ov-103), or OVOC7453 related (Ov-CPI-2M). In some embodiments, the immunogenic composition includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins. The immunogenic composition can comprise the filarial parasite proteins in a mixture or as fusion proteins which include sequences from two or more filarial parasite proteins assembled in a single polypeptide sequence. If multiple filarial proteins are assembled into a fusion protein, one or more linker sequences can be included.

[0067] In other embodiments, the filarial parasite is D. immitis, and the immunogenic composition comprises at least one protein having at least 85%, at least 90%, at least 95%, or at least 98% sequence identity to the full length mature protein of OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), OVOC8227 (SEQ ID NO:30), OVOC9988 (SEQ ID NO:31), or OVOC4230 (SEQ ID NO:32), an ortholog thereof, or a nucleic acid encoding the protein or ortholog. In some embodiments, the ortholog comprises one of the proteins of Table 6, or an immunogenic fragment thereof. In some embodiments, the immunogenic composition includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins. The immunogenic composition can comprise the filarial parasite proteins in a mixture or as fusion proteins which include sequences from two or more filarial parasite proteins assembled in a single polypeptide sequence.

[0068] As used herein, the term "immunogenic composition" refers to a substance which induces a specific immune response against an immunogen (protein) in an individual who is in need of an immune response to the immunogen. As used herein the term "immunogen" refers to any substrate that elicits an immune response in a host. Thus, the disclosed immunogenic compositions comprising filarial parasite proteins are useful for inducing an immune response against a filarial parasite. In certain embodiments, the immune response is a protective immune response. In other embodiments, the immune response is a therapeutic immune response. A non-limiting example of an immunogenic composition is a vaccine.

[0069] In certain embodiments, the immunogenic composition comprises a protein disclosed herein along with additional sequences to enhance immunogenicity.

[0070] In certain embodiments, the immunogenic composition is a fusion protein which includes several filarial parasite proteins. In some embodiments, the immunogenic composition is a mixture of one or more filarial parasite proteins. In some embodiments, the immunogenic composition includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

[0071] Certain amino acid sequences disclosed herein may include a signal sequence. As is understood by a person of skill in the art, expressed filarial proteins useful as immunogenic composition or biomarkers disclosed herein will not include a signal sequence. Thus, in certain embodiments, the amino acid sequences are referred to as "mature" proteins, which are proteins without a signal sequence. In some embodiments, the protein sequence will be recited as less than the entire amino acid sequence disclosed herein to reflect the absence of the signal sequence.

[0072] The disclosed filarial parasite immunogenic compositions and proteins include conservative variants of the proteins and fusion proteins. A conservative variant refers to a peptide or protein that has at least one amino acid substituted by another amino acid, or an amino acid analog, that has at least one property similar to that of the original amino acid from an exemplary reference peptide. Examples of properties include, without limitation, similar size, topography, charge, hydrophobicity, hydrophilicity, lipophilicity, covalent-bonding capacity, hydrogen-bonding capacity, physicochemical property, or the like, or any combination thereof. A conservative substitution can be assessed by a variety of factors, such as, e.g., the physical properties of the amino acid being substituted (Table 1) or how the original amino acid would tolerate a substitution (Table 2). The selections of which amino acid can be substituted for another amino acid in a peptide disclosed herein are known to a person of ordinary skill in the art. A conservative variant can function in substantially the same manner as the exemplary reference peptide, and can be substituted for the exemplary reference peptide in any aspect of the present specification.

TABLE-US-00001 TABLE 1 Amino Acid Properties Property Amino Acids Aliphatic G, A, I, L, M, P, V Aromatic F, H, W, Y C-beta branched I, V, T Hydrophobic C, F, I, L, M, V, W Small polar D, N, P Small non-polar A, C, G, S, T Large polar E, H, K, Q, R, W, Y Large non-polar F, I, L, M, V Charged D, E, H, K, R Uncharged C, S, T Negative D, E Positive H, K, R Acidic D, E Basic K, R Amide N, Q

TABLE-US-00002 TABLE 2 Amino Acid Substitutions Amino Favored Acid Substitution Neutral Substitutions Disfavored substitution A G, S, T C, E, I, K, M, L, P, Q, R, V D, F, H, N, Y, W C F, S, Y, W A, H, I, M, L, T, V D, E, G, K, N, P, Q, R D E, N G, H, K, P, Q, R, S, T A, C, I, L, E D, K, Q A, H, N, P, R, S, T C, F, G, I, L, M, V, W, Y F M, L, W, Y C, I, V A, D, E, G, H, K, N, P, Q, R, S, T G A, S D, K, N, P, Q, R C, E, F, H, I, L, M, T, V, W, Y H N, Y C, D, E, K, Q, R, S, T, W A, F, G, I, L, M, P, V I V, L, M A, C, T, F, Y D, E, G, H, K, N, P, Q, R, S, W K Q, E, R A, D, G, H, M, N, P, S, T C, F, I, L, V, W, Y L F, I, M, V A, C, W, Y D, E, G, H, K, N, P, Q, R, S, T M F, I, L, V A, C, R, Q, K, T, W, Y D, E, G, H, N, P, S N D, H, S E, G, K, Q, R, T A, C, F, I, L, M, P, V, W, Y P -- A, D, E, G, K, Q, R, S, T C, F, H, I, L, M, N, V, W, Y Q E, K, R A, D, G, H, M, N, P, S, T C, F, I, L, V, W, Y R K, Q A, D, E, G, H, M, N, P, S, T C, F, I, L, V, W, Y S A, N, T C, D, E, G, H, K, P, Q, R, T F, I, L, M, V, W, Y T S A, C, D, E, H, I, K, M, N, P, Q, F, G, L, W, Y R, V V I, L, M A, C, F, T, Y D, E, G, H, K, N, P, Q, R, S, W W F, Y H, L, M A, C, D, E, G, I, K, N, P, Q, R, S, T, V Y F, H, W C, I, L, M, V A, D, E, G, K, N, P, Q, R, S, T Matthew J. Betts and Robert, B. Russell, Amino Acid Properties and Consequences of Substitutions, pp. 289-316, In Bioinformatics for Geneticists, (eds Michael R. Barnes, Ian C. Gray, Wiley, 2003).

[0073] A filarial parasite immunogenic composition can also comprise conservative variants to the disclosed proteins or fusion proteins. In aspects of this embodiment, a conservative variant of a filarial parasite protein or fusion protein can be, for example, an amino acid sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, or at least 99% amino acid sequence identity to the filarial parasite protein or fusion protein. In other aspects of this embodiment, a conservative variant of a filarial parasite protein or fusion protein can be, for example, an amino acid sequence having at most 75%, at most 80%, at most 85%, at most 90%, at most 95%, at most 97%, at most 98%, or at most 99% amino acid sequence identity to the filarial parasite protein or fusion protein.

[0074] In other embodiments, a conservative variant of a filarial parasite protein or fusion protein amino acid sequence can have, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or more conservative substitutions, to the amino acid sequence of the filarial parasite protein or fusion protein. In other embodiments, a conservative variant of a filarial parasite protein or fusion protein amino acid sequence can be, for example, an amino acid sequence having at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 15 conservative substitutions to the amino acid sequence of the filarial parasite protein or fusion protein. In yet other embodiments, a conservative variant of a filarial parasite protein or fusion protein amino acid sequence can be, for example, an amino acid sequence having at most 1, at most 2, at most 3, at most 4, at most 5, at most 6, at most 7, at most 8, at most 9, at most 10, at most 11, at most 12, at most 13, at most 14, or at most 15 conservative substitutions to the amino acid sequence of the filarial parasite protein or fusion protein. In further embodiments, a conservative variant of a filarial parasite protein or fusion protein amino acid sequence can be, for example, an amino acid sequence having from 1 to 15, 2 to 15, 3 to 15, 4 to 15, 5 to 15, 6 to 15, 7 to 15, 1 to 12, 2 to 12, 3 to 12, 4 to 12, 5 to 12, 6 to 12, 7 to 12, 1 to 10, 2 to 10, 3 to 10, 4 to 10, 5 to 10, 6 to 10, 7 to 10, 1 to 8, 2 to 8, 3 to 8, 4 to 8, 5 to 8, 6 to 8, 1 to 6, 2 to 6, 3 to 6, 4 to 6, 1 to 4, 2 to 4, or 1 to 3 conservative substitutions to the amino acid sequence of the filarial parasite protein or fusion protein.

[0075] In certain embodiments, the immunogenic compositions further comprise or are administered with an adjuvant. Adjuvants suitable for use in animals include, but are not limited to, Freund's complete or incomplete adjuvants, Sigma Adjuvant System (SAS), and Ribi adjuvants. Adjuvants suitable for use in humans include, but are not limited to, MF59.RTM. (an oil-in-water emulsion adjuvant, Novartis AG); MONTANIDE.RTM. ISA 51 or 720 (a mineral oil-based or metabolizable oil-based adjuvant, SEPPIC); aluminum hydroxide, -phosphate, or -oxide; HAVLOGEN.RTM. (an acrylic acid polymer-based adjuvant, Intervet Inc.); polyacrylic acids; oil-in-water or water-in-oil emulsion based on, for example a mineral oil, such as BAYOL.TM. or MARCOL.TM. (Esso Imperial Oil Limited), or a vegetable oil such as vitamin E acetate; a saponin; CpG oligodeoxynucleotide adjuvants; or a glucagon-like peptide (GLP) adjuvant. However, components with adjuvant activity are widely known and, generally, any adjuvant may be utilized that does not adversely interfere with the efficacy or safety of the immunogenic composition.

[0076] Immunogenic compositions according to the various embodiments disclosed herein can be prepared and/or marketed in the form of a liquid, frozen suspension, or in a lyophilized form. Typically, vaccines and/or immunogenic compositions contain a pharmaceutically acceptable carrier or diluent customarily used for such compositions. Carriers include, but are not limited to, stabilizers, preservatives, and buffers. Suitable stabilizers are, for example SPGA, TWEEN.RTM. compositions (such as are available from A.G. Scientific, Inc.), carbohydrates (such as sorbitol, mannitol, starch, sucrose, dextran, glutamate, or glucose), proteins (such as dried milk serum, albumin, or casein), or degradation products thereof. Examples of suitable buffers include alkali metal phosphates. Suitable preservatives include thimerosal, merthiolate, and gentamicin. Diluents include water, aqueous buffer (such as buffered saline), alcohols, and polyols (such as glycerol).

[0077] Also disclosed herein are methods for inducing an immune response to a filarial parasite using the disclosed proteins. Generally, the vaccine and/or immunogenic composition may be administered subcutaneously, intradermally, submucosally, intranasally, or intramuscularly in an effective amount to prevent infection from the filarial parasite and/or treat an infection from the filarial parasite. An effective amount to prevent infection is an amount of immunizing protein that will induce immunity in the immunized animals against challenge by infective stage larvae or microfilariae such that infection is prevented or the severity is reduced. Immunity is defined herein as the induction of a significant higher level of protection in a subject after immunization compared to an unimmunized group. An effective amount to treat an infection is an amount of immunizing protein that induces an appropriate immune response against filarial parasite such that severity of the infection is reduced.

[0078] Protective immune responses can include humoral immune responses and cellular immune responses. Protection against filarial parasite is believed to be conferred through serum antibodies (humoral immune response) directed to the surface proteins and/or proteins secreted during the early development in the human host, probably through antibody-dependent cellular cytotoxicity (ADCC) and cell-mediated immune responses. Cellular immune responses are useful in protection against filarial parasite infection with CD4+ T cell responses of the Th1, Th2 and/or Th17 type being particularly important. Additionally, the disclosed proteins and/or immunogenic compositions can be administered using immunization schemes known by persons of ordinary skill in the art to induce protective immune responses. These include a single immunization or multiple immunizations in a prime-boost strategy. A boosting immunization can be administered at a time after the initial, prime, immunization that is days, weeks, months, or even years after the prime immunization. In certain embodiments, a boost immunization is administered 2 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, or 6 months or more after the initial prime immunization. Additional multiple boost immunizations can be administered such as weekly, every other week, monthly, every other month, every third month, or more. In other embodiments, the boost immunization can be administered every 3 weeks, every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, or every 12 weeks. In certain embodiments, boosting immunizations can continue until a protective anti-filarial parasite antibody titer is seen in the subject's serum. In certain embodiments, a subject is given one boost immunization, two boost immunizations, three boost immunizations, or four or more boost immunizations, as needed to obtain a protective antibody titer. In other embodiments, the adjuvant in the initial prime immunization and the adjuvant in the boost immunizations are different.

[0079] Further, in various formulations of the proteins and/or immunogenic compositions, suitable excipients, stabilizers, and the like may be added as are known by persons of ordinary skill in the art.

[0080] The disclosed proteins, immunogenic compositions, and methods may be used to prevent filarial parasite infection in a subject susceptible thereto such as, but not limited to, a human, or a domesticated animal.

EXAMPLES

Example 1. Transcriptome and Proteome of Onchocerca volvulus

[0081] Parasite and Serum Samples.

[0082] Parasite materials used for RNAseq and proteomic analyses were collected at the research facility at the Tropical Research Station, Kumba, Cameroon, and in Ecuador and Guatemala. Written informed consent was obtained. In cases of illiteracy, the participant made a thumbprint and a literate witness signed. Institutional Review Board (IRB) approvals were obtained from the National Institutes of Health, the New York Blood Center and the Tropical Research Station, Kumba. The individuals who consented to participate in the study were born, or had resided for more than 10 years, in endemic areas, and were confirmed to have, or not, microfilaria in their skin snips as well as any other clinical symptoms of disease, such as dermatitis, nodules and/or ocular lesions. In order to identify the putatively immune individuals, biopsies of the mf-individuals were also tested for the presence of the 150-mer DNA repeats specific for O. volvulus using PCR and Southern blot. Samples were collected before the introduction of ivermectin or from subjects that had not received ivermectin treatment prior to the studies. Adult worm samples were obtained from nodules excised during nodulectomies. Briefly, individual and cleaned freshly obtained nodules were immersed in 0.5% collagenase (Sigma Aldrich, grade IV) in RPMI-1640 containing 10% FCS supplemented with 200 units of penicillin and 200 .mu.g/mL streptomycin. The flat tubes containing the nodule were then placed in a rocking water bath and incubated at 35.degree. C. until the tissue was digested completely. Alternatively, frozen nodules were thawed, cleaned and digested with LIBERASE.RTM. TL (Roche) in Hanks Balanced Salt Solution (HBSS) supplemented with 3 mM CaCl.sub.2. When digested, the liberated adult worms were unraveled from residual tissue with mounted needles under a dissecting scope, and then washed in several changes of RPMI-1640 or HBSS. The cleaned adult worms were stored at -80.degree. C. until use.

[0083] L3 larvae were produced at the Tropical Medicine Research Station, Kumba, Cameroon. They were obtained from Simulium damnosum flies 7-8 days after infection with skin microfilariae. After dissection and washing, the larvae were cryopreserved and shipped to the USA. Fresh L3 larvae were also cultured in vitro in groups of 10 larvae in 96-well plates containing a 1:1 mixture of Iscove's modified Dulbecco medium and NCTC-135, 20% FCS and antibiotic-antimycotic (Life Technologies, Gaithersburg, Md.) for 3-days at 37.degree. C. Larvae were collected after 1, 2 or 3 days in culture, washed with Tris-EDTA and snap frozen in liquid nitrogen.

[0084] Nodular and skin microfilariae were also purified. Embryonic stages were purified from mf and eggs that were extruded into the medium during the cleaning process. The medium was collected and centrifuged at 1000 rpm for 10 min at room temperature. The pellet containing the mix of microfilariae and embryonic stages was resuspended and layered on LSM (MP Biomedicals, CA) and centrifuged at 500 rpm for 15 min with the brake off. The purified embryonic stages that formed the pellet were washed and stored at -80.degree. C. until use.

[0085] Transcriptome Sequencing, Assembly and Analyses.

[0086] High-throughput transcriptome data were generated from the RNA of O. volvulus stages: nodular microfilariae (NodMF), skin microfilariae (SknMF), L2 (OvL2), L3 (OvL3), L3 day 1 (OvL3D1), L3 day 3 (OvL3D3), adult male (OvAM), and adult female worms (OvAF). For all larval stages and adult worms, RNA was prepared using TRIzol.RTM. and lysing matrix D (1.4-mm ceramic spheres) and a FASTPREP24.RTM. (MP Biomedicals). RNA-seq libraries were prepared following the RNAseq protocols of the Illumina mRNA-Seq Sample Prep kit and the Illumina TRUSEQ.RTM. kit. Transcriptome libraries were sequenced on Illumina HiSeq 2000 machines. De novo assembly was done and is reproduced here with slight modifications. Reads were trimmed of low quality regions (<13), and only those with an average quality of 20 or more were used. Illumine primers were removed from the sequences following a parallel BLASTN of the reads against HiSeq TrueSeq adapters. Resulting reads were assembled with the ABySS software (Genome Sciences Centre) using various kmer (k) values (every fifth from 21 to 91). Because the ABySS assembler tends to miss highly expressed transcripts, the SOAPdenovo-Trans assembler was also used, again with odd kmers from 21-91. The resulting assemblies were joined by an iterative BLAST and cap3 assembler. Coding sequences (CDS) were extracted using an automated pipeline based on similarities to known proteins or by obtaining CDS containing a signal peptide. CDS and their protein sequences were mapped into a hyperlinked Excel spreadsheet. Signal peptide, transmembrane domains, furin cleavage sites, and mucin-type glycosylation were determined with software from the Center for Biological Sequence Analysis (Technical University of Denmark). Reads were mapped into the contigs using BLASTN with a word size of 25, masking homonucleotide decamers and allowing mapping up to three different CDS if the BLAST results had the same score values. Genes that had blast scores <30% of max possible score (self blast) in other nematodes with an e-value greater than 1E-05 were considered as `unique`. To be O. volvulus unique, the genes were compared with the genomes of O. flexuosa and O. ochengi. Automated annotation of proteins was based on a vocabulary of nearly 290 words found in matches to various databases, including Swissprot, Gene Ontology, KOG, Pfam, and SMART, Refseq-invertebrates and a subset of the GenBank sequences containing nematode protein sequences, as well as the presence or not of signal peptides and transmembrane domains. Protein repeats were analyzed using repseq and reptile (www.reptile.unibe.ch) algorithms. Further manual annotation was done as required.

[0087] Transcriptome data (RPKM) from Excel spreadsheets was imported into JMP Genomics (SAS, Inc.) for general assessment of distribution analyses, correlations, principal component analyses, analysis of variation (ANOVA), hierarchical clustering, and heatmap generation, parallel co-ordinate plots. Heatmaps of clustering analyses were also done in R using array of packages. Differential expressing of genes was analysed using DESeq. Two replicate samples Ov1F (male) and Ov4F (female), were observed to not be exclusively male or female (pre-analyses) and were excluded from all stage-specific analyses. However, they were used for differential expression analyses with the rationale that any contaminating female transcripts present in the male sample would result in the differentially expressed genes with lower adjusted p-values to drop off and thus enriching for highly expressed genes. Likewise, any male transcripts in the female (including contributions from stored sperm and embryos) would lead to drop-off of lower range of genes and selecting for the most highly regulated genes.

[0088] Protein Depletion, Denaturation, Digestion, and Desalting.

[0089] For proteomic analyses, additional stages of embryos (OvEMB), L3D2 (OvL3D2), and L4 larvae (OvL4) were also analyzed. Total soluble proteins from all the stages were extracted using the UPX universal protein extraction kit (Protein Discovery) as per manufacturer's instructions and quantified using PIERCE.RTM. BCA assay (ThermoFisher Scientific). Extracted protein samples were prepared for digestion using the filter-assisted sample preparation (FASP) method. Briefly, the samples were suspended in 1% SDC, 50 mM Tris-HCl, pH 7.6, 3 mM DTT, sonicated briefly, and incubated in a Thermo-Mixer at 40.degree. C., 1000 RPM for 20 min. Samples were centrifuged to clarify and the supernatant was transferred to a 30 kD MWCO device (Millipore) and centrifuged at 13000.times.g for 30 min. The remaining sample was buffer exchanged with 1% SDC, 100 mM Tris-HCl, pH 7.6, then alkylated with 15 mM iodoacetamide. The SDC concentration was then reduced to 0.1%. Samples were digested overnight using trypsin at an enzyme:substrate ratio of 1:100 at 37.degree. C. in a Thermo-Mixer at 1000 RPM. Digested peptides were collected by centrifugation. Twenty micrograms of the digested peptides were desalted using reversed phase stop-and-go extraction (STAGE) tips. Peptides were eluted with 80% acetonitrile, 0.2% trifluoroacetic acid and lyophilized in a SPEEDVAC.RTM. (ThermoFisher) to near dryness, approximately 1 hr.

[0090] Protein Array Construction.

[0091] The following cDNA libraries from OvAM (SAW98MLW-OvAM), OvAF (SAW98MLW-OvAF), OvL2 (SAW98MLW-OvL2), OvL3 (SAW94WL-OvL3), molting L3 (SL96MLW-OvML3), and MF (SAW98MLW-OvMf) were obtained from the NIH/NIAID Filariasis Research Reagent Resource Center (www.filariasiscenter.org) and used to amplify selected gene products. Molting larvae transcripts that were not amplified successfully from the cDNA libraries were subsequently obtained from oligodT cDNA prepared from RNA purified from OvL3D1, OvL3D2 or OvL3D3 (SUPERSCRIPT.RTM. III First-Strand Synthesis System, Invitrogen). In vivo recombination cloning was performed. Briefly, PCR primers were designed as 40 mer oligonucleotides with 20 sequence specific bases and a 20-base adapter sequence. The adapter sequences were designed to be homologous to the cloning site of the linearized T7 expression vector pXT7 and allow the PCR products to be cloned by homologous recombination in E. coli DH5a cells. PCR reactions were set up using HOT MASTER MIX.RTM. (5 Prime) plus DMSO (5%). The recommended cycling conditions were used and PCR products were checked for correct size using an agarose gel. PCR products were mixed with linearized pXT7 vector and were transformed into DH5a competent cells. DNA was purified using QIAPREP 96 Turbo.RTM. Miniprep Kit (Qiagen). Resulting clones were checked for insert on an agarose gel and were sent for sequencing (Retrogen).

[0092] Chip Fabrication.

[0093] Proteins were expressed using a coupled in vitro transcription and translation (IVTT) system, E. coli based cell-free Rapid Translation System (RTS) 100 High Yield Kit (5 Prime), from the O. volvulus expressible clone library following the manufacturer's instructions with the exception of adding detergent to the IVTT master mix at a final concentration of 0.1% Brij 78. Shortlisted O. volvulus proteins were synthesized using IVTT in disulfide-bond folded formats and printed onto an array. Known immunogenic proteins (purified recombinant proteins) were also printed as positive controls.

[0094] Approximately 1 nL of unpurified IVTT reactions were spotted onto 8-pad nitrocellulose coated ONCYTE.RTM. Avid Slides (GraceBio Labs) using an OmniGrid Accent microarray printer (Digilab) equipped with a Avid.TM. 946 Printhead and 946MP4 Spotting Pins (Arraylt). Each IVTT expressed protein includes an N-terminal 10.times. polyhistidine (HIS) epitope tag and C-terminal hemagglutinin (HA) epitope tag. Microarray chip printing and protein expression were quality checked by probing random slides with mouse anti-polyHIS (Sigma), rat anti-HA (Roche) and rabbit anti-E. coli (LifeSpan BioSciences). Antibodies were diluted 1:1,000 in a 3 mg/mL E. coli DH5a lysate solution in protein arraying buffer (GVS Filter Technology) and incubated at room temperature for 30 min. Chips, FAST.RTM. Slide Holders (GVS Filter Technology) and FAST.RTM. Slide Incubation Chambers (GVS Filter Technology) were assembled and nitrocellulose pads were hydrated using 100 .mu.L blocking buffer for 30 min at room temperature with rocking. Blocking buffer was removed, pre-incubated antibodies were added and chips were incubated for 2 hr at room temperature, washed three times with 1.times.TBS-0.05% TWEEN.RTM. 20, followed by incubation with Cy5-conjugated goat anti-mouse IgG Fc.gamma., Cy5-conjugated goat anti-rat IgG Fc.gamma. or Cy5-conjugated goat anti-rabbit IgG Fc.gamma. (Jackson ImmunoResearch) diluted 1:400 in blocking buffer for 1 hr at room temperature with agitation. Chips were washed three times with 1.times.TBS-0.05% Tween 20, three times with 1.times.TBS, and once with water. Chips were air dried by centrifugation at 500.times.g for 10 min, stored in a light proof desiccator for at least 2 hr and scanned on a GENEPIX.RTM. 4300 with Autoloader (Molecular Devices) using the 635 nm laser. Resulting 16-bit TIFF images were quantified using GENEPIX.RTM. Pro Microarray Analysis Software (Molecular Devices) and a GENEPIX.RTM. Array List (GAL) file. Spot and background intensities were measured and median spot values minus local background (M635-B) values were exported as comma delimited file (CSV).

[0095] Probing Samples.

[0096] Serum samples were diluted 1:100 for IgG and 1:50 for IgE in a 3 mg/mL E. coli DH5.alpha. lysate solution in protein arraying buffer and incubated at room temperature for 30 min. Chips, FAST.RTM. Slide Holders and FAST.RTM. Slide Incubation Chambers were assembled and nitrocellulose pads were hydrated using 250 .mu.L blocking buffer for 30 min at room temperature with rocking. Blocking buffer was removed, pre-incubated serum samples were added and chips were incubated overnight at 4.degree. C. with agitation. The following day, chips were washed three times with 1.times.TBS-0.05% TWEEN.RTM. 20, followed by incubation with biotin-conjugated anti-human secondary antibodies against IgG1, IgG3, IgG4 or IgE (Sigma Aldrich) diluted (1:1,000 for IgG, 1:500 for IgE) in blocking buffer for 1 hr at room temperature with agitation for one hour. Chips were washed three times with 1.times.TBS-0.05% TWEEN.RTM. 20, followed by incubation with streptavidin-conjugated SURELIGHT.TM. P-3 (Columbia Biosciences) at room temperature protected from light with agitation. Chips were washed three times with 1.times.TBS-0.05% TWEEN.RTM. 20, three times with 1.times.TBS, and once with water. Chips were air dried by centrifugation at 500.times.g for 10 min, stored in a light proof desiccator for at least 2 hr and scanned on a GENEPIX.RTM. 4300 with Autoloader using the 635 nm laser. Resulting 16-bit TIFF images were quantified using Innopsys Mapix Software and a GAL file. M635-B values were exported for each slide as GPR files.

[0097] Data Analysis.

[0098] Software developed in R (Antigen Discovery Inc) was used to process the individual GPR files in batch to create a single matrix of the raw data and to perform automated data quality checks. The raw data were normalized by dividing the IVTT protein spot intensity by the sample specific median of the IVTT control spots printed throughout the chip, then taking the base-2 logarithm of the ratio. The normalized data provides a relative measure of the specific antibody binding to the non-specific antibody binding to the IVTT controls. Normalized data was imported into JMP Genomics (SAS) and analyzed for antigen reactivity and significance (ANOVA) between the clinical groups and isotypes, and adjusted for multiple comparisons. Significant proteins were graphed in Prism 6.0 (GraphPad).

[0099] Transcriptional profiling by RNAseq resulted in the identification of transcripts corresponding to 99% of the predicted genes (FIG. 1A) across all the stages in the parasite lifecycle. Over 75% of the genes had 100% transcript coverage in all the stages except the adult female which may have been related to the age/condition of the worm(s) inside the nodule, degradation of RNA during the digestion of the nodules, or the fact that the majority of the adult female worm is comprised of uterine tissue and embryos. Several transcripts with less than 1 RPKM were subsequently identified and verified by mass spectrometry (in proteomic analyses) and thus have not been excluded. Shotgun proteomics identified proteins with a median coverage range of -10-15% from each of the stages profiled. A total of 7,774 O. volvulus proteins were identified across all the stages (FIG. 1A) resulting in the validation of over 64% of all predicted proteins. Though there were no differences in the number of transcripts identified in each of the stages, maximal proteomic coverage was observed during the L3 to L4 development and in the adult male and female worms (FIG. 1A). This approach also resulted in the identification of 465 of the 785 putative Wolbachia proteins (FIG. 1B). Wolbachia is a genus of bacteria which infects some nematodes. Wolbachia species have been found to be endosymbionts of O. volvulus adults and microfilariae, and are thought to be the driving force behind most of O. volvulus morbidity. Overall r-values for correlations across all of the stages between the transcriptome's RPKMs and proteome abundance ranged from 0.25-0.39, values that are considered acceptable for global comparisons.

[0100] Multivariate analysis revealed stage specific transcript profiles that segregated the vector-derived stages (OvL2, OvL3), the early human developmental stages performed in vitro (L3 to L4 molting: L3D1, L3D3), the adult male (OvAM), the adult female (OvAF) and the microfilarial stages (SknMF--mf obtained from skin, NodMF--mf obtained from nodules). Infected nodules often contain more male worms than female worms which has been attributed to adult male migration between nodules. The proteomic analyses indicate a probable bias towards a male-like expression profile as the worms develop from L3 to L4 and to young adults. Hence it was hypothesized that it is also likely that proportionately more male worms develop from a single infection. Indeed, structural gender differentiation can be observed in in vitro developing L4 larvae. Notable among transcriptional and proteomic profiles was the observation that, compared to all other stages, the adult males have higher transcript abundance levels with many differentially expressed genes.

Example 2. Stage-Specific Functional Enrichment

[0101] Functionally, the total putative proteome was classified into functional categories. Forty-four percent of O. volvulus genes have no yet known function. The distinctive biology of O. volvulus is likely to be underpinned by genes with potentially novel functions and with relatively few homologues in other helminth parasites. Approximately 9% (1173) of the predicted genes in the O. volvulus genome encode unique genes with less than 30% homology with other nematodes. 92% of these `unique` genes are hypothetical or genes of unknown function of which 7% are potentially secreted. Clustering of these unique and divergent genes based on transcript and protein abundances indicates distinct subsets that are enriched in specific stages, and that these clusters have signatures of being able to be secreted ("secreted-divergent"). Although largely uncharacterized, the stage specificity of their expression is an indication of their developmental regulation and may allow for functional assignments in the future.

[0102] Gene Set Enrichment Analysis (GSEA) demonstrated that the female stages were associated with pathways linked to detoxification and the extracellular matrix. This enriched subset of extracellular matrix related genes was primarily comprised of collagens and chitin. Although the microfilariae are an integral part of the fertile adult female, genes corresponding to NADH dehydrogenase activity (GO:0008137) and cytochrome-c oxidase activity (GO:0004129) were highly represented in adult females. In contrast, the microfilarial stages showed significant enrichment for processes associated with protein synthesis (ribosomal proteins) and protein modification with cyclophilins and chaperones (heat shock proteins) being the major contributors. These are likely the machinery required for cellular morphogenesis that occurs after being ingested by the blackfly vector.

Example 3. Secretome and Host-Parasite Interactions

[0103] The O. volvulus genome encodes .about.20% of genes predicted to be secreted by classical secretion and about .about.42% through non-classical secretion. All filarial helminths are known to release excretory/secretory (E/S) products that are critical components in the helminth arsenal of proteins that perform diverse functions that include: 1) modulating the host immune response; 2) host tissue remodeling; 3) alteration in host tissue nutritional status; or 4) enhancement of larval tissue migration. The O. volvulus genome encodes many of these immunologically relevant genes. Among the examples of the stage-specific enrichment of these immune-related gene products are the L3-enriched or mf-enriched cystatins and serpins that have been shown to interfere with antigen processing and presentation to T cells; the OvAM-enriched expression of indoleamine 2,3 dioxygenase (IDO); and the developmentally regulated L3/L4-enriched homolog of suppressor of cytokine signaling 7 (SOCS7; OVOC678). Proteases (serine, aspartic, cysteine and metallo-) are integral to host invasion, developmental molts and migration in a number of nematodes. Serine protease inhibitors also play an important role in controlling the molting process and immune evasion. The analysis of the Ov genome revealed the presence of 18 serine protease inhibitors, nine of which are highly expressed during the L3 to L4 molt. Four of these are SPI-like, probably having resulted from a duplication event of Ov-SPI-1 and Ov-SPI-2; their marked expression during the L3 to L4 molt is consistent with not only their role in early larval development but also in their putative role in immune evasion during their early adaptation to their human host. Interestingly, two of the serpins are also highly expressed in adult males indicating a potential role in spermatogenesis, while one is highly expressed in both nodular and skin mf.

Example 4. Immunomics

[0104] Using an immunomics approach, host antibody responses to candidate parasite antigens were profiled. Selected (397) proteins (based on their elevated expression in infective stage larvae and during molting and/or in microfilariae and/or adult stages) were printed as protein microarrays, quality checked, and assessed for isotype-specific responses (IgG1, IgG3, IgG4, IgE) with 52 individual sera comprising O. volvulus-infected, putatively immune, and control individuals from Ecuador, Guatemala, and Cameroon. After normalization, clusters specific for IgG4, IgG3 and/or IgG1, and with or without IgE reactivity were identified. Heretofore unrecognized biomarkers of active infection were identified (e.g. OVOC10469, OVOC10602, OVOC11950, OVOC3261, OVOC5127, OVOC8491, OVOC9988) as seen in FIG. 2A and Table 3. Further analyses led to the identification of potential novel vaccine candidates (e.g. OVOC10819, OVOC5395, OVOC11598, OVOC12235, OVOC8619, OVOC7083) (FIG. 2B and Table 4), based on IgG1 and/or IgG3 reactivity (with little to no IgE reactivity).

TABLE-US-00003 TABLE 3 O. volvulus biomarker sequences SEQ Protein ID NO. Description Sequence 1 OVOC10469 NIAFAPNPKDSNNELFADAE Secreted SALGSEYAQFVEQSKQHKPV protein YFSDNQNTLETIKLESIPNP ETETAYPMFICGFLGCMKKM NSVEEYLEHFKMHEKQGY 2 OVOC11950 YPTEKETVEPIDTMVKDDID Secreted LVKAEVAEAEEADVEKEVAE protein LTEEEAAEIAEVLDEMEEEF FAFLLFDFILDLFRETLEKN SESQEASIDEVMPEIQGVSA EEA 3 OVOC10602 FRTQSIGIRGRLMCGSKPAS Conserved NERIKLWEEDSDDLLDQGYT secreted DENGEFLLKGDTVELTPIDP protein VFKVYHDCDDGIKPGKRKVK FKIPKSYITEGKTPKKIFDL GTLNLETIFNDEERELIVT 4 OVOC3261 SYCEDWDPEDFPSFVLKLSQ Secreted NATEEFCELYEMEMEVPINK protein FYDMLRKWAEKYSVQAETNR FIAEEMNYDKMQSKVLMERL QASNGTTEVKGVLEKALKLQ ESMHLSPDYIQNVIDTMMEN LPIDKQNEATLLWNSLYPDD IYNECGPRF 5 OVOC5127 APPNRDTADDLQNADMQRQW DNA binding EQEQRQREEVQKEEIAKYVK protein YMRYLETVLNILQATPQWKE AMQSMTQEEMRGGKIAEMVD KLEPHIIEQLAKAKILELQR LEQEIKDQLNADGGATHNIK VSEILTVCLKEGFKKPSNLG IPEHLDFNNWETFSQEDLRK LIVKIVTDMDELEEQRKQDF KQYEMKKKAEEDHKMQAKMI QTEREEYIRQMEEQRRRHNK HEPLKHPGSRNQLRKVWEDT DKLDKDAYDPTTLFGLHDRN NDGYWSYDELNTIFLPEIEK LNNFSDVERLEELYRMRDHV MKQMDTDGDHRISRAEFLAD REAQEEKPDQGWEDIGDKDQ YTKEELEIFEKEYAKQQGWG EYAYSTPAPTPDPSRMIQPD QAPMQRLDAPSDQVGDMFAQ QSHQIPVKHVEPIQSVQQQQ MDEVNS 6 OVOC8491 FPTEANPAVGTDNAHEDNLT Fatty acid TEEKMQLKKFAKTNAANFSL retinoid TDPEFIDGLKNEAAGLFSKL binding TGLRDIINAKLDTMQPESRL protein 2 FIEKLLRRFLAAFSHDGLMN ILESLKGFGKEVIDMFDGLS RPIQNDILNAFPLVGSYITS DIARLMLRKLAELDLLSRKS TLTPTVDQFNDDSGKHFPRP QVIEPEEPENSDPEDAQSTD YGKKKVVTTTTFPIITGEED EILVKKIVENK 7 OVOC6759 IPLPEELDYDGEIPNCRDGE Conserved KPLLAADIGVYTCDKNCPKG secreted FRCEYRTMDSTSKKGICCPN protein LKELAKIYSEDEEVDKSIKK SNI 8 OVOC451 MISCFALPFPHVCYMAYCTQ Filarial VIASIMKGWNQNFRFSTVIY antigen LFRNIFSSSVISCVNMILSS Av33 TFYALLFVSAVVIVEAMPAS ESTYSVIIIRINDTTCKIED GVVSVNGQVIGNLTEEQKEE LEAYNVQTQGWFQQLHQKIE ELFKTFFGSIKSMWKHSPIS GSESSPQSSTPDNIITDKLD DQDRRLKDQGDSENSSLFGL KLPSFCKVN 9 OVOC12329 FRSLKIGRKQSTAVKGVLTC Conserved NGKPAVNVKVKLYNDSQGRY secreted VENSMDEGKTDSEGRFLLQG protein HETSITSIDPILKLYHNCDV ENAQCLKRFSILIPNDFVSE GLEPKKTFDMGTLNLGGKFF DEGRECAS 10 OVOC3337 QIIGSFNGNYAGDGSLNNNA Glycine-rich NSFGERTTTTRSTSRPSLPP cell wall RPGYPSRPGYPFKPGFPPRG structural PPIPYPHGKPSGPRYPCYGG protein-like YGGYGHPGYGPFGGNGYLGY TVCSGRGEFGGYGPGLGGGT GLGGLGPGEFGGYGPGLGGG TGLGGLGPGGFGGIGPGLGG GGGLGGPGRGGFAGYGPGLG GGRGLGGPGPGGFDGYGPGL GGRPYPGGYGRFYGPGPYPG DRLDPRGLSESGRPRTRLAS YNRNDRGTQFSYIRDR 11 OVOC10264 MTNEYETNYPVPYRSKLTES Beta- FEPGQTLLVKGKTAEDSVRF galactoside- TINLHNTSADFSGNDVPLHI binding SVRFDEGKIVFNTFSKGEWG lectin KEERKSNPYKKGDDIDIRIR AHDSKYTIYVDQKEVKEYEH RVPLSSVTHFSIDGDVLVTY IHWGGKYYPVPYESGLSGEG LVPGKSLLIFATPEKKGKRF HINLLKKNGDIALHFNPRFD EKAIVRNSLIAGEWGNEERE GKMILEKGIGFDLEIKNEEY AFQIFINGERYATYAHRLDP REINGLQIGGDLEVSGIQMR 12 OVOC4230 DLLSEAGDFFTKHFTDIKSL Conserved FAKDEKQLQQSVDRVKDLLA secreted TIQDKMSMLQPLANDMQKTT protein LGKIGDLISQVNSFRETMSN PKMDFTNKENKWEELLKKIF VTEGLNKVIPLLQKLKNSAP TTFATYLFTCIVPVLINTLR E 13 OVOC10384 MARINRLNFLLCIVHANITS APNPKDSNDELFADAESALG SEYAQFVEQSKQHKPVYFSD NQNTLETIKLESIPNPETET AYPMFICGFLGCMKKMNSVE EYLEHFKMHEKQGY 14 OVOC8422 FSWKFGERLDEPVLMLRDLR secreted AKEISPPSYMKRFESDTNEQ protein LLRYILHPKMLRRHDLSNAL FYQPLWKMR 15 OVOC6395 MEGSPIKETRGLEATPVFEM Protein VRSATLTFLLAVSTVLVVSR LOAG_00657 PNVLLPPKLPWDSDWRQKPP PFPPEPPPEFKGILPPEIFA KLTAIHQDQSLTIPQKIVKI EEIMNSLPEDVLQRLPLPPV FRLLPQNVQEMIKTVRTTKN LTMEEKWLQMIILIESLPKQ QHRLLQQMLPKFSLGPLPDF QDIIPKEDWDKLTAVYQDTN LDNIEKLRRVDEIIDALPDS IRQKIPLSPPFQKLPDHIQQ QLQIIHTERGLTTEQRFRKM KAIIESLPWDMKKLMFQP 51 OVOC10469_ EQSKQHKPVYFSDNQNTLET Pep2 IKLESIPNPETETA 52 OVOC3261_ CPSLSSYCEDWDPEDFPSFV Pep1 53 OVOC3261_ LPIDKQNEATLLWNSLYPDD Pep3 IYNECGPRF 54 OVOC10469_ AFAPNPKDSNNELFADAESA Pep1 LGSEY 55 OVOC10469_ GCMKKMNSVEEYLEHFKMHE Pep3 KQGY 56 OVOC3261_ INKFYDMLRKWAEKYSVQAE Pep2 TNRFIAEEMNYDKMQS 57 OVOC5127_ APPNRDTADDLQNADMQRQW Pep1 EQEQRQREEVQKEEI 58 OVOC5127_ TDMDELEEQRKQDFKQYEMK Pep2 KKAEEDHKMQAIQTEREEYI RQMEEQRRRHNKHEPLKHPG SRNQLR 59 OVOC5127_ DREAQEEKPDQGWEDIGDKD Pep4 QYTKEELE 60 OVOC5127_ TPAPTPDPSRMIQPDQAPMQ Pep5 RLDAPSDQVG 61 OVOC5127_ VSEILTVCLKEGFKKPSNLG PepX I

TABLE-US-00004 TABLE 4 Immunogenic composition O. volvulus protein sequences SEQ Protein ID NO. Description Sequence 16 OVOC8619 LIKVFPEISANMSVMFANSRSNQ Adhesion- ANNGYLVEFKAGRSNLQAGSTVD regulating RRKVVADKTKGLVFIKQSSDQLM molecule HFCWKNRETGAVVDDLIIFPGDT EFLRVRECTDGRVYMLKFKSTDE KRLFWMQDGKTDKDDENCKKVNE TLNNPPAPRAAARGGADRADVSS FGTLAALGSAGAESELGALGNLD QSQLMQLLSLMNHTNSTSASEAT NLLPQLPLVADTSHPMTSEDSGT TSTHGATPSNTPANGIVADSSSN NAMQLSQLKEIIASITPPDGSGR KPSIDFTDVLCCADKINDVLRKY AEQLIPHLPSQEPIYNNQEELQQ TLRTPQFRQAADIFGHALQTGQL APVLRQFGIDGNTATAAGNGDMV AWAAQFTTAENGKEITAKTETSP SQPGMESDVEDEETNEKAIRETE KNRTDDHMDLD 17 OVOC7083 MNYKAPIELQQLLSITKMLSLSV Secreted LLLFTSMAIMARPPNSDEIKELR protein QQQLNESKDDYDTLPDVNHIPES FKESLKKQKMLYLDMLRQHNL 18 OVOC4111 LSVPAGLRPAKKVGDPKEQIVPG Mediator of KQQQLQQQQQQQLLQQQQQQQQQ RNA QQQQQQQQQQQQQQQQQQQQQEQ polymerase II QEELQQLQESEETGEEHRQQQQQ transcription QHDEALTLSPTPKVPPNLSIRSR subunit 15 MMAALSASVGESNKEKNSSNDET DNSSKSTNSPSKPPIIFPKANKK TVVGKIAPSGISKGSARVIVAPP SKLGTNNFGLNTVLQTNLVDSRG RIMKNVNSVPIKVPSSAEMRNAR TRHTARQVESDADKVVPIKFGST SRRR 19 OVOC1808 NNSNLDISMREKNAVNAIEKQDL PRSHRFKRQYSCGQCGGGGGPPV VVSPCQQCKGGGAGVSAIGGAGG ISAIGGGVSAIGGGFGGGGGDTV AVVCCGATGLKGMFRNWWLHIPL LLLPMSMSWIKALFL 20 OVOC11598 YYVPDNYWPLRIIGYHHIPVMIN Secreted MWYLFQTEISNIGVDAVLVQSPL protein YRTLTPDVVHDIISINVEPNHTV VVEQSNPMLQASSVEQAPAAAPL SITLIAPGITISRTHKVDTYKST MEMYDADKLHSNEIFKRRVRKMV LPPSRGEEVRKPPSSTDGYESEN VESYGQKGVEQAPPEIEQYVKKK K 21 OVOC3901 MKYCLSSIIAATIATTTTTATAI Immuno- IATTITAATISVAPFHASSPSSS globulin LSSSSFSSFFLVLPLITTILLIV I-set PEQAHSTATVTEHRSPPDLSIPS domain QTEFRVPVGTKQFRLICPVKEKN containing DDLLMIQWKKNDEPIGFDFNNRF protein KLARSDRELKIRNPQLSDGGIYQ CQVVNGFGHRELNFTVTFYDPAM ENDQNTDSTLTLTTKASPPIWKN ETEIRNWMINPVRITIGGALLLK CPAKGNPLPHITWLRDGKVLERE ITYHYSSAILYLSDVQPSEGGKY ICKLENEHGSIEASFHVYVENFF EGLDGESWSIDQTNAQLYPVIDE PFNNTVRVGRTAQFQCKVKNQQQ PLIKWLKRVEDPNAIRQTNANAT LIHANNMHLLLLEKPETSAELSD GISLNRLIIPNVRYEHSGTYLCV VTNARGDIAYRSAYLNVIARSDH GELSNLYFYGGLLVLIVVFTLIT YAVHFLRKNQAAKSTESAPGITN IRYSFSLRPPPPNLPPPKAPALP SERQQLMPNNQPCDRYTVNSAAA TYYPQFATPDKKLQKIITESGTR PTPIRRTNGGDTKYRLKDDYISS PKWVHAKGDNIEVEMDQNLLKNR STHCHNPVSIAYGRIDNIDRQQQ KSFLTIGNLQKR 22 OVOC10819 KEIIWDCYGDYEECVAESSKMDH Secreted VDVNNVESRNIIEFCSDHTQNIL protein PCLATKLGLIKSMSVSMFSLLLT ICEAETRNNRPAATEVQQILKHL ARLYAYFCAYSNVIDLRYNKECF RYLKKRCILNKPDDSCIFHHCGE KNLNLSESSPFIQQHKTTIINQL NQSATFKNYHHRITTIFTVIITF ISMIQ 23 OVOC5395 MYNQENHDKRRNDDRFILSLPFG Protein TNVENKSYFKPIKLSNPYSDKYL Bm1_06245 EVNKKSSDDSDQNLNQALSVPQS NYDQSSESLSIDDSDLIDDSTSA AQLSTSSPISVTSASTSSFYPTL NIGNGMEISAKYAKLEQSQGIKS DQSTSRVSDRYKKYTAVKRRLSE LYGIIEEKDEQLRVVRNELNGKD LEIGKLCDKIRALEYNCGRLQSM IESAGDESDQNQVKLHEIINERD GLLIRNASLSRQIEFEKREWSIE RERLSMDLDDVTRELELQKMILN GESISEIVQRWQTKVFELEGMIT DRDRAIRAQQVQISKLKESIAET DRISCADSSESQTKFDFPSFTYI KRLLLQYLTRLADLHFSSDEERM QLVRNMSSILHLSDEEQRQVWAN LKSKIQIS 24 OVOC12235 QCPTGSVSLLSGYRCTSSIQCQT Conserved IIPGSYCYYGVCCTGGSDVLSKT secreted VSYGGYCTMTVQCSTTGATCISN protein ICQCDINSHYNGHSCVSISNFCP SNQVFIKGECYRKVTYGFLCNYT QQCGYIGAFCIGNICSCQLDYTF DGSKCIPRSRICPANQIAIGGQC YPSARFGERCLYSEQCIDRWYRS LSCVNGFCNIRNDDDISKPKCRN PRAEVEYVNGTAKNCLYWPCTVG YFCEYAGGMNGGRYICCGTNANK IYGKVQLYPGTGTPLQCTEIGRC PFPDTPNCVMSYRYGYKVCCSTL NC 25 OVOC7908 QETSEQPGLTVEIIAEQQDATTA Lateral DQEVTTTVDTHHQHQHQTDKVVK signaling SRQITGDEQTTTTTTAINLNETI target protein TNSTTDSNSTIITTTLDLQESTT 2 homolog TGTTDNHHHHHHHHHHHE 26 OVOC7430 MKQTTAWGNALCVLCNCHQPQII RhoA GTPase CPPPPPAVCPRVVCPPPRPPVCP effector DIA/ PIYCPPPVVCPPPPVCPPVPFCH Diaphanous SQICPPCGTHTVPVAVVGCCKGC ACSVRFKRDSSSVNGLMLKKNLL CNNDQLMTIMEKKIGTNATEAAF AIKKEADSELKAKFSVFCAMNDL IYVAHAESFCQHKKGDIICFAYK S 27 OVOC8936 MDCKLILPFYILLANLEANAFHL Microfilarial SGYRSRSYLQGIQPYDIQPLDVQ sheath protein PQFIRVQTLKSQDIQPYSIQSRS EDQPCEGCKITISCGSKNCKSKK LPYVYKPIFKLLSTRSTKKPVFT LPTQPPAQWDCPCPCHVPQRCRM CSACHESYI 28 OVOC5806 NRIISRRLSLFIQQYCCNNISQI Conserved YRLNDCKYSKVKMEIDKKIFIIV plasma SKTEWCNEAIKVVFGKSAEAIRN membrane NSDAISWLASYNYTGSMDLRSKW protein PYDAYFDNVTRTAHGLARIDLLC HKKRPQLGPRIWKRSVQKIKQKK DRPFAVNTYGNNKGLFTITVGVL LYAAFGTCFLIANLAYLFGIYII YDASIIDEVS 29 OVOC4665 IGENPMDVNAIAGIIGGISNMMQ Conserved NNVETIDVPSSQIMGRWYQVYKA secreted AIAFDVYRTDIFCPVAYFKPNSV protein MGEDGFSIEEAYRVITKNGPVET YKRDLNKVGTGQYWMYTEEYFYP RQFNIISVGPNYTNTTDGSEEEK QYQYMVVTDGNRLSLSVYARHPM IFYQKYNEEVVKFLEHAGFGGKV FWNSPKPIYQGADCEWPSEKEVF ARRVLKNQELAKNGGLDTATKSG SFGGSSQATDVRSSITEILQNPQ LALQKLVQGH 30 OVOC8227 MTIIKSMLKITHVIFDLDGLLID HAD- TEVVFSKVNQCLLSKYDKKFTPH superfamily LRGLVTGMPKKAAVTYMLEHEKL hydrolase SGKVDVDEYCKKYDEMAEEMLPK CSLMPGVMKLVRHLKTHRIPMAI CTGATKKEFEIKTRHHKELLDLI SLWVLSGDDPAIKRGKPAPDPFL VTMDRFKQKPEKAENVLVFEDAT NGVCAAIAAGMNVVMVPDLTYMK IPEGLENKINSVLKSLEDFKPES VGLPAYDASSNE 31 OVOC9988 IPQRRQQQQQQQQQQQRDEREIP Serine/ PFLEGAPPSVIDEFYNLLKTDEN threonine KTDQQTEADVEAFINRLGGSYKV protein kinase RFTQFMEEVKKARADYERIHQQA DDB_G0280133 VARFSPAAKDADARMSAIADSPH LTTRQKSQQIQAIMDSLSESVRR EIINALSPQE 32 OVOC4230 DLLSEAGDFFTKHFTDIKSLFAK Conserved DEKQLQQSVDRVKDLLATIQDKM secreted SMLQPLANDMQKTTLGKIGDLIS protein QVNSFRETMSNPKMDFINKENKW EELLKKIFVTEGLNKVIPLLQKL KNSAPTTFATYLFTCIVPVLINT LRE 62 OVOC7453 KNPSKMESKTGENQDRPVLLGGW (CPI2M) EDRDPKDEEILELLPSILMKVNE QSKDEYHLMPIKLLKVSSQVVAG VKYKMDVQVARSQCKKSSNEKVD LTKCKKLEGHPEKVMTLEVWEKP WENFMRVEILGTKEV

[0105] Natural immunity against O. volvulus can be acquired in a few individuals in affected populations and these individuals are known as putatively immune. Consequently, they exhibit protective immune response against L3 larvae, suggesting that E/S products released by molting larvae and/or surface proteins of L3 larvae are an important source of protective antigens. The identification of proteins that are highly expressed by the mf and that are specifically recognized by sera from protected individuals who never developed a patent infection opens up new possibilities for also developing a safe anti-transmission or therapeutic vaccine. The identification of Ov-unique proteins that are adult and/or mf stage-specific that are recognized by sera of Ov-infected individuals provided additional novel biomarkers needed for better mapping the prevalence of infection and for post-control surveillance.

[0106] It is anticipated that O. volvulus proteins, or orthologs thereof, will provide protection against infection with D. immitis. D. immitis orthologs of O. volvulus proteins are provided in Table 5.

TABLE-US-00005 TABLE 5 D. immitis orthologs of O. volvulus proteins SEQ ID Protein Ortholog NO. Description of Sequence 33 nDi.2.2.2.t OVOC8619 MRTASQLTFMLFLVLKKKFK 00004 NIDKLFSQISVNMSVMFANS Proteasomal RSSQANSGYLVEFKAGRSNL ubiquitin QAGSTVDKRKVVADKTKGLI receptor FIKQSSDQLMHFCWKNRETG ADRM1 TVVDDLIIFPGDTEFLRVKE homolog CTDGRVYMLKFKSTDEKRLF WMQDGKTDKDDENCKKINET LNNPPAPRAAARGGADRAGA SSFGTLAALGSAGADSELGA LGNLDQNQLMQLLSLMNHTN SASASEAANLLPQLPLVADT PNPVASEESGTTSTQGATPS NTPANGIIAGSSSNNAVQLS QLKEIIASITPPDGSIRKPS VDFTDVLCCADKINDVLGKY AERLIPHLPNQEPIYNNQEE LQQTLRTPQFRQAVDIFGHA LQTGQLAPILRQFGIDSNTA IAAGNGDLIAWATQFTTSEN EKEIAVKTETLPFHPGMESD VEDEETNEKAVRESDKNRTD DHMDLD 34 nDi.2.2.2.t OVOC7083 MLPTLYINNAVIRPVLSETK 03357 KVKVQNISSPFLIFLLLSIT KMLSLSVLLLFISMATMARP PNPDEIKELHEQQLNDSKDD YDMLPDVGHIPESFKESLKK QKMLYLDMLRQQSL 35 nDi.2.2.2.t OVOC4111 MISSRLRITIPESIVIFGIF 05919 CFFIFFCFLSFFFFFTLWSH RDTINFQTDFMTETIKFIVY AVVILRMMFFDIVCFYSFLM MTIVLINTSNGLSVPAGLRP AKKVGDPREQIVPGKEQQQQ REQQQQQQQQLQEEEQQQQQ QHDEVSNLRPTPKVPPNLSI RSRMMAALSASPVEPNKEKN SSKVETDSFSKPPIIFSKGN KKTVPGKIAPSGSSKGNARV IVAPPADLGKNNYGLNTVLQ TNLVDSHGRIMKNVNSVPIK VPSSAEMKNARTRHTARQVE SDADKVVPIKFGSTSRRR 36 nDi.2.2.2.t OVOC1808 MMRIKWIILLLLLLLPIITA 07753 EFSAPVGINSSLTIFDKDKQ VLLRSDRLKRQCGPCGVAPS PVIVCCGAAGLKEIFRSWWL HIPLLLLPMSTSWLKTMVC 37 nDi.2.2.2.t OVOC11598 MFRLLIAIQILRFCQANYIN 06812 DVYWKRSIIGYQHIPIILNI CYLLQTEVSNKGVVDALFLH SPTYHRVEMSEETDNIESIA DKSNITVANKPNLMIYPADF QVSSNERASASIPITITITS SGDTIIKSFKHKHQSNEIFK RRVAKMAIAPVNAPEVENLA PEVENPSPSTAGYESKTEEQ APSESGQYGKRRK 38 Fibroblast OVOC3901 MYNLAKLLENEHGSIEASFH growth VYVENFFEGLDGESWSIDQT factor NAQLYPIIDEPFNNTVRVGR receptor- TAQFQCKVKNQQQPLIKWLK like 1 RIDDPNAIRQANANATLIHA nDi.2.2.2.t NNMHLLLLEKPETSAELSDG 10368 ISLNRLIIPNVRYEHSGTYL CVVTNAHGDIAYRSAYLHVI ARSDHGMLSNIYFYGGILVL IVVFTLITYAVYFLRKNQAA KNSESAQDITNTRYSFSLRP PPPNLPPPKAPALPSERQQL MSDNQPCDRYAVNSAATTYY PQFATPDKKLQKIITESGGT RPTPIRRTNGGDTKYRLKDE YINSPKWVHTKGDNIEVEMD QNLLKNRSSHCYNPISGAYG RTDNIDRQQQKSFLTIGNLQ KR 39 nDi.2.2.2.t OVOC10819 MLKLANTEIFFIAFLVYSKE 02919 IILNCYEDYKECVATSNKTN HVNMDNVNPQNLIEFCFDHT QNILPCLVTKLGLTKGISVS IFSLFLSTCELEAQNNKSSS TTEMQQILRHLLRLYAYFCA YSNIIDLHRNRECFRYLMKR CVLNKPDESCMFYHCGKIHF NLSKSSRKILFTRQHDTTKI VNLGNKMNQLATFNNHQVRS AVVVTLIITFIDMIQ 40 nDi.2.2.2.t OVOC5395 MYSQENQDDKRRNDERIALS 01093 VPYNNTNIMDRSYFKPIKLS YPYSDECLEVNKKSSDDSDQ RLSQNSSTPQSNYDQSSERL SIDDSDLIDDSTSAAQLSTS SPISVTSASTSSFYPTLNIG NGMEMNAKYAKIEQSEGIRS DQSSTLRISDKYKKYTAIKR RLSELCGIIEEKDKQLRVVR NGLNEKDLEIGKLCDKIRAL EYNCGRLQAVIESVGDESDQ NQIKLHEIINERDGLLVRNA SLSRQIEFEKREWSIERERL SMDLDDVTRELELQKMILNG ENISEIVQRWQTKVFELEGM IADRDRAIRAQQVRISKLKQ SLAEADRISCDDSSESQTKL DSPSFTCIKRLLLQYLTSSD EERIQLLRNVSTMLHLSDDE QHQVLTNLKSRIQIS 41 nDi.2.2.2.t OVOC12235 XKCRDQRAEVEYVNGSAKNC 11596 LYWPCTVGYFCEYTESRNGG HYICCGTNANNIYGKVKVYP GTNKPLHCSIMNTCPFLDTP NCVMSHRYGYKVCCSTMNC 42 nDi.2.2.2.t OVOC7908 MLMKQSDSCVDYFYDQYKGQ 05701 EYVKDDAFNTQNITDNFRKS SSDIAQLMNSQIELISQPEK VNEDSAKSSHYNDDLQKSIE DDTVEATQRKKDEKLLEFLH SLIVSTIPKTIHLEGNSVNL LTLTTTITPIAIITTKNTSG TANAITTRKYKKYKLNAFVN ISSDTLTELPKFLPENFNST NFANVEKTEKFSNSKQVATD SIFSLKESAYLETPVIRDFS SANDSAKTDPLFTRNYVDKQ IDMNTTKFNKNLKKSRLTTI STSNLTTVLSQLQTTTSIST TTSVTTTISTSITIPELTLV SQSHRHLHHYHHHHHHQYEN YDHESPIIVTALFDIGRGKW PRYTRTYEQYMNYLKHLLKL ENCLVIYTDSRGAEFVRQTR NVHNTQIFEISMHDLPLYRY REEMKGIIQREQKDWQFSPK TRYHPEANSADYNIIVNSKP YFLYNATQNVRFRTSDRMFV WIDAGYGHGRKGIIPDHCHW RPRLQRDRMTIIQLTPKHDK VSRYSITDLYRVDWVVLSGG FIAGDSHTINRFYRFYQKLF MELLDSGRIDDDQTILTLML KHYTTLFNPISSNGDWYALF RLFPCHDRQ 43 nDi.2.2.2.t OVOC7430 MKQATTWGSICEMCPCAAKP 04336 ICPPPVICPPRICPPPVICP PQICPPCPPRICPPPVICPP QICPPCPPQICPPCPKPQPP PPPPPPPVLPSLPPTSFKPM ITCCRTCICYIRRKRDSLND YDRIHDINPVCNNDQLMMIM KKKIRTNVTESTIAIKKAAD SMLQAEFNVFCAINDLTHVA HAEHFCQYKKONSVFDSFLF RSTLKGLIEECREGVRWWPG SLGDLDFSHISLYRAHKYIG NEEMNRSTKTKISFTRINKK WRLGHTGKKYNKVRFSRNIA KKFIGVCNIIRLKKSVSRSV RPFENQKSTSFNVFQLLVPK EKVEIVVDDTQAEEMNSETA QEVQLFNVRKSNADSKTDGE KDTADLDVILLTNEECSSSR QENLNKDEPEIVILDDSAPS KSDLNTSDEIICLQDLKMVN EVPTFSVTPKQKTVKELPRE TRTYGTRRGRQSRAYCEDLR KFPSIRNPVSSSSSSIHAKN MPEFVDLLTQGTLLICKKWL RRWDIVQSGVIGGNPLRICS YNVLCQQTAYKTPELYIHLT KPGRAYELTWENRWRLLTRE FSMIGADIFCLQEVQYDHYD QFFRPYFEAAGFFGKYKKRT NNLLDGCAIFYKSHLQLLHY RYIEYFLNIDSVLNRDNVGQ LIRLKDMRSGREFCVVNTHL LFNKRRGDVKLAQLAILLAN IDQECGPESGQECPYILCGD FNFHPYSPIYNFIMNGEICF TNLRRGDISGQGNAGGPFVS VNLLPEDVKIARNCRFNYLK NRTMLLPSLNCWSHPLCFNS VYQNMNGETRPMISTYHSIE AVNPDFIFYSVKSKRVQQSM LPHSVPAMNVSEREIRLIRR LSLPDMNELAGTLGPWPNST TPSDHIPLIADFVLQ 44 nDi.2.2.2.t OVOC8936 MYCKLIISFYMLLSIANMTH 10647 LVGYRPQIYLQGIPQNIQSH DIQRLDMQQQSLKLPDTELY SIPSHDNQLQGLQLYDMQFQ GKQSKGSEKLCSGCKISINC SGKKCVPMRTRKPIVTTPSP LSTQRPVLTRPRLLADCPCP CHVSRQCRICQPCQESFI 45 nDi.2.2.2.t OVOC5806 MFVGMRLYLAIDVLLLLVLR 03537 IKSNRIILHRFSLFIQQHCC NNISQIHRLNDCKYSKVRMK IDKKILIIVSKTEWCNEAIK VVFGKSAEARRNRSDAISWV TPYNFTGLMNLHSKWRYDAY FDNVIRTAHGLARIDLLCPK RRSHSGRRILKRSIQENKQE KSRRSFTVNIYGSSKGIFTI TVGVVIYAIFGVCFLITNMA YLSGIYTVHNTSVIPEDKKR KETSKRKEIL 46 nDi.2.2.2.t OVOC4665 MISVFLLLTVIVSYVETIGE 01073 NPMDINALAGIIGGISNMMQ NNVETIDVPSSQIMGQWYQV YKAAISFDAYKTDMFCPVAY FKPNSVMGEDGFSIEEAYRV ITKNGPVETFKRDLNKVGTG QYWMYTEEYFYPRQFNIIGV GPNYTNATDGREKENLYEYM IVTDANRLSLSVYARHPMIF YQKYNEEVVKFLEHAGFGGR VFWNSPRPIYQGTDCEWPSE KEVFARRVLKNQEAARNTGL ETATKSGLFGSSLTTDAYNP IKEMLQNPQLALQKLVQGH 47 nDi.2.2.2.t OVOC8227 MTVIKSMLNITHVIFDLDGL 00378 LINTEIVFSQVNQCLLSKYG KKFTSHLRGLVTGMPKKAAV AHILEHERLSEKIDVDEYCK KYDEMAEEMLPKCSLMPGVM KLVRHLKAHSIPMAICTGAT KKEFELKTRCHKELLDLISL

RVLSGDDPAVKRGKPAPDPF LVTMERFKQKPEKAENVLVF EDATNGVYAAIAAEESKIVK 48 nDi.2.2.2.t OVOC9988 MILEQLEVPPFLVGAPQSVI 01674 KQFYDLLKADETKTDAQTEA DVEAFINRLGGTYKTRFDQF KQEIKQGKAAYERLHQQAVA KFSKEAREADAKMSAIADSP SLITQQKTQQIQAIMD 49 nDi.2.2.2.t OVOC4230 MLKYGILLILITVGAYCDLL 06953 SEAGDFFSKHFTDFKSLFAS DEKQLQQNMDRVKDLLATIQ DKMTILKQLADNSQKSTLEK ITDIISQVNDFRENVFNSNV DFNQKKTKWEEVVTKIFVTD GLNKVIPLLQKAKNSAPATF ITYLLTCIVPLLINALRE

Example 5. Immunoreactivity of O. volvulus Proteins

[0107] Among the biomarker sequences listed in Table 3, OVOC3261, OVOC5127 and OV10469 were tested for their individual immunoreactivity using a variety of Onchocerca microfilaria positive infected sera (truly infected) and a variety of control sera (non-infected (EN_all, BB), infected with unrelated human filarial pathogens or S. stercoralis) in immunoassays. As can be seen in FIG. 3, when using a cutoff that gave 100% specificity, OVOC10469 had 78% sensitivity (FIG. 3B), OVOC3261 had 78% sensitivity (FIG. 3C), and OVOC5127 had a sensitivity of 49% (FIG. 3D). Combinations of these newly identified proteins were tested in combination with the known Ov16 (KISAENANCKKCTPMLVDSAFKEHGIVPDVVSTAPTKLVNVSYNNLTVNLGNELTPTQVKN QPTKVSWDAEPGALYTLVMTDPDAPSRKNPVFREWHHWLIINISGQNVSSGTVLSDYIGSG PRKGTGLHRYVFLVYKQPGSITDTQHGGNRRNFKVMDFANKHHLGNPVAGNFFQAKHED; SEQ ID NO:63) (Table 6), it can be seen that the sensitivity increases for all of these combination compared to Ov16 alone. Each individual positive sera was tested against each of the antigens and in combination with all four. As can be seen in Table 6, the combination gets to 97 percent sensitivity (8/245 mf positives being false negatives).

TABLE-US-00006 TABLE 6 Reactivity of Ov-infected samples with O. volvulus biomarkers Combination Ov16 + of 4 Ov16 OVOC10469 OVOC3261 OVOC5127 OVOC3261 biomarkers Positive 187 (77%)* 167 (81%) 219 (91%) 172 (70%) 225 (94%) 235 (97%) Negative 53 (23%) 40 (19%) 22 (9%) 71 (29%) 16 (6%) 8 (3%) Not tested 2 38 4 2 4 2 *number of samples (percent of samples tested)

[0108] Further analysis demonstrated that OVOC3261, OVOC10469, OVOC8491, OVOC11950, OVOC10602 are microfilaria-specific. Moreover, most of these antigens are relatively invariant based on non-synonymous SNPs and that antibodies of the IgG and IgG.sub.4 isotypes of two of these (OVOC3261 and OVOC10469) only appear after microfilariae appear in the skin of experimentally infected chimpanzees.

Example 6. Immunization of Mice with O. volvulus Proteins

[0109] Yeast codon optimized DNAs encoding for O. volvulus proteins Ov-CPI-2M (OVOC7453), Ov-103 (OVOC4230), and Ov-RAL-2 (OVOC9988), minus the signal peptides at the N-terminus, were synthesized and subsequently subcloned in-frame into the yeast expression vector pPink.alpha.-HC (Life Technologies) with XhoI/KpnI sites and E. coli expression vector pET41a (EMDMillipore) with the fusion GST deleted (NdeI/XhoI). The correct open reading frame (ORF) was confirmed by double-stranded sequencing using the vector flanking primers (5'AOX1/CYC1 for pPink.alpha.-HC and T7 promoter/T7 terminator for pET41a). For expression in yeast, the recombinant plasmids were linearized with AflII digestion and then transformed by electroporation into PichiaPink strain#4 with protease A and B knockout (pep4/prb1.sup.-) to prevent P. pastoris-derived protease degradation. Yeast transformants were selected on P. pastoris adenine dropout (PAD) selection plates. The expression of recombinant filarial antigens with hexahistidine (6His)-tag at the C-terminus was induced with 0.5% methanol and the soluble recombinant proteins secreted into the culture were purified with immobilized metal ion affinity chromatography (IMAC). For expression in E. coli, the recombinant constructs cloned into pET41a were transformed into BL21(DE3) (EMDMillipore) and recombinant proteins were induced with 1 mM isopropyl-.beta.-thiogalactoside (IPTG) and purified with IMAC.

[0110] In order to test the synergistic protection of two or three O. volvulus protective antigen combinations, protective O. volvulus antigens Ov-103, Ov-RAL-2 and Ov-CPI-2M, were fused together as a triple antigen (Ov-103-RAL-2-CPI2-M) or as two double antigens (Ov-103-RAL-2 and Ov-RAL-2-CPI2-M) by using a flexible linker (KGPDVPETNQQCPSNTGMTD; SEQ ID NO:50) obtained from Na-ASP-1 structure between two pathogenesis-related (RP) domains. The yeast codon optimized fusion DNAs were subcloned into either yeast expression vector pPICZ.alpha.A (Life Technologies) or E. coli expression vector pET41a with GST knockout. The recombinant fusion proteins were expressed and purified using the same methods described above except for the use of yeast strain P. pastoris X-33.

TABLE-US-00007 TABLE 7 SEQ ID Protein NO. Description Sequence 64 Ov-103- DLLSEAGDFFTKHFTDIKSLFAKDEKQLQQSVDRVKDLLATIQDKMSMLQ RAL2-CPI2M PLANDMQKTTLGKIGDLISQVNSFRETMSNPKMDFTNKENKWEELLKKIF fusion ##STR00001## protein QQQQQQRDEREIPPFLEGAPPSVIDEFYNLLKTDENKTDQQTEADVEAFIN RLGGSYKVRFTQFMEEVKKARADYERIHQQAVARFSPAAKDADARMSAIA ##STR00002## ##STR00003## NEQSKDEYHLMPIKLLKVSSQVVAGVKYKMDVQVARSQCKKSSNEKVDLT KCKKLEGHPEKVMTLEVWEKPWENFMRVEILGTKEV 65 Ov103- DLLSEAGDFFTKHFTDIKSLFAKDEKQLQQSVDRVKDLLATIQDKMSMLQ RAL2 PLANDMQKTTLGKIGDLISQVNSFRETMSNPKMDFTNKENKWEELLKKIF fusion ##STR00004## protein QQQQQQRDEREIPPFLEGAPPSVIDEFYNLLKTDENKTDQQTEADVEAFIN RLGGSYKVRFTQFMEEVKKARADYERIHQQAVARFSPAAKDADARMSAIA DSPHLTTRQKSQQIQAIMDSLSESVRREIINALSPQE 66 OvRAL2- PQRRQQQQQQQQQQQRDEREIPPFLEGAPPSVIDEFYNLLKTDENKTDQ CPI2M QTEADVEAFINRLGGSYKVRFTQFMEEVKKARADYERIHQQAVARFSPAA fusion ##STR00005## protein ##STR00006## LELLPSILMKVNEQSKDEYHLMPIKLLKVSSQVVAGVKYKMDVQVARSQCK KSSNEKVDLTKCKKLEGHPEKVMTLEVWEKPWENFMRVEILGTKEV Ov103 sequence - Bold; Linkers - Highlighted; OvRAL2 sequence - underlined; OVCPI-2M sequence - Italics.

[0111] The purity and the molecular weight of purified recombinant proteins were analyzed by SDS-PAGE using pre-cast 4-20% Tris-glycine gels (Life Technologies) and stained with Coomassie brilliant blue R-250 (Fisher Scientific).

[0112] Male BALB/cByJ mice were purchased from The Jackson Laboratory at 6-8 weeks of age. All mice were housed in micro-isolator boxes in a room that was pathogen-free and under temperature, humidity and light cycle controlled conditions. Mice were fed autoclavable rodent chow and given water ad libitum. All protocols using mice were approved by the Institutional Animal Care and Use Committee.

[0113] Mice were immunized with 25 .mu.g of the produced vaccine antigens (Ov-CPI-2M, Ov-103, or Ov-RAL-2, or the two- or three-antigen fusion proteins) in 0.1 ml of Tris-buffered saline (TBS) formulated with 0.1 ml of 1:5 Rehydragel LV (alum) in PBS (General Chemical). Mice were immunized s.c. in the nape of the neck, followed by two booster injections 14 and 28 days later.

[0114] The mice were challenged 14 days after the final booster as previously described (Hess et al., Int. J. Parasitology 44:637-646, 2014) with 25 L3 larvae delivered within a diffusion chamber. The diffusion chambers were implanted in a s.c. pocket on a rear flank of each mouse. Recovery of the chambers was performed 21 days later and larval survival was determined based on mobility and morphology of the remaining larvae. Protective immunity was calculated in two ways: (i) percentage of reduction in larvae was calculated as follows: reduction=((average worm survival in control mice--average worm survival in immunized mice)/average worm survival in control mice).times.100; and (ii) host protection was calculated as follows: (number of immunized mice with parasite recovery levels below the lower S.D. of parasite recovery in control mice/ total number of immunized mice).times.100). Host cells within the diffusion chamber were collected and analyzed by centrifugation onto slides using a Cytospin 3 (Shandon Inc.) and then stained for differential cell counts using Hemastain 3 (Fisher Scientific).

[0115] Serum was collected at the time of recovery for antigen-specific IgG analysis. Maxisorp 96-well plates (Nunc Nalgene) were coated with 2 .mu.g/ml of the immunizing recombinant antigen in 50 mM Tris-CI coating buffer, pH 8.8, overnight at 4.degree. C. Plates were washed with deionized water between each step. Plates were blocked with borate buffer solution (BBS) (0.17 M boric acid, 0.12 M NaCl, 0.5% TWEEN 20, 0.025% BSA, 1 mM EDTA, pH 8.2) at room temperature for 30 min. Individual sera were diluted to an appropriate starting concentration with BBS and serially diluted; plates were sealed and incubated at 4.degree. C. overnight. Biotinylated IgG (eBioscience) was diluted 1:250 in BBS and incubated for 1 hr at room temperature, followed by ExtrAvidin Px (Sigma) which was diluted 1:1000 in BBS and incubated for 30 min at room temperature. One component ABTS peroxidase substrate (KPL) was added and O.D.s were read after 30 min at 405 nm in a Bio-Rad iMark Microplate reader (Bio-Rad). ELISA data are presented as endpoint titers which were calculated as the serum dilution from experimental animals that had an O.D. reading three times higher than the O.D. recorded for control serum.

[0116] Onchocerca volvulus proteins were expressed as soluble recombinant proteins in high yield in P. pastoris and E. coli BL21(DE3) after being induced with 0.5% methanol for P. pastoris and 1 mM IPTG for E. coli, and purified with IMAC. Purified recombinant Ov-103, Ov-RAL-2 and Ov-CPI-2M expressed in P. pastoris or in E. coli migrated at the same molecular mass as calculated by the coding sequence (14.5 kDa, 17.9 kDa and 16.0 kDa, respectively) on SDS-PAGE and Coomassie staining. The fusion recombinant proteins of two or three antigen combination (Ov-103-RAL2, Ov-RAL2-CPI2M and Ov-103-RAL2-CPI2M) were also expressed in P. pastoris and E. coli expression systems as soluble proteins and the purified recombinant fusions were shown at the correct molecular weight as estimated by sequences on SDS-PAGE (50.6 kDa, 32.5 kDa and 35.2 kDa, respectively).

[0117] In BALB/cByJ mice immunized with Ov-103 with alum prepared in both P. pastoris and E. coli expression systems, E. coli-expressed protein induced an 8% reduction in larval survival and a 50% level of host protection, whereas mice immunized with the P. pastoris-expressed protein had a statistically significant 30% reduction in parasite survival and a 63% level of host protection (FIG. 4A). Differential cell counts were performed at the conclusion of the experiments on the diffusion chamber contents. Comparable numbers of total cells (1.4.times.10.sup.6.+-.1.3.times.10.sup.6), and percentages of lymphocytes (5.+-.7%), neutrophils (52.+-.20%), macrophages (37.+-.15%) and eosinophils (12.+-.14%) were seen in the control and immunized mice. Parasite-specific antibody titers show equivalent endpoint titers for mice immunized with P. pastoris and E. coli expressed Ov-103 when measured against both the P. pastoris and E. coli expressed proteins (Table 8). Correlation analyses were performed between parasite survival and antibody endpoints titers and there were no significant relationships between the amount of antibody produced and the survival of the larvae.

[0118] Mice immunized with E. coli-expressed Ov-RAL-2 induced a statistically significant 39% reduction in larval survival and a 64% level of host protection, whereas mice immunized with the P. pastoris-expressed protein induced a 24% reduction in parasite survival and a 55% level of host protection (FIG. 4B). As with Ov-103, differential cell counts showed comparable numbers of total cells, lymphocytes, neutrophils, macrophages and eosinophils in the control and immunized mice. Parasite-specific antibody titers show equivalent endpoint titers for both the P. pastoris and E. coli expressed proteins (Table 8). Again, correlations between parasite survival and antibody endpoints titers did not reveal any significant relationship between the amount of antibody produced and parasite survival.

[0119] Immunization of mice with Ov-CPI-2M expressed in both E. coli and P. pastoris induced statistically significant reductions of 30% in larval survival and 17% levels of host protection (FIG. 4C). As with the other two antigens, differential cell counts showed comparable numbers of total and specific cells in the control and immunized mice, and parasite-specific antibody titers had equivalent endpoints (Table 8). There were no significant correlations between antibody endpoint titers and parasite survival.

TABLE-US-00008 TABLE 8 Geometric mean of IgG endpoint titers following immunization with individual, fusion, or concurrent antigen formulations. Endpoint titer to antigen Ov-RAL-2/ Ov-RAL-2/ Immunizing Antigen Ov-103 Ov-RAL-2 Ov-CPI-2M Ov-RAL-2/103 CPI-2M 103/CPI-2M Ov-103 E. coli 33,064 Ov-103 P. pastoris 35,882 Ov-RAL-2 E. coli 571,055 Ov--RAL-2 P. pastoris 519,490 Ov-CPI-2M E. coli 431,803 Ov-CPI-2M P. pastoris 462,057 Ov-103/RAL-2 fusion 317,320 439,250 1,509,278 Ov-RAL-2/CPI-2M fusion 187,884 266,079 691,063 Ov-RAL-2/103/CPI-2M fusion 90,464 146,607 165,510 1,112,542 OV-RAL-2, 103, CPI-2M concurrent 16,019 271,416 392,676

[0120] In mice immunized with Ov-RAL-2/103 fusion protein expressed in P. pastoris and E. coli, E. coli-expressed protein significantly reduced larval survival by 21% and provided a 58% level of host protection, whereas immunization with P. pastoris-expressed protein only reduced larval survival by 11% and provided a 45% level of host protection (FIG. 5A). Immunization with Ov-RAL-2/CPI-2M E. coli fusion protein induced protective immunity with parasite reduction at 34% and a 50% level of host protection (FIG. 5B). Analysis of the cells within the diffusion chamber contents showed similar numbers of total cells, lymphocytes, neutrophils, macrophages and eosinophils. Parasite-specific antibody titer endpoints were measured against the individual antigens and the fusion protein. Antibody endpoint titers for the two fusion proteins were significantly higher than the antibody responses in these mice to the individual antigens of which the fusion was composed. The antibody response to Ov-RAL-2 and Ov-CPI-2M by mice immunized with these antigens as part of a fusion were equivalent to the responses seen in mice immunized with antigen individually. However, the parasite-specific antibody titer endpoint to Ov-103 was approximately eight-fold higher in mice immunized with the antigen as part of a fusion compared with immunization with the individual antigen (Table 8). Once again, there were no significant correlations between antibody endpoints and parasite survival.

[0121] A fusion protein consisting of Ov-103, Ov-RAL-2 and Ov-CPI-2M was created to determine whether enhanced protective immunity would be achieved with this triple fused antigen. The Ov-RAL-2/103/CPI-2M E. coli fusion was tested in comparison with concurrent immunization consisting of the three antigens injected simultaneously but at different locations on the mice. Immunization with the three-antigen fusion protein and the concurrent immunization resulted in significant levels of protective immunity, with the fusion inducing a 20% reduction in larval survival and a 45% level of host protection and the concurrent immunization resulting in a 25% reduction in parasite survival and a 64% level of host protection (FIG. 7). Analysis of the cells within the diffusion chamber contents showed similar numbers of total cells, lymphocytes neutrophils, macrophages and eosinophils. Antibody titer endpoints were measured against the individual antigens and the fusion protein. Mice immunized with the three antigens concurrently had antibody endpoint titers to the three antigens that were comparable with those seen in mice immunized with the three individual antigens (Table 8). Mice immunized with the three-antigen fusion protein had endpoint titers to the single antigens that were comparable with the titers seen in mice immunized with individual antigens. Antibody endpoint titers for the three-antigen fusion protein were significantly higher than the antibody responses in these mice to the individual antigens of which the fusion was composed (Table 8). There were no significant correlations between antibody endpoints and parasite survival.

Example 7. Orthologs of O. volvulus Proteins Induce Protective Immunity to Other Filarial Parasites

[0122] Brugia malayi is a filarial parasite, one of the three causative agents of lymphatic filariasis in humans. Lymphatic filariasis, also known as elephantiasis, is a condition characterized by swelling of the lower limbs. The B. malayi Bm-103 and Bm-RAL-2 proteins are orthologous to O. volvulus Ov-103 and Ov-RAL-2 which are candidates for development of an O. volvulus immunogenic composition (Table 9). The B. malayi gerbil model was used to confirm the efficacy of these O. volvulus orthologs, alone or in combination, against adult worms. Efficacy of recombinant Bm-103 and Bm-RAL-2 administered individually, concurrently, or as a fusion protein were tested in gerbils using alum as adjuvant. Immunization with Bm-103 resulted in worm reductions of 39%, 34%, and 22% on 42, 120 and 150 days post infection (dpi), respectively, and immunization with Bm-RAL-2 resulted in worm reductions of 42%, 22%, and 46% on 42, 120, and 150 dpi, respectively. Immunization with a fusion protein comprised of Bm-103 and Bm-RAL-2 resulted in improved efficacy with significant reduction of worm burden of 51% and 49% at 90 dpi, as did the concurrent immunization with Bm-103 and Bm-RAL-2, with worm reduction of 61% and 56% at 90 dpi. Immunization with Bm-103 and Bm-RAL-2 as a fusion protein or concurrently not only induced a significant worm reduction of 61% and 42%, respectively, at 150 dpi, but also significantly reduced the fecundity of female worms as determined by embryograms. Elevated levels of antigen-specific IgG were observed in all immunized gerbils. Serum from gerbils immunized with Bm-103 and Bm-RAL-2 individually, concurrently, or as a fusion protein killed third stage larvae in vitro when combined with peritoneal exudate cells.

[0123] Thus, immunization with Bm-103 and Bm-RAL-2 individually conferred protection against B. malayi infection in gerbils.

[0124] Unless otherwise indicated, all numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term "about." As used herein the terms "about" and "approximately" means within 10 to 15%, preferably within 5 to 10%. Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present invention. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0125] The terms "a," "an," "the" and similar referents used in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[0126] Groupings of alternative elements or embodiments of the invention disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in any combination with other members of the group or other elements found herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and/or patentability. When any such inclusion or deletion occurs, the specification is deemed to contain the group as modified thus fulfilling the written description of all Markush groups used in the appended claims.

[0127] Certain embodiments of this invention are described herein, including the best mode known to the inventors for carrying out the invention. Of course, variations on these described embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects skilled artisans to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[0128] Specific embodiments disclosed herein may be further limited in the claims using consisting of or consisting essentially of language. When used in the claims, whether as filed or added per amendment, the transition term "consisting of" excludes any element, step, or ingredient not specified in the claims. The transition term "consisting essentially of" limits the scope of a claim to the specified materials or steps and those that do not materially affect the basic and novel characteristic(s). Embodiments of the invention so claimed are inherently or expressly described and enabled herein.

[0129] Furthermore, numerous references have been made to patents and printed publications throughout this specification. Each of the above-cited references and printed publications are individually incorporated herein by reference in their entirety.

[0130] In closing, it is to be understood that the embodiments of the invention disclosed herein are illustrative of the principles of the present invention. Other modifications that may be employed are within the scope of the invention. Thus, by way of example, but not of limitation, alternative configurations of the present invention may be utilized in accordance with the teachings herein. Accordingly, the present invention is not limited to that precisely as shown and described.

Sequence CWU 1

1

66198PRTOnchocerca volvulus 1Asn Ile Ala Phe Ala Pro Asn Pro Lys Asp Ser Asn Asn Glu Leu Phe1 5 10 15Ala Asp Ala Glu Ser Ala Leu Gly Ser Glu Tyr Ala Gln Phe Val Glu 20 25 30Gln Ser Lys Gln His Lys Pro Val Tyr Phe Ser Asp Asn Gln Asn Thr 35 40 45Leu Glu Thr Ile Lys Leu Glu Ser Ile Pro Asn Pro Glu Thr Glu Thr 50 55 60Ala Tyr Pro Met Phe Ile Cys Gly Phe Leu Gly Cys Met Lys Lys Met65 70 75 80Asn Ser Val Glu Glu Tyr Leu Glu His Phe Lys Met His Glu Lys Gln 85 90 95Gly Tyr2103PRTOnchocerca volvulus 2Tyr Pro Thr Glu Lys Glu Thr Val Glu Pro Ile Asp Thr Met Val Lys1 5 10 15Asp Asp Ile Asp Leu Val Lys Ala Glu Val Ala Glu Ala Glu Glu Ala 20 25 30Asp Val Glu Lys Glu Val Ala Glu Leu Thr Glu Glu Glu Ala Ala Glu 35 40 45Ile Ala Glu Val Leu Asp Glu Met Glu Glu Glu Phe Phe Ala Phe Leu 50 55 60Leu Phe Asp Phe Ile Leu Asp Leu Phe Arg Glu Thr Leu Glu Lys Asn65 70 75 80Ser Glu Ser Gln Glu Ala Ser Ile Asp Glu Val Met Pro Glu Ile Gln 85 90 95Gly Val Ser Ala Glu Glu Ala 1003119PRTOnchocerca volvulus 3Phe Arg Thr Gln Ser Ile Gly Ile Arg Gly Arg Leu Met Cys Gly Ser1 5 10 15Lys Pro Ala Ser Asn Glu Arg Ile Lys Leu Trp Glu Glu Asp Ser Asp 20 25 30Asp Leu Leu Asp Gln Gly Tyr Thr Asp Glu Asn Gly Glu Phe Leu Leu 35 40 45Lys Gly Asp Thr Val Glu Leu Thr Pro Ile Asp Pro Val Phe Lys Val 50 55 60Tyr His Asp Cys Asp Asp Gly Ile Lys Pro Gly Lys Arg Lys Val Lys65 70 75 80Phe Lys Ile Pro Lys Ser Tyr Ile Thr Glu Gly Lys Thr Pro Lys Lys 85 90 95Ile Phe Asp Leu Gly Thr Leu Asn Leu Glu Thr Ile Phe Asn Asp Glu 100 105 110Glu Arg Glu Leu Ile Val Thr 1154149PRTOnchocerca volvulus 4Ser Tyr Cys Glu Asp Trp Asp Pro Glu Asp Phe Pro Ser Phe Val Leu1 5 10 15Lys Leu Ser Gln Asn Ala Thr Glu Glu Phe Cys Glu Leu Tyr Glu Met 20 25 30Glu Met Glu Val Pro Ile Asn Lys Phe Tyr Asp Met Leu Arg Lys Trp 35 40 45Ala Glu Lys Tyr Ser Val Gln Ala Glu Thr Asn Arg Phe Ile Ala Glu 50 55 60Glu Met Asn Tyr Asp Lys Met Gln Ser Lys Val Leu Met Glu Arg Leu65 70 75 80Gln Ala Ser Asn Gly Thr Thr Glu Val Lys Gly Val Leu Glu Lys Ala 85 90 95Leu Lys Leu Gln Glu Ser Met His Leu Ser Pro Asp Tyr Ile Gln Asn 100 105 110Val Ile Asp Thr Met Met Glu Asn Leu Pro Ile Asp Lys Gln Asn Glu 115 120 125Ala Thr Leu Leu Trp Asn Ser Leu Tyr Pro Asp Asp Ile Tyr Asn Glu 130 135 140Cys Gly Pro Arg Phe1455426PRTOnchocerca volvulus 5Ala Pro Pro Asn Arg Asp Thr Ala Asp Asp Leu Gln Asn Ala Asp Met1 5 10 15Gln Arg Gln Trp Glu Gln Glu Gln Arg Gln Arg Glu Glu Val Gln Lys 20 25 30Glu Glu Ile Ala Lys Tyr Val Lys Tyr Met Arg Tyr Leu Glu Thr Val 35 40 45Leu Asn Ile Leu Gln Ala Thr Pro Gln Trp Lys Glu Ala Met Gln Ser 50 55 60Met Thr Gln Glu Glu Met Arg Gly Gly Lys Ile Ala Glu Met Val Asp65 70 75 80Lys Leu Glu Pro His Ile Ile Glu Gln Leu Ala Lys Ala Lys Ile Leu 85 90 95Glu Leu Gln Arg Leu Glu Gln Glu Ile Lys Asp Gln Leu Asn Ala Asp 100 105 110Gly Gly Ala Thr His Asn Ile Lys Val Ser Glu Ile Leu Thr Val Cys 115 120 125Leu Lys Glu Gly Phe Lys Lys Pro Ser Asn Leu Gly Ile Pro Glu His 130 135 140Leu Asp Phe Asn Asn Trp Glu Thr Phe Ser Gln Glu Asp Leu Arg Lys145 150 155 160Leu Ile Val Lys Ile Val Thr Asp Met Asp Glu Leu Glu Glu Gln Arg 165 170 175Lys Gln Asp Phe Lys Gln Tyr Glu Met Lys Lys Lys Ala Glu Glu Asp 180 185 190His Lys Met Gln Ala Lys Met Ile Gln Thr Glu Arg Glu Glu Tyr Ile 195 200 205Arg Gln Met Glu Glu Gln Arg Arg Arg His Asn Lys His Glu Pro Leu 210 215 220Lys His Pro Gly Ser Arg Asn Gln Leu Arg Lys Val Trp Glu Asp Thr225 230 235 240Asp Lys Leu Asp Lys Asp Ala Tyr Asp Pro Thr Thr Leu Phe Gly Leu 245 250 255His Asp Arg Asn Asn Asp Gly Tyr Trp Ser Tyr Asp Glu Leu Asn Thr 260 265 270Ile Phe Leu Pro Glu Ile Glu Lys Leu Asn Asn Phe Ser Asp Val Glu 275 280 285Arg Leu Glu Glu Leu Tyr Arg Met Arg Asp His Val Met Lys Gln Met 290 295 300Asp Thr Asp Gly Asp His Arg Ile Ser Arg Ala Glu Phe Leu Ala Asp305 310 315 320Arg Glu Ala Gln Glu Glu Lys Pro Asp Gln Gly Trp Glu Asp Ile Gly 325 330 335Asp Lys Asp Gln Tyr Thr Lys Glu Glu Leu Glu Ile Phe Glu Lys Glu 340 345 350Tyr Ala Lys Gln Gln Gly Trp Gly Glu Tyr Ala Tyr Ser Thr Pro Ala 355 360 365Pro Thr Pro Asp Pro Ser Arg Met Ile Gln Pro Asp Gln Ala Pro Met 370 375 380Gln Arg Leu Asp Ala Pro Ser Asp Gln Val Gly Asp Met Phe Ala Gln385 390 395 400Gln Ser His Gln Ile Pro Val Lys His Val Glu Pro Ile Gln Ser Val 405 410 415Gln Gln Gln Gln Met Asp Glu Val Asn Ser 420 4256231PRTOnchocerca volvulus 6Phe Pro Thr Glu Ala Asn Pro Ala Val Gly Thr Asp Asn Ala His Glu1 5 10 15Asp Asn Leu Thr Thr Glu Glu Lys Met Gln Leu Lys Lys Phe Ala Lys 20 25 30Thr Asn Ala Ala Asn Phe Ser Leu Thr Asp Pro Glu Phe Ile Asp Gly 35 40 45Leu Lys Asn Glu Ala Ala Gly Leu Phe Ser Lys Leu Thr Gly Leu Arg 50 55 60Asp Ile Ile Asn Ala Lys Leu Asp Thr Met Gln Pro Glu Ser Arg Leu65 70 75 80Phe Ile Glu Lys Leu Leu Arg Arg Phe Leu Ala Ala Phe Ser His Asp 85 90 95Gly Leu Met Asn Ile Leu Glu Ser Leu Lys Gly Phe Gly Lys Glu Val 100 105 110Ile Asp Met Phe Asp Gly Leu Ser Arg Pro Ile Gln Asn Asp Ile Leu 115 120 125Asn Ala Phe Pro Leu Val Gly Ser Tyr Ile Thr Ser Asp Ile Ala Arg 130 135 140Leu Met Leu Arg Lys Leu Ala Glu Leu Asp Leu Leu Ser Arg Lys Ser145 150 155 160Thr Leu Thr Pro Thr Val Asp Gln Phe Asn Asp Asp Ser Gly Lys His 165 170 175Phe Pro Arg Pro Gln Val Ile Glu Pro Glu Glu Pro Glu Asn Ser Asp 180 185 190Pro Glu Asp Ala Gln Ser Thr Asp Tyr Gly Lys Lys Lys Val Val Thr 195 200 205Thr Thr Thr Phe Pro Ile Ile Thr Gly Glu Glu Asp Glu Ile Leu Val 210 215 220Lys Lys Ile Val Glu Asn Lys225 230783PRTOnchocerca volvulus 7Ile Pro Leu Pro Glu Glu Leu Asp Tyr Asp Gly Glu Ile Pro Asn Cys1 5 10 15Arg Asp Gly Glu Lys Pro Leu Leu Ala Ala Asp Ile Gly Val Tyr Thr 20 25 30Cys Asp Lys Asn Cys Pro Lys Gly Phe Arg Cys Glu Tyr Arg Thr Met 35 40 45Asp Ser Thr Ser Lys Lys Gly Ile Cys Cys Pro Asn Leu Lys Glu Leu 50 55 60Ala Lys Ile Tyr Ser Glu Asp Glu Glu Val Asp Lys Ser Ile Lys Lys65 70 75 80Ser Asn Ile8209PRTOnchocerca volvulus 8Met Ile Ser Cys Phe Ala Leu Pro Phe Pro His Val Cys Tyr Met Ala1 5 10 15Tyr Cys Thr Gln Val Ile Ala Ser Ile Met Lys Gly Trp Asn Gln Asn 20 25 30Phe Arg Phe Ser Thr Val Ile Tyr Leu Phe Arg Asn Ile Phe Ser Ser 35 40 45Ser Val Ile Ser Cys Val Asn Met Ile Leu Ser Ser Thr Phe Tyr Ala 50 55 60Leu Leu Phe Val Ser Ala Val Val Ile Val Glu Ala Met Pro Ala Ser65 70 75 80Glu Ser Thr Tyr Ser Val Ile Ile Ile Arg Ile Asn Asp Thr Thr Cys 85 90 95Lys Ile Glu Asp Gly Val Val Ser Val Asn Gly Gln Val Ile Gly Asn 100 105 110Leu Thr Glu Glu Gln Lys Glu Glu Leu Glu Ala Tyr Asn Val Gln Thr 115 120 125Gln Gly Trp Phe Gln Gln Leu His Gln Lys Ile Glu Glu Leu Phe Lys 130 135 140Thr Phe Phe Gly Ser Ile Lys Ser Met Trp Lys His Ser Pro Ile Ser145 150 155 160Gly Ser Glu Ser Ser Pro Gln Ser Ser Thr Pro Asp Asn Ile Ile Thr 165 170 175Asp Lys Leu Asp Asp Gln Asp Arg Arg Leu Lys Asp Gln Gly Asp Ser 180 185 190Glu Asn Ser Ser Leu Phe Gly Leu Lys Leu Pro Ser Phe Cys Lys Val 195 200 205Asn9128PRTOnchocerca volvulus 9Phe Arg Ser Leu Lys Ile Gly Arg Lys Gln Ser Thr Ala Val Lys Gly1 5 10 15Val Leu Thr Cys Asn Gly Lys Pro Ala Val Asn Val Lys Val Lys Leu 20 25 30Tyr Asn Asp Ser Gln Gly Arg Tyr Val Glu Asn Ser Met Asp Glu Gly 35 40 45Lys Thr Asp Ser Glu Gly Arg Phe Leu Leu Gln Gly His Glu Thr Ser 50 55 60Ile Thr Ser Ile Asp Pro Ile Leu Lys Leu Tyr His Asn Cys Asp Val65 70 75 80Glu Asn Ala Gln Cys Leu Lys Arg Phe Ser Ile Leu Ile Pro Asn Asp 85 90 95Phe Val Ser Glu Gly Leu Glu Pro Lys Lys Thr Phe Asp Met Gly Thr 100 105 110Leu Asn Leu Gly Gly Lys Phe Phe Asp Glu Gly Arg Glu Cys Ala Ser 115 120 12510256PRTOnchocerca volvulus 10Gln Ile Ile Gly Ser Phe Asn Gly Asn Tyr Ala Gly Asp Gly Ser Leu1 5 10 15Asn Asn Asn Ala Asn Ser Phe Gly Glu Arg Thr Thr Thr Thr Arg Ser 20 25 30Thr Ser Arg Pro Ser Leu Pro Pro Arg Pro Gly Tyr Pro Ser Arg Pro 35 40 45Gly Tyr Pro Phe Lys Pro Gly Phe Pro Pro Arg Gly Pro Pro Ile Pro 50 55 60Tyr Pro His Gly Lys Pro Ser Gly Pro Arg Tyr Pro Cys Tyr Gly Gly65 70 75 80Tyr Gly Gly Tyr Gly His Pro Gly Tyr Gly Pro Phe Gly Gly Asn Gly 85 90 95Tyr Leu Gly Tyr Thr Val Cys Ser Gly Arg Gly Glu Phe Gly Gly Tyr 100 105 110Gly Pro Gly Leu Gly Gly Gly Thr Gly Leu Gly Gly Leu Gly Pro Gly 115 120 125Glu Phe Gly Gly Tyr Gly Pro Gly Leu Gly Gly Gly Thr Gly Leu Gly 130 135 140Gly Leu Gly Pro Gly Gly Phe Gly Gly Ile Gly Pro Gly Leu Gly Gly145 150 155 160Gly Gly Gly Leu Gly Gly Pro Gly Arg Gly Gly Phe Ala Gly Tyr Gly 165 170 175Pro Gly Leu Gly Gly Gly Arg Gly Leu Gly Gly Pro Gly Pro Gly Gly 180 185 190Phe Asp Gly Tyr Gly Pro Gly Leu Gly Gly Arg Pro Tyr Pro Gly Gly 195 200 205Tyr Gly Arg Phe Tyr Gly Pro Gly Pro Tyr Pro Gly Asp Arg Leu Asp 210 215 220Pro Arg Gly Leu Ser Glu Ser Gly Arg Pro Arg Thr Arg Leu Ala Ser225 230 235 240Tyr Asn Arg Asn Asp Arg Gly Thr Gln Phe Ser Tyr Ile Arg Asp Arg 245 250 25511280PRTOnchocerca volvulus 11Met Thr Asn Glu Tyr Glu Thr Asn Tyr Pro Val Pro Tyr Arg Ser Lys1 5 10 15Leu Thr Glu Ser Phe Glu Pro Gly Gln Thr Leu Leu Val Lys Gly Lys 20 25 30Thr Ala Glu Asp Ser Val Arg Phe Thr Ile Asn Leu His Asn Thr Ser 35 40 45Ala Asp Phe Ser Gly Asn Asp Val Pro Leu His Ile Ser Val Arg Phe 50 55 60Asp Glu Gly Lys Ile Val Phe Asn Thr Phe Ser Lys Gly Glu Trp Gly65 70 75 80Lys Glu Glu Arg Lys Ser Asn Pro Tyr Lys Lys Gly Asp Asp Ile Asp 85 90 95Ile Arg Ile Arg Ala His Asp Ser Lys Tyr Thr Ile Tyr Val Asp Gln 100 105 110Lys Glu Val Lys Glu Tyr Glu His Arg Val Pro Leu Ser Ser Val Thr 115 120 125His Phe Ser Ile Asp Gly Asp Val Leu Val Thr Tyr Ile His Trp Gly 130 135 140Gly Lys Tyr Tyr Pro Val Pro Tyr Glu Ser Gly Leu Ser Gly Glu Gly145 150 155 160Leu Val Pro Gly Lys Ser Leu Leu Ile Phe Ala Thr Pro Glu Lys Lys 165 170 175Gly Lys Arg Phe His Ile Asn Leu Leu Lys Lys Asn Gly Asp Ile Ala 180 185 190Leu His Phe Asn Pro Arg Phe Asp Glu Lys Ala Ile Val Arg Asn Ser 195 200 205Leu Ile Ala Gly Glu Trp Gly Asn Glu Glu Arg Glu Gly Lys Met Ile 210 215 220Leu Glu Lys Gly Ile Gly Phe Asp Leu Glu Ile Lys Asn Glu Glu Tyr225 230 235 240Ala Phe Gln Ile Phe Ile Asn Gly Glu Arg Tyr Ala Thr Tyr Ala His 245 250 255Arg Leu Asp Pro Arg Glu Ile Asn Gly Leu Gln Ile Gly Gly Asp Leu 260 265 270Glu Val Ser Gly Ile Gln Met Arg 275 28012141PRTOnchocerca volvulus 12Asp Leu Leu Ser Glu Ala Gly Asp Phe Phe Thr Lys His Phe Thr Asp1 5 10 15Ile Lys Ser Leu Phe Ala Lys Asp Glu Lys Gln Leu Gln Gln Ser Val 20 25 30Asp Arg Val Lys Asp Leu Leu Ala Thr Ile Gln Asp Lys Met Ser Met 35 40 45Leu Gln Pro Leu Ala Asn Asp Met Gln Lys Thr Thr Leu Gly Lys Ile 50 55 60Gly Asp Leu Ile Ser Gln Val Asn Ser Phe Arg Glu Thr Met Ser Asn65 70 75 80Pro Lys Met Asp Phe Thr Asn Lys Glu Asn Lys Trp Glu Glu Leu Leu 85 90 95Lys Lys Ile Phe Val Thr Glu Gly Leu Asn Lys Val Ile Pro Leu Leu 100 105 110Gln Lys Leu Lys Asn Ser Ala Pro Thr Thr Phe Ala Thr Tyr Leu Phe 115 120 125Thr Cys Ile Val Pro Val Leu Ile Asn Thr Leu Arg Glu 130 135 14013114PRTOnchocerca volvulus 13Met Ala Arg Ile Asn Arg Leu Asn Phe Leu Leu Cys Ile Val His Ala1 5 10 15Asn Ile Thr Ser Ala Pro Asn Pro Lys Asp Ser Asn Asp Glu Leu Phe 20 25 30Ala Asp Ala Glu Ser Ala Leu Gly Ser Glu Tyr Ala Gln Phe Val Glu 35 40 45Gln Ser Lys Gln His Lys Pro Val Tyr Phe Ser Asp Asn Gln Asn Thr 50 55 60Leu Glu Thr Ile Lys Leu Glu Ser Ile Pro Asn Pro Glu Thr Glu Thr65 70 75 80Ala Tyr Pro Met Phe Ile Cys Gly Phe Leu Gly Cys Met Lys Lys Met 85 90 95Asn Ser Val Glu Glu Tyr Leu Glu His Phe Lys Met His Glu Lys Gln 100 105 110Gly Tyr1469PRTOnchocerca volvulus 14Phe Ser Trp Lys Phe Gly Glu Arg Leu Asp Glu Pro Val Leu Met Leu1 5 10 15Arg Asp Leu Arg Ala Lys Glu Ile Ser Pro Pro Ser Tyr Met Lys Arg 20 25 30Phe Glu Ser Asp Thr Asn Glu Gln Leu Leu Arg Tyr Ile Leu His Pro 35 40 45Lys Met Leu Arg Arg His Asp Leu Ser Asn Ala Leu Phe Tyr Gln Pro 50 55 60Leu Trp Lys Met Arg6515278PRTOnchocerca volvulus 15Met Glu Gly Ser Pro Ile Lys Glu Thr Arg Gly Leu Glu Ala Thr Pro1 5 10 15Val Phe Glu Met Val Arg Ser Ala Thr Leu Thr Phe Leu Leu Ala

Val 20 25 30Ser Thr Val Leu Val Val Ser Arg Pro Asn Val Leu Leu Pro Pro Lys 35 40 45Leu Pro Trp Asp Ser Asp Trp Arg Gln Lys Pro Pro Pro Phe Pro Pro 50 55 60Glu Pro Pro Pro Glu Phe Lys Gly Ile Leu Pro Pro Glu Ile Phe Ala65 70 75 80Lys Leu Thr Ala Ile His Gln Asp Gln Ser Leu Thr Ile Pro Gln Lys 85 90 95Ile Val Lys Ile Glu Glu Ile Met Asn Ser Leu Pro Glu Asp Val Leu 100 105 110Gln Arg Leu Pro Leu Pro Pro Val Phe Arg Leu Leu Pro Gln Asn Val 115 120 125Gln Glu Met Ile Lys Thr Val Arg Thr Thr Lys Asn Leu Thr Met Glu 130 135 140Glu Lys Trp Leu Gln Met Ile Ile Leu Ile Glu Ser Leu Pro Lys Gln145 150 155 160Gln His Arg Leu Leu Gln Gln Met Leu Pro Lys Phe Ser Leu Gly Pro 165 170 175Leu Pro Asp Phe Gln Asp Ile Ile Pro Lys Glu Asp Trp Asp Lys Leu 180 185 190Thr Ala Val Tyr Gln Asp Thr Asn Leu Asp Asn Ile Glu Lys Leu Arg 195 200 205Arg Val Asp Glu Ile Ile Asp Ala Leu Pro Asp Ser Ile Arg Gln Lys 210 215 220Ile Pro Leu Ser Pro Pro Phe Gln Lys Leu Pro Asp His Ile Gln Gln225 230 235 240Gln Leu Gln Ile Ile His Thr Glu Arg Gly Leu Thr Thr Glu Gln Arg 245 250 255Phe Arg Lys Met Lys Ala Ile Ile Glu Ser Leu Pro Trp Asp Met Lys 260 265 270Lys Leu Met Phe Gln Pro 27516425PRTOnchocerca volvulus 16Leu Ile Lys Val Phe Pro Glu Ile Ser Ala Asn Met Ser Val Met Phe1 5 10 15Ala Asn Ser Arg Ser Asn Gln Ala Asn Asn Gly Tyr Leu Val Glu Phe 20 25 30Lys Ala Gly Arg Ser Asn Leu Gln Ala Gly Ser Thr Val Asp Arg Arg 35 40 45Lys Val Val Ala Asp Lys Thr Lys Gly Leu Val Phe Ile Lys Gln Ser 50 55 60Ser Asp Gln Leu Met His Phe Cys Trp Lys Asn Arg Glu Thr Gly Ala65 70 75 80Val Val Asp Asp Leu Ile Ile Phe Pro Gly Asp Thr Glu Phe Leu Arg 85 90 95Val Arg Glu Cys Thr Asp Gly Arg Val Tyr Met Leu Lys Phe Lys Ser 100 105 110Thr Asp Glu Lys Arg Leu Phe Trp Met Gln Asp Gly Lys Thr Asp Lys 115 120 125Asp Asp Glu Asn Cys Lys Lys Val Asn Glu Thr Leu Asn Asn Pro Pro 130 135 140Ala Pro Arg Ala Ala Ala Arg Gly Gly Ala Asp Arg Ala Asp Val Ser145 150 155 160Ser Phe Gly Thr Leu Ala Ala Leu Gly Ser Ala Gly Ala Glu Ser Glu 165 170 175Leu Gly Ala Leu Gly Asn Leu Asp Gln Ser Gln Leu Met Gln Leu Leu 180 185 190Ser Leu Met Asn His Thr Asn Ser Thr Ser Ala Ser Glu Ala Thr Asn 195 200 205Leu Leu Pro Gln Leu Pro Leu Val Ala Asp Thr Ser His Pro Met Thr 210 215 220Ser Glu Asp Ser Gly Thr Thr Ser Thr His Gly Ala Thr Pro Ser Asn225 230 235 240Thr Pro Ala Asn Gly Ile Val Ala Asp Ser Ser Ser Asn Asn Ala Met 245 250 255Gln Leu Ser Gln Leu Lys Glu Ile Ile Ala Ser Ile Thr Pro Pro Asp 260 265 270Gly Ser Gly Arg Lys Pro Ser Ile Asp Phe Thr Asp Val Leu Cys Cys 275 280 285Ala Asp Lys Ile Asn Asp Val Leu Arg Lys Tyr Ala Glu Gln Leu Ile 290 295 300Pro His Leu Pro Ser Gln Glu Pro Ile Tyr Asn Asn Gln Glu Glu Leu305 310 315 320Gln Gln Thr Leu Arg Thr Pro Gln Phe Arg Gln Ala Ala Asp Ile Phe 325 330 335Gly His Ala Leu Gln Thr Gly Gln Leu Ala Pro Val Leu Arg Gln Phe 340 345 350Gly Ile Asp Gly Asn Thr Ala Thr Ala Ala Gly Asn Gly Asp Met Val 355 360 365Ala Trp Ala Ala Gln Phe Thr Thr Ala Glu Asn Gly Lys Glu Ile Thr 370 375 380Ala Lys Thr Glu Thr Ser Pro Ser Gln Pro Gly Met Glu Ser Asp Val385 390 395 400Glu Asp Glu Glu Thr Asn Glu Lys Ala Ile Arg Glu Thr Glu Lys Asn 405 410 415Arg Thr Asp Asp His Met Asp Leu Asp 420 4251790PRTOnchocerca volvulus 17Met Asn Tyr Lys Ala Pro Ile Glu Leu Gln Gln Leu Leu Ser Ile Thr1 5 10 15Lys Met Leu Ser Leu Ser Val Leu Leu Leu Phe Thr Ser Met Ala Ile 20 25 30Met Ala Arg Pro Pro Asn Ser Asp Glu Ile Lys Glu Leu Arg Gln Gln 35 40 45Gln Leu Asn Glu Ser Lys Asp Asp Tyr Asp Thr Leu Pro Asp Val Asn 50 55 60His Ile Pro Glu Ser Phe Lys Glu Ser Leu Lys Lys Gln Lys Met Leu65 70 75 80Tyr Leu Asp Met Leu Arg Gln His Asn Leu 85 9018257PRTOnchocerca volvulus 18Leu Ser Val Pro Ala Gly Leu Arg Pro Ala Lys Lys Val Gly Asp Pro1 5 10 15Lys Glu Gln Ile Val Pro Gly Lys Gln Gln Gln Leu Gln Gln Gln Gln 20 25 30Gln Gln Gln Leu Leu Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 35 40 45Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 50 55 60Gln Gln Gln Glu Gln Gln Glu Glu Leu Gln Gln Leu Gln Glu Ser Glu65 70 75 80Glu Thr Gly Glu Glu His Arg Gln Gln Gln Gln Gln Gln His Asp Glu 85 90 95Ala Leu Thr Leu Ser Pro Thr Pro Lys Val Pro Pro Asn Leu Ser Ile 100 105 110Arg Ser Arg Met Met Ala Ala Leu Ser Ala Ser Val Gly Glu Ser Asn 115 120 125Lys Glu Lys Asn Ser Ser Asn Asp Glu Thr Asp Asn Ser Ser Lys Ser 130 135 140Thr Asn Ser Pro Ser Lys Pro Pro Ile Ile Phe Pro Lys Ala Asn Lys145 150 155 160Lys Thr Val Val Gly Lys Ile Ala Pro Ser Gly Ile Ser Lys Gly Ser 165 170 175Ala Arg Val Ile Val Ala Pro Pro Ser Lys Leu Gly Thr Asn Asn Phe 180 185 190Gly Leu Asn Thr Val Leu Gln Thr Asn Leu Val Asp Ser Arg Gly Arg 195 200 205Ile Met Lys Asn Val Asn Ser Val Pro Ile Lys Val Pro Ser Ser Ala 210 215 220Glu Met Arg Asn Ala Arg Thr Arg His Thr Ala Arg Gln Val Glu Ser225 230 235 240Asp Ala Asp Lys Val Val Pro Ile Lys Phe Gly Ser Thr Ser Arg Arg 245 250 255Arg19130PRTOnchocerca volvulus 19Asn Asn Ser Asn Leu Asp Ile Ser Met Arg Glu Lys Asn Ala Val Asn1 5 10 15Ala Ile Glu Lys Gln Asp Leu Pro Arg Ser His Arg Phe Lys Arg Gln 20 25 30Tyr Ser Cys Gly Gln Cys Gly Gly Gly Gly Gly Pro Pro Val Val Val 35 40 45Ser Pro Cys Gln Gln Cys Lys Gly Gly Gly Ala Gly Val Ser Ala Ile 50 55 60Gly Gly Ala Gly Gly Ile Ser Ala Ile Gly Gly Gly Val Ser Ala Ile65 70 75 80Gly Gly Gly Phe Gly Gly Gly Gly Gly Asp Thr Val Ala Val Val Cys 85 90 95Cys Gly Ala Thr Gly Leu Lys Gly Met Phe Arg Asn Trp Trp Leu His 100 105 110Ile Pro Leu Leu Leu Leu Pro Met Ser Met Ser Trp Ile Lys Ala Leu 115 120 125Phe Leu 13020185PRTOnchocerca volvulus 20Tyr Tyr Val Pro Asp Asn Tyr Trp Pro Leu Arg Ile Ile Gly Tyr His1 5 10 15His Ile Pro Val Met Ile Asn Met Trp Tyr Leu Phe Gln Thr Glu Ile 20 25 30Ser Asn Ile Gly Val Asp Ala Val Leu Val Gln Ser Pro Leu Tyr Arg 35 40 45Thr Leu Thr Pro Asp Val Val His Asp Ile Ile Ser Ile Asn Val Glu 50 55 60Pro Asn His Thr Val Val Val Glu Gln Ser Asn Pro Met Leu Gln Ala65 70 75 80Ser Ser Val Glu Gln Ala Pro Ala Ala Ala Pro Leu Ser Ile Thr Leu 85 90 95Ile Ala Pro Gly Ile Thr Ile Ser Arg Thr His Lys Val Asp Thr Tyr 100 105 110Lys Ser Thr Met Glu Met Tyr Asp Ala Asp Lys Leu His Ser Asn Glu 115 120 125Ile Phe Lys Arg Arg Val Arg Lys Met Val Leu Pro Pro Ser Arg Gly 130 135 140Glu Glu Val Arg Lys Pro Pro Ser Ser Thr Asp Gly Tyr Glu Ser Glu145 150 155 160Asn Val Glu Ser Tyr Gly Gln Lys Gly Val Glu Gln Ala Pro Pro Glu 165 170 175Ile Glu Gln Tyr Val Lys Lys Lys Lys 180 18521633PRTOnchocerca volvulus 21Met Lys Tyr Cys Leu Ser Ser Ile Ile Ala Ala Thr Ile Ala Thr Thr1 5 10 15Thr Thr Thr Ala Thr Ala Ile Ile Ala Thr Thr Ile Thr Ala Ala Thr 20 25 30Ile Ser Val Ala Pro Phe His Ala Ser Ser Pro Ser Ser Ser Leu Ser 35 40 45Ser Ser Ser Phe Ser Ser Phe Phe Leu Val Leu Pro Leu Ile Thr Thr 50 55 60Ile Leu Leu Ile Val Pro Glu Gln Ala His Ser Thr Ala Thr Val Thr65 70 75 80Glu His Arg Ser Pro Pro Asp Leu Ser Ile Pro Ser Gln Thr Glu Phe 85 90 95Arg Val Pro Val Gly Thr Lys Gln Phe Arg Leu Ile Cys Pro Val Lys 100 105 110Glu Lys Asn Asp Asp Leu Leu Met Ile Gln Trp Lys Lys Asn Asp Glu 115 120 125Pro Ile Gly Phe Asp Phe Asn Asn Arg Phe Lys Leu Ala Arg Ser Asp 130 135 140Arg Glu Leu Lys Ile Arg Asn Pro Gln Leu Ser Asp Gly Gly Ile Tyr145 150 155 160Gln Cys Gln Val Val Asn Gly Phe Gly His Arg Glu Leu Asn Phe Thr 165 170 175Val Thr Phe Tyr Asp Pro Ala Met Glu Asn Asp Gln Asn Thr Asp Ser 180 185 190Thr Leu Thr Leu Thr Thr Lys Ala Ser Pro Pro Ile Trp Lys Asn Glu 195 200 205Thr Glu Ile Arg Asn Trp Met Ile Asn Pro Val Arg Ile Thr Ile Gly 210 215 220Gly Ala Leu Leu Leu Lys Cys Pro Ala Lys Gly Asn Pro Leu Pro His225 230 235 240Ile Thr Trp Leu Arg Asp Gly Lys Val Leu Glu Arg Glu Ile Thr Tyr 245 250 255His Tyr Ser Ser Ala Ile Leu Tyr Leu Ser Asp Val Gln Pro Ser Glu 260 265 270Gly Gly Lys Tyr Ile Cys Lys Leu Glu Asn Glu His Gly Ser Ile Glu 275 280 285Ala Ser Phe His Val Tyr Val Glu Asn Phe Phe Glu Gly Leu Asp Gly 290 295 300Glu Ser Trp Ser Ile Asp Gln Thr Asn Ala Gln Leu Tyr Pro Val Ile305 310 315 320Asp Glu Pro Phe Asn Asn Thr Val Arg Val Gly Arg Thr Ala Gln Phe 325 330 335Gln Cys Lys Val Lys Asn Gln Gln Gln Pro Leu Ile Lys Trp Leu Lys 340 345 350Arg Val Glu Asp Pro Asn Ala Ile Arg Gln Thr Asn Ala Asn Ala Thr 355 360 365Leu Ile His Ala Asn Asn Met His Leu Leu Leu Leu Glu Lys Pro Glu 370 375 380Thr Ser Ala Glu Leu Ser Asp Gly Ile Ser Leu Asn Arg Leu Ile Ile385 390 395 400Pro Asn Val Arg Tyr Glu His Ser Gly Thr Tyr Leu Cys Val Val Thr 405 410 415Asn Ala Arg Gly Asp Ile Ala Tyr Arg Ser Ala Tyr Leu Asn Val Ile 420 425 430Ala Arg Ser Asp His Gly Glu Leu Ser Asn Leu Tyr Phe Tyr Gly Gly 435 440 445Leu Leu Val Leu Ile Val Val Phe Thr Leu Ile Thr Tyr Ala Val His 450 455 460Phe Leu Arg Lys Asn Gln Ala Ala Lys Ser Thr Glu Ser Ala Pro Gly465 470 475 480Ile Thr Asn Ile Arg Tyr Ser Phe Ser Leu Arg Pro Pro Pro Pro Asn 485 490 495Leu Pro Pro Pro Lys Ala Pro Ala Leu Pro Ser Glu Arg Gln Gln Leu 500 505 510Met Pro Asn Asn Gln Pro Cys Asp Arg Tyr Thr Val Asn Ser Ala Ala 515 520 525Ala Thr Tyr Tyr Pro Gln Phe Ala Thr Pro Asp Lys Lys Leu Gln Lys 530 535 540Ile Ile Thr Glu Ser Gly Thr Arg Pro Thr Pro Ile Arg Arg Thr Asn545 550 555 560Gly Gly Asp Thr Lys Tyr Arg Leu Lys Asp Asp Tyr Ile Ser Ser Pro 565 570 575Lys Trp Val His Ala Lys Gly Asp Asn Ile Glu Val Glu Met Asp Gln 580 585 590Asn Leu Leu Lys Asn Arg Ser Thr His Cys His Asn Pro Val Ser Ile 595 600 605Ala Tyr Gly Arg Ile Asp Asn Ile Asp Arg Gln Gln Gln Lys Ser Phe 610 615 620Leu Thr Ile Gly Asn Leu Gln Lys Arg625 63022189PRTOnchocerca volvulus 22Lys Glu Ile Ile Trp Asp Cys Tyr Gly Asp Tyr Glu Glu Cys Val Ala1 5 10 15Glu Ser Ser Lys Met Asp His Val Asp Val Asn Asn Val Glu Ser Arg 20 25 30Asn Ile Ile Glu Phe Cys Ser Asp His Thr Gln Asn Ile Leu Pro Cys 35 40 45Leu Ala Thr Lys Leu Gly Leu Ile Lys Ser Met Ser Val Ser Met Phe 50 55 60Ser Leu Leu Leu Thr Ile Cys Glu Ala Glu Thr Arg Asn Asn Arg Pro65 70 75 80Ala Ala Thr Glu Val Gln Gln Ile Leu Lys His Leu Ala Arg Leu Tyr 85 90 95Ala Tyr Phe Cys Ala Tyr Ser Asn Val Ile Asp Leu Arg Tyr Asn Lys 100 105 110Glu Cys Phe Arg Tyr Leu Lys Lys Arg Cys Ile Leu Asn Lys Pro Asp 115 120 125Asp Ser Cys Ile Phe His His Cys Gly Glu Lys Asn Leu Asn Leu Ser 130 135 140Glu Ser Ser Pro Phe Ile Gln Gln His Lys Thr Thr Ile Ile Asn Gln145 150 155 160Leu Asn Gln Ser Ala Thr Phe Lys Asn Tyr His His Arg Ile Thr Thr 165 170 175Ile Phe Thr Val Ile Ile Thr Phe Ile Ser Met Ile Gln 180 18523399PRTOnchocerca volvulus 23Met Tyr Asn Gln Glu Asn His Asp Lys Arg Arg Asn Asp Asp Arg Phe1 5 10 15Ile Leu Ser Leu Pro Phe Gly Thr Asn Val Glu Asn Lys Ser Tyr Phe 20 25 30Lys Pro Ile Lys Leu Ser Asn Pro Tyr Ser Asp Lys Tyr Leu Glu Val 35 40 45Asn Lys Lys Ser Ser Asp Asp Ser Asp Gln Asn Leu Asn Gln Ala Leu 50 55 60Ser Val Pro Gln Ser Asn Tyr Asp Gln Ser Ser Glu Ser Leu Ser Ile65 70 75 80Asp Asp Ser Asp Leu Ile Asp Asp Ser Thr Ser Ala Ala Gln Leu Ser 85 90 95Thr Ser Ser Pro Ile Ser Val Thr Ser Ala Ser Thr Ser Ser Phe Tyr 100 105 110Pro Thr Leu Asn Ile Gly Asn Gly Met Glu Ile Ser Ala Lys Tyr Ala 115 120 125Lys Leu Glu Gln Ser Gln Gly Ile Lys Ser Asp Gln Ser Thr Ser Arg 130 135 140Val Ser Asp Arg Tyr Lys Lys Tyr Thr Ala Val Lys Arg Arg Leu Ser145 150 155 160Glu Leu Tyr Gly Ile Ile Glu Glu Lys Asp Glu Gln Leu Arg Val Val 165 170 175Arg Asn Glu Leu Asn Gly Lys Asp Leu Glu Ile Gly Lys Leu Cys Asp 180 185 190Lys Ile Arg Ala Leu Glu Tyr Asn Cys Gly Arg Leu Gln Ser Met Ile 195 200 205Glu Ser Ala Gly Asp Glu Ser Asp Gln Asn Gln Val Lys Leu His Glu 210 215 220Ile Ile Asn Glu Arg Asp Gly Leu Leu Ile Arg Asn Ala Ser Leu Ser225 230 235 240Arg Gln Ile Glu Phe Glu Lys Arg Glu Trp Ser Ile Glu Arg Glu Arg 245 250 255Leu Ser Met Asp Leu Asp Asp Val Thr Arg Glu Leu Glu Leu Gln Lys 260 265 270Met Ile Leu Asn Gly Glu Ser Ile Ser Glu Ile Val Gln Arg Trp Gln 275 280

285Thr Lys Val Phe Glu Leu Glu Gly Met Ile Thr Asp Arg Asp Arg Ala 290 295 300Ile Arg Ala Gln Gln Val Gln Ile Ser Lys Leu Lys Glu Ser Ile Ala305 310 315 320Glu Thr Asp Arg Ile Ser Cys Ala Asp Ser Ser Glu Ser Gln Thr Lys 325 330 335Phe Asp Phe Pro Ser Phe Thr Tyr Ile Lys Arg Leu Leu Leu Gln Tyr 340 345 350Leu Thr Arg Leu Ala Asp Leu His Phe Ser Ser Asp Glu Glu Arg Met 355 360 365Gln Leu Val Arg Asn Met Ser Ser Ile Leu His Leu Ser Asp Glu Glu 370 375 380Gln Arg Gln Val Trp Ala Asn Leu Lys Ser Lys Ile Gln Ile Ser385 390 39524301PRTOnchocerca volvulus 24Gln Cys Pro Thr Gly Ser Val Ser Leu Leu Ser Gly Tyr Arg Cys Thr1 5 10 15Ser Ser Ile Gln Cys Gln Thr Ile Ile Pro Gly Ser Tyr Cys Tyr Tyr 20 25 30Gly Val Cys Cys Thr Gly Gly Ser Asp Val Leu Ser Lys Thr Val Ser 35 40 45Tyr Gly Gly Tyr Cys Thr Met Thr Val Gln Cys Ser Thr Thr Gly Ala 50 55 60Thr Cys Ile Ser Asn Ile Cys Gln Cys Asp Ile Asn Ser His Tyr Asn65 70 75 80Gly His Ser Cys Val Ser Ile Ser Asn Phe Cys Pro Ser Asn Gln Val 85 90 95Phe Ile Lys Gly Glu Cys Tyr Arg Lys Val Thr Tyr Gly Phe Leu Cys 100 105 110Asn Tyr Thr Gln Gln Cys Gly Tyr Ile Gly Ala Phe Cys Ile Gly Asn 115 120 125Ile Cys Ser Cys Gln Leu Asp Tyr Thr Phe Asp Gly Ser Lys Cys Ile 130 135 140Pro Arg Ser Arg Ile Cys Pro Ala Asn Gln Ile Ala Ile Gly Gly Gln145 150 155 160Cys Tyr Pro Ser Ala Arg Phe Gly Glu Arg Cys Leu Tyr Ser Glu Gln 165 170 175Cys Ile Asp Arg Trp Tyr Arg Ser Leu Ser Cys Val Asn Gly Phe Cys 180 185 190Asn Ile Arg Asn Asp Asp Asp Ile Ser Lys Pro Lys Cys Arg Asn Pro 195 200 205Arg Ala Glu Val Glu Tyr Val Asn Gly Thr Ala Lys Asn Cys Leu Tyr 210 215 220Trp Pro Cys Thr Val Gly Tyr Phe Cys Glu Tyr Ala Gly Gly Met Asn225 230 235 240Gly Gly Arg Tyr Ile Cys Cys Gly Thr Asn Ala Asn Lys Ile Tyr Gly 245 250 255Lys Val Gln Leu Tyr Pro Gly Thr Gly Thr Pro Leu Gln Cys Thr Glu 260 265 270Ile Gly Arg Cys Pro Phe Pro Asp Thr Pro Asn Cys Val Met Ser Tyr 275 280 285Arg Tyr Gly Tyr Lys Val Cys Cys Ser Thr Leu Asn Cys 290 295 30025110PRTOnchocerca volvulus 25Gln Glu Thr Ser Glu Gln Pro Gly Leu Thr Val Glu Ile Ile Ala Glu1 5 10 15Gln Gln Asp Ala Thr Thr Ala Asp Gln Glu Val Thr Thr Thr Val Asp 20 25 30Thr His His Gln His Gln His Gln Thr Asp Lys Val Val Lys Ser Arg 35 40 45Gln Ile Thr Gly Asp Glu Gln Thr Thr Thr Thr Thr Thr Ala Ile Asn 50 55 60Leu Asn Glu Thr Ile Thr Asn Ser Thr Thr Asp Ser Asn Ser Thr Ile65 70 75 80Ile Thr Thr Thr Leu Asp Leu Gln Glu Ser Thr Thr Thr Gly Thr Thr 85 90 95Asp Asn His His His His His His His His His His His Glu 100 105 11026185PRTOnchocerca volvulus 26Met Lys Gln Thr Thr Ala Trp Gly Asn Ala Leu Cys Val Leu Cys Asn1 5 10 15Cys His Gln Pro Gln Ile Ile Cys Pro Pro Pro Pro Pro Ala Val Cys 20 25 30Pro Arg Val Val Cys Pro Pro Pro Arg Pro Pro Val Cys Pro Pro Ile 35 40 45Tyr Cys Pro Pro Pro Val Val Cys Pro Pro Pro Pro Val Cys Pro Pro 50 55 60Val Pro Phe Cys His Ser Gln Ile Cys Pro Pro Cys Gly Thr His Thr65 70 75 80Val Pro Val Ala Val Val Gly Cys Cys Lys Gly Cys Ala Cys Ser Val 85 90 95Arg Phe Lys Arg Asp Ser Ser Ser Val Asn Gly Leu Met Leu Lys Lys 100 105 110Asn Leu Leu Cys Asn Asn Asp Gln Leu Met Thr Ile Met Glu Lys Lys 115 120 125Ile Gly Thr Asn Ala Thr Glu Ala Ala Phe Ala Ile Lys Lys Glu Ala 130 135 140Asp Ser Glu Leu Lys Ala Lys Phe Ser Val Phe Cys Ala Met Asn Asp145 150 155 160Leu Ile Tyr Val Ala His Ala Glu Ser Phe Cys Gln His Lys Lys Gly 165 170 175Asp Ile Ile Cys Phe Ala Tyr Lys Ser 180 18527147PRTOnchocerca volvulus 27Met Asp Cys Lys Leu Ile Leu Pro Phe Tyr Ile Leu Leu Ala Asn Leu1 5 10 15Glu Ala Asn Ala Phe His Leu Ser Gly Tyr Arg Ser Arg Ser Tyr Leu 20 25 30Gln Gly Ile Gln Pro Tyr Asp Ile Gln Pro Leu Asp Val Gln Pro Gln 35 40 45Phe Ile Arg Val Gln Thr Leu Lys Ser Gln Asp Ile Gln Pro Tyr Ser 50 55 60Ile Gln Ser Arg Ser Glu Asp Gln Pro Cys Glu Gly Cys Lys Ile Thr65 70 75 80Ile Ser Cys Gly Ser Lys Asn Cys Lys Ser Lys Lys Leu Pro Tyr Val 85 90 95Tyr Lys Pro Ile Phe Lys Leu Leu Ser Thr Arg Ser Thr Lys Lys Pro 100 105 110Val Phe Thr Leu Pro Thr Gln Pro Pro Ala Gln Trp Asp Cys Pro Cys 115 120 125Pro Cys His Val Pro Gln Arg Cys Arg Met Cys Ser Ala Cys His Glu 130 135 140Ser Tyr Ile14528194PRTOnchocerca volvulus 28Asn Arg Ile Ile Ser Arg Arg Leu Ser Leu Phe Ile Gln Gln Tyr Cys1 5 10 15Cys Asn Asn Ile Ser Gln Ile Tyr Arg Leu Asn Asp Cys Lys Tyr Ser 20 25 30Lys Val Lys Met Glu Ile Asp Lys Lys Ile Phe Ile Ile Val Ser Lys 35 40 45Thr Glu Trp Cys Asn Glu Ala Ile Lys Val Val Phe Gly Lys Ser Ala 50 55 60Glu Ala Ile Arg Asn Asn Ser Asp Ala Ile Ser Trp Leu Ala Ser Tyr65 70 75 80Asn Tyr Thr Gly Ser Met Asp Leu Arg Ser Lys Trp Pro Tyr Asp Ala 85 90 95Tyr Phe Asp Asn Val Thr Arg Thr Ala His Gly Leu Ala Arg Ile Asp 100 105 110Leu Leu Cys His Lys Lys Arg Pro Gln Leu Gly Pro Arg Ile Trp Lys 115 120 125Arg Ser Val Gln Lys Ile Lys Gln Lys Lys Asp Arg Pro Phe Ala Val 130 135 140Asn Thr Tyr Gly Asn Asn Lys Gly Leu Phe Thr Ile Thr Val Gly Val145 150 155 160Leu Leu Tyr Ala Ala Phe Gly Thr Cys Phe Leu Ile Ala Asn Leu Ala 165 170 175Tyr Leu Phe Gly Ile Tyr Ile Ile Tyr Asp Ala Ser Ile Ile Asp Glu 180 185 190Val Ser29263PRTOnchocerca volvulus 29Ile Gly Glu Asn Pro Met Asp Val Asn Ala Ile Ala Gly Ile Ile Gly1 5 10 15Gly Ile Ser Asn Met Met Gln Asn Asn Val Glu Thr Ile Asp Val Pro 20 25 30Ser Ser Gln Ile Met Gly Arg Trp Tyr Gln Val Tyr Lys Ala Ala Ile 35 40 45Ala Phe Asp Val Tyr Arg Thr Asp Ile Phe Cys Pro Val Ala Tyr Phe 50 55 60Lys Pro Asn Ser Val Met Gly Glu Asp Gly Phe Ser Ile Glu Glu Ala65 70 75 80Tyr Arg Val Ile Thr Lys Asn Gly Pro Val Glu Thr Tyr Lys Arg Asp 85 90 95Leu Asn Lys Val Gly Thr Gly Gln Tyr Trp Met Tyr Thr Glu Glu Tyr 100 105 110Phe Tyr Pro Arg Gln Phe Asn Ile Ile Ser Val Gly Pro Asn Tyr Thr 115 120 125Asn Thr Thr Asp Gly Ser Glu Glu Glu Lys Gln Tyr Gln Tyr Met Val 130 135 140Val Thr Asp Gly Asn Arg Leu Ser Leu Ser Val Tyr Ala Arg His Pro145 150 155 160Met Ile Phe Tyr Gln Lys Tyr Asn Glu Glu Val Val Lys Phe Leu Glu 165 170 175His Ala Gly Phe Gly Gly Lys Val Phe Trp Asn Ser Pro Lys Pro Ile 180 185 190Tyr Gln Gly Ala Asp Cys Glu Trp Pro Ser Glu Lys Glu Val Phe Ala 195 200 205Arg Arg Val Leu Lys Asn Gln Glu Leu Ala Lys Asn Gly Gly Leu Asp 210 215 220Thr Ala Thr Lys Ser Gly Ser Phe Gly Gly Ser Ser Gln Ala Thr Asp225 230 235 240Val Arg Ser Ser Ile Thr Glu Ile Leu Gln Asn Pro Gln Leu Ala Leu 245 250 255Gln Lys Leu Val Gln Gly His 26030242PRTOnchocerca volvulus 30Met Thr Ile Ile Lys Ser Met Leu Lys Ile Thr His Val Ile Phe Asp1 5 10 15Leu Asp Gly Leu Leu Ile Asp Thr Glu Val Val Phe Ser Lys Val Asn 20 25 30Gln Cys Leu Leu Ser Lys Tyr Asp Lys Lys Phe Thr Pro His Leu Arg 35 40 45Gly Leu Val Thr Gly Met Pro Lys Lys Ala Ala Val Thr Tyr Met Leu 50 55 60Glu His Glu Lys Leu Ser Gly Lys Val Asp Val Asp Glu Tyr Cys Lys65 70 75 80Lys Tyr Asp Glu Met Ala Glu Glu Met Leu Pro Lys Cys Ser Leu Met 85 90 95Pro Gly Val Met Lys Leu Val Arg His Leu Lys Thr His Arg Ile Pro 100 105 110Met Ala Ile Cys Thr Gly Ala Thr Lys Lys Glu Phe Glu Ile Lys Thr 115 120 125Arg His His Lys Glu Leu Leu Asp Leu Ile Ser Leu Trp Val Leu Ser 130 135 140Gly Asp Asp Pro Ala Ile Lys Arg Gly Lys Pro Ala Pro Asp Pro Phe145 150 155 160Leu Val Thr Met Asp Arg Phe Lys Gln Lys Pro Glu Lys Ala Glu Asn 165 170 175Val Leu Val Phe Glu Asp Ala Thr Asn Gly Val Cys Ala Ala Ile Ala 180 185 190Ala Gly Met Asn Val Val Met Val Pro Asp Leu Thr Tyr Met Lys Ile 195 200 205Pro Glu Gly Leu Glu Asn Lys Ile Asn Ser Val Leu Lys Ser Leu Glu 210 215 220Asp Phe Lys Pro Glu Ser Val Gly Leu Pro Ala Tyr Asp Ala Ser Ser225 230 235 240Asn Glu31148PRTOnchocerca volvulus 31Ile Pro Gln Arg Arg Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln1 5 10 15Arg Asp Glu Arg Glu Ile Pro Pro Phe Leu Glu Gly Ala Pro Pro Ser 20 25 30Val Ile Asp Glu Phe Tyr Asn Leu Leu Lys Thr Asp Glu Asn Lys Thr 35 40 45Asp Gln Gln Thr Glu Ala Asp Val Glu Ala Phe Ile Asn Arg Leu Gly 50 55 60Gly Ser Tyr Lys Val Arg Phe Thr Gln Phe Met Glu Glu Val Lys Lys65 70 75 80Ala Arg Ala Asp Tyr Glu Arg Ile His Gln Gln Ala Val Ala Arg Phe 85 90 95Ser Pro Ala Ala Lys Asp Ala Asp Ala Arg Met Ser Ala Ile Ala Asp 100 105 110Ser Pro His Leu Thr Thr Arg Gln Lys Ser Gln Gln Ile Gln Ala Ile 115 120 125Met Asp Ser Leu Ser Glu Ser Val Arg Arg Glu Ile Ile Asn Ala Leu 130 135 140Ser Pro Gln Glu14532141PRTOnchocerca volvulus 32Asp Leu Leu Ser Glu Ala Gly Asp Phe Phe Thr Lys His Phe Thr Asp1 5 10 15Ile Lys Ser Leu Phe Ala Lys Asp Glu Lys Gln Leu Gln Gln Ser Val 20 25 30Asp Arg Val Lys Asp Leu Leu Ala Thr Ile Gln Asp Lys Met Ser Met 35 40 45Leu Gln Pro Leu Ala Asn Asp Met Gln Lys Thr Thr Leu Gly Lys Ile 50 55 60Gly Asp Leu Ile Ser Gln Val Asn Ser Phe Arg Glu Thr Met Ser Asn65 70 75 80Pro Lys Met Asp Phe Thr Asn Lys Glu Asn Lys Trp Glu Glu Leu Leu 85 90 95Lys Lys Ile Phe Val Thr Glu Gly Leu Asn Lys Val Ile Pro Leu Leu 100 105 110Gln Lys Leu Lys Asn Ser Ala Pro Thr Thr Phe Ala Thr Tyr Leu Phe 115 120 125Thr Cys Ile Val Pro Val Leu Ile Asn Thr Leu Arg Glu 130 135 14033446PRTDirofilaria immitis 33Met Arg Thr Ala Ser Gln Leu Thr Phe Met Leu Phe Leu Val Leu Lys1 5 10 15Lys Lys Phe Lys Asn Ile Asp Lys Leu Phe Ser Gln Ile Ser Val Asn 20 25 30Met Ser Val Met Phe Ala Asn Ser Arg Ser Ser Gln Ala Asn Ser Gly 35 40 45Tyr Leu Val Glu Phe Lys Ala Gly Arg Ser Asn Leu Gln Ala Gly Ser 50 55 60Thr Val Asp Lys Arg Lys Val Val Ala Asp Lys Thr Lys Gly Leu Ile65 70 75 80Phe Ile Lys Gln Ser Ser Asp Gln Leu Met His Phe Cys Trp Lys Asn 85 90 95Arg Glu Thr Gly Thr Val Val Asp Asp Leu Ile Ile Phe Pro Gly Asp 100 105 110Thr Glu Phe Leu Arg Val Lys Glu Cys Thr Asp Gly Arg Val Tyr Met 115 120 125Leu Lys Phe Lys Ser Thr Asp Glu Lys Arg Leu Phe Trp Met Gln Asp 130 135 140Gly Lys Thr Asp Lys Asp Asp Glu Asn Cys Lys Lys Ile Asn Glu Thr145 150 155 160Leu Asn Asn Pro Pro Ala Pro Arg Ala Ala Ala Arg Gly Gly Ala Asp 165 170 175Arg Ala Gly Ala Ser Ser Phe Gly Thr Leu Ala Ala Leu Gly Ser Ala 180 185 190Gly Ala Asp Ser Glu Leu Gly Ala Leu Gly Asn Leu Asp Gln Asn Gln 195 200 205Leu Met Gln Leu Leu Ser Leu Met Asn His Thr Asn Ser Ala Ser Ala 210 215 220Ser Glu Ala Ala Asn Leu Leu Pro Gln Leu Pro Leu Val Ala Asp Thr225 230 235 240Pro Asn Pro Val Ala Ser Glu Glu Ser Gly Thr Thr Ser Thr Gln Gly 245 250 255Ala Thr Pro Ser Asn Thr Pro Ala Asn Gly Ile Ile Ala Gly Ser Ser 260 265 270Ser Asn Asn Ala Val Gln Leu Ser Gln Leu Lys Glu Ile Ile Ala Ser 275 280 285Ile Thr Pro Pro Asp Gly Ser Ile Arg Lys Pro Ser Val Asp Phe Thr 290 295 300Asp Val Leu Cys Cys Ala Asp Lys Ile Asn Asp Val Leu Gly Lys Tyr305 310 315 320Ala Glu Arg Leu Ile Pro His Leu Pro Asn Gln Glu Pro Ile Tyr Asn 325 330 335Asn Gln Glu Glu Leu Gln Gln Thr Leu Arg Thr Pro Gln Phe Arg Gln 340 345 350Ala Val Asp Ile Phe Gly His Ala Leu Gln Thr Gly Gln Leu Ala Pro 355 360 365Ile Leu Arg Gln Phe Gly Ile Asp Ser Asn Thr Ala Ile Ala Ala Gly 370 375 380Asn Gly Asp Leu Ile Ala Trp Ala Thr Gln Phe Thr Thr Ser Glu Asn385 390 395 400Glu Lys Glu Ile Ala Val Lys Thr Glu Thr Leu Pro Phe His Pro Gly 405 410 415Met Glu Ser Asp Val Glu Asp Glu Glu Thr Asn Glu Lys Ala Val Arg 420 425 430Glu Ser Asp Lys Asn Arg Thr Asp Asp His Met Asp Leu Asp 435 440 44534114PRTDirofilaria immitis 34Met Leu Pro Thr Leu Tyr Ile Asn Asn Ala Val Ile Arg Pro Val Leu1 5 10 15Ser Glu Thr Lys Lys Val Lys Val Gln Asn Ile Ser Ser Pro Phe Leu 20 25 30Ile Phe Leu Leu Leu Ser Ile Thr Lys Met Leu Ser Leu Ser Val Leu 35 40 45Leu Leu Phe Ile Ser Met Ala Thr Met Ala Arg Pro Pro Asn Pro Asp 50 55 60Glu Ile Lys Glu Leu His Glu Gln Gln Leu Asn Asp Ser Lys Asp Asp65 70 75 80Tyr Asp Met Leu Pro Asp Val Gly His Ile Pro Glu Ser Phe Lys Glu 85 90 95Ser Leu Lys Lys Gln Lys Met Leu Tyr Leu Asp Met Leu Arg Gln Gln 100 105 110Ser Leu35298PRTDirofilaria immitis 35Met Ile Ser Ser Arg Leu Arg Ile Thr Ile Pro Glu Ser Ile Val Ile1 5 10 15Phe Gly Ile Phe Cys Phe Phe Ile Phe Phe Cys Phe Leu Ser Phe Phe 20 25

30Phe Phe Phe Thr Leu Trp Ser His Arg Asp Thr Ile Asn Phe Gln Thr 35 40 45Asp Phe Met Thr Glu Thr Ile Lys Phe Ile Val Tyr Ala Val Val Ile 50 55 60Leu Arg Met Met Phe Phe Asp Ile Val Cys Phe Tyr Ser Phe Leu Met65 70 75 80Met Thr Ile Val Leu Ile Asn Thr Ser Asn Gly Leu Ser Val Pro Ala 85 90 95Gly Leu Arg Pro Ala Lys Lys Val Gly Asp Pro Arg Glu Gln Ile Val 100 105 110Pro Gly Lys Glu Gln Gln Gln Gln Arg Glu Gln Gln Gln Gln Gln Gln 115 120 125Gln Gln Leu Gln Glu Glu Glu Gln Gln Gln Gln Gln Gln His Asp Glu 130 135 140Val Ser Asn Leu Arg Pro Thr Pro Lys Val Pro Pro Asn Leu Ser Ile145 150 155 160Arg Ser Arg Met Met Ala Ala Leu Ser Ala Ser Pro Val Glu Pro Asn 165 170 175Lys Glu Lys Asn Ser Ser Lys Val Glu Thr Asp Ser Phe Ser Lys Pro 180 185 190Pro Ile Ile Phe Ser Lys Gly Asn Lys Lys Thr Val Pro Gly Lys Ile 195 200 205Ala Pro Ser Gly Ser Ser Lys Gly Asn Ala Arg Val Ile Val Ala Pro 210 215 220Pro Ala Asp Leu Gly Lys Asn Asn Tyr Gly Leu Asn Thr Val Leu Gln225 230 235 240Thr Asn Leu Val Asp Ser His Gly Arg Ile Met Lys Asn Val Asn Ser 245 250 255Val Pro Ile Lys Val Pro Ser Ser Ala Glu Met Lys Asn Ala Arg Thr 260 265 270Arg His Thr Ala Arg Gln Val Glu Ser Asp Ala Asp Lys Val Val Pro 275 280 285Ile Lys Phe Gly Ser Thr Ser Arg Arg Arg 290 2953699PRTDirofilaria immitis 36Met Met Arg Ile Lys Trp Ile Ile Leu Leu Leu Leu Leu Leu Leu Pro1 5 10 15Ile Ile Thr Ala Glu Phe Ser Ala Pro Val Gly Thr Asn Ser Ser Leu 20 25 30Thr Ile Phe Asp Lys Asp Lys Gln Val Leu Leu Arg Ser Asp Arg Leu 35 40 45Lys Arg Gln Cys Gly Pro Cys Gly Val Ala Pro Ser Pro Val Ile Val 50 55 60Cys Cys Gly Ala Ala Gly Leu Lys Glu Ile Phe Arg Ser Trp Trp Leu65 70 75 80His Ile Pro Leu Leu Leu Leu Pro Met Ser Thr Ser Trp Leu Lys Thr 85 90 95Met Val Cys37193PRTDirofilaria immitis 37Met Phe Arg Leu Leu Ile Ala Ile Gln Ile Leu Arg Phe Cys Gln Ala1 5 10 15Asn Tyr Ile Asn Asp Val Tyr Trp Lys Arg Ser Ile Ile Gly Tyr Gln 20 25 30His Ile Pro Ile Ile Leu Asn Ile Cys Tyr Leu Leu Gln Thr Glu Val 35 40 45Ser Asn Lys Gly Val Val Asp Ala Leu Phe Leu His Ser Pro Thr Tyr 50 55 60His Arg Val Glu Met Ser Glu Glu Thr Asp Asn Ile Glu Ser Ile Ala65 70 75 80Asp Lys Ser Asn Ile Thr Val Ala Asn Lys Pro Asn Leu Met Ile Tyr 85 90 95Pro Ala Asp Phe Gln Val Ser Ser Asn Glu Arg Ala Ser Ala Ser Ile 100 105 110Pro Ile Thr Ile Thr Ile Thr Ser Ser Gly Asp Thr Ile Ile Lys Ser 115 120 125Phe Lys His Lys His Gln Ser Asn Glu Ile Phe Lys Arg Arg Val Ala 130 135 140Lys Met Ala Ile Ala Pro Val Asn Ala Pro Glu Val Glu Asn Leu Ala145 150 155 160Pro Glu Val Glu Asn Pro Ser Pro Ser Thr Ala Gly Tyr Glu Ser Lys 165 170 175Thr Glu Glu Gln Ala Pro Ser Glu Ser Gly Gln Tyr Gly Lys Arg Arg 180 185 190Lys38362PRTDirofilaria immitis 38Met Tyr Asn Leu Ala Lys Leu Leu Glu Asn Glu His Gly Ser Ile Glu1 5 10 15Ala Ser Phe His Val Tyr Val Glu Asn Phe Phe Glu Gly Leu Asp Gly 20 25 30Glu Ser Trp Ser Ile Asp Gln Thr Asn Ala Gln Leu Tyr Pro Ile Ile 35 40 45Asp Glu Pro Phe Asn Asn Thr Val Arg Val Gly Arg Thr Ala Gln Phe 50 55 60Gln Cys Lys Val Lys Asn Gln Gln Gln Pro Leu Ile Lys Trp Leu Lys65 70 75 80Arg Ile Asp Asp Pro Asn Ala Ile Arg Gln Ala Asn Ala Asn Ala Thr 85 90 95Leu Ile His Ala Asn Asn Met His Leu Leu Leu Leu Glu Lys Pro Glu 100 105 110Thr Ser Ala Glu Leu Ser Asp Gly Ile Ser Leu Asn Arg Leu Ile Ile 115 120 125Pro Asn Val Arg Tyr Glu His Ser Gly Thr Tyr Leu Cys Val Val Thr 130 135 140Asn Ala His Gly Asp Ile Ala Tyr Arg Ser Ala Tyr Leu His Val Ile145 150 155 160Ala Arg Ser Asp His Gly Met Leu Ser Asn Ile Tyr Phe Tyr Gly Gly 165 170 175Ile Leu Val Leu Ile Val Val Phe Thr Leu Ile Thr Tyr Ala Val Tyr 180 185 190Phe Leu Arg Lys Asn Gln Ala Ala Lys Asn Ser Glu Ser Ala Gln Asp 195 200 205Ile Thr Asn Thr Arg Tyr Ser Phe Ser Leu Arg Pro Pro Pro Pro Asn 210 215 220Leu Pro Pro Pro Lys Ala Pro Ala Leu Pro Ser Glu Arg Gln Gln Leu225 230 235 240Met Ser Asp Asn Gln Pro Cys Asp Arg Tyr Ala Val Asn Ser Ala Ala 245 250 255Thr Thr Tyr Tyr Pro Gln Phe Ala Thr Pro Asp Lys Lys Leu Gln Lys 260 265 270Ile Ile Thr Glu Ser Gly Gly Thr Arg Pro Thr Pro Ile Arg Arg Thr 275 280 285Asn Gly Gly Asp Thr Lys Tyr Arg Leu Lys Asp Glu Tyr Ile Asn Ser 290 295 300Pro Lys Trp Val His Thr Lys Gly Asp Asn Ile Glu Val Glu Met Asp305 310 315 320Gln Asn Leu Leu Lys Asn Arg Ser Ser His Cys Tyr Asn Pro Ile Ser 325 330 335Gly Ala Tyr Gly Arg Thr Asp Asn Ile Asp Arg Gln Gln Gln Lys Ser 340 345 350Phe Leu Thr Ile Gly Asn Leu Gln Lys Arg 355 36039215PRTDirofilaria immitis 39Met Leu Lys Leu Ala Asn Thr Glu Ile Phe Phe Ile Ala Phe Leu Val1 5 10 15Tyr Ser Lys Glu Ile Ile Leu Asn Cys Tyr Glu Asp Tyr Lys Glu Cys 20 25 30Val Ala Thr Ser Asn Lys Thr Asn His Val Asn Met Asp Asn Val Asn 35 40 45Pro Gln Asn Leu Ile Glu Phe Cys Phe Asp His Thr Gln Asn Ile Leu 50 55 60Pro Cys Leu Val Thr Lys Leu Gly Leu Thr Lys Gly Ile Ser Val Ser65 70 75 80Ile Phe Ser Leu Phe Leu Ser Thr Cys Glu Leu Glu Ala Gln Asn Asn 85 90 95Lys Ser Ser Ser Thr Thr Glu Met Gln Gln Ile Leu Arg His Leu Leu 100 105 110Arg Leu Tyr Ala Tyr Phe Cys Ala Tyr Ser Asn Ile Ile Asp Leu His 115 120 125Arg Asn Arg Glu Cys Phe Arg Tyr Leu Met Lys Arg Cys Val Leu Asn 130 135 140Lys Pro Asp Glu Ser Cys Met Phe Tyr His Cys Gly Lys Ile His Phe145 150 155 160Asn Leu Ser Lys Ser Ser Arg Lys Ile Leu Phe Thr Arg Gln His Asp 165 170 175Thr Thr Lys Ile Val Asn Leu Gly Asn Lys Met Asn Gln Leu Ala Thr 180 185 190Phe Asn Asn His Gln Val Arg Ser Ala Val Val Val Thr Leu Ile Ile 195 200 205Thr Phe Ile Asp Met Ile Gln 210 21540395PRTDirofilaria immitis 40Met Tyr Ser Gln Glu Asn Gln Asp Asp Lys Arg Arg Asn Asp Glu Arg1 5 10 15Ile Ala Leu Ser Val Pro Tyr Asn Asn Thr Asn Ile Met Asp Arg Ser 20 25 30Tyr Phe Lys Pro Ile Lys Leu Ser Tyr Pro Tyr Ser Asp Glu Cys Leu 35 40 45Glu Val Asn Lys Lys Ser Ser Asp Asp Ser Asp Gln Arg Leu Ser Gln 50 55 60Asn Ser Ser Thr Pro Gln Ser Asn Tyr Asp Gln Ser Ser Glu Arg Leu65 70 75 80Ser Ile Asp Asp Ser Asp Leu Ile Asp Asp Ser Thr Ser Ala Ala Gln 85 90 95Leu Ser Thr Ser Ser Pro Ile Ser Val Thr Ser Ala Ser Thr Ser Ser 100 105 110Phe Tyr Pro Thr Leu Asn Ile Gly Asn Gly Met Glu Met Asn Ala Lys 115 120 125Tyr Ala Lys Ile Glu Gln Ser Glu Gly Ile Arg Ser Asp Gln Ser Ser 130 135 140Thr Leu Arg Ile Ser Asp Lys Tyr Lys Lys Tyr Thr Ala Ile Lys Arg145 150 155 160Arg Leu Ser Glu Leu Cys Gly Ile Ile Glu Glu Lys Asp Lys Gln Leu 165 170 175Arg Val Val Arg Asn Gly Leu Asn Glu Lys Asp Leu Glu Ile Gly Lys 180 185 190Leu Cys Asp Lys Ile Arg Ala Leu Glu Tyr Asn Cys Gly Arg Leu Gln 195 200 205Ala Val Ile Glu Ser Val Gly Asp Glu Ser Asp Gln Asn Gln Ile Lys 210 215 220Leu His Glu Ile Ile Asn Glu Arg Asp Gly Leu Leu Val Arg Asn Ala225 230 235 240Ser Leu Ser Arg Gln Ile Glu Phe Glu Lys Arg Glu Trp Ser Ile Glu 245 250 255Arg Glu Arg Leu Ser Met Asp Leu Asp Asp Val Thr Arg Glu Leu Glu 260 265 270Leu Gln Lys Met Ile Leu Asn Gly Glu Asn Ile Ser Glu Ile Val Gln 275 280 285Arg Trp Gln Thr Lys Val Phe Glu Leu Glu Gly Met Ile Ala Asp Arg 290 295 300Asp Arg Ala Ile Arg Ala Gln Gln Val Arg Ile Ser Lys Leu Lys Gln305 310 315 320Ser Leu Ala Glu Ala Asp Arg Ile Ser Cys Asp Asp Ser Ser Glu Ser 325 330 335Gln Thr Lys Leu Asp Ser Pro Ser Phe Thr Cys Ile Lys Arg Leu Leu 340 345 350Leu Gln Tyr Leu Thr Ser Ser Asp Glu Glu Arg Ile Gln Leu Leu Arg 355 360 365Asn Val Ser Thr Met Leu His Leu Ser Asp Asp Glu Gln His Gln Val 370 375 380Leu Thr Asn Leu Lys Ser Arg Ile Gln Ile Ser385 390 3954199PRTDirofilaria immitismisc_feature(1)..(1)Xaa can be any naturally occurring amino acid 41Xaa Lys Cys Arg Asp Gln Arg Ala Glu Val Glu Tyr Val Asn Gly Ser1 5 10 15Ala Lys Asn Cys Leu Tyr Trp Pro Cys Thr Val Gly Tyr Phe Cys Glu 20 25 30Tyr Thr Glu Ser Arg Asn Gly Gly His Tyr Ile Cys Cys Gly Thr Asn 35 40 45Ala Asn Asn Ile Tyr Gly Lys Val Lys Val Tyr Pro Gly Thr Asn Lys 50 55 60Pro Leu His Cys Ser Ile Met Asn Thr Cys Pro Phe Leu Asp Thr Pro65 70 75 80Asn Cys Val Met Ser His Arg Tyr Gly Tyr Lys Val Cys Cys Ser Thr 85 90 95Met Asn Cys42589PRTDirofilaria immitis 42Met Leu Met Lys Gln Ser Asp Ser Cys Val Asp Tyr Phe Tyr Asp Gln1 5 10 15Tyr Lys Gly Gln Glu Tyr Val Lys Asp Asp Ala Phe Asn Thr Gln Asn 20 25 30Ile Thr Asp Asn Phe Arg Lys Ser Ser Ser Asp Ile Ala Gln Leu Met 35 40 45Asn Ser Gln Ile Glu Leu Ile Ser Gln Pro Glu Lys Val Asn Glu Asp 50 55 60Ser Ala Lys Ser Ser His Tyr Asn Asp Asp Leu Gln Lys Ser Ile Glu65 70 75 80Asp Asp Thr Val Glu Ala Thr Gln Arg Lys Lys Asp Glu Lys Leu Leu 85 90 95Glu Phe Leu His Ser Leu Ile Val Ser Thr Ile Pro Lys Thr Ile His 100 105 110Leu Glu Gly Asn Ser Val Asn Leu Leu Thr Leu Thr Thr Thr Ile Thr 115 120 125Pro Ile Ala Ile Ile Thr Thr Lys Asn Thr Ser Gly Thr Ala Asn Ala 130 135 140Ile Thr Thr Arg Lys Tyr Lys Lys Tyr Lys Leu Asn Ala Phe Val Asn145 150 155 160Ile Ser Ser Asp Thr Leu Thr Glu Leu Pro Lys Phe Leu Pro Glu Asn 165 170 175Phe Asn Ser Thr Asn Phe Ala Asn Val Glu Lys Thr Glu Lys Phe Ser 180 185 190Asn Ser Lys Gln Val Ala Thr Asp Ser Ile Phe Ser Leu Lys Glu Ser 195 200 205Ala Tyr Leu Glu Thr Pro Val Ile Arg Asp Phe Ser Ser Ala Asn Asp 210 215 220Ser Ala Lys Thr Asp Pro Leu Phe Thr Arg Asn Tyr Val Asp Lys Gln225 230 235 240Ile Asp Met Asn Thr Thr Lys Phe Asn Lys Asn Leu Lys Lys Ser Arg 245 250 255Leu Thr Thr Ile Ser Thr Ser Asn Leu Thr Thr Val Leu Ser Gln Leu 260 265 270Gln Thr Thr Thr Ser Ile Ser Thr Thr Thr Ser Val Thr Thr Thr Ile 275 280 285Ser Thr Ser Ile Thr Ile Pro Glu Leu Thr Leu Val Ser Gln Ser His 290 295 300Arg His Leu His His Tyr His His His His His His Gln Tyr Glu Asn305 310 315 320Tyr Asp His Glu Ser Pro Ile Ile Val Thr Ala Leu Phe Asp Ile Gly 325 330 335Arg Gly Lys Trp Pro Arg Tyr Thr Arg Thr Tyr Glu Gln Tyr Met Asn 340 345 350Tyr Leu Lys His Leu Leu Lys Leu Glu Asn Cys Leu Val Ile Tyr Thr 355 360 365Asp Ser Arg Gly Ala Glu Phe Val Arg Gln Thr Arg Asn Val His Asn 370 375 380Thr Gln Ile Phe Glu Ile Ser Met His Asp Leu Pro Leu Tyr Arg Tyr385 390 395 400Arg Glu Glu Met Lys Gly Ile Ile Gln Arg Glu Gln Lys Asp Trp Gln 405 410 415Phe Ser Pro Lys Thr Arg Tyr His Pro Glu Ala Asn Ser Ala Asp Tyr 420 425 430Asn Ile Ile Val Asn Ser Lys Pro Tyr Phe Leu Tyr Asn Ala Thr Gln 435 440 445Asn Val Arg Phe Arg Thr Ser Asp Arg Met Phe Val Trp Ile Asp Ala 450 455 460Gly Tyr Gly His Gly Arg Lys Gly Ile Ile Pro Asp His Cys His Trp465 470 475 480Arg Pro Arg Leu Gln Arg Asp Arg Met Thr Ile Ile Gln Leu Thr Pro 485 490 495Lys His Asp Lys Val Ser Arg Tyr Ser Ile Thr Asp Leu Tyr Arg Val 500 505 510Asp Trp Val Val Leu Ser Gly Gly Phe Ile Ala Gly Asp Ser His Thr 515 520 525Ile Asn Arg Phe Tyr Arg Phe Tyr Gln Lys Leu Phe Met Glu Leu Leu 530 535 540Asp Ser Gly Arg Ile Asp Asp Asp Gln Thr Ile Leu Thr Leu Met Leu545 550 555 560Lys His Tyr Thr Thr Leu Phe Asn Pro Ile Ser Ser Asn Gly Asp Trp 565 570 575Tyr Ala Leu Phe Arg Leu Phe Pro Cys His Asp Arg Gln 580 58543875PRTDirofilaria immitis 43Met Lys Gln Ala Thr Thr Trp Gly Ser Ile Cys Glu Met Cys Pro Cys1 5 10 15Ala Ala Lys Pro Ile Cys Pro Pro Pro Val Ile Cys Pro Pro Arg Ile 20 25 30Cys Pro Pro Pro Val Ile Cys Pro Pro Gln Ile Cys Pro Pro Cys Pro 35 40 45Pro Arg Ile Cys Pro Pro Pro Val Ile Cys Pro Pro Gln Ile Cys Pro 50 55 60Pro Cys Pro Pro Gln Ile Cys Pro Pro Cys Pro Lys Pro Gln Pro Pro65 70 75 80Pro Pro Pro Pro Pro Pro Pro Val Leu Pro Ser Leu Pro Pro Thr Ser 85 90 95Phe Lys Pro Met Ile Thr Cys Cys Arg Thr Cys Ile Cys Tyr Ile Arg 100 105 110Arg Lys Arg Asp Ser Leu Asn Asp Tyr Asp Arg Ile His Asp Ile Asn 115 120 125Pro Val Cys Asn Asn Asp Gln Leu Met Met Ile Met Lys Lys Lys Ile 130 135 140Arg Thr Asn Val Thr Glu Ser Thr Ile Ala Ile Lys Lys Ala Ala Asp145 150 155 160Ser Met Leu Gln Ala Glu Phe Asn Val Phe Cys Ala Ile Asn Asp Leu 165 170 175Thr His Val Ala His Ala Glu His Phe Cys Gln Tyr Lys Lys Asp Asn 180 185 190Ser Val Phe Asp Ser Phe Leu Phe Arg Ser Thr Leu Lys Gly Leu Ile 195 200 205Glu Glu Cys Arg Glu Gly Val Arg Trp Trp Pro Gly Ser Leu Gly Asp 210 215

220Leu Asp Phe Ser His Ile Ser Leu Tyr Arg Ala His Lys Tyr Ile Gly225 230 235 240Asn Glu Glu Met Asn Arg Ser Thr Lys Thr Lys Ile Ser Phe Thr Arg 245 250 255Ile Asn Lys Lys Trp Arg Leu Gly His Thr Gly Lys Lys Tyr Asn Lys 260 265 270Val Arg Phe Ser Arg Asn Ile Ala Lys Lys Phe Ile Gly Val Cys Asn 275 280 285Ile Ile Arg Leu Lys Lys Ser Val Ser Arg Ser Val Arg Pro Phe Glu 290 295 300Asn Gln Lys Ser Thr Ser Phe Asn Val Phe Gln Leu Leu Val Pro Lys305 310 315 320Glu Lys Val Glu Ile Val Val Asp Asp Thr Gln Ala Glu Glu Met Asn 325 330 335Ser Glu Thr Ala Gln Glu Val Gln Leu Phe Asn Val Arg Lys Ser Asn 340 345 350Ala Asp Ser Lys Thr Asp Gly Glu Lys Asp Thr Ala Asp Leu Asp Val 355 360 365Ile Leu Leu Thr Asn Glu Glu Cys Ser Ser Ser Arg Gln Glu Asn Leu 370 375 380Asn Lys Asp Glu Pro Glu Ile Val Ile Leu Asp Asp Ser Ala Pro Ser385 390 395 400Lys Ser Asp Leu Asn Thr Ser Asp Glu Ile Ile Cys Leu Gln Asp Leu 405 410 415Lys Met Val Asn Glu Val Pro Thr Phe Ser Val Thr Pro Lys Gln Lys 420 425 430Thr Val Lys Glu Leu Pro Arg Glu Thr Arg Thr Tyr Gly Thr Arg Arg 435 440 445Gly Arg Gln Ser Arg Ala Tyr Cys Glu Asp Leu Arg Lys Phe Pro Ser 450 455 460Ile Arg Asn Pro Val Ser Ser Ser Ser Ser Ser Ile His Ala Lys Asn465 470 475 480Met Pro Glu Phe Val Asp Leu Leu Thr Gln Gly Thr Leu Leu Ile Cys 485 490 495Lys Lys Trp Leu Arg Arg Trp Asp Ile Val Gln Ser Gly Val Ile Gly 500 505 510Gly Asn Pro Leu Arg Ile Cys Ser Tyr Asn Val Leu Cys Gln Gln Thr 515 520 525Ala Tyr Lys Thr Pro Glu Leu Tyr Ile His Leu Thr Lys Pro Gly Arg 530 535 540Ala Tyr Glu Leu Thr Trp Glu Asn Arg Trp Arg Leu Leu Thr Arg Glu545 550 555 560Phe Ser Met Ile Gly Ala Asp Ile Phe Cys Leu Gln Glu Val Gln Tyr 565 570 575Asp His Tyr Asp Gln Phe Phe Arg Pro Tyr Phe Glu Ala Ala Gly Phe 580 585 590Phe Gly Lys Tyr Lys Lys Arg Thr Asn Asn Leu Leu Asp Gly Cys Ala 595 600 605Ile Phe Tyr Lys Ser His Leu Gln Leu Leu His Tyr Arg Tyr Ile Glu 610 615 620Tyr Phe Leu Asn Ile Asp Ser Val Leu Asn Arg Asp Asn Val Gly Gln625 630 635 640Leu Ile Arg Leu Lys Asp Met Arg Ser Gly Arg Glu Phe Cys Val Val 645 650 655Asn Thr His Leu Leu Phe Asn Lys Arg Arg Gly Asp Val Lys Leu Ala 660 665 670Gln Leu Ala Ile Leu Leu Ala Asn Ile Asp Gln Glu Cys Gly Pro Glu 675 680 685Ser Gly Gln Glu Cys Pro Tyr Ile Leu Cys Gly Asp Phe Asn Phe His 690 695 700Pro Tyr Ser Pro Ile Tyr Asn Phe Ile Met Asn Gly Glu Ile Cys Phe705 710 715 720Thr Asn Leu Arg Arg Gly Asp Ile Ser Gly Gln Gly Asn Ala Gly Gly 725 730 735Pro Phe Val Ser Val Asn Leu Leu Pro Glu Asp Val Lys Ile Ala Arg 740 745 750Asn Cys Arg Phe Asn Tyr Leu Lys Asn Arg Thr Met Leu Leu Pro Ser 755 760 765Leu Asn Cys Trp Ser His Pro Leu Cys Phe Asn Ser Val Tyr Gln Asn 770 775 780Met Asn Gly Glu Thr Arg Pro Met Ile Ser Thr Tyr His Ser Ile Glu785 790 795 800Ala Val Asn Pro Asp Phe Ile Phe Tyr Ser Val Lys Ser Lys Arg Val 805 810 815Gln Gln Ser Met Leu Pro His Ser Val Pro Ala Met Asn Val Ser Glu 820 825 830Arg Glu Ile Arg Leu Ile Arg Arg Leu Ser Leu Pro Asp Met Asn Glu 835 840 845Leu Ala Gly Thr Leu Gly Pro Trp Pro Asn Ser Thr Thr Pro Ser Asp 850 855 860His Ile Pro Leu Ile Ala Asp Phe Val Leu Gln865 870 87544158PRTDirofilaria immitis 44Met Tyr Cys Lys Leu Ile Ile Ser Phe Tyr Met Leu Leu Ser Ile Ala1 5 10 15Asn Met Thr His Leu Val Gly Tyr Arg Pro Gln Ile Tyr Leu Gln Gly 20 25 30Ile Pro Gln Asn Ile Gln Ser His Asp Ile Gln Arg Leu Asp Met Gln 35 40 45Gln Gln Ser Leu Lys Leu Pro Asp Thr Glu Leu Tyr Ser Ile Pro Ser 50 55 60His Asp Asn Gln Leu Gln Gly Leu Gln Leu Tyr Asp Met Gln Phe Gln65 70 75 80Gly Lys Gln Ser Lys Gly Ser Glu Lys Leu Cys Ser Gly Cys Lys Ile 85 90 95Ser Ile Asn Cys Ser Gly Lys Lys Cys Val Pro Met Arg Thr Arg Lys 100 105 110Pro Ile Val Thr Thr Pro Ser Pro Leu Ser Thr Gln Arg Pro Val Leu 115 120 125Thr Arg Pro Arg Leu Leu Ala Asp Cys Pro Cys Pro Cys His Val Ser 130 135 140Arg Gln Cys Arg Ile Cys Gln Pro Cys Gln Glu Ser Phe Ile145 150 15545230PRTDirofilaria immitis 45Met Phe Val Gly Met Arg Leu Tyr Leu Ala Ile Asp Val Leu Leu Leu1 5 10 15Leu Val Leu Arg Ile Lys Ser Asn Arg Ile Ile Leu His Arg Phe Ser 20 25 30Leu Phe Ile Gln Gln His Cys Cys Asn Asn Ile Ser Gln Ile His Arg 35 40 45Leu Asn Asp Cys Lys Tyr Ser Lys Val Arg Met Lys Ile Asp Lys Lys 50 55 60Ile Leu Ile Ile Val Ser Lys Thr Glu Trp Cys Asn Glu Ala Ile Lys65 70 75 80Val Val Phe Gly Lys Ser Ala Glu Ala Arg Arg Asn Arg Ser Asp Ala 85 90 95Ile Ser Trp Val Thr Pro Tyr Asn Phe Thr Gly Leu Met Asn Leu His 100 105 110Ser Lys Trp Arg Tyr Asp Ala Tyr Phe Asp Asn Val Thr Arg Thr Ala 115 120 125His Gly Leu Ala Arg Ile Asp Leu Leu Cys Pro Lys Arg Arg Ser His 130 135 140Ser Gly Arg Arg Ile Leu Lys Arg Ser Ile Gln Glu Asn Lys Gln Glu145 150 155 160Lys Ser Arg Arg Ser Phe Thr Val Asn Ile Tyr Gly Ser Ser Lys Gly 165 170 175Ile Phe Thr Ile Thr Val Gly Val Val Ile Tyr Ala Ile Phe Gly Val 180 185 190Cys Phe Leu Ile Thr Asn Met Ala Tyr Leu Ser Gly Ile Tyr Thr Val 195 200 205His Asn Thr Ser Val Ile Pro Glu Asp Lys Lys Arg Lys Glu Thr Ser 210 215 220Lys Arg Lys Glu Ile Leu225 23046279PRTDirofilaria immitis 46Met Ile Ser Val Phe Leu Leu Leu Thr Val Ile Val Ser Tyr Val Glu1 5 10 15Thr Ile Gly Glu Asn Pro Met Asp Ile Asn Ala Leu Ala Gly Ile Ile 20 25 30Gly Gly Ile Ser Asn Met Met Gln Asn Asn Val Glu Thr Ile Asp Val 35 40 45Pro Ser Ser Gln Ile Met Gly Gln Trp Tyr Gln Val Tyr Lys Ala Ala 50 55 60Ile Ser Phe Asp Ala Tyr Lys Thr Asp Met Phe Cys Pro Val Ala Tyr65 70 75 80Phe Lys Pro Asn Ser Val Met Gly Glu Asp Gly Phe Ser Ile Glu Glu 85 90 95Ala Tyr Arg Val Ile Thr Lys Asn Gly Pro Val Glu Thr Phe Lys Arg 100 105 110Asp Leu Asn Lys Val Gly Thr Gly Gln Tyr Trp Met Tyr Thr Glu Glu 115 120 125Tyr Phe Tyr Pro Arg Gln Phe Asn Ile Ile Gly Val Gly Pro Asn Tyr 130 135 140Thr Asn Ala Thr Asp Gly Arg Glu Lys Glu Asn Leu Tyr Glu Tyr Met145 150 155 160Ile Val Thr Asp Ala Asn Arg Leu Ser Leu Ser Val Tyr Ala Arg His 165 170 175Pro Met Ile Phe Tyr Gln Lys Tyr Asn Glu Glu Val Val Lys Phe Leu 180 185 190Glu His Ala Gly Phe Gly Gly Arg Val Phe Trp Asn Ser Pro Arg Pro 195 200 205Ile Tyr Gln Gly Thr Asp Cys Glu Trp Pro Ser Glu Lys Glu Val Phe 210 215 220Ala Arg Arg Val Leu Lys Asn Gln Glu Ala Ala Arg Asn Thr Gly Leu225 230 235 240Glu Thr Ala Thr Lys Ser Gly Leu Phe Gly Ser Ser Leu Thr Thr Asp 245 250 255Ala Tyr Asn Pro Ile Lys Glu Met Leu Gln Asn Pro Gln Leu Ala Leu 260 265 270Gln Lys Leu Val Gln Gly His 27547200PRTDirofilaria immitis 47Met Thr Val Ile Lys Ser Met Leu Asn Ile Thr His Val Ile Phe Asp1 5 10 15Leu Asp Gly Leu Leu Ile Asn Thr Glu Ile Val Phe Ser Gln Val Asn 20 25 30Gln Cys Leu Leu Ser Lys Tyr Gly Lys Lys Phe Thr Ser His Leu Arg 35 40 45Gly Leu Val Thr Gly Met Pro Lys Lys Ala Ala Val Ala His Ile Leu 50 55 60Glu His Glu Arg Leu Ser Glu Lys Ile Asp Val Asp Glu Tyr Cys Lys65 70 75 80Lys Tyr Asp Glu Met Ala Glu Glu Met Leu Pro Lys Cys Ser Leu Met 85 90 95Pro Gly Val Met Lys Leu Val Arg His Leu Lys Ala His Ser Ile Pro 100 105 110Met Ala Ile Cys Thr Gly Ala Thr Lys Lys Glu Phe Glu Leu Lys Thr 115 120 125Arg Cys His Lys Glu Leu Leu Asp Leu Ile Ser Leu Arg Val Leu Ser 130 135 140Gly Asp Asp Pro Ala Val Lys Arg Gly Lys Pro Ala Pro Asp Pro Phe145 150 155 160Leu Val Thr Met Glu Arg Phe Lys Gln Lys Pro Glu Lys Ala Glu Asn 165 170 175Val Leu Val Phe Glu Asp Ala Thr Asn Gly Val Tyr Ala Ala Ile Ala 180 185 190Ala Glu Glu Ser Lys Ile Val Lys 195 20048116PRTDirofilaria immitis 48Met Ile Leu Glu Gln Leu Glu Val Pro Pro Phe Leu Val Gly Ala Pro1 5 10 15Gln Ser Val Ile Lys Gln Phe Tyr Asp Leu Leu Lys Ala Asp Glu Thr 20 25 30Lys Thr Asp Ala Gln Thr Glu Ala Asp Val Glu Ala Phe Ile Asn Arg 35 40 45Leu Gly Gly Thr Tyr Lys Thr Arg Phe Asp Gln Phe Lys Gln Glu Ile 50 55 60Lys Gln Gly Lys Ala Ala Tyr Glu Arg Leu His Gln Gln Ala Val Ala65 70 75 80Lys Phe Ser Lys Glu Ala Arg Glu Ala Asp Ala Lys Met Ser Ala Ile 85 90 95Ala Asp Ser Pro Ser Leu Thr Thr Gln Gln Lys Thr Gln Gln Ile Gln 100 105 110Ala Ile Met Asp 11549158PRTDirofilaria immitis 49Met Leu Lys Tyr Gly Ile Leu Leu Ile Leu Ile Thr Val Gly Ala Tyr1 5 10 15Cys Asp Leu Leu Ser Glu Ala Gly Asp Phe Phe Ser Lys His Phe Thr 20 25 30Asp Phe Lys Ser Leu Phe Ala Ser Asp Glu Lys Gln Leu Gln Gln Asn 35 40 45Met Asp Arg Val Lys Asp Leu Leu Ala Thr Ile Gln Asp Lys Met Thr 50 55 60Ile Leu Lys Gln Leu Ala Asp Asn Ser Gln Lys Ser Thr Leu Glu Lys65 70 75 80Ile Thr Asp Ile Ile Ser Gln Val Asn Asp Phe Arg Glu Asn Val Phe 85 90 95Asn Ser Asn Val Asp Phe Asn Gln Lys Lys Thr Lys Trp Glu Glu Val 100 105 110Val Thr Lys Ile Phe Val Thr Asp Gly Leu Asn Lys Val Ile Pro Leu 115 120 125Leu Gln Lys Ala Lys Asn Ser Ala Pro Ala Thr Phe Ile Thr Tyr Leu 130 135 140Leu Thr Cys Ile Val Pro Leu Leu Ile Asn Ala Leu Arg Glu145 150 1555020PRTArtificial SequenceFlexible linker 50Lys Gly Pro Asp Val Pro Glu Thr Asn Gln Gln Cys Pro Ser Asn Thr1 5 10 15Gly Met Thr Asp 2051114PRTOnchocerca volvulus 51Met Ala Arg Ile Asn Arg Leu Asn Phe Leu Leu Cys Ile Val His Ala1 5 10 15Asn Ile Thr Ser Ala Pro Asn Pro Lys Asp Ser Asn Asp Glu Leu Phe 20 25 30Ala Asp Ala Glu Ser Ala Leu Gly Ser Glu Tyr Ala Gln Phe Val Glu 35 40 45Gln Ser Lys Gln His Lys Pro Val Tyr Phe Ser Asp Asn Gln Asn Thr 50 55 60Leu Glu Thr Ile Lys Leu Glu Ser Ile Pro Asn Pro Glu Thr Glu Thr65 70 75 80Ala Tyr Pro Met Phe Ile Cys Gly Phe Leu Gly Cys Met Lys Lys Met 85 90 95Asn Ser Val Glu Glu Tyr Leu Glu His Phe Lys Met His Glu Lys Gln 100 105 110Gly Tyr5220PRTOnchocerca volvulus 52Cys Pro Ser Leu Ser Ser Tyr Cys Glu Asp Trp Asp Pro Glu Asp Phe1 5 10 15Pro Ser Phe Val 205329PRTOnchocerca volvulus 53Leu Pro Ile Asp Lys Gln Asn Glu Ala Thr Leu Leu Trp Asn Ser Leu1 5 10 15Tyr Pro Asp Asp Ile Tyr Asn Glu Cys Gly Pro Arg Phe 20 255425PRTOnchocerca volvulus 54Ala Phe Ala Pro Asn Pro Lys Asp Ser Asn Asn Glu Leu Phe Ala Asp1 5 10 15Ala Glu Ser Ala Leu Gly Ser Glu Tyr 20 255524PRTOnchocerca volvulus 55Gly Cys Met Lys Lys Met Asn Ser Val Glu Glu Tyr Leu Glu His Phe1 5 10 15Lys Met His Glu Lys Gln Gly Tyr 205636PRTOnchocerca volvulus 56Ile Asn Lys Phe Tyr Asp Met Leu Arg Lys Trp Ala Glu Lys Tyr Ser1 5 10 15Val Gln Ala Glu Thr Asn Arg Phe Ile Ala Glu Glu Met Asn Tyr Asp 20 25 30Lys Met Gln Ser 355735PRTOnchocerca volvulus 57Ala Pro Pro Asn Arg Asp Thr Ala Asp Asp Leu Gln Asn Ala Asp Met1 5 10 15Gln Arg Gln Trp Glu Gln Glu Gln Arg Gln Arg Glu Glu Val Gln Lys 20 25 30Glu Glu Ile 355866PRTOnchocerca volvulus 58Thr Asp Met Asp Glu Leu Glu Glu Gln Arg Lys Gln Asp Phe Lys Gln1 5 10 15Tyr Glu Met Lys Lys Lys Ala Glu Glu Asp His Lys Met Gln Ala Ile 20 25 30Gln Thr Glu Arg Glu Glu Tyr Ile Arg Gln Met Glu Glu Gln Arg Arg 35 40 45Arg His Asn Lys His Glu Pro Leu Lys His Pro Gly Ser Arg Asn Gln 50 55 60Leu Arg655928PRTOnchocerca volvulus 59Asp Arg Glu Ala Gln Glu Glu Lys Pro Asp Gln Gly Trp Glu Asp Ile1 5 10 15Gly Asp Lys Asp Gln Tyr Thr Lys Glu Glu Leu Glu 20 256030PRTOnchocerca volvulus 60Thr Pro Ala Pro Thr Pro Asp Pro Ser Arg Met Ile Gln Pro Asp Gln1 5 10 15Ala Pro Met Gln Arg Leu Asp Ala Pro Ser Asp Gln Val Gly 20 25 306121PRTOnchocerca volvulus 61Val Ser Glu Ile Leu Thr Val Cys Leu Lys Glu Gly Phe Lys Lys Pro1 5 10 15Ser Asn Leu Gly Ile 2062130PRTOnchocerca volvulus 62Lys Asn Pro Ser Lys Met Glu Ser Lys Thr Gly Glu Asn Gln Asp Arg1 5 10 15Pro Val Leu Leu Gly Gly Trp Glu Asp Arg Asp Pro Lys Asp Glu Glu 20 25 30Ile Leu Glu Leu Leu Pro Ser Ile Leu Met Lys Val Asn Glu Gln Ser 35 40 45Lys Asp Glu Tyr His Leu Met Pro Ile Lys Leu Leu Lys Val Ser Ser 50 55 60Gln Val Val Ala Gly Val Lys Tyr Lys Met Asp Val Gln Val Ala Arg65 70 75 80Ser Gln Cys Lys Lys Ser Ser Asn Glu Lys Val Asp Leu Thr Lys Cys 85 90 95Lys Lys Leu Glu Gly His Pro Glu Lys Val Met Thr Leu Glu Val Trp 100 105 110Glu Lys Pro Trp Glu Asn Phe Met Arg Val Glu Ile Leu Gly Thr Lys 115 120 125Glu Val 13063181PRTOnchocerca volvulus 63Lys Ile Ser Ala Glu Asn Ala Asn Cys Lys Lys Cys Thr Pro Met Leu1 5 10 15Val Asp Ser Ala Phe Lys Glu His Gly Ile Val Pro Asp Val Val Ser 20 25

30Thr Ala Pro Thr Lys Leu Val Asn Val Ser Tyr Asn Asn Leu Thr Val 35 40 45Asn Leu Gly Asn Glu Leu Thr Pro Thr Gln Val Lys Asn Gln Pro Thr 50 55 60Lys Val Ser Trp Asp Ala Glu Pro Gly Ala Leu Tyr Thr Leu Val Met65 70 75 80Thr Asp Pro Asp Ala Pro Ser Arg Lys Asn Pro Val Phe Arg Glu Trp 85 90 95His His Trp Leu Ile Ile Asn Ile Ser Gly Gln Asn Val Ser Ser Gly 100 105 110Thr Val Leu Ser Asp Tyr Ile Gly Ser Gly Pro Arg Lys Gly Thr Gly 115 120 125Leu His Arg Tyr Val Phe Leu Val Tyr Lys Gln Pro Gly Ser Ile Thr 130 135 140Asp Thr Gln His Gly Gly Asn Arg Arg Asn Phe Lys Val Met Asp Phe145 150 155 160Ala Asn Lys His His Leu Gly Asn Pro Val Ala Gly Asn Phe Phe Gln 165 170 175Ala Lys His Glu Asp 18064436PRTArtificial SequenceOv-103-RAL2-CPI2M fusion protein 64Asp Leu Leu Ser Glu Ala Gly Asp Phe Phe Thr Lys His Phe Thr Asp1 5 10 15Ile Lys Ser Leu Phe Ala Lys Asp Glu Lys Gln Leu Gln Gln Ser Val 20 25 30Asp Arg Val Lys Asp Leu Leu Ala Thr Ile Gln Asp Lys Met Ser Met 35 40 45Leu Gln Pro Leu Ala Asn Asp Met Gln Lys Thr Thr Leu Gly Lys Ile 50 55 60Gly Asp Leu Ile Ser Gln Val Asn Ser Phe Arg Glu Thr Met Ser Asn65 70 75 80Pro Lys Met Asp Phe Thr Asn Lys Glu Asn Lys Trp Glu Glu Leu Leu 85 90 95Lys Lys Ile Phe Val Thr Glu Gly Leu Asn Lys Val Ile Pro Leu Leu 100 105 110Gln Lys Leu Lys Asn Ser Ala Lys Gly Pro Asp Val Pro Glu Thr Asn 115 120 125Gln Gln Cys Pro Ser Asn Thr Gly Met Thr Asp Pro Gln Arg Arg Gln 130 135 140Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Arg Asp Glu Arg Glu Ile145 150 155 160Pro Pro Phe Leu Glu Gly Ala Pro Pro Ser Val Ile Asp Glu Phe Tyr 165 170 175Asn Leu Leu Lys Thr Asp Glu Asn Lys Thr Asp Gln Gln Thr Glu Ala 180 185 190Asp Val Glu Ala Phe Ile Asn Arg Leu Gly Gly Ser Tyr Lys Val Arg 195 200 205Phe Thr Gln Phe Met Glu Glu Val Lys Lys Ala Arg Ala Asp Tyr Glu 210 215 220Arg Ile His Gln Gln Ala Val Ala Arg Phe Ser Pro Ala Ala Lys Asp225 230 235 240Ala Asp Ala Arg Met Ser Ala Ile Ala Asp Ser Pro His Leu Thr Thr 245 250 255Arg Gln Lys Ser Gln Gln Ile Gln Ala Ile Met Asp Ser Leu Ser Glu 260 265 270Ser Val Arg Arg Glu Ile Ile Asn Ala Leu Ser Pro Gln Glu Lys Gly 275 280 285Pro Asp Val Pro Glu Thr Asn Gln Gln Cys Pro Ser Asn Thr Gly Met 290 295 300Thr Asp Lys Asn Pro Ser Lys Met Glu Ser Lys Thr Gly Glu Asn Gln305 310 315 320Asp Arg Pro Val Leu Leu Gly Gly Trp Glu Asp Arg Asp Pro Lys Asp 325 330 335Glu Glu Ile Leu Glu Leu Leu Pro Ser Ile Leu Met Lys Val Asn Glu 340 345 350Gln Ser Lys Asp Glu Tyr His Leu Met Pro Ile Lys Leu Leu Lys Val 355 360 365Ser Ser Gln Val Val Ala Gly Val Lys Tyr Lys Met Asp Val Gln Val 370 375 380Ala Arg Ser Gln Cys Lys Lys Ser Ser Asn Glu Lys Val Asp Leu Thr385 390 395 400Lys Cys Lys Lys Leu Glu Gly His Pro Glu Lys Val Met Thr Leu Glu 405 410 415Val Trp Glu Lys Pro Trp Glu Asn Phe Met Arg Val Glu Ile Leu Gly 420 425 430Thr Lys Glu Val 43565286PRTArtificial SequenceOv103-RAL2 fusion protein 65Asp Leu Leu Ser Glu Ala Gly Asp Phe Phe Thr Lys His Phe Thr Asp1 5 10 15Ile Lys Ser Leu Phe Ala Lys Asp Glu Lys Gln Leu Gln Gln Ser Val 20 25 30Asp Arg Val Lys Asp Leu Leu Ala Thr Ile Gln Asp Lys Met Ser Met 35 40 45Leu Gln Pro Leu Ala Asn Asp Met Gln Lys Thr Thr Leu Gly Lys Ile 50 55 60Gly Asp Leu Ile Ser Gln Val Asn Ser Phe Arg Glu Thr Met Ser Asn65 70 75 80Pro Lys Met Asp Phe Thr Asn Lys Glu Asn Lys Trp Glu Glu Leu Leu 85 90 95Lys Lys Ile Phe Val Thr Glu Gly Leu Asn Lys Val Ile Pro Leu Leu 100 105 110Gln Lys Leu Lys Asn Ser Ala Lys Gly Pro Asp Val Pro Glu Thr Asn 115 120 125Gln Gln Cys Pro Ser Asn Thr Gly Met Thr Asp Pro Gln Arg Arg Gln 130 135 140Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Arg Asp Glu Arg Glu Ile145 150 155 160Pro Pro Phe Leu Glu Gly Ala Pro Pro Ser Val Ile Asp Glu Phe Tyr 165 170 175Asn Leu Leu Lys Thr Asp Glu Asn Lys Thr Asp Gln Gln Thr Glu Ala 180 185 190Asp Val Glu Ala Phe Ile Asn Arg Leu Gly Gly Ser Tyr Lys Val Arg 195 200 205Phe Thr Gln Phe Met Glu Glu Val Lys Lys Ala Arg Ala Asp Tyr Glu 210 215 220Arg Ile His Gln Gln Ala Val Ala Arg Phe Ser Pro Ala Ala Lys Asp225 230 235 240Ala Asp Ala Arg Met Ser Ala Ile Ala Asp Ser Pro His Leu Thr Thr 245 250 255Arg Gln Lys Ser Gln Gln Ile Gln Ala Ile Met Asp Ser Leu Ser Glu 260 265 270Ser Val Arg Arg Glu Ile Ile Asn Ala Leu Ser Pro Gln Glu 275 280 28566297PRTArtificial SequenceOvRAL2-CPI2M fusion protein 66Pro Gln Arg Arg Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Arg1 5 10 15Asp Glu Arg Glu Ile Pro Pro Phe Leu Glu Gly Ala Pro Pro Ser Val 20 25 30Ile Asp Glu Phe Tyr Asn Leu Leu Lys Thr Asp Glu Asn Lys Thr Asp 35 40 45Gln Gln Thr Glu Ala Asp Val Glu Ala Phe Ile Asn Arg Leu Gly Gly 50 55 60Ser Tyr Lys Val Arg Phe Thr Gln Phe Met Glu Glu Val Lys Lys Ala65 70 75 80Arg Ala Asp Tyr Glu Arg Ile His Gln Gln Ala Val Ala Arg Phe Ser 85 90 95Pro Ala Ala Lys Asp Ala Asp Ala Arg Met Ser Ala Ile Ala Asp Ser 100 105 110Pro His Leu Thr Thr Arg Gln Lys Ser Gln Gln Ile Gln Ala Ile Met 115 120 125Asp Ser Leu Ser Glu Ser Val Arg Arg Glu Ile Ile Asn Ala Leu Ser 130 135 140Pro Gln Glu Lys Gly Pro Asp Val Pro Glu Thr Asn Gln Gln Cys Pro145 150 155 160Ser Asn Thr Gly Met Thr Asp Lys Asn Pro Ser Lys Met Glu Ser Lys 165 170 175Thr Gly Glu Asn Gln Asp Arg Pro Val Leu Leu Gly Gly Trp Glu Asp 180 185 190Arg Asp Pro Lys Asp Glu Glu Ile Leu Glu Leu Leu Pro Ser Ile Leu 195 200 205Met Lys Val Asn Glu Gln Ser Lys Asp Glu Tyr His Leu Met Pro Ile 210 215 220Lys Leu Leu Lys Val Ser Ser Gln Val Val Ala Gly Val Lys Tyr Lys225 230 235 240Met Asp Val Gln Val Ala Arg Ser Gln Cys Lys Lys Ser Ser Asn Glu 245 250 255Lys Val Asp Leu Thr Lys Cys Lys Lys Leu Glu Gly His Pro Glu Lys 260 265 270Val Met Thr Leu Glu Val Trp Glu Lys Pro Trp Glu Asn Phe Met Arg 275 280 285Val Glu Ile Leu Gly Thr Lys Glu Val 290 295

Claims

1. An immunogenic composition for preventing or treating infection with a filarial parasite, wherein the filarial parasite is Onchocerca volvulus, and wherein the immunogenic composition comprises at least one filarial parasite protein having at least 85% sequence identity to full length mature protein OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), or OVOC8227 (SEQ ID NO:30).

2. An immunogenic composition for preventing or treating infection with a filarial parasite, wherein the filarial parasite is Dirofilaria immitis, and wherein the immunogenic composition comprises at least one filarial parasite protein having at least 85% sequence identity to full length mature protein of OVOC8619 (SEQ ID NO:16), OVOC7083 (SEQ ID NO:17), OVOC4111 (SEQ ID NO:18), OVOC1808 (SEQ ID NO:19), OVOC11598 (SEQ ID NO:20), OVOC3901 (SEQ ID NO:21), OVOC10819 (SEQ ID NO:22), OVOC5395 (SEQ ID NO:23), OVOC12235 (SEQ ID NO:24), OVOC7908 (SEQ ID NO:25), OVOC7430 (SEQ ID NO:26), OVOC8936 (SEQ ID NO:27), OVOC5806 (SEQ ID NO:28), OVOC4665 (SEQ ID NO:29), OVOC8227 (SEQ ID NO:30), OVOC9988 (SEQ ID NO:31), or OVOC4230 (SEQ ID NO:32), or an ortholog thereof.

3. The immunogenic composition of claim 2, wherein the ortholog comprises a filarial parasite protein having at least 85% sequence identity to the full length of SEQ ID NO:33-49.

4. The immunogenic composition of either of claim 1, wherein the immunogenic composition further comprises an adjuvant.

5. The immunogenic composition of either of claim 1, wherein the immunogenic composition comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

6. The immunogenic composition of claim 5, wherein the at least two filarial parasite proteins are present in the immunogenic composition as a mixture.

7. The immunogenic composition of claim 5, wherein the at least two filarial parasite proteins are present in the immunogenic composition as a fusion protein comprising the amino acid sequences of the at least two filarial parasite proteins.

8. The immunogenic composition of claim 7, wherein the fusion protein optionally further comprises at least one linker sequence separating the at least two filarial parasite amino acid sequences.

9. A method of preventing infection with, or transmission of, O. volvulus, the method comprising administering an immunogenic composition of claim 1 to a subject in need thereof, wherein the immunogenic composition prevents or treats the infection.

10. The method of claim 9, wherein the immunogenic composition is administered to a subject at risk of O. volvulus infection, and the administration prevents infection with O. volvulus and/or prevents transmission of O. volvulus.

11. The method of claim 9, wherein the subject is a human.

12. A method of preventing an infection with D. immitis, the method comprising administering an immunogenic composition of claim 2 to a canine subject in need thereof, wherein the immunogenic composition prevents or treats the infection.

13. The method of claim 12, wherein the immunogenic composition is administered to a subject at risk of D. immitis infection, and the administration prevents infection with D. immitis.

14. A method of detecting infection with O. volvulus, comprising identifying in a specimen from a subject at least one filarial protein having at least 85% sequence identity to OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC10384 (SEQ ID NO:13), OVOC8422 (SEQ ID NO:14), OVOC9988 (SEQ ID NO:31), or OVOC6395 (SEQ ID NO:15), or an immunoreactive fragment thereof.

15. The method of claim 14, wherein the specimen comprises blood, a skin biopsy, or urine.

16. The method of claim 14, wherein the immunoreactive fragment comprises the amino acid sequence of OVOC10469_Pep2 (SEQ ID NO:51), OVOC3261_Pep1 (SEQ ID NO:52), OVOC3261_Pep3 (SEQ ID NO:53), OVOC10469_Pep1 (SEQ ID NO:54), OVOC10469_Pep3 (SEQ ID NO:55), OVOC3261_Pep2 (SEQ ID NO:56), OVOC5127_Pep1 (SEQ ID NO:57), OVOC5127_Pep2 (SEQ ID NO:58), OVOC5127_Pep4, (SEQ ID NO:59), OVOC5127_Pep5 (SEQ ID NO:60), and OVOC5127_PepX (SEQ ID NO:61).

17. The method of claim 14, wherein the method further comprises detecting at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

18. The method of claim 17, wherein the at least two filarial parasite proteins comprise Ov16 and OVOC3261.

19. The method of claim 17, wherein the at least four filarial parasite proteins comprise Ov16, OVOC3261, OVOC10469, and OVOC5127.

20. The method of claim 14, wherein the filarial parasite protein is detected by a method selected from the group consisting of ELISA, dipstick tests, lateral flow, microfluidic devices, luciferase immunoprecipitation systems, luminex, multiplex-formats, and microarrays.

21. A method of detecting infection with O. volvulus, comprising identifying in the blood of a subject, antibodies to at least one filarial protein having at least 85% sequence identity to OVOC10469 (SEQ ID NO:1), OVOC11950 (SEQ ID NO:2), OVOC10602 (SEQ ID NO:3), OVOC3261 (SEQ ID NO:4), OVOC5127 (SEQ ID NO:5), OVOC8491 (SEQ ID NO:6), OVOC6759 (SEQ ID NO:7), OVOC451 (SEQ ID NO:8), OVOC12329 (SEQ ID NO:9), OVOC3337 (SEQ ID NO:10), OVOC10264 (SEQ ID NO:11), OVOC4230 (SEQ ID NO:12), OVOC10384 (SEQ ID NO:13), OVOC8422 (SEQ ID NO:14), OVOC9988 (SEQ ID NO:31), or OVOC6395 (SEQ ID NO:15), or an immunoreactive fragment thereof.

22. The method of claim 21, wherein the immunoreactive fragment comprises the amino acid sequence of OVOC10469_Pep2 (SEQ ID NO:51), OVOC3261_Pep1 (SEQ ID NO:52), OVOC3261_Pep3 (SEQ ID NO:53), OVOC10469_Pep1 (SEQ ID NO:54), OVOC10469_Pep3 (SEQ ID NO:55), OVOC3261_Pep2 (SEQ ID NO:56), OVOC5127_Pep1 (SEQ ID NO:57), OVOC5127_Pep2 (SEQ ID NO:58), OVOC5127_Pep4, (SEQ ID NO:59), OVOC5127_Pep5 (SEQ ID NO:60), and OVOC5127_PepX (SEQ ID NO:61).

23. The method of claim 21, wherein the method comprises detecting at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

24. The method of claim 23, wherein the method further comprises detecting antibodies to at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten filarial parasite proteins.

25. The method of claim 24, wherein the at least two filarial parasite proteins comprise Ov16 and OVOC3261.

26. The method of claim 24, wherein the at least four filarial parasite proteins comprise Ov16, OVOC3261, OVOC10469, and OVOC5127.

27. The method of claim 21, wherein the filarial protein or antibody to the filarial protein are detected by a method selected from the group consisting of ELISA, dipstick tests, lateral flow, microfluidic devices, luciferase immunoprecipitation systems, luminex, multiplex-formats, and microarrays.

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