Histolytica Protein Research Articles

The unusual transhydrogenase of Entamoeba histolytica

We have expressed and purified a protein fragment from Entamoeba histolytica. It catalyses transhydrogenation between analogues of NAD(H) and NADP(H). The characteristics of this reaction resemble those of the reaction catalysed by a complex of the NAD(H)- and NADP(H)-binding subunits of proton-translocating transhydrogenases from bacteria and mammals. It is concluded that the complete En. histolytica protein, which, along with similar proteins from other protozoan parasites, has an unusual subunit organisation, is also a proton-translocating transhydrogenase. The function of the transhydrogenase, thought to be located in organelles which do not have the enzymes of oxidative phosphorylation, is not clear.

Systemic antiamebic antibodies induced orally in mice: detection by an ELISA test using preserved Entamoeba histolytica-antigens without enzymatic inhibitors.

We have previously reported a method to preserve amebic antigens without using enzymatic inhibitors (1,2). The fraction obtained by this method shows reduced proteolytic activity even in the presence of SDS and 2-ME, which usually stimulate amebic enzymatic activity. This method preserves more E. histolytica proteins than preparations containing iodoacetamide, as demonstrated by SDS-PAGE. In addition, these proteins retain their antigenicity, and are recognized by antibodies present in sera of patients with amebic liver abscess in Western blots (WB) (Flores-Castaneda et al., submitted). Moreover, the protein fraction can withstand several cycles of freeze thawing without losing antigenicity even for several years. As a result of this stability, the preserved fraction can be used as antigen in immunoassay tests (U.S. Patents 5,459,042 and 5,861,263). Asymptomatic individuals without previous history of invasive amebiasis, stool-negative, and living in amebic endemic zones, may still have systemic antiamebic antibodies. Such antibodies could be explained by previous E. histolytica subclinical infections, previous E. dispar infections with crossreacting antibodies, or ingestion of E. histolytica antigens that stimulate the production of systemic antibodies in the absence of invasive infection. Animal models have helped in our understanding of many clinical observations. Although mice are not the best model to study E. histolytica infection, their immune system is well known, and they provide advantages over other animal models (3). The dual goal of our study was to develop an ELISA test using preserved amebic antigens without protease inhibitors, and to stimulate the production of systemic antibodies by oral immunization with freeze-dried trophozoites.

DNA vaccination with the serine rich Entamoeba histolytica protein (SREHP) prevents amebic liver abscess in rodent models of disease

Amebiasis remains one of the leading parasitic causes of death worldwide. A vaccine that prevented amebic liver abscess would significantly reduce mortality from this disease. To test the feasibility of a DNA vaccine to prevent amebic liver abscess, we immunized both mice and gerbils with plasmid DNA encoding the serine rich Entamoeba histolytica protein (SREHP). Animals receiving the SREHP DNA vaccine developed both antibody and cell mediated immune responses that recognized amebic trophozoites. A single dose of the SREHP DNA vaccine protected 80% of vaccinated mice and 60% of vaccinated gerbils from developing amebic liver abscess after direct hepatic inoculation of amebic trophozoites. Our study indicates that DNA vaccination with SREHP can provide high levels of protection against amebic liver abscess in animal models of disease.

Diagnosis and management of amebiasis.

s of the annual meeting of the American Society of Tropical Medicine and Hygiene, San Juan, Puerto Rico, 18-22 October1998. North- brook, IL: 1998. 56. Gilman R, Islam M, Paschi S, Goleburn J, Ahmad F Comparison of con- ventional and immunofluorescent techniques for the detection of Entamoeba histolytica in rectal biopsies. Gastroenterology 1980;78:435-9. 57. Rails PW. Focal inflammatory disease of the liver. Radiol Clin North Am 1998; 36:377-89. 58. Kimura K, Stoopen M, Reeder MM, Moncada R. Amebiasis: modern diagnostic imaging with pathological and clinical correlation. Semin Roent- genol 1997; 32:250-75. 59. Huston CD, Petri WA Jr. Amebiasis. In: Rakel RE, ed. Conn's current therapy. New York: W.B. Saunders (in press). 60. Rajak CL, Gupta S, Jain S, Chawla Y, Gulati M, Suri S. Percutaneous treatment of liver abscesses: needle aspiration versus catheter drainage. AJR Am J Roentgenol 1998; 170:1035-9. 61. Tandon A, Jain AK, Dixit VK, Agarwal AK, Gupta JP. Needle aspiration in large amoebic liver abscess. Trop Gastroenterol 1997; 18:19-21. 62. Petri WA Jr, Ravdin JI. Protection of gerbils from amebic liver abscess by immunization with the galactose-specific adherence lectin of Entamoeba histolytica. Infect Immun 1991;59:97-101. 63. Zhang T, Stanley SL Jr. Protection of gerbils from amebic liver abscess by This content downloaded from 157.55.39.23 on Sun, 26 Mar 2017 18:12:48 UTC All use subject to http://about.jstor.org/terms CID 1999;29 (November) Amebiasis1125 immunization with a recombinant protein derived from the 170-kilodalton surface adhesin of Entamoeba histolytica. Infect Immun 1994; 62:2605-8. 64. Zhang T, Cieslak PR, Foster L, Kuntz-Jenkins C, Stanley SL Jr. Antibodies to the serine rich Entamoeba histolytica protein (SREHP) prevent amoebic liver abscess in severe combined immunodeficient (SCID) mice. Parasite Immunol 1994; 16:225-30. 65. Soong CJG, Kain KC, Abd-Alla M, Jackson TFHG, Ravdin JI. A recombinant cysteine-rich section of the Entamoeba histolytica galactose inhib- itable lectin is efficacious as a subunit vaccine in the Gerbil model of amebic liver abscess. J Infect Dis 1995; 171:645-51. 66. Mann BJ, Burkholder BV, Lockhart LA. Protection in a gerbil model of amebiasis by oral immunization with Salmonella expressing the galactose N-acetyl D-galactosamine inhibitable lectin of Entamoeba histolytica. Vac- cine 1997; 15:659-63. 67. Huston CD, Petri WA Jr. The host-pathogen interaction in amebiasis: Is a vaccine feasible? Eur J Clin Microbiol Infect Dis 1998; 17:601-14. 68. Salata RA, Pearson RP, Murphy CF, Ravdin JI. Interaction of human leu- kocytes with Entamoeba histolytica: killing of virulent amebae by the activated macrophage. J Clin Invest 1985; 76:491-9. 69. Denis M, Chadee K. Human neutrophils activated by interferon-gamma and tumor necrosis factor-alpha kill Entamoeba histolytica trophozoites in vitro. J Leukoc Biol 1989;46:270-4. This content downloaded from 157.55.39.23 on Sun, 26 Mar 2017 18:12:48 UTC All use subject to http://about.jstor.org/terms

Induction of a spectroscopically defined transition by guanidinium hydrochloride on a recombinant calcium binding protein from Entamoeba histolytica

Sequence analysis and metal ion binding studies reported earlier have established that the calcium binding protein (CaBP) from the parasitic ameboid Entamoeba histolytica protein has four canonical EF hand motifs which bind calcium. Equilibrium denaturation studies on both the apo and the holo forms of this protein indicate the presence of stable transition intermediates at low denaturant concentrations as revealed by the binding of the non-specific hydrophobic dye ANS. Fast reaction kinetics shows that the binding of the Gdn + ions at or near the Ca 2+ sites in the N-terminal domain influences metal ion binding to the sites in the C-terminal domain. Isothermal calorimetric titrations performed using low GdnHCl concentrations reveal the presence of two binding sites of low affinity, both being endothermic in nature. Thus the stabilization of CaBP observed at low GdnHCl concentration represents a native-like intermediate, with the Gdn + ions mimicking Ca 2+ binding at the N-terminal domain of this protein.

The small GTP-binding protein RacG regulates uroid formation in the protozoan parasite Entamoeba histolytica.

Entamoeba histolytica is a protozoan parasite that invades human intestine leading to ulceration and destruction of tissue. Amoebic movement and phagocytosis of human cells is accompanied by characteristic changes in cell morphology. Amoebae become polarized, developing a frontal pseudopod and a well-defined rear zone of membrane accumulation designated the uroid. In motile eukaryotic cells, a phenomenon that contributes to movement is the capping of receptors at the cell surface. During the capping process, E. histolytica concentrates ligand-receptor complexes in the uroid. Interestingly, some of these surface receptors are involved in the survival of the parasite. While looking for regulators of capping and uroid formation, we identified RacG, an E. histolytica protein that is homologous to human Rac1. This protein belongs to the Rac subfamily of small GTPases implicated in interactions between the actin cytoskeleton and the membrane of mammalian cells. Cloning of the EhracG gene and analysis of the protein activity either in murine fibroblasts or in E. histolytica revealed that EhRacG induces a characteristic Rac phenotype. When expressed in amoebae, an EhRacG-V12 mutant protein not only deregulated cell polarity, but also caused a defect in cytokinesis. Analysis of the cytoskeleton in amoebae bearing this mutant revealed that F-actin concentrated at the periphery of the cell. In addition, the number and localization of uroids were modified. These results suggest a role for EhRacG in amoebic morphogenesis and cytokinesis.

Longitudinal study of the antibody response to recombinant Entamoeba histolytica antigens in patients with amebic liver abscess.

Serology is a critical component in the diagnosis of amebic liver abscess. However, in areas endemic for amebiasis there is a high background level of seropositivity for amebiasis (owing to previous infection with Entamoeba histolytica), which may complicate the interpretation of a positive serologic test result. Recently, we reported that serologic tests based on recombinant E. histolytica antigens might offer improved diagnosis of current invasive amebiasis because they apparently differentiated active infection from past exposure to the parasite. To confirm this finding, we have performed a longitudinal study on 20 patients with amebic liver abscess by examining their seroreactivity over time with recombinant versions of two major E. histolytica proteins, the serine rich E. histolytica protein (SREHP), and the 170-kD subunit of the galactose-specific adhesin. We found that more than 50% of the patients examined had become seronegative by one or both recombinant tests within 180 days of their diagnosis of amebic liver abscess. In the case of the recombinant SREHP-based tests, 12 patients had become seronegative 90 days after presentation. In contrast, all patients remained seropositive by a standard conventional test, an indirect hemagglutination test, at more than six months after presentation. Our study shows that patients lose seroreactivity with the recombinant SREHP or 170-kD antigen-based tests more rapidly than with a conventional serologic test; this may make them useful for the serologic diagnosis of amebiasis in endemic areas.

Mucosal Immunogenicity of a Holotoxin-Like Molecule Containing the Serine-RichEntamoeba histolyticaProtein (SREHP) Fused to the A2Domain of Cholera Toxin

One strategy for the induction of mucosal immune responses by oral immunization is to administer the antigen in conjunction with cholera toxin. Cholera toxin consists of one A polypeptide (CTA) which is noncovalently linked to five B subunits (CTB) via the A2 portion of the A subunit (CTA2). Coupling of antigens to the nontoxic B subunit of cholera toxin may improve the immunogenicity of antigens by targeting them to GM1 ganglioside on M cells and intestinal epithelial cells. Here, we describe the construction of a translational fusion protein containing the serine-rich Entamoeba histolytica protein (SREHP), a protective amebic antigen, fused to a maltose binding protein (MBP) and to CTA2. When coexpressed in Escherichia coli with the CTB gene, these proteins assembled into a holotoxin-like chimera containing MBP-SREHP-CTA2 and CTB. This holotoxin-like chimera (SREHP-H) inhibited the binding of cholera toxin to GM1 ganglioside. Oral vaccination of mice with SREHP-H induced mucosal immunoglobulin A (IgA) and serum IgG antiamebic antibodies and low levels of mucosal anti-CTB antibodies. Our studies confirm that the genetic coupling of antigens to CTA2 and their coexpression in E. coli can produce holotoxin-like molecules that are mucosally immunogenic without the requirement for supplemental cholera toxin, and they establish the SREHP-H protein as a candidate for evaluation as a vaccine to prevent amebiasis.

Short Report: Identification of B Cell Epitopes in the Serine-Rich Entamoeba histolytica Protein

The serine-rich Entamoeba histolytica protein (SREHP) has been shown to be a protective antigen in animal models of amebic liver abscess when delivered by either parenteral or oral routes of immunization, and antibodies to SREHP can prevent amebic liver abscess in severe combined immunodeficient mice. To identify B cell epitopes of the SREHP molecule that could serve as the basis for a peptide-based vaccine, we synthesized overlapping peptides spanning the amino acid sequence of SREHP, and looked at the reactivity of serum samples from five individuals with amebic liver abscess to the overlapping peptides. We found that most of the epitopes recognized by serum samples from patients with amebic liver abscess map to the hydrophilic dodecapeptide or octapeptide repeats of SREHP, but there was no universal epitope recognized by all five serum samples. In addition, we show that synthetic peptides that include the epitopes of SREHP recognized in the mapping study are immunogenic in animals and can generate antibodies that recognize SREHP.

Progress towards development of a vaccine for amebiasis.

The application of molecular biologic techniques over the past decade has seen a tremendous growth in our knowledge of the biology of Entamoeba histolytica, the causative agent of amebic dysentery and amebic liver abscess. This approach has also led to the identification and structural characterization of three amebic antigens, the serine-rich Entamoeba histolytica protein (SREHP), the 170-kDa subunit of the Gal/GalNAc binding lectin, and the 29-kDa cysteine-rich protein, which all show promise as recombinant antigen-based vaccines to prevent amebiasis. In recent studies, an immunogenic dodecapeptide derived from the SREHP molecule has been genetically fused to the B subunit of cholera toxin, to create a recombinant protein capable of inducing both antiamebic and anti-cholera toxin antibodies when administered by the oral route. Continued progress in this area will bring us closer to the goal of a cost-effective oral combination "enteric pathogen" vaccine, capable of inducing protective mucosal immune responses to several clinically important enteric diseases, including amebiasis.

Oral immunization with attenuated vaccine strains of Vibrio cholerae expressing a dodecapeptide repeat of the serine-rich Entamoeba histolytica protein fused to the cholera toxin B subunit induces systemic and mucosal antiamebic and anti-V. cholerae antibody responses in mice.

Entamoeba histolytica is a significant cause of morbidity and mortality worldwide. The serine-rich E. histolytica protein (SREHP) is a surface-expressed trophozoite protein that includes multiple hydrophilic tandem repeats. A purified fusion protein between the dodecapeptide repeat of SREHP and cholera toxin B subunit (CTB) has previously been shown to be immunogenic in mice after oral inoculation when cholera toxin is coadministered as an immunoadjuvant. We engineered a live attenuated El Tor Vibrio cholerae vaccine strain, Peru2, to express the SREHP-12-CTB fusion protein to the supernatant from either a plasmid [Peru2 (pETR5.1)] or from a chromosomal insertion (ETR3). Vector strains were administered orally to germfree mice that were subsequently housed under nongermfree conditions; mice received one (day 0) or two (days 0 and 14) inoculations. No immunoadjuvant or cholera holotoxin was administered. Mice that received two inoculations of Peru2(pETR5.1) had the most pronounced antiamebic systemic and mucosal immunologic responses. Less marked, but significant, anti-SREHP serum immunoglobulin G antibody responses were also induced in mice that received either one or two oral inoculations of strain ETR3. Anti-V. cholerae responses were also induced, as measured by the induction of serum vibriocidal antibodies and by serum and mucosal anti-CTB antibody responses. These results suggest that V. cholerae vector strains can be successful delivery vehicles for the SREHP-12-CTB fusion protein, to induce mucosal and systemic antiamebic and anti-V. cholerae immune responses. The magnitude of these responses is proportional to the amount of SREHP-12-CTB produced by the vector strain.

Expression of the serine rich Entamoeba histolytica protein (SREHP) in the avirulent vaccine strain Salmonella typhi TY2 χ4297 ( Δcya Δcrp Δasd): safety and immunogenicity in mice

Infection by the intestinal protozoan parasite Entamoeba histolytica remains a significant threat to health in much of the world. Here we describe the successful expression of the serine rich Entamoeba histolytica protein (SREHP), a protective antigen of ameba, in an attenuated vaccine strain Salmonella typhi TY2 χ4297 ( Δcya Δcrp Δasd). The attenuation of S. typhi TY2 χ4297 was not altered by expression of the SREHP-maltose binding protein (MBP) fusion protein and mice parenterally vaccinated with S. typhi TY2 χ4297 expressing SREHP-MBP developed serum anti-amebic and anti-LPS antibodies. S. typhi TY2 χ4297 expressing SREHP-MBP represents a prototype combination vaccine designed to prevent both amebiasis and typhoid fever.

Progress towards development of a vaccine for amebiasis.

The application of molecular biologic techniques over the past decade has seen a tremendous growth in our knowledge of the biology of Entamoeba histolytica, the causative agent of amebic dysentery and amebic liver abscess. This approach has also led to the identification and structural characterization of three amebic antigens, the serine-rich Entamoeba histolytica protein (SREHP), the 170-kDa subunit of the Gal/GalNAc binding lectin, and the 29-kDa cysteine-rich protein, which all show promise as recombinant antigen-based vaccines to prevent amebiasis. In recent studies, an immunogenic dodecapeptide derived from the SREHP molecule has been genetically fused to the B subunit of cholera toxin, to create a recombinant protein capable of inducing both antiamebic and anti-cholera toxin antibodies when administered by the oral route. Continued progress in this area will bring us closer to the goal of a cost-effective oral combination "enteric pathogen" vaccine, capable of inducing protective mucosal immune responses to several clinically important enteric diseases, including amebiasis.

Immunodominant Entamoeba histolytica antigens recognized by serum and intestinal antibodies after local or systemic immunization of mice with glutaraldehyde fixed trophozoites.

We performed an immunoblot analysis of the main E. histolytica proteins recognized by immune sera and intestinal fluids of Balb/c mice immunized with glutaraldehyde fixed trophozoites (GFT) by intragastric, rectal and intraperitoneal routes, to determine if there were differences in the amebic antigens immunodominantly recognized at mucosal and systemic levels. The antigen patterns recognized by mice immunized via intraperitoneal and rectal routes were complex and similar suggesting that the immunization route (systemic or local), does not influence the recognition pattern elicited at mucosal or systemic levels. However, the number of amebic bands recognized after intragastric immunization was very low. The molecular weights of the principal amebic proteins recognized by serum antibodies were 150-130, 116, 104, 84, 56, 42, 18, and 16 kDa. The intestinal fluids of mice immunized via intraperitoneal and rectal routes contained antibodies that recognized five bands of 220-200, 150-134, 93-84, 43-41, and 16-14 kDa. These results suggest that there are differences in the number of immunodominant amebic antigens recognized at mucosal and systemic levels. Moreover we found that the bands of 150, 39 and 19 kDa. were mainly recognized by IgG, whereas the bands of 116, 93, and 16 were mainly recognized by IgM, indicating differences between the antigens immunodominantly recognized by serum antibodies from different isotypes.

Oral immunization with an attenuated vaccine strain of Salmonella typhimurium expressing the serine-rich Entamoeba histolytica protein induces an antiamebic immune response and protects gerbils from amebic liver abscess

Attenuated salmonellae represent attractive candidates for the delivery of foreign antigens by oral vaccination. In this report, we describe the high-level expression of a recombinant fusion protein containing the serine-rich Entamoeba histolytica protein (SREHP), a protective antigen derived from virulent amebae, and a bacterially derived maltose-binding protein (MBP) in an attenuated strain of Salmonella typhimurium. Mice and gerbils immunized with S. typhimurium expressing SREHP-MBP produced mucosal immunoglobulin A antiamebic antibodies and serum immunoglobulin G antiamebic antibodies. Gerbils vaccinated with S typhimurium SREHP-MBP were protected against amebic liver abscess, the most common extraintestinal complication of amebiasis. Our findings indicate that the induction of mucosal and immune responses to the amebic SREHP antigen is dependent on the level of SREHP-MBP expression in S. typhimurium and establish that oral vaccination with SREHP can produce protective immunity to invasive amebiasis.

Oral immunization with the dodecapeptide repeat of the serine-rich Entamoeba histolytica protein (SREHP) fused to the cholera toxin B subunit induces a mucosal and systemic anti-SREHP antibody response

The intestinal protozoan parasite Entamoeba histolytica causes amebic dysentery, a major cause of morbidity worldwide. The induction of a mucosal antibody response capable of blocking amebic adhesion to intestinal cells could represent an approach to preventing E. histolytica infection and disease. Here we describe the expression of a chimeric protein containing an immunogenic dodecapeptide derived from the serine-rich E. histolytica protein (SREHP), fused to the cholera toxin B subunit (CtxB). The CtxB-SREHP-12 chimeric protein was purified from Escherichia coli lysates and retained the critical GM1 ganglioside-binding activity of the CtxB moiety. Mice fed the CtxB-SREHP-12 fusion protein along with a subclinical dose of cholera toxin developed mucosal immunoglobulin A and immunoglobulin G and systemic antibody responses that recognized recombinant and native SREHP. Our study confirms the feasibility of inducing mucosal immune responses to immunogenic peptides by their genetic fusion to the CtxB subunit and identifies the CtxB-SREHP-12 chimeric protein as a candidate oral vaccine to prevent E. histolytica infection.

Entamoeba histolytica suppresses gamma interferon-induced macrophage class II major histocompatibility complex Ia molecule and I-A beta mRNA expression by a prostaglandin E2-dependent mechanism.

The surface expression of class II major histocompatibility complex immune-associated (Ia) antigen is a principal accessory function of macrophages for antigen-specific T-cell activation and immunoregulation. To explore the mechanisms of impaired cell-mediated immunity in invasive amebiasis, we investigated the effects of Entamoeba histolytica proteins on gamma interferon (IFN-gamma)-induced Ia expression by murine bone marrow-derived macrophages. Pretreatment of macrophages with secreted (conditioned medium) or whole soluble amebic proteins inhibited the induction of IFN-gamma-induced surface Ia antigen expression by 30 to 61% but had no effect on surface Ia molecules already expressed. By Northern (RNA) blot analysis, amebic proteins suppressed IFN-gamma-induced macrophage I-A beta mRNA accumulation by 36% but did not alter the constitutive levels of actin mRNA expression. E. histolytica stimulated macrophages to produce high levels of prostaglandin E2 (PGE2) as determined by reverse-phase high-pressure liquid chromatography and quantification by PGE2-specific radioimmunoassay. Inhibition of PGE2 biosynthesis with the cyclooxygenase inhibitor indomethacin abrogated ameba-induced suppression of Ia antigen by 60%, whereas exogenously added PGE2 decreased IFN-gamma-induced macrophage Ia expression by 44%. Our results suggest that the mechanism whereby E. histolytica suppresses IFN-gamma-induced macrophage surface Ia molecule synthesis and I-A beta mRNA expression is by stimulating the production of PGE2, which acts in an autocrine fashion for immunoregulation. E. histolytica subverting critical macrophage accessory function via PGE2 biosynthesis is a novel strategy which the parasite uses to suppress host defenses.

The Serine-rich Entamoeba histolytica Protein Is a Phosphorylated Membrane Protein Containing O-Linked Terminal N-Acetylglucosamine Residues

Previously, we described the isolation of a cDNA clone and the gene encoding a protective antigen of the protozoan parasite Entamoeba histolytica, the serine-rich Entamoeba histolytica protein (SREHP). The derived amino acid sequence of the SREHP cDNA clone was remarkable for a high serine content (52/233 amino acids), a putative signal sequence, multiple hydrophilic dodecapeptide and octapeptide tandem repeats, and a hydrophobic C-terminal putative membrane-spanning region. Here, we show that SREHP is modified by the addition of phosphate at serine residues, O-linked terminal N-acetylglucosamine residues, and by acylation. When the SREHP gene is expressed in baculovirus transformed Sf-9 cells, the product is also phosphorylated and glycosylated and is localized to the plasma membrane of the insect cells. The native SREHP molecule also serves as a potent chemoattractant for amebic trophozoites. The data presented here suggest that SREHP is a unique membrane protein with phosphorylation and glycosylation patterns usually associated with nuclear or cytoplasmic proteins.

Passive immunization during experimental amebic liver-abscess development

The role of antibodies during invasive amebic infection in humans or in experimental animals has not been clearly established. Early investigations by Swartzwelder and Avant (1952) found partial protection in dogs passively immunized with whole blood from animals made resistant to rectal inoculation of Entamoeba histolytica trophozoites. Sepflveda et al. (1971) also induced partial protection against ameba challenge in hamsters injected intraperitoneally with serum from a patient with amebic liver abscess. Virtually no further report appeared until 1992, when Cieslak et al. carried out a passive immunization study in severe combined immunodeficient mice (SCID), suggesting that antiamebic antibodies are protective, in contrast to several clinical findings that tend to support the opposite (Kretschmer 1986). Thus, although specific circulating antibodies can be detected at a relatively early stage of invasive amebiasis and last for months or even years after the completion of treatment (Knobloch and Mannweiler 1983; Krupp and Powell 1971; Osinsaya and Warhust 1980), antibodies neither cure nor prevent the establishment of disease (Krupp 1970). On the other hand, under certain experimental conditions a partial protection can be induced in rodents that have been actively immunized with purified amebic proteins; however, in spite of the presence of circulating antiamebic antibodies, larger liver lesions have developed in immunized animals than in controls (Petri and Ravdin 1991). Recently, Zhang et al. (1994) reported the induction of protective immunity in gerbils immunized with a recombinant E. histolytica protein, although antibody titers and the degree of protection were unrelated. We performed passive immunization studies in hamsters using purified and characterized antiamebic immunoglobulins obtained from rabbits or hamsters.

Immunogenicity of the recombinant serine rich Entamoeba histolytica protein (SREHP) amebiasis vaccine in the African Green Monkey

We report the first study in non-human primates of the safety and immunogenicity of a recombinant vaccine designed to prevent amebic liver abscess. In a pilot study, a recombinant vaccine containing the serine rich Entamoeba histolytica protein (SREHP) attached to a maltose binding protein ( SREHP MBP ), which has been shown to be effective in preventing amebic liver abscess in rodent models of infection, was used to immunize two African Green Monkeys. Vaccination with SREHP MBP resulted in no systemic side-effects. The monkeys receiving the SREHP MBP protein developed antibodies that recognized the recombinant SREHP MBP molecule, the native SREHP protein, and the surface of amebic trophozoites. Antiserum from SREHP MBP -vaccinated monkeys could block the adhesion of E. histolytica trophozoites to mammalian cells, a feature that may correlate with vaccine efficacy. Attempts to produce amebic liver abscess in naive African Green Monkeys by direct hepatic inoculation with virulent E. histolytica trophozoites was not successful, suggesting this species is probably not suitable for vaccine efficacy studies.