Interaction of laminin with Entamoeba histolytica cysteine proteinases and its effect on amebic pathogenesis.

The Entamoeba histolytica 27-kDa cysteine proteinases exhibit striking binding specificities for immobilized laminin over other components of the extracellular matrix, such as collagen and fibronectin. Inactivation of the proteinase with the active-site inhibitor L-trans-epoxysuccinyl-leucylamido(4-guanidino)butane abolishes laminin binding by the enzyme, and conversely, laminin inhibits cleavage of a fluorogenic dipeptide substrate of the amebic cysteine proteinase, suggesting that the substrate binding pocket of the enzyme is involved in the binding of laminin. The addition of laminin but not fibronectin or collagen to E. histolytica trophozoites significantly reduces amebic liver abscess formation in severe combined immunodeficient mice, further supporting the hypothesis that E. histolytica cysteine proteinases play an important role in amebic pathogenesis. The specific interaction of amebic proteinases with laminin may be exploited in designing new inhibitors of these enzymes.     

Serologic reactivity to purified recombinant and native 29-kilodalton peripheral membrane protein of pathogenic Entamoeba histolytica.

The 29-kDa peripheral membrane protein of Entamoeba histolytica has recently been demonstrated to have epitopes on pathogenic clinical isolates which were not detected by monoclonal antibodies on nonpathogenic isolates. To analyze the serological response to this protein, we tested 93 serum specimens (from 33 patients with amebic liver abscess, 7 patients with colitis, 2 patients with ameboma, 18 individuals harboring a nonpathogenic zymodeme strain, 10 healthy Mexican migrant workers, and 23 healthy controls) by enzyme-linked immunosorbent assay (ELISA) using immunoaffinity-purified native or recombinant protein. When tested by ELISA with the native antigen, 79% (26 of 33) of the serum specimens from patients with amebic liver abscess, 4 of 9 serum specimens from symptomatic patients with colitis or ameboma, and serum from one migrant worker were positive. None of the 18 subjects harboring a nonpathogenic strain or 23 control individuals were seropositive to the native antigen (sensitivity, 71%; specificity, 98%). Of 30 serum specimens from patients with amebic liver abscess tested with recombinant antigen, 27 were seropositive (90%). In addition, six patients with colitis or ameboma and two individuals who harbored a nonpathogenic strain were seropositive to the recombinant antigen. One healthy Mexican migrant worker tested positive by both ELISAs (sensitivity, 87%; specificity, 94%). Immunoblotting of 51 serum specimens to sodium dodecyl sulfate-denatured native 29-kDa protein was less sensitive (65%) than ELISA in detecting serum antibodies to the antigen. These results suggest a similar antibody response to native and recombinant antigens (r = 0.86) and support the potential utility of a quantitative assay with defined recombinant antigen for the serodiagnosis of invasive amebiasis in nonendemic areas in conjunction with other diagnostic tools.     

Isolation of antigens with proteolytic activity from Coccidioides immitis.

Three antigens with proteolytic activity have been isolated from crude, water-soluble fractions of the saprobic phase of the fungal pathogen Coccidioides immitis. Two proteinases, identified in our immunoelectrophoresis reference system as Ag11 and AgCS, were isolated from the soluble conidial wall fraction (SCWF). Ag11 was previously shown to be a serine proteinase and was characterized in this study as a 60-kilodalton (kDa) fraction by gel filtration (GF). The purified proteinase demonstrated little or no reactivity with 21 serum samples from coccidioidomycosis patients in the enzyme-linked immunosorbent assay; this may be due to limited presentation of this antigen to the host during the course of coccidioidomycosis. AgCS was separated by GF chromatography into two fractions identified by molecular masses of 39 and 19 kDa. Most proteolytic activity was shown by substrate gel electrophoresis to be associated with the lower-molecular-mass fraction. AgCS was reactive with 18 of the 21 serum samples and shown to be the major component of a heat-stable antigen previously reported to be immunospecific for C. immitis. The third antigen with proteolytic activity was isolated from the 5-day mycelial culture filtrate and identified by GF as a 56-kDa fraction. Uniformly high levels of immunoreactivity between 18 of the 21 patient sera and the 56-kDa antigen were demonstrated. Antigens with proteolytic activity may play important roles in fungus-host interactions as well as morphogenesis of the pathogen.     

Inactivation of various proteinase inhibitors and the complement system in human plasma by the 56-kilodalton proteinase from Serratia marcescens.

The interaction of the 56-kilodalton (kDa) proteinase from Serratia marcescens with human plasma activated C1 (C1) inhibitor, alpha 2-antiplasmin, and antithrombin III was investigated. The 56-kDa proteinase was not affected by these inhibitors; on the contrary, all the inhibitors were inactivated by the 56-kDa proteinase within 2 to 6 h. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that all three inhibitors showed decreases in molecular weight of approximately 8,000 to 10,000 as a result of proteolytic cleavage by the 56-kDa proteinase. The 56-kDa proteinase also inactivated serum complement within 2 to 6 h. The loss of inhibitory activity caused by the 56-kDa proteinase, together with the effects of endogenous serine proteinases, may facilitate tissue destruction and inflammation.     

Use of a recombinant 170-kilodalton surface antigen of Entamoeba histolytica for serodiagnosis of amebiasis and identification of immunodominant domains of the native molecule.

We expressed the gene that encodes one of the major surface antigens of Entamoeba histolytica, the 170-kDa protein (1,270 amino acids), as a glutathione S-transferase fusion protein containing amino acids 1 to 1202 (lacking the putative transmembrane and cytoplasmic regions) and as separate fusion proteins containing each of three major domains of the 170-kDa molecule. Lysates from bacteria induced to express one of these proteins were used as the target antigens in a Western blot (immunoblot) analysis to determine whether a recombinant 170-kDa antigen could serve as the basis for a serologic test used to detect invasive amebiasis and whether there are differences in humoral immunogenicity among the three major domains of the 170-kDa antigen. Among patients with invasive amebiasis from three major areas where the disease is endemic and two sites in the United States, 54 (90%) of 60 had antibodies to the recombinant 170-kDa protein. Among 37 patients from regions where the disease is endemic and 20 patients from the United States without amebic disease, 1 (2%) of 57 had antibodies to the recombinant 170-kDa protein. We found significant differences in seroreactivity to each of three major domains of the molecule among patients seropositive for the complete construct, ranging from 100% seroreactivity with the fusion protein containing the domain designated cysteine rich and 89% seropositivity with the fusion protein incorporating a portion of the region designated cysteine poor to only 9% seropositivity for the fusion protein containing the pseudorepeat domain. Our study indicates that a serologic test based on the recombinant 170-kDA antigen could serve as a highly sensitive and specific test for acute invasive amebiasis.     

Role of the Entamoeba histolytica cysteine proteinase in amebic liver abscess formation in severe combined immunodeficient mice.

Evidence from in vitro studies suggest that the Entamoeba histolytica cysteine proteinase plays a role in the tissue lysis and cytopathic effects seen in invasive amebiasis. We used affinity-purified antibodies against a recombinant E. histolytica cysteine proteinase to demonstrate that the proteinase is present extracellularly in amebic liver abscesses in mice with severe combined immunodeficiency (SCID mice). Treatment of E. histolytica trophozoites with specific cysteine proteinase inhibitor E-64 blocked or greatly reduced liver abscess formation at 48 h in SCID mice. Our study suggests an important role for a functional cysteine proteinase in amebic liver abscess formation.     

Investigation of extracellular elastase, acid proteinase and phospholipase activities as putative virulence factors in clinical isolates of Aspergillus species

We investigated the production of extracellular elastase, acid proteinase and phospholipase enzyme activities in clinical isolates of Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger and any possible correlation between the existence of these enzymes and development of invasive aspergillosis. A total of 73 strains (45 A. fumigatus, 23 A. flavus, 5 A. niger) isolated from patients with invasive aspergillosis (n = 55), superficial aspergillosis (n = 5), allergic bronchopulmonary aspergillosis (n = 1), and from those colonized with Aspergillus (n = 12) were included in the study. The enzymatic activities were tested on solid media supplemented with the corresponding substrates. Elastase activity was detected in 95.6, 82.6, and 0% of A. fumigatus, A. flavus, and A. niger isolates, respectively. Acid proteinase activity was detected only for A. fumigatus and in 20 of 45 isolates belonging to this species. Phospholipase activity was present in all strains of A. fumigatus and A. niger but in none of the isolates of A. flavus. No statistical correlation could be established between the existence of elastase or acid proteinase activity and development of invasive disease (p > 0.05). While high phospholipase production was found to be associated with development of invasive aspergillosis (p < 0.01), not all isolates that caused invasive diseases showed high phospholipase activities.     

A surface amebic cysteine proteinase inactivates interleukin-18

Amebiasis is a major cause of morbidity and mortality worldwide. Invasion by Entamoeba histolytica trophozoites causes secretion of proinflammatory cytokines from host epithelial cells, leading to a local acute inflammatory response, followed by lysis of colonic cells. Extracellular cysteine proteinases from amebic trophozoites are key virulence factors and have a number of important interactions with host defenses, including cleavage of immunoglobulin G (IgG), IgA, and complement components C3 and C5. Amebic lysates have also been shown to activate the precursor to interleukin 1-beta (proIL-1beta), mimicking the action of caspase-1. IL-18 is also a central cytokine, which induces gamma interferon (IFN-gamma) and activates macrophages, one of the main host defenses against invading trophozoites. Because proIL-18 is also activated by caspase-1, we evaluated whether amebic proteinases had a similar effect. Instead, we found that recombinant proIL-18 was cleaved into smaller fragments by the complex of surface-associated and released amebic proteinases. To evaluate the function of an individual proteinase from the complex pool, we expressed an active surface proteinase, EhCP5, which is functional only in E. histolytica. Recombinant EhCP5 expressed in Pichia pastoris had kinetic properties similar to those of the native enzyme with respect to substrate specificity and sensitivity to proteinase inhibitors. In contrast to the activation of proIL-1beta by amebic lysates, the purified proteinase cleaved proIL-18 and mature IL-18 to biologically inactive fragments. These studies suggest that the acute host response and amebic invasion result from a complex interplay of parasite virulence factors and host defenses. E. histolytica may block the host inflammatory response by a novel mechanism, inactivation of IL-18.     

Protection of gerbils from amebic liver abscess by immunization with recombinant Entamoeba histolytica 29-kilodalton antigen.

The goal of our study was to obtain a highly conserved Entamoeba histolytica recombinant antigen for study as a subunit amebiasis vaccine. We screened a Uni-Zap cDNA library of E. histolytica (strain HM1:IMSS) with human immune sera and isolated a dominant 804-bp cDNA clone. A 33-kDa fusion protein expressed from the cDNA clone was determined by monoclonal antibody binding, DNA hybridization, and nucleotide sequence to be the complete E. histolytica 29-kDa antigen. Serum antibodies to the recombinant protein were detected by enzyme-linked immunosorbent assay in 80% of subjects from Egypt and South Africa with amebic liver abscess. Similar results were found with the native 29-kDa protein. Native and recombinant 29-kDa antigens induced proliferation of lymphocytes harvested from patients with amebic liver abscess (P < 0.01 compared with controls). Intraperitoneal immunization of gerbils with the recombinant fusion protein (10 micrograms) with Titermax adjuvant elicited an antigen-specific serum immunoglobulin G antibody response and was partially protective (54%) against intrahepatic challenge with 5 x 10(5) virulent axenic trophozoites (strain HM1:IMSS). In summary, the recombinant form of the E. histolytica 29-kDa antigen demonstrated serologic specificity for amebic liver abscess, exhibited conserved T-cell epitopes, and was effective as a subunit vaccine in an experimental animal model of amebic liver abscess.     

CP30, a cysteine proteinase involved in Trichomonas vaginalis cytoadherence

We describe here the participation of a Trichomonas vaginalis 30-kDa proteinase (CP30) with affinity to the HeLa cell surface in attachment of this parasite to host epithelial cells. The CP30 band is a cysteine proteinase because its activity was inhibited by E-64, a thiol proteinase inhibitor. In two-dimensional substrate gel electrophoresis of total extracts of the trichomonad isolate CNCD 147, three spots with proteolytic activity were detected in the 30-kDa region, in the pI range from 4.5 to 5.5. Two of the spots (pI 4.5 and 5.0) bound to the surfaces of fixed HeLa cells corresponding to the CP30 band. The immunoglobulin G fraction of the rabbit anti-CP30 antiserum that recognized a 30-kDa band by Western blotting and immunoprecipitated CP30 specifically inhibited trichomonal cytoadherence to HeLa cell monolayers in a concentration-dependent manner and reacted with CP30 at the parasite surface. CP30 degraded proteins found on the female urogenital tract, including fibronectin, collagen IV, and hemoglobin. Interestingly, CP30 digested fibronectin and collagen IV only at pH levels between 4.5 and 5.0. Moreover, trichomonosis patients whose diagnosis was confirmed by in vitro culture possessed antibody to CP30 in both sera and vaginal washes, and CP30 activity was found in vaginal washes. Our results suggest that surface CP30 is a cysteine proteinase necessary for trichomonal adherence to human epithelial cells.     

Purification and immunologic characterization of a 30-kDa cysteine proteinase ofEntamoeba histolytica

A 30-kDa cysteine proteinase was purified from extracts of axenically grown trophozoites ofEntamoeba histolytica strain HM1:IMSS. The purification procedure involved two consecutive chromatographic steps. Sequence analysis revealed high similarity with histolysin and with other 27-kDa cysteine proteinase. Western-blot analysis using F(ab)₂ fragments of a polyclonal antibody raised against the purified enzyme revealed that when the amebic extract was prepared in the absence of proteinase inhibitors there were many positive bands ranging in relative molecular weight from 115 to 12.5kDa, but when the extract was prepared in the presence of proteinase inhibitors there was only a single 30-kDa positive band. Similar results were obtained with immunoprecipitates. This phenomenon would suggest the formation of multimer aggregates of the 30-kDa cysteine proteinase after partial proteolysis.     

Increased expression of Candida albicans secretory proteinase, a putative virulence factor, in isolates from human immunodeficiency virus-positive patients.

The increased prevalence and the severity of oropharyngeal candidiasis in human immunodeficiency virus (HIV)-positive patients are attributed exclusively to the virus-induced immune deficiency of the host. The present study was aimed at answering the question of whether Candida albicans secretory proteinase, a putative virulence factor of the opportunistic C. albicans yeast, has any potential influence on the clinical manifestation of oropharyngeal candidiasis in HIV-positive patients. We measured the secretory proteinase activities of clinical C. albicans isolates from the oropharynges of either HIV-positive individuals (n = 100) or a control group (n = 122). The mean secretory proteinase activity of C. albicans isolates from the HIV-positive group (4,255 +/- 2,372 U/liter) was significantly higher compared with that of isolates from the control group (2,324 +/- 1,487 U/liter) (P < 0.05). The higher level of secretory proteinase activity in the culture supernatants of individual C. albicans isolates correlated with the increased level of proteinase expression on the cell surface, as revealed by cytofluorometry, and with higher levels of secretion of the immunodetectable protein, as shown by Western blotting (immunoblotting). Proteinase activity within the population of C. albicans isolates from HIV-positive individuals was independent of the patient's clinical disease stage and the CD4+/CD8+ cell numbers. Furthermore, no correlation of the proteinase activities with the C. albicans serotype was found, although C. albicans serotype B was significantly more frequent in the HIV-positive group (40%) compared with that in the control group (12%). However, a positive correlation of proteinase activity to antifungal susceptibility was evident.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mutational analysis of the tobacco etch potyviral 35-kDa proteinase: identification of essential residues and requirements for autoproteolysis.

The tobacco etch potyvirus (TEV) polyprotein is processed by three virus-encoded proteinases, termed Nla, HC-Pro, and the 35-kDa proteinase. The 35-kDa proteinase is derived from the amino-terminal region of the polyprotein. Analysis of polyproteins containing beta-glucuronidase fused to the expected carboxy terminus of the 35-kDa proteinase confirmed the previously identified Tyr304-Ser305 dipeptide as the cleavage site between the 35-kDa proteinase and HC-Pro. The 35-kDa proteinase of TEV was unable to catalyze proteolysis when synthetic substrate polyproteins were supplied in a bimolecular or trans reaction, suggesting that processing occurs by an autolytic mechanism. The results of a mutational analysis within the 35-kDa proteolytic domain indicated that His214, Asp223, Ser256, and Asp288 were required for optimal autoproteolytic activity. Replacement of Ser256 with either Thr or Cys resulted in low but detectable proteinase activity, as did substitution of Asp223 and Asp288 with Glu. These results are consistent with the hypothesis that the 35-kDa proteinase resembles cellular serine-type proteinases, with Ser256 functioning as the nucleophilic residue within the active site. Cleavage mediated by the 35-kDa proteinase has been shown previously to occur after polyprotein synthesis in wheat germ extracts and transgenic plants, but not in rabbit reticulocyte lysate. We were able to demonstrate that processing in vitro may require a heat-labile factor present in wheat germ extracts.     

Biochemical characterization of Porphyromonas (Bacteroides) gingivalis collagenase.

A protease was purified from Porphyromonas gingivalis 1101, a clinical isolate, by sequential sodium dodecyl sulfate-polyacrylamide gel electrophoresis, substrate diffusion gel electrophoresis, and electroelution. The enzyme cleaved radiolabeled human basement membrane type IV collagen and the synthetic collagen peptide substrate for eukaryotic collagenases. It was inactivated by the thiol protease inhibitor N-ethylmaleimide but not by EDTA or EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] and activated by reducing agents such as beta-mercaptoethanol. The enzyme exists as an active precursor protein of molecular mass 94 kDa and undergoes proteolytic cleavage to 75-, 56-, and 19-kDa forms. Biotin-labeled collagen bound specifically to the 94-kDa form of the protein and to its cleavage products in ligand blots, suggesting a role for this enzyme not only in collagen degradation but also in adhesion to collagenous substrata.     

Inverse relation between disease severity and expression of the streptococcal cysteine protease, SpeB, among clonal M1T1 isolates recovered from invasive group A streptococcal infection cases

The streptococcal cysteine protease (SpeB) is one of the major virulence factors produced by group A streptococci (GAS). In this study we investigated if differences exist in SpeB production by clonally related M1T1 clinical isolates derived from patients with invasive infections. Twenty-nine of these isolates were from nonsevere cases and 48 were from severe cases, including streptococcal toxic shock syndrome (STSS) and necrotizing fasciitis (NF) cases. The expression and amount of the 28-kDa SpeB protein produced were determined by quantitative Western blotting, and protease activity was measured by a fluorescent enzymatic assay. A high degree of variation in SpeB expression was seen among the isolates, and this variation seemed to correlate with the severity and/or clinical manifestation of the invasive infection. The mean amount of 28-kDa SpeB protein and cysteine protease activity produced by isolates from nonsevere cases was significantly higher than that from STSS cases (P = 0.001). This difference was partly due to the fact that 41% of STSS isolates produced little or no SpeB compared to only 14% of isolates recovered in nonsevere cases. Moreover, the cysteine protease activity among those isolates that expressed SpeB was significantly lower for STSS isolates than for isolates from nonsevere cases (P = 0.001). Increased SpeB production was also inversely correlated with intact M protein expression, and inhibition of cysteine protease activity blocked the cleavage of the surface M protein. Together, the data support the existence of both an "on-off" and a posttranslational regulatory mechanism(s) controlling SpeB production, and they suggest that isolates with the speB gene in the "off" state are more likely to spare the surface M protein and to be isolated from cases of severe rather than nonsevere invasive infection. These findings may have important implications for the role of SpeB in host-pathogen interactions via regulation of the expression of GAS virulence genes and the severity of invasive disease.     

The 96-kilodalton antigen as an integral membrane protein in pathogenic Entamoeba histolytica: potential differences in pathogenic and nonpathogenic isolates.

A surface antigen (EH-96) of Entamoeba histolytica was demonstrated to be a plasma membrane antigen by immunoprecipitation of metabolically 35S-labeled antigen from live trophozoites, Triton X-114 detergent extracts, and plasma membrane-enriched fractions prepared by concanavalin A membrane stabilization and differential centrifugation. In addition, the antigen was localized to the plasma membrane by electron microscopy with colloidal gold. Antigen from E. histolytica strains immunoprecipitated with specific immunoglobulin M (IgM) or IgG2b monoclonal antibody was identical by one-dimensional peptide mapping with N-chlorosuccinimide. Additionally, antigen from different axenically cultivated amebae was demonstrated to be identical by N-chlorosuccinimide peptide mapping, as were peptide maps of IgG and IgM monoclonal antibody-purified antigen. The 96-kilodalton (kDa) surface antigen was identified on four axenically cultivated pathogenic isolates and on three polyxenically cultivated pathogenic isolates (zymodeme II) of E. histolytica but was absent or present in lesser quantity on six nonpathogenic polyxenically cultivated isolates. The 96-kDa antigen was detected in liver abscess fluid from four patients with amebic abscesses by enzyme-linked immunosorbent assay (ELISA) and immunoprecipitation. Two-dimensional gel electrophoresis profiles of the 96-kDa antigen purified from abscess material or from polyxenically cultivated trophozoites demonstrated that the antigens were related to the 96-kDa antigen found in axenically cultivated organisms.     

Recognition of the galactose- or N-acetylgalactosamine-binding lectin of Entamoeba histolytica by human immune sera.

Cure of amebic liver abscess is associated with resistance to recurrent invasive amebiasis and the development of a humoral and cell-mediated immune response. We determined whether human immune sera contain blocking antibody for the 170-kilodalton (kDa) galactose or N-acetylgalactosamine (Gal/GalNAc)-binding lectin of Entamoeba histolytica. By Western blot (immunoblot) of whole amebae subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all eight immune sera studied here prominently recognized a 170-kDa amebic protein. Western blot of the purified Gal/GalNAc lectin with pooled human immune sera (PHIS) confirmed that the 170-kDa band was the adherence lectin. Immunoprecipitation of [35S]methionine-metabolically-labeled amebae with the antilectin monoclonal antibody H8-5 and with PHIS demonstrated that the 170-kDa lectin was the major antigen recognized by PHIS. The in vitro adherence of E. histolytica trophozoites to CHO cells at 4 degrees C was inhibited by prior exposure of amebae to greater than or equal to 1.0% PHIS. The humoral response to the Gal/GalNAc-binding lectin of the parasite may contribute to the development of protective immunity against invasive amebiasis.     

Proteinase excretion ability of Candida albicans species isolates in clinical specimens

AIM: The aim of the study was to establish whether the Candida (C.) albicans species isolates from samples of the patient digestive, respiratory and genitourinary systems differed in the ability of proteinase excretion. METHODS: A total of 1009 isolates of the C. albicans species obtained from 1009 clinical specimens of the digestive, respiratory and genitourinary systems of 666 patients were examined. All samples were inoculated onto Sabourauds glucose agar and incubated aerobically at 37 degrees C for 3-7 days. Identification of C. albicans was done by standard and commercial tests. To test the proteinase excretion ability, we employed the Odds and Abbott method. The isolates were inoculated on a culture medium containing agar, glucose, vitamins and beef albumin fraction V (pH = 3.2), and stored for 7 days at 30 degrees C. Each isolate was tested twice, with the results read by the same person. Development of thready colonies in a milky-white field was considered as positive finding. Study results are shown in easy-to-consult tables. Distribution differences were assessed by chi2-test. RESULTS: Of 1009 C. albicans species isolates obtained from clinical specimens, 695 (68.9%) had the proteinase excreting ability. The presence of this enzyme was demonstrated in 72.7% of the species isolates obtained from samples of the digestive system, 65.8% of isolates from respiratory system and 59.6% of isolates from genitourinary system. Analysis of chi2-test results showed no statistically significant difference in the ability of C. albicans species isolates from specimens obtained from digestive, respiratory and genitourinary systems to excrete proteinase. DISCUSSION: The results of the present study are in agreement with the results of most other researchers reporting on the proteinase excreting ability to be demonstrated in 40%-80% of C. albicans isolates. In the present study, the ability to excrete proteinase was demonstrated in 68.9% of C. albicans species isolates obtained from clinical samples. CONCLUSION: The ability to excrete proteinase was demonstrated in 59.6%-72.7% of C. albicans species isolates obtained from the patient digestive, respiratory and genitourinary systems. Analysis of the results yielded no statistically significant difference in the proteinase excreting ability among the isolates obtained from digestive, respiratory and genitourinary systems.     

Protection of gerbils from amebic liver abscess by vaccination with a 25-mer peptide derived from the cysteine-rich region of Entamoeba histolytica galactose-specific adherence lectin

The protozoan parasite Entamoeba histolytica causes extensive morbidity and mortality through intestinal infection and amebic liver abscess. Here we show that immunization of gerbils with a single keyhole limpet hemocyanin-coupled 25-mer peptide derived from the 170-kDa subunit of the E. histolytica galactose-binding adhesin is sufficient to confer substantial protection against experimentally induced amebic liver abscesses. Vaccination provided total protection in 5 of 15 immunized gerbils, and abscesses were significantly smaller (P < 0.01) in the remaining vaccinated animals. The degree of protection correlated with the titer of antibodies to the peptide, and results of passive transfer experiments performed with SCID mice were consistent with a role for antibodies in protection. In addition, parenteral or oral vaccination of gerbils with 13-amino-acid subfragments of the peptide N-terminally fused to the B subunit of cholera toxin also significantly inhibited liver abscess formation (P < 0.05). These data indicate that small peptides derived from the galactose-binding adhesin administered by the parenteral or oral route can provide protection against amebic liver abscess and should be considered as components of a subunit vaccine against invasive amoebiasis.     

Clinical features and management of amebic liver abscess. Experience from 29 patients

Summary 29 patients with amebic liver abscess were evaluated in a study to examine clinical picture, laboratory data, epidemiology, radiologic methods, and therapy. Since the clinical picture was unspecific a considerable amount of misdiagnoses occurred, and often originated from pulmonary symptoms. To establish diagnosis one should rely on the triad with positive amebic serology, intrahepatic scanning defects, and clinical picture with fever, right upper quadrant pain, and hepatomegaly. Nearly all patients had an exposure history of travel or immigration from endemic areas in the tropics. Medical therapy with metronidazole alone is highly effective and leads to defervescence and clinical improvement usually within 3–5 days. Invasive procedures, such as needle aspiration or surgical drainage of the abscess are rarely needed; these invasive methods neither shorten the course of the disease nor improve prognosis.Abbreviations E. histolytica Entamoeba histolytica - ALA Amebic liver abscess - ESR Erythrocyte sedimentation rate