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PubMed2003

Ultrastructural study of Entamoeba invadens encystation and excystation.

In the life cycle of Entamoeba species, the cyst and all the processes associated to it have been poorly studied. Entamoeba invadens, a serpent's parasite, has been commonly accepted as a model for the study of encystation and excystation. Here we analyzed through scanning and transmission electron microscopy the in vitro morphological differentiation of both processes. During encystation, the formation of an irregular net of fibrillar material on the surface of precysts was observed. In thin sections of cryofixed and cryosubstituted specimens, abundant vacuoles containing a microfibrillar material of similar appearance to the structural components of the cyst wall were found in the cytoplasm. Assays with a calcofluor probe on cryosections of encysting trophozoites and precysts showed the presence of fluorescent circular cytoplasmic structures. In the cyst stage, the fluorescence was located on the surface. During excystation, the detachment of the metacyst from the cyst wall was observed through scanning electron microscopy. Metacysts endocyting amorphous material which may correspond to cyst wall residues were commonly found. By transmission electron microscopy the formation of a crescent-shaped space between the plasma membrane and the cyst wall was observed. Abundant small electrondense bodies were found in the cytoplasm. Many of them were in close apposition to the plasma membrane and frequently some of them were seen projecting towards this newly formed space. Our results suggest that the microfibrillar content of the vacuoles corresponds to the cyst wall material, that the electrondense bodies may be involved in the excystation process, and that part of the cyst wall residues may be endocyted by the parasite.

Animals↗
PubMedDate not supplied

[Evaluation of the activity of polymorphonuclear neutrophils against Entamoeba histolytica antigens].

This paper evaluates the production of superoxide as an indicator of the activity of neutrophils from healthy individuals, stimulated with Entamoeaba histolytica membrane antigens, chemical stimulators and antigens from other parasites. The kinetic nitroblue tetrazolium reduction test was used. No significant differences were observed in the control group as to superoxide production by neutrophils primed with Entamoeba histolytica antigens or Cisticercus cellulosae, Toxoplasma gondii and Giardia lamblia antigens. These results allow us to conclude that Entamoeba histolytica membrane antigens do not induce superoxide production by neutrophils, which contradicts previous statements on suppresed activity of neutrophils after they get in contact with Entamoeaba histolytica antigens.

Animals↗
PubMed2005

High prevalence rate of Entamoeba histolytica asymptomatic infection in a rural Mexican community.

The frequency of Entamoeba histolytica and Entamoeba dispar infection was analyzed in a rural community in the state of Morelos, Mexico, using polymerase chain reaction (PCR). Sociodemographic variables as risk factors for the infection were assessed. Results highlighted the number of individuals with intestinal parasites (43.1%) in the community, indicating extensive fecalism. A high frequency of E. histolytica asymptomatic infection, higher than E. dispar infection (13.8% versus 9.6%), was detected by PCR. Anti-amebic antibody levels (IgG) in serum and saliva (IgA) samples were not associated with E. histolytica intestinal infection. These findings suggest a predominant distribution of E. histolytica strains of low invasive potential in this community.

Animals↗
PubMedDate not supplied

Detection of Entamoeba histolytica using polymerase chain reaction in pus samples from amebic liver abscess.

BACKGROUND AND OBJECTIVE: Direct demonstration of Entamoeba histolytica by conventional microscopy and in vitro culture in pus obtained from amebic liver abscess (ALA) is often unsuccessful. We evaluated polymerase chain reaction (PCR) for detection of E. histolytica DNA in such pus. METHODS: Species-specific primers were used for the amplification of E. histolytica DNA from liver pus obtained from 30 patients with ALA. Patients with pyogenic liver abscess and sterile (autoclaved) pus spiked with Entamoeba dispar and bacteria (Escherichia coli, Klebsiella spp. and Bacteroides spp.) were used as negative controls. RESULTS: PCR was positive in 83% of pus specimens from patients with ALA, and was negative in all 25 pus specimens obtained from pyogenic abscess and autoclaved pus spiked with known bacteria. Sensitivity and specificity of PCR were 83% and 100%, respectively. The overall positivity of PCR was higher compared to serological tests. CONCLUSION: PCR may be a more reliable and better alternative diagnostic modality for ALA.

Animals↗
PubMedDate not supplied

Relationship between free radicals produced by entamoeba histolytica and its proteases complex activity.

Entamoeba histolytica is a parasite which causes health problems and there has been many approaches to know what is the factor causing its pathogenicity. In the present work, we assayed if the production of free radicals by the amoeba, has a relationship with the proteases activity. When we test the DMSO action (free radicals quenching activity) the specific activity of the proteases complex of the parasite were affected also. At 33.3% (V/V) concentration of DMSO it was present a maximal decrease of the initial activity (about 46% decrease), for to a higher concentrations existing a trend to recuperate the original activity, suggesting that the free radicals are an important factor for the hydrolysis grade of the protein substrate. All the differences except those between 46.7 and 66.6%, were significantly different compared with the control without any addition. The effects of Probucol and Probucol plus DMSO, compared to those caused by Metronidazol (MZ). We can observe that the quenchers caused a decrease on proteases activity similar to that of MZ (which is an antiparasite drug) and it was of c.a. 58% of activity decrease. These data suggest that the action of both, free radicals and proteases complex of Entamoeba histolytica, can account for the pathogenicity of the parasite.

Animals↗
PubMed1990

[Prevalence of Entamoeba histolytica and other intestinal parasites in a neighborhood of the Mara Municipality, Zulia State, Venezuela].

A parasitologic survey of a low socioeconomic level community from Mara County, Zulia State, Venezuela was done. A single stool specimen per person from 327 individuals was obtained and examined by hematoxylin stained fecal smears and formol-ether concentrates. The overall parasitic infection rate was 92%. Multiple infections were present in 89% of the population. The amebic prevalence rate (APR) was 41.5%. Entamoeba histolytica was found in 9.2% of the individuals, most of them being asymptomatic. Haematophagous trophozoites were observed in two symptomatic cases. Entamoeba coli was the most frequent protozoan (23.5%). The APR was high in all groups. In males it was higher in 7-12 year old children and adolescents, and in females it was higher in adults. Giardia lamblia was the most frequent flagellate (18%) and among the helminths, Trichuris trichiura (82.8%) and Ascaris lumbricoides (74.6%). The high rates of parasitic infection, multiple parasitosis and amebic prevalence reflect the low socioeconomic conditions in the population.

Adolescent↗
PubMed1978

[Culture and establishment of new Mexican Strains of Entamoeba histolytica].

Mixed and monoxenic cultures of rectal exudates from three patients with amebiasis of colon, and of feces from three asymptomatic carriers of Entamoeba histolytica obtained after administration of a saline cathartic, and from a case of exudate of the perianal cutaneous lesion border produced by this parasite, were done. The culture media used were for: mixed culture, Boeck and Drbohlav modified culture and, for monoxenic culture, Diamond's monophasic TP-S-1 medium without vitamin mixture and using bovine serum. In order to inhibit the bacterial growth in the mixed culture, streptomycin sulphate and pencillin G were used. In order to eliminate the bacterial flora kanamicin sulphate, chloramphenicol, gentamycin and disodic carbenicillin were added. Mixed cultures of seven strains of Entamoeba histolytica were obtained: HM22:IMSS, HM23:IMSS, HM24:IMSS, HM26:IMSS, HM27:IMSS, HM28:IMSS y HM29:IMSS, and monoxenic culture with Bacteroids symbiosus of the strains HM22:IMSS and HM27:IMSS, with a previous adaptation in mixed culture. The adaptation of HM26:IMSS strain in monoxenic culture was obtained by means of direct seeding of the rectal exudate with Fusobacterium symbiosus.

Animals↗
PubMed1990

[The mechanism of natural resistance in the rat to Entamoeba histolytica].

The data presented in this paper may be summarized as follows: 1) intraportal injection of a virulent strain (HM-1) of Entamoeba histolytica in Wistar rats, of both sexes, gives way to non-progressive microscopic changes, characterized by a rapid leukocytic reaction surrounding the amebas, disappearance of the parasites within 5 hours, and total lack of hepatic damage; 2) leucopenia only modifies the previous description by the fact that there are no leucocytes around the amebas, although these disappear in the same time, showing, at this moment, an early and prominent vacuolar degeneration; 3) hypocomplementaemia shows the same results as leucopenia; 4) fragments and extracts from various tissues from the rat and hamster show variable degrees of interference with the viability of axenic amebas of Entamoeba histolytica conserved in culture; only a minimal part of the interference to be due to the activity of the complement which is present in the tissue extracts.

Animals↗
PubMed1990

[Proteins associated with actin: profilin in the trophozoites of Entamoeba histolytica].

The presence and distribution of profilin, an actin binding protein that regulates the actin organization, in trophozoites of Entamoeba histolytica, strain HM1-IMSS was examined using polyclonal antibodies against calf spleen profilin. The results shows: (1) in Entamoeba extracts a peptide that comigrates with spleen profilin and representing 0.86% of the total cells proteins, (2), this peptide is immunoprecipitated by antibodies antiprofilin, and (3) stained cells with the antibodies shows the protein distributed in cortical region associated to plasma membrane and th cytoplasm. These results show that a peptide similar to spleen profilin is present in the trophozoites of E. histolytica, supporting the idea that the profilin has been conserved through evolution. The presence of this protein would be crucial in the reversible control of soluble and polymerized actin required in the amoeboid movement.

Actins↗
PubMed1990

An Entamoeba histolytica antigen devoid of enzymatic activity, but endowed with immunogenic and protective properties.

One of the most relevant features of Entamoeba histolytica extracts is their great enzymatic activity. The consequence of this peculiarity is that these extracts digest not only extrinsic proteins but also those within the parasite, causing instability during extract preparation which makes their characterization more difficult and hinders their use in immunoprophylaxis. This report presents evidence which indicates that treatment of E. histolytica extracts with a pH2 acid shock for 30 min at 37 degrees C irreversibly, as shown through its reactivity with polyclonal E. histolytica antisera and through its capacity to induce antibody formation and to protect against intrahepatic challenge with E. histolytica. Among the foremost activities in the field of immunoparasitology are the identification, isolation, purification and characterization of parasite antigens, particularly because of their potential use in immunoprophylaxis. Given the world wide distribution of amebiasis, particularly in undeveloped countries, various researchers have given special importance to the antigenic characterization of the causal agent, Entamoeba histolytica, and to its ability to induce protective immunity in several animal models. Initial studies demonstrated that the parasite produces a variety of proteolytic enzymes which play an important role in the immune response and pathogenesis of the infected host. Additionally, these enzymes cause autodegradation of the amebic extracts, their use in immunoprophylaxis, since proteins germane in inducing protection could be degraded. Even though enzymatic activity may be abolished through specific inhibitors, these are very toxic and therefore not adequate for biological material to be tried on humans.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Purification and partial characterization of the major cysteine protease from Entamoeba invadens.

The purification and partial characterization of a major protease from the parasitic protozoon of reptiles, Entamoeba invadens, is described. The enzyme has a molecular mass of 28 kDa, and three distinct isoelectric points at pH 4.7, 5.7 and 6.3, respectively. As an endopeptidase the enzyme digests denatured protein substrates, such as azocasein with an optimal turnover rate at pH 4.8 with a temperature optimum of 48 degrees C. The protease exhibits exopeptidase activity towards arginine containing dipeptide derivatives. Thus, it splits the chromogenic substrates N-benzyloxycarbonyl-arginine-arginine-4-methoxy-beta-naphthylamide and arginine-arginine-4-methoxy-beta-naphthylamide in the ratio of velocities of 3:1. The kinetic constants for the hydrolysis of N-benzyloxycarbonyl-arginine-arginine-4-methoxy-beta-naphthylamide are: Km 22 microM and kcat 172 s-1. The enzyme is activated by the thiol reagents cysteine and dithiothreitol and is inhibited by typical cysteine protease inhibitors, such as cystatin, E-64, iodoacetamide and p-chloromercuribenzoate. Although in many of its characteristics it resembles the cathepsin B-like cysteine protease from Entamoeba histolytica, the two enzymes were found to be immunologically different.

Animals↗
PubMedDate not supplied

[Importance of Trichomonas tenax and Entamoeba gingivalis protozoa in the human oral cavity].

Trichomonas tenax and Entamoeba gingivalis are protozoa found in the human oral cavity. Morphological and ultrastructural characteristics of those parasites were reviewed and then studied with the scanning electron microscope in this paper. Based on previous epidemiological studies, the relationship between periodontal disease and those protozoa was analysed. Evaluation of the pathogenicity of Trichomonas tenax and Entamoeba gingivalis was also part of the discussion of this study.

Adolescent↗
PubMedDate not supplied

[Periodontal diseases with Entamoeba gingivalis].

At the beginning of our century Entamoeba gingivalis was considered to be a pathogenic bacteria, capable to induce parodontal lesions. Later on it was also found in healthy persons, and the germ was less interesting from the medical view-point. In the present study the authors report their findings concerning E. gingivalis in 135 patients with various stomatological affections including: dental caries, parodontopathies, pulpitis, gangrene, ulcero-necrotic stomatitis etc. The study was started following the discovery of the amoeba in the gingival exsudate of a male aged 19 years with chronic superficial marginal parodontopathy, who, after a treatment with metronidazol, was cured. Entamoeba gingivalis belongs to the Rhizopoda class, together with E. dysenteriae, and E. coli, but, in contrast with these strains it does not have resistance forms (cysts). Oral amoeba were evidenced in 18 out of 78 patients with parodontal lesions (23.07%), in the gingival exsudate, the purulent secretion from parodontal pouches, in the dental tartar, the alveolar fluid following extraction etc. In 117 students from the Faculty of Stomatology, and in 57 patients with various other stomatological affections these germs were not found in any of the abovementioned products. Microscopic examination of fresh preparations, and of Giemsa-stained smears was the main method for the detection of the amoeba. The etiopathogenic role of E. gingivalis is re-examined in discussions regarding certain parodontopathies.

Adult↗
PubMed1989

Chemoattractant activity of Entamoeba histolytica for human polymorphonuclear neutrophils.

The ability of axenic Entamoeba histolytica trophozoites and amoebic protein preparations to stimulate chemotaxis of human polymorphonuclear neutrophils (PMN) was evaluated. Virulent E. histolytica (strain HM1:IMSS) stimulated chemotaxis (delta distance = 0.55 +/- 0.02 mm, P less than 0.01 vs. control medium). Sonicated (100 micrograms protein/ml) or homogenized (500 micrograms protein/ml) virulent amoebae also significantly stimulated PMN chemotaxis, whereas preparations of the nonpathogenic "Entamoeba-like" Laredo strain did not stimulate chemotaxis. Preparations of subcellular fractions of E. histolytica demonstrated maximal stimulation of PMN chemotaxis existed in nonvesiculated membranes and the supernatant from plasma membranes.

Animals↗
PubMed1989

Studies on the microflora associated with xenic cultures of Entamoeba gingivalis.

The microflora associated with xenic stock cultures (ATCC 30927) of Entamoeba gingivalis, the major protozoan of the human oral cavity, were isolated and identified as Citrobacter diversus, Yersinia enterocolitica, Acinetobacter anitratus and Pseudomonas maltophilia. In studies to determine whether the bacterial isolates were able to utilize rice starch as a sole carbon source, Y. enterocolitica exhibited excellent growth in rice starch minimal medium and TYSGM-9 medium (with rice starch), but growth was weak in TYSGM-9 medium (without rice starch). C. diversus, A. anitratus and P. maltophilia exhibited poor growth in rice starch minimal medium, but they produced excellent growth in TYSGM-9 medium with or without rice starch. In order to determine the effect of the rice starch hydrolysis on Entamoeba growth, the filtrate from each isolate grown in rice starch minimal medium was added to an E. gingivalis culture grown in TYSGM-9 medium. The filtrate from a Y. enterocolitica culture grown in rice starch minimal medium enhanced E. gingivalis growth, but the filtrates from cultures of C. diversus, A. anitratus and P. maltophilia suppressed E. gingivalis growth. This supported the concept that Y. enterocolitica is capable of metabolizing rice starch into intermediate products, which in turn can be utilized by the amoeba.

Acinetobacter↗
PubMed1987

A double-antibody sandwich ELISA for the detection of Entamoeba histolytica antigen in stool samples of humans.

A double-antibody sandwich ELISA was developed to detect detergent-solubilized antigens of Entamoeba histolytica in stool samples of humans. The test system was evaluated for its methodical and diagnostic sensitivity and specificity. In recovery experiments the lower limit of detection was 400 ng E. histolytica (HK9) protein/ml stool, corresponding to approximately 2000 amoebic trophozoites/ml stool. Samples of 97 patients with suspected intestinal amoebiasis were examined. Specific antigens were detected by ELISA (= positive reaction) in 14 (93%) out of 15 stool samples containing trophozoites of E. histolytica. In contrast, 68 (93%) of 73 samples with other protozoa, including Entamoeba coli, E. hartmanni, Endolimax nana, Iodamoeba buetschlii and Giardia lamblia, did not react in the test system (= negative reaction). The test was shown to detect only trophozoites of E. histolytica and not the cyst stage. This fact could facilitate the differentiation between cyst carriers and persons excreting trophozoites. The results of this preliminary study justify a further large scale evaluation of the test system.

Antigens, Protozoan↗
PubMed1980

Comparison of conventional and immunofluorescent techniques for the detection of Entamoeba histolytica in rectal biopsies.

Two comparative studies of six staining techniques in detection of Entamoeba histolytica in rectal biopsies were performed. The techniques evaluated were direct and indirect fluorescent antibody technique, and four conventional stains: PAS, H & E, Trichrome, and PTAH). On ethanol-fixed tissue, the direct fluorescent antibody technique was the most sensitive, detecting amoebas in 84% of rectal biopsies from 19 aspirate-positive dysentery patients. PAS staining was specific, almost as sensitive, and amoebas could be rapidly detected. The other three conventional methods, however, detected amoebas in only 53--58% of the aspirate-positive patients and required considerable efforts to find the amoebas. The indirect fluorescent antibody technique performed on ethanol-fixed tissues was comparable to the direct technique. The same technique used on formalin-fixed tissues had high background staining, but amoebas could still be detected more efficiently than with conventional staining. Following 20 hr or more treatment with metronidazole, no patient had amoebas detectable in their rectal biopsies by any technique used. Patients with amoebic dysentery and necrotic ulcers on protoscopy had amoebas detected more frequently in rectal biopsy than did patients with mild disease as determined by proctoscopy. Entamoeba histolytica in rectal biopsies can be detected most easily in ethanol-fixed tissue stained with either a fluorescent antibody or with PAS. If amoebas are not detected in the rectal biopsy, amoebic colitis cannot be excluded.

Adult↗
PubMed1980

Dynamic changes on the surface of Entamoeba induced by antibodies.

Antibody and Concanavalin A bound to the cell surface of Entamoeba induced segregation of these complexes towards the uroid, followed by an evagination of the membrane portion that contained the components redistributed. This vesiculated cap was released into the medium. Con A polarized all Con A-binding glycoproteins from the surface of the ameba while with antibodies only some of the surface antigens were movilized towards the cap area. This phenomenon was temperature dependent as shown in E. invadens. It was also shown that most of surface antigens that form the cap could not be removed from the cell by proteolysis, suggesting that they were tightly bound to plasma membrane. The interest to define the process of capping in Entamoeba, is because its unique fashion to loss a portion of plasma membrane, and to determine if this phenomenon is related with the ability of ameba to outcome immune injury.

Antibodies, Bacterial↗