Entamoeba

Entamoeba histolytica , the protozoan parasite that phagocytoses bacteria and host cells, has a vesicle/vacuole-filled cytosol like that of macrophages. In contrast, the infectious cyst form has four nuclei and a chitin wall. Here, anti-chitinase antibodies identified hundreds of small secretory vesicles in encysting E. invadens parasites and in E. histolytica trophozoites overexpressing chitinase under an actin gene promoter. Abundant small secretory vesicles were also identified with antibodies to the surface antigen Ariel and with a fluorescent substrate of cysteine proteinases. Removal of an N-terminal signal sequence directed chitinase to the cytosol. Addition of a C-terminal KDEL peptide, identified on amebic BiP, retained chitinase in a putative endoplasmic reticulum, which was composed of a few vesicles of mixed sizes. A putative Golgi apparatus, which was Brefeldin A sensitive and composed of a few large, perinuclear vesicles, was identified with antibodies to ADP-ribosylat...

Enteric protozoan parasites, which are spread by the fecal-oral route, are important causes of diarrhea (Giardia duodenalis) and amebic dysentery (Entamoeba histolytica). Cyst walls ofGiardiaandEntamoebahave a single layer composed of fibrils of β-1,3-linked GalNAc and β-1,4-linked GlcNAc (chitin), respectively. The goal here was to determine whether hand sanitizers that contain ethanol or isopropanol as the active microbicide might reduce transmission of these parasites. We found that treatment with these alcohols with or without drying in a rotary evaporator (to model rapid evaporation of sanitizers on hands) kills 85 to 100% of cysts ofG. duodenalisand 90 to 100% of cysts ofEntamoeba invadens(a nonpathogenic model forE. histolytica), as shown by nuclear labeling with propidium iodide and failure to excystin vitro. Alcohols with or without drying collapsed the cyst walls ofGiardiabut did not collapse the cyst walls ofEntamoeba. To validate thein vitroresults, we showed that treatm...

The Jacob lectin, the most abundant glycoprotein in the cyst wall of Entamoeba invadens , contains five unique 6-Cys chitin-binding domains (CBDs). We identified Entamoeba histolytica and Entamoeba dispar genes encoding Jacob homologues, each of which contains two predicted 6-Cys CBDs. A unique 8-Cys CBD was found at the N termini of the E. histolytica chitinase and three other predicted lectins, called Jessie 1 to Jessie 3. The CBDs of four E. histolytica lectins (Jacob, chitinase, and Jessies 2 and 3) were expressed in secretory vesicles of transfected amebae and shown to bind to particulate chitin.

The infectious stage of amebae is the chitin-walled cyst, which is resistant to stomach acids. In this study an extraordinarily abundant, encystation-specific glycoprotein (Jacob) was identified on two-dimensional protein gels of cyst walls purified from Entamoeba invadens . Jacob, which was acidic and had an apparent molecular mass of ∼100 kDa, contained sugars that bound to concanavalin A and ricin. The jacob gene encoded a 45-kDa protein with a ladder-like series of five Cys-rich domains. These Cys-rich domains were reminiscent of but not homologous to the Cys-rich chitin-binding domains of insect chitinases and peritrophic matrix proteins that surround the food bolus in the insect gut. Jacob bound purified chitin and chitin remaining in sodium dodecyl sulfate-treated cyst walls. Conversely, the E. histolytica plasma membrane Gal/GalNAc lectin bound sugars of intact cyst walls and purified Jacob. In the presence of galactose, E. invadens formed wall-less cysts, which were quadran...

Cysts of Giardia lamblia and Entamoeba histolytica and oocysts of Toxoplasma gondii and Cryptosporidium parvum are the infectious and sometimes diagnostic forms of these parasites. To discover the structural components of cyst and oocyst walls, we have developed strategies based upon a few simple assumptions. Briefly, the most abundant wall proteins are identified by monoclonal antibodies or mass spectrometry. Structural components include a sugar polysaccharide (chitin for Entamoeba , β-1,3-linked glucose for Toxoplasma , and β-1,3-linked GalNAc for Giardia ) and/or acid-fast lipids ( Toxoplasma and Cryptosporidium ). Because Entamoeba cysts and Toxoplasma oocysts are difficult to obtain, studies of walls of nonhuman pathogens ( E. invadens and Eimeria , respectively) accelerate discovery. Biochemical methods to dissect fungal walls work well for cyst and oocyst walls, although the results are often unexpected. For example, echinocandins, which inhibit glucan synthases and kill fun...

Entamoeba histolytica , which causes amebic dysentery and liver abscesses, is spread via chitin-walled cysts. The most abundant protein in the cyst wall of Entamoeba invadens , a model for amebic encystation, is a lectin called EiJacob1. EiJacob1 has five tandemly arrayed, six-Cys chitin-binding domains separated by low-complexity Ser- and Thr-rich spacers. E. histolytica also has numerous predicted Jessie lectins and chitinases, which contain a single, N-terminal eight-Cys chitin-binding domain. We hypothesized that E. invadens cyst walls are composed entirely of proteins with six-Cys or eight-Cys chitin-binding domains and that some of these proteins contain sugars. E. invadens genomic sequences predicted seven Jacob lectins, five Jessie lectins, and three chitinases. Reverse transcription-PCR analysis showed that mRNAs encoding Jacobs, Jessies, and chitinases are increased during E. invadens encystation, while mass spectrometry showed that the cyst wall is composed of an ∼30:70 m...

We sought to establish an ex vivo model for examining the interaction of E. histolytica with human tissue, using precision-cut liver slices (PCLS) from donated organs. E. histolytica- or E. dispar-infected PCLS were analyzed at different post-infection times (0, 1, 3, 24 and 48 h) to evaluate the relation between tissue damage and the expression of genes associated with three factors: a) parasite survival (peroxiredoxin, superoxide dismutase and 70 kDa heat shock protein), b) parasite virulence (EhGal/GalNAc lectin, amoebapore, cysteine proteases and calreticulin), and c) the host inflammatory response (various cytokines). Unlike E. dispar (non-pathogenic), E. histolytica produced some damage to the structure of hepatic parenchyma. Overall, greater expression of virulence genes existed in E. histolytica-infected versus E. dispar-infected tissue. Accordingly, there was an increased expression of EhGal/GalNAc lectin, Ehap-a and Ehcp-5, Ehcp-2, ehcp-1 genes with E. histolytica, and a d...

This study included the investigation of Bifidobacterium spp antagonistic
activity against Staphylococcus aureus isolates, and the effect of these
antimicrobial agent on virulence factors produce by MRSA (Methicillin
Resistance Staphylococcus aureus) such as Biofilm and Slime layer .
One handered different samples were collected to isolate Bifidobacterium spp
and 50 clinical isolates of Staphylococcus aureus from clinical specimens in ALNajaf
AL-Asharaf governorate during the period of (10/10/2013-20/1/2014) by
the cultural characteristic colonies, microscopic for cells, biochemical tests and
polymerase chain reaction PCR technique to detect 16SrDNA, the results
showed that 13 isolates belong to Bifidobacterium spp and there are divergent
gene content between these isolates and they have been bearing lm26/ lm3 gene.
Carbohydrate fermentation test was used to distinguish the species of
Bifidobacterium, the results revealed that the B.bifidum is more predominant
than the other species; B.thermoacidophilum, B.adolescentis, B.breve, B.longum.
Antibiotics susceptibility test to Staphylococcus aureus using antibiotic disc
diffusion assay showed that Methicillin Resistant Staphylococcus aureus formed
70% and 30% of isolates were sensitive to the same antibiotics, also the ability
of MRSA isolates to produce Biofilm was investigated by tube method, then 10
isolates of Staphylococcus aureus were selected as efficient isolates according
to the resistance of Methicillin and the high ability to produce biofilm, and the
ability of bacteriocin production of Bifidobacterium spp against MRSA have
been conducted by well diffusion broth method, B.bifidum3 was greater than
other species in producing bacteriocin, Furthermore the impact of bacteriocin
production to inhibit the development and formation of biofilm was investigated,
the results showed that the bacteriocin has ahigh activity against MRSA.

Bu calismada erkeklerin perine bolgesinde Demodex folliculorum arastirilmasi amaclandi. Calismaya cesitli sikayetlerle uroloji poliklinigine basvuran 122 erkek hasta dahil edildi. Hastalarin perine bolgeleri lokal deri bulgusu acisindan degerlendirildi ve standart yuzeyel deri biyopsisi yontemiyle ornekler alindi. Mikroskobik incelemede cm2’de 5 ve daha fazla Demodex sp. gorulmesi pozitif olarak degerlendirildi. Calismaya dahil edilen hastalarin yas ortalamasi 46.4 ± 18.3 yil ( 18-83 yil) olarak belirlendi. Hastalarin 15’inde (%12.3) D. folliculorum pozitif olarak saptandi. D. folliculorum pozitif bulunan hastalarin yas ortalamasi 61.93 ± 17.72 yil, negatif olgularin ise 44.26 ± 17.40 yil olarak bulundu (p= 0.002). D. folliculorum pozitifligine en sik benign prostat hipertrofisi olan hastalarda rastlandi. Uroloji hastalarinda perine bolgesinde D. folliculorum akarlari gorulebilmekle birlikte etkenin varligi ozellikle cesitli klinik bulgulari olan hastalarin yonetiminde goz onunde bu...

Background and objective: Intestinal and extraintestinal amebiasis is caused by the protozoan parasite Entamoeba (E.) histolytica, it is of considerable morbidity and mortality in developing countries. E.histoltica complex species includes E.histolytica, E. moshkovskii and E. dispar are morphologically indistinctable. The current study goal was to use molecular assays to detect the true prevalence of E. histolytica  complex species among a cohort of Egyptians. Methods:  A single stool specimen was collected from 133 patients, examined coproscopically before and after concentration. DNA was extracted from microscopically positive stool specimens for E. histolytica complex species were molecularly idenitified using multiplex PCR. Results: The  coroscopic prevalence of intestinal parasites was 51.1% (68/133) of them 30 cases had E.histolytica complex (22.6%; 30/133). Among coproscopically positive samples, E. dispar was the most common parasite (63.3%;  19/30),  followed by  E.  histol...

Entamoeba histolytica, an intestinal parasite that is the causative agent of amoebiasis, is exposed to elevated amounts of highly toxic reactive oxygen and nitrogen species during tissue invasion. A flavodiiron protein and a rubrerythrin have been characterized in this human pathogen, although their physiological reductants have not been identified. The present work deals with biochemical studies performed to reach a better understanding of the kinetic and structural properties of rubredoxin reductase and two ferredoxins from E. histolytica. We complemented the characterization of two different metabolic pathways for O2 and H2O2 detoxification in E. histolytica. We characterized a novel amoebic protein with rubredoxin reductase activity that is able to catalyze the NAD(P)H-dependent reduction of heterologous rubredoxins, amoebic rubrerythrin and flavodiiron protein but not ferredoxins. In addition, the protein exhibited NAD(P)H oxidase activity, which generates hydrogen peroxide fro...

Diarrhoea associated illness presents with mortality and morbidity in rural communities in most low income countries especially in children < 5 years of age. The continuous emergence of several opportunistic infections in immuno-compromised individuals has worsened the burden of diarrhoea in most of these countries. Protozoan infections caused by species of Cryptosporidium, Entamoeba spp. Giardia intestinalis, Blastocystis hominis and Trichomonas vaginalis have received insufficient attention because data on their prevalence and incidence are scanty. The commonly used drugs to treat infections caused by these organisms are becoming less effective due to the development of drug resistance. Evidence from literature has shown that natural products from medicinal plants are likely to be suitable alternatives and complimentary therapeutic drugs to combat most protozoan infections. Natural products and their bioactive compounds could be the solution to treat most protozoan infections t...

There is a clear need to perform epidemiological studies to find the true prevalence of Entamoeba histolytica around the world. The evaluation of this prevalence has been hindered by the existence of two different species which are morphologically identical, but genetically different, namely E. histolytica, which causes amebiasis, and E. dispar, which is non-pathogenic. In Brazil, the E. dispar has been detected in communities in the Southeastern (SE) and Northeastern (NE) regions with poor sanitation. However, individuals infected with E. histolytica have been identified in other regions. There is an absence of reports on the prevalence of these parasites in the state of Paraíba, which also has areas with poor sanitary conditions where a high prevalence of the E. histolytica/E. dispar complex has been detected in children from urban slums. The present study evaluated the prevalence of E. histolytica and E. dispar in 1,195 asymptomatic children between two and 10 years of age, livin...

Background: Entamoeba gingivalis was the first commensal parasite detected in the oral cavity of humans, and a high incidence has been reported in patients with poor oral hygiene. The current study aimed to investigate the association of Entamoeba gingivalis with gingivitis and periodontitis among Egyptian subjects. Methods: A total of 120 plaque samples were collected for this case-control study and were divided as follows: 40 plaque samples from gingivitis patients (group 1), 40 from stage II grade A and B periodontitis patients (group 2), and 40 samples from healthy volunteers (group 3). Diagnosis of parasitic stages relied on direct microscopic detection using permanent stains, trichrome stain, and hematoxylin and eosin (H&E) stain, in addition to ocular micrometry to confirm the diagnosis. Results: The occurrence of Entamoeba gingivalis within the gingivitis group was significantly higher (40%) than that observed in the control group (22.5%), whereas the occurrence within the p...

Infections with Giardia lamblia , Entamoeba histolytica , and Trichomonas vaginalis , which cause diarrhea, dysentery, and vaginitis, respectively, are each treated with metronidazole. Here we show that Giardia , Entamoeba , and Trichomonas have oxygen-insensitive nitroreductase ( ntr ) genes which are homologous to those genes that have nonsense mutations in metronidazole-resistant Helicobacter pylori isolates. Entamoeba and Trichomonas also have nim genes which are homologous to those genes expressed in metronidazole-resistant Bacteroides fragilis isolates. Recombinant Giardia , Entamoeba , and Trichomonas nitroreductases used NADH rather than the NADPH used by Helicobacter , and two recombinant Entamoeba nitroreductases increased the metronidazole sensitivity of transformed Escherichia coli strains. Conversely, the recombinant nitroimidazole reductases (NIMs) of Entamoeba and Trichmonas conferred very strong metronidazole resistance to transformed bacteria. The Eh ntr1 gene of th...

A diagnostic test for intestinal amebiasis was developed by using an ELISA that incorporated a well‐defined monoclonal antibody specific for a membrane antigen of Entamoeba histolytica. The ELISA was found to be sensitive and quite specific for E. histolytica, since there were no cross reactions with other parasites, especially Entamoeba coli, with which Entamoeba histolytica is often confused. In addition, this test detected not only E. histolytica trophozoites but also cysts. This technique will be of great value in the rapid and accurate diagnosis of E. histolytica in human fecal material.

The aim of this study was to determine if the protozoa that cause dysentery might have been present in Jerusalem, the capital of the Kingdom of Judah, during the Iron Age. Sediments from 2 latrines pertaining to this time period were obtained, 1 dating from the 7th century BCE and another from the 7th to early 6th century BCE. Microscopic investigations have previously shown that the users were infected by whipworm (Trichuris trichiura), roundworm (Ascaris lumbricoides), Taenia sp. tapeworm and pinworm (Enterobius vermicularis). However, the protozoa that cause dysentery are fragile and do not survive well in ancient samples in a form recognizable using light microscopy. Enzyme-linked immunosorbent assay kits designed to detect the antigens of Entamoeba histolytica, Cryptosporidium sp. and Giardia duodenalis were used. Results for Entamoeba and Cryptosporidium were negative, while Giardia was positive for both latrine sediments when the analysis was repeated three times. This provid...

The protozoan parasite Entamoeba histolytica colonizes the human large bowel. Invasion of the intestinal epithelium causes amoebic colitis and opens the route for amoebic liver abscesses. The parasite relies on its dynamic actomyosin cytoskeleton and on surface adhesion molecules for dissemination in the human tissues. Here we show that the galactose/ N -acetylgalactosamine (Gal/GalNAc) lectin clusters in focal structures localized in the region of E. histolytica that contacts monolayers of enterocytes. Disruption of myosin II activity impairs the formation of these structures and renders the trophozoites avirulent for liver abscess development. Production of the cytoplasmic domain of the E. histolytica Gal/GalNAc lectin in engineered trophozoites causes reduced adhesion to enterocytes. Intraportal delivery of these parasites to the liver leads to the formation of a large number of small abscesses with disorganized morphology that are localized in the vicinity of blood vessels. The ...

Entamoeba histolytica causes amebic colitis and liver abscess in developing countries such as Mexico and India. Entamoeba dispar is morphologically identical but is not associated with disease. Here we determined the ploidy of E. histolytica and developed PCR-based methods for distinguishing field isolates of E. histolytica or E. dispar . Fluorescence in situ hybridization showed that E. histolytica trophozoites are diploid for five “single-copy” probes tested. Intergenic sequences between superoxide dismutase and actin 3 genes of clinical isolates of E. histolytica from the New and Old Worlds were identical, as were those of E. dispar . These results suggest a bottleneck or demographic sweep in entamoebae which infect humans. In contrast, E. histolytica and E. dispar genes encoding repeat antigens on the surface of trophozoites (Ser-rich protein) or encysting parasites (chitinase) were highly polymorphic. chitinase alleles suggested that the early axenized strains of E. histolytica...

As an alternative to microscopic identification of Entamoeba histolytica parasites isolated from stool, a sensitive and species-specific DNA hybridization probe was made for rapid diagnosis of E. histolytica parasites in clinical samples directly applied to nylon membranes. The DNA hybridization probe was made by screening a genomic library of a virulent HM-1:IMSS strain of E. histolytica to detect recombinant plasmids containing highly repeated parasite DNA sequences. Four plasmid clones that reacted across Entamoeba species coded for highly repeated rRNA genes of E. histolytica. Four other plasmid clones were E. histolytica specific in that they bound to four axenized and nine xenic strains of E. histolytica but did not recognize closely related E. histolytica-like Laredo, Entamoeba moshkovskii, or Entamoeba invadens parasites. The diagnostic clones detected as few as eight cultured amoebae and did not distinguish between pathogenic and nonpathogenic zymodemes of E. histolytica. T...

A diagnostic test for intestinal amebiasis was developed by using an ELISA that incorporated a well‐defined monoclonal antibody specific for a membrane antigen of Entamoeba histolytica. The ELISA was found to be sensitive and quite specific for E. histolytica, since there were no cross reactions with other parasites, especially Entamoeba coli, with which Entamoeba histolytica is often confused. In addition, this test detected not only E. histolytica trophozoites but also cysts. This technique will be of great value in the rapid and accurate diagnosis of E. histolytica in human fecal material.

Entamoeba histolytica, an intestinal parasite that is the causative agent of amoebiasis, is exposed to elevated amounts of highly toxic reactive oxygen and nitrogen species during tissue invasion. A flavodiiron protein and a rubrerythrin have been characterized in this human pathogen, although their physiological reductants have not been identified. The present work deals with biochemical studies performed to reach a better understanding of the kinetic and structural properties of rubredoxin reductase and two ferredoxins from E. histolytica. We complemented the characterization of two different metabolic pathways for O2 and H2O2 detoxification in E. histolytica. We characterized a novel amoebic protein with rubredoxin reductase activity that is able to catalyze the NAD(P)H-dependent reduction of heterologous rubredoxins, amoebic rubrerythrin and flavodiiron protein but not ferredoxins. In addition, the protein exhibited NAD(P)H oxidase activity, which generates hydrogen peroxide fro...

Background Surveys for intestinal parasitic infections (IPIs) often involve samples from three sampling dates analysed by various microscopy techniques. However, analysis of three samples per individual is a huge burden on time and resources. We compared the value from analysing three or fewer samples. Methods In this cross-sectional study, three faecal samples were collected every other day from 332 children from two locations in Guantanamo province, Cuba. Samples were analysed by wet mount with Lugol stain, Willis flotation method and Kato–Katz thick smear. Results Most parasites were detected by wet mount, although helminth eggs not found by wet smear were detected by the Willis flotation method (in particular) and Kato–Katz smear. Blastocystis spp. was the most commonly detected parasite (about 65%), then Giardia duodenalis and then Entamoeba spp. Although analysis of two stool samples significantly increased occurrence data for Blastocystis, this was not so for the other parasi...

In developing countries, the prevalence of intestinal parasitic infection is still significant, particularly due to geographical and socioeconomic variables. The objective of this study was to map the distribution pattern of intestinal parasitic infection in a cohort of the Egyptian population, as well as to assess associated risk factors. A cross-sectional hospital-based study was conducted on 386 patients. A single fecal specimen was collected from the study individual and examined microscopically for the detection of parasitic infection. DNA was extracted from all samples and utilized to amplify Entamoeba histolytica complex species, Cryptosporidium species, Giardia intestinalis assemblages, and Blastocystis species using PCRs. Typing of Cryptosporidium species and Giardia intestinalis assemblages was performed using restriction enzymes RasI and HaeIII respectively. While Blastocystis spp. subtypes (ST) were identified through sequencing of PCR products and phylogenetic analysis....