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At least 19 recordsLinked to original sources
PubMed2000

Inter- and intra-strain variation in the 5.8S ribosomal RNA and internal transcribed spacer sequences of Entamoeba histolytica and comparison with Entamoeba dispar, Entamoeba moshkovskii and Entamoeba invadens.

The ribosomal RNA genes in Entamoeba histolytica are located on circular DNA molecules in about 200 copies per genome equivalent. Nucleotide sequence analysis of the 5.8S rRNA gene and the flanking internal transcribed spacers was carried out to determine the degree of sequence divergence in the multiple rRNA gene copies of a given strain; amongst three different E. histolytica strains (HM-1:IMSS, Rahman and HK-9); and amongst four species of Entamoeba (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii and Entamoeba invadens). The results show that all rRNA gene copies of a given strain are identical. Few nucleotide positions varied between strains of a species but the differences were very pronounced amongst species. In general, the internal transcribed spacer 2 sequence was more variable and may be useful for strain- and species-identification. The 5.8S rRNA gene and the internal transcribed spacer 2 of E. invadens were unusually small in size.

Animals↗
PubMed2006

Differential detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii by a single-round PCR assay.

A single-round PCR assay was developed for detection and differential diagnosis of the three Entamoeba species found in humans, Entamoeba moshkovskii, Entamoeba histolytica, and Entamoeba dispar, that are morphologically identical as both cysts and trophozoites. A conserved forward primer was derived from the middle of the small-subunit rRNA gene, and reverse primers were designed from signature sequences specific to each of these three Entamoeba species. PCR generates a 166-bp product with E. histolytica DNA, a 752-bp product with E. dispar DNA, and a 580-bp product with E. moshkovskii DNA. Thirty clinical specimens were examined, and the species present were successfully detected and differentiated using this assay. It was possible to detect as little as 10 pg of E. moshkovskii and E. histolytica DNA, while for E. dispar the sensitivity was about 20 pg of DNA. Testing with DNA from different pathogens, including bacteria and other protozoa, confirmed the high specificity of the assay. We propose the use of this PCR assay as an accurate, rapid, and effective diagnostic method for the detection and discrimination of these three morphologically indistinguishable Entamoeba species in both routine diagnosis of amoebiasis and epidemiological surveys.

Animals↗
PubMedDate not supplied

Lipophosphoglycan is present in distinctly different form in different Entamoeba histolytica strains and absent in Entamoeba moshkovskii and Entamoeba invadens.

Lipophosphoglycan has recently been demonstrated on the cell surface of Entamoeba histolytica strain HM-1:IMSS. A monoclonal antibody against this molecule had failed to react with some other strains of E. histolytica, including the strain Rahman. To determine if a structurally distinct lipophosphoglycan existed in Rahman, [3H]galactose-labeled glycoconjugates were electrophoresed through sodium dodecyl sulfate polyacrylamide gel electrophoresis. The electrophoretic pattern in Rahman was very different compared to that obtained with strains HM-1:IMSS and 200:NIH. A number of experiments including sensitivity to mild acid, nitrous acid and phosphoinositol-specific phospholipase C suggest that the Rahman glycoconjugate is indeed a lipophosphoglycan-like molecule but distinctly different from that of HM-1:IMSS. Mild acid-treated glycoconjugates from Rahman and HM-1:IMSS revealed the presence of neutral trisaccharides and monosaccharides in Rahman but not in HM-1:IMSS. Human immune sera from amoebiasis patients and a polyclonal antibody against HM-1:IMSS lipophosphoglycan both recognized Rahman glycoconjugate. Thus, while lipophosphoglycan molecules from the two strains share common epitopes, they are clearly distinct from each other. Molecules bearing resemblance to lipophosphoglycan could not be detected in other Entamoeba species, namely Entamoeba invadens and Entamoeba moshkovskii.

Animals↗
PubMed2007

PCR detection of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in stool samples from Sydney, Australia.

This study investigated the presence of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in stool samples from a patient population in Sydney, Australia. Stool samples were tested by microscopy and PCR. Five patients were found with E. histolytica infections, while E. dispar and E. moshkovskii were observed in 63 (70.8%) and 55 (61.8%) patients, respectively, by PCR. This is the first study in Australia using molecular techniques to determine the presence of E. histolytica, E. dispar, and E. moshkovskii.

Adult↗
PubMed1995

Rapid diagnosis of Entamoeba infection by using Entamoeba and Entamoeba histolytica stool antigen detection kits.

Humans are infected by two morphologically identical species of Entamoeba: Entamoeba histolytica causes amebic colitis and liver abscess, and Entamoeba dispar is noninvasive. Several weeks of culture and isoenzyme (zymodeme) analysis are required to differentiate E. histolytica from E. dispar. Here we report a field trial of commercial antigen detection kits designed to rapidly detect and differentiate E. histolytica from E. dispar in stool specimens. Stool specimens from 202 patients with diarrhea were examined for E. histolytica and E. dispar by microscopy, culture, and antigen detection. Compared with culture, microscopic identification of the E. histolytica-E. dispar complex was 60% sensitive and 79% specific, while the screening antigen detection test for the E. histolytica-E. dispar complex was 80% sensitive and 99% specific. Differentiation of E. dispar from E. histolytica by the E. histolytica-specific test was 95% sensitive and 93% specific compared with zymodeme analysis. We conclude that the antigen detection test for the E. histolytica-E. dispar complex is more sensitive and specific than microscopy and that the E. histolytica-specific antigen detection test is as reliable and much more rapid than zymodeme analysis for the differentiation of E. histolytica from E. dispar.

Animals↗
PubMed1999

Characterization of the gene encoding serine acetyltransferase, a regulated enzyme of cysteine biosynthesis from the protist parasites Entamoeba histolytica and Entamoeba dispar. Regulation and possible function of the cysteine biosynthetic pathway in Entamoeba.

The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites.

Acetyltransferases↗
PubMed1980

[Staining of cysts of Entamoeba invadens, Entamoeba histolytica and Entamoeba coli with wheat germ agglutinin labelled with colloidal gold].

Cysts of Entamoeba invadens obtained under axenic conditions were stained with wheat germ agglutinin labelled with colloidal gold. Microscopic observation of encysting cultures revealed that upon staining, cysts acquired a red coloration, while trophozoites remained unstained. E. histolytica and E. coli cysts obtained from asymptomatic patients were also stained red by this technique. Electronmicroscopic examination of stained cells showed a layer of colloidal gold granules attached to the surface of cysts, while trophozoites remained free of gold granules. The results of the staining procedure developed, agreed with the known facts that wheat germ agglutinin is a lectin that binds specifically to chitin and its oligomers, and that chitin is present in the cyst wall of E. invadens. The results suggested that chitin is also present in the wall of cysts of E. histolytica and E. coli.

Entamoeba↗
PubMedDate not supplied

Axenic cultivation of Entamoeba dispar Brumpt 1925, Entamoeba insolita Geiman and Wichterman 1937 and Entamoeba ranarum Grassi 1879.

Three species of Entamoeba have been grown in axenic culture for the first time. In two cases, novel methods for adapting the organisms to growth without bacteria were employed. While E. ranarum was axenized by the classic technique of Diamond, from a monoxenic culture with Trypanosoma cruzi as the associate, both E. dispar and E. insolita were first grown in axenic culture medium supplemented with lethally irradiated bacteria. From there, E. insolita was axenized directly, but E. dispar initially required the presence of fixed bacteria. After prolonged culture under this technically axenic but unwieldy culture system, E. dispar was eventually adapted to growth in the absence of added bacteria.

Animals↗
PubMed1997

Molecular biology of the hexokinase isoenzyme pattern that distinguishes pathogenic Entamoeba histolytica from nonpathogenic Entamoeba dispar.

The electrophoretic patterns of hexokinase and phosphoglucomutase have been widely used to distinguish Entamoeba histolytica from Entamoeba dispar isolates. Although E. histolytica and E. dispar, previously called pathogenic and nonpathogenic Entamoeba histolytica, differ clearly in sequences of many homologous genes, a conversion between the two has been reported by several laboratories, in each case showing the conversion of hexokinase (ATP, D-hexose 6-phosphotransferase, EC 2.7.1.1) isoenzyme patterns. An apparent mobility shift of this enzyme may either be due to posttranslational modification or processing, or to the appearance of a new isoform encoded by a second gene. In this study we observed that the four observed bands in the isoenzyme patterns of pathogenic and nonpathogenic forms of Entamoeba were correlated with four different cDNAs, and that the four recombinant hexokinases produced in Escherichia coli comigrated with their natural counterparts. Polymerase chain reaction (PCR) experiments did not reveal hidden genes which might be responsible for conversion phenomena. These results strongly support the redefinition of pathogenic and nonpathogenic Entamoeba histolytica as two closely related species Entamoeba histolytica and Entamoeba dispar.

Amino Acid Sequence↗
PubMed2000

Differentiation of Entamoeba histolytica and Entamoeba dispar cysts using polymerase chain reaction on DNA isolated from faeces with spin columns.

Since Entamoeba histolytica and Entamoeba dispar were formally recognized as two different species at the World Health Organization (WHO)/Pan American Health Organization (PAHO)/United Nations Educational, Scientific and Cultural Organization (UNESCO) meeting in Mexico City in 1997, the specific differentiation of the two morphologically identical species would seem relevant in clinical diagnosis. Several polymerase chain reaction (PCR)-based methods have been described and used successfully, but methods for DNA isolation from cysts in stool samples are time-consuming and problematic due to inhibitory factors in faeces. The use of the slightly modified QIAamp tissue method (Qiagen, Germany) for DNA isolation was evaluated in 657 unpreserved faecal samples from cases of suspected Entamoeba histolytica/Entamoeba dispar infection. In only 1.7% of the cases was PCR hampered by inhibitors present in the faeces. The DNA isolation procedure was found to be rapid, simple and one that could easily be implemented in a routine diagnostic setting. In 98.8% of Entamoeba histolytica/Entamoeba dispar cyst-positive faecal samples, the true identity of the cysts could be determined using PCR specific for Entamoeba histolytica and Entamoeba dispar, respectively.

Animals↗
PubMed2002

Real-time PCR for detection and differentiation of Entamoeba histolytica and Entamoeba dispar in fecal samples.

A closed-tube, real-time PCR assay was developed for sensitive and specific detection and differentiation of the two closely related intestinal protozoan parasites Entamoeba histolytica and Entamoeba dispar directly from human feces. The assay is performed with the LightCycler system using fluorescence-labeled detection probes and primers amplifying a 310-bp fragment from the high-copy-number, ribosomal DNA-containing ameba episome. The assay was able to detect as little as 0.1 parasite per g of feces. The two pairs of primers used were specific for the respective ameba species, and results were not influenced by the presence of other Entamoeba species even when present in exceeding amounts. PCR was evaluated using several hundred stool samples from areas of amebiasis endemicity in Vietnam and South Africa, and results were compared with those of microscopy and ameba culture. PCR was found to be significantly more sensitive than microscopy or culture, as all samples positive by microscopy and 22 out of 25 (88%) samples positive by culture were also positive by PCR, but PCR revealed a considerable number of additional E. histolytica- or E. dispar-positive samples. Compared to culture and subsequent ameba differentiation by isoenzyme analysis, PCR was 100% specific for each of the two Entamoeba species. Interestingly, the comparison with PCR revealed that culture, in particular, underestimates E. histolytica infections. Given the high sensitivity and specificity of the developed PCR assay, the inability of microscopy to distinguish between the two ameba species, and the time it takes to culture and subsequently differentiate entamoebae by isoenzyme analysis, this assay is more suitable than microscopy or culture to correctly diagnose intestinal E. histolytica or E. dispar infection.

Animals↗
PubMed2026

Comparative proteomics reveals distinct functions and localization of invasive Entamoeba histolytica and non-invasive Entamoeba moshkovskii proteins.

BACKGROUND: Entamoeba histolytica is a pathogenic protozoan accountable for amoebiasis, while Entamoeba moshkovskii is considered non-invasive. Despite morphological similarity, the molecular mechanisms underlying their different pathogenicity remain largely undefined. METHODS: Trophozoite proteins from axenic cultures of E. histolytica and E. moshkovskii were separated and identified using GeLC-MS/MS, and classified using Gene Ontology. Selected and differentially expressed proteins were validated by peptide-specific antibody production, ELISA, and immunofluorescence to determine cellular localization. RESULTS AND DISCUSSION: A total of 1,077 and 1,201 proteins were identified from E. histolytica and E. moshkovskii, respectively. The 801 of Entamoeba common proteins included kinases, GTPase-activating proteins, and heat shock proteins, reflecting conserved cellular processes. E. histolytica-unique proteins involved in nitrogen compound metabolism, vesicle-mediated transport, and catalytic activities, whereas E. moshkovskii proteins were related to lipid metabolism and environmental resilience. Subcellular localization revealed species-specific distribution of MmpL and AIG1-family proteins, suggesting potential roles in pathogenicity and host-immune response. A large proportion of hypothetical proteins was identified, highlighting gaps and opportunities for future study. CONCLUSIONS: Our study highlights conserved and divergent functions and cellular locations of Entamoeba species-specific proteins as insights for distinct pathogenicity and adaptation. MmpL and AIG1 proteins were proposed as potential targets for further diagnostic and therapeutic development.

Proteomics↗
PubMed1988

Comparison of repeated DNA from strains of Entamoeba histolytica and other Entamoeba.

Restriction enzyme digestion patterns of total genomic DNA revealed the presence of highly repeated DNA in Entamoeba histolytica and E. histolytica-like (Laredo type) amoebae of humans, E. invadens and E. invadens and E. terrapinae of reptiles, and E. moshkovskii isolated from sewage polluted waters. Homology with Plasmodium berghei ribosomal DNA was striking. Northern blot analysis of E. histolytica RNA provided further evidence that the highly repeated DNA codes for ribosomal RNA. The distribution of restriction enzyme sites on repeated DNA of the Entamoeba was highly polymorphic. Thus it was possible, based on EcoRI digestion patterns, to distinguish between strains of E. histolytica and the other species of Entamoeba. Additionally, these investigations at the molecular level corroborate previously reported physiological and biochemical evidence indicating the E. histolytica-like amoebae and E. histolytica are not conspecific. EcoRI fragments from repeated DNA of E. histolytica (strain HM-1: IMSS) were cloned in the plasmid vector pTZ18R. A subfragment from one of these clones proved to be a useful hybridization probe in distinguishing E. histolytica from the other Entamoeba.

Animals↗
PubMed2005

Species- and strain-specific probes derived from repetitive DNA for distinguishing Entamoeba histolytica and Entamoeba dispar.

Entamoeba histolytica and Entamoeba dispar are two morphologically indistinguishable species that are found in the human gut. Of the two, E. histolytica is considered to be pathogenic while E. dispar is nonpathogenic. To generate molecular probes to detect and distinguish between the two species, we utilized repeat sequences present in Entamoeba genome. We have developed probes and primers from rDNA episomes, and unidentified Entamoeba EST1 repeat for this purpose, and used them for dot blot hybridization and PCR amplification. To investigate the possible existence of invasive and noninvasive strains of E. histolytica, the ability to differentiate individual isolates is necessary. For this purpose, we have utilized a modification of the AFLP procedure called 'Transposon display,' which generates and displays large number of genomic bands associated with a transposon. We have used the abundant retrotransposon, EhSINE1, for this purpose,and demonstrated its potential as a marker to study strain variation in E. histolytica. This technique could suitably be employed in carrying out significant molecular epidemiological studies and large-scale typing of this parasite.

Animals↗
PubMed1978

Clonal growth of Entamoeba histolytica and other species of Entamoeba in agar.

This is the first description of a method for growing axenized Entamoeba histolytica as colonies from single cells in agar suspension. Factors affecting the efficiency of colony formation included unknown characteristics of the amebal strain, age of cells used, and the type and concentration of agar employed. Colony formation was dependent upon filling deep vessels (culture tubes or tissue culture flasks) with agar, probably to maintain reduced oxygen tension, and upon solidifying the agar rapidly at 0 C. It was demonstrated in a study with the drug metronidazole that the agar colony method was useful for quantifying cell viability in drug testing. Eleven of 12 E. histolytica strains tested, as well as one Entamoeba terrapinae, one Entamoeba barreti, and 3 Entamoeba invadens strains formed colonies in agar suspension. E. histolytica-like amebae (Laredo type) and E. moshkovskii formed only tiny colonies in agar.

Agar↗