A novel transcribed repeat element from Entamoeba histolytica.

We have identified an unusual 0.55-kb DNA repeat element specific to Entamoeba histolytica (Eh) which we call interspersed element (IE). The IE is a common feature in independently isolated genomic and cDNA fragments. Hybridization of labeled IE sequences to trophozoite DNA, RNA and first-strand cDNA prepared from poly(A)-enriched mRNA indicate that the IE are reiterated about 500 times per Eh trophozoite and that one or more can be found as RNA transcripts. These features and the degree of conservation of IE suggest a possible role for these sequences.

Influence of bacteria on electrophoretic proteinase patterns of Entamoeba histolytica isolates.

Gelatin SDS- polyacrylamide gel electrophoresis was used to compare proteinase banding patterns under reducing conditions from whole cell lysates of four axenic and four xenic pathogenic strains of Entamoeba histolytica. All strains shared major bands in the 34 and 66-68 kDa regions, whereas only the axenic strains produced major bands at 26, 28, 30 and 45 kDa. One axenic strain, NIH 200, when reassociated with mixed bacterial flora, reverted to an electrophoretic banding pattern characteristic of other xenic strains. These results suggest that the 26-30 kDa and 45 kDa proteinases of E. histolytica are induced by bacterial starvation while others are constitutively expressed. It is also proposed that the axenic bands of 45 and 34 kDa represent respectively, the reduced forms of the 56 and 40 kDa bands reported elsewhere under non-reducing conditions.

The effect of axenic versus xenic culture conditions on the total and secreted proteolytic activity of Entamoeba histolytica strains.

Proteolytic activity was measured in lysates of four axenic and five xenic cultures of different strains of pathogenic E. histolytica, and in five non-pathogenic strains ("E. dispar") growing in xenic culture. There was no significant difference in total proteolytic activity, using either gelatin or albumin as substrate, between the two sets of xenic cultures; the axenic pathogens however had significantly higher levels of activity than the xenic pathogens, the levels appearing to correlate with virulence. An axenic strain (NIH 200) reassociated with mixed bacterial flora reverted to close to xenic levels of activity. There was no evidence of secretion of an elevated proportion of the total enzymic activity by pathogenic strains, and expression levels may owe more to culture conditions than to genotype.

The amoeba enigma.

After more than 70 years of intermittent debate over the true relationship between the 'pathogenic' and 'non-pathogenic' forms of Entamoeba histolytica, the application of molecular biology has finally yielded an unambiguous answer: these are not interconvertible phenotypes of the same parasite, a kind of unicellular Jekyll and Hyde, but two quite distinct genetic entities that just happen to look the same. But given the overwhelming evidence now available from gene sequences, pointing to an evolutionary divergence some tens of millions of years ago, why is it that certain eminent workers in the field are still claiming that, at least in vitro, conversion between the two phenotypes can take place? In this article Bill Spice and John Ackers review recent developments in the molecular biology of E. histolytica and assess the continuing controversy over the status of this enigmatic parasite.

The effects of Entamoeba histolytica lysates on human colonic mucins.

Detergent lysates of Entamoeba histolytica trophozoites contained high levels of beta-N acetyl-D-glucosaminidase, beta-N acetyl-D-galactosaminidase and alpha-D-galactosidase activity, and lower but significant levels of five other glycosidases. Although these activities should have been capable of largely degrading the oligosaccharide side-chains of human colonic mucin, in fact only about one third of high MW mucin was degraded in 72 h and trypsin alone produced a similar effect. There was no evidence that these glycosidases were excreted and we conclude that they are unlikely to represent significant virulence factors for E. histolytica.

Ribosomal DNA sequences in the differentiation of pathogenic and non-pathogenic isolates of Entamoeba histolytica.

Recombinant ribosomal DNA sequences were amplified by PCR and used as probes to perform a fingerprint analysis of total DNA from different Entamoeba histolytica isolates. RFLPs obtained with one of the probes, R-1, support previous proposals that pathogenic and non-pathogenic E. histolytica are closely related, yet genotypically distinct. Another probe, R-2, while not distinguishing between the two forms of E. hystolytica, was able to differentiate between them and E. moshkovskii, which has morphologically identical cysts and trophozoites. A third probe, BR-1, identified strain-specific RFLPs.

Immunological differentiation of pathogenic and non-pathogenic isolates of Entamoeba histolytica.

Entamoeba histolytica can act as a harmless commensal organism in the lumen of the large intestine, or can cause invasive amoebiasis. Some workers have suggested that there are two distinct subspecies of this organism, and that only one of these is associated with invasive disease. Present isoenzyme tests to identify the subspecies take several days to analyse: we report a technique that uses immunofluorescence with monoclonal antibodies, takes two days to perform, and may, therefore, assist in the clinical management of patients infected with this organism.