Sucker-like structures on the pathogenic amoeba Naegleria fowleri.

Using scanning electron microscopy, we observed sucker-like structures on amoebae of 13 human isolates of Naegleria fowleri. The number of suckers per amoeba seemed to vary according to the virulence of the strain. We propose the term amoebastome to describe this unique sucker-like structure of N. fowleri.     

neo-inositol polyphosphates in the amoeba Entamoeba histolytica

We have reexamined the structure of inositol phosphates present in trophozoites of the parasitic amoeba Entamoeba histolytica and show here that, rather than being myo-inositol derivatives (Martin, J.-B., Bakker-Grunwald, T., and Klein, G. (1993) Eur. J. Biochem. 214, 711-718), these compounds belong to a new class of inositol phosphates in which the cyclitol isomer is neo-inositol. The structures of neo-inositol hexakisphosphate, 2-diphospho-neo-inositol pentakisphosphate, and 2, 5-bisdiphospho-neo-inositol tetrakisphosphate, which are present in E. histolytica at concentrations of 0.08-0.36 mM, were solved by two-dimensional (31)P-(1)H NMR spectroscopy. No evidence for the co-existence of their myo-inositol counterparts has been found. These neo-inositol compounds were not substrates of 6-diphospho-inositol pentakisphosphate 5-kinase, an enzyme purified from Dictyostelium discoideum that phosphorylates 6-diphospho-myo-inositol pentakisphosphate and more slowly also myo-inositol hexakisphosphate, specifically on position 5. Because preliminary data indicate that large amounts of the same neo-inositol phosphate and diphosphate esters are also present in another primitive amoeba, Phreatamoeba balamuthi, the occurrence of high concentrations of neo-inositol polyphosphates may be much more general than previously thought.     

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.     

Characterization of the genome of the small free-living amoeba Acanthamoeba castellanii.

The cellular DNAs of Acanthamoeba castellanii have been characterized by their behaviour in CsC1 density gradients, by their thermal denaturation and by their renaturation kinetics. Whole-cell DNA exhibits, on CsC1 density gradients, a major peak with a density of 1.717 g/cm3 (major component) and a minor peak with a density of 1.692 g/cm3 (minor component). The major component is nuclear and the minor component is of cytoplasmic origin. The latter contains mitochondrial DNA as well as an extramitochondrial DNA fraction. Reiterated sequences make up approximately 14% of the total and are mainly cytoplasmic. They are characterized by three families of nucleotide sequences. The mitochondrial DNA exhibits a complex renaturation pattern. The fast renaturing component has a calculated complexity of 4.107 daltons. The slower renaturing component has a kinetic complexity tentatively estimated as 1.1010 daltons. The melting profile of mitochondrial DNA suggests heterogeneity in base composition.     

A phylogenetic study of the free-living amoeba

Immunological distances were determined for four strains of the free-living amoeba classified as Amoeba proteus, two strains classified as Polychaos dubia and a single strain classified as Chaos carolinensis. The data show that the _ShP strain does not belong to the proteus group; that A. proteus is more closely related to C. carolinensis and is derived from Chaos as is the _ShP strain; that P. dubia and C. carolinensis are the more distantly related species and appear to be the first of the above to have diverged from a common ancestor; and that the amoebae have a long evolutionary history. The accuracy of the phylogenetic tree and the distance Wagner network was discussed. Since the amoebae may be polyphyletic in origin, the latter was assumed to be a more accurate representation of the immunological distance data.     

Effect of near ultraviolet and visible light on amoeba.

The near ultraviolet and visible light (VL) impinging at an intensity of 2-5 x 10(2) J s-1 m-2 for 2-5 h kills the mitotic and the early S-phase (0- to 15-min-old) amoebae. At the mid- and late S-period only a fraction of cells are killed by VL and G2 phase cells are quite resistant. Amoebae of all cell cycle stages show a delay in the first mitotic division. DNA synthesis, as measured by [3H]thymidine incorporation, is depressed in the VL-exposed early-S amoebae. A concurrent but temporary inhibition in [3H]leucine incorporation also occurs in these cells. However, no significant change in [3H]uridine incorporation has been found. To localize the site of lethal damage, nuclear transplantation studies were undertaken between the control amoebae and the amoebae treated with VL. The nucleus of a VL-exposed early S-phase cell recovers when transplanted immediately after VL exposure into an enucleate G2 cytoplasm but dies if grafted into an enucleat S-phase cytoplasm. The therapeutic effect of the G2 cytoplasm, although at a lower level, is also evident even when the treated early S-phase nucleus is implanted 20 h later, but not after 48 h, into the G2 cytoplasm. The amoeba cytoplasm shows resistance to VL-irradiation, can accept a control nucleus from any cell cycle stage, and function normally. The G2 nucleus also remains apparently unaffected to VL exposure and can survive when it is transfered to the control cytoplasm of any cell-cycle phase. All these findings are discussed in the light of the possible existence of a repair system against VL-induced damage in the G2-phase amoeba.     

A study of histones in amoeba nuclei by indirect immunofluorescence method.

Nuclear proteins of four species of free-living Amoebidae (Amoeba proteus, A. discoides, Chaos carolinensis and Polychaos dubia) have been studied by indirect immunofluorescence technique using specific antisera to H1, H2A, H2B, H3 and H4 histone fractions from the calf thymus. It has been shown that the nuclei of the species examined have all these five histone fractions. However, the degree of similarity between homologous fractions from amoebae and the calf thymus varies and can be expressed in terms of immunological distance. Immunological differences between amoebic and calf thymus histones are the most pronounced in H1, being least in H3 and H4. Judged by its immunochemical characteristics, the histone fraction H2A from P. dubia is closer to the corresponding fraction from the calf thymus than is H2A from the other three amoeba species.     

A pathway for the interconversion of hexose and pentose in the parasitic amoeba Entamoeba histolytica.

The following three potent inhibitors of hepatocytic proteolysis were investigated to see if they would inhibit the intracellular inactivation of enzymes: chymostatin and leupeptin (proteinase inhibitors) and methylamine (a lysosomotropic weak base). Chymostatin inhibited the inactivation of two of the three enzymes tested: tyrosine aminotransferase (EC 2.6.1.5) and tryptophan oxygenase (tryptophan 2,3-dioxygenase, EC 1.13.11.11). Leupeptin had no effect on any of the enzymes, whereas methylamine had only a weak inhibitory effect on tyrosine aminotransferase inactivation. Apparently proteolytic cleavage (probably by a non-lysosomal proteinase, since only chymostatin is effective) is involved in the inactivation of tyrosine aminotransferase and tryptophan oxygenase. The third enzyme, benzopyrene hydroxylase (flavoprotein-linked mono-oxygenase, EC 1.14.14.1), is probably inactivated by a non-proteolytic mechanism.     

Digestion of seaweeds by the marine amoeba Trichosphaerium

A crude enzyme preparation from the marine amoeba Trichosphaerium was used to produce protoplasts from Sargassum muticum, Macrocystis pyrifera Porphyra perforata, and other red and brown marcroalgae. Cortical and medullary protoplasts of Sargassum, which were impossible to obtain using mixtures of previously available enzymes have now been prepared. Intact inner cortical and medullary protoplasts of Macrocystis, which were not observed in past isolations were obtained. Improved protoplast yields of as much as 500 fold resulted from feeding the amoebae on specific seaweeds. Cuticles of live Sargassum and Macrocystis were digested easily by the amoebae. However cuticles of autoclaved Macrocystis and those of Porphyra (fresh or autoclaved) were eaten last. In spite of the absence of identifiable extracellular enzymatic activity in the medium the amoebae were able to cut and consume live fronds and blocks of gelled agars carrageenans, and alginates.     

Endosymbiotic methanogenic bacteria of the sapropelic amoeba Mastigella

Abstract Large numbers of Gram-positive methanogenic bacteria, which morphologicallt resembled Methanobacterium formicicum , were found in the flagellated amoeba Mastigella from fresh water sapropel. A non-methanogenic Gram-positive bacterium with a characteristic axial cleft was found to live in close association with the amoebal nucleus. The similarity between the symbiotic system of Mastigella and that of the giant amoeba Pelomyxa is discussed, as well as the contribution of sapropelic amoebae to the methanogenesis of the sapropel ecosystem.