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PubMed1989

Mechanics and control of the cytoskeleton in Amoeba proteus.

Many models of the cytoskeletal motility of Amoeba proteus can be formulated in terms of the theory of reactive interpenetrating flow (Dembo and Harlow, 1986). We have devised numerical methodology for testing such models against the phenomenon of steady axisymmetric fountain flow. The simplest workable scheme revealed by such tests (the minimal model) is the main preoccupation of this study. All parameters of the minimal model are determined from available data. Using these parameters the model quantitatively accounts for the self assembly of the cytoskeleton of A. proteus: for the formation and detailed morphology of the endoplasmic channel, the ectoplasmic tube, the uropod, the plasma gel sheet, and the hyaline cap. The model accounts for the kinematics of the cytoskeleton: the detailed velocity field of the forward flow of the endoplasm, the contraction of the ectoplasmic tube, and the inversion of the flow in the fountain zone. The model also gives a satisfactory account of measurements of pressure gradients, measurements of heat dissipation, and measurements of the output of useful work by amoeba. Finally, the model suggests a very promising (but still hypothetical) continuum formulation of the free boundary problem of amoeboid motion. by balancing normal forces on the plasma membrane as closely as possible, the minimal model is able to predict the turgor pressure and surface tension of A. proteus. Several dynamical factors are crucial to the success of the minimal model and are likely to be general features of cytoskeletal mechanics and control in amoeboid cells. These are: a constitutive law for the viscosity of the contractile network that includes an automatic process of gelation as the network density gets large; a very vigorous cycle of network polymerization and depolymerization (in the case of A. proteus, the time constant for this reaction is approximately 12 s); control of network contractility by a diffusible factor (probably calcium ion); and control of the adhesive interaction between the cytoskeleton and the inner surface of the plasma membrane.

Amoeba↗

PubMed1985

Sterol biosynthesis de nova via cycloartenol by the soil amoeba Acanthamoeba polyphaga.

The soil amoeba Acanthamoeba polyphaga is capable of synthesizing its sterols de novo from acetate. The major sterols are ergosterol and poriferasta-5,7,22-trienol. Furthermore C28 and C29 sterols of still unknown structure with an aromatic B-ring are also synthesized by the amoeba. The first cyclic sterol precursor is cycloartenol, which is the sterol precursor in all photosynthetic phyla. No trace of lanosterol, which is the sterol precursor in animals and fungi, could be detected. These results show that at least some of the biochemical processes of Acanthamoeba polyphaga might be phylogenetically related to those of unicellular algae. Addition of exogenous sterols to the culture medium does not influence the sterol biosynthesis and the sterol composition of the cells.

Acetates↗

PubMed2006

Towards accurate solvation dynamics of divalent cations in water using the polarizable amoeba force field: From energetics to structure.

Molecular dynamics simulations were performed using a modified amoeba force field to determine hydration and dynamical properties of the divalent cations Ca2+ and Mg2+. The extension of amoeba to divalent cations required the introduction of a cation specific parametrization. To accomplish this, the Thole polarization damping model parametrization was modified based on the ab initio polarization energy computed by a constrained space orbital variation energy decomposition scheme. Excellent agreement has been found with condensed phase experimental results using parameters derived from gas phase ab initio calculations. Additionally, we have observed that the coordination of the calcium cation is influenced by the size of the periodic water box, a recurrent issue in first principles molecular dynamics studies.

Amoeba↗

PubMed2002

The analysis of 100 genes supports the grouping of three highly divergent amoebae: Dictyostelium, Entamoeba, and Mastigamoeba.

The phylogenetic relationships of amoebae are poorly resolved. To address this difficult question, we have sequenced 1,280 expressed sequence tags from Mastigamoeba balamuthi and assembled a large data set containing 123 genes for representatives of three phenotypically highly divergent major amoeboid lineages: Pelobionta, Entamoebidae, and Mycetozoa. Phylogenetic reconstruction was performed on approximately 25,000 aa positions for 30 species by using maximum-likelihood approaches. All well-established eukaryotic groups were recovered with high statistical support, validating our approach. Interestingly, the three amoeboid lineages strongly clustered together in agreement with the Conosa hypothesis [as defined by T. Cavalier-Smith (1998) Biol. Rev. Cambridge Philos. Soc. 73, 203-266]. Two amitochondriate amoebae, the free-living Mastigamoeba and the human parasite Entamoeba, formed a significant sister group to the exclusion of the mycetozoan Dictyostelium. This result suggested that a part of the reductive process in the evolution of Entamoeba (e.g., loss of typical mitochondria) occurred in its free-living ancestors. Applying this inexpensive expressed sequence tag approach to many other lineages will surely improve our understanding of eukaryotic evolution.

Amoeba↗

PubMed2000

A molecular reassessment of the Leptomyxid amoebae.

Leptomyxid amoebae encompass a diverse assemblage of amoeboid protists that have been implicated as encephalitis-causing agents. This characteristic is attributed to recent studies identifying new members of the Leptomyxidae, in particular, Balamuthia mandrillaris, that cause the disease. Their morphologies range from limax to plasmodial, as well as reticulated and polyaxial. Although systematic studies have identified B. mandrillaris as a new member of the Leptomyxidae, its precise placement within the leptomyxids is uncertain. To further assess the taxonomic placement of Balamuthia among the leptomyxid amoebae and to determine whether the members of the Leptomyxida form a monophyletic assemblage, we have sequenced 16S-like rRNA genes from representatives of three leptomyxid families. Our phylogenetic analyses revealed that current members of the order Leptomyxida do not constitute a monophyletic assemblage. Our analyses clearly show that Gephyramoeba, as well as Balamuthia do not belong in the order Leptomyxida. We highlight where molecular data give differing insights than taxonomic schemes based on traditional characters.

Amoeba↗

PubMed1983

Recovery of soil amoebae from the air during the harmattan in Zaria, Nigeria.

An attempt was made to recover soil amoebae from the air during the harmattan in Zaria, Nigeria. Non-nutrient agar plates seeded with Escherichia coli were used as settle plates and exposed to the air for 30 minutes to four hours, after which they were incubated at 27 degrees C or at 37 degrees C. A total of 38 strains of amoebae were cultured: 21 of the genus Hartmannella, eight Naegleria, four Schizopyrenus, three Didascalus and two Tetramitus. Three pathogenic species, H. culbertsoni, N. fowleri and H. rhysodes, were also recovered and all killed mice. These results suggest the possibility of airborne primary amoebic meningoencephalitis infections in Nigeria.

Air Microbiology↗

PubMed2002

Free-living amoebae promote growth and survival of Helicobacter pylori.

Transmission routes of Helicobacter pylori remain poorly understood. The finding of bacterial DNA in water suggests the involvement of environmental factors. Here we report successful co-cultivation of H. pylori with Acanthamoeba castellanii, which circumvents the requirement of this bacterium for precise microaerobic conditions and a large supply of nutrients in order to grow. H. pylori was able to propagate and remain viable for several weeks in the presence of amoebae under experimental conditions. Intact, metabolically active bacteria could be demonstrated in vacuoles. The putative dependence of H. pylori on free-living amoebae in nature could be important with respect to transmission and prevalence, as shown for some other pathogenic bacteria.

Acanthamoeba↗

PubMed1974

Chemical characterization of the isolated cell surface of Amoeba.

The cell surface has been isolated from uninucleate, freshwater, phagocytic amoebae by a new procedure. Several criteria were employed to demonstrate purity of the cell surface fraction. All morphological components of the tripartite surface were present in the isolated surface and the weight of the isolated surface was quantitatively accounted for by the components analyzed. Chemical analyses showed the presence of lipid, protein, and carbohydrate. Mannose was the predominant neutral sugar. Analyses for three different strains of Amoeba were similar. Phosphate was found to be the major anionic group in the cell surface material. Sulfate, uronic acid, sialic acid, muramic acid, and nonamidated glutamic acid and aspartic acid were absent. Evidence is presented suggesting that the phosphate is associated with an unidentified nonreducing polyol.

Acid Phosphatase↗

PubMed1979

Calcium distribution in Amoeba proteus.

A preliminary investigation of the distribution of cellular calcium in Amoeba proteus was undertaken. Total cellular calcium under control conditions was found to be 4.59 mmol/kg of cells. When the external Ca++ concentration is increased from the control level of 0.03 to 20 mM, a net Ca++ influx results with a new steady-state cellular calcium level being achieved in integral of 3 h. At steady state the amount of calcium per unit weight of cells is higher than the amount of calcium per unit weight of external solution when the external concentration of Ca++ is below 10 mM. At external Ca++ concentrations above this level, total cellular calcium approaches the medium level of Ca++. Steady-state calcium exchange in Amoeba proteus was determined with 45Ca. There is an immediate and rapid exchange of integral of 0.84 mmol/kg of cells or 18% of the total cellular calcium with the labelled Ca++. Following this initial exchange, there was very little if any further exchange observed. Most of this exchanged calcium could be eliminated from the cell with 1 mM La+++, suggesting that the exchanged calcium is associated with the surface of the cell. Increase in either the external Ca++ concentration of pH raise the amount of exchangeable calcium associated with the cell. Calcium may be associated with the cell surface as a co-ion in the diffuse double layer or bound to fixed negative sites on the surface of the cell. If Ca++-binding sites do exist on the cell surface, there may be more than one type and they may have different dissociation constants. The cytoplasmic Ca++ ion activity is probably maintained at very low levels.

Amoeba↗

PubMed1987

Amoebae and humidifier fever.

One hundred and nineteen sera from workers at four different work sites exposed to different contaminated humidifiers were examined by the immunofluorescent antibody (IFA) technique for antibodies to the amoebae Acanthamoeba polyphaga and Naegleria gruberi. Twenty-five of the sera were from workers with humidifier fever (HF) and six from workers with work related asthma (WRA) shown to be due to the contaminated humidifiers. A positive IFA test was found to correlate precipitin reaction to humidifier antigen, but did not correlate with smoking habit, work related symptoms (detected by standard questionnaire) or with HF or WRA. Amoebae were identified in all humidifiers studied.

Amoeba↗

PubMed1978

Interference reflection microscopy of adhesion of Amoeba proteus.

A simple method of observing the adhesive behaviour of large cells by means of interference reflection microscope is described, and some observations of monopodial Amoeba proteus are presented. Amoebae may contact with the substratum at any point along the long axis of the cells. Points of contact are usually few, small and temporary. Frequently the cell surface within the contact points oscillates, changing the separation distance from the substratum.

Amoeba↗

PubMed1981

Some effects of chemical irritants on the membrane of the giant amoeba.

1 The effects of chemical irritants on the membrane potential and input resistance of the giant amoeba, Chaos carolinense, have been investigated. The membrane potential and input resistance were -111.5 mV and 8.6 M pi respectively. 2 In the resting state the cell membrane of Chaos carolinense was found to be impermeable to Na+ but permeable to K+. The distribution of K+ across the cell membrane conformed to a Donnan equilibrium with the resting membrane potential being the K+ equilibrium potential. 3 The chemical irritants dibenzoxazepine and its 2-chloro- and 3-chloro-analogues and o-chlorobenzylidene malononitrile produced a fall in input resistance but no change in membrane potential. It is suggested that these effects are caused by an increase in K+ permeability. 4 The potencies of a series of chemical irritants with respect to dibenzoxazepine were measured on the giant amoeba. These potencies did not reflect those found in mammalian preparations.

Amoeba↗

PubMed1987

Seasonal distribution of thermotolerant free-living amoebae. II. Lake Issaqueena.

A quantitative study of the seasonal distribution of thermotolerant (37 degrees C and 45 degrees C), small free-living amoebae (FLA) was conducted in Lake Issaqueena, a warm, monomictic lake with steep, sloping banks and a maximum basin depth of 10 m in the Piedmont region of South Carolina. Naegleria and Vahlkampfia were the most frequently encountered FLA in littoral sediment and surface water samples whereas Acanthamoeba was most commonly isolated from profundal sediment, especially during late summer. In the water column, FLA populations were highest in a persistent detrital layer; however, few amoebae were isolated from a massive (approximately 1.5 m thick) layer of Oscillatoria. The only N. fowleri isolated in this study was from the detrital layer. Discussion of the influence of differences in watershed and basin morphology on variations in the size and generic composition of FLA populations for the aquatic ecosystems of Lake Issaqueena and Willard's Pond is included.

Amoeba↗

PubMedDate not supplied

Antigenic reactivity of a monoclonal antibody against Amoeba proteus actin.

The reactivity of a monoclonal antibody against actin of Amoeba proteus with actins from other sources was examined. The monoclonal antibody cross-reacted with actins from vertebrate muscles, human erythrocytes, and Acanthamoeba castellanii, but it did not react with Naegleria gruberi actin. The amoeba actin was resolved into 3 bands with isoelectric points of 5.96, 6.03 and 6.10 in electrofocusing gels and they corresponded to 3 peptide spots reacting with the antibody on 2-dimensional immunoblots.

Actins↗

PubMedDate not supplied

Phylogenetic analyses of small subunit ribosomal RNA coding regions reveal a monophyletic lineage of euglyphid testate amoebae (Order Euglyphida).

The Testaceafilosia includes amoebae with filopodia and with a proteinaceous, agglutinated or siliceous test. To explore the deeper phylogeny of this group, we sequenced the small subunit ribosomal RNA coding region of 13 species, including the first sequence of an amoeba with an agglutinated test, Pseudodifflugia sp. Phylogenetic analyses using maximum parsimony and maximum likelihood methods as well as neighbor joining method yielded the following results: the order Euglyphida forms a monophyletic lineage with the sarcomonads as sister group. The next related taxa are the Chlorarachnea and the unidentified filose strain N-Por. In agreement with the previous studies the Phytomyxea branch off at the base of this lineage. The Monadofilosa (Testaceafilosia and Sarcomonadea) appear monophyletic. The Testaceafilosia are polyphyletic, because Pseudodifflugia sp. is positioned as the sister taxon to the sarcomonads. Within the order Euglyphida Paulinella branches off first, together with Cyphoderia followed by Tracheleuglypha. In maximum likelihood and neighbor joining analyses, the genus Euglypha is monophyletic. The branching pattern within the order Euglyphida reflects the evolution of shell morphology from simple to complex built test.

Amoeba↗

PubMedDate not supplied

Bacterial endosymbionts of free-living amoebae.

The occurrence of bacterial endosymbionts in free-living amoebae has been known for decades, but their obligate intracellular lifestyle hampered their identification. Application of the full cycle rRNA approach, including 16S rRNA gene sequencing and fluorescence in-situ hybridization with 16S rRNA-targeted oligonucleotide probes, assigned the symbionts of Acanthamoeba spp. and Hartmannella sp. to five different evolutionary lineages within the Proteobacteria, the Bacteroidetes, and the Chlamydiae, respectively. Some of these bacterial symbionts are most closely related to bacterial pathogens of humans, and it has been suggested that they should be considered potential emerging pathogens. Complete genome sequence analysis of a chlamydia-related symbiont of Acanthamoeba sp. showed that this endosymbiont uses similar mechanisms for interaction with its eukaryotic host cell as do the well-known bacterial pathogens of humans. Furthermore, phylogenetic analysis suggested that these mechanisms have been evolved by the ancestor of these amoeba symbionts in interplay with ancient unicellular eukaryotes.

Amoeba↗

PubMedDate not supplied

A method for estimating cell volume of amoebae based on measurements of cell length of motile forms: physiological and ecological applications.

A novel method of estimating the cell volume of non-testate, free-living amoebae is presented. Measurements of the length (L) of motile forms of some amoebae, obtained by applying some basic rules, can be used to calculate diameter (D) using a relatively simple formula, derived from regression analysis (N=39): D=0.6L. Important physiological and ecological variables, including carbon content and respiratory rate, can then be easily calculated using this estimated cell volume.

Amoeba↗

PubMed1975

Membrane potential and conductance during pinocytosis induced in Amoeba proteus with alkali metal ions.

An investigation of the relationship between the polarized state of the membrane and the onset and the intensity of pinocytosis was made in Amoeba proteus. Membrane potential and input resistance was in all instances found to decrease in approximate proportion to the number of channels when pinocytosis was induced by a variety of alkali metal ions at varying pH. Channels began to appear when the membrane was depolarized to -30 mV by the inducer of pinocytosis. With all inducers the maximum pinocytosis was encountered at membrane potentials close to zero. No positive potentials were recorded when the chloride salts of the inducing cations were used. At high concentrations of alkali ions a transient increase of the chloride permeability caused short-lasting hyperolarizations of the membrane. Inhibition of pinocytosis by Ca++ was accompanied by an increase of input resistance and membrane potential. The selectivity of the membrane to different alkali metal ions observed as changes in pinocytosis intensity, membrane potential and input resistance was found to vary with the concentration of the inducer and with the Ca++ concentration of the extracellular solution. Displacement of membrane bound Ca++ appeared to decrease the field strength of charged groups in the membrane altering its selectivity among alkali cations. The formation of pinocytotic channels is suggested to require translocation of Ca++ from the membrane into the cell and would therefore be closely related to the electrical properties of the amoeba.

Alkalies↗