-
The Korean Journal of Parasitology Dec 1996The taxonomic validity of morphological group III Acanthamoeba spp. is uncertain. In the present study, six type strains of group III Acanthamoeba spp., A. culbertsoni,...
The taxonomic validity of morphological group III Acanthamoeba spp. is uncertain. In the present study, six type strains of group III Acanthamoeba spp., A. culbertsoni, A. healyi, A. pustulosa, A. palestinensis, A. royreba and A. lenticulata were subjected for the evaluation of their taxonomic validity by comparison of the isoenzyme patterns by isoelectic focusing on polyacrylamide gels, mitochondrial DNA (Mt DNA) restriction fragment length polymorphism (RFLP), and small subunit ribosomal DNA (ssu rDNA) PCR-RFLP patterns. The Mt DNA RFLP patterns were heterogeneous between the species. The type strains of A. palestinensis and A. pustulosa showed almost identical patterns of isoenzymes and rDNA PCR RFLP with an estimated sequence divergence of 2.6%. The other species showed heterogeneous patterns of isoenzymes and rDNA PCR-RFLP. It is likely that A. pustulosa is closely related with A. palestinensis and that the former may be regarded as a junior synonym of the latter.
Topics: Acanthamoeba; Animals; DNA, Mitochondrial; DNA, Protozoan; DNA, Ribosomal; Isoelectric Focusing; Isoenzymes; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length
PubMed: 9017912
DOI: 10.3347/kjp.1996.34.4.259 -
Iranian Journal of Parasitology Mar 2010Acanthamoeba keratitis develops by pathogenic Acanthamoeba such as A. palestinensis. Indeed this species is one of the known causative agents of amoebic keratitis in...
BACKGROUND
Acanthamoeba keratitis develops by pathogenic Acanthamoeba such as A. palestinensis. Indeed this species is one of the known causative agents of amoebic keratitis in Iran. Mannose Binding Protein (MBP) is the main pathogenicity factors for developing this sight threatening disease. We aimed to characterize MBP gene in pathogenic Acanthamoeba isolates such as A. palestinensis.
METHODS
This experimental research was performed in the School of Public Health, Tehran University of Medical Sciences, Tehran, Iran during 2007-2008. A. palestinensis was grown on 2% non-nutrient agar overlaid with Escherichia coli. DNA extraction was performed using phenol-chloroform method. PCR reaction and amplification were done using specific primer pairs of MBP. The amplified fragment were purified and sequenced. Finally, the obtained fragment was deposited in the gene data bank.
RESULTS
A 900 bp PCR-product was recovered after PCR reaction. Sequence analysis of the purified PCR product revealed a gene with 943 nucleotides. Homology analysis of the obtained sequence showed 81% similarity with the available MBP gene in the gene data bank. The fragment was deposited in the gene data bank under accession number EU678895,
CONCLUSION
MBP is known as the most important factor in Acanthamoeba pathogenesis cascade. Therefore, characterization of this gene can aid in developing better therapeutic agents and even immunization of high-risk people.
PubMed: 22347229
DOI: No ID Found -
Development, Growth & Differentiation Oct 1987Acanthamoeba palestinensis grown in monolayer cultures encysted almost synchronously at a stationary phase, with a yield of about 80% cysts. Under these growth...
Acanthamoeba palestinensis grown in monolayer cultures encysted almost synchronously at a stationary phase, with a yield of about 80% cysts. Under these growth conditions an encystmentinducing factor was released into the medium by transforming amoebae. Cell-free supernatants induced encystment of amoebae from early-log phase cultures. The not dialyzable encystment factor was resistant to nuclease, protease and trypsin digestion, as well as to boiling, but the activity was almost completely destroyed by autoclaving. Isolation and further characterization of the factor will enable clarification of the mode of its action as a regulator of amoeba-cyst transformation.
PubMed: 37282095
DOI: 10.1111/j.1440-169X.1987.00547.x -
Journal of Applied Microbiology Nov 2009To advance our understanding of the mechanisms involved in the RLP068 phthalocyanine-photosensitized inactivation of Acanthamoeba palestinensis trophozoites through a...
AIMS
To advance our understanding of the mechanisms involved in the RLP068 phthalocyanine-photosensitized inactivation of Acanthamoeba palestinensis trophozoites through a precise identification of the targets of the photoprocess in both the cytosolic and mitochondrial compartments.
METHODS AND RESULTS
We followed the activities of selected marker enzymes as well as we performed fluorescence and transmission electron microscopy investigations of the alterations induced by the photoprocess in the fine structure of subcellular compartments. RLP068 is preferentially located in the contractile vacuole: the fluorescence in that site is particularly evident in the unirradiated cells and becomes more diffused after irradiation. Electron microscopic analysis of photosensitized A. palestinensis cells clearly shows that the swelling of trophozoites and the appearance of vacuoles spread throughout the cytoplasm after phototreatment. The activity of a typical cytoplasmic enzyme, such as lactate dehydrogenase, underwent a 35% decrease as a consequence of the photoprocess, reflecting the photodamage induced by migrating phthalocyanine molecules in their micro-environment.
CONCLUSIONS
The presence of multiple targets for the phthalocyanine-photosensitized process is of utmost importance because this pattern of cell damage makes it unlikely that photoresistant A. palestinensis strains are gradually selected or mutagenic phenomena are developed as a consequence of the photoinduced damage.
SIGNIFICANCE AND IMPACT OF THE STUDY
Photosensitization via phthalocyanines appears to represent an efficient and safe approach for achieving a close control of the population of a potentially pathogenic protozoan such as A. palestinensis, opening new perspectives for the disinfection of microbiologically polluted waters.
Topics: Acanthamoeba; Caspase 3; Indoles; L-Lactate Dehydrogenase; Lactic Acid; Microscopy, Electron, Transmission; Microscopy, Fluorescence; Mitochondria; Organometallic Compounds; Photochemotherapy; Photosensitizing Agents; Spectrophotometry; Succinic Acid; Trophozoites
PubMed: 19457022
DOI: 10.1111/j.1365-2672.2009.04348.x -
Photochemical & Photobiological... Jun 2003Incubation of Acanthamoeba palestinensis cells with a tetracationic phthalocyanine (RLP068) at concentrations ranging between 0.2 and 1.0 microM, caused a ready uptake...
Incubation of Acanthamoeba palestinensis cells with a tetracationic phthalocyanine (RLP068) at concentrations ranging between 0.2 and 1.0 microM, caused a ready uptake of the photosensitizer with recoveries of the order of 0.5-2.5 nmol per mg of cell protein. The amount of cell-bound phthalocyanine did not appreciably change with incubation times ranging between 0.5 and 3 h. Fluorescence microscopic investigations showed an obvious accumulation of the phthalocyanine at the level of the vacuolar membranes. A nearly complete photoinduced cell death occurred upon irradiating A. palestinensis cells with 600-700 nm light with a total energy of 15-30 J cm(-2) using 1.0 microM RLP068 in the incubation medium. DAPI staining of the photosensitized cells indicates significant damage of the nucleus. On the other hand, photosensitization of the protozoan cells does not directly involve the mitochondria as shown by the lack of photoinduced decrease in the activity of typical mitochondrial enzymes, such as NADH dehydrogenase and citrate synthase.
Topics: Acanthamoeba; Animals; Cell Death; Cell Nucleus; Citrate (si)-Synthase; Dose-Response Relationship, Drug; Indoles; Isoindoles; Microscopy, Fluorescence; Mitochondria; NADH Dehydrogenase; Organometallic Compounds; Photochemotherapy; Photosensitivity Disorders; Photosensitizing Agents
PubMed: 12859151
DOI: 10.1039/b300293d -
Journal of Applied Microbiology Jul 2006To develop alternative approaches for medical and environmental control of pathogenic Acanthamoeba spp. by means of photodynamic treatment with a tetracationic...
AIMS
To develop alternative approaches for medical and environmental control of pathogenic Acanthamoeba spp. by means of photodynamic treatment with a tetracationic Zn(II)-phthalocyanine (RLP068).
METHODS AND RESULTS
Incubation of cyst cultures with RLP068 for 1 h caused an accumulation of readily detectable concentrations of the phthalocyanine, even at doses as low as 0.5 micromol l(-1). RLP068 exhibited no dark toxicity towards cysts up to 5 micromol l(-1) concentration. A decrease of c. 50% in cyst survival in comparison with controls was measured upon incubation of the cysts with 0.5 micromol l(-1) RLP068, followed by exposure to light (600-700 nm) for 20 min at a fluence rate of 50 mW cm(-2) (60 J cm(-2)). After incubation with 3 and 5 micromol l(-1) RLP068 and irradiation, the cysts lost their excystment ability as early as day 5 and up to day 10, and were clearly damaged when observed under an interference contrast microscope.
CONCLUSIONS
These data indicate the promising use of RLP068 in phototreatment of diseases caused by pathogenic amoebae and in initial disinfection of wastewaters.
SIGNIFICANCE AND IMPACT OF THE STUDY
Rapid and extensive photodamage may be induced in the highly resistant cystic stages by means of 600- to 700-nm light sources.
Topics: Acanthamoeba; Animals; Disinfectants; Environmental Microbiology; Indoles; Infection Control; Isoindoles; Light; Microscopy, Fluorescence; Microscopy, Phase-Contrast; Oocysts; Organometallic Compounds; Photosensitizing Agents; Spectrophotometry
PubMed: 16834608
DOI: 10.1111/j.1365-2672.2006.02893.x -
Canadian Journal of Microbiology Dec 1989The hypothesis of positive interactions between Francisella tularensis LVS (live vaccine strain) and Acanthamoeba palestinensis was tested. Pregrowth of the amoebae, in...
The hypothesis of positive interactions between Francisella tularensis LVS (live vaccine strain) and Acanthamoeba palestinensis was tested. Pregrowth of the amoebae, in a rich (autoclaved and filtered) medium, from which they were subsequently removed by filtration, conditioned the medium so that growth of the live vaccine strain of F. tularensis occurred and the growth rate of one other strain was increased.
Topics: Acanthamoeba; Animals; Culture Media; Francisella tularensis; Humans; Regression Analysis; Sterilization
PubMed: 2630032
DOI: 10.1139/m89-184 -
Life Sciences Mar 1986Preincubation of Acanthamoeba palestinensis homogenates in 0.25M sucrose-TM (2mM MgSO4 and 5mM Tris-HCl, pH 7.4) at 0 degree C for increasing periods of time up to 3 h,...
Preincubation of Acanthamoeba palestinensis homogenates in 0.25M sucrose-TM (2mM MgSO4 and 5mM Tris-HCl, pH 7.4) at 0 degree C for increasing periods of time up to 3 h, leads to a progressive increase in the activity of adenylated cyclase. In contrast, preincubation of isolated membrane fractions enriched in enzyme activity in the same medium results in no activation. However, preincubation of membrane fractions in medium containing a high density of sugars (sucrose, glucose or fructose) mimics the activation obtained with homogenates. The high density sugar activation is time and temperature dependent, and reversible upon return to a low density medium. The high osmotic pressure of the sugars utilized may be a factor, since high concentrations of the sucrose polymer, Ficoll, which has low osmotic activity, causes not activation. Soluble activators, protein synthesis and changes in cyclic nucleotide phosphodiesterase activity were all eliminated as possible effectors of the apparent activation of adenylate cyclase. In contrast to mammalian adenylate cyclase, the endoplasmic reticulum localized enzyme of Acanthamoeba is inhibited by NaF and is unaffected by GTP, adenosine, epinephrine, prostaglandin E1, propranolol, and meclofenamic acid. These data indicate that the adenylate cyclase of Acanthamoeba is structurally different from that of most mammalian cells.
Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Adenosine Triphosphate; Adenylyl Cyclases; Amoeba; Animals; Caffeine; Cold Temperature; Enzyme Activation; Fructose; Glucose; Osmotic Pressure; Sodium Fluoride; Sucrose; Time Factors
PubMed: 3005797
DOI: 10.1016/0024-3205(86)90602-8 -
The Journal of Cell Biology Sep 1971The membranes of Acanthamoeba palestinensis were studied by examination in fixed cells, and then by following the movements of glycerol-(3)H-labeled phospholipids by...
The membranes of Acanthamoeba palestinensis were studied by examination in fixed cells, and then by following the movements of glycerol-(3)H-labeled phospholipids by cell fractionation. Two previously undescribed structures were observed: collapsed cytoplasmic vesicles of cup shape, and plaques in food vacuole and plasma membrane similar in size to the collapsed vesicles. It appeared that the plaques formed by insertion of collapsed vesicles into membranes and/or that collapsed vesicles formed by pinching off of plaques. Fractions were isolated, enriched with nuclei, rough endoplasmic reticulum (RER), plasma membrane, Golgi-like membranes, and collapsed vesicles. The changes in specific activity of glycerol-(3)H-labeled phospholipids in these membranes during incorporation, turnover, and after pulse-labeling indicated an ordered sequence of appearances of newly synthesized phospholipids, first in nuclei and RER, then successively in Golgi membranes, collapsed vesicles, and finally, plasma membrane. In previous work we had found no large nonmembranous phospholipid pool in A. palestinensis. These observations are consistent with the hypothesis that membrane phospholipids are synthesized, perhaps as integral parts of membranes, in RER and nuclei. Subsequently, some of the newly synthesized phospholipids are transported to the Golgi complex to become integrated into the membranes of collapsed vesicles, which are precursors of the plasma membrane. Collapsed vesicles from the plasma membrane by inserting into it as plaques. When portions of the plasmalemma from food vacuoles, collapsed vesicles pinch off from their membranes and are recycled back to the cell surface.
Topics: Amoeba; Animals; Cell Fractionation; Cell Membrane; Cell Nucleus; Centrifugation, Zonal; Endoplasmic Reticulum; Glycerol; Golgi Apparatus; Histocytochemistry; Inclusion Bodies; Kinetics; Microscopy, Electron; Microscopy, Phase-Contrast; Microsomes; Phospholipids; Time Factors; Tritium
PubMed: 4329152
DOI: 10.1083/jcb.50.3.634 -
The Journal of Cell Biology Sep 1971In order to determine the feasibility of using radioactive precursors as markers for membrane phospholipids in Acanthamoeba palestinensis, the characteristics of...
Assembly of lipids into membranes in Acanthamoeba palestinensis. I. Observations on the specificity and stability of choline- 14 C and glycerol- 3 H as labels for membrane phospholipids.
In order to determine the feasibility of using radioactive precursors as markers for membrane phospholipids in Acanthamoeba palestinensis, the characteristics of phospholipids labeled with choline-(14)C and glycerol-(3)H were examined. Choline-(14)C was found to be a specific label for phosphatidyl choline. There was a turnover of the radioactive moiety of phosphatidyl choline at a rate that varied with the concentration of nonradioactive choline added to the growth medium. Radioactivity was lost from labeled phosphatidyl choline into the acid-soluble intracellular pool and from the pool into the extracellular medium. This loss of radioactivity from cells leveled off and an equilibrium was reached between the label in the cells and in the medium. Radioactive choline was incorporated into phosphatidyl choline by cell-free microsomal suspensions. This incorporation leveled off with the attainment of an equilibrium between the choline-(14)C in the reaction mixture and the choline-(14)C moiety of phosphatidyl choline in the microsomal membranes. Therefore, a choline exchange reaction may occur in cell-free membranes, as well as living A. palestinensis. In contrast to choline-(14)C, the apparent turnover of glycerol-(3)H-labeled phospholipids was not affected by large concentrations of nonradioactive choline or glycerol in the medium. The radioactivity in lipids labeled with glycerol-(3)H consisted of 33% neutral lipids and 67% phospholipids. Phospholipids labeled with glycerol-(3)H turned over slowly, with a concomitant increase in the percentage of label in neutral lipids, indicating a conversion of phospholipids to neutral lipids. Because most ( approximately 96%) of the glycerol-(3)H recovered from microsomal membranes was in phospholipids, whereas only a minor component ( approximately 2%) of the glycerol-(3)H was in the phospholipids isolated from nonmembrane lipids, glycerol-(3)H was judged to be a specific marker for membrane phospholipids.
Topics: Amoeba; Animals; Carbon Isotopes; Cell Fractionation; Cell Membrane; Cell-Free System; Choline; Culture Media; Germ-Free Life; Glycerides; Glycerol; Lipids; Microsomes; Phosphatidylcholines; Phospholipids; Postural Balance; Time Factors; Tritium
PubMed: 5098863
DOI: 10.1083/jcb.50.3.625