scholarly journals Diversity of Amoeba-Associated Giant Viruses Isolated in Algeria

Diversity ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 215
Author(s):  
Hadjer Boudjemaa ◽  
Julien Andreani ◽  
Idir Bitam ◽  
Bernard La Scola
Keyword(s):  
Electron Microscopy ◽  
Flow Cytometry ◽  
Environmental Samples ◽  
Acanthamoeba Castellanii ◽  
Vermamoeba Vermiformis ◽  
Giant Viruses

The discovery of several giant amoeba viruses has opened up a novel area in the field of virology. Despite this, knowledge about ecology of these viruses remains patchy. In this study, we aimed to characterize the diversity of giant viruses in Algeria by inoculating 64 environmental samples on various amoeba strains. After isolation by co-culture with nine amoeba supports, flow cytometry and electron microscopy were used to putatively identify viruses. Definitive identification was performed by PCR and sequencing. Mimiviruses, marseilleviruses, faustoviruses and cedratviruses were the main viruses isolated in this study. Moreover, a new virus, which we named fadolivirus, was also isolated and was found to belong to the recent metagenomic descriptions of Klosneuvirinae. Despite the use of 9 amoeba supports for co-culture, most of the isolates were obtained from two amoebas: Acanthamoeba castellanii Neff and Vermamoeba vermiformis CDC 19. Finally, the viruses most frequently isolated were marseilleviruses (55.5%) and Mimiviruses (22.2%). This work shows that the isolation of viruses previously detected by metagenomic analyses can be tedious, but possible.

scholarly journals Cellular Response of the Amoeba Acanthamoeba castellanii to Chlorine, Chlorine Dioxide, and Monochloramine Treatments

2011 ◽  
Vol 77 (14) ◽  
pp. 4974-4980 ◽  
Author(s):  
Emerancienne Mogoa ◽  
Charles Bodet ◽  
Franck Morel ◽  
Marie-Hélène Rodier ◽  
Bernard Legube ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Flow Cytometry ◽  
Chlorine Dioxide ◽  
Mode Of Action ◽  
Cellular Response ◽  
Acanthamoeba Castellanii ◽  
Size Reduction ◽  
Resistant Bacteria ◽  
Free Living ◽  
Content Type

ABSTRACTAcanthamoeba castellaniiis a free-living amoebae commonly found in water systems. Free-living amoebae might be pathogenic but are also known to bear phagocytosis-resistant bacteria, protecting these bacteria from water treatments. The mode of action of these treatments is poorly understood, particularly on amoebae. It is important to examine the action of these treatments on amoebae in order to improve them. The cellular response to chlorine, chlorine dioxide, and monochloramine was tested onA. castellaniitrophozoites. Doses of disinfectants leading to up to a 3-log reduction were compared by flow cytometry and electron microscopy. Chlorine treatment led to size reduction, permeabilization, and retraction of pseudopods. In addition, treatment with chlorine dioxide led to a vacuolization of the cytoplasm. Monochloramine had a dose-dependent effect. At the highest doses monochloramine treatment resulted in almost no changes in cell size and permeability, as shown by flow cytometry, but the cell surface became smooth and dense, as seen by electron microscopy. We show that these disinfectants globally induced size reduction, membrane permeabilization, and morphological modifications but that they have a different mode of action onA. castellanii.

scholarly journals A metallo-beta-lactamase with both beta-lactamase and ribonuclease activity is linked with traduction in giant viruses

2019 ◽  
Author(s):  
Philippe Colson ◽  
Lucile Pinault ◽  
Said Azza ◽  
Nicholas Armstrong ◽  
Eric Chabriere ◽  
...  
Keyword(s):  
Acanthamoeba Castellanii ◽  
Deep Ocean ◽  
Penicillin G ◽  
Beta Lactamase ◽  
Beta Lactam ◽  
Strong Activity ◽  
Beta Lactam Antibiotics ◽  
Double Stranded Dna ◽  
Giant Viruses ◽  
Hydrolysis Of

ABSTRACTEnzymatic proteins with a metallo-beta-lactamase (MBL) fold have been essentially studied in bacteria for their activity on beta-lactam antibiotics. However, the MBL fold is ancient and highly conserved, and these proteins are capable of cleaving a broad range of substrates. It has recently been shown that MBLs are present in a wide array of cellular organisms, including eukaryotes and archaea. We show here that Tupanvirus deep ocean, a giant virus, also encodes a protein with a MBL fold. Phylogeny showed its clustering with transfer ribonucleases (RNases) and the presence of orthologs in other giant viruses, mainly those harboring the largest sets of translation components. In addition, it suggests an ancient origin for these genes and a transfer between giant viruses and Acanthamoeba spp., a host of many giant viruses. Biologically, after its expression in Escherichia coli, the tupanvirus protein was found to hydrolyse nitrocefin, a chromogenic beta-lactam. We also observed an hydrolysis of penicillin G (10 μg/mL) and detected the metabolite of penicillin G hydrolysis, benzylpenilloic acid. This was inhibited by sulbactam, a beta-lactamase inhibitor. In addition, we tested the degradation of single-stranded DNA, double-stranded DNA, and RNAs, and observed a strong activity on RNAs from seven bacteria with G+C varying from 42% to 67%, and from Acanthamoeba castellanii, the tupanvirus host. This was not inhibited by sulbactam or ceftriaxone. RNase activity was estimated to be 0.45±0.15 mU/mg using a fluorescence-based assay. Our results still broaden the range of hosts of MBL fold proteins and demonstrate that such protein can have dual beta-lactamase/nuclease activities. We suggest that they should be annotated according to this finding to avoid further confusion.

Characterization of cellular development and fatty acid accumulation in thraustochytrid Aurantiochytrium strains of Taiwan

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Natarajan Velmurugan ◽  
Yesupatham Sathishkumar ◽  
Shashanka Sonowal ◽  
Ka-Lai Pang ◽  
Yang Soo Lee
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Flow Cytometry ◽  
Cell Growth ◽  
Intracellular Lipid ◽  
Fatty Acid Accumulation ◽  
Transmission Electron ◽  
Intracellular Lipid Accumulation ◽  
High Level

Abstract Long-chain saturated and polyunsaturated fatty acids of two new thraustochytrid isolates cultured from Taiwan mangroves, Aurantiochytrium sp. IMB169 and Aurantiochytrium sp. IMB171, were characterized through their cell growth and development in relation to their intracellular lipid accumulation using transmission electron microscopy. Flow cytometry in combination with the lipophilic fluorescent dye BODIPY 505/515 was used to stain and characterize intracellular lipid bodies in the two isolates. The transmission electron microscopy and flow cytometry analyses revealed a progressive accumulation of lipid products in IMB169 and IMB171. Further, selective BODIPY stained cells were successfully separated and enriched using flow cytometry at single cell level. Among the two isolates, IMB169 was found to produce a high level of docosahexaenoic acid. The qualitative and analytical results obtained using electron microscopy and flow cytometry studies were validated by gas chromatography (GC). In addition, a quantitative baseline was established using cell growth, flow cytometry and GC analyses for developing an efficient bioprocessing methodology to selectively enrich thraustochytrids phenotypes with desirable characteristics.

scholarly journals Endlicheria bracteolata (Meisn.) Essential Oil as a Weapon Against Leishmania amazonensis: In Vitro Assay

Molecules ◽  
2019 ◽  
Vol 24 (14) ◽  
pp. 2525 ◽  
Author(s):  
Mariana Margatto Rottini ◽  
Ana Claudia Fernandes Amaral ◽  
José Luiz Pinto Ferreira ◽  
Edinilze Souza Coelho Oliveira ◽  
Jefferson Rocha de Andrade Silva ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Flow Cytometry ◽  
Essential Oil ◽  
Cutaneous Leishmaniasis ◽  
Leishmania Amazonensis ◽  
Vitro Assay ◽  
Transmission Electron ◽  
Biological Potential ◽  
Intracellular Amastigote

The difficulties encountered and the numerous side effects present in the treatment of cutaneous leishmaniasis have encouraged the research for new compounds that can complement or replace existing treatment. The growing scientific interest in the study of plants, which are already used in folk remedies, has led our group to test Endlicheria bracteolata essential oil against Leishmania amazonensis. Several species of the Lauraceae family, or their compounds, have relevant antiprotozoal activities Therefore, the biological potential on L. amazonensis forms from the essential oil of Endlicheria bracteolata leaves was verified for the first time in that work. The antileishmanial activity was evaluated against promastigotes and intracellular amastigotes, and cytotoxicity were performed with J774.G8, which were incubated with different concentrations of E. bracteolata essential oil. Transmission electron microscopy and flow cytometry were performed with E. bracteolata essential oil IC50. Promastigote forms showed E. bracteolata essential oil IC50 of 7.945 ± 1.285 µg/mL (24 h) and 6.186 ± 1.226 µg/mL (48 h), while for intracellular amastigote forms it was 3.546 ± 1.184 µg/mL (24 h). The CC50 was 15.14 ± 0.090 µg/mL showing that E. bracteolata essential oil is less toxic to macrophages than to parasites. Transmission electron microscopy showed that E. bracteolata essential oil treatment is capable of inducing mitochondrial damage to promastigote and intracellular amastigote forms, while flow cytometry showed ΔѰm disruption in treated parasites. These results could bring about new possibilities to develop products based on E. bracteolata essential oil to treat cutaneous leishmaniasis, especially for people who cannot receive the conventional therapy.

scholarly journals Thrombin induces a rapid redistribution of glycoprotein Ib-IX complexes within the membrane systems of activated human platelets

Blood ◽  
1990 ◽  
Vol 76 (8) ◽  
pp. 1503-1513 ◽  
Author(s):  
P Hourdille ◽  
E Heilmann ◽  
R Combrie ◽  
J Winckler ◽  
KJ Clemetson ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Flow Cytometry ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Ionophore A23187 ◽  
Membrane Systems ◽  
Immunogold Staining ◽  
Thin Sections ◽  
Surface Areas ◽  
Lowicryl K4m

Abstract Previous studies have shown a decreased binding of monoclonal antibodies (MoAbs) to glycoprotein (GP) Ib-IX complexes on thrombin- stimulated platelets, but the reason for this is poorly understood. We have used (1) immunofluorescence procedures and flow cytometry, and (2) immunogold staining and electron microscopy to investigate this phenomenon. Washed platelets were incubated with alpha-thrombin, adenosine diphosphate, or ionophore A23187 for increasing lengths of time. For alpha-thrombin, but not the other agonists, flow cytometry confirmed a dose- and time-dependent decrease in the binding of MoAbs specific for GP Ib alpha (AP-1, Bx-1), GP IX (FMC 25), or to the complex itself (SZ 1). Immunoglold staining performed using standard transmission or scanning electron microscopy high-lighted surface areas devoid of bound antibody. However, a quantitatively normal immunofluorescence was restored if paraformaldehyde-fixed, thrombin- stimulated platelets were permeabilized with Triton X-100 (Sigma Chemical Co, St Louis, MO) before MoAb addition, while immunogold staining was now seen to be concentrated within the interior of the platelet. Glutaraldehyde-fixed samples were then embedded in the resin Lowicryl K4M (Taab Laboratories Equipment Ltd, Aldermaston, England) and immunogold staining performed on thin sections using a polyclonal antibody to glycocalicin. An increased presence of GP Ib-IX complexes within surface-connected membrane systems of the thrombin-stimulated platelets was confirmed. Interestingly, GP Ib-IX movement was opposite to the thrombin-induced externalization of internal pools of GP IIb- IIIa complexes and of the alpha-granule membrane GP, GMP-140.

scholarly journals Proeryptotic Activity of 4-Hydroxynonenal: A New Potential Physiopathological Role for Lipid Peroxidation Products

Biomolecules ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 770
Author(s):  
Mario Allegra ◽  
Ignazio Restivo ◽  
Alberto Fucarino ◽  
Alessandro Pitruzzella ◽  
Sonya Vasto ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Lipid Peroxidation ◽  
Scanning Electron Microscopy ◽  
Flow Cytometry ◽  
Endothelial Cells ◽  
Caspase 3 ◽  
Morphological Alterations ◽  
Signalling Molecule ◽  
Free Haemoglobin ◽  
Scanning Electron

Background: Eryptosis is a physiological, apoptosis-like death of injured erythrocytes crucial to prevent premature haemolysis and the pathological sequalae generated by cell-free haemoglobin. When dysregulated, the process is associated to several inflammatory-based pathologies. 4-Hydroxy-trans-2-nonenal (HNE) is an endogenous signalling molecule at physiological levels and, at higher concentrations, is involved in the pathogenesis of several inflammatory-based diseases. This work evaluated whether HNE could induce eryptosis in human erythrocytes. Methods: Measurements of phosphatidylserine, cell volume, intracellular oxidants, Ca++, glutathione, ICAM-1, and ceramide were assessed by flow cytometry. Scanning electron microscopy evaluated morphological alterations of erythrocytes. Western blotting assessed caspases. PGE2 was measured by ELISA. Adhesion of erythrocytes on endothelial cells was evaluated by gravity adherence assay. Results: HNE in the concentration range between 10–100 µM induces eryptosis, morphological alterations correlated to caspase-3 activation, and increased Ca++ levels. The process is not mediated by redox-dependent mechanisms; rather, it strongly depends on PGE2 and ceramide. Interestingly, HNE induces significant increase of erythrocytes adhesion to endothelial cells (ECs) that are in turn dysfunctionated as evident by overexpression of ICAM-1. Conclusions: Our results unveil a new physiopathological role for HNE, provide mechanistic details of the HNE-induced eryptosis, and suggest a novel mechanism through which HNE could exert pro-inflammatory effects.

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