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Parasitology Research Jan 2024Acanthamoeba castellanii, a ubiquitous protozoan, is responsible for significant diseases such as Acanthamoeba keratitis and granulomatous amoebic encephalitis. A...
Acanthamoeba castellanii, a ubiquitous protozoan, is responsible for significant diseases such as Acanthamoeba keratitis and granulomatous amoebic encephalitis. A crucial survival strategy of A. castellanii involves the formation of highly resistant cysts during adverse conditions. This study delves into the cellular processes underpinning encystment, focusing on gene expression changes related to reactive oxygen species (ROS) balance, with a particular emphasis on mitochondrial processes. Our findings reveal a dynamic response within the mitochondria during encystment, with the downregulation of key enzymes involved in oxidative phosphorylation (COX, AOX, and NADHalt) during the initial 48 h, followed by their overexpression at 72 h. This orchestrated response likely creates a pro-oxidative environment, facilitating encystment. Analysis of other ROS processing enzymes across the cell reveals differential expression patterns. Notably, antioxidant enzymes, such as catalases, glutaredoxins, glutathione S-transferases, peroxiredoxins, and thioredoxins, mirror the mitochondrial trend of downregulation followed by upregulation. Additionally, glycolysis and gluconeogenesis are downregulated during the early stages in order to potentially balance the metabolic requirement of the cyst. Our study underscores the importance of ROS regulation in Acanthamoeba encystment. Understanding these mechanisms offers insights into infection control and identifies potential therapeutic targets. This work contributes to unraveling the complex biology of A. castellanii and may aid in combatting Acanthamoeba-related infections. Further research into ROS and oxidase enzymes is warranted, given the organism's remarkable respiratory versatility.
Topics: Humans; Acanthamoeba castellanii; Reactive Oxygen Species; Catalase; Acanthamoeba Keratitis; Amebiasis; Cysts
PubMed: 38289423
DOI: 10.1007/s00436-024-08138-9 -
Mikrobiyoloji Bulteni Jan 2024Free-living amoebae belonging to the genus Acanthamoeba are microorganisms that live in air, soil and aquatic environments. In humans, they cause infections such as...
Free-living amoebae belonging to the genus Acanthamoeba are microorganisms that live in air, soil and aquatic environments. In humans, they cause infections such as amoebic keratitis, graulamotous amoebic encephalitis that are difficult to treat and can be fatal. In addition, it is known that they contribute to the replication of bacteria and increase their pathogenicity by being a host for various bacteria. However, information on its inhibitory properties against bacteria and its production of antimicrobial agents is very limited. In this context, in this study, it was aimed to investigate whether cell-free supernatants of Acanthamoeba strains have antibacterial effects against Pseudomonas aeruginosa isolates. Four different Acanthamoeba strains (A10, A13, A14, U.GÖL) isolated from aquatic environments in our country were selected and used in the study, P.aeruginosa isolates (PA2, PA3, PA4, PA5) were selected from clinical strains belonging to patients in our country. Acanthamoeba castellanii ATCC 50373 and P.aeruginosa ATCC 27853 were used as standard strains. P.aeruginosa isolates were grown on nutrient agar at 37 °C and Acanthamoeba strains were grown on E.coli spread non-nutrient agar at 30 °C under aerobic conditions. Pepton yeast extract glucose (PYG) medium supplemented with penicillin and streptomycin was used to obtain axenic cultures of Acanthamoeba strains. After the centrifugation of axenic cultures at 3000 rpm for five minutes, Acanthamoeba-cell-free supernatants were obtained by filtering the supernatant part through a sterile filter with a pore diameter of 0.22 µm. The antibacterial activities of these supernatants against P.aeruginosa isolates were determined using the colony counting method. Analysis of each Acanthamoeba-cell-free supernatants was performed according to the GC-MS method. Acanthamoeba-cell-free supernatants were found to have varying degrees of inhibitory effects (3.9-91.5%) against tested P.aeruginosa isolates. It was determined that the cell-free supernatant of A.castellanii ATCC 50373 strain showed the highest antibacterial effect (91.5%) against PA5 isolate. A14 strain showed similar inhibitory effects (89.4%) against the same Pseudomonas isolate. The average inhibitory effect of most of the Acanthamoeba strains of our country was found to be higher than that of the reference strain A.castellanii ATCC 50492. It is thought that the compounds responsible for the anti-Pseudomonas activity of the tested Acanthamoeba strains may be fructose, phosphoric acid, galactose, N-Acetylphenylalanine and glucopyranose determined as major compounds. This is the first study showing the anti-Pseudomonas activity of microorganisms of the genus Acanthamoeba living in the waters of our country. Acanthamoeba, which is widely found in nature, appears to be a good source for new antimicrobial agents.
Topics: Humans; Pseudomonas aeruginosa; Agar; Pseudomonas; Acanthamoeba castellanii; Anti-Bacterial Agents; Escherichia coli
PubMed: 38263942
DOI: 10.5578/mb.20249950 -
Applied and Environmental Microbiology Feb 2024In this study, we conducted an in-depth analysis to characterize potential () proteins capable of recognizing fungal β-1,3-glucans. specifically anchors curdlan or...
In this study, we conducted an in-depth analysis to characterize potential () proteins capable of recognizing fungal β-1,3-glucans. specifically anchors curdlan or laminarin, indicating the presence of surface β-1,3-glucan-binding molecules. Using optical tweezers, strong adhesion of laminarin- or curdlan-coated beads to was observed, highlighting their adhesive properties compared to controls (characteristic time τ of 46.9 and 43.9 s, respectively). Furthermore, () G217B, possessing a β-1,3-glucan outer layer, showed significant adhesion to compared to a G186 strain with an α-1,3-glucan outer layer (τ of 5.3 s vs τ 83.6 s). The addition of soluble β-1,3-glucan substantially inhibited this adhesion, indicating the involvement of β-1,3-glucan recognition. Biotinylated β-1,3-glucan-binding proteins from exhibited higher binding to G217B, suggesting distinct recognition mechanisms for laminarin and curdlan, akin to macrophages. These observations hinted at the β-1,3-glucan recognition pathway's role in fungal entrance and survival within phagocytes, supported by decreased fungal viability upon laminarin or curdlan addition in both phagocytes. Proteomic analysis identified several proteins capable of binding β-1,3-glucans, including those with lectin/glucanase superfamily domains, carbohydrate-binding domains, and glycosyl transferase and glycosyl hydrolase domains. Notably, some identified proteins were overexpressed upon curdlan/laminarin challenge and also demonstrated high affinity to β-1,3-glucans. These findings underscore the complexity of binding via β-1,3-glucan and suggest the existence of alternative fungal recognition pathways in .IMPORTANCE () and macrophages both exhibit the remarkable ability to phagocytose various extracellular microorganisms in their respective environments. While substantial knowledge exists on this phenomenon for macrophages, the understanding of 's phagocytic mechanisms remains elusive. Recently, our group identified mannose-binding receptors on the surface of that exhibit the capacity to bind/recognize fungi. However, the process was not entirely inhibited by soluble mannose, suggesting the possibility of other interactions. Herein, we describe the mechanism of β-1,3-glucan binding by and its role in fungal phagocytosis and survival within trophozoites, also using macrophages as a model for comparison, as they possess a well-established mechanism involving the Dectin-1 receptor for β-1,3-glucan recognition. These shed light on a potential parallel evolution of pathways involved in the recognition of fungal surface polysaccharides.
Topics: Acanthamoeba castellanii; Amoeba; Mannose; Proteomics; beta-Glucans; Glucans; Histoplasma
PubMed: 38259076
DOI: 10.1128/aem.01736-23 -
Microbiology Spectrum Feb 2024Carbapenem-resistant causes one of the most difficult-to-treat nosocomial infections. Polycationic drugs like polymyxin B or colistin and tetracycline drugs such as...
Carbapenem-resistant causes one of the most difficult-to-treat nosocomial infections. Polycationic drugs like polymyxin B or colistin and tetracycline drugs such as doxycycline or minocycline are commonly used to treat infections caused by carbapenem-resistant . Here, we show that a subpopulation of cells associated with the opaque/translucent colony phase variation by AB5075 displays differential tolerance to subinhibitory concentrations of colistin and tetracycline. Using a variety of microscopic techniques, we demonstrate that extracellular polysaccharide moieties mediate colistin tolerance to opaque at single-cell level and that mushroom-shaped biofilm structures protect opaque bacteria at the community level. The colony switch phenotype is found to alter several traits of , including long-term survival under desiccation, tolerance to ethanol, competition with , and intracellular survival in the environmental model host . Additionally, our findings suggest that extracellular DNA associated with membrane vesicles can promote colony switching in a DNA recombinase-dependent manner.IMPORTANCEAs a WHO top-priority drug-resistant microbe, significantly contributes to hospital-associated infections worldwide. One particularly intriguing aspect is its ability to reversibly switch its colony morphotype on agar plates, which has been remarkably underexplored. In this study, we employed various microscopic techniques and phenotypic assays to investigate the colony phase variation switch under different clinically and environmentally relevant conditions. Our findings reveal that the presence of a poly N-acetylglucosamine-positive extracellular matrix layer contributes to the protection of bacteria from the bactericidal effects of colistin. Furthermore, we provide intriguing insights into the multicellular lifestyle of , specifically in the context of colony switch variation within its predatory host, .
Topics: Humans; Colistin; Acinetobacter baumannii; Phase Variation; Acinetobacter Infections; Microbial Sensitivity Tests; Anti-Bacterial Agents; Minocycline; Carbapenems; Biofilms; DNA; Drug Resistance, Multiple, Bacterial
PubMed: 38205963
DOI: 10.1128/spectrum.02956-23 -
Heliyon Jan 2024A rare but lethal central nervous system disease known as granulomatous amoebic encephalitis (GAE) and potentially blinding keratitis are diseases caused by free-living...
A rare but lethal central nervous system disease known as granulomatous amoebic encephalitis (GAE) and potentially blinding keratitis are diseases caused by free-living . Currently, no therapeutic agent can completely eradicate or prevent GAE. Synthetic compounds are a likely source of bioactive compounds for developing new drugs. This study synthesized seventeen 1,4-benzothiazine derivatives (I -XVII) by a base-catalyzed one-pot reaction of 2-amino thiophenol with substituted bromo acetophenones. Different spectroscopic techniques, such as EI-MS, H-, and C NMR (only for the new compounds), were used for the structural characterization and conformation of compounds. These compounds were assessed for the first time against All compounds showed anti-amoebic potential against , reducing its ability to encyst and excyst at 100 μM. Compounds , , and showed the most potent activities among all derivatives and significantly reduced the viability to 5.3 × 10 ( < 0.0003), 2 × 10 ( < 0.006), and 2.4 × 10 ( < 0.002) cells/mL, respectively. The cytotoxicity profile revealed that these molecules showed lower to moderate cytotoxicity, i.e., 36 %, 2 %, and 21 %, respectively, against human keratinocytes . These results indicate that 1,4-benzothiazines showed potent activity against trophozoites and cysts of . Hence, these 1,4-benzothiazine derivatives should be considered to develop new potential therapeutic agents against infections.
PubMed: 38205285
DOI: 10.1016/j.heliyon.2023.e23258 -
Journal of Medicinal Chemistry Jan 2024is an amoeba that inhabits soil and water in every part of the world. Acanthamoeba infection of the eye causes keratitis and can lead to a loss of vision. Current...
is an amoeba that inhabits soil and water in every part of the world. Acanthamoeba infection of the eye causes keratitis and can lead to a loss of vision. Current treatment options are only moderately effective, have multiple harmful side effects, and are tedious. In our study, we developed a novel drug screening method to define the inhibitory properties of potential new drugs against in vitro. We found that the clinically used carbonic anhydrase inhibitors, acetazolamide, ethoxzolamide, and dorzolamide, have promising antiamoebic properties.
Topics: Acanthamoeba castellanii; Carbonic Anhydrase Inhibitors; Amoeba; Drug Evaluation, Preclinical
PubMed: 38150360
DOI: 10.1021/acs.jmedchem.3c01020 -
Antioxidants (Basel, Switzerland) Dec 2023is a ubiquitous genus of amoebae that can act as opportunistic parasites in both humans and animals, causing a variety of ocular, nervous and dermal pathologies....
is a ubiquitous genus of amoebae that can act as opportunistic parasites in both humans and animals, causing a variety of ocular, nervous and dermal pathologies. Despite advances in therapy, the management of patients with infections remains a challenge for health services. Therefore, there is a need to search for new active substances against Acanthamoebae. In the present study, we evaluated the amoebicidal activity of nitroxoline against the trophozoite and cyst stages of six different strains of . The strain showed the lowest IC value in the trophozoite stage (0.69 ± 0.01 µM), while the strain L-10 showed the lowest IC value in the cyst stage (0.11 ± 0.03 µM). In addition, nitroxoline induced in treated trophozoites of features compatibles with apoptosis and autophagy pathways, including chromatin condensation, mitochondrial malfunction, oxidative stress, changes in cell permeability and the formation of autophagic vacuoles. Furthermore, proteomic analysis of the effect of nitroxoline on trophozoites revealed that this antibiotic induced the overexpression and the downregulation of proteins involved in the apoptotic process and in metabolic and biosynthesis pathways.
PubMed: 38136200
DOI: 10.3390/antiox12122081 -
Microbiology Spectrum Jan 2024At the National Cheng Kung University Hospital, numerous cases of amoebic keratitis had been identified with concurrent bacterial infections. Among these bacterial...
At the National Cheng Kung University Hospital, numerous cases of amoebic keratitis had been identified with concurrent bacterial infections. Among these bacterial coinfections, accounted for 50% of the reported cases. However, the impact of pathogenic bacteria on amoeba-induced corneal damage remains unclear. In our study, we successfully demonstrated that accumulated on the surface and caused more severe corneal damage. We also indicated that the exposure of to amoeba-soluble antigens enhanced its adhesion ability, promoted biofilm formation, and led to more severe corneal cell damage. These findings significantly contributed to our understanding of the risk associated with coinfection in the progression of amoeba keratitis.
Topics: Humans; Pseudomonas aeruginosa; Coinfection; Cornea; Keratitis; Corneal Injuries
PubMed: 38095463
DOI: 10.1128/spectrum.02683-23 -
ELife Dec 2023Identifying virulence-critical genes from pathogens is often limited by functional redundancy. To rapidly interrogate the contributions of combinations of genes to a...
Identifying virulence-critical genes from pathogens is often limited by functional redundancy. To rapidly interrogate the contributions of combinations of genes to a biological outcome, we have developed a ltiplex, andomized RISPR nterference equencing (MuRCiS) approach. At its center is a new method for the randomized self-assembly of CRISPR arrays from synthetic oligonucleotide pairs. When paired with PacBio long-read sequencing, MuRCiS allowed for near-comprehensive interrogation of all pairwise combinations of a group of 44 virulence genes encoding highly conserved transmembrane proteins for their role in pathogenesis. Both amoeba and human macrophages were challenged with bearing the pooled CRISPR array libraries, leading to the identification of several new virulence-critical combinations of genes. and were particularly fascinating for their apparent redundant functions during human macrophage infection, while alone was essential for virulence in the amoeban host . Thus, MuRCiS provides a method for rapid genetic examination of even large groups of redundant genes, setting the stage for application of this technology to a variety of biological contexts and organisms.
Topics: Humans; Macrophages; Legionella pneumophila; Acanthamoeba castellanii; Virulence; Legionnaires' Disease; Bacterial Proteins
PubMed: 38095310
DOI: 10.7554/eLife.86903 -
Scientific Reports Dec 2023Phthalates constitute a family of anthropogenic chemicals developed to be used in the manufacture of plastics, solvents, and personal care products. Their dispersion and...
Phthalates constitute a family of anthropogenic chemicals developed to be used in the manufacture of plastics, solvents, and personal care products. Their dispersion and accumulation in many environments can occur at all stages of their use (from synthesis to recycling). However, many phthalates together with other accumulated engineered chemicals have been shown to interfere with hormone activities. These compounds are also in close contact with microorganisms that are free-living, in biofilms or in microbiota, within multicellular organisms. Herein, the activity of several phthalates and their substitutes were investigated on the opportunistic pathogen Legionella pneumophila, an aquatic microbe that can infect humans. Beside showing the toxicity of some phthalates, data suggested that Acetyl tributyl citrate (ATBC) and DBP (Di-n-butyl phthalate) at environmental doses (i.e. 10 M and 10 M) can modulate Legionella behavior in terms of motility, biofilm formation and response to antibiotics. A dose of 10 M mostly induced adverse effects for the bacteria, in contrast to a dose of 10 M. No perturbation of virulence towards Acanthamoeba castellanii was recorded. These behavioral alterations suggest that L. pneumophila is able to sense ATBC and DBP, in a cross-talk that either mimics the response to a native ligand, or dysregulates its physiology.
Topics: Humans; Legionella pneumophila; Legionella; Phthalic Acids; Biofilms
PubMed: 38092873
DOI: 10.1038/s41598-023-49426-1