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Scientific Reports Nov 2021Balamuthia mandrillaris, a pathogenic free-living amoeba, causes cutaneous skin lesions as well as granulomatous amoebic encephalitis, a 'brain-eating' disease. As with...
Balamuthia mandrillaris, a pathogenic free-living amoeba, causes cutaneous skin lesions as well as granulomatous amoebic encephalitis, a 'brain-eating' disease. As with the other known pathogenic free-living amoebas (Naegleria fowleri and Acanthamoeba species), drug discovery efforts to combat Balamuthia infections of the central nervous system are sparse; few targets have been validated or characterized at the molecular level, and little is known about the biochemical pathways necessary for parasite survival. Current treatments of encephalitis due to B. mandrillaris lack efficacy, leading to case fatality rates above 90%. Using our recently published methodology to discover potential drugs against pathogenic amoebas, we screened a collection of 85 compounds with known antiparasitic activity and identified 59 compounds that impacted the growth of Balamuthia trophozoites at concentrations below 220 µM. Since there is no fully annotated genome or proteome of B. mandrillaris, we sequenced and assembled its transcriptome from a high-throughput RNA-sequencing (RNA-Seq) experiment and located the coding sequences of the genes potentially targeted by the growth inhibitors from our compound screens. We determined the sequence of 17 of these target genes and obtained expression clones for 15 that we validated by direct sequencing. These will be used in the future in combination with the identified hits in structure guided drug discovery campaigns to develop new approaches for the treatment of Balamuthia infections.
Topics: Acanthamoeba; Amebiasis; Amoeba; Balamuthia mandrillaris; Base Sequence; Brain; Drug Design; Drug Discovery; Encephalitis; Gene Expression; Naegleria fowleri; Transcriptome; Trophozoites
PubMed: 34737367
DOI: 10.1038/s41598-021-99903-8 -
Journal of Fish Diseases Sep 2023Nodular gill disease (NGD) is an infectious condition characterized by proliferative gill lesions leading to respiratory problems, oxygen deficiency and mortality in...
Nodular gill disease (NGD) is an infectious condition characterized by proliferative gill lesions leading to respiratory problems, oxygen deficiency and mortality in fish. Globally, NGD primarily impacts freshwater salmonids in intensive aquaculture systems. In recent years, numerous outbreaks of severe gill disease have affected more than half of the larger rainbow trout (Oncorhynchus mykiss) farms in Switzerland, mainly during spring and early summer. Mortality has reached up to 50% in cases where no treatment was administered. Freshwater amoeba are the presumed aetiologic agent of NGD. The gross gill score (GS) categorising severity of gill pathology is a valuable first-line diagnostic tool aiding fish farmers in identifying and quantifying amoebic gill disease (AGD) in farmed marine salmonids. In this study, the GS was adapted to the NGD outbreak in farmed trout in Switzerland. In addition to scoring disease severity, gill swabs from NGD-affected rainbow trout were sampled and amoeba were cultured from these swabs. Morphologic and molecular methods identified six amoeba strains: Cochliopodium sp., Naegleria sp., Vannella sp., Ripella sp., Saccamoeba sp. and Mycamoeba sp. However, the importance of the different amoeba species for the onset and progression of NGD still has to be evaluated. This paper presents the first description of NGD with associated amoeba infection in farmed rainbow trout in Switzerland.
Topics: Animals; Amoeba; Oncorhynchus mykiss; Gills; Switzerland; Fish Diseases; Aquaculture
PubMed: 37294659
DOI: 10.1111/jfd.13819 -
ACS Biomaterials Science & Engineering Feb 2020In many situations, cells migrate through tiny orifices. Examples include the extravasation of immune cells from the bloodstream for fighting infections, the...
In many situations, cells migrate through tiny orifices. Examples include the extravasation of immune cells from the bloodstream for fighting infections, the infiltration of cancer cells during metastasis, and the migration of human pathogens. An extremely motile and medically relevant type of human pathogen is . In the study presented here, we investigated how a combination of microparticles and microstructured interfaces controls the migration of trophozoites. The microinterfaces comprised well-defined micropillar arrays, and the trophozoites easily migrated through the given constrictions by adapting the shape and size of their intracellular vacuoles and by adapting intracellular motion. After feeding the trophozoite cells in microinterfaces with synthetic, stiff microparticles of various sizes and shapes, their behavior changed drastically: if the particles were smaller than the micropillar gap, migration was still possible. If the cells incorporated particles larger than the pillar gap, they could become immobilized but could also display remarkable problem-solving capabilities. For example, they turned rod-shaped microparticles such that their short axis fit through the pillar gap or they transported the particles above the structure. As migration is a crucial contribution to pathogenicity and is also relevant to other biological processes in microenvironments, such as cancer metastasis, our results provide an interesting strategy for controlling the migration of cells containing intracellular particles by microstructured interfaces that serve as migration-limiting environments.
Topics: Acanthamoeba castellanii; Amoeba; Animals; Humans; Trophozoites
PubMed: 32215319
DOI: 10.1021/acsbiomaterials.9b00496 -
Journal of Chemical Information and... May 2023A grand challenge of computational biophysics is accurate prediction of interactions between molecules. Molecular dynamics (MD) simulations have recently gained much...
A grand challenge of computational biophysics is accurate prediction of interactions between molecules. Molecular dynamics (MD) simulations have recently gained much interest as a tool to directly compute rigorous intermolecular binding affinities. The choice of a fixed point-charge or polarizable multipole force field used in MD is a topic of ongoing discussion. To compare alternative methods, we participated in the SAMPL7 and SAMPL8 Gibb octaacid host-guest challenges to assess the Atomic Multipole Optimized Energetics for Biomolecular Applications (AMOEBA) polarizable multipole force field. Advantages of AMOEBA over fixed charge models include improved representation of molecular electrostatic potentials and better description of water occupying the unligated host cavity. Prospective predictions for 26 host-guest systems exhibit a mean unsigned error experiment of 0.848 kcal/mol across all absolute binding free energies, demonstrating excellent agreement between computational and experimental results. In addition, we explore two topics related to the inclusion of ions in MD simulations: use of a neutral co-alchemical protocol and the effect of salt concentration on binding affinity. Use of the co-alchemical method minimally affects computed energies, but salt concentration significantly perturbs our binding results. Higher salt concentration strengthens binding through classical charge screening. In particular, added Na ions screen negatively charged carboxylate groups near the binding cavity, thereby diminishing repulsive coulomb interactions with negatively charged guests. Overall, the AMOEBA results demonstrate the accuracy available through a force field providing a detailed energetic description of the four octaacid hosts and 13 charged organic guests. Use of the AMOEBA polarizable atomic multipole force field in conjunction with an alchemical free energy protocol can achieve chemical accuracy in application to realistic molecular systems.
Topics: Thermodynamics; Amoeba; Prospective Studies; Molecular Dynamics Simulation; Sodium Chloride; Water
PubMed: 37075788
DOI: 10.1021/acs.jcim.3c00155 -
Frontiers in Cellular and Infection... 2017Infections with filamentous fungi are common to all animals, but attention is rising especially due to the increasing incidence and high mortality rates observed in... (Review)
Review
Infections with filamentous fungi are common to all animals, but attention is rising especially due to the increasing incidence and high mortality rates observed in immunocompromised human individuals. Here, and other members of its genus are the leading causative agents. Attributes like their saprophytic life-style in various ecological niches coupled with nutritional flexibility and a broad host range have fostered the hypothesis that environmental predators could have been the actual target for some of their virulence determinants. In this mini review, we have merged the recent findings focused on the potential dual-use of fungal defense strategies against innate immune cells and soil amoebae as natural phagocytes. Well-established virulence attributes like the melanized surface of fungal conidia or their capacity to produce toxic secondary metabolites have also been found to be protective against the model amoeba . Some of the recent advances during interaction studies with human cells have further promoted the adaptation of other amoeba infection models, including the wide-spread generalist , or less prominent representatives like . We further highlight prospects and limits of these natural phagocyte models with regard to the infection biology of filamentous fungi and in comparison to the phagocytes of the innate immune system.
Topics: Acanthamoeba castellanii; Amoeba; Animals; Aspergillus fumigatus; Biological Evolution; Dictyostelium; Disease Models, Animal; Fungi; Host-Pathogen Interactions; Immunity, Innate; Phagocytes; Phagocytosis; Soil; Spores, Fungal; Virulence; Virulence Factors
PubMed: 29259922
DOI: 10.3389/fcimb.2017.00497 -
ACS Omega Mar 2023Primary amoebic meningoencephalitis and granulomatous amoebic encephalitis are distressing infections of the central nervous system caused by brain-eating amoebae,...
Primary amoebic meningoencephalitis and granulomatous amoebic encephalitis are distressing infections of the central nervous system caused by brain-eating amoebae, namely, and spp., respectively, and present mortality rates of over 90%. No single drug has been approved for use against these infections, and current therapy is met with an array of obstacles including high toxicity and limited specificity. Thus, the development of alternative effective chemotherapeutic agents for the management of infections due to brain-eating amoebae is a crucial requirement to avert future mortalities. In this paper, we synthesized a conducting polymer-based nanocomposite entailing polyaniline (PANI) and molybdenum disulfide (MoS) and explored its anti-trophozoite and anti-cyst potentials against and . The intracellular generation of reactive oxygen species (ROS) and ultrastructural appearances of amoeba were also evaluated with treatment. Throughout, treatment with the 1:2 and 1:5 ratios of PANI/MoS at 100 μg/mL demonstrated significant anti-amoebic effects toward as well as , appraised to be ROS mediated and effectuate physical alterations to amoeba morphology. Further, cytocompatibility toward human keratinocyte skin cells (HaCaT) and primary human corneal epithelial cells (pHCEC) was noted. For the first time, polymer-based nanocomposites such as PANI/MoS are reported in this study as appealing options in the drug discovery for brain-eating amoebae infections.
PubMed: 36910978
DOI: 10.1021/acsomega.2c06050 -
PLoS Pathogens Nov 2023The "Amoeboid Predator-Fungal Animal Virulence Hypothesis" posits that interactions with environmental phagocytes shape the evolution of virulence traits in fungal...
The "Amoeboid Predator-Fungal Animal Virulence Hypothesis" posits that interactions with environmental phagocytes shape the evolution of virulence traits in fungal pathogens. In this hypothesis, selection to avoid predation by amoeba inadvertently selects for traits that contribute to fungal escape from phagocytic immune cells. Here, we investigate this hypothesis in the human fungal pathogens Cryptococcus neoformans and Cryptococcus deneoformans. Applying quantitative trait locus (QTL) mapping and comparative genomics, we discovered a cross-species QTL region that is responsible for variation in resistance to amoeba predation. In C. neoformans, this same QTL was found to have pleiotropic effects on melanization, an established virulence factor. Through fine mapping and population genomic comparisons, we identified the gene encoding the transcription factor Bzp4 that underlies this pleiotropic QTL and we show that decreased expression of this gene reduces melanization and increases susceptibility to amoeba predation. Despite the joint effects of BZP4 on amoeba resistance and melanin production, we find no relationship between BZP4 genotype and escape from macrophages or virulence in murine models of disease. Our findings provide new perspectives on how microbial ecology shapes the genetic architecture of fungal virulence, and suggests the need for more nuanced models for the evolution of pathogenesis that account for the complexities of both microbe-microbe and microbe-host interactions.
Topics: Animals; Humans; Mice; Amoeba; Metagenomics; Predatory Behavior; Cryptococcus neoformans; Cryptococcosis
PubMed: 37956179
DOI: 10.1371/journal.ppat.1011763 -
International Journal of Hygiene and... May 2024Free living amoeba (FLA) are among the organisms commonly found in wastewater and are well-established hosts for diverse microbial communities. Despite its clinical...
Free living amoeba (FLA) are among the organisms commonly found in wastewater and are well-established hosts for diverse microbial communities. Despite its clinical significance, there is little knowledge on the FLA microbiome and resistome, with previous studies relying mostly on conventional approaches. In this study we comprehensively analyzed the microbiome, antibiotic resistome and virulence factors (VFs) within FLA isolated from final treated effluents of two wastewater treatment plants (WWTPs) using shotgun metagenomics. Acanthamoeba has been identified as the most common FLA, followed by Entamoeba. The bacterial diversity showed no significant difference (p > 0.05) in FLA microbiomes obtained from the two WWTPs. At phylum level, the most dominant taxa were Proteobacteria, followed by Firmicutes and Actinobacteria. The most abundant genera identified were Enterobacter followed by Citrobacter, Paenibacillus, and Cupriavidus. The latter three genera are reported here for the first time in Acanthamoeba. In total, we identified 43 types of ARG conferring resistance to cephalosporins, phenicol, streptomycin, trimethoprim, quinolones, cephalosporins, tigecycline, rifamycin, and kanamycin. Similarly, a variety of VFs in FLA metagenomes were detected which included flagellar proteins, Type IV pili twitching motility proteins (pilH and rpoN), alginate biosynthesis genes AlgI, AlgG, AlgD and AlgW and Type VI secretion system proteins and general secretion pathway proteins (tssM, tssA, tssL, tssK, tssJ, fha, tssG, tssF, tssC and tssB, gspC, gspE, gspD, gspF, gspG, gspH, gspI, gspJ, gspK, and gspM). To the best of our knowledge, this is the first study of its kind to examine both the microbiomes and resistome in FLA, as well as their potential pathogenicity in treated effluents. Additionally, this study showed that FLA can host a variety of potentially pathogenic bacteria including Paenibacillus, and Cupriavidus that had not previously been reported, indicating that their relationship may play a role in the spread and persistence of antibiotic resistant bacteria (ARBs) and antibiotic resistance genes (ARGs) as well as the evolution of novel pathogens.
Topics: Wastewater; Anti-Bacterial Agents; Amoeba; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Microbiota; Bacteria; Genes, Bacterial; Drug Resistance, Microbial; Cephalosporins
PubMed: 38471337
DOI: 10.1016/j.ijheh.2024.114345 -
The Journal of Eukaryotic Microbiology Jan 2018Class Ascetosporea (Rhizaria; Endomyxa) comprises many parasites of invertebrates. Within this group, recent group-specific environmental DNA (eDNA) studies have...
Class Ascetosporea (Rhizaria; Endomyxa) comprises many parasites of invertebrates. Within this group, recent group-specific environmental DNA (eDNA) studies have contributed to the establishment of the new order Mikrocytida, a new phylogeny and characterization of Paramyxida, and illuminated the diversity and distribution of haplosporidians. Here, we use general and lineage-specific PCR primers to investigate the phylogenetic "gap" between haplosporidians and their closest known free-living relatives, the testate amoeba Gromia and reticulate amoeba Filoreta. Within this gap are Paradinium spp. parasites of copepods, which we show to be highly diverse and widely distributed in planktonic and benthic samples. We reveal a robustly supported radiation of parasites, ENDO-3, comprised of Paradinium and three further clades (ENDO-3a, ENDO-3b and SPP). A further environmental group, ENDO-2, perhaps comprising several clades, branches between this radiation and the free-living amoebae. Early diverging haplosporidians were also amplified, often associated with bivalves or deep-sea samples. The general primer approach amplified an overlapping set of novel lineages within ENDO-3 and Haplosporida, whereas the group-specific primer strategy, targeted to amplify from the earliest known divergent haplosporidians to Gromia, generated greater sequence diversity across part of this phylogenetic range.
PubMed: 29336517
DOI: 10.1111/jeu.12501 -
PeerJ 2022Curcumin is an active compound derived from turmeric, , and is known for its benefits to human health. The amoebicidal activity of curcumin against was recently...
BACKGROUND
Curcumin is an active compound derived from turmeric, , and is known for its benefits to human health. The amoebicidal activity of curcumin against was recently discovered. However, a physiological change of intracellular pathways related to encystation mechanism, including autophagy in the surviving amoeba after curcumin treatment, has never been reported. This study aims to investigate the effect of curcumin on the survival of under nutrient starvation and nutrient-rich condition, as well as to evaluate the encystation and a physiological change of autophagy at the mRNA level.
METHODS
In this study, amoebas were treated with a sublethal dose of curcumin under nutrient starvation and nutrient-rich condition and the surviving amoebas was investigated. Cysts formation and vacuolization were examined by microscopy and transcriptional expression of autophagy-related genes and other encystation-related genes were evaluated by real-time PCR.
RESULTS
cysts were formed under nutrient starvation. However, in the presence of the autophagy inhibitor, 3-methyladenine (3-MA), the percentage of cysts was significantly reduced. Interestingly, in the presence of curcumin, most of the parasites remained in the trophozoite stage in both the starvation and nutrient-rich condition. In vacuolization analysis, the percentage of amoebas with enlarged vacuole was increased upon starvation. However, the percentage was significantly declined in the presence of curcumin and 3-MA. Molecular analysis of autophagy-related (ATG) genes showed that the mRNA expression of the ATG genes, ATG3, ATG8b, ATG12, ATG16, under the starvation with curcumin was at a basal level along the treatment. The results were similar to those of the curcumin-treated amoebas under a nutrient-rich condition, except ATG16 which increased later. On the other hand, mRNA expression of encystation-related genes, cellulose synthase and serine proteinase, remained unchanged during the first 18 h, but significantly increased at 24 h post treatment.
CONCLUSION
Curcumin inhibits cyst formation in surviving trophozoites, which may result from its effect on mRNA expression of key ATG-related genes. However, further investigation into the mechanism of curcumin in trophozoites arrest and its association with autophagy or other encystation-related pathways is needed to support the future use of curcumin.
Topics: Animals; Humans; Acanthamoeba; Curcumin; Trophozoites; Amoeba
PubMed: 35811814
DOI: 10.7717/peerj.13657