Two protists isolated simultaneously from the same sample of gill tissue of Psetta maxima (L.) were identified as Thecamoeba hilla Schaeffer, 1926 and Labyrinthula sp. A Labyrinthula strain (LTH) derived from a mixed culture of both organisms was well established in a short time, while subcultures of T. hilla continued to be associated with Labyrinthula cells despite all efforts to eliminate them. Ultrastructural examination, repeated several times in the course of long-lasting subculturing of amoebae, revealed that trophozoites of T. hilla host in their cytoplasm multiplying labyrinthulid cells. Comparison of SSU rDNA sequences of the Labyrinthula strain LTH and those from labyrinthulid endosymbionts from T. hilla verified the assumption that the extra- and intra-cellularly multiplying Labyrinthula cells are identical organisms. The association of the marine amoeba T. hilla and Labyrinthula sp. displayed signs of mutualistic symbiosis.
- MeSH
- Amoeba genetics physiology ultrastructure MeSH
- Phylogeny MeSH
- Symbiosis * MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
In brain sections of the Naegleria-caused cases of primary amoebic meningoencephalitis, extensive demyelinization was found in the white matter, besides the severe histopathological changes and large clusters of trophozoites in the grey matter. The myelinoclasis appeared to be a result of a specific phospholipolytic effect, unlike that in post-viral encephalomyelitis, which has been attributed to vascular blockade or hemorrhages. In monkey kidney cell cultures a very early cytopathic effect was observed and traced to the cytolytic property of the seeding culture fluid. Rat brain slices inoculated with Naegleria culture exhibited amoebic growth and demyelinization in 28-52 hours incubation at 35 degrees C. In a chemically defined medium containing sphingomyelin, casein and glucose, the Naegleria produced a limited growth parallelling the clearance of the lipid turbidity during a 72 hour incubation at 35 degrees C. Chromatographic analysis of the turbidity-cleared cultures revealed decomposition of sphingomyeline with liberation of choline, sphingosine and fatty acids. It is, hence, concluded that the pathogenicity of cytopathic effect of pathogenic Naegleria can be attributed to the latter's capacity to liberate a phospholipolytic enzyme or factor during active growth, which "makes holes" in the lipid-rich cytoplasmic membrane of cells as well as demyelinizes nerve tissue.
- MeSH
- Amebiasis pathology MeSH
- Amoeba growth & development metabolism pathogenicity MeSH
- Species Specificity MeSH
- Haplorhini MeSH
- Rats MeSH
- Culture Techniques MeSH
- Kidney MeSH
- Humans MeSH
- Meningoencephalitis pathology MeSH
- Brain pathology MeSH
- Sphingomyelins metabolism MeSH
- Animals MeSH
- Check Tag
- Rats MeSH
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Sphingomyelins MeSH
Copper is a trace metal that is necessary for all organisms but toxic when present in excess. Different mechanisms to avoid copper toxicity have been reported to date in pathogenic organisms such as Cryptococcus neoformans and Candida albicans. However, little if anything is known about pathogenic protozoans despite their importance in human and veterinary medicine. Naegleria fowleri is a free-living amoeba that occurs naturally in warm fresh water and can cause a rapid and deadly brain infection called primary amoebic meningoencephalitis (PAM). Here, we describe the mechanisms employed by N. fowleri to tolerate high copper concentrations, which include various strategies such as copper efflux mediated by a copper-translocating ATPase and upregulation of the expression of antioxidant enzymes and obscure hemerythrin-like and protoglobin-like proteins. The combination of different mechanisms efficiently protects the cell and ensures its high copper tolerance, which can be advantageous both in the natural environment and in the host. Nevertheless, we demonstrate that copper ionophores are potent antiamoebic agents; thus, copper metabolism may be considered a therapeutic target.
- Keywords
- Copper, Copper-translocating ATPase, Hemerythrin, Ionophores, Naegleria fowleri, Oxidative stress,
- MeSH
- Adenosine Triphosphatases metabolism MeSH
- Amoeba MeSH
- Antioxidants physiology MeSH
- Humans MeSH
- Copper metabolism MeSH
- Brain MeSH
- Naegleria fowleri * physiology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Adenosine Triphosphatases MeSH
- Antioxidants MeSH
- Copper MeSH
We present a fascinating triad relationship between a eukaryotic amoeba and its two bacterial symbionts. The morphological characteristics of the amoeba allowed for a confident assignment to the genus Nuclearia (Opisthokonta, Nucleariidae), but species identification resulted in an ambiguous result. Sequence analysis indicated an affiliation to the species N. thermophila, however, several morphological features contradict the original description. Amoebal isolates were cultured for several years with their preferred food source, the microcystin-producing harmful cyanobacterium Planktothrix rubescens. Symbioses of the amoeba with ecto- and endosymbiotic bacteria were maintained over this period. Several thousand cells of the ectosymbiont are regularly arranged inside a layer of extracellular polymeric substances produced by the amoeba. The ectosymbiont was identified as Paucibacter toxinivorans (Betaproteobacteria), which was originally isolated by enrichment with microcystins. We found indications that our isolated ectosymbiont indeed contributed to toxin-degradation. The endosymbiont (Gammaproteobacteria, 15-20 bacteria per amoeba) is enclosed in symbiosomes inside the host cytoplasm and represents probably an obligate symbiont. We propose the name "Candidatus Endonucleariobacter rarus" for this bacterium that was neither found free-living nor in a symbiotic association. Nucleariidae are uniquely suited model organisms to study the basic principles of symbioses between opisthokonts and prokaryotes.
- Keywords
- Bacteria-protist symbioses, Nucleariidae, Paucibacter toxinivorans., ectosymbionts, endosymbionts, feeding,
- MeSH
- Amoeba classification cytology isolation & purification microbiology MeSH
- Betaproteobacteria classification isolation & purification physiology MeSH
- DNA, Bacterial chemistry genetics MeSH
- Gammaproteobacteria classification isolation & purification physiology MeSH
- Genes, rRNA MeSH
- Lakes parasitology MeSH
- DNA, Ribosomal Spacer chemistry genetics MeSH
- Molecular Sequence Data MeSH
- DNA, Protozoan chemistry genetics MeSH
- DNA, Ribosomal chemistry genetics MeSH
- RNA, Protozoan genetics MeSH
- RNA, Ribosomal, 16S genetics MeSH
- RNA, Ribosomal, 18S genetics MeSH
- Sequence Analysis, DNA MeSH
- Symbiosis * MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Switzerland MeSH
- Names of Substances
- DNA, Bacterial MeSH
- DNA, Ribosomal Spacer MeSH
- DNA, Protozoan MeSH
- DNA, Ribosomal MeSH
- RNA, Protozoan MeSH
- RNA, Ribosomal, 16S MeSH
- RNA, Ribosomal, 18S MeSH
BACKGROUND: Mitochondria and peroxisomes are the two organelles that are most affected during adaptation to microoxic or anoxic environments. Mitochondria are known to transform into anaerobic mitochondria, hydrogenosomes, mitosomes, and various transition stages in between, collectively called mitochondrion-related organelles (MROs), which vary in enzymatic capacity. Anaerobic peroxisomes were identified only recently, and their putatively most conserved function seems to be the metabolism of inositol. The group Archamoebae includes anaerobes bearing both anaerobic peroxisomes and MROs, specifically hydrogenosomes in free-living Mastigamoeba balamuthi and mitosomes in the human pathogen Entamoeba histolytica, while the organelles within the third lineage represented by Pelomyxa remain uncharacterized. RESULTS: We generated high-quality genome and transcriptome drafts from Pelomyxa schiedti using single-cell omics. These data provided clear evidence for anaerobic derivates of mitochondria and peroxisomes in this species, and corresponding vesicles were tentatively identified in electron micrographs. In silico reconstructed MRO metabolism harbors respiratory complex II, electron-transferring flavoprotein, a partial TCA cycle running presumably in the reductive direction, pyruvate:ferredoxin oxidoreductase, [FeFe]-hydrogenases, a glycine cleavage system, a sulfate activation pathway, and an expanded set of NIF enzymes for iron-sulfur cluster assembly. When expressed in the heterologous system of yeast, some of these candidates localized into mitochondria, supporting their involvement in the MRO metabolism. The putative functions of P. schiedti peroxisomes could be pyridoxal 5'-phosphate biosynthesis, amino acid and carbohydrate metabolism, and hydrolase activities. Unexpectedly, out of 67 predicted peroxisomal enzymes, only four were also reported in M. balamuthi, namely peroxisomal processing peptidase, nudix hydrolase, inositol 2-dehydrogenase, and D-lactate dehydrogenase. Localizations in yeast corroborated peroxisomal functions of the latter two. CONCLUSIONS: This study revealed the presence and partially annotated the function of anaerobic derivates of mitochondria and peroxisomes in P. schiedti using single-cell genomics, localizations in yeast heterologous systems, and transmission electron microscopy. The MRO metabolism resembles that of M. balamuthi and most likely reflects the state in the common ancestor of Archamoebae. The peroxisomal metabolism is strikingly richer in P. schiedti. The presence of myo-inositol 2-dehydrogenase in the predicted peroxisomal proteome corroborates the situation in other Archamoebae, but future experimental evidence is needed to verify additional functions of this organelle.
- Keywords
- Anaerobic peroxisome, Anaerobiosis, FeS cluster assembly, Hydrogenosome, Mitochondrion-related organelle, Pelomyxa, Single-cell genomics,
- MeSH
- Amoeba * genetics metabolism MeSH
- Anaerobiosis MeSH
- Archamoebae * genetics metabolism MeSH
- Genomics MeSH
- Humans MeSH
- Mitochondria metabolism MeSH
- Peroxisomes metabolism MeSH
- Saccharomyces cerevisiae MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- MeSH
- Amebiasis diagnosis MeSH
- Amoeba isolation & purification MeSH
- Adult MeSH
- Humans MeSH
- Water Microbiology MeSH
- Nasal Mucosa microbiology MeSH
- Carrier State diagnosis MeSH
- Military Medicine MeSH
- Environmental Pollution MeSH
- Check Tag
- Adult MeSH
- Humans MeSH
- Male MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czechoslovakia MeSH
A new species of amphizoic amoeba, Nuclearia pattersoni sp. n., isolated from gills of Rutilus rutilus L. is described. It is characterised by elongate flattened trophozoites of irregular shape. The longer dimension of their bodies is 13.2 (11.0-15.7) microm. Filopodia radiating mostly from the poles are 2 to 2.5 times longer than the body. The diameter of less frequently observed spherical trophozoites is 8.2-10.8 microm; their filopodia radiate to all directions. Cyst-like stages have shorter pseudopodia that arise from one pole only. The surface of locomotive forms from agar plate cultures has a thin amorphous glycocalyx, while most cells are covered by two layers of extracellular matrix. Mitochondria have flattened cristae, dictyosomes are located in the perinuclear zone. A conspicuous ultrastructural feature of the morphologically similar N. simplex, perinuclear striated band, is not present. Light microscopic and ultrastructural data are completed with the sequence of SSU rRNA gene and phylogenetic analysis including sequences of related taxa. The bacterial endosymbiont found in N. pattersoni type strain RR2G2 is assigned to the genus Rickettsia.
- MeSH
- Amoeba classification genetics isolation & purification ultrastructure MeSH
- Cyprinidae microbiology parasitology MeSH
- DNA, Bacterial genetics MeSH
- DNA genetics MeSH
- Microscopy, Electron MeSH
- Phylogeny MeSH
- Rickettsia classification genetics isolation & purification MeSH
- Base Sequence MeSH
- Symbiosis MeSH
- Gills microbiology parasitology MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- DNA, Bacterial MeSH
- DNA MeSH
- MeSH
- Amoeba pathogenicity MeSH
- Humans MeSH
- Meningoencephalitis epidemiology etiology mortality therapy MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Czechoslovakia MeSH
- MeSH
- Amebiasis * MeSH
- Amoeba pathogenicity MeSH
- Adult MeSH
- Culture Media MeSH
- Humans MeSH
- Meningoencephalitis etiology MeSH
- Nasal Mucosa microbiology MeSH
- Rhinitis etiology MeSH
- Check Tag
- Adult MeSH
- Humans MeSH
- Male MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Culture Media MeSH
Nodular gill disease (NGD) is an emerging condition associated with amoeba trophozoites in freshwater salmonid farms. However, unambiguous identification of the pathogens still must be achieved. This study aimed to identify the amoeba species involved in periodic NGD outbreaks in two rainbow trout (Oncorhynchus mykiss) farms in Northeastern Italy. During four episodes (February-April 2023), 88 fish were euthanized, and their gills were evaluated by macroscopic, microscopic and histopathological examination. The macroscopic and microscopic severity of the lesions and the degree of amoebae infestation were scored and statistically evaluated. One gill arch from each animal was put on non-nutrient agar (NNA) Petri dishes for amoeba isolation, cultivation and subsequent identification with SSU rDNA sequencing. Histopathology confirmed moderate to severe lesions consistent with NGD and mild to moderate amoeba infestation. The presence of amoebae was significantly correlated with lesion severity. Light microscopy of cultured amoebae strains and SSU rDNA analysis revealed the presence of a previously characterized amoeba Naegleria sp. strain GERK and several new strains: two strains from Hartmannelidae, three vannelid amoebae from the genus Ripella and cercozoan amoeba Rosculus. Despite the uncertainty in NGD etiopathogenesis and amoebae pathogenic role, identifying known and new amoebae leans towards a possible multi-aetiological origin.
- Keywords
- Oncorhynchus mykiss, Italy, SSU rDNA, amphizoic amoeba, nodular gill disease (NGD), rainbow trout,
- MeSH
- Amebiasis * veterinary parasitology MeSH
- Amoeba genetics isolation & purification classification MeSH
- Amoebozoa genetics isolation & purification classification physiology MeSH
- Phylogeny MeSH
- Fish Diseases * parasitology epidemiology MeSH
- Oncorhynchus mykiss * parasitology MeSH
- Aquaculture MeSH
- Gills * parasitology pathology MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Geographicals
- Italy MeSH
