Meningoencephalitis due to Hartmannella (Acanthamoeba).
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Amebiasis↗
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Hartmannella (Acanthamoeba) experiments in preservation and detection of trophozoites and amebic cysts in infected tissue.
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Amoeba↗
[Research on Hartmannella castellanii. I. Cytochemical study on polysaccharides].
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Amoeba↗
18S ribosomal RNA gene sequences of Cochliopodium (Himatismenida) and the phylogeny of Amoebozoa.
Cochliopodium is a very distinctive genus of discoid amoebae covered by a dorsal tectum of carbohydrate microscales. Its phylogenetic position is unclear, since although sharing many features with naked "gymnamoebae", the tectum sets it apart. We sequenced 18S ribosomal RNA genes from three Cochliopodium species (minus, spiniferum and Cochliopodium sp., a new species resembling C. minutum). Phylogenetic analysis shows Cochliopodium as robustly holophyletic and within Amoebozoa, in full accord with morphological data. Cochliopodium is always one of the basal branches within Amoebozoa but its precise position is unstable. In Bayesian analysis it is sister to holophyletic Glycostylida, but distance trees mostly place it between Dermamoeba and a possibly artifactual long-branch cluster including Thecamoeba. These positions are poorly supported and basal amoebozoan branching ill-resolved, making it unclear whether Discosea (Glycostylida, Himatismenida, Dermamoebida) is holophyletic; however, Thecamoeba seems not specifically related to Dermamoeba. We also sequenced the small-subunit rRNA gene of Vannella persistens, which constantly grouped with other Vannella species, and two Hartmannella strains. Our trees suggest that Vexilliferidae, Variosea and Hartmannella are polyphyletic, confirming the existence of two very distinct Hartmannella clades: that comprising H. cantabrigiensis and another divergent species is sister to Glaeseria, whilst Hartmannella vermiformis branches more deeply.
Animals↗
Comparison of free-living amoebae in hot water systems of hospitals with isolates from moist sanitary areas by identifying genera and determining temperature tolerance.
Legionella-contaminated hot water systems and moist sanitary areas in six hospitals were sampled for amoebae by following a standardized collection protocol. Genus identifications and temperature tolerance determinations were made. Amoebae identified as Hartmannella vermiformis (65%), Echinamoebae spp. (15%), Saccamoebae spp. (12%), and Vahlkampfia spp. (9%) were detected in 29 of 56 (52%) hot water samples. Twenty-three of 49 (47%) swabs obtained from moist areas were amoeba positive. The following genera were identified: Acanthamoeba (22%), Naegleria (22%), Vahlkampfia (20%), Hartmannella (15%), and Vanella (7%). The temperature tolerance of amoebae from hot water systems was strikingly different from that of amoebae from moist areas. At 44 degrees C on agar, 59% of amoebic isolates sampled from hot water systems showed growth. The corresponding value for isolates from moist areas was only 17%. Six Acanthamoeba isolates from the moist areas were considered potential pathogens. Four Hartmannella and two Saccamoeba isolates from hot water could be cultured at 53 degrees C.
Acanthamoeba↗
Electrophoretic characterization of small free-living Amoebae.
Disc electrophoresis was employed in an attempt to determine its usefulness as a means of differentiating and classifying some free-living species of amoebae. The Rp values and the percentage of similarity of the protein bands were used as taxonomic criteria. Some species of the genera Hartmanella and Naegleria were examined. The protein patterns of all species studied were highly reproducible. Four strains of Hartmannella castellanii showed an absolute identity to the original H. castellanii strain. One strain isolated by Neff and H. rhysodes were similar to all the other strains although their protein patten were not completely identical. On the other hand, H. culbertsoni, H. astronyxis and H. lammellipodia were different from all the other examined species of Hartmannella. Pathogenic N. fowleri-strains were identical to each other and were different in their protein patterns from non-pathogenic N. gruberi strain. The similarities and differences of protein bands of strains of ameobae and their correlations with other attributes were discussed. The results have shown that the disc electrophoresis appears useful in differentiating between species of Hartmannella and Naegleria.
Amoeba↗
Detection of novel chlamydiae in cats with ocular disease.
OBJECTIVE: To detect and characterize the full range of chlamydial infections in cats with ocular disease by use of polymerase chain reaction (PCR) assays, cytologic examination, immunohistochemical analysis, and evaluation of clinical information including status for feline herpesvirus-1 (FeHV-1). SAMPLE POPULATION: DNA extracted from 226 conjunctival samples obtained from cats with clinically diagnosed keratitis or conjunctivitis and 30 conjunctival samples from healthy cats. PROCEDURE: PCR assays for the 16S rRNA gene specific for the order Chlamydiales and a new Chlamydophila felis (formerly Chlamydia psittaci) species-specific 23S rRNA gene were performed. Seventy-four conjunctival samples were prepared with Romanowsky-type stain, grouped on the basis of inflammatory pattern, and screened for chlamydial inclusions by use of immunohistochemical analysis. Clinical information and FeHV-1 status were recorded. RESULTS: 26 (12%) specimens had positive results for the only known feline chlamydial pathogen, C felis. Surprisingly, an additional 88 (39%) were positive for non-C felis chlamydial DNA. Identification of non-C felis chlamydial DNA by direct sequencing revealed 16S rRNA gene sequences that were 99% homologous to the sequence for Neochlamydia hartmannellae, an amebic endosymbiont. Chlamydial prevalence was significantly higher in cats with ocular disease. CONCLUSIONS AND CLINICAL RELEVANCE: Application of a broad-range detection method resulted in identification of a new agent associated with ocular disease in cats. Finding chlamydia-like agents such as N hartmannellae in coinfections with their obligate amebic host, Hartmannella vermiformis, raises questions about the potential role of these microorganisms in causation or exacerbation of ocular disease in cats.
Animals↗
Flagella are a positive predictor for virulence in Legionella.
Pathogenesis of Legionnaires>> disease is strictly related to the ability of the legionellae to infect phagocytic cells, yet surface markers of virulence in Legionella isolates are currently unknown. Rabbit antibodies raised against purified flagella of Legionella pneumophila serogroup 1 recognized a total of 24 of 30 laboratory-maintained isolates of L. pneumophila serogroups 1-15 and 16 of 24 other Legionella species tested by rapid immunoblot and indirect immunofluorescence assay. All isolates possessing flagella detectable with these anti-flagella antibodies, regardless of species, were capable of infecting Hartmannella vermiformis. Isolates lacking immunologic cross-reactivity were shown to lack purifiable flagella. The majority of aflagellate isolates were not motile and failed to multiply intracellularly in co-culture with Hartmannella vermiformis. Some isolates characterized as aflagellate when harvested from BCYE agar were able to multiply in amoebae, and flagella were subsequently detectable by immunologic methods. These data suggest that lack of immunologic recognition of flagella in laboratory-maintained isolates of Legionella is due to their attenuation and a corresponding loss of expression of flagella. More importantly, the presence of flagella can serve as a positive predictive marker for strain virulence and is useful in determining the virulence status of Legionella isolates.
Animals↗
The identification of free-living environmental isolates of amoebae from Bulgaria.
A survey was carried out in Bulgaria to determine the presence of free-living amoebae (FLA) from environmental sources. In 171 (61.1%) of 280 samples, isolates of Acanthamoeba with group II or III morphology, as well as Hartmannella spp. were recovered. Five isolates named "6" (artificial lake), Ep (lake), G2 (soil), R4* (river) and PK (spring water)--all exhibiting a highly efficient proliferation in axenic cultures--were subsequently cloned and subjected to molecular analyses for identification and genotyping In accordance with morphological findings, PCR-based analyses identified four isolates (6, Ep, G2, R4*) belonging to the genus Acanthamoeba. Confirmation of these findings was obtained by phylogenetic analysis using partial sequencing of the 18S rDNA (ASA.S1) Acanthamoeba-gene. Comparison of these sequences with corresponding regions from other Acanthamoeba strains available from GenBank sorted all four isolates into the sequence type group T4 that contains most of the pathogenic Acanthamoeba strains already identified. The fifth isolate (PK) exhibited morphological characteristics matching those of Hartmannella, and scored negative in the Naegleria fowleri and Acanthamoeba PCRs.
Acanthamoeba↗
The presence of free-living amoebae in portable and stationary eye wash stations.
Portable and stationary eye wash stations were tested for the presence of free-living amoebae. Such amoebae may be found in potable waters, and at least one genera, the Acanthamoebae, can cause severe infections when introduced into traumatized eyes. Concentrates of filtrates of water from eye wash stations were placed on nonnutrient agar plates seeded with Escherichia coli. Resultant outgrowths of free-living amoebae, which were morphologically identified as mixtures of Hartmannella and Acanthamoebae, were inoculated intranasally into weanling mice. Subsequently, brain and lung tissues from injected mice were tested for amoebae as an indication of persistent infection. Acanthamoebae and Hartmannella were detected in some eye wash stations at each of four test sites. Both portable and stationary stations harbored Acanthamoebae. Some of the isolates caused persistent pulmonary infection but were not isolated from brain tissue. Flushing stationary eye wash stations temporarily reduced the number of stations positive for amoebae. Treatment of portable stations with 25 ppm of free chlorine also reduced the number of stations harboring amoebae but caused corrosion in some of the stations.
Acanthamoeba↗
Comparative studies on related free-living and pathogenic amebae with special reference to Acanthamoeba.
Comparative studies were conducted on the structure, nutrition, protein composition, immunology, and effect on cell cultures of Acanthamoeba sp. (Lilly A-1 strain), A. castellanii (Singh and Neff strains), A. astronyxis, A. comandoni, A. polyphaga, A. terricola, Hartmannella vermiformis, and Naegleria gruberi. Lilly A-1 strain of Acanthamoeba received special attention owing to its pathogenicity for experimental animals. Distinct differences were noted in structure, nutrition, and antigenic composition of Acanthamoeba spp. and Hartmannella, and it was concluded that their recognition as separate genera is justified. With the exception of A. terricola, all species of Acanthamoeba could be differentiated by cyst structure. Cysts of A. terricola closely resembled those of A. castellanii Singh strain, and close antigenic relationships between these 2 species were demonstrated by gel diffusion and immunoelectrophoresis (IEP); it was concluded that the 2 amebae belong in the same species. The pathogenic Acanthamoeba sp, Lilly strain differed from the nonpathogenic A. castellanii Singh strain (a) cyst structure; (b) protein distribution patterns (on disc electrophoresis);(c)soluble and particulate antigens (on gel diffusion, IEP, complement fixation, and immobilization tests); (d) capacity to induce cell-free plaques and other cytopathic effect (CPE) in mammalian monlayer cell cultures; (e) elimination of a phospholipase, responsible for some of the CPE, into the culture medium. Acanthamoeba sp. Lilly strain, which liberated more phospholipase, produced more CPE. Acanthamoeba sp. Lilly strain differed also from other species of this genus in cyst structure and antigenic composition. It was concluded, therefore, that, following the recommendation of Singh & Das, it ought to be placed in a separate species, Acanthamoeba culbertsoni.
Amphotericin B↗
Identification of antigens of pathogenic free-living amoebae by protein immunoblotting with rabbit immune and human sera.
Prominent antigens of pathogenic and nonpathogenic free-living amoebae were identified by using polyclonal rabbit immune sera in immunoblot assays. The intent was to determine if prominent epitopes identified with rabbit immune sera could also be recognized by human sera. With rabbit sera, the development of immunoreactive bands was restricted to molecular masses of greater than 18.5 kDa for Naegleria, Hartmannella, and Vahlkampfia antigens. Two or more broad bands of less than 18.5 kDa were prominent features in three different Acanthamoeba species. Few cross-reactive antibodies could be detected between representative species of the three different subgroups of Acanthamoeba. Naegleria antigen was likewise serologically distinct, as were Hartmannella and Vahlkampfia antigens. The relative lack of cross-reacting antibodies between the pathogenic amoebae suggested that i would be desirable to use a panel of amoebic antigens to represent the range of serologically distinct antigens for assessing reactive antibodies in human sera. In pooled human sera (10 serum specimens per pool), the appearance of minimally reactive bands ranging from 32.5 to 106 kDa was a common feature of all six antigens. A prominent band of less than 18.5 kDa was identified in the Acanthamoeba culbertsoni antigen lane in 2 of the 10 human serum specimen pools. When sera from each of the two groups were tested individually by immunoblotting, the reaction with A. culbertsoni antigen could be associated with one individual. By using a panel of amoebic antigens, this method could prove useful in recognizing undiagnosed amoebic infections by revealing specific reactive antibodies.
Acanthamoeba↗
A brief survey of free-living amebae in Thailand and Hamamatsu District, Japan.
The aim of this study was to determine the presence of free-living amebae in aquatic habitats of human environments in Thailand and Hamamatsu district, Japan. Genus identification was based on the morphology of cyst and trophozoite forms and a flagellation test for genus Naegleria. The pathogenic potential was tested in mice by nasal instillation for genus Naegleria and Acanthameba. In 14 provinces of Thailand, amebae were isolated in 43 from 95 water samples and 67 from 120 soil swabs. Amebae of 49 isolates from waters were identified as Acanthameba (36.7%), Naegleria (28.6%), Hartmannella (20.4%), Vahlkampfia (12.2%) and Vannella (2%). Soil samples have significantly higher levels of Acanthameba and Hartmannella (p<0.05) but lower for Naegleria (p<0.05) and 7 unidentified amebae were found. In Hamamatsu district, Japan, 62 amebae of the same genera were isolated from 47 of 95 water samples. There were significantly higher levels of Acanthameba (22.6%) (p<0.05) but lower for Naegleria (4.8%) (p<0.05) than those of Thailand which each of them caused death in mice. Three unidentified amebae were isolated. This finding serves as additional evidence for the presence of free-living amebae under natural and the difference in distribution between tropic and subtropic areas.
Acanthamoeba↗
[Studies of limax-amoebae in a physiotherapeutical indoor-swimming-pool (author's transl)].
A physiotherapeutical indoor swimmingpool in a hospital was inspected for the presence of potentially pathogenic Limax-amoebae. In cultures inoculated with material from water samples and from cotton swabs taken from the bottom and walls of the pool the following species of Limax-amoebae could be cloned and identified: Naegleria gruberi, Hartmannella vermiformis, Acanthamoeba castellamii, A. polyphaga, A. palestinesis, Vannella mira, V. platypodia, Platyamoeba stenopodia, Echinamoebia exundans and Vexillifera bacilipedes. Two further strains of unidentified Acanthamoeba and 1 of the genus Vanella were observed. 3 to 5 species remained unidentified. Beyond this numerous monotrichous flagellates and Colpoda steini, a holotrichous ciliate, multiplied in water samples and agar plates. The concentration of the total number of pelagic amoebae increased from 10 to 20/liter one week after exchange of the pool-water to 100-1000/liter after 3 weeks and finally to more than 10(4) per liter after a period of 2 months. The isolated strains of Nageleria, Acanthamoeba and Hartmannella were tested for pathogenicity by means of the intranasal mice inoculation test. The clones of Naegleria were submitted to a temperature- and salt-tolerance test. All of the strains tested by these methods proved to be nonpathogenic.
Amoeba↗
Chronic amebic meningoencephalitis.
A patient had a five-year history of sleep disturbance that culminated in a confusional illness with convulsions from which he made a partial recovery. On three separate occasions, the same ameba (Hartmannella rhysodes) was seen in and cultured from the patient's CSF. There was a rising serum titer of immobilizing antibody against the isolated strain of the ameba. The diagnosis of chronic amebic meningoencephalitis was considered in this patient.
Adult↗
Heterogeneity in intracellular replication and cytopathogenicity of Legionella pneumophila and Legionella micdadei in mammalian and protozoan cells.
In contrast to Legionella pneumophila, little is known about the pathogenesis of other legionellae species that are capable of causing Legionnaires' disease. In this report, we contrast L. pneumophila and L. micdadei for their cytopathogenicity and intracellular replication within mammalian and protozoan cells. We show by transmission electron microscopy that L. micdadei replicates within an endoplasmic reticulum (RER)-free phagosome within human macrophages, alveolar epithelial cells, and within the protozoan Hartmannella vermiformis. In contrast, L. pneumophila replicates within a RER-surrounded phagosome within the same host cells. In contrast to replication of L. pneumophila within Acanthamoebae polyphaga, L. micdadei does not replicate within this protozoan host. Despite the prolific intracellular replication, L. micdadei is less cytopathogenic to all host cells than L. pneumophila. Since both species replicate intracellularly to a similar level, we have examined whether the reduced cytopathogenicity of L. micdadei is due to a reduced capacity to induce apoptosis or pore formation-mediated necrosis, both of which contribute to killing of the host cell by L. pneumophila. The data show that both species induced apoptosis-mediated killing of mammalian cells to a similar level. In contrast to L. pneumophila, expression of the pore-forming toxin by L. micdadei and its necrotic effect on macrophages and alveolar epithelial cells is undetectable. This has been further confirmed showing that L. micdadei is completely defective in contact-dependent haemolysis of RBCs, an activity mediated by the pore-forming toxin. Finally, in contrast to L. pneumophila, there was no significant intrapulmonary replication of L. micdadei in the A/J mice animal model. Our data show dramatic differences between L. pneumophila and L. micdadei in intracellular replication, cytopathogenicity, and infectivity to mammalian and protozoan cells.
Acanthamoeba↗
The Legionella pneumophila prp locus; required during infection of macrophages and amoebae.
Transposon mutagenesis was performed using mTn 10phoA to identify Legionella pneumophila genes that are expressed under certain in vitro conditions, and are required for intracellular replication. Of the 1653 PhoA fusions examined, 19 PhoA(+)fusion mutants were isolated and screened for differential expression of fusion proteins after growth at 30 or 37 degrees C, in the presence of low iron, or increased magnesium concentrations. The mutants were examined for their cytopathogenicity and intracellular replication within U937 macrophage-like cells and the protozoan Hartmannella vermiformis. One of the mutants generated, BS10, was defective in its multiplication within U937 macrophage-like cells and H. vermiformis. The defect in BS10 was complemented with a cosmid clone containing the wild type locus. The open reading frame interrupted by the insertion was homologous to prpD of Salmonella typhimurium and mmgE of Bacillus subtilis.
Alkaline Phosphatase↗
The frequency of eubacterium-to-eukaryote lateral gene transfers shows significant cross-taxa variation within amoebozoa.
Single-celled bacterivorous eukaryotes offer excellent test cases for evaluation of the frequency of prey-to-predator lateral gene transfer (LGT). Here we use analysis of expressed sequence tag (EST) data sets to quantify the extent of LGT from eubacteria to two amoebae, Acanthamoeba castellanii and Hartmannella vermiformis. Stringent screening for LGT proceeded in several steps intended to enrich for authentic events while at the same time minimizing the incidence of false positives due to factors such as limitations in database coverage and ancient paralogy. The results were compared with data obtained when the same methodology was applied to EST libraries from a number of other eukaryotic taxa. Significant differences in the extent of apparent eubacterium-to-eukaryote LGT were found between taxa. Our results indicate that there may be substantial inter-taxon variation in the number of LGT events that become fixed even between amoebozoan species that have similar feeding modalities.
Acanthamoeba castellanii↗