Acanthamoeba spp. are free-living amebae that inhabit a variety of air, soil, and water environments. However, these amebae can also act as opportunistic as well as nonopportunistic pathogens. They are the causative agents of granulomatous amebic encephalitis and amebic keratitis and have been associated with cutaneous lesions and sinusitis. Immuno compromised individuals, including AIDS patients, are particularly susceptible to infections with Acanthamoeba. The immune defense mechanisms that operate against Acanthamoeba have not been well characterized, but it has been proposed that both innate and acquired immunity play a role. The ameba's life cycle includes an active feeding trophozoite stage and a dormant cyst stage. Trophozoites feed on bacteria, yeast, and algae. However, both trophozoites and cysts can retain viable bacteria and may serve as reservoirs for bacteria with human pathogenic potential. Diagnosis of infection includes direct microscopy of wet mounts of cerebrospinal fluid or stained smears of cerebrospinal fluid sediment, light or electron microscopy of tissues, in vitro cultivation of Acanthamoeba, and histological assessment of frozen or paraffin-embedded sections of brain or cutaneous lesion biopsy material. Immunocytochemistry, chemifluorescent dye staining, PCR, and analysis of DNA sequence variation also have been employed for laboratory diagnosis. Treatment of Acanthamoeba infections has met with mixed results. However, chlorhexidine gluconate, alone or in combination with propamidene isethionate, is effective in some patients. Furthermore, effective treatment is complicated since patients may present with underlying disease and Acanthamoeba infection may not be recognized. Since an increase in the number of cases of Acanthamoeba infections has occurred worldwide, these protozoa have become increasingly important as agents of human disease.

Acanthamoeba spp. as agents of disease in humans.

Among the many genera of free-living amoebae that exist in nature, members of only four genera have an association with human disease: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri and Sappinia diploidea. Acanthamoeba spp. and B. mandrillaris are opportunistic pathogens causing infections of the central nervous system, lungs, sinuses and skin, mostly in immunocompromised humans. Balamuthia is also associated with disease in immunocompetent children, and Acanthamoeba spp. cause a sight-threatening infection, Acanthamoeba keratitis, mostly in contact-lens wearers. Of more than 30 species of Naegleria, only one species, N. fowleri, causes an acute and fulminating meningoencephalitis in immunocompetent children and young adults. In addition to human infections, Acanthamoeba, Balamuthia and Naegleria can cause central nervous system infections in animals. Because only one human case of encephalitis caused by Sappinia diploidea is known, generalizations about the organism as an agent of disease are premature. In this review we summarize what is known of these free-living amoebae, focusing on their biology, ecology, types of disease and diagnostic methods. We also discuss the clinical profiles, mechanisms of pathogenesis, pathophysiology, immunology, antimicrobial sensitivity and molecular characteristics of these amoebae.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1574-695X.2007.00232.x

Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea

Acanthamoeba is an opportunistic protozoan that is widely distributed in the environment and is well recognized to produce serious human infections, including a blinding keratitis and a fatal encephalitis. This review presents our current understanding of the burden of Acanthamoeba infections on human health, their pathogenesis and pathophysiology, and molecular mechanisms associated with the disease, as well as virulence traits of Acanthamoeba that may be targets for therapeutic interventions and/or the development of preventative measures.

Acanthamoeba: biology and increasing importance in human health

Free-living amoebae as opportunistic and non-opportunistic pathogens of humans and animals.

Summary-The capacity of bacteria and protozoa to mineralize soil nitrogen was studied in microcosms with sterilized soil with or without wheat plants. The effect of small additions of glucose or ammonium nitrate or both, twice a week was also tested. Plant dry weight and N-content, number of microorganisms and biomass plus inorganic N were determined after 6 weeks. The introduction of plants profoundly influenced the N tr~sfo~ations. In the presence of root-derived carbon, much more N was mineralized from the organic matter and immobilized mainly in plant biomass. “Total observable change in biomass N plus inorganic N” was negative in the unvegetated soils without additions, while a mmeralization of 1.7 mg N microcosm-’ was observed in microcosms with wheat plants grown with bacteria only. When protozoa were included, the N taken up by plants increased by 75%. Sugar additions resulted in an 18% increase of total N in the shoots when protozoa were present, but had no significant effect in the absence of grazers. Plants with the same root weight were more efficient in their uptake of inorganic N when protozoa were present. Plants grown with protozoa also had a lower R/S ratio, indicating a less stressed N availabiIitv situatron. The lowest ratio was found with N additions m the presence of protozoa. The results indicate that, with energy supplied by plant roots or with external glucose additions, soil bacteria can mineralize N from the soil organic matter to support their own growth. Grazing of the bacteria is necessary to make bacterial biomass N available for plant uptake.

Interactions of bacteria, protozoa and plants leading to mineralization of soil nitrogen.

SYNOPSIS. Seven species of limax amoebae were isolated into clonal, monoxenic cultures with Aerobacter aerogenes from material collected from freshwater habitats. Studies were made of their trophic structure, nuclear division, cyst structure, some aspects of cytochemistry, and other characteristics. Six new species are described: Vahlkampfia inornata, V. avara, V. jugosa, Hartmannella limacoides, H. vermiformis, and H. exundans. The well-known species Naegleria gruberi (Schardinger, 1899) is re-described on the basis of 8 strains; its flagellated phase was found to be biflagellate, with rare exceptions.



A correlation exists between the manner of locomotion and the pattern of nuclear division in the limax amoebae in the family Vahlkampfiidae and those in the genus Hartmannella.



Trophic amoebae of all species had a PAS-positive surface layer, altho results with H. vermiformis and H. exundans were less definite than with other species. All species except H. limacoides formed cysts in culture. The cyst walls of all cyst-forming species were strongly PAS-positive, but results of the zinc chloroiodide test for cellulose were negative with the method used.



The genus Hartmannella Alexeieff, 1912, is re-defined to include those species which assume a simple, monopodial limax-like form during locomotion and have nuclear division similar to that of metazoan cells and to distinguish it from the genus Acanthamoeba Volkonsky, 1931.

Taxonomic Criteria for Limax Amoebae, with Descriptions of 3 New Species of Hartmannella and 3 of Vahlkampfia*

SYNOPSIS. Ten strains of Acanthamoeba from freshwater habitats were isolated in clonal cultures. Studies were made of trophic structure, nuclear division, cyst structure, some aspects of cytochemistry, and other characteristics. One strain was identified as A. castellanii (Douglas, 1930), one as A. astronyxis (Ray and Hayes, 1954), and 8 as A. polyphaga (Puschkarew, 1913). Strains of Acanthamoeba isolated by other workers were also examined comparatively.

The pattern of nuclear division in all strains resembled that in metazoan cells, with the exception that centrioles were never found. Trophic amoebae had a PAS-positive surface outline. Cyst walls were strongly PAS-positive and also gave a positive test for cellulose with zinc chloroiodide.

The genus Acanthamoeba Volkonsky, 1931 is re-defined, being distinguished from Hartmannella Alexeieff, 1912, emend. Volkonsky, chiefly by the formation of tapering, hyaline pseudopods (acanthopodia) and by a cyst made up of an ectocyst and a polyhedral or stellate endocyst, with excystment by removal of opercula. Other characteristics found in all strains include a distinctive food cup, the presence of many small refractile globules in the cytoplasm of trophic amoebae, and a cyst wall containing cellulose. The degree of spindle convergence, employed by Volkonsky as a generic criterion, was unusable.

Differential diagnoses based principally on cyst structure are offered for A. castellanii, A. astronyxis, and A. polyphaga. The strain previously called Mayorella palestinensis Reich, 1933 is a distinct species of Acanthamoeba.

Re-definition of the genus Acanthamoeba with descriptions of three species.

SYNOPSIS. Six strains of amoebae isolated from freshwater habitats and having broad, flattened locomotive forms were studied in clonal cultures, with attention to trophic structure, nuclear division, cyst structure in those strains which formed cysts, some aspects of cytochemistry, and other characteristics.



Three strains were found to belong to the genus Flabellula, one being identified as F. platypodia (Glaser, 1912) and the other 2 as F. mira Schaeffer, 1926. The new species Rugipes placidus is described. One strain, identified as belonging to the species originally called Amoeba actinophora Auerbach, 1855, is re-described as Hyalodiscus actinophorus, and a similar strain is described as H. actinophorus var. minor n. var.



All 3 strains of Flabellula had the unusual type of mitotic figure reported by Glaser for F. platypodia but not previously studied with the Feulgen technic. The genus Rugipes has the mitotic pattern more common to small amoebae and is distinguished from Flabellula partly on that basis. Both strains of Hyalodiscus have division figures of this common type.



Of the strains included in this investigation, only R. placidus and H. actinophorus formed cysts in culture. The cysts of neither species were viable after one year at room temperature and humidity.



The status of Rugipes as distinct from Flabellula is discussed. The genus Hyalodiscus is interpreted in a manner compatible with the original definition of Hertwig and Lesser and is included within the order Amoebida. The strains of this genus isolated have a doubly contoured, strongly PAS-positive covering over the endoplasmic hump.



Some criteria for the taxonomy of amoebae are evaluated in the light of work reported in this and 2 previous papers.

Generic Criteria for Flabellula, Rugipes and Hyalodiscus, With Descriptions of Species*

In its amoeboid stage, Protacanthamoeba caledonica n. g., n. sp. closely resembles the genus Acanthamoeba, on both light- and electron-microscopical levels, including possession of a centrosphere with a plaque-shaped centriole-like body. The cyst wall differs from that of Acanthamoeba in lack of preformed exit pores and in fine structure; the occasional apparent division into exocyst and endocyst is due to irregular splitting. The strain isolated from a Scottish estuary did not grow at 37°C and did not grow normally on agar made with 25% sea water, but cysts remained viable after a week in full-strength sea water. Protacanthamoeba n. g. is distinguished from Acanthamoeba on the basis of cyst structure, but it is assigned to the family Acanthamoebidae.

Frederick C. Page

Papers

Taxonomic Criteria for Limax Amoebae, with Descriptions of 3 New Species of Hartmannella and 3 of Vahlkampfia*

Re-definition of the genus Acanthamoeba with descriptions of three species.

Generic Criteria for Flabellula, Rugipes and Hyalodiscus, With Descriptions of Species*

A Light- and Electron-Microscopical Study of Protacanthamoeba caledonica n. sp., Type-Species of Protacanthamoeba n. g. (Amoebida, Acanthamoebidae)†