Rapid Growth of Acanthamoeba In Defined Media; Induction of Encystment By Glucose‐Acetate Starvation

Byers, Thomas J.; Akins, Robert A.; Maynard, Barbara; Lefken, Richard A.; Martin, Scott M.

doi:10.1111/j.1550-7408.1980.tb04684.x

Cited by 64 publications

(27 citation statements)

References 15 publications

(5 reference statements)

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“…Any interference in the maturation process will unduly affect the resistance of the endocyst because resistance to biocides develops during the cellulose synthesis phase of encystment. Previous studies have shown that inadequate aeration and improper control of pH may also hamper encystment (e.g., 8% encystment versus Ͼ80% encystment with aeration and no pH control [6,27]), leading to imperfect cyst wall synthesis. Variation in buffers and the inclusion of a chelating agent (EDTA) or the use of dimethyl sulfoxide in the test solutions may also adversely affect the efficacies of the biocides (18,42).…”

Section: Discussionmentioning

confidence: 99%

Resistance of Acanthamoeba Cysts to Disinfection in Multiple Contact Lens Solutions

Johnston¹,

Sriram²,

Qvarnström³

et al. 2009

J Clin Microbiol

114

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Acanthamoebae are free-living amoebae found in the environment, including soil, freshwater, brackish water, seawater, hot tubs, and Jacuzzis. Acanthamoeba species can cause keratitis, a painful vision-threatening infection of the cornea, and fatal granulomatous encephalitis in humans. More than 20 species of Acanthamoeba belonging to morphological groups I, II, and III distributed in 15 genotypes have been described. Among these, Acanthamoeba castellanii, A. polyphaga, and A. hatchetti are frequently identified as causing Acanthamoeba keratitis (AK). Improper contact lens care and contact with nonsterile water while wearing contact lenses are known risk factors for AK. During a recent multistate outbreak, AK was found to be associated with the use of Advanced Medical Optics Complete MoisturePlus multipurpose contact lens solution, which was hypothesized to have had insufficient anti-Acanthamoeba activity. As part of the investigation of that outbreak, we compared the efficacies of 11 different contact lens solutions against cysts of A. castellanii, A. polyphaga, and A. hatchetti (the isolates of all species were genotype T4), which were isolated in 2007 from specimens obtained during the outbreak investigation. The data, generated with A. castellanii, A. polyphaga, and A. hatchetti cysts, suggest that the two contact lens solutions containing hydrogen peroxide were the only solutions that showed any disinfection ability, with 0% and 66% growth, respectively, being detected with A. castellanii and 0% and 33% growth, respectively, being detected with A. polyphaga. There was no statistically significant difference in disinfection efficacy between the 11 solutions for A. hatchetti.Acanthamoebae, which are free-living amoebae, occur worldwide in soil and water. It has been isolated from ponds, lakes, brackish water. and seawater; filters of heating, ventilating, and air-conditioning units; medical equipment, such as gastric wash tubing, dental irrigation units, contact lenses, and contact lens solutions; as well as vegetables, cell cultures, and even human and animal tissues (7,23,39). It has also been isolated from toxic waste dumpsites with high levels of pesticides, herbicides, pharmaceuticals, heavy metals, and polychlorinated biphenyls (35). Acanthamoeba species have two stages in their life cycle: a vegetative or trophozoite stage that reproduces by binary fission and that feeds voraciously on the bacteria and detritus present in the environment and a nondividing, cyst stage that is resistant to environmental stress. Acanthamoeba amoebae cause different types of human disease, including central nervous system infections (granulomatous amebic encephalitis, cutaneous infections) Acanthamoeba dermatitis, and ocular infections (Acanthamoeba keratitis [AK]). Granulomatous amebic encephalitis and cutaneous infections principally occur in immunocompromised individuals, including patients with human immunodeficiency virus infection or AIDS (17,23,37,43). In contrast, AK principally occurs in immunocompetent individuals.

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Section: Discussionmentioning

confidence: 99%

Resistance of Acanthamoeba Cysts to Disinfection in Multiple Contact Lens Solutions

Johnston¹,

Sriram²,

Qvarnström³

et al. 2009

J Clin Microbiol

114

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“…Cultures representing the three morphological groups (30,38) and the 12 18S ribosomal DNA (rDNA) sequence types (termed genotypes here) of Acanthamoeba (35) were maintained in liquid broth (OGM) as previously described (5). The Scottish corneal scrapes were obtained in a population-based longitudinal study of keratitis in the West of Scotland (33) and were stored in sterile saline.…”

Section: Methodsmentioning

confidence: 99%

Use of Subgenic 18S Ribosomal DNA PCR and Sequencing for Genus and Genotype Identification of Acanthamoebae from Humans with Keratitis and from Sewage Sludge

et al. 2001

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This study identified subgenic PCR amplimers from 18S rDNA that were (i) highly specific for the genus Acanthamoeba, (ii) obtainable from all known genotypes, and (iii) useful for identification of individual genotypes. A 423-to 551-bp Acanthamoeba-specific amplimer ASA.S1 obtained with primers JDP1 and JDP2 was the most reliable for purposes i and ii. A variable region within this amplimer also identified genotype clusters, but purpose iii was best achieved with sequencing of the genotype-specific amplimer GTSA.B1. Because this amplimer could be obtained from any eukaryote, axenic Acanthamoeba cultures were required for its study. GTSA.B1, produced with primers CRN5 and 1137, extended between reference bp 1 and 1475. Genotypic identification relied on three segments: bp 178 to 355, 705 to 926, and 1175 to 1379. ASA.S1 was obtained from single amoeba, from cultures of all known 18S rDNA genotypes, and from corneal scrapings of Scottish patients with suspected Acanthamoeba keratitis (AK). The AK PCR findings were consistent with culture results for 11 of 15 culture-positive specimens and detected Acanthamoeba in one of nine culturenegative specimens. ASA.S1 sequences were examined for 6 of the 11 culture-positive isolates and were most closely associated with genotypic cluster T3-T4-T11. A similar distance analysis using GTSA.B1 sequences identified nine South African AK-associated isolates as genotype T4 and three isolates from sewage sludge as genotype T5. Our results demonstrate the usefulness of 18S ribosomal DNA PCR amplimers ASA.S1 and GTSA.B1 for Acanthamoeba-specific detection and reliable genotyping, respectively, and provide further evidence that T4 is the predominant genotype in AK.The demonstrated pathogenicity for humans and animals of organisms belonging to the genus Acanthamoeba (17, 26), coupled with the difficulty of using morphological criteria for subgeneric identification of isolates (30,38), has stimulated a number of laboratories to pursue molecular methods for detection and identification. The objective is to develop methods that are suitable for both clinical and environmental applications. The identification of amoebic isolates should be very reliable and, at least for clinical use, the detection system should be very sensitive. Several research groups, including our own, have demonstrated the usefulness of PCR methods for detection of acanthamoebae (10,15,21,25,27,40). As few as 1 to 10 trophozoites can be detected. It also is possible to enhance detection of individual amoeba in very dilute liquid clinical samples with fluorescent in situ hybridization (FISH) (36). Several molecular approaches increase the reliability of specimen identification, but the use of DNA sequence variation appears to be the most promising. The variation is observed in restriction fragment length polymorphisms of complete or partial nuclear 18S rRNA genes (8,20,21,22), of complete mitochondrial 16S rRNA genes (7, 46), and of the complete mitochondrial genome (3,7,13,18,22,45). It also is observed in the DNA seq...

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“…Adam and Blewett (2) compared carbohydrate utilization of different strains of A. castellanii in a basal medium containing 5 (arginine, leucine, isoleucine, methionine, and valine) of the 10 essential amino acids and found variations in use of sucrose, melibiose, mannitol, and raffinose as carbon energy sources. These defined media supported growth, but often with extended generation times from 40 to Ͼ60 h. In order to produce a higher growth rate, Byers et al (9) formulated a defined medium (Table 3) based on these earlier studies. Their media, DGM-21A and DGM-21B, gave generation times of about 13 and 16 h, respectively, and cell yields of 2 ϫ 10 6 to 3 ϫ 10 6 amebas/ml.…”

Section: Acanthamoeba Sppmentioning

confidence: 99%

Cultivation of Pathogenic and Opportunistic Free-Living Amebas

2002

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Free-living amebas are widely distributed in soil and water, particularly members of the genera Acanthamoeba and Naegleria. Since the early 1960s, they have been recognized as opportunistic human pathogens, capable of causing infections of the central nervous system (CNS) in both immunocompetent and immunocompromised hosts. Naegleria is the causal agent of a fulminant CNS condition, primary amebic meningoencephalitis; Acanthamoeba is responsible for a more chronic and insidious infection of the CNS termed granulomatous amebic encephalitis, as well as amebic keratitis. Balamuthia sp. has been recognized in the past decade as another ameba implicated in CNS infections. Cultivation of these organisms in vitro provides the basis for a better understanding of the biology of these amebas, as well as an important means of isolating and identifying them from clinical samples. Naegleria and Acanthamoeba can be cultured axenically in cell-free media or on tissue culture cells as feeder layers and in cultures with bacteria as a food source. Balamuthia, which has yet to be isolated from the environment, will not grow on bacteria. Instead, it requires tissue culture cells as feeder layers or an enriched cell-free medium. The recent identification of another ameba, Sappinia diploidea, suggests that other free-living forms may also be involved as causal agents of human infections

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Rapid Growth of Acanthamoeba In Defined Media; Induction of Encystment By Glucose‐Acetate Starvation

Cited by 64 publications

References 15 publications

Resistance of Acanthamoeba Cysts to Disinfection in Multiple Contact Lens Solutions

Resistance of Acanthamoeba Cysts to Disinfection in Multiple Contact Lens Solutions

Use of Subgenic 18S Ribosomal DNA PCR and Sequencing for Genus and Genotype Identification of Acanthamoebae from Humans with Keratitis and from Sewage Sludge

Cultivation of Pathogenic and Opportunistic Free-Living Amebas

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