+ Site Statistics
+ Search Articles
+ PDF Full Text Service
How our service works
Request PDF Full Text
+ Follow Us
Follow on Facebook
Follow on Twitter
Follow on LinkedIn
+ Subscribe to Site Feeds
Most Shared
PDF Full Text
+ Translate
+ Recently Requested

Purification of Enterocytozoon bieneusi spores from stool specimens by gradient and cell sorting techniques



Purification of Enterocytozoon bieneusi spores from stool specimens by gradient and cell sorting techniques



Journal of Clinical Microbiology 42(7): 3256-3261



A three-step method for the purification of Enterocytozoon bieneusi spores from stool specimens was developed. The primary process of purification of the spores from bacterial contaminants involved Percoll gradient centrifugation followed by additional separation using cesium chloride density gradient centrifugation. The cesium chloride-isolated spores were further purified using a flow cytometer with cell sorting capabilities. Sorting was performed without the use of antibodies, fluorochromes, or dyes, leaving the sorted spores in their native state, which appears to be less destructive for spores. When quantified by flow cytometry using tubes with known numbers of highly fluorescent polystyrene beads, the sorted material showed a slight decrease in light scatter characteristics compared with the slightly larger Encephalitozoon species spores. Although the overall recovery of the E. bieneusi spores was low, calcofluor and Gram chromotrope staining, indirect immunofluorescence assay, and transmission electron microscopy revealed that the sorted material was highly purified and contained large numbers of E. bieneusi spores and relatively few bacteria and other debris. The sorted material appeared to be sufficiently pure and could be used for in vitro culture and for the development of a variety of diagnostic reagents as well as in studying the genome of E. bieneusi and host-parasite interactions.

Please choose payment method:






(PDF emailed within 0-6 h: $19.90)

Accession: 004288611

Download citation: RISBibTeXText

PMID: 15243090

DOI: 10.1128/JCM.42.7.3256-3261.2004


Related references

Identification of spores of Enterocytozoon bieneusi in stool and duodenal fluid from AIDS patients. Lancet 336(Nov. 3): 1127-1128, 1990

Prevalence of Enterocytozoon bieneusi spores in the stool of AIDS patients and African children not infected by HIV. Bulletin de la Societe de Pathologie Exotique 86(5): 351-357, 1993

Real-time PCR method for detection of the microsporidian Enterocytozoon bieneusi from stool specimens. Abstracts of the General Meeting of the American Society for Microbiology 103: C-283, 2003

Detection and identification of Enterocytozoon bieneusi and Encephalitozoon species in stool and urine specimens by PCR and differential hybridization. Journal of Clinical Microbiology 43(2): 610-614, 2005

Diagnosis of enterocytozoon bieneusi microsporidiosis by detection of spores in human fecal specimens. Program & Abstracts of the Interscience Conference on Antimicrobial Agents & Chemotherapy 32: 244, 1992

Diagnosis of infections caused by Enterocytozoon bieneusi and Encephalitozoon intestinalis using polymerase chain reaction in stool specimens. Aids 11(6): 723-726, 1997

Quantitative light microscopic detection of Enterocytozoon bieneusi in stool specimens: a longitudinal study of human immunodeficiency virus-infected microsporidiosis patients. Journal of Clinical Microbiology 34(3): 520-523, 1996

One-step purification of Enterocytozoon bieneusi spores from human stools by immunoaffinity expanded-bed adsorption. Journal of Clinical Microbiology 39(5): 1947-1951, 2001

Development of a real-time polymerase-chain-reaction assay for quantitative detection of Enterocytozoon bieneusi DNA in stool specimens from immunocompromised patients with intestinal microsporidiosis. Journal of Infectious Diseases 187(9): 1469-1474, 2003

Diagnosis of Enterocytozoon bieneusi by PCR in stool samples eluted from filter paper disks. Clinical and Diagnostic Laboratory Immunology 7(3): 504-506, 2000

Genotype identification of Enterocytozoon bieneusi isolates from stool samples of HIV-infected Tunisian patients. Parasite 19(2): 147-151, 2012

Diagnosis of enterocytozoon bieneusi spores in feces of aids patients. Micron & Microscopica Acta 21(4): 251-252, 1990

Genotype Identification and Phylogenetic Analysis of Enterocytozoon bieneusi Isolates from Stool Samples of Diarrheic Children. Journal of Parasitology 104(3): 297-301, 2018

Flow cytometry as a possible method of isolation of spores of the microsporidian Enterocytozoon bieneusi. Journal of Eukaryotic Microbiology 41(5): 27s, 1994

Identification of genotypes of Enterocytozoon bieneusi from stool samples from human immunodeficiency virus-infected patients in Thailand. Journal of Clinical Microbiology 44(8): 3001-3004, 2006