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. 1997 Mar;35(3):724–729. doi: 10.1128/jcm.35.3.724-729.1997

Simple diagnosis of Encephalitozoon sp. microsporidial infections by using a panspecific antiexospore monoclonal antibody.

F J Enriquez 1, O Ditrich 1, J D Palting 1, K Smith 1
PMCID: PMC229658  PMID: 9041420

Abstract

Microsporidia (phylum Microsproa) have recently become recognized as common opportunistic protozoans in the United States and worldwide, particularly affecting immunodeficient patients. Microsporidian organisms within the genus Encephalitozoon are the cause of nephrologic, ophthalmic, pneumologic, gastroenteric, and systemic infections. However, diagnosis of the small spores by light microscopy is difficult, even with newly developed and improved staining techniques. We have developed an anti-Encephalitozoon species monoclonal antibody-based immunoassay for easy diagnosis. A hybridoma was produced and selected following one main criterion: recognition by immunofluorescence of all known Encephalitozoon spores affecting humans. The selected monoclonal antibody-secreting hybridomas were characterized by enzyme-linked immunosorbent assay, immunofluorescence, Western blot, and immunoelectron microscopy using Encephalitozoon species from fresh and fixed samples from patients and from in vitro cultures. In the immunofluorescence assay, one monoclonal antibody, termed 3B6, strongly recognized Encephalitozoon cuniculi, E. hellem, and E. intestinalis. Monoclonal antibody 3B6 bound to other microsporidia (Nosema and Vairimorpha spp.) without cross-reacting with any other parasite, including Enterocytozoon bieneusi, fungus, or bacterium tested. In immunoelectron microscopy assays, monoclonal antibody 3B6 bound to the exospore of Encephalitozoon species, while in Western blot assays, it recognized three to seven antigens with molecular masses ranging from 34 to 117 kDa. We have developed a sensitive and specific monoclonal antibody-based immunoassay to diagnose common microsporidian infections, particularly with Encephalitozoon species. This is a new tool for identifying spores in bodily fluids and biopsy samples and is an efficient diagnostic test. Additionally, monoclonal antibody 3B6 can serve to assess the prevalence of microsporidial infections in immunodeficient and immunocompetent patients.

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Selected References

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  1. Aldras A. M., Orenstein J. M., Kotler D. P., Shadduck J. A., Didier E. S. Detection of microsporidia by indirect immunofluorescence antibody test using polyclonal and monoclonal antibodies. J Clin Microbiol. 1994 Mar;32(3):608–612. doi: 10.1128/jcm.32.3.608-612.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Asmuth D. M., DeGirolami P. C., Federman M., Ezratty C. R., Pleskow D. K., Desai G., Wanke C. A. Clinical features of microsporidiosis in patients with AIDS. Clin Infect Dis. 1994 May;18(5):819–825. doi: 10.1093/clinids/18.5.819. [DOI] [PubMed] [Google Scholar]
  3. Beckers P. J., Derks G. J., Gool T., Rietveld F. J., Sauerwein R. W. Encephalocytozoon intestinalis-specific monoclonal antibodies for laboratory diagnosis of microsporidiosis. J Clin Microbiol. 1996 Feb;34(2):282–285. doi: 10.1128/jcm.34.2.282-285.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Becnel J. Life cycles and host-parasite relationships of Microsporidia in culicine mosquitoes. Folia Parasitol (Praha) 1994;41(2):91–96. [PubMed] [Google Scholar]
  5. Cali A., Kotler D. P., Orenstein J. M. Septata intestinalis N. G., N. Sp., an intestinal microsporidian associated with chronic diarrhea and dissemination in AIDS patients. J Eukaryot Microbiol. 1993 Jan-Feb;40(1):101–112. doi: 10.1111/j.1550-7408.1993.tb04889.x. [DOI] [PubMed] [Google Scholar]
  6. Clarridge J. E., 3rd, Karkhanis S., Rabeneck L., Marino B., Foote L. W. Quantitative light microscopic detection of Enterocytozoon bieneusi in stool specimens: a longitudinal study of human immunodeficiency virus-infected microsporidiosis patients. J Clin Microbiol. 1996 Mar;34(3):520–523. doi: 10.1128/jcm.34.3.520-523.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Desportes I., Le Charpentier Y., Galian A., Bernard F., Cochand-Priollet B., Lavergne A., Ravisse P., Modigliani R. Occurrence of a new microsporidan: Enterocytozoon bieneusi n.g., n. sp., in the enterocytes of a human patient with AIDS. J Protozool. 1985 May;32(2):250–254. doi: 10.1111/j.1550-7408.1985.tb03046.x. [DOI] [PubMed] [Google Scholar]
  8. Didier E. S., Didier P. J., Friedberg D. N., Stenson S. M., Orenstein J. M., Yee R. W., Tio F. O., Davis R. M., Vossbrinck C., Millichamp N. Isolation and characterization of a new human microsporidian, Encephalitozoon hellem (n. sp.), from three AIDS patients with keratoconjunctivitis. J Infect Dis. 1991 Mar;163(3):617–621. doi: 10.1093/infdis/163.3.617. [DOI] [PubMed] [Google Scholar]
  9. Didier E. S., Rogers L. B., Brush A. D., Wong S., Traina-Dorge V., Bertucci D. Diagnosis of disseminated microsporidian Encephalitozoon hellem infection by PCR-Southern analysis and successful treatment with albendazole and fumagillin. J Clin Microbiol. 1996 Apr;34(4):947–952. doi: 10.1128/jcm.34.4.947-952.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Enriquez F. J., Bradley-Dunlop D., Joens L. Increased proportion of antigen-specific antibody-producing hybridomas following an in vitro immunization with in vivo immunized mouse spleen cells. Hybridoma. 1991 Dec;10(6):745–751. doi: 10.1089/hyb.1991.10.745. [DOI] [PubMed] [Google Scholar]
  11. Field A. S., Marriott D. J., Hing M. C. The Warthin-Starry stain in the diagnosis of small intestinal microsporidiosis in HIV-infected patients. Folia Parasitol (Praha) 1993;40(4):261–266. [PubMed] [Google Scholar]
  12. Hartskeerl R. A., Van Gool T., Schuitema A. R., Didier E. S., Terpstra W. J. Genetic and immunological characterization of the microsporidian Septata intestinalis Cali, Kotler and Orenstein, 1993: reclassification to Encephalitozoon intestinalis. Parasitology. 1995 Apr;110(Pt 3):277–285. doi: 10.1017/s0031182000080860. [DOI] [PubMed] [Google Scholar]
  13. Moura H., Da Silva J. L., Sodré F. C., Brasil P., Wallmo K., Wahlquist S., Wallace S., Croppo G. P., Visvesvara G. S. Gram-chromotrope: a new technique that enhances detection of microsporidial spores in clinical samples. J Eukaryot Microbiol. 1996 Sep-Oct;43(5):94S–95S. doi: 10.1111/j.1550-7408.1996.tb05019.x. [DOI] [PubMed] [Google Scholar]
  14. Pleshinger J., Weidner E. The microsporidian spore invasion tube. IV. Discharge activation begins with pH-triggered Ca2+ influx. J Cell Biol. 1985 Jun;100(6):1834–1838. doi: 10.1083/jcb.100.6.1834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Ryan N. J., Sutherland G., Coughlan K., Globan M., Doultree J., Marshall J., Baird R. W., Pedersen J., Dwyer B. A new trichrome-blue stain for detection of microsporidial species in urine, stool, and nasopharyngeal specimens. J Clin Microbiol. 1993 Dec;31(12):3264–3269. doi: 10.1128/jcm.31.12.3264-3269.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Visvesvara G. S., Leitch G. J., da Silva A. J., Croppo G. P., Moura H., Wallace S., Slemenda S. B., Schwartz D. A., Moss D., Bryan R. T. Polyclonal and monoclonal antibody and PCR-amplified small-subunit rRNA identification of a microsporidian, Encephalitozoon hellem, isolated from an AIDS patient with disseminated infection. J Clin Microbiol. 1994 Nov;32(11):2760–2768. doi: 10.1128/jcm.32.11.2760-2768.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Weber R., Bryan R. T. Microsporidial infections in immunodeficient and immunocompetent patients. Clin Infect Dis. 1994 Sep;19(3):517–521. doi: 10.1093/clinids/19.3.517. [DOI] [PubMed] [Google Scholar]
  18. Weber R., Bryan R. T., Owen R. L., Wilcox C. M., Gorelkin L., Visvesvara G. S. Improved light-microscopical detection of microsporidia spores in stool and duodenal aspirates. The Enteric Opportunistic Infections Working Group. N Engl J Med. 1992 Jan 16;326(3):161–166. doi: 10.1056/NEJM199201163260304. [DOI] [PubMed] [Google Scholar]
  19. Weber R., Bryan R. T., Schwartz D. A., Owen R. L. Human microsporidial infections. Clin Microbiol Rev. 1994 Oct;7(4):426–461. doi: 10.1128/cmr.7.4.426. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Weiss L. M., Cali A., Levee E., LaPlace D., Tanowitz H., Simon D., Wittner M. Diagnosis of Encephalitozoon cuniculi infection by western blot and the use of cross-reactive antigens for the possible detection of microsporidiosis in humans. Am J Trop Med Hyg. 1992 Oct;47(4):456–462. doi: 10.4269/ajtmh.1992.47.456. [DOI] [PubMed] [Google Scholar]
  21. van Gool T., Hollister W. S., Schattenkerk W. E., Van den Bergh Weerman M. A., Terpstra W. J., van Ketel R. J., Reiss P., Canning E. U. Diagnosis of Enterocytozoon bieneusi microsporidiosis in AIDS patients by recovery of spores from faeces. Lancet. 1990 Sep 15;336(8716):697–698. doi: 10.1016/0140-6736(90)92198-q. [DOI] [PubMed] [Google Scholar]
  22. van Gool T., Snijders F., Reiss P., Eeftinck Schattenkerk J. K., van den Bergh Weerman M. A., Bartelsman J. F., Bruins J. J., Canning E. U., Dankert J. Diagnosis of intestinal and disseminated microsporidial infections in patients with HIV by a new rapid fluorescence technique. J Clin Pathol. 1993 Aug;46(8):694–699. doi: 10.1136/jcp.46.8.694. [DOI] [PMC free article] [PubMed] [Google Scholar]

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