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Detection by an immunofluorescence test of Encephalitozoon intestinalis spores in routinely formalin-fixed stool
H Moura1, F C Sodre, F J Bornay-Llinares
1Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, Department of Health and Human Services, Atlanta, Georgia 30341-3724, USA.
Abstract:
Of the several microsporidia that infect humans, Enterocytozoon bieneusi is known to cause a gastrointestinal disease whereas Encephalitozoon intestinalis causes both a disseminated and an intestinal disease. Although several different staining techniques, including the chromotrope technique and its modifications, Uvitex 2B, and the quick-hot Gram-chromotrope procedure, detect microsporidian spores in fecal smears and other clinical samples, they do not identify the species of microsporidia. A need for an easily performed test therefore exists. We reevaluated 120 stool samples that had been found positive for microsporidia previously, using the quick-hot Gram-chromotrope technique, and segregated them into two groups on the basis of spore size. We also screened the smears by immunofluorescence microscopy, using a polyclonal rabbit anti-E. intestinalis serum at a dilution of 1:400. Spores in 29 (24.1%) of the 120 samples fluoresced brightly, indicating that they were E. intestinalis spores. No intense background or cross-reactivity with bacteria, yeasts, or other structures in the stool samples was seen. Additionally, the numbers of spores that fluoresced in seven of these samples were substantially smaller than the numbers of spores that were present in the stained smears, indicating that these samples were probably derived from patients with mixed infections of Enterocytozoon bieneusi and E. intestinalis. Because a 1:400 dilution of this serum does not react with culture-grown Encephalitozoon hellem, Encephalitozoon cuniculi, or Vittaforma corneae or with Enterocytozoon bieneusi spores in feces, we concluded that an immunofluorescence test using this serum is a good alternative for the specific identification of E. intestinalis infections.
Insights
A new immunofluorescence test accurately identifies Encephalitozoon intestinalis (E. intestinalis) in stool samples. This method aids in diagnosing microsporidiosis, distinguishing it from Enterocytozoon bieneusi infections.
Area of Science:
- Medical Parasitology
- Clinical Microbiology
- Immunology
Background:
- Microsporidia are opportunistic pathogens causing gastrointestinal and disseminated diseases in humans.
- Current staining techniques for microsporidia lack species-specific identification.
- Enterocytozoon bieneusi causes gastrointestinal disease, while Encephalitozoon intestinalis causes both intestinal and disseminated disease.
Purpose of the Study:
- To develop and evaluate an immunofluorescence assay for the specific identification of Encephalitozoon intestinalis in clinical samples.
- To differentiate E. intestinalis from other microsporidia species, particularly Enterocytozoon bieneusi.
Main Methods:
- Re-evaluation of 120 stool samples previously identified as positive for microsporidia using the quick-hot Gram-chromotrope technique.
- Screening of samples using immunofluorescence microscopy with a polyclonal rabbit anti-E. intestinalis serum at a 1:400 dilution.
- Comparison of immunofluorescence results with spore size and known reactivity profiles of the anti-serum.
Main Results:
- Spores of E. intestinalis were detected by immunofluorescence in 29.1% (29/120) of the samples.
- The assay showed no cross-reactivity with bacteria, yeasts, or other fecal structures.
- Seven samples indicated mixed infections of E. intestinalis and E. bieneusi based on differential spore counts.
- The anti-E. intestinalis serum did not react with other Encephalitozoon species or E. bieneusi spores.
Conclusions:
- Immunofluorescence microscopy using a specific polyclonal rabbit anti-E. intestinalis serum is a reliable method for identifying E. intestinalis infections.
- This assay provides a valuable tool for differentiating E. intestinalis from E. bieneusi and diagnosing microsporidiosis.
- The method can also detect mixed infections, improving diagnostic accuracy.