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Chapter 10 : Laboratory Diagnosis of Microsporidiosis

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Abstract:

The most robust and widely practicable technique for the diagnosis of microsporidial infection is light microscopic morphological demonstration of the organisms themselves. Evaluation of patients with suspected microsporidiosis should begin with light microscopic examination of stool specimens and urine or cytological examination of other body fluids. The most common clinical finding of ocular microsporidiosis is keratoconjunctivitis. In patients with suspected ocular microsporidiosis, urine and respiratory secretions also should be examined for microsporidia. In patients with ocular microsporidiosis, urine and respiratory secretions should also be examined for the presence of microsporidial spores. Serologic assays (including carbon immunoassay [CIA], IFAT, enzyme-linked immunosorbent assay [ELISA], and Western blot [WB]) have been applied in detecting specific IgG antibodies directed to spores in humans and to spores in humans and several animal species. Aside from studies on ocular microsporidiosis in presumably otherwise healthy persons, detailed histopathologic investigation of human microsporidial infection has been performed only in immunodeficient individuals. One clue that microsporidiosis may be present is based on the observation that infected epithelial cells containing mature spores are often shed intact into the lumen from the underlying basement membrane. Upper and lower respiratory tract infections due to microsporidia are associated almost exclusively with disseminated disease produced by all three members of the genus . Microsporidia have been found in almost every organ system. Molecular techniques have been used to compare microsporidial isolates obtained from humans and animals in order to study possible sources of infection and reservoir hosts.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10

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Lower Respiratory Tract Infections
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Upper Respiratory Tract Infections
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Image of FIGURE 1
FIGURE 1

Stool specimen from a patient with AIDS and chronic diarrhea, showing pinkish red-stained spores of measuring 0.7 to 1.0 by 1.1 to 1.6 μm. Chromotrope stain was used. Magnification, ×1,000 (oil immersion).

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 2
FIGURE 2

Stool specimen from a patient with AIDS and chronic diarrhea, showing microsporidial spores stained with a chemofluorescent optical brightening agent (Fungi-Fluor Kit, Polysciences, Inc., Warrington, Pa.). Magnification, ×1,000 (oil immersion).

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 3
FIGURE 3

Urine sediment from a patient with AIDS and disseminated infection, showing pinkish red-stained nricrosporidial spores measuring 1.0 to 1.5 by 2.0 to 3.0 μm. Chromotrope stain was used. Magnification, ×1,000 (oil immersion).

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 4
FIGURE 4

Cytospin preparation of bronchoalveolar lavage fluid from a patient with AIDS and intestinal infection, showing intracellular gram-positive microsporidial spores. Gram stain was used. Magnification ×1,000 (oil immersion).

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 5
FIGURE 5

Cytospin preparation of bronchoalveolar lavage fluid from a patient with AIDS and intestinal infection, showing intracellular microsporidia. Giemsa stain was used. Magnification, ×1,000 (oil immersion).

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 6
FIGURE 6

Cytospin preparation of cerebrospinal fluid, showing intracellular clusters of pink-stained microsporidial spores measuring 2 to 3 µm. Chromotrope stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 7
FIGURE 7

Immunofluorescence staining of sputum from a patient with disseminated infection (Weber et al., 1997a). Polyclonal rabbit antibody to was used. Spore walls and extruded polar tubes are seen. Magnification, ×600.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 8
FIGURE 8

Cytological preparation of conjunctival smear with superficial epithelial cells containing numerous gram-positive spores of . Brown and Hopps stain was used. Magnification, ×1,000.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 9
FIGURE 9

Stool specimen from a patient with AIDS and diarrhea, showing dark violet-stained spores of measuring 1.0 to 1.2 by 2.0 to 2.5 μm. Quick hot Gram-chromotrope stain was used. Magnification, ×1,000 (oil immersion). (Reprinted with permission from G. S.Visvesvara, Centers for Disease Control, Atlanta, Ga.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 10
FIGURE 10

Stool specimen from an immunocompetent Mexican patient with diarrhea, showing microsporidial spores stained with a monoclonal antibody to (Mab 3B6) ( ). The microsporidian was identified as , as described by Bornay-Llinares et al. (1998). Immunofluorescence detection was used. (Reprinted with permission from J. Enriquez, University of Arizona, Tucson,Ariz.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 11
FIGURE 11

Section of duodenal villus tip, showing epithelial cells infected with . Clusters of black-stained spores and plasmodia at various stages of development are shown. Modified Warthin-Starry stain was used. Magnification, ×600. (Reprinted with permission from A. S. Field, St.Vincent's Hospital, Darlinghurst, Australia.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 12
FIGURE 12

Section of duodenal villus tip, showing enterocytes and macrophages infected with blackstained clusters of . Modified Warthin-Starry stain was used. Magnification, ×600. (Reprinted with permission from A. S. Field, St. Vincent's Hospital, Darlinghurst, Australia.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 13
FIGURE 13

Transmission electron micrograph showing duodenal epithelium from an HIV-infected patient infected with . The different developmental stages between the enterocyte nuclei and the microvillus border include a proliferative plasmodium (1), late sporogonial plasmodia (2), and mature spores (3). A sloughing enterocyte containing mature spores is also shown (arrow). Magnification, ×3,700. (Reprinted with permission from M. A. Spycher, University Hospital, Zurich, Switzerland.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 14
FIGURE 14

Transmission electron micrograph of a microsporidial spore.The polar tubes (arrow) lie in two rows characteristic of . . Bar, 0.5 μm. Magnification, ×51,000.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 15
FIGURE 15

Immature and mature spores of within a parasitophorous vacuole in an infected renal tubular epithelial cell. Note the high concentration of spores within this vacuole. Original magnification, ×19,200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 16
FIGURE 16

Transmission electron micrograph of meronts and developing spores within enterocytes separated by a fibrillar matrix. develops within parasitophorous vacuoles. Magnification, ×5,300. (Reprinted with permission from M. A. Spycher, University Hospital, Zurich , Switzerland.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 17
FIGURE 17

Typical ultrastructural appearance of a mature spore of a sp. A diplokaryotic nucleus (with a dividing membrane) and 10 turns of the polar tubule are evident. Original magnification, ×38,000.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 18
FIGURE 18

Mucosal biopsy specimen, showing duodenal epithelial cells infected with . A cluster of gram-positive spores is located in the supranuclear portion of the cytoplasm. Brown and Brenn stain was used. Magnification, ×630.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 19
FIGURE 19

infection of the small intestine. The plastic-embedded semithin section shows spores larger than those of . The parasites are not in direct contact with the host cell cytoplasm but instead develop within a parasitophorous vacuole. Toluidine blue stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 20
FIGURE 20

A solitary corneal epithelial cell infected with from an enucleated globe obtained at autopsy. The spores are gram-positive. Brown and Hopps stain was used. Magnification, ×200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 21
FIGURE 21

Conjunctival biopsy specimen, demonstrating shedding and necrotic conjunctival epithelial cells containing gram-positive microsporidial spores admixed with neutrophils and fibrin from a patient with AIDS and disseminated infection. Brown and Hopps stain was used. Magnification, ×1,000.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 22
FIGURE 22

Plastic embedded semithin section of a nasal mucosal biopsy specimen, showing epithelial infection with spores. Toluidine blue stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 23
FIGURE 23

Microsporidial sinusitis due to an sp. Numerous infected epithelial cells can be seen to contain faintly gram-positive clusters of spores. Brown and Hopps stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 24
FIGURE 24

Plastic-embedded semithin section of tracheal mucosa with numerous spores of present in respiratory lining epithelium. Toluidine blue stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 25
FIGURE 25

Open lung biopsy specimen, showing bronchiolitis and bronchopneumonia due to . Microsporidia cannot be seen in this section, but the identity of this microsporidian was confirmed with both fluorescent antibody and PCR methods. Hematoxylin and eosin stain was used. Magnification, ×200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 26
FIGURE 26

Alveolar macrophage containing phagocytosed spores of . Brown and Hopps stain was used. Magification, ×1,000.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 27
FIGURE 27

Chronic interstitial nephritis in a patient with disseminated infection. The glomeruli are not involved. Hematoxylin and eosin stain was used. Magnification, ×200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 28
FIGURE 28

Renal tubule with necrosis, infected with . The granular material filling the lumen represents microsporidial spores and necrotic cellular debris. Hematoxylin and eosin stain was used. Magnification, ×400.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 29
FIGURE 29

The lumen of this renal tubule is filled with . spores. Steiner stain was used. Magnification, ×200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 30
FIGURE 30

Microsporidial prostate abscess. The abscess material contained numerous spores which cannot be seen with routine hematoxylin and eosin staining. Hematoxylin and eosin stain was used. Magnification, ×200.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 31
FIGURE 31

Muscle fiber with a sporont of containing several sporoblasts. The sporont has a thick, electron dense pansporoblastic membrane (large arrows). Sporoblasts have a nucleus (N), electron-dense exospore (arrowheads), and a coiled polar tubule (small arrows) with 9 to 12 turns. (Reprinted with permission from L. S. Adelman and from J. Alroy, Department of Pathology, Tufts University School of Medicine, Boston, Mass.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 32
FIGURE 32

Section of brain tissue from a rabbit with torticollis, showing a microgranuloma with a central focus of necrosis but without detectable spores. Hematoxylin and eosin stain was used. Magnification, ×250. (Reprinted with permission from F. Guscetti, University of Zurich, Zurich, Switzerland.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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Image of FIGURE 33
FIGURE 33

Section of brain tissue from an emperor tamarin with lethal neonatal encephalitozoonosis. Immunohistochemical detection of spores in a granuloma was done with a polyclonal rabbit antibody to . Visualization was by the labeled streptavidin-biotin method. Magnification, ×250. (Reprinted with permission from F. Guscetti, University of Zurich, Zurich, Switzerland.)

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
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References

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Tables

Generic image for table
TABLE 1

Microsporidial species pathogenic in humans and their clinical manifestations

Species not yet classified.

is a collective generic name for microsporidia that cannot be classified because available information is not sufficient.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
Generic image for table
TABLE 2

Sites of detection of microsporidia

−, not reported; +, case report(s) of detection; ++, consistently reliable specimens for detection of microsporidia.

Pituitary gland.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
Generic image for table
TABLE 3

Diagnostic techniques and identification of microsporidia

The table has been modified from Weber et al. (1994a).

+, recommended;(+), useful, but not widely available (research technique or commercially not available); −, not recommended.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
Generic image for table
TABLE 4

Chromotrope stains and their modifications

Laboratory procedures are described in the appendix to this chapter.

FEA, formalin-ethyl acetate; SAF, sodium acetate-acetic acid-formalin.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10
Generic image for table
TABLE 5

Murine monoclonal antibodies with diagnostic potential for the detection of microsporidia in humans

MAbs, monoclonal antibodies.

Citation: Weber R, Schwartz D, Deplazes P. 1999. Laboratory Diagnosis of Microsporidiosis, p 315-362. In Wittner M, Weiss L (ed), The Microsporidia and Microsporidiosis. ASM Press, Washington, DC. doi: 10.1128/9781555818227.ch10

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