TABLE 1. Most Common Clinical Presentation of Human Pathogenic Microsporidia

In 1973, Ashton and Wirasinha² published the first report of a well-documented case of human ocular microsporidial infection, which had been identified in the cornea of a young boy who had been injured by a goat 6 years earlier. Since then, cases of ocular microsporidiosis have been reported in both immunocompetent persons^3–5 and immunocompromised persons.^3,6–8 Ocular microsporidial infection typically presents either as chronic epithelial keratoconjunctivitis or as deep stromal keratitis, but one case of sclerouveitis with retinal detachment has also been reported.⁹

As eukaryotes lacking identifiable mitochondria, microsporidia were later considered to be ancient protists that had diverged before the evolution of mitochondria. This theory placed microsporidia together with other amitochondriate protists, and they were collectively known as Archezoa.¹⁰ Early molecular genetic analyses of ribosomal RNA also supported the Archezoan hypothesis, as did the absence of organelles such as peroxisomes and Golgi apparati.

However, further analysis of microsporidian phylogeny contradicts the notion that microsporidia derive from a primitive origin. Recent scientific discoveries, including the sequencing of the full genome of Encephalitozoon cuniculi, support the relation of microsporidia to fungi.^11,12 Rather than emerging from primitive origins, microsporidia may be highly reduced, sophisticated fungi that contained mitochondria at one time.^10,13

Although the first microsporidial organism was not discovered until the mid-1800s, since then approximately 150 genera with more than 1,200 different species have been described within the phylum Microspora. Fourteen microsporidian species have been identified as human pathogens (Table 1): Brachiola algerae, B. connori, B. vesicularum, Encephalitozoon cuniculi, E. hellem, E. intestinalis, Enterocytozoon bieneusi, Microsporidium africanum, M. ceylonensis, N. ocularum, Pleistophora ronneafiei, Trachipleistophora anthropophthera, T. hominis, and Vittaforma corneae. Species from several genera have caused documented ocular infection (Table 1), with E. hellem being the most commonly identified organism in reported cases.

Because both domestic and wild animals are common hosts for microsporidia, zoonotic transmission from animal reservoirs is likely. However, recent evidence suggests that some strains of microsporidia may also be waterborne pathogens. Cotte et al¹⁴ reported an outbreak of intestinal microsporidiosis that was linked to local water distribution subsystems. In 1998, scientists in Arizona identified three different pathogenic species of microsporidia in surface water, groundwater, and tertiary effluent by using polymerase chain reaction (PCR) amplification.¹⁵ That same year, Enriquez et al¹⁶ reported a link between community water sources in Mexico and the presence of E. intestinalis spores in the feces of local residents.

Molecular methods have similarly confirmed the presence of E. intestinalis in water used for drinking.¹⁷ Although the evidence is accumulating for waterborne transmission of microsporidiosis, respiratory tract infection with microsporidia suggests potential airborne transmission and ocular infection may be caused by autoinoculation with contaminated fingers.

Although microsporidia can infect immunocompetent persons, microsporidiosis is more common in immunosuppressed populations. In 1985, E. bieneusi was detected in a patient with AIDS with chronic diarrhea.¹ Since then, several published studies have identified microsporidial infection as a cause of chronic diarrhea in patients with AIDS.¹⁸ Ocular microsporidial infection has also been reported in both immunocompromised and immunocompetent patients.^4–8,19,20 Risk factors for ocular disease include AIDS, prednisone use, and trauma. However, in 2003, Lewis et al⁴ reported a case of microsporidial keratoconjunctivitis in an immunocompetent patient without a history of trauma or contact lens wear. A case of corneal microsporidiosis has also been reported in a healthy patient who had undergone laser in situ keratomileusis 3 years earlier.²¹

The production and distribution of the microsporidian spore define the life cycle of microsporidia. Microsporidian spores are unicellular, ranging in size from 1 μm to 10 μm. The size and morphologic characteristics of these spores are the criteria for the differentiation and classification of the various species of microsporidia. The complex structure of the cell wall of the microsporidian spore provides resistance to many environmental stresses. Each mature microsporidian spore comprises an extrusion apparatus consisting of a polaroplast, a polar tube or filament, and a posterior vacuole. The unique microsporidial mechanism of infection involves the rapid extrusion and uncoiling of the polar tube through the anterior end of the spore. The tube can then penetrate an adjacent host cell (Fig. 1) and inoculate the host cytoplasm directly with its infective sporoplasm.^22,23

Host cells can also be infected with microsporidian spores by ingesting spores via phagocytosis. Microsporidia have been identified as persisting within macrophages. Confined to a specialized compartment known as a parasitophorous vacuole, microsporidia can avoid the destructive enzymes of a macrophage's lysosomal network. A spore can also escape hostile lysosomal enzymes by using its polar tube to penetrate the wall of the lysosome and injecting its sporoplasm into the free cytoplasm of its host macrophage. Once inside the cytoplasm of the host cell, the microsporidium goes through two major life cycle stages: multiplication (merogony or schizogony) and maturation (sporogony). These developmental phases are species- and host-specific.

Many microsporidian genera multiply and mature while floating free among the intracellular organelles within the host cell's cytoplasm. Other genera develop within either a host-produced vacuole (Encephalitozoon) (Fig. 2) or a parasite-produced envelope (Pleistophora).

Numerous animal models exist for the human pathogenic species of microsporidia. Nonhuman primates, pigs, mice, rabbits, fish, birds, and insects have all been used as experimental hosts for microsporidia, but most of the immunologic information about microsporidiosis is derived from research with mice.²⁵ Studies of cell-mediated immunity suggest that T lymphocytes are key players in the defense against microsporidiosis. Evidence from experiments with severe combined immunodeficient mice points to CD8⁺ T lymphocytes as central to the prevention of microsporidial infection.^24,26 However, the mechanisms of both cell-mediated and humoral immunity may vary, depending on the species of both the parasite and the host as well as the route of infection.

E. hellem is the most commonly reported cause of ocular infection. In fact, it was first isolated from the corneas of patients with AIDS and keratoconjunctivitis. Athymic mice can serve as an animal model for this disease after they are infected with E. hellem spores either by inhalation or by intraperitoneal injection. Although E. hellem is a common cause of microsporidial keratoconjunctivitis, it is primarily a respiratory pathogen that causes disseminated disease. Athymic mice can also serve as a model for infection with another reported ocular pathogen, V. corneae.

The role of humoral immunity in human microsporidiosis is unclear. Studies of antibody responses to microsporidial infection in humans have produced variable results. This variability may be attributable to differences in the immune status of the populations being studied, subclinical microsporidial infection, or cross reactivity with other pathogens. Although a specific antibody response may contribute to host resistance to microsporidial infection, antibodies alone are not protective. In fact, in some domestic dogs, the antibody response to microsporidiosis may contribute to disease by causing immune complex renal failure.²⁴

In the first reported case of ocular microsporidiosis, Ashton and Wirasinha² described the histopathologic characteristics of spores within the cornea as “refractile oval bodies” having an average dimension of 3.5 × 1.5 μm. They reported that microsporidia from ocular tissue can be well visualized with Giemsa staining, but that Giemsa stain is less useful with stool specimens because of the presence of debris and artifact material.

On Gram's stain, microsporidia will stain weakly positive. Spores can also be visualized using acid-fast stains or phase-contrast microscopy, but they are sometimes missed on routine hematoxylin-and-eosin stain. If microsporidial infection is suspected, a modified trichrome stain is quite reliable and usually readily available. Chemofluorescent agents such as Calcofluor White 2MR (American Cyanamid Corp, Princeton, NJ) can also enhance detection sensitivity in cases missed by modified trichrome staining.²⁸

Electron microscopy can identify microsporidia when traditional light microscopy fails to reveal spores. Although ultrastructural detail provided by electron microscopy can help distinguish between microsporidia of different genera, it is time consuming and not always readily available. Molecular methods of identifying microsporidia that use PCR have been incorporated in research, but their clinical application is not yet widespread. In one recent case report, a PCR-based assay was used to facilitate the diagnosis of microsporidial keratitis in an HIV-positive patient.²⁹ PCR-based diagnostic assays hold much promise for improvement in the sensitivity of tests for microsporidiosis.

In 1993, there were at least two reports about the treatment of microsporidial keratoconjunctivitis with topical fumagillin.^30,31 This crystalline antibiotic is produced from the fungus Aspergillus fumigatus. Although it originally was used to treat microsporidiosis in honeybees infected with N. apis, a fumagillin analogue has been studied as an angiogenesis inhibitor and as a potential anticancer agent.³² Topical fumagillin appears to be the treatment of choice for microsporidial keratoconjunctivitis, but its systemic use has been limited because of the medication's side effect of bone marrow toxicity (e.g., thrombocytopenia and neutropenia). Nonetheless, oral fumagillin is a promising treatment of recalcitrant intestinal E. bieneusi infection in patients with AIDS.³³

Albendazole is used to treat microsporidiosis because it inhibits microtubule polymerization and interferes with cellular division. It has been used to counteract infections with echinococcus, Taenia solium, and Strongyloides stercoralis. Albendazole is administered orally and has shown particular efficacy in treating E. intestinalis infection.³² For ocular microsporidiosis, albendazole is effective in both immunocompromised and immunocompetent patients. The side effects of albendazole are usually reversible but can include hepatotoxicity, neutropenia, and alopecia. In cases of ocular microsporidiosis when albendazole is ineffective, the antifungal agent itraconazole may also be considered.³⁴

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