Introduction

 

Microsporidia are minute, unicellular, obligate intracellular parasites, capable of invading any animal cell. They are characterized by the production of resistant spores that vary in size, depending on the species. They possess a unique organelle, the polar filament, which is coilded inside the spore and ensures the inoculation of the nonmotile infective agent into the host cell. Microsporidia are found in a variety of vertebrates and invertebrates and is considered a common opportunistic pathogen in human patients with AIDS.

Electron micrograph of an Enterocytozoon bieneusi spore. Arrows indicate the double rows of polar tubule coils in cross section which characterize a mature E. bieneusi spore.

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Agent

 

Microsporidia belong to the phylum Microspora in the subkingdom Protozoa. These obligate intraccellular parasites are considered ancient eukaryotes.

Microsporidia are currently classified based on ultrastructural featrues, including size, and morphology of their spores, developmental life cycle, number of coils of the polar tube, and host-parasite relationship.

 

Taxonomy of microsporidia parasites in man and their targets. Involvement of different organs occur in immunodeficient patients.

Family

Genus

Species

Main Localization

Pleistophoridae

 

 

Trachipleistophora

 

Pleistophora

hominis

anthropophthera

 

skeletal muscle

brain, heart, kidney, liver

skeletal muscle

Encephalitozoonidae

 

 

Encephalitozoon

 

 

cuniculi

hellem

intestinalis

brain, kidney, heart, trachea, adrenals, liver

cornea, conjunctiva, pulonary tract, genitourinary tract

small intestine, kidney, lung

Enterocytozoonidae Enterocytozoon bieneusi small intestine

Nosematidae

 

 

Nosema

Brachiola

 

Vittaforma

ocularum

vesicularum

connori

corneae

corneal stroma

skeletal muscle

lung, adrenals, diaphram, myocardium, liver, gastrointestinal tract

corneal stroma

Undetermined

 

Microsporidium

 

africanum

ceylonensis

corneal stroma

corneal stroma

Deportes-Livage, 2000

 

 

 

 

Synonyms

None

 

 

 

 

History of Discovery

 

Nosema bombycis, a parasite of silkworms, was the first organism to be identified as a microsporidia in 1857. The first well documented human case was reported in 1959, over one hundred years later. Microsporidia remained a rather uncommon human infection until the Human Immuno Defficiency Virus (HIV) pandemic with the first HIV/AIDS related case reported in 1985.

 

 

 

 

Clinical Presentation in Humans

 

The clinical presentation of microsporidiosis in humans if very diverse, varying according to the causal species and immune system of the host. Microsporidiosis is a rapidly growing opportunistic disease in humans mainly but not exclusively affecting immunocompromised patients with AIDS. The most common clinical manifestation of microsporidiosis is diarrhea.

Microsporidian Species Clinical Manifestation
Enterocytozoon bieneusi Diarrhea, acalculus cholecystitis
Encephalitozoon intestinalis (syn. Septata intestinalis) Infection of the GI tract causing diarrhea, and dissemination to ocular, genitourinary and respiratory tracts
Encephalitozoon hellem and Enchepalitozoon cuniculi Keratoconjunctivitis, infection of respiratory and genitourinary tract, disseminated infection
Vittaforma corneae (syn. Nosema corneum), Nosema spp. (N. connori and N. ocularum) Ocular infection
Trachipleistophora hominis and Pleistophora sp. Muscular infection
Microsporidium (M. ceylonensis and M. africanum) Infection of the cornea

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Transmission

 

Transmission electron micrograph of a microsporidian spore with an extruded polar tubule inserted into a eukaryotic cell. The spore injects the infective sporoplasms through its polar tubule.

Figure contributed by Dr. Massimo Scaglia, Laboratory of Clinical Parasitology, Institute of Infectious Diseases, University-IRCCS San Matteo, Pavia, Italy.

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Microsporidia form characterisitc unicellular spores that are environmentally resistant. As mentioned previously, the spore possesses a polar filament that pierces the target host cell and inoculates it with the sporoplasm, (infective agent of microsporidian spore). Conditions that promote germination vary widely between species, which suggests the organisms' great ability to adapt to their hosts and external environment. Conditions that promote spore discharge include pH shifts, dehydration followed by rehydration, various cations and anions, mucin or polyanions, hydrogen peroxide, radiation, and the calcium ionophore A2387. Inhibitors of spore discharge include magnesium chloride, ammonium chloride, low salt concentrations, sodium fluoride, ultraviolet light, temperatures >40°C, calcium channel antagonists, calmodulin inhibitors, cytochalasin D, demecolcine, and itraconazole. All microsporidia exhibit the same response to stimuli regardless of the specific activation channel. The response being and increase in intrasporal osmotic pressure. This results in an influx of water into the spore accompanied by swelling of the spolaroplast and posterior vacuole prior to spore discharge. The polar tube discharges from the anterior pole of the spore in an explosive reaction ocurring in less than 2 seconds and is thought to form a hollow tube by a process of eversion.

Deportes-Livage, 2000

 

 

 

 

Reservoir

 

Microsporidia have been found in vertebrates and invertebrates including myxosporidia, cnidaria, annelids, nematodes, arthropods, and molluscs. Human forms of the pathogen have been identified in dogs, birds, fish, rabbits, and lab rodents. Specifically, Encephalitozoonidae is widely distributed in mammals and birds. The onset of microsporidia linked to this species has been associated with exposure livestock, fowl and pets.

 

 

 


Vector

None

 

 

 

 

Incubation Period

Unknown

 

 

 

 

Morphology

 

Microsporidia are true eukaryotes possessing a nucleus, nuclear envelope, intracytoplasmic membrane system, and chromosome separation mitotic spindles as well as Golgi. They do not have centrioles or mitochondria.

Perhaps the defining characteristic for all microsporidia is the polar filament. Attached to the inside of the anterior end of the spore, the polar filament ensures enoclulation of the nonmotile infective agent (sporoplasm) into the host cell. The polar filament, also known as the polar tubule, forms between 4 and 30 coils around the sporoplasm within the spore, the actual number depending on the species. During germination, the polar filament quickly extends from the spore, forming a hollow tube that pierces the adjacent host cell. With the polar filament essentially functioning as a hypodermic needle, this process transports the sporoplasm directly into the host cell.



Scanning electron micrograph of a microsporidian spore with an extruded polar tubule inserted into a eukaryotic cell. The spore injects the infective sporoplasms through its polar tubule.
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Transmission electron micrograph showing ultrastructural features of a mature microsporidian spores.

Legend:
Pt: Portion of coiled polar tubule
Ex: Electron dense exospore
En: Electron lucent endospore
Pv: Posterior vacuole

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Life Cycle

 

1. The infective form of microsporidia is the resistant spore and it can survive for a long time in the environment .
2. The spore extrudes its polar tubule and infects the host cell .
3. The spore injects the infective sporoplasm into the eukaryotic host cell through the polar tubule .
4. Inside the cell, the sporoplasm undergoes extensive multiplication either by merogony (binary fission) or schizogony (multiple fission) . This development can occur either in direct contact with the host cell cytoplasm (e.g., E. bieneusi) or inside a vacuole termed parasitophorous vacuole (e.g., E. intestinalis).
5. Either free in the cytoplasm or inside a parasitophorous vacuole, microsporidia develop by sporogony to mature spores . During sporogony, a thick wall is formed around the spore, which provides resistance to adverse environmental conditions.
6. When the spores increase in number and completely fill the host cell cytoplasm, the cell membrane is disrupted and releases the spores to the surroundings . These free mature spores can infect new cells thus continuing the cycle.

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Scanning electron micrograph showing an eukaryotic cell bursting and releasing spores of Encephalitozoon hellem to the extracellular medium.

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Diagnostic Tests

 

Test
Utility
Urine Examination This is often positive in cases of microsporidia caused by microsporidia other than Enterocytozoonidae and should be done in all suspected microsporidia cases.
Stool Examination This is useful for gastrointestinal presentations. At least three stools should be examined. The combination of Chromotrope and chemofluorescent stains provides the highest sensitivity and specificity.
Endoscopy This should be considered for all patients with chronic diarrhea of greater than 2 months duration and negative stool and urine examinations. In this group endoscopy has yielded a diagnosis of microsporidia in up to 30% of patients. Tissue should be examined by a microsporidial stain. Touch preparations are useful for rapid diagnosis (within 24 hours). If microsporidia are demonstrated to invade the lamina propria, then urine examination should be repeated because Encephalitozoonidae are the most likely etiologic agents. In this setting albendazole has a high treatment efficacy.
PCR Available as a research technique. Species identification can be performed on stool, urine, and tissue samples.
Electron microscopy Provides identification of the species of microsporidia involved and is crucial for the identification of new species. It is essential for the characterization of microsporidia in unusual or new locations.
Conjunctival scrapings This has a high diagnostic yield in microsporidian keratoconjunctivitis. Urine examination should also be performed in suspected cases to screen for disseminated microsporidiosis.
Nasal scrapings This can be useful for the diagnosis of microsporidian sinusitis. Because most of the microsporidia associated with sinusitis are present in the kidneys, examination of urine should be routine for suspected sinusitis cases. If these tests are negative, biopsy of nasl mucosa may be useful for diagnosis.
Serology Not useful for diagnosis but may be useful for epidemiolgic surveys
Weiss, 1999

 

Stool smear stained with Chromotrope 2R containing Enterocytozoon bieneusi spores.

Black arrows indicate E. bieneusi spores with their belt-like stripe visible. Red arrow indicates an unidentified yeast. The yellow arrow indicates a vacuolated spore.

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Management and Therapy

 

Fumagilin and albendazole have demonstrated consistent activity agains microsporidia in vitro and in vivo. Albendazole is recommended as the primary drug treatment for all microsporidial infections.

Organism Drug Dosage*
Enterocytozoon bieneusi No effective reatment: albendazole has resulted in clinical improvement in up to 50% of patients. Oral fumagillin is promising.  

Encephalitzoonidae infection (systemic, sinusitis, encephalitis, hepatitis, etc.)

Enc. cuniculi

Enc. hellem

Enc. intestinalis

 

Albendazole

Albendazole

Albendazole

 

400 mg bid

400 mg bid

400 mg bid

Encephalitozoonidae keratoconjunctivitis

Fumagillin solution

Patients may also need albendazole if systemic infection is present

2 drops every 2 hr for 4 d. then 2 drops 4 times/d.
Trachipleistophora hominis Albendazole 400 mg bid
Nosema-like myositis (Brachiola vesiculatum) +/- itraconazole 400 mg qd

*The duration of treatment for microsporidia has not been established. Relapse of infection has occurred upon stopping treatment. Patients should be maintained on treatment for at least 4 weeks, and most patients should be on treatment indefinitely.

Weiss, 1999

 

 

 

 

Epidemiology



Monoclonal antibody-based immunofluorescence identification of Encephalitozoon hellem. Spores are present in a bronchoalveolar lavage sample of a 30 year-old AIDS patient from Georgia. Note the bright fluorescent spores, which have extruded their polar tubules.
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Research suggests that microsporidia is a common infection in man, however is self-limited or asymptomatic in normal hosts. As mentioned previously, microsporidia have gained more attention in their recent association with the AIDS pandemic. It is now recognized as a common pathogen in HIV immunocompramised patients. Reported prevalence rates in AIDS have varied between 2 and 70 percent depending on the population studied and the diagnostic technique employed.

 

 

 

 

Country Information

 

Cases of microsporidiosis have been reported in the developed and developing world. Identified countries include Argentina, Australia, Botswana, Brazil, Canada, Czech Republic, France, Germany, India, Italy, Japan, The Netherlands, New Zealand, Spain, Sri Lanka, Sweden, Switzerland, Thailand, Uganda, United Kingdom, United States of America, and Zambia.

 

 

 

 

Public Health and Prevention

 

There is currently no anti-parasitic agent identified for microsporidia. Although the direct transmission is unknown, it is likely that microsporidia are food or waterborne pathogens. Standard sanitary measures that prevent food and water contamination by animal feces or urine should decrease risk of infection. Hand washing and general hygiene practice should prevent infection of the conjunctiva or cornea.

 

 

 

 

Reference

 

http://www.dpd.cdc.gov/DPDx/HTML/ImageLibrary/Microsporidiosis_il.htm

http://www.dpd.cdc.gov/DPDx/HTML/Microsporidiosis.htm

Desportes-Livage, I. Biology of Microsporidia

Cryptosporidiosis and microsporidiosis, 2000

Series: Contributions to microbiology; v.6 2000

Weiss, L. Microsporidiosis

M. AIDS therapy 1999; (27): 336-49


Fedorko, D. Hijazi, Y. Application of Molecular Techniques to the Diagnosis of Microsporidial Infection

http://www.cdc.gov/ncidod/eid/vol2no3/fedorko.htm