Infection disease

I2 I2 洪俊斌洪俊斌

CASE11CASE11 A 32-year-old woman with pain in her right eye was seen by her oA 32-year-old woman with pain in her right eye was seen by her o

phthalmologist, who diagnosed a corneal ulcer in the affected eyphthalmologist, who diagnosed a corneal ulcer in the affected eye. Her physical examination was unremarkable otherwise; she hae. Her physical examination was unremarkable otherwise; she had no history of eye problems and had recently been examined by d no history of eye problems and had recently been examined by her primary-care physician, who declared her to be in good healther primary-care physician, who declared her to be in good health.h.

Bacterial cultures taken from a discharge from her eye were negBacterial cultures taken from a discharge from her eye were neg

ative. Viral cultures were also negative. Specimens were collecteative. Viral cultures were also negative. Specimens were collected for culture for free-living amebae. No amebic parasite was recod for culture for free-living amebae. No amebic parasite was recovered in culture. Figure 11.1 shows the parasite in a smear stainevered in culture. Figure 11.1 shows the parasite in a smear stained by the chromotrope Gram stain method. The Warthin-Starry sild by the chromotrope Gram stain method. The Warthin-Starry silver stain was used to confirm the diagnosis.ver stain was used to confirm the diagnosis.

PicturePicture

Chromotrope Gram stain methodChromotrope Gram stain methodIn this method, the samples are stained in heaIn this method, the samples are stained in heated (50 degrees C to 55 degrees C) solutions of ted (50 degrees C to 55 degrees C) solutions of crystal violet and iodinecrystal violet and iodine used in Gram's stain, f used in Gram's stain, followed by a modified ollowed by a modified chromotrope solutionchromotrope solution (heated to 50 degrees C to 55 degrees C). The (heated to 50 degrees C to 55 degrees C). The modified stain is composed of modified stain is composed of chromotrope 2chromotrope 2R (1%), fast green (0.15%), and phosphotungstR (1%), fast green (0.15%), and phosphotungstic acid (0.25%).ic acid (0.25%). Gram-chromotrope" staining technique can bGram-chromotrope" staining technique can be used to detect e used to detect microsporidian sporesmicrosporidian spores in cli in clinical specimens(stool).nical specimens(stool).

How to thinkHow to thinkRight eye corneal ulcerRight eye corneal ulcerImmunocompetent patientImmunocompetent patientChromotrope Gram stain(+)Chromotrope Gram stain(+)The Warthin-Starry silver stain(+)The Warthin-Starry silver stain(+)In microscopic view: Many spores like mIn microscopic view: Many spores like materials was seen in this specien: blue or aterials was seen in this specien: blue or blackblackMicrosporidian spores was highly suspeMicrosporidian spores was highly suspectedcted

QUESTIONSQUESTIONS Which group of Which group of obligate intracellular proobligate intracellular pro

tozoan parasitestozoan parasites fits the description gi fits the description given and might be causing this infection ven and might be causing this infection in this immunocompetent woman?in this immunocompetent woman?

ANSANSMicrosporidiaMicrosporidia are obligately intracellular, spor are obligately intracellular, spore-forming parasites belonging to the phylum e-forming parasites belonging to the phylum Microspora and the order Microsporida. They aMicrospora and the order Microsporida. They are eukaryotic organisms containing 70S ribosore eukaryotic organisms containing 70S ribosomes but lacking mitochondria, peroxisomes, Gmes but lacking mitochondria, peroxisomes, Golgi membranes, and other typically eukaryotiolgi membranes, and other typically eukaryotic organelles. The phylum Microspora contains c organelles. The phylum Microspora contains over 1000 species. The host range is extensive over 1000 species. The host range is extensive and includes honeybees, fish, mosquitoes, fleaand includes honeybees, fish, mosquitoes, fleas, grasshoppers, rodents, rabbits, and other fus, grasshoppers, rodents, rabbits, and other fur-bearing mammals.r-bearing mammals.

QUESTIONSQUESTIONS

List several members of this group List several members of this group that may be causing this patient's that may be causing this patient's infection infection

ANSANSseven genera seven genera

Nosema corneaeNosema corneae Vittaforma ocularumVittaforma ocularum EnterocytozoonEnterocytozoon EncephalitozoonEncephalitozoon PleistophoraPleistophora TrachipleistophoraTrachipleistophora BrachiolaBrachiola

QUESTIONSQUESTIONS

Which laboratory procedure may be Which laboratory procedure may be used to identify the parasite to used to identify the parasite to genus level?genus level?

ANSANS

Polymerase chain reaction is available in Polymerase chain reaction is available in some research laboratories and can be usome research laboratories and can be used to diagnose the following 3 sed to diagnose the following 3 EncephaEncephalitozoonlitozoon species: (1) species: (1) E bieneusi,E bieneusi, (2) (2) V corV corneae,neae, and (3) and (3) NosemaNosema species species

QUESTIONSQUESTIONS

What are the structures seen in the What are the structures seen in the smear?smear?

ANSANSThe spore wall should stain The spore wall should stain pinkish to red, with the pinkish to red, with the interior of the spore being interior of the spore being clear or perhaps showing a clear or perhaps showing a horizontal or diagonal horizontal or diagonal stripe, which represents stripe, which represents the polar tube. The the polar tube. The background will appear background will appear green or blue, depending green or blue, depending on the methodon the methodArrow shows coiled polarArrow shows coiled polar

filament.filament.Silver stainSilver stain

Nosema connori

QUESTIONSQUESTIONS

Which other laboratory procedWhich other laboratory procedures are available to diagnose iures are available to diagnose infections with this group of pronfections with this group of protozoans?tozoans?

ANSANSBody fluid specimensBody fluid specimens– Microscopic examination of stained stool samples Microscopic examination of stained stool samples – Examine stools for other parasites (ova and parasite exam) anExamine stools for other parasites (ova and parasite exam) an

d bacteria. d bacteria. – The modified trichrome stain (chromotrope 2R) commonly is The modified trichrome stain (chromotrope 2R) commonly is

used to detect microsporidia in urine, stool, or mucus. used to detect microsporidia in urine, stool, or mucus. – The rapid Gram chromotrope method can be performed morThe rapid Gram chromotrope method can be performed mor

e quickly (about 11 min) and combines the chromotrope mete quickly (about 11 min) and combines the chromotrope method with a Gram-staining step. The spores stain dark violet, ahod with a Gram-staining step. The spores stain dark violet, and the equatorial stripe is enhanced. nd the equatorial stripe is enhanced.

– Cytologic and histologic examinations are quite useful in diagCytologic and histologic examinations are quite useful in diagnosis. A conjunctival scraping or swab frequently can reveal tnosis. A conjunctival scraping or swab frequently can reveal the organism after a Gram stain (organisms usually stain grahe organism after a Gram stain (organisms usually stain gram-positive) or chromotrope stain.m-positive) or chromotrope stain.

– Stains that are used to detect microsporidia includStains that are used to detect microsporidia include the Brown Brenn Gram stain, Warthin-Starry silvee the Brown Brenn Gram stain, Warthin-Starry silver stain, Giemsa, and trichrome blue.r stain, Giemsa, and trichrome blue.

– Fluorochrome stains, including calcofluor white anFluorochrome stains, including calcofluor white and uvitex 2B, have a high affinity for chitin. d uvitex 2B, have a high affinity for chitin.

– Microsporidia stain poorly with hematoxylin-eosin.Microsporidia stain poorly with hematoxylin-eosin.Immunofluorescence assays using monoclonal or polyImmunofluorescence assays using monoclonal or polyclonal antibodies are available in some research laboclonal antibodies are available in some research laboratories and can be performed on most specimens, ratories and can be performed on most specimens,

QUESTIONSQUESTIONS

Describe the pathogenesis of this Describe the pathogenesis of this infectioninfection

Humans acquire infection through ingestion or inhalatHumans acquire infection through ingestion or inhalation of spores. Studies isolating Encephalitozoon speciion of spores. Studies isolating Encephalitozoon species in the urinary tract in those with disseminated infeces in the urinary tract in those with disseminated infections suggest that sexual transmission is possible. The tions suggest that sexual transmission is possible. The spore is the infective form. It is environmentally resistspore is the infective form. It is environmentally resistant and can survive for several months. The spore extrant and can survive for several months. The spore extrudes its polar tubule and injects the infective sporoplaudes its polar tubule and injects the infective sporoplasm into the host cell. Once inside the cell, it multiplies sm into the host cell. Once inside the cell, it multiplies by binary fission or schizogony. Development can occby binary fission or schizogony. Development can occur directly inside the host cell cytoplasm or inside parur directly inside the host cell cytoplasm or inside parasitophorous vacuoles. As mature spores develop and asitophorous vacuoles. As mature spores develop and accumulate, the cell expands and eventually ruptures, accumulate, the cell expands and eventually ruptures, releasing the spores. releasing the spores.

QUESTIONSQUESTIONS

How does this infection differ in immuHow does this infection differ in immunocompromised (AIDS) patients?nocompromised (AIDS) patients?

ANSANS– Immunocompetent patients with ocular involvement due to Immunocompetent patients with ocular involvement due to

EncephalitozoonEncephalitozoon species can have conjunctival hyperemia. A species can have conjunctival hyperemia. A slit lamp examination can reveal keratoconjunctivitis, which islit lamp examination can reveal keratoconjunctivitis, which is characterized by diffuse, superficial, punctate keratopathy. Is characterized by diffuse, superficial, punctate keratopathy. Infection frequently is bilateral.nfection frequently is bilateral.

– Corneal ulceration is rare in patients with AIDS. Retinal involvCorneal ulceration is rare in patients with AIDS. Retinal involvement is not reported. But chronic diarrhea and wasting can ement is not reported. But chronic diarrhea and wasting can be found in some immunocompromised patients.be found in some immunocompromised patients.

– The symptom of corneal ulcer in immunocompromised patieThe symptom of corneal ulcer in immunocompromised patients is mild and the destruction of corneal is less serious than ints is mild and the destruction of corneal is less serious than immunocompetent pateints due the low response of immunitmmunocompetent pateints due the low response of immunity,so the ocular symptoms are always neglected in some HIV iy,so the ocular symptoms are always neglected in some HIV infected patients.nfected patients.

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