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Cyclosporiasis

[Cyclospora cayetanensis]

Causal Agents

Cyclospora cayetanensis, a coccidian protozoan. It appears that all human cases are caused by this species.

References:
  1. Herwaldt BL. Cyclospora cayetanensis: a review, focusing on the outbreaks of cyclosporiasis in the 1990s. Clin Infect Dis 2000;31:1040-1057.
  2. Ortega YR, Gilman RH, Sterling CR. A new coccidian parasite (Apicomplexa: Eimeriidae) from humans. J Parasitol 1994;80:625-629.
  3. Pieniazek NJ, Herwaldt BL. Reevaluating the molecular taxonomy: Is human-associated Cyclospora a mammalian Eimeria species? Emerg Infect Dis 1997;3:381-383.

Life Cycle

lifecycle

When freshly passed in stools, the oocyst is not infective The number 1. (thus, direct fecal-oral transmission cannot occur; this differentiates Cyclospora from another important coccidian parasite, Cryptosporidium). In the environment The number 2., sporulation occurs after days or weeks at temperatures between 22°C to 32°C, resulting in division of the sporont into two sporocysts, each containing two elongate sporozoites The number 3.. Fresh produce and water can serve as vehicles for transmission The number 4. and the sporulated oocysts are ingested (in contaminated food or water) The number 5.. The oocysts excyst in the gastrointestinal tract, freeing the sporozoites which invade the epithelial cells of the small intestine The number 6.. Inside the cells they undergo asexual multiplication and sexual development to mature into oocysts, which will be shed in stools The number 7.. The potential mechanisms of contamination of food and water are still under investigation.

Geographic Distribution

Cyclosporiasis has been reported in many countries, but is most common in tropical and subtropical areas. Since 1990, at least 11 foodborne outbreaks of cyclosporiasis, affecting approximately 3600 persons, have been documented in the United States and Canada.

Clinical Presentation

After an average incubation period of 1 week, symptomatic infections typically manifest as watery diarrhea, which can be severe. Other symptoms include anorexia, weight loss, abdominal pain, nausea and vomiting, myalgias, low-grade fever, and fatigue. Untreated infections typically last for 10-12 weeks and may follow a relapsing course. Infections, especially in disease-endemic settings can be asymptomatic.

Cyclospora cayetanensis oocysts in wet mounts.

 

Cyclospora cayetanensis oocysts are spherical, 7.5-10 µm in diameter and unsporulated when passed in feces. Sporulation in the environment is temperature-dependent and may take one to several weeks for an infective oocyst to contain two sporocysts, each containing two sporozoites. Oocysts of C. cayetanensis will autofluoresce under ultraviolet (UV) microscopy.
	Figure A

Figure A: Oocyst of C. cayetanensis in an unstained wet mount. Image courtesy of the Oregon State Public Health Laboratory.

	Figure B

Figure B: Oocyst of C. cayetanensis in an unstained wet mount. Image courtesy of the Oregon State Public Health Laboratory.

	Figure C

Figure C: Oocyst of C. cayetanensis in an unstained wet mount of stool. Image taken at 1000x magnification.

	Figure D

Figure D: Oocyst of C. cayetanensis in an unstained wet mount of stool. Image taken at 1000x magnification.

C. cayetanensis oocysts stained with trichrome.

	Figure A

Figure A: Oocyst of C. cayetanensis? stained with trichrome; while the oocyst is visible, the staining characteristics are inadequate for a reliable diagnosis.

	Figure B

Figure B: Oocysts of C. cayetanensis stained with trichrome; while the oocyst is visible, the staining characteristics are inadequate for a reliable diagnosis.

C. cayetanensis oocysts viewed under ultraviolet (UV) microscopy.

	Figure A

Figure A: Oocyst of C. cayetanensis viewed under UV microscopy.

	Figure B

Figure B: Oocyst of C. cayetanensis viewed under UV microscopy.

	Figure C

Figure C: Oocyst of C. cayetanensis viewed under UV microscopy.

	Figure D

Figure D: Oocyst of C. cayetanensis viewed under UV microscopy.

	Figure E

Figure E: Oocyst of C. cayetanensis viewed under UV microscopy.

	Figure F

Figure F: Oocyst of C. cayetanensis viewed under UV microscopy.

C. cayetanensis oocysts stained with modified acid-fast.

	Figure A

Figure A: Oocysts of C. cayetanensis stained with modified acid-fast stain. Note the variability of staining in the four oocysts.

	Figure B

Figure B: Two oocysts of C. cayetanensis stained with modified acid-fast stain. Both oocysts failed to take up the carbol fuschin stain. Image courtesy of the Arizona State Public Health Laboratory.

	Figure C

Figure C: Oocysts of C. cayetanensis stained with modified acid-fast stain. Note the wrinkled edge and the lack of stain in the two oocysts. Image courtesy of the Arizona State Public Health Laboratory.

	Figure D

Figure D: Oocyst of C. cayetanensis stained with modified acid-fast stain.

	Figure E

Figure E: Oocysts of C. cayetanensis stained with modified acid-fast stain.

	Figure F

Figure F: Oocysts of C. cayetanensis stained with modified acid-fast stain.

C. cayetanensis oocysts stained with safranin (SAF).

	Figure A

Figure A: Oocyst of C. cayetanensis stained with safranin (SAF).

	Figure B

Figure B: Oocyst of C. cayetanensis stained with safranin (SAF).

	Figure C

Figure C: Oocyst of C. cayetanensis stained with safranin (SAF).

	Figure D

Figure D: Oocyst of C. cayetanensis stained with safranin (SAF).

	Figure E

Figure E: A pair of oocysts of C. cayetanensis stained with safranin (SAF).

	Figure F

Figure F: Oocyst of C. cayetanensis (yellow arrow) along with an oocyst of Cryptosporidium parvum (red arrow), stained with safranin (SAF). Cryptosporidium spp. also stain with the safranin and modified acid-fast stains.

C. cayetanensis oocysts viewed under differential interference contrast (DIC) microscopy.

	Figure A

Figure A: Oocyst of C. cayetanensis viewed under differential interference contrast (DIC) microscopy. The refractile globules are easily visible under DIC.

	Figure B

Figure B: Oocyst of C. cayetanensis viewed under differential interference contrast (DIC) microscopy. The refractile globules are easily visible under DIC.

	Figure C

Figure C: Oocyst of C. cayetanensis viewed under DIC microscopy. There are two sporocysts are visible in this image.

	Figure D

Figure D: Oocyst of C. cayetanensis viewed under DIC microscopy.

	Figure E

Figure E: A pair of oocysts of C. cayetanensis viewed under DIC microscopy.

	Figure F

Figure F: Rupturing oocyst of C. cayetanensis viewed under DIC microscopy. One sporocyst has has been released from the mature oocyst; the second sporocyst is still contained within the oocyst wall.

Laboratory Diagnosis

Currently, the most practical diagnostic method consists of the identification of oocysts in stool specimens by light microscopy. Other methods are also available or under investigation.

Specimen Processing

Specimens should be refrigerated and sent to the diagnostic laboratory as rapidly as possible. If it is not possible to send the specimen to the laboratory promptly, it should be preserved. Ideally, because a range of tests might be desired, each of which has different requirements of the specimen, the latter should be split in portions which should be respectively:

  • fixed in 10% formalin (for direct microscopy, concentration procedures, and preparation of stained smears);
  • fixed in 2.5% potassium dichromate (for sporulation assays and molecular diagnosis); and
  • frozen without fixation (for molecular diagnosis).

Note: Specimens fixed in sodium acetate-acetic acid formalin can be handled in the same manner as specimens fixed in formalin; however, specimens fixed in polyvinyl alcohol (PVA) are of limited value because they are not usable for concentration procedures.

Cyclospora oocysts can be excreted intermittently and in small numbers. Thus:

  • a single negative stool specimen does not rule out the diagnosis; three or more specimens at 2- or 3-day intervals may be required
  • concentration procedures should be used to maximize recovery of oocysts. The method most familiar to laboratorians is the formalin-ethyl acetate sedimentation technique (centrifuge for 10 minutes at 500 × g). Other methods can also be used (such as the Sheather's flotation procedure).
Microscopic Examination

The sediment can be examined microscopically with different techniques:

Sporulation Assay

Because of the morphologic similarity between freshly passed, unsporulated Cyclospora oocysts and blue-green algae (cyanobacterium-like bodies), it has been advocated that to confirm the diagnosis of cyclosporiasis, unfixed oocysts should be examined over a 2- to 3-week period for evidence of sporulation. This is accomplished by placing an aliquot of fresh stool in 2.5% potassium dichromate (which reduces bacterial overgrowth), and keeping it under observation for sporulation of the oocysts.*

*As the ability of laboratorians to accurately diagnose cyclosporiasis has improved, the need to do sporulation assays has decreased.

Procedure

Mix stool with 2-3 volumes of potassium dichromate (depending on the stool consistency) and agitate specimen occasionally during the incubation period. Leave an air space over the specimen, for example by turning the specimen tube on its side to increase surface area, to promote aeration. At room temperature, oocysts generally sporulate in 5-14 days, although the percentage of oocysts that sporulate can vary. If the number of oocysts is small, the material held in potassium dichromate can be centrifuged and the pellet examined. Prior to examination, the potassium dichromate solution should be diluted (it quenches fluorescence): add to the specimen an equal volume of distilled/deionized water; or remove the dichromate by centrifugation and washing (use distilled/deionized water or various solutions such as PBS, culture medium). Optimal examination of the specimen, using a wet mount, would include epifluorescence (to identify the oocysts) followed by DIC, phase contrast microscopy, or conventional bright-field microscopy (to identify the sporocysts/sporozoites). Sporulated mature oocysts contain two sporocysts, each of which contains two sporozoites. (Because the sporozoites are tightly packed inside the sporocysts, they are difficult to visualize.)

Sporulation of Cyclospora oocysts.

Sporulation of Cyclospora oocysts. The sequence shows, as observed by DIC microscopy of wet mounts: an oocyst passed in fresh stool (Day 0); sporulated oocysts at days 5 (Day 5) and 10 (Day 10), which both contain 2 sporocysts; and a ruptured oocyst (Rupture), with a sporocyst still inside the oocyst and the other sporocyst just outside—the coiled sporozoites are barely visible inside the sporocysts.

Molecular diagnosis

A nested PCR assay targeting the small-subunit ribosomal RNA has been developed.1  Preliminary results indicate that the Cyclospora-specific PCR primers cross react with some Eimeria species and the sensitivity of the assay may be low (62%).2

Agarose gel (2%) analysis of a PCR diagnostic test for detection of Cyclospora DNA. PCR was performed using nested primers CYCF1E/CYCR2B (first round) and CYCF3E/CYCR4B (second round).1,2

  • Lane S: Molecular base pair standard (100-bp ladder). Black arrows show the size of standard bands.
  • Lane 1: Cyclospora positive fecal specimen. The red arrow shows the diagnostic band for Cyclospora cayetanensis (size: 308 bp).
Reference:

Eberhard ML, Pieniazek NJ, and Arrowood MJ. Laboratory diagnosis of Cyclospora infections. Arch Pathol Lab Med 1997;121:792-797.

Agarose gel for a PCR diagnostic test for detection of Cyclospora DNA

Treatment Information

Trimethoprim-sulfamethoxazole (TMP-SMX), or Bactrim*, Septra*, or Cotrim*, is the treatment of choice. The typical regimen for immunocompetent adults is TMP 160 mg plus SMX 800 mg (one double-strength tablet), orally, twice a day, for 7-10 days. HIV-infected patients may need longer courses of therapy.

No highly effective alternatives have been identified for persons who are allergic to (or are intolerant of) TMP-SMX. Approaches to consider for such persons include observation and symptomatic treatment, use of an antibiotic whose effectiveness against Cyclospora is based on limited data, or desensitization to TMP-SMX. The latter approach should be considered only for selected patients who require treatment, have been evaluated by an allergist, and do not have a life-threatening allergy.

Anecdotal or unpublished data suggest that the following drugs are ineffective: albendazole, trimethoprim (when used as a single agent), azithromycin, nalidixic acid, tinidazole, metronidazole, quinacrine, tetracycline, doxycycline, and diloxanide furoate. Although data from a small study among HIV-infected patients in Haiti suggested that ciprofloxacin might have modest activity against Cyclospora, substantial anecdotal experience among many immunocompetent persons suggests that ciprofloxacin is ineffective.

Trimethoprim–sulfamethoxazole

Trimethoprim–sulfamethoxazole (TMP–SMX) is available for human use in the United States.

Trimethoprim–sulfamethoxazole (TMP–SMX) is in pregnancy category C. TMP–SMX should be used during pregnancy only if the potential benefit justifies the potential risk to the fetus. TMP-SMX should be avoided near-term because of the potential for hyperbilirubinemia and kernicterus in the newborn.

Pregnancy Category C: Either studies in animals have revealed adverse effects on the fetus (teratogenic or embryocidal, or other) and there are no controlled studies in women or studies in women and animals are not available. Drugs should be given only if the potential benefit justifies the potential risk to the fetus.

Trimethoprim–sulfamethoxazole (TMP–SMX) is excreted in breast milk. TMP–SMX generally is compatible with breastfeeding of healthy, full-term infants after the newborn period. However, TMP-SMX generally should be avoided by women when nursing infants who are premature, jaundiced, ill, or stressed, or who have glucose-6-phosphate dehydrogenase deficiency.

The safety of trimethoprim–sulfamethoxazole (TMP–SMX) in children has not been systematically evaluated. Use in children less than 2 months of age generally is not recommended.

 

DPDx is an education resource designed for health professionals and laboratory scientists. For an overview including prevention and control visit www.cdc.gov/parasites/.

  • Page last reviewed: May 3, 2016
  • Page last updated: May 3, 2016
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