Journal of Postgraduate Medicine Medknow Publications and Staff Society of Seth GS Medical College and KEM Hospital, Mumbai, India ISSN: 0022-3859 EISSN: 0972-2823 Vol. 48, Num. 3, 2002, pp. 217

Journal of Postgraduate Medicine, Vol. 48, Issue 3, 2002 pp. 217

Laboratory Diagnosis of Cryptosporidiosis

Mehta P

Department of Microbiology, Seth G. S. Medical College and K. E. M. Hospital, Parel, Mumbai 400070, India.

Code Number: jp02075

The diagnosis of Cryptosporidiosis in the laboratory is achieved by one of the following:

1. Demonstration of Cryptosporidium parvum oocysts in stool
2. Demonstration of Cryptosporidium in intestinal fluid or small bowel biopsy specimens
3. Demonstration of Cryptosporidium antigen in stool
4. Molecular methods

Demonstration of Cryptosporidium parvum Oocysts in Stool

Mature C. parvum oocysts recovered from stool are 4-6 mm in size, round and contain four sporozoites within the thick walled oocysts. These can be identified easily using the staining methods such as modified Kinyoun's acid-fast method; hot Safranin stain and fluorescent dyes such as auramine/carbol fuchsine fluorescence method.

To maximize recovery of oocysts, stool samples should be concentrated (formol ethyl acetate- FEA) prior to microscopic examination. Multiple stool samples (at least three) should be tested before a negative diagnostic interpretation is reported. However, some studies have shown that the first sample is usually enough to provide accurate diagnosis in 90% of the cases.¹

Demonstration of Cryptosporidium in Intestinal Fluid or Small Bowel Biopsy Specimens

This method is not used commonly now. It was used earlier when the staining and antigen detection methods were not available. Also, due to the patchy nature of the intestinal parasitic infection, false negatives are commonly encountered with this method.²

Demonstration of Cryptosporidium Antigen in Stool

ELISA: The specimens should not be concentrated prior to testing. This is a highly sensitive and specific technique, and is useful for screening large numbers of specimens in a short time period. Also, it does not rely on skills in microscopy. However, controls are necessary to determine the quality of commercially available reagents.

Immunofluorescence assay: This technique offers the highest combination of sensitivity and specificity and is considered the gold standard by many laboratories. However, it does not provide a permanent record such as a stained slide, which can be archived. The stool specimen should be concentrated using the FEA before using this test.

For antigen detection, commercially available kit sensitivities and specificities range from 66.3% to 100% and 93% to 100%, respectively.³

Molecular Methods

PCR is used to detect C. parvum in stool specimens. Faecal material must be stored in potassium dichromate or be frozen for detecting the DNA of the organism by PCR.

To conclude, in Indian set up, acid-fast staining methods can be used in most clinical laboratories. For greatest sensitivity and specificity, immunofluorescence microscopy is the method of choice (followed closely by ELISA). Molecular methods are to be used mainly as a research tool.

Mehta P

Department of Microbiology, Seth G. S. Medical College and K. E. M. Hospital, Parel, Mumbai 400070, India.

References

1. Centers for Disease Control & Prevention. Cryptosporidium: Key points for laboratory diagnosis I. 2002. http:/www.dpd.cdc.gov/dpdx/HTML/PDF_Files/Crypto_bench.pdf. [accessed on August 30, 2002]
2. Flanigan TP, Soave R. Cryptosporidiosis. Prog Clin Parasitol 1993;3:1-20.
3. Centers for Disease Control & Prevention, National Center for Infectious Diseases, Division of Parasitic Diseases. Cryptosporidiosis. 2002. http://www.dpd.cdc.gov/DPDx/HTML/Cryptosporidiosis.htm. [accessed on August 30, 2002]

Brief Report - Bioline Code: jp02062

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