option, but some organisms may be missed. Another testing option for three pooled specimens is to perform a single concentration and three separate permanent stained smears. This approach would probably increase the yield of intestinal protozoa over the previous option. Another suggestion involves placing a sample of each of three stools into a single vial of preservative. This collection approach would require only a single vial, but it is very likely that the vial would be overfilled and that mixing and the ratio of fixative to stool would be inaccurate.
Fecal immunoassays could also be used for G. lamblia, the E. histolytica/E. dispar group, E. histolytica, or Cryptosporidium spp. However, information about patient history is rarely received with the clinical specimen; fecal testing based on the risk group or recent activity of the patient is impossible without sufficient information. Testing by immunoassay procedures should be performed on request; however, client education is critical for successful implementation of this approach. Collection and testing options and their pros and cons can be seen in Tables 3.1 and 3.2.
The Clinical Laboratory Standards Institute/National Committee for Clinical Laboratory Standards document Procedures for the Recovery and Identification of Parasites from the Intestinal Tract (approved guideline M28-2A) was updated in 2005. Various stool collection, processing, and testing options are also included in that publication.
In summary, laboratories performing diagnostic parasitology testing must decide on the test methods that are relevant for their own operations based on a number of variables mentioned above. It is unrealistic to assume or state that one approach is applicable for every laboratory; however, it is important to thoroughly understand the options within your test menu and to convey this information to your clients once your approach has been selected for implementation. Prior discussion with clients, written educational memos, meetings, and examples of revised report formats are highly recommended prior to implementation. Based on available testing options, it is recommended that laboratories include both the O&P exam and various fecal immunoassays in their test menus.
Collection Times
A series of three specimens should be collected on separate days. If possible, the specimens should be collected every other day; otherwise, the series of three specimens should be collected within no more than 10 days. If a series of six specimens is requested, these specimens should also be collected on separate days or within no more than 14 days. Many organisms, particularly the intestinal protozoa, are shed sporadically and do not appear in the stool in consistent numbers on a daily basis; thus, the series of two (minimum) or three specimens is recommended for an adequate examination. Multiple specimens should not be collected from the same patient on the same day. One possible exception would be a patient who has severe, watery diarrhea such that any organisms present might be missed because of a tremendous dilution factor related to fluid loss. These specimens should be accepted only after consultation with the physician. It is also not recommended for the three specimens to be submitted one each day for three consecutive days; however, use of this collection time frame would not be sufficient to reject the specimens.
Although three stool specimens are recommended, laboratories have been more willing to accept two specimens, primarily because of cost savings and the assumption that if the patient is symptomatic, the presence of any organisms is likely to be confirmed by testing two specimens. However, it is important that clients understand the pros and cons of the two approaches. Both collection approaches are being used by diagnostic laboratories.
Posttherapy Collection
Patients who have received treatment for a protozoan infection should be checked 3 to 4 weeks after therapy, and those treated for Taenia infections should be checked 5 to 6 weeks after therapy; these recommendations have been used for many years. If the patient remains asymptomatic, the posttherapy specimens may not be collected, often as a cost containment measure. If the patient becomes symptomatic again, additional specimens can be submitted. If fecal immunoassays are ordered and parasites are present, the patient should be tested 7 to 10 days posttherapy. It usually takes approximately a week for antigen to be eliminated from the stool. It is important to remember that if the post-therapy specimens are collected too long after therapy, the presence of parasites or parasite antigen may represent a reinfection.
Specimen Type, Stability, and Need for Preservation
Fresh specimens are required for the recovery of motile trophozoites (amebae, flagellates, or ciliates). The protozoan trophozoite stage is found in patients with diarrhea; the gastrointestinal tract contents are moving through the system too rapidly for cyst formation to occur. Once the stool specimen is passed from the body, trophozoites do not encyst but may disintegrate if not examined or preserved within a short time after passage. However, most helminth eggs and larvae, coccidian oocysts, and microsporidian spores can survive for extended periods. Since it is impossible to know which organisms might be present, it is recommended that the most conservative time limits be used for parasite preservation and recovery. Liquid specimens should be examined within 30 min of passage, not 30 min from the time they reach the laboratory. If this general time recommendation of 30 min is not possible, the specimen should be placed in one of the available fixatives. Soft (semiformed) specimens may contain a mixture of protozoan trophozoites and cysts and should be examined within 1 h of passage; again, if this time frame is not possible, preservatives should be used. Immediate examination of formed specimens is not as critical; in fact, if the specimen is examined any time within 24 h after passage, the protozoan cysts should still be intact (Table 3.3).
Currently, fresh or frozen fecal specimens are required for the following fecal immunoassays (either as a single-organism test or when combined with other organisms such as G. lamblia or Cryptosporidium spp.): E. histolytica/E. dispar group and E. histolytica.
Summary: Collection of Fresh Stool Specimens
1. Occupational Safety and Health Act regulations (Standard Precautions) should be used for handling all specimens.
2. Interfering substances (e.g., barium, mineral oil, or antibiotics) should be avoided when stool specimens are collected.
3. Contamination with urine or water should be avoided.
4. Recommendation for collection: two (minimum) or three specimens collected, one every other day or within a 10-day time frame; see Table 3.1 for options and pros/cons.
5. Liquid stool should be examined or preserved within 30 min of passage (trophozoites). Soft stool should be examined or preserved within 1 h of passage (trophozoites and cysts*). Formed stool should be examined or preserved within 24 h of passage.
6. Fresh or frozen fecal specimens are required for the following fecal immunoassays (either as a single-organism test or combined with other organisms such as G. lamblia or Cryptosporidium spp.): E. histolytica/E. dispar group and E. histolytica. Fresh, frozen, or preserved specimens can be used for G. lamblia and Cryptosporidium spp.; specimens submitted in Cary-Blair transport medium are also acceptable. Note: Due to the freeze-thaw cycle, frozen specimens cannot be used for the FA procedures for G. lamblia and/or Cryptosporidium spp. (destruction of the actual
