A 6-year old boy was brought into the clinic after complaining of diarrhea over a two week period. His other symptoms were unremarkable and he had no relevant travel history.
Complete stool examinations were performed (O&P exam: direct wet mount, concentration, and permanent stained slide).
Laboratory results revealed the following:
These images were photographed from permanent stained smears stained with trichrome. Both organisms measure approximately 9 by 12 microns.
Please identify the organisms.
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Answer and Discussion of Quiz #23
The images presented in Diagnostic Quiz #23 are the following:
Comment: D. fragilis was first seen in 1909 and described in 1918. On the basis of electron microscopy studies, it has been reclassified as an ameba-flagellate rather than an ameba and is closely related to Histomonas and Trichomonas spp. It has a cosmopolitan distribution, and past surveys provide incidence rates of from 1.4 to 19%. Much higher incidence figures have been reported for mental institution inmates, missionaries, and Indians in Arizona. D. fragilis tends to be common in some pediatric populations, and in some studies, incidence figures are higher for patients under 20 years of age. No cyst form has been identified for this organism.
Although its pathogenic status is still not totally defined, D. fragilis has been associated with a wide range of symptoms. Case reports of children infected with D. fragilis reveal a number of symptoms, including intermittent diarrhea, abdominal pain, nausea, anorexia, malaise, fatigue, poor weight gain, and unexplained eosinophilia. The most common symptoms in patients infected with this parasite appear to be intermittent diarrhea and fatigue. In some patients, both the organism and the symptoms may persist or reappear until appropriate treatment is initiated.
Diagnosis of D. fragilis infections depends on proper collection and processing techniques (a minimum of three fecal specimens). Although survival time for this parasite has been reported as 24 to 48 h, the survival time in terms of morphology is limited, and stool specimens must be examined immediately or preserved in a suitable fixative soon after defecation. It is particularly important that permanently stained smears of stool material be examined with an oil immersion lens (1,000 x). These organisms have been recovered in formed stool; thus, a permanent stained smear must be prepared for every stool submitted for a parasite examination.
If preserved specimens are submitted to the laboratory, it may be more cost effective to begin the stool examination with the concentration procedure rather than the direct wet mount, particularly since motile protozoa will not be viable because of the prior addition of preservative. Even if parasites are seen on a direct mount of preserved stool, they would almost certainly be seen on the concentration examination as well as on the permanent stained smear (protozoa in particular). With few exceptions, intestinal protozoa should never be identified on the basis of a wet mount alone; permanent stained smears should be examined to confirm the specific identification of suspected organisms.
(Consistent with CAP Inspection Checklist): Every stool submitted for an O&P examination must be examined using the concentration and permanent stained smear procedures.
O&P Exam (Fresh Stool Specimen/liquid and/or very soft): Direct wet smear, concentration, permanent stained smear.
O&P Exam (Fresh Stool Specimen/formed stool): Concentration, permanent stained smear.
O&P Exam (Preserved Stool Specimen): Concentration, permanent stained smear.
Remember that all intestinal protozoan infections can be missed if the concentration ONLY is performed. The permanent stained smear is much more sensitive than the concentration alone. To date, there are no commercial immunoassay products available for the confirmation of infection with D. fragilis; however, they are under development
Dientamoeba fragilis, Key Points - Laboratory Diagnosis
Life Cycle: The life cycle and mode of transmission for D. fragilis are not known, although transmission via helminth eggs such as those of Ascaris lumbricoides and Enterobius vermicularis has been postulated. The cyst stage has not been confirmed to date. The trophozoite is characterized as having one (20 to 40%) or two (60 to 80%) nuclei. The nuclear chromatin is usually fragmented into three to five granules, and there is normally no peripheral chromatin on the nuclear membrane. In some organisms, the nuclear chromatin may tend to mimic that of Endolimax nana, Entamoeba hartmanni, or even Chilomastix mesnili, particularly if the organisms are overstained. The cytoplasm is usually vacuolated and may contain ingested debris as well as some large, uniform granules. The cytoplasm can also appear uniform and clean with few inclusions. There can also be considerable size and shape variation among organisms, even on a single smear.
Epidemiology and Control: As reported for many of the intestinal protozoa, D. fragilis is worldwide in distribution. Since fecal-oral transmission has not been documented, it is difficult to speculate on preventive measures. However, if transmission does occur from the ingestion of certain helminth eggs, then the appropriate hygienic and sanitary measures to prevent contamination with fecal material would be appropriate. There is speculation that D. fragilis may be infrequently recovered and identified; low incidence or absence from survey studies may be due to poor laboratory techniques and a general lack of knowledge concerning the organism.
Most experts agree that the single most effective practice that prevents the spread of infection in the child care setting is good handwashing by the children, staff, and visitors. Rubbing hands together under running water is the most important part of washing away infectious organisms. Premoistened towelettes or wipes and waterless hand cleaners should not be used as a substitute for washing hands with soap and running water. Certainly these guidelines are not limited to infections with D. fragilis, but include all potentially infectious organisms.
Treatment: Clinical improvement has been observed in adults receiving tetracycline; symptomatic relief was reported in children receiving either diiodohydroxyquin, metronidazole, or tetracycline. Current recommendations include iodoquinol, paromomycin or tetracycline. Since symptomatic relief has been observed to follow appropriate therapy, D. fragilis is probably pathogenic in infected individuals who are symptomatic.
The image on the left is a D. fragilis trophozoite; note the clear area within the karyosome. This suggests that the karyosome is about to fragment into several different dots; the image can easily mimic an E. nana trophozoite.
The image in the center is a D. fragilis trophozoite containing two nuclei, both of which have fragmented into several chromatin dots.
The image on the right is an E. nana trophozoite in which the nucleus is very pleomorphic; this organism might be confused with D. fragilis.