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Parasitology – First Quadrimester, 2021 AAB PARASITOLOGY – FIRST QUADRIMESTER, 2021 American Association of Bioanalysts Proficiency Testing 11931 Wickchester Ln., Ste. 200 Houston, TX 77043 800-234-5315 ♦ 281-436-5357 Q1 2021 Parasitology Sample 1 Referees Extent 1 Extent 2 Total Code - Organism Frequency % No. % No. % No. % No. 525 – No parasites found None 50% 5 11.1% 1 44.4% 8 33.3% 9 544 – Endolimax nana 44.4% 4 11.1% 2 22.2% 6 533 – Dientamoeba fragilis Few 40.0% 4 0.0% 0 27.8% 5 18.5% 5 546 – Entamoeba hartmanii Few 10.0% 1 11.1% 1 11.1% 2 11.1% 3 524 – parasite(s) found referred for ID 33.3% 3 0.0% 0 11.1% 3 553 – Cryptosporidium sp. 0.0% 0 5.6% 1 3.7% 1 Due to a lack of consensus, Sample 1 was not evaluated this event. The intended organism was 533-Dientamoeba fragilis. SPECIMEN 1: FORMALIN: Specimen 1A was a fecal suspension in 10% formalin for direct wet mount examination; concentration was not necessary. The specimen was to be examined for all parasites unstained, with iodine or other acceptable wet mount stain. The specimen contains Dientamoeba fragilis trophozoites. Typically, it is very difficult if not impossible to identify these organisms from a wet mount examination. SPECIMEN 1: PERMANENT SMEAR FOR STAINING: Specimen 1B was the smear to be stained and examined. This specimen contains Dientamoeba fragilis. Due to a lack of participant consensus, Specimen 1 was not evaluated for this event. However, 40% of the Referees correctly reported Dientamoeba fragilis. Many Referees (50%) and Participants (33.3%) reported the specimen as negative. Entamoeba coli trophozoites Entamoeba coli cysts (shrunk and normal) Sample 2 Referees Extent 1 Extent 2 Total Code - Organism Frequency % No. % No. % No. % No. 534 – Gardia lamblia Few to Many 100.0% 10 80.0% 8 89.5% 17 86.2% 25 541 – Blastocystis hominis Few 10.0% 1 10.0% 1 5.3% 1 6.9% 2 545 – Entamoeba coli 0.0% 0 5.3% 1 3.4% 1 524 – parasite(s) found referred for ID 10.0% 1 0.0% 0 3.4% 1 Extent 1 flagging appears for failure to report 534 or 524. Extent 2 flagging appears for failure to report 534. Flagging appears in both extents for reporting other than 534, 541 or 524. 1 AAB 3rd Quadrimester Parasitology, 2020 AAB PARASITOLOGY – FIRST QUADRIMESTER, 2021 SPECIMEN 2: FORMALIN: The specimen was a fecal suspension in 10% formalin for direct wet mount examination; concentration was not necessary. The specimen was to be examined for all parasites unstained, with iodine or other acceptable wet mount stain. This specimen contained Giardia lamblia. The referees reported 100% Giardia lamblia. Flagging occurs in Extent 1 for failure to report Giardia lamblia or Parasite(s) found, referred for ID. Extent 2 flagging occurs for failure to report Giardia lamblia. Flagging also appears in both extents for reporting other than Giardia lamblia, Blastocystis spp. or Parasite(s) found, referred for ID. Participants reported 86.2% Giardia lamblia. The G. lamblia cyst morphology was very typical with a round to oval shape and the presence of multiple nuclei, curved median bodies, and linear axonemes. Representative images can be seen below. The internal structures often appear very refractile in the wet preparation examination. Although some of the cysts are shrunk within the cyst wall, there are plenty of organisms that can be easily identified. Very high magnification G. lamblia cyst G. lamblia trophozoite G. lamblia cyst G. lamblia trophozoite Wet Mounts Permanent Stained Smear Not used in this challenge. The Blastocystis spp. central body forms were very typical with a round to somewhat ovoid shape, the presence of a central area with multiple small nuclei arranged around the central body area within the cell membrane. See comparative images below. The Referees (10%) and the Participants (6.9%) also correctly reported Blastocystis. Sample 3 Referees Extent 1 Extent 2 Total Code - Organism Frequency % No. % No. % No. % No. 525 – No parasites found None 100.0% 10 88.9% 8 100.0% 17 96.2% 25 524 – parasite(s) found referred for ID 11.1% 1 0.0% 0 3.8% 1 Extent 1 flagging appears for failure to report 525. Extent 2 flagging appears for failure to report 525. There are no other acceptable codes. 2 AAB 3rd Quadrimester Parasitology, 2020 AAB PARASITOLOGY – FIRST QUADRIMESTER, 2021 SPECIMEN 3 FORMALIN: The specimen was a fecal suspension in 10% formalin for direct wet mount examination; concentration was not necessary. The specimen was to be examined for all parasites unstained, with iodine or other acceptable wet mount stain. There are no parasites in this specimen. Artifact material and/or yeast cells can be somewhat confusing when reviewing the wet preparation using the low power and even high dry power objectives. However, there is nothing present that can be specifically identified at 100X and 400X magnifications as a parasite, either helminth or protozoan. When having trouble seeing possible internal structures and/or morphologic details, tap the coverslip and get things to move around a bit. Also, reduce the light intensity if you’re not using iodine and drop the condenser to increase contrast. Iodine can be used to provide a bit more contrast; some laboratories routinely use iodine, while others do not. Too much light for wet preparations may prevent you from seeing parasites, particularly protozoa, which might be present in the specimen. Although occasionally a formalin preparation may contain very rare organisms, positive specimens selected for proficiency testing tend to have moderate to many organisms that are present for identification. Flagging appears in both extents for reporting other than “No Parasites Seen” – participants performed very well in the examination of this specimen with an overall 96.2% correct response. Sample 4 Referees Extent 1 Extent 2 Total Code - Organism Frequency % No. % No. % No. % No. 549 – Iodamoeba butschlii 100.0% 10 66.7% 4 100.0% 15 90.5% 19 524 – parasite(s) found referred for ID 33.3% 2 0.0% 0 9.5% 2 Extent 1 flagging appears for failure to report 549 or 524. Extent 2 flagging appears for failure to report 549. Flagging also appears in both extents for reporting other than 549 or 524. SPECIMEN 4 (Digital Image): This specimen (digital image of routine trichrome stain) contains Iodamoeba bütschlii trophozoites and cysts. The referees (10/10) reported correctly, identifying Iodamoeba bütschlii. Extent 1 flagging appears for failure to report Iodamoeba bütschlii or parasite(s) found referred for ID. Extent 2 flagging appears for failure to report Iodamoeba bütschlii. Flagging also appears in both extents for reporting other than Iodamoeba bütschlii or parasite(s) found referred for ID. The participants performed well with an overall percent of 90.5%. Because the trophozoite can resemble that of Endolimax nana, most identifications of Iodamoeba bütschlii will be based on the typical cyst morphology. When examining the permanent stained smears, it is important to read at least 300 fields using the oil immersion objective (100X objective) for a total magnification of X1000). This examination is in contrast to the concentration sediment wet preparation, for which at least 1/3 to ½ of the coverslip should be examined using the high dry objective (40X) and the entire 22x22 mm coverslip should be examined using the low power objective (10X). NOTE: MANY OF THE CYSTS CONTAIN A “BASKET NUCLEUS” – THIS MAY NOT BE TYPICAL FOR CLINICAL SPECIMENS (VARIES TREMENDOUSLY AMONG CYSTS). Example 1 contains a single cyst, which has the typical large karyosome. Note the fine granules at the bottom of the karyosome; this represents the “basket” nucleus, which can also occur in the trophozoite although is more commonly seen in the cyst form. Because the trophozoite can often resemble that of Endolimax nana, identification of Iodamoeba bütschlii is normally based on the cyst form. In this cyst, the vacuole (left) is somewhat obscured by debris. Example 2 contains one typical cyst; note the large red/purple karyosome and the pale glycogen vacuole (bottom of image). Example 3 contains one typical cyst with the clear glycogen vacuole lying to the left of the karyosome and large red/purple karyosome (granules to the left of the karyosome represent the “basket” nucleus). Example 4 contains one cyst; note that the pale glycogen vacuole is distorted and appears collapsed and the large karyosome with many granules at the top (“basket nucleus”). 3 AAB 3rd Quadrimester Parasitology, 2020 AAB PARASITOLOGY – FIRST QUADRIMESTER, 2021 Example 5 contains a single cyst, which has a very large karyosome and typical glycogen vacuole. The nucleus is a typical “basket” nucleus with extra pale granules at the top of the karyosome. Example 6 contains one typical cyst with a glycogen vacuole; note the basket nucleus (extra chromatin is at the bottom of the karyosome). Example 7 contains a single cyst (glycogen vacuole is somewhat out of focus – left of karyosome). The karyosome is very typical; note the large number of granules right below/left of the karyosome (“basket nucleus”). Example 8 contains a typical trophozoite (random shape). The large karyosome is typical. Example 9 contains one cyst with a glycogen vacuole and typical large karyosome; no vacuole. Example 10 contains a single cyst with an excellent karyosome and large glycogen vacuole (somewhat distorted). Example 11 contains a single trophozoite containing much debris in the cytoplasm; the karyosome is large and typical. Note the trophozoite outline is somewhat difficult to see. Example 12 contains a typical cyst with a large karyosome and glycogen vacuole. Example 13 contains one cyst with a typical glycogen vacuole and typical large karyosome. Note the granules above the karyosome (“basket nucleus”).
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