Detection of Pathogenic Protozoa in the Diagnostic Laboratory: Result Reproducibility, Specimen Pooling, and Competency Assessment

Abstract

Stool microscopy as performed in clinical parasitology laboratories is a complex procedure with subjective interpretation. Quality assurance (QA) programs often emphasize proficiency testing as an assessment tool. We describe a result reproducibility assessment tool, which can form part of a broader QA program, and which is based on the blinded resubmission of selected clinical samples, using concordance between the reports of the initial and resubmitted specimen as an indicator. Specimens preserved in sodium acetate-acetic acid-formalin can be stored for several months for use in such a program. The presence of multiple protozoa in one specimen does not affect concordance. Some dilution of specimens occurs in this process, and this may explain poor concordance when specimens with low protozoal concentrations are resubmitted. Evaluation of this tool in a large parasitology laboratory revealed concordance rates for pathogenic protozoa (Entamoeba histolytica/Entamoeba dispar, Giardia lamblia, and Dientamoeba fragilis) of about 80%, which may be considered for use as a benchmark value. We also used this tool to demonstrate that when pairs of specimens from one patient are pooled to create a single specimen, concordance between the results of the individual and pooled specimens is high.Clinical parasitology laboratories, in contrast to most other diagnostic laboratories, utilize many complex manual technical procedures which are subject to individual variation and subjective interpretations (8). As a result, quality assurance (QA) programs are crucial for maintaining confidence in the accuracy of test results. Meaningful QA programs can be difficult to design and expensive to implement. In addition, indicators of diagnostic accuracy can be difficult to assess given the inherent variability among testing procedures (19) and technologists (11). Therefore, not only does the routine testing of stool specimens for intestinal protozoa by microscopy remain a challenge for the modern diagnostic laboratory (3, 19), but so does the assessment of the accuracy of this testing.To date, no studies have been published on the value of a QA program undertaken to directly evaluate the quality of microscopy in the detection of intestinal protozoa, although several have reviewed this for other types of procedures and microorganisms (e.g., bacterial counts [2]; human immunodeficiency virus [4]). QA programs, such as that undertaken by the College of American Pathologists, have typically relied on results from proficiency tests (17). In some areas of clinical microbiology, proficiency testing has been used to evaluate multiple steps in a complex process and reveal widespread systematic problems (e.g., for Streptococcus pneumoniae antimicrobial susceptibility testing [5]). In the parasitology laboratory, proficiency tests are based on sending "unknowns" to each participating laboratory, and the evaluation is based on the number of parasite species correctly identified. A potential weakness of proficiency testing is the possibility for the...

ACG Clinical Guideline: Diagnosis, Treatment, and Prevention of Acute Diarrheal Infections in Adults

“…Th e limitations of this method are that it is labor and time intensive, requires technical expertise, and lacks sensitivity and reproducibility. Multiple specimens are oft en required to reduce the day-to-day variability in parasite shedding ( 34 ).…”

Dientamoeba fragilis diagnosis by fecal screening: relative effectiveness of traditional techniques and molecular methods

“…The suspension was filtered through two layers of gauze and centrifuged at 800 × g for 5 min. Sediments were re-suspended in ~2 mL of PBS before combining with culture medium and fixing in either sodium acetate-acetic acid-formalin (SAF) or 80% ethanol [27][28][29].…”

Comparison of Individual Differences in the Direct Microscopic Examination in the Diagnosis of Intestinal Parasites

“…Gaitanın mikroskobik incelenmesi ile elde edilen sonuçların kullanılan yönteme, örneğin kıvamına, kan ve mukus içerip içermedi-ğine, laboratuvar çalışanlarının niteliklerine göre farklılıklar gös-terdiği bildirilmektedir (2,4,6). Gerçekten bağırsak parazitlerinin doğru tanı ve tedavisinde, gaita örneklerinin doğru şekilde toplanması, uygun şekilde ve sürede taşınması, saklanması, örnekle-rin yeterli miktarda olması gibi laboratuvar öncesi uygulamaların yanı sıra, laboratuvara ulaşan örneklerin doğru yöntemlerle işle-me alınması, deneyimli uzmanlar tarafından değerlendirilmesi ve doğru şekilde raporlanması da son derece önemlidir (8,9).…”

“…Ayrıca test prosedürleri ve uygulayıcılar arasındaki farklılıklar nedeniyle bu tür sistemlerin değerlendirilmesinde de zorluklar yaşanmaktadır (4,5). Modern tanı laboratuvarlarının tek sorunu bağırsak parazitlerinin rutin tanı-sında kullanılan yöntemin mikroskobik incelenmesi değil, aynı zamanda bu testin doğruluğunun da yeterince değerlendirileme-mesidir (6). Geleneksel tanı yöntemlerinde deneyimli personel ve zaman önemli bir faktördür.…”