Chapter 135 : General Approaches for Detection and Identification of Parasites

Processing liquid stool for O&P examination. Either PVA or Schaudinn’s fixative can be used for the preparation of the permanent stained smear. Organism motility is seen when saline is used; iodine kills the organisms, so motility will no longer be visible. The use of ethyl acetate may remove the entire specimen and pull it into the layer of debris that will be discarded (liquid specimen normally contains mucus); centrifuge at 500 × g for 10 min (normal centrifugation time), but do not use ethyl acetate in the procedure. In general, laboratories have switched to nonmercury substitutes; the original Schaudinn’s fixative contains mercuric chloride. However, in some instances the term “Schaudinn’s fixative” is still used to describe not the original fixative but a formulation that is prepared with a copper or zinc base or other proprietary compounds. When fixatives are selected, it is important to know the contents in order to comply with disposal regulations. Reprinted from reference 9 . doi:10.1128/9781555817381.ch135.f1

Processing preserved stool for O&P examination (two-vial collection kit). The formalin can be buffered or nonbuffered, depending on the laboratory protocol in use. Fixative prepared with mercuric chloride provides the best organism preservation. Alternatives are available, including zinc-based PVA, copper sulfate-based PVA, SAF, and the one of the single vial fixatives that requires no adhesive (PVA or albumin). Reprinted from reference 9 . doi:10.1128/9781555817381.ch135.f2

Processing preserved stool for O&P examination (one-vial collection kit). There are single-vial collection systems for which the formulas are proprietary; however, many contain zinc sulfate as one of the key ingredients. With the exception of SAF, compatibility of these fixatives with immunoassay reagents is not always possible, particularly if the fixative contains PVA. Other good options include Unifix, Total-Fix (single-vial fixative with no mercury, formalin, or PVA) (Medical Chemical Corp.), or Z-PVA (Medical Chemical Corp.) with trichrome stain, as well as Ecofix with Ecostain (Meridian Biosciences, Cincinnati, OH). If the iron hematoxylin method containing the carbol fuchsin step is used, the coccidian oocysts will stain pink (Cryptosporidium spp., Cyclospora cayetanensis, or Cystoisospora belli). Also, when using SAF, some recommend the use of an adhesive such as egg albumin to “glue” the fecal material onto the slide prior to staining. It is highly recommended that special stains be performed for the detection and identification of the coccidia (modified acid-fast stains) and the microsporidia (modified trichrome stains) from concentrated sediment to enhance organism recovery. Coccidian oocysts of Cystoisospora belli can easily be detected in the concentration sediment wet mount; however, unless a very heavy infection is present, Cryptosporidium spp. oocysts may not be seen without special modified acid-fast stains. The small size of the microsporidian spores prevents identification without the use of special modified trichrome stains and microscopic examination with a 100× oil immersion objective. Reprinted from reference 9 . doi:10.1128/9781555817381.ch135.f3