Method and Organism Discussions

Discussion #2A: Diagnosis of Blastocystis spp. Infection

The standard O&P exam is recommended for recovery and identification of Blastocystis spp. in stool specimens. Microscopic examination of a direct saline wet mount may reveal small to large ‘‘central body’’ forms. An asymptomatic individual may have few organisms present, all of which are the same central-body forms. Although other forms are present in the gut (primarily in patients with diarrhea), the morphology is difficult to identify; identification relies almost exclusively on the presence of the central body form. While the concentration technique is helpful in demonstrating these organisms, the most important technique for the recovery and identification of protozoan organisms is the permanent stained smear (normally stained with trichrome or iron hematoxylin). A minimum of three specimens collected over not more than 10 days is often recommended.

This organism can be easily confused with other small protozoa and/or yeast cells. There is a tremendous size range in Blastocystis organisms, particularly the central body form. Typical morphology of the peripheral nuclei and the central body area may be difficult to see, especially in the smaller organisms. Also, there tends to be great variation in staining intensity and/or differentiation between the nuclei, limited cytoplasm and central body contents.

The classic form (Central Body Form) that is usually seen in the human stool specimen varies tremendously in size, from 6 to 40 µm, and is characterized by a large central body, which may be involved with carbohydrate and lipid storage (visually like a large vacuole). The more amebic form is occasionally seen in diarrheal fluid but may be extremely difficult to recognize. Generally, B. hominis is identified on the basis of the more typical round form with the central body.

Genetic Signatures' EasyScreen™ Gastrointestinal Parasite Detection Kit and GS1 automated workflow has been FDA 510(k) cleared for sale in the United States. This system can detect and identify 8 important GI parasites in a single test, including the following organisms:

Giardia lamblia / intestinalis (Varies with D. fragilis between #2 and #3 most common parasites found in stool specimens)
Cryptosporidium spp
Entamoeba histolytica
Cyclospora cayetanensis
Dientamoeba fragilis
Blastocystis spp. (Most common parasite found in stool specimens)
Enterocytozoon bieneusi (Microsporidia)
Encephalitozoon intestinalis (Microsporidia)

The EasyScreen™ Gastrointestinal Parasite Detection Kit and GS1 automated workflow offers a significant advancement in clinical diagnostics for expanded gastrointestinal (GI) parasite testing, providing healthcare professionals with a powerful, innovative tool for accurate and timely diagnosis.

General Comments

Blastocystis is a stramenopile (strict anaerobe protist) of worldwide significance due to its capacity to colonize several hosts. It is estimated that more than a billion individuals are infected with Blastocystis. Based on its high level of genetic diversity, Blastocystis is classified into global ribosomal subtypes (STs). Currently, 17 subtypes are known, of which ST1 to ST9 and ST12 have been identified in humans. In humans from Europe, STs 1, 2, 3 and 4 reportedly occur most commonly, whereas ST1, 2 and 3 commonly occur in South America. More than one ST can reportedly colonize humans, and infections with mixed STs have been reported. Blastocystis may cause clinical manifestations such as diarrhea, abdominal pain, irritable bowel syndrome, constipation and flatulence, along with extraintestinal manifestations such as chronic urticaria. However, these symptoms are non-specific. Different Blastocystis STs could exhibit different growth rates, drug susceptibilities, host ranges, and other biological characteristics. These differences could therefore influence the protist’s influence on the gut microbiota. It is possible that microbiota composition in relation to Blastocystis may be dependent on the organism’s subtype identity. Various studies have confirmed that the subtype differences play an important role in the pathogenesis of Blastocystis spp. Approximately half of the human subtypes may cause symptoms; relevant factors include trophozoite adhesion to epithelium and release of cysteine proteases leading to mucosal sloughing, increased intestinal permeability, and pro-inflammatory cytokine responses. There also tends to be distinct links between parasite colonization and the gut microbial flora profiles. As classification discussions continue, this organism may be reclassified with the protozoa, and the subtypes may eventually be separated into different species.

Additional Information

Terminology for Blastocystis may change, depending on the outcome of molecular studies attempting to classify this organism more accurately. Some use the term ‘‘cyst’’ to denote the central-body form, while others continue to use ‘‘central-body form.’ Either term is acceptable at present.

The organisms may resemble yeast cells, debris, and/ or amebic cysts. In some preparations, the organisms appear to be dividing, a finding not consistent with a cyst form of the intestinal protozoa.

In the absence of other organisms, Blastocystis may cause diarrhea, cramps, nausea, fever, vomiting, and abdominal pain and may require therapy. It has also been linked to infective arthritis. In patients with other underlying conditions, the symptoms may be more pronounced. The incidence of this organism in stools submitted for parasite examination appears to be higher than suspected. In symptomatic patients in whom no other etiologic agent has been identified, Blastocystis should be considered the possible pathogen. When a symptomatic Blastocystis infection responds to therapy, the improvement may represent elimination of some other undetected pathogenic organism (E. histolytica, G. lamblia, D. fragilis). However, it is also clear that some organisms within the Blastocystis group of subtypes are almost certainly pathogenic. In symptomatic patients in whom no other etiologic agent has been identified, B. hominis should be considered the possible pathogen.

Blastocystis spp. (Pathogenicity/Clinical Relevance

Statement	Comment	Reference
Genetic diversity in morphologically identical organisms; intra subtype (ST) variations; may become different species	9/17 human subtypes; half pathogenic; pathogenicity data inconsistent due to various subtyping methods in use; ST 1-ST4 >90% human carriage	Stensvold, CR et al. 2007. Trends Parasitol 23:93-96. Parija. SC, Jeremiah SS. 2013. Trop Parasitol 3:17-25 Stensvold, CR 2013. Trop Parasitol 3:26-34. Vargas-Sanchez, GB et al. 2015. PLoS One 10:e0124006 Tan, K et al. 2010. Curr Infect Dis Rep 12:28-35
Prevalence higher than expected worldwide	Accurate ID via molecular methods = clinical importance	Stensvold, CR2013. Trop Parasitol 3:26-34 Fletcher, SM et al. 2012. Clin Microbiol Rev 25:420-449
Treatment: yes or no TMP-SMX or nitazoxanide rather than metronidazole (resistance documented)	Asymptomatic: no Symptomatic with many organisms: yes Search for other causes	Coyle, CM. et al. 2012. CID 54:105-110 Roberts, T. et al. 2015. Antimicrob Agents Chemother 59:4417-4423 Tan, KSW. 2008. Clin Microbiol Rev 21:639-665 Jones, MS et al. 2009. Parasitol Res104:341-345 Sekar, U., Shanthi, M. 2013. Trop Parasitol 3:35-39.
Molecular testing See 3A, 3B, 3C in this section.	Increased sensitivity, specificity	Verweij, JJ, Stensvold, CR. 2014. Clin Microbiol Rev 27:371-418

Permanent stained slide: Central Body form

Iodine: Central Body form

References

Garcia, L.S. 2016. Diagnostic Medical Parasitology, 6^th ed. ASM Press, Washington, DC, 1388 pp.

Garcia, LS, 2021. Practical Guide to Diagnostic Medical Parasitology, 3^rd ed., ASM Press, Washington, DC