Dientamoeba fragilis

Edited by [Huiqian Han, Lu Bai, Suzy Zhao, Wenjing Jiang], student of Jennifer Talbot for BI 311 General Microbiology, 2015, Boston University.

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Classification

Higher order taxa

Kingdom Excavata, Subkingdom Metamonada, Phylum Parabasalia, Class Tritrichomonadidae, Order Trichomonadida, Family Dientamoebidae

Genus

Dientamoeba

Species

Dientamoeba fragilis

Description and significance

Dientamoeba fragilis (D. fragilis) is a bacterium regarded as a human intestinal parasite. There is a possible link between D. fragilis colonization and abnormal gastrointestinal symptoms (1) ; however, some studies show that there is no causal relationship (2). D. fragilis has a worldwide distribution in both urban and rural areas, with infection rates ranging from 0.5% to 16%, where higher rates were reported in outbreaks and associated to the lack of personal hygiene(3). Similar to some other parasites, D. fragilis causes disease in humans regardless of their immune status, by which common symptoms include abdominal pain and diarrhea. D. fragilis has been increasingly recognized as a relatively common cause of human diarrhea and long-term chronic infections since the late twentieth century(3).

Genome structure

The complete genome of D. fragilis strains haven’t been sequenced. To date, only the coding region of the small subunit ribosomal (SSU rRNA) gene has been published(15). So far, genotypes 1 and 2 of D. fragilis are recognized through comparing the sequence of their coding regions of the ssu rRNA gene. Genotype 1 has a strong predominance in both humans and few animal hosts. RFLP (Restriction Fragment Length Polymorphism) analysis of ssu rRNA PCR products enables the distinction of these genotypes, a more applicable direct detection method for this parasite in human stool(4). By sequencing a large number of PCR products and examining across the investigated loci, a very low level of polymorphism was observed, indicating the existence of major clone of D. fragilis with a widespread geographical distribution(5). In other words, there is little genetic viability found in the SSU rRNA sequences: the two genotypes share ~97% similarity at the locus, and scarce variability is found among the isolates of the predominant genotype 1. Thus, studies have been conducted to find other portions on the ribosomal cluster that can be used for detection and typing of D. fragilis. Variation in the sequence of the internal transcribed spacers (ITS) within the AT rich region showed extensive genetic variability, due to the variation in number of A and T bases. However, since such variability was also found within single isolate, it can’t be used as a subtyping tool. Analysis done on the C-profile, which are the peaks for C nucleotides from a sequencing chromatogram, allows comparing isolates and identifying intra-genomic variability. Besides, 5.8S ribosomal RNA gene sequence is used for detection but not genotyping of D. fragilis due to its short length and limited genetic variability(4). Variations in D. fragilis’ rRNA gene sequence are used only as markers to analyze speciation and search for phylogenetic association in closely related species. They don’t have any influence on the pathogenic potential of the parasites. Further studies of variation in potentially pathogenicity involving genes of D. fragilis could be valuable(15). For instance, D. fragilis encode key virulence proteins such as cysteine proteases, lectins, and calmodulin, and the difference in expression level could play a potential role in determining their pathogenicity(16).

Cell structure

Interesting features of cell structure. Can be combined with “metabolic processes” The cell shape of D. fragilis varies from spherical to ovoidal, sometimes amoeboid with the range of size between 4 to 10 m. Flagella or undulating membrane were not found. There were two types of cell surface structure found under the scanning electron microscopy (SEM): smooth and ruffled surfaces. The ruffled cells are more common.

Nucleus: The nucleus, which is normally detected in the central area, ranges in size from 0.86 to 3.52 um in mononucleated trophozoites (a growing stage of some parasites when they absorb nutrients from the host) and from 1.12 to 2.06 um in binucleated cells. Both mono- and bi-nucleated trophozoites have fragmented nuclei. But only mononucleated cells divide their nucleus. When dividing nuclei appears, the nuclei size could be larger than binucleated cells. A double membrane with plenty of nuclear pores consists the nuclear envelope. Two to eight chromatin bodies, without peripheral chromatin, could be seen in both of the trophozoites. The nucleolus, a rounded structure on the periphery of the nucleus, consisted of a dense fibrillar component but there is no membrane appeared(6).

The predominant stage observed in vitro culture is mononucleated cells. D. fragilis seems reproduce by binary fission, which helps to constrict the body. The shape of non-dividing cells could be spherical, oval and even irregular shaped. There is a extracellular spindle between the nuclei in binucleated cells, which emanating from one polar complex adjacent to one nuclei. The spindle microtubules originate in pairs and non-periodic structures. The types of microtubules observed in the cell are: pole-to-pole, pole-to-nucleus, and pole-to-cytosol(6).

Golgi: the well-developed golgi complex was observed as a vesicular structure in perinuclear region, near endoplasmic reticulum(ER) and near microtubule bundles. The complex is a single and prominent structure with 7-10 cisternae. The length of golgi in D. fragilis is around 450nm. In binucleated cells, golgi seems to be mostly fragmented and not in an organism form(6).

Endoplasmic Reticulum (ER): Smooth and rough ER are well developed in both mononucleated and binucleated trophozoites, appearing clearly around nucleus. Rough ER was more commonly showed in the mononucleated cells. They were closely connected with the hydrogenosomes, food vacuoles and microtubules(6).

Hydrogenosomes: They are electron-bound organelles with double layered membrane, which appear when peroxisomes and mitochondria were absent. The shape of the organelle could be spherical, oval-shaped or slightly elongated. The hydrogenosomes located in cytoplasm with the smooth membrane and homogeneously granular matrix. They were observed associated with cytoplasmic inclusions and with digestive vacuoles(6).

Digestive vacuole (DV)/food vacuole: DV was normally found in cytoplasm, and it might contain bacteria, rice starch and myelin configurations. The size varied between 0.59 to 4.2um. Some DVs with bacteria and rice starch were recognisable at the early stage of digestion. D. fragilis feeds by phagocytosis, and the waste was released in DVs by exocytosis(6).

Lysosomes: Lysosomes were presented in size ranging between 0.50-2.0um, and concentrated in posterior region and close to the cell membrane. It could digest internalised bacteria and rice starch(6).

Cytoplasm and other cytoplasmic inclusions: Cytoplasm is surrounded by a double layered cell membrane. A large amount of glycogen granules and electron dense materials are usually distributed throughout the cytoplasm. It is hard to identify rough ER in perinuclear region.

Phagocytosis: D. fragilis changes their shapes to engulf bacteria and rice starch. The formation of phagocytosis allows food digestion(6). [JMB8] Virus-like particle (VLP): They are often seen in cytoplasm of trophozoites, with sizes ranging between 40 and 200 nm. VLPs are normally in spherical, with a dense core, a middle electron-lucent layer and an outer coat(6).

Metabolic processes

Enzymes associated with glycolysis/gluconeogenesis, pyruvate metabolism, the tricarboxylic acid (TCA) cycle, the pentose phosphate pathway and starch and sucrose metabolism were detectioned(7). D. fragilis lacks the ability to carry out de novo purine and pyrimidine nucleotide biosynthesis; instead uses a pathway that is able to break down the purines and pyrimidines that does not require the enzyme to convert 5-phospho--D-ribose 1-diphosphate into inosine monophosphate (IMP) and uridine monophosphate (UMP) (7). Another process used is the arginine dihydrolase pathway, which can use arginine as a direct form of energy metabolism, it is a direct and efficient way of generating ATP (7).

Ecology

D. fragilis is often regarded as a human parasite which causes abnormal gastrointestinal symptoms such as abdominal pain and diarrhea(8). It is commonly found in human bile and feces, which suggests that it thrives in small and large intestines, as well as the gallbladder(9). D. fragilis is anaerobic bacteria; therefore, it is very sensitive to aerobic environment(8). It may not survive and reproduce after it leaves the host body, so it nearly does not exist in the outside environment. Besides, similar to many other kinds of microbes, D. fragilis does not thrive alone: it is likely to build a dependent relationship with Entamoeba histolytica(8).

Pathology

Ever since Dientamoeba fragilis’ first discovery in 1918, the parasite has struggled to be recognized as a pathogen. There exists a growing body of case reports and studies from countries all over the world, linking this protozoan parasite to clinical symptoms such as diarrhea, abdominal pain, flatulence, and anorexia(10). On the other hand, there are other research suggesting that the parasite is also commonly found in healthy subjects with no presence of any gastrointestinal symptoms(4). Those studies argue that the colonization of D. fragilis does not increase risks for pathogenic gastrointestinal symptoms(11).

Current Research

The pathogenicity of D. fragilis is controversial. Some research provides evidence that D. fragilis is a human pathogenic bacterium, which causes gastrointestinal symptoms(8), while others argue that D. fragilis is just a common asymptomatic parasite(11).

           (a) D. fragilis is a pathogenic parasite

D. fragilis was usually regarded as a harmless parasite that does not cause abnormal gastrointestinal symptoms. However, its pathogenicity was explored through recent researches. The colonization of D. fragilis among patients with gastrointestinal symptoms was evaluated through stool analysis, and it was compared to a better-known intestinal parasite Giardia lamblia (G. lambila). The result showed that the prevalence and pathogenicity of D. fragilis is similar to G. lambila: D. fragilis is a potential cause of gastrointestinal symptoms(1). Moreover, D. fragilis is considered as a pathogen based on a report which focuses on a family cluster of D. fragilis-related illness and asymptomatic carriage(12). The report conducted five case studies within one family regarding complete blood count (CBC) and fecal specimen analysis. The results found linkage between detection of D.fragilis and marked peripheral eosinophilia, often associated as response to allergens and indicates activation of the immune system. Besides, D. fragilis was the only consistent finding in all family members involved in this cluster, all shown gastrointestinal symptoms(3).

           (b) D. fragilis is a harmless parasite

Though many researches provide evidence about the pathogenicity of D. fragilis, some researchers insist that D. fragilis is a harmless intestinal parasite. One study explores the causal relationship between the presence of D. fragilis and gastrointestinal symptoms among children in primary care setting(11). A cross-sectional study among children with D. fragilis infection using logistic regression analysis as well as a control analysis using asymptomatic siblings of the subjects was conducted. The conclusion is that D. fragilis is not a pathogenic parasite which causes abdominal pain and diarrhea; instead, it is a common intestinal parasite which does not cause gastrointestinal symptoms(11). Furthermore, a study compares several reports based on the prevalence of D. fragilis in individuals with and without diarrhea. Based on the statistic results, it shows that only a proportion of subjects with D. fragilis infection is present with diarrhea symptoms. Vice versa, out of the entire study population with diarrhea symptoms, only a small percent of them was detected with D. fragilis infection. These research states that the pathogenicity of D. fragile isn’t conclusive due to lack of a systematic study on the organism’s treatment and resolution(13).

           (c) Possible Treatments for D. fragilis infection

Diiodohydroxyquin, tetracycline, metronidazole and paromomycin are common treatments for eradicating D. fragilis colonization, and treatment period is between 5 to 21 days. In particular, study on paromomycin show promising efficiency in treatment. It was evaluated by examining feces for D. fragilis and other intestinal parasites after 28 days of treatment, and 80% and 87% infected study subjects shown parasitologically and clinically recovered. D. fragilis infection symptoms such as abdominal pain and diarrhea were reported to be relieved after taking paromomycin(14). Furthermore, Secnidazole is suggested as a relatively novel, more effective treatment compared to the other drugs. It has a short half-life, therefore only a single-dose is required in most of the cases to eradicate D. fragilis infection(1). This drug is suggested to take after dinner to avoid possible nausea and vomiting, and it does not severe side-effects except for occasional mild nausea(1).

References

[1][Girginkardeşler, N., Coşkun, Ş, Balcioğlu, I. C., Ertan, P., & Ok, Ü Z. (2003). Dientamoeba fragilis, a neglected cause of diarrhea, successfully treated with secnidazole. Clinical Microbiology and Infection,9(2), 110-113.]

[2][Holtman, G. A., Kranenberg, J. J., Blanker, M. H., Ott, A., Leeuwen, Y. L., & Berger, M.Y. (2016). Dientamoeba fragilis colonization is not associated with Gastrointestinal symptoms in children at primary care level. Family Practice, 34(1), 25-29. doi:10.1093/fampra/cmw111]

[3][El-Gayar, E. K., Mokhtar, A. B., & Hassan, W. A. (2016). Study of the pathogenic potential of Dientamoeba fragilis in experimentally infected mice. Parasite Epidemiology and Control,1(2), 136-143.]

[4][Cacciò, S. M. (2018). Molecular epidemiology of Dientamoeba fragilis. Acta Tropica,184, 73-77. doi:10.1016/j.actatropica.2017.06.029]

[5][Cacciò, S. M., Sannella, A. R., Bruno, A., Stensvold, C. R., David, E. B., Guimarães, S., ...Pozio, E. (2016). Multilocus sequence typing of Dientamoeba fragilis identified a major clone with widespread geographical distribution. International Journal for Parasitology,46(12), 793-798. doi:10.1016/j.ijpara.2016.07.002]

[6][Banik, G.R., Birch, D., Stark, D., Ellos J. T. (2011). A microscopic description and ultrastructural characterisation of Dientamoeba fragilis: An emerging cause of human enteric disease.International Journal for Parasitology, 42(2), 139-153.]

[7][Baratt, J.L.N., Cao M., Stark D.J., & Ellis J.T. (2015). The Transcriptome Sequence of Dientamoeba fragilis Offers New Biological Insights on its Metabolism, Kinome, Degradome and Potential Mechanisms of Pathogenicity. Protist, 166 (4), 389-408.]

[8][Stark, D., Roberts, T., Marriott, D., Harkness, J., & Ellis, J. T. (2012). Detection and Transmission of Dientamoeba fragilis from Environmental and Household Samples. The American Journal of Tropical Medicine and Hygiene,86(2), 233-236. doi:10.4269/ajtmh.2012.11-0526]

[9][Talis, B., Stein, B., & Lengy, J. (1971). Dientamoeba fragilis in Human Feces and Bile. Israel Journal of Medical Sciences, 7(9), 1063-1069.]

[10][Johnson, E. H., Windsor, J. J., & Clark, C. G. (2004). Emerging from obscurity biological, clinical, and diagnostic aspects of Dientamoeba fragilis. Clinical microbiology reviews, 17(3), 553-570.]

[11][Holtman, G. A., Kranenberg, J. J., Blanker, M. H., Ott, A., Leeuwen, Y. L., & Berger, M. Y. (2016). Dientamoeba fragilis colonization is not associated with gastrointestinal symptoms in children at primary care level. Family Practice, 34(1), 25-29. doi:10.1093/fampra/cmw111]

[12][Gray, T. J., et al. (2013). Dientamoeba fragilis: a family cluster of disease associated with marked peripheral eosinophilia.Clin Infect Dis,57(6), 845-848.]

[13][Wong, ZW., Faulder, K. & Robinson, J.L. (2018) Does Dientamoeba fragilis cause diarrhea? A systematic review. Parasitology research, 117, 4, 971-980.]

[14][Vandenberg, O, et al. (2007) Treatment of Dientamoeba Fragilis infection with Paromomycin.The Pediatric Infectious Disease,26:88-90 Peek, R., Reedeker, F. R., & van Gool, T. (2004). Direct amplification and genotyping of Dientamoeba fragilis from human stool specimens. Journal of clinical microbiology, 42(2), 631-5.]

[15][Peek, R., Reedeker, F. R., & van Gool, T. (2004). Direct amplification and genotyping of Dientamoeba fragilis from human stool specimens. Journal of clinical microbiology, 42(2), 631-5.]

[16][Stark, D., Barratt, J., Chan, D., & Ellis, J. T. (2016). Dientamoeba fragilis, the Neglected Trichomonad of the Human Bowel. Clinical microbiology reviews, 29(3), 553-80.]

Edited by [Jennifer Talbot], student of Jennifer Talbot for BI 311 General Microbiology, 2016, Boston University.