E. cuniculi

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1. Classification

a. Higher order taxa

Eukaryota; Fungi; Microsporidia; Apansporoblastina; Unikaryonidae; Encephalitozoon; Encephalitozoon cuniculi

2. Description and significance

Encephalitozoon cuniculi is a common spore-forming zoonotic microsporidian parasite that causes encephalitozoonosis infection in mammals [1]. Though it is most commonly diagnosed in pet rabbits, it is also an opportunistic pathogen of humans that causes disease in immunocompromised, and specifically, HIV positive individuals [1]. Infected animals release spores in their urine, and E. cuniculi is transmitted when other animals ingest these spores in urine-contaminated food or water [1].. Although many rabbits are asymptomatic carriers, clinical signs of E. cuniculi include head tilt, torticollis, eye lesions, kidney disease, and ocular disease [1]. A 2011 study in Taiwan found that up to 67% of healthy pet rabbits had antibodies against Encephalitozoon cuniculi, confirming that asymptomatic carriers are prevalent for this disease. [2]. Encephalitozoon cuniculi is fatal in undiagnosed and untreated individuals but symptoms are nonspecific and there is no consensus for how to diagnose the disease in living animals [3]. Encephalitozoon cuniculi poses significant public health concern in both human and veterinary medicine [1].

3. Genome structure

The full genome of Encephalitozoon cuniculi has been sequenced [4]. It contains eleven chromosomes with a total of 2.9 Mbp, which code for about 1,915 proteins [5]. It has a median GC (guanine: cytosine) content of 46.9%, and its genome is very compact. It has few introns, short intergenic regions (space between coding genes), and shorter protein genes compared to eukaryotic orthologs [6] [7]. The spliceosome (protein complex responsible for removal of introns from pre-mRNA) is made of thirty smaller protein components [8].

SWP1, EnP1, and EnP2 are proteins that are unique to Encephalitozoon cuniculi. SWP1 is a chitin deacetylase-like protein that is localized to the Encephalitozoon cuniculi endospore (9). SWP1 is involved in endospore cell wall rigidity [9]. EnP1 and EnP2 are also endospore cell wall proteins [9]. The genes for EnP1 and EnP2 are located on Chromosome I and do not have homologues [9]. These proteins are important targets for treatment development because destroying endospore cell walls could stop spread of disease both within and between animals and humans [9].

Three strains of this species have been identified, but intraspecies genetic and morphological diversity has not been substantially studied [10].

4. Cell structure

Encephalitozoon cuniculi, like many other microsporidia, lacks mitochondria and peroxisomes, and produces extremely resistant endospores [7].. It is unclear what the cell structure of non-pathogenic cells looks like, but several studies have been done to characterize the structure of the pathogenic endospores. One challenge of studying these spores is that in histological diagnoses they are easily mistaken for other microorganisms such as yeasts, protozoa, or bacteria [12]. Nonetheless, an extensive staining study done in 2017 found that the spores are ovoid (oval in shape), approximately 1.5–2.5 μm long, and have a thick wall composed of an outer proteinaceous exospore, an endospore made of chitin, and an inner plasma membrane. The spore also contains a nucleus and a coiled polar tube, which is important in its pathogenesis [7]. E. cuniculi spores also contain a polaroplast (an organelle that contributes to rupturing mature spores to release more endospores) and a posterior vacuole (where immature spores are contained) [7].

5. Metabolic processes

Encephalitozoon cuniculi is a chemoorganoheterotroph and acquires organic material from its host that it cannot synthesize itself for energy. The Encephalitozoon cuniculi genome contains four genes for a nucleotide transporter that is used to import ATP from the cytosol of the eukaryotic host cell. Three of these transporters are located on the surface of E. cuniculi while the fourth transporter is located on a remnant form of a mitochondrion called a mitosome. Encephalitozoon cuniculi is host-dependent and lacks genes coding for the tricarboxylic acid cycle (TCA cycle) or the electron transport chain [11].

6. Ecology

Encephalitozoon cuniculi is an obligate, endospore-forming parasite that cannot reproduce outside of a host [12]. Rabbits are the main reservoir for E. cuniculi, although other animals serve as hosts as well [2]. Encephalitozoon cuniculi persist in wild and domestic animals: it has been found in rodents, domestic animals like cats and dogs, farm animals like pigs and sheep, large mammalian carnivores, non-human primates, and humans [13]. The seroprevalence of E. cuniculi is higher in wild species compared to domestic animals [13] [14]..

Encephalitozoon cuniculi endospores are found in the excreted urine and stool of an infected host [15]. These spores can survive for months because of their high resistance to environmental stressors. Ingestion or inhalation of Encephalitozoon cuniculi spores leads to the intra-generational horizontal transmission of this pathogen [16].

7. Pathology

Encephalitozoon cuniculi is a zoonotic pathogen that primarily affects wild and domestic animals. Rabbits are most commonly affected by this pathogen, and symptoms include head tilt, paralysis, renal failure, incontinence, tremors, and ocular issues ([16]. Rabbits remain asymptomatic as long as the immune response and parasite reproduction are balanced [16]. Proliferative stage Encephalitozoon cuniculi and Encephalitozoon cuniculi spores are found in the intestine, mesenteric lymph nodes, spleen, liver, kidneys, lungs, heart, and nervous tissue of an infected host [16] [17]. Encephalitozoon cuniculi is extremely virulent and has important implications in human medicine, especially among immunocompromised individuals. Known vulnerable groups are HIV positive patients and kidney transplant patients [18] [19].

E. cuniculi has evolved a unique invasion mechanism based on the explosive discharge of a hollow polar tube. The infectious endospores are released through this tubule and injected into the host cell [20]. Inside the host cell, endospores undergo a full cell cycle of development and division to spread throughout the host’s body [3] [20]. Transmission between rabbits primarily occurs through ingestion (fecal-to-oral route) or inhalation (through the respiratory tract) of Encephalitozoon cuniculi endospores [16]. There is also evidence of ocular (through the eyes) and uterine transmission (through the uterus from mother to offspring) [16]. Human-to-human transmission of E. cuniculi is possible through solid organ transplantation [22]. Certain sources of contamination, such as water, food, or soil are modes of transmission of Encephalitozoon cuniculi spores to humans because microsporidian endospores are resistant to desiccation (drying out), so they are able to survive outside of a host for several weeks or months [28]. Close contact between rabbits and their owners increases risk of exposure to Encephalitozoon cuniculi. Encephalitozoon cuniculi is a zoonotic pathogen, and precautionary actions should be taken in order to prevent the spread of Encephalitozoon cuniculi in humans [23].

8. Current Research

Encephalitozoon cuniculi infection in humans is one focus of current research. One study found that the seroprevalence (prevalence of the pathogen in blood serum samples) of Encephalitozoon cuniculi in asymptomatic humans was 9.67%, indicating that this pathogen lies dormant in many humans [23]. Immunocompromised patients are at higher risk (seven times more likely) of being infected and symptomatic with Encephalitozoon cuniculi [24].

Rabbits are the main host of Encephalitozoon cuniculi in animals and veterinary research on Encephalitozoon cuniculi primarily focuses around pet rabbits [24]. Encephalitozoon cuniculi is difficult to diagnose due to its similar appearance to other microbial pathogens in animals, and because clinical symptoms are not always indicative of current infection. Much of the current research is geared towards improving diagnosis accuracy and efficiency. In microscopic analysis of tissue samples, the best way to detect Encephalitozoon cuniculi endospores for diagnosis is by modified trichrome stains and Gram stains under light microscopy, and calcofluor white stain under UV light microscopy [17].

9. Treatments

There is no standardized treatment protocol and there are no vaccines for Encephalitozoon cuniculi in humans or rabbits [24]. The typical treatment for Encephalitozoon cuniculi in rabbits is fenbendazole (a broad-spectrum dewormer). Fenbendazole is only effective as a preventative method, and not as treatment during infection [17]. In humans, the typical treatment is albendazole, an anthelmintic used for treating parasitic worm infections [26]. Albendazole treatment only works when the host has a functioning immune system and is not effective in immunocompromised individuals [27]. Most research on the prevalence of this pathogen among immunocompromised patients focuses on screening, and treatments have yet to be suggested and tested.

10. References

[1] [E. Keeble. 2011. Encephalitozoonosis in rabbits—what we do and don’t know. In Practice 33(9): 426-435] [2] [K. Y. Tee, J. P. Kao, H. Y. Chiu, et. al. 2011. Serological survey for antibodies to Encephalitozoon cuniculi in rabbits in Taiwan. Vet Parasitol 183(1-2): 68-71] [3] [M. Kicia, M. Szydlowicz, K. Cebulski, et al. 2019. Symptomatic respiratory Encephalitozoon cuniculi infection in renal transplant recipients. International Journal of Infectious Diseases 79: 21-5] [4] [Encephalitozoon cuniculi. NCBI Genome] [5] [E. Peyretaillade, O. Goncalves, S. Terrat, et al. 2009. Identification of transcriptional signals in Encephalitozoon cuniculi widespread among microsporidia phylum: support for accurate structural genome annotation. BMC Genomics 10: 607] [6] [ J. Pombert, J. Xu, D. R. Smith, et al. 2013. Complete genome sequences from three genetically distinct strains reveal high intraspecies genetic diversity in the microsporidian Encephalitozoon cuniculi. Eukaryotic Cell 12(4): 503-11] [7] [M. D. Katinka, S. Duprat, E. Cornillot, et al. 2001. Genome sequence and gene compaction of the eukaryotic parasite Encephalitozoon cuniculi. Nature 414(6862): 450-3] [8] [C. J. Grisdale, L. C. Bowers, E. S. Didier, et al. 2013. Transcriptome analysis of the parasite Encephalitozoon cuniculi: an in-depth examination of pre-mRNA splicing in a reduced eukaryote. BMC Genomics 14: 207] [9] [I. Peuvel-Fanget, V. Polonais, D. Brosson, et al. 2006. EnP1 and EnP2, two proteins associated with the Encephalitozoon cuniculi endospore, the chitin-rich inner layer of the microsporidian spore wall. International Journal of Parasitology 36(3): 309-18] [10] [A. Pelin, H. Moteshareie, B. Sak, et al. 2016. The genome of an Encephalitozoon cuniculi type III strain reveals insights into the genetic diversity and mode of reproduction of a ubiquitous vertebrate pathogen. Heredity (Edinb) 116(5): 458-65.] [11] [P. Deplazes, A. Mathis, R. Baumgartner et. al. 1996. Immunologic and molecular characteristics of Encephalitozoon-like microsporidia isolated from humans and rabbits indicate that Encephalitozoon cuniculi is a zoonotic parasite. Clin Infect Dis 22(3): 557-9 [12] [L. E. Rodríguez-Tovar, A. Villarreal-Marroquín, A. M. Nevárez-Garza, et al. 2017.] Histochemical study of Encephalitozoon cuniculi spores in the kidneys of naturally infected New Zealand rabbits. Journal of Veterinary Diagnostic Investigation 29(3): 269–277] [13] [K. Wasson and R. L. Peper. 2000. Mammalian microsporidiosis. Vet Pathol 37(2): 113-28] [14] [A. L. Meredith, S. C. Cleaveland, J. Brown, et al. 2015. Seroprevalence of Encephalitozoon cuniculi in wild rodents, foxes and domestic cats in three sites in the United Kingdom. Transboundary and Emerging Diseases 62(2): 148–156] [15] [R. Tsukada, Y. Osaka, T. Takano, et al. 2016. Serological survey of Encephalitozoon cuniculi infection in cats in Japan. Journal of Veterinary Medical Science 78(10): 1615–1617] [16] [E. Jeklova, L. Leva, J. Matiasovic, et al. 2020. Characterization of humoral and cell-mediated immunity in rabbits orally infected with Encephalitozoon cuniculi. Veterinary Research 51(1)] [17] [L. Rodríguez-Tovar, A. Nevárez-Garza, A. Trejo-Chávez, et al. 2016. Encephalitozoon cuniculi: grading the histological lesions in brain, kidney, and liver during primoinfection outbreak in rabbits. Journal of Pathogens 5768428–5768429] [18] [P. Deplazes, A. Mathis, R. Baumgartner, et al. 1996. Serological survey for antibodies to Encephalitozoon cuniculi in pet rabbits in Italy. Clinical Infectious Diseases 22(3): 557-9] [19] [W. Bohne, K. Bottcher, U. Gross. 2011. The parasitophorous vacuole of Encephalitozoon cuniculi: biogenesis and characteristics of the host cell-pathogen interface. Int J Med Microbiol 301(5): 395-9] [20] [S. P. Pakes, J. A. Shadduck, A. Cali. 1975. Fine structure of Encephalitozoon cuniculi from rabbits, mice and hamsters. J Protozool 22(4): 481-8] [21] [C. Jordan, A. M. Zajac, D. S. Lindsay. 2006. Encephalitozoon cuniculi infection in rabbits. VetFolio Parasitology 28(2)] [22] [S. Hocevar, C. Paddock, C. Spak, et al. 2014. Microsporidiosis acquired through solid organ transplantation a public health investigation. Annals of Internal Medicine 160(4): 213–220] [23] [Y. Pan, S. Wang, X. Liu, et al. 2015. Seroprevalence of Encephalitozoon cuniculi in humans and rabbits in China. Iran J Parasitol 10(2): 290–295] [24] [S. S. Abu-Akkada, E. D. H. El Kerdany, R. F. Mady, et al. 2015. Encephalitozoon cuniculi infection among immunocompromised and immunocompetent humans in Egypt. Iran J Parasitol 10(4): 561–570] [25] [L. Dipineto, L. Rinaldi, A. Santaniello, et al. 2008. Serological survey for antibodies to Encephalitozoon cuniculi in pet rabbits in Italy. Zoonoses and Public Health 55(3):173-175] [26] [S. Hong. 2018. Albendazole and praziquantel: review and safety monitoring in Korea. Infection and Chemotherapy 50(1): 1-10] [27] [M. Kotkova, B. Sak, D. Kvetonova, et. al. 2013. Latent microsporidiosis caused by Encephalitozoon cuniculi in immunocompetent hosts: a murine model demonstrating the ineffectiveness of the immune system and treatment with albendazole. PLoS One 8(4)] [28] [K. I. Ashmawy, S. S. Abuakkada, A. M. Awad. 2011. Seroprevalence of antibodies to Encephalitozoon cuniculi and Toxoplasma gondii in farmed domestic rabbits in Egypt. Zoonoses and Public Health 58: 357-364]

Edited by Hannah Steinbach, student of Jennifer Bhatnagar for BI 311 General Microbiology, 2021, Boston University.