Encephalitozoonosis is a common infectious disease widely spread among rabbits. Encephalitozoon cuniculi, is considered as a zoonotic and emerging pathogen capable of infecting both immunocompetent and immunocompromised hosts. The aim of the study was to describe in detail the spread of the E. cuniculi in a rabbit organism after experimental infection and the host humoral and cellular immune response including cytokine production. For that purpose, healthy immunocompetent rabbits were infected orally in order to simulate the natural route of infection and euthanised at 2, 4, 6 and 8-weeks post-infection. Dissemination of E. cuniculi in the body of the rabbit was more rapid than previously reported. As early as 2 weeks post-infection, E. cuniculi was detected using immunohistochemistry not only in the intestine, mesenteric lymph nodes, spleen, liver, kidneys, lungs and heart, but also in nervous tissues, especially in medulla oblongata, cerebellum, and leptomeninges. Based on flow cytometry, no conspicuous changes in lymphocyte subpopulations were detected in the examined lymphoid organs of infected rabbits. Cell-mediated immunity was characterized by ability of both CD4+ and CD8+ T cells to proliferate after stimulation with specific antigens. Th1 polarization of immune response with a predominance of IFN-γ expression was detected in spleen, mesenteric lymph nodes and Peyer's patches. The increased expression of IL-4 and IL-10 mRNA in mixed samples from the small intestine is indicative of balanced control of IFN-γ, which prevents tissue damage. On the other hand, it can enable E. cuniculi to survive and persist in the host organism in a balanced host-parasite relationship. The Th17 immunity lineage seems to play only a minor role in E. cuniculi infection in rabbits.

Veterinary Research Institute Hudcova 296 70 621 00 Brno Czech Republic

See more in PubMed

Didier ES, Didier PJ, Snowden KF, Shadduck JA. Microsporidiosis in mammals. Microb Infect. 2000;2:709–720. PubMed

Katinka MD, Duprat S, Cornillot E, Méténier G, Thomarat F, Prensier G, Barbe V, Peyretaillade E, Brottier P, Wincker P, Delbac F, El Alaoui H, Peyret P, Saurin W, Gouy M, Weissenbach J, Vivarès CP. Genome sequence and gene compaction of the eukaryote parasite Encephalitozoon cuniculi. Nature. 2001;414:450–4533. PubMed

Deplazes P, Mathis A, Baumgartner R, Tanner I, Weber R. Immunologic and molecular characteristics of Encephalitozoon—like microsporidia isolated form humans and rabbits indicate that Encephalitozoon cuniculi is a zoonotic parasite. Clin Infect Dis. 1996;22:557–559. PubMed

Mathis A, Weber R, Deplazes P. Zoonotic potential of the microsporidia. Clin Microbiol Rev. 2005;18:423–445. PubMed PMC

Hunt RD, King NW, Foster HL. Encephalitozoonosis: evidence for vertical transmission. J Infect Dis. 1972;126:212–214. PubMed

Baneux PJ, Pognan F. In utero transmission of Encephalitozoon cuniculi strain type I in rabbits. Lab Anim. 2003;37:132–138. PubMed

Jeklova E, Leva L, Kovarcik K, Matiasovic J, Kummer V, Kummer V, Maskova J, Skoric M, Faldyna M. Experimental oral and ocular Encephalitozoon cuniculi infection in rabbits. Parasitology. 2010;13:1749–1757. PubMed

Ozkan O, Karagoz A, Kocak N. First molecular evidence of ocular transmission of Encephalitozoonosis during the intrauterine period in rabbits. Parasitol Int. 2019;71:1–4. PubMed

Jeklova E, Leva L, Kummer V, Jekl V, Faldyna M. Immunohistochemical detection of Encephalitozoon cuniculi in ocular structures of immunocompetent rabbits. Animals (Basel) 2019;18:988. PubMed PMC

Didier ES, Stovall ME, Green LC, Brindley PJ, Sestak K, Didier PJ. Epidemiology of microsporidiosis: sources and modes of transmission. Vet Parasitol. 2004;126:145–166. PubMed

Cox JC, Hamilton RC, Attwood HD. An investigation of the route and progression of Encephalitozoon cuniculi infection in adult rabbits. J Protozool. 1979;26:260–265. PubMed

Schmidt EC, Shadduck JA. Murine encephalitozoonosis model for studying the host-parasite relationship of the chronic infection. Infect Immun. 1983;40:936–942. PubMed PMC

Shadduck JA, Orenstein JM. Comparative pathology of microsporidiosis. Arch Pathol Lab Med. 1993;117:1215–1219. PubMed

Harcourt-Brown FM, Holloway HKR. Encephalitozoon cuniculi in pet rabbits. Vet Rec. 2003;152:427–431. PubMed

Künzel F, Joachim A. Encephalitozoonosis in rabbits. Parasitol Res. 2010;106:299–309. PubMed

Braunfuchsová P, Salát J, Kopecký J. Comparison of the significance of CD4+ and CD8+ T lymphocytes in the protection of mice against Encephalitozoon cuniculi infection. J Parasitol. 2002;88:797–799. PubMed

Salát J, Sak B, Le T, Kopecký J. Susceptibility of IFN-gamma or IL-12 knock-out and SCID mice to infection with two microsporidian species, Encephalitozoon cuniculi and E. intestinalis. Folia Parasitol (Praha) 2004;51:275–282. PubMed

Khan IA, Schwartzman JD, Kasper LH, Moretto M. CD8 + CTLs are essential for protective immunity against Encephalitozoon cuniculi infection. J Immunol. 1999;162:6086–6089. PubMed

Rodríguez-Tovar LE, Castillo-Velázquez U, Arce-Mendoza AY, Nevárez-Garza AM, Zarate-Ramos JJ, Hernández-Vidal G, Rodríguez-Ramírez HG, Trejo-Chávez A. Interferon γ and interleukin 10 responses in immunocompetent and immunosuppressed New Zealand White rabbits naturally infected with Encephalitozoon cuniculi. Dev Comp Immunol. 2019;62:82–88. PubMed

Schmidt EC, Shadduck JA. Mechanisms of resistance to the intracellular protozoan Encephalitozoon cuniculi in mice. J Immunol. 1984;133:2712–2719. PubMed

Sak B, Saková K, Ditrich O. Effects of a novel anti-exospore monoclonal antibody on microsporidial development in vitro. Parasitol Res. 2004;92:74–80. PubMed

Visvesvara GS, Moura H, Leitch J, Schwartz DA. Culture and propagation of microsporidia. In: Wittner M, Weiss LM, editors. The microsporidia and microsporidiosis. Washington: ASM Press; 1999. pp. 363–392.

Mo L, Drancourt M. Monoclonal antibodies for specific detection of Encephalitozoon cuniculi. Clin Diagn Lab Immunol. 2004;11:1060–1063. PubMed PMC

Jass A, Matiasek K, Hartmann K, Kuchenhoff H, Fischer A. Evaluierung von Liquoruntersuchung und PCR zur Diagnose der Enzephalitozoonose beim Kaninchen. Prakt Tierartz. 2006;87:518–524.

Jeklova E, Jekl V, Kovarcik K, Hauptman K, Koudela B, Neumayerova H, Knotek Z, Faldyna M. Usefulness of detection of specific IgM and IgG antibodies for diagnosis of clinical encephalitozoonosis in pet rabbits. Vet Parasitol. 2010;170:143–148. PubMed

Jeklova E, Leva L, Faldyna M. Lymphoid organ development in rabbits: major lymphocyte subsets. Dev Comp Immunol. 2007;31:632–644. PubMed

Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol. 2002;3:0034. PubMed PMC

Didier ES. Mammalian animal models of human microsporidiosis. In: Weiss LM, Becnel JJ, editors. Microsporidia. Pathogens of Opportunity. New Jersey: Wiley; 2014. pp. 307–325.

Kunstýr I, Lev L, Naumannm S. Humoral antibody response of rabbits to experimental infection with Encephalitozoon cuniculi. Vet Parasitol. 1986;21:223–232. PubMed

Wicher V, Baughn RE, Fuentealba C, Shadduck JA, Abbruscato F, Wicher K. Enteric infection with an obligate intracellular parasite, Encephalitozoon cuniculi, in an experimental model. Infect Immun. 1991;59:2225–2231. PubMed PMC

Csokai J, Joachim A, Gruber A, Tichy A, Pakozdy A, Künzel F. Diagnostic markers for encephalitozoonosis in pet rabbits. Vet Parasitol. 2009;163:18–26. PubMed

Künzel F, Gruber A, Tichy A, Edelhofer R, Nell B, Hassan J, Leschnik M, Thalhammer JG, Joachim A. Clinical symptoms and diagnosis of encephalitozoonosis in pet rabbits. Vet Parasitol. 2008;151:115–124. PubMed

Sak B, Salát J, Horká H, Saková K, Ditrich O. Antibodies enhance the protective effect of CD4+ T lymphocytes in SCID mice perorally infected with Encephalitozoon cuniculi. Parasite Immunol. 2006;28:95–99. PubMed

Moretto M, Weiss LM, Khan IA. Induction of a rapid and strong antigen-specific intraepithelial lymphocyte response during oral Encephalitozoon cuniculi infection. J Immunol. 2004;172:4402–4409. PubMed PMC

Didier ES, Shadduck JA. Modulated immune responsiveness associated with experimental Encephalitozoon cuniculi infection in BALB/c mice. Lab Anim Sci. 1988;38:680–684. PubMed

Khan IA, Moretto M. Role of gamma interferon in cellular immune response against murine Encephalitozoon cuniculi infection. Infect Immun. 1999;67:1887–1893. PubMed PMC

Khan IA, Schwartzman JD, Fonseka S, Kasper LH. Neospora caninum: role for immune cytokines in host immunity. Exp Parasitol. 1997;85:24–34. PubMed

Sternberg J, McGuigan F. Nitric oxide mediates suppression of T cell responses in murine Trypanosoma brucei infection. Eur J Immunol. 1992;22:2741–2744. PubMed

Moretto MM, Khan IA. IL-21 is important for induction of KLRG1+ effector CD8 T cells during acute intracellular infection. J Immunol. 2016;196:375–384. PubMed PMC

Salát J, Jelínek J, Chmelař J, Kopecký J. Efficiency of gamma interferon and specific antibody for treatment of microsporidiosis caused by Encephalitozoon cuniculi in SCID mice. Antimicrob Agents Chemother. 2008;52:2169–2174. PubMed PMC

Salát J, Horka H, Sak B, Kopecky J. Pure CD4+ T lymphocytes fail to protect perorally infected SCID mice from lethal microsporidiosis caused by Encephalitozoon cuniculi. Parasite Res. 2006;99:682–686. PubMed

da Costa LFV, Alvares-Saraiva AM, Dell’Armelina Rocha PR, Spadacci-Morena DD, Perez EC, Mariano M, Lallo MA. B-1 cell decreases susceptibility to encephalitozoonosis in mice. Immunobiology. 2017;222:218–227. PubMed

Didier ES, Shadduck JA. IFN-gamma and LPS induce murine macrophages to kill Encephalitozoon cuniculi in vitro. J Eukaryot Microbiol. 1994;41:34S. PubMed

Didier ES. Reactive nitrogen intermediates implicated in the inhibition of Encephalitozoon cuniculi (phylum Microspora) replication in murine peritoneal macrophages. Parasite Immunol. 1995;17:405–412. PubMed

Moretto MM, Lawlor EM, Khan IA. Lack of interleukin-12 in p40-deficient mice leads to poor CD8+ T-cell immunity against Encephalitozoon cuniculi infection. Infect Immun. 2010;78:2505–2511. PubMed PMC

Nevárez-Garza AM, Castillo-Velázquez U, Soto-Domínguez A, Montes-de-Oca-Luna R, Zamora-Ávila DE, Wong-González A, Rodríguez-Tovar LE. Quantitative analysis of TNF-α, IL-4, and IL-10 expression, nitric oxide response, and apoptosis in Encephalitozoon cuniculi-infected rabbits. Dev Comp Immunol. 2018;81:235–243. PubMed

Sak B, Ditrich O. Humoral intestinal immunity against Encephalitozoon cuniculi (Microsporidia) infection in mice. Folia Parasitol (Praha) 2005;52:158–162. PubMed

Curtis MM, Way SS. Interleukin-17 in host defence against bacterial, mycobacterial and fungal pathogens. Immunology. 2009;126:177–185. PubMed PMC

Mensikova M, Stepanova H, Faldyna M. Interleukin-17 in veterinary animal species and its role in various diseases: a review. Cytokine. 2013;64:11–17. PubMed

Francisco Neto A, Dell’Armelina Rocha PR, Perez EC, Xavier JG, Peres GB, Spadacci-Morena DD, Alvares-Saraiva AM, Lallo MA. Diabetes mellitus increases the susceptibility to encephalitozoonosis in mice. PLoS One. 2017;12:e0186954. PubMed PMC

Langanke Dos Santos D, Alvares-Saraiva AM, Xavier JG, Spadacci-Morena DD, Peres GB, Dell’Armelina Rocha PR, Perez EC, Lallo MA. B-1 cells upregulate CD8 T lymphocytes and increase proinflammatory cytokines serum levels in oral encephalitozoonosis. Microbes Infect. 2018;20:196–204. PubMed