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Article

Method for Isolation of Myxozoan Proliferative Stages from Fish at High Yield and Purity: An Essential Prerequisite for In Vitro, In Vivo and Genomics-Based Research Developments

1
Institute of Parasitology, Biology Centre of the Czech Academy of Sciences, 37005 České Budějovice, Czech Republic
2
Laboratory of Ecological Plant Physiology, Global Change Research Institute of the Czech Academy of Sciences, 60300 Brno, Czech Republic
3
South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses, Faculty of Fisheries and Protection of Waters, University of South Bohemia, 37005 České Budějovice, Czech Republic
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
Academic Editors: Alexander E. Kalyuzhny and Annemarie H. Meijer
Cells 2022, 11(3), 377; https://doi.org/10.3390/cells11030377
Received: 23 November 2021 / Revised: 11 January 2022 / Accepted: 17 January 2022 / Published: 23 January 2022
(This article belongs to the Special Issue 10th Anniversary of Cells—Advances in Cell Techniques)
Myxozoans are a diverse group of microscopic cnidarian parasites and some representatives are associated with important diseases in fish, in both marine and freshwater aquaculture systems. Research on myxozoans has been largely hampered by the inability to isolate myxozoan parasites from their host tissues. In this study, we developed and optimized a method to isolate the myxozoan proliferative stages of different size and cellularity from fish blood, using DEAE-cellulose ion exchange chromatography. We optimized several parameters and obtained 99–100% parasite purity, as well as high survival and infectivity. Using polyclonal pan-carp blood cell-specific antibodies, we further developed a rapid cytometric assay for quantification of the proliferative stages, not only in highly concentrated DEAE-C isolates but also in dilute conditions in full blood. Early developmental stages of myxozoans are key to parasite proliferation, establishment, and pathology in their hosts. The isolation of these stages not only opens new possibilities for in vivo and in vitro studies, but also for obtaining purified DNA and protein extracts for downstream analyses. Hence, we provide a long-desired tool that will advance the functional research into the mechanisms of host exploitation and immune stimulation/evasion in this group, which could contribute greatly to the development of therapeutic strategies against myxozoans. View Full-Text
Keywords: diethylaminoethyl (DEAE) cellulose; cell separation; cytometry; anti-carp antibody; Sphaerospora; parasite; blood stages; teleost; common carp diethylaminoethyl (DEAE) cellulose; cell separation; cytometry; anti-carp antibody; Sphaerospora; parasite; blood stages; teleost; common carp
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MDPI and ACS Style

Born-Torrijos, A.; Kosakyan, A.; Patra, S.; Pimentel-Santos, J.; Panicucci, B.; Chan, J.T.H.; Korytář, T.; Holzer, A.S. Method for Isolation of Myxozoan Proliferative Stages from Fish at High Yield and Purity: An Essential Prerequisite for In Vitro, In Vivo and Genomics-Based Research Developments. Cells 2022, 11, 377. https://doi.org/10.3390/cells11030377

AMA Style

Born-Torrijos A, Kosakyan A, Patra S, Pimentel-Santos J, Panicucci B, Chan JTH, Korytář T, Holzer AS. Method for Isolation of Myxozoan Proliferative Stages from Fish at High Yield and Purity: An Essential Prerequisite for In Vitro, In Vivo and Genomics-Based Research Developments. Cells. 2022; 11(3):377. https://doi.org/10.3390/cells11030377

Chicago/Turabian Style

Born-Torrijos, Ana, Anush Kosakyan, Sneha Patra, Joana Pimentel-Santos, Brian Panicucci, Justin T.H. Chan, Tomáš Korytář, and Astrid S. Holzer. 2022. "Method for Isolation of Myxozoan Proliferative Stages from Fish at High Yield and Purity: An Essential Prerequisite for In Vitro, In Vivo and Genomics-Based Research Developments" Cells 11, no. 3: 377. https://doi.org/10.3390/cells11030377

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