Cargando…
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
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...
Autores principales: | , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8833907/ https://www.ncbi.nlm.nih.gov/pubmed/35159187 http://dx.doi.org/10.3390/cells11030377 |
_version_ | 1784649051737686016 |
---|---|
author | Born-Torrijos, Ana Kosakyan, Anush Patra, Sneha Pimentel-Santos, Joana Panicucci, Brian Chan, Justin Tze Ho Korytář, Tomáš Holzer, Astrid S. |
author_facet | Born-Torrijos, Ana Kosakyan, Anush Patra, Sneha Pimentel-Santos, Joana Panicucci, Brian Chan, Justin Tze Ho Korytář, Tomáš Holzer, Astrid S. |
author_sort | Born-Torrijos, Ana |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-8833907 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-88339072022-02-12 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 Born-Torrijos, Ana Kosakyan, Anush Patra, Sneha Pimentel-Santos, Joana Panicucci, Brian Chan, Justin Tze Ho Korytář, Tomáš Holzer, Astrid S. Cells Article 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. MDPI 2022-01-23 /pmc/articles/PMC8833907/ /pubmed/35159187 http://dx.doi.org/10.3390/cells11030377 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Born-Torrijos, Ana Kosakyan, Anush Patra, Sneha Pimentel-Santos, Joana Panicucci, Brian Chan, Justin Tze Ho Korytář, Tomáš Holzer, Astrid 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 |
title | 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 |
title_full | 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 |
title_fullStr | 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 |
title_full_unstemmed | 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 |
title_short | 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 |
title_sort | 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 |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8833907/ https://www.ncbi.nlm.nih.gov/pubmed/35159187 http://dx.doi.org/10.3390/cells11030377 |
work_keys_str_mv | AT borntorrijosana methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT kosakyananush methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT patrasneha methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT pimentelsantosjoana methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT panicuccibrian methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT chanjustintzeho methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT korytartomas methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments AT holzerastrids methodforisolationofmyxozoanproliferativestagesfromfishathighyieldandpurityanessentialprerequisiteforinvitroinvivoandgenomicsbasedresearchdevelopments |