Development of an Optimized Protocol for the Cytological Evaluation of Glass Anemones (Aiptasia pulchella)
IAAAM 2018
Sarah J. Wahlstrom1*+; Thierry M. Work2; Nicole I. Stacy3; Renee Breeden2
1College of Veterinary Medicine, The Ohio State University, Columbus, OH, USA; 2US Geological Survey, National Wildlife Health Center, Honolulu Field Station, Honolulu, HI, USA; 3Department of Large Animal Clinical Sciences, Department of Aquatic Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL, USA

Abstract

Coral reefs are known as some of the most biodiverse and complex ecosystems in the world, containing an estimated 25% of the world’s marine fish species in less than 0.1% of the ocean’s floor,1 with significant ecological, economic, and culture significance. As of 2008, 19% of coral reefs had died with continually increasing rates of decline.2 Research on stony corals is hampered by the inability to remove coral cells from their calcium carbonate skeleton and difficulty maintaining them in a laboratory environment. A closely related symbiotic Anthozoan, the glass anemone Aiptasia provides an ideal model for studying coral as they lack a skeleton, are easily maintained in a laboratory and are capable of both asexual and sexual reproduction.3 To dissociate the organism for cytology, Aiptasia were first relaxed in a bath of 2.5% MgCl2 in artificial sea water (ASW) for euthanasia then macerated with sharp scissors. Aiptasia fragments were then placed in one of three maceration solutions: 1:1:13 glycerin:glacial acetic acid:3% NaCl in DI H2O (GAS), 1:13 glycerin:3% NaCl in DI H2O (GS), or 1:13 glacial acetic acid:3% NaCl in DI H2O (AS).4,5 After dissociation, the cellular monolayer was stained with modified Wright-Giemsa and evaluated under compound light microscopy. While fine cellular detail was best preserved in GS, cellular dissociation was most complete in GAS and AS indicating that acetic acid may play a role in dissociation of Aiptasia tissue. Identifiable cellular components included epithelial cells, secretory cells, zooxanthellae, bacteria, spirocysts, and several stages of nematocysts including microbasic p-mastigophores, B-mastigophores, and holotrichous isorhiza. Here we present an optimized protocol that will be useful in obtaining high quality diagnostic results in the cytologic evaluation of Aiptasia.

Acknowledgements

The authors wish to thank the staff of the Kewalo Marine Laboratory for their support of the project.

* Presenting author
+ Student presenter

Literature Cited

1.  Spalding MD, Ravilious C, Green EP, United Nations Environment Programme, World Conservation Monitoring Centre. 2001. World Atlas of Coral Reefs. Berkeley, CA: University of California Press; 416.

2.  Global Coral Reef Monitoring Network, Reef and Rainforest Research Centre. Wilkinson C, ed. 2008. Status of coral reefs of the world. Townsville, Australia. 296.

3.  Hambleton EA, Guse A, Pringle JR. 2014. Similar specificities of symbiont uptake by adults and larvae in an anemone model system for coral biology. J Exp Biol. 217(9):1613–1619.

4.  David CN. 1973. A quantitative method for maceration of hydra tissue. Roux Arch Dev Biol. 171:259–268.

5.  David CN. 1983. Dissociating hydra tissue into single cells by the maceration technique. In: Lenhoff HM, ed. Hydra: Research Methods. New York, NY: Plenum Press; 153–156.

Speaker Information
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Sarah J. Wahlstrom
College of Veterinary Medicine
The Ohio State University
Columbus, OH, USA


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