J Fish Dis. 2020;00:1–15. wileyonlinelibrary.com/journal/jfd | 1 © 2020 John Wiley & Sons Ltd 1 | INTRODUCTION Myxozoans are morphologically diverse metazoan parasites. They exhibit a two-host life cycle involving an actinospore stage released by an invertebrate definitive host, typically an annelid, and a myxo- spore stage in a vertebrate intermediate host (Kent et al., 2001). The myxospore stage is predominantly found in fish, with limited reports from amphibians, birds and small mammals (Atkinson, Bartholomew, & Lotan, 2018). Myxospores consist of 1–13 polar capsules containing polar filaments, 2–13 shell valves with or without caudal processes, a nucleated sporoplasm and occasionally a vacuole. During sporo- genesis, the polar capsules and shell valves are produced from cap- sulogenic and valvulogenic cells, respectively (Current & Janovy Jr., 1977). Their morphology reflects a markedly simplified form of their evolutionary origins within the phylum Cnidaria (Atkinson et al., 2018). In particular, the myxozoan polar capsule is compared to the cnidarian nematocyst in form and function (Ben-David et al., 2016; Cannon & Wagner, 2003). The polar filament, ejected from the cap- sule upon contact with host mucin, attaches to the host and retracts to bring the spore and host into close proximity (Ben-David et al., 2016; Kallert, Ponader, Eszterbauer, El-Matbouli, & Haas, 2007). Received: 17 January 2020 | Revised: 19 February 2020 | Accepted: 21 February 2020 DOI: 10.1111/jfd.13152 ORIGINAL ARTILCE Insights into myxozoan composition and physiology revealed by histochemical properties of myxospores Justin M. Stilwell 1 | Matt J. Griffin 2 | Thomas G. Rosser 3 | Haitham H. Mohammed 4 | Inga F. Sidor 5 | Alvin C. Camus 1 1 Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA, USA 2 Department of Pathobiology and Population Medicine, College of Veterinary Medicine, Mississippi State University, Stoneville, MS, USA 3 Department of Basic Sciences, College of Veterinary Medicine, Mississippi State University, Stoneville, MS, USA 4 School of Fisheries, Aquaculture, and Aquatic Sciences, Auburn University, Auburn, AL, USA 5 New Hampshire Veterinary Diagnostic Laboratory, Department of Molecular, Cellular, and Biomedical Sciences, University of New Hampshire, Durham, NH, USA Correspondence Justin M. Stilwell, Department of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA, USA. Email: stilwellj@uga.edu Present address Haitham H. Mohammed, Department of Biological Sciences, University of Wisconsin–Milwaukee, Milwaukee, WI, USA and Department of Aquatic Animals Medicine and Management, Faculty of Veterinary Medicine, Assiut University, Assiut, Egypt Abstract Myxozoa (phylum Cnidaria) are a diverse group of metazoan parasites that predomi- nately infect fish. Little is known regarding the composition and physiology of their myxospore life stage. The objective of this work was to investigate the composition of myxospores and extrasporogonic stages of nine myxozoan species infecting various teleost fish using histochemical staining techniques. Thirty histochemical stains were applied to formalin-fixed, paraffin-embedded tissues processed routinely for light mi- croscopic evaluation. The polar capsules were the most consistent stain target across the taxa examined. Polar capsule staining with Alizarin red, von Kossa and methyl green-pyronin suggests the presence of intracapsular calcium and phosphate, which may contribute to polar filament discharge or pathogenesis of host invasion. The shell valves and suture lines of most myxozoans were stained with Luna and phosphotung- stic acid haematoxylin stains, consistent with the presence of chitin and microfibrils, respectively. Vacuoles were consistently highlighted by diastase-susceptible periodic acid-Schiff and Grocott's methenamine silver staining, indicating glycogen. Other histochemical stains exhibited inconsistent staining across the taxa, suggesting dif- ferences in myxospore composition potentially reflective of physiologic variations and tissue tropisms. This work provides some information on conserved features and taxa-associated composition of myxospores and lends insight into myxozoan physiol- ogy and host–parasite interactions. KEYWORDS histochemical, histopathology, myxospore, myxozoa