Bivalve digestive epithelial virosis (DEV): A cause of disease or a natural process?

Epithelial hyperplasia and sloughing of the digestive gland in bivalve mollusks are a global phenomenon and occur in species of commercial interest and cultural significance to indigenous peoples. Where hemocytosis, hyperplasia, and necrosis of digestive tubule cells have been observed associated with electron-dense uncoated virus-like particles (VLPs) 25-45 nm in diameter, the condition has been named digestive epithelial virosis (DEV). This condition has been associated with mortalities of some bivalve species in New Zealand. Similar digestive gland alterations, but without detection of associated VLPs, have been reported in other bivalve species worldwide and are termed "DEV-like" since no virus link has been demonstrated. It remains unclear if DEV is an infectious condition and whether associated VLPs are the cause, a contributor, or simply associated with the observed condition. It is also unclear whether DEV or DEV-like conditions pose a biosecurity or economic threat, or alternatively, whether they reflect a natural cyclic event that does not require disease management. In this mini-review, we summarize the history of digestive epithelial alteration with VLPs (i.e., DEV) or without observation of VLPs (i.e., DEV-like), and we examine the evidence for and against viral-like particles as the cause of DEV in bivalves. We also explore other viral afflictions of bivalves and non-infectious agents, such as harmful algae and xenotoxins, that could elicit similar tissue alterations. Future recommendations for approaches to identify key risk factors that lead to the development of digestive epithelial alterations such as DEV include histological characterization of the digestive gland of marine mollusks; the use of metagenome analysis to design primers that could be used for detection of VLPs and to study host microbiota; disease challenges demonstrating that DEV causes pathology and the relationship between DEV intensity and morbidity/mortality.

Flow cytometric characterization of hemocytes of the flat oyster, Ostrea chilensis.

The flat oyster, Ostrea chilensis, native to New Zealand (NZ) and Chile is considered an important ecological, cultural and fisheries resource. Currently, commercial landings of this species in NZ are restricted due to low population numbers caused by ongoing mortalities resulting from the presence of the haplosporidian parasite, Bonamia exitiosa. More recently, the arrival of B. ostreae in NZ led to major mortalities in farmed stocks. To understand how diseases caused by Bonamia spp. affect this oyster species, a more complete understanding of its biology, physiology and immune system is needed. The present study characterized, for the first time, hemocytes of adult O. chilensis, from the Foveaux Strait, NZ, using flow cytometry (FCM) and histology. Based on the internal complexity of the hemocytes, two main circulating hemocyte populations were identified: granulocytes and hyalinocytes (accounting for ~30% and ~70% of the total circulating hemocyte population, respectively). These were further divided into two sub-populations of each cell type using FCM. A third sub-population of granulocytes was identified using histology. Using FCM, functional and metabolic characteristics were investigated for the two main hemocyte types. Granulocytes showed higher phagocytic capabilities, lysosomal content, neutral lipid content and reactive oxygen species production compared to hyalinocytes, indicating their important role in cellular immune defence in this species. Methods of hemocyte sampling and storage were also investigated and flow cytometric protocols were detailed and verified to allow effective future investigations into the health status of this important species.

Bonamia ostreae in the New Zealand oyster Ostrea chilensis: a new host and geographic record for this haplosporidian parasite.

Previous reports of the haplosporidian parasite Bonamia ostreae have been restricted to the Northern Hemisphere, including Europe, and both eastern and western North America. This species is reported for the first time in New Zealand infecting the flat oyster Ostrea chilensis. Histological examination of 149 adult oysters identified 119 (79.9%) infected with Bonamia microcells. Bonamia generic PCR of several oysters followed by DNA sequencing of a 300 bp portion of the 18S rDNA gene produced a 100% match with that of B. ostreae. All DNA-sequenced products also produced a B. ostreae PCR-restriction fragment length polymorphism (PCR-RFLP) profile. Bonamia species-specific PCRs further detected single infections of B. exitiosa (2.7%), B. ostreae (40.3%), and concurrent infections (53.7%) with these 2 Bonamia species identifying overall a Bonamia prevalence of 96.6%. Detailed histological inspection revealed 2 microcell types. An infection identified by PCR as B. ostreae histologically presented small microcells (mean ± SE diameter = 1.28 ± 0.16 µm, range = 0.9-2 µm, n = 60) commonly with eccentric nuclei. A B. exitiosa infection exhibited larger microcells (mean ± SE diameter = 2.12 ± 0.27 µm, range = 1.5-4 µm, n = 60) with more concentric nuclei. Concurrent infections of both Bonamia species, as identified by PCR, exhibited both types of microcells. DNA barcoding of the B. ostreae-infected oyster host confirmed the identification as O. chilensis. A suite of other parasites that accompany O. chilensis are reported here for the first time in mixed infection with B. ostreae including apicomplexan X (76.5%), Microsporidium rapuae (0.7%) and Bucephalus longicornutus (30.2%).