Implementation of various approaches to study the prevalence, incidence and progression of disseminated neoplasia in mussel stocks

Abstract

Marine mussel production is of substantial economic interest in numerous coastal areas worldwide, making crucial the study of pathologies that affect them. Disseminated neoplasia (DN) has recently been suggested to be linked to blue mussel, Mytilus edulis, mortality outbreaks observed in France since 2014, although the evidence remains indirect. In order to improve DN detection and monitoring, we compared the sensitivity of four diagnostic tools, namely haemocytology, histology, flow cytometry, and genetics. Haemocytological examination gave the best results in sensitivity and had the advantage of being non-invasive, allowing disease progression to be followed in affected mussels. Using this approach, we showed that DN progression is usually slow, and we provide evidence of remission events. We observed a high diversity of forms and mitotic features of neoplastic cells located in the vesicular connective tissue but rarely in the haemolymph. Circulating cells occur as four main types but are homogenous in morphology and DNA content within a single individual. Polyploidy proved very high, from 8N to 18N. Genetic analysis of haemolymph DNA showed that a Mytilus trossulus genetic signal was associated with almost all the DN cases here diagnosed by haemocytological examination, regardless of the DN type. This result corroborates DN is a transmissible cancer that first originated in a M. trossulus host and subsequently crossed into M. edulis. No pre-neoplastic conditions were detectable. The prevalence of the disease was quite low, 1 which, together with the low morbidity observed in the lab, suggest DN is unlikely to be the direct cause of mortality outbreaks in France.

The root of the transmissible cancer: first description of a widespreadMytilus trossulus-derived cancer lineage inM. trossulus

“…The features of the neoplastic hemocytes in our study-a rounded shape, a large nucleus, a high nucleus-to-cytoplasm ratio and an increased ploidy-agree with the previous descriptions of DN in Mytilus 14,22,23 . What seems unusual is their low proliferation level.…”

“…Neoplastic hemocytes of mussels are generally assumed to have a high proliferation activity 20 . However, a low proliferation rate of neoplastic hemocytes in mussels with DN was reported in two other studies: of BTN (supposedly BTN2) in France 22 and of DN 66 in the same Argentinean population where BTN2 was recognized later 10 . A possible explanation is that in case of BTN2 the proliferation site of the neoplastic cells is located not in the hemolymph.…”

“…Normal hemocytes of healthy bivalves (agranulocytes and granulocytes) are diploid and have a low mitotic activity 18,19 . Neoplastic hemocytes are aneuploid [18][19][20][21][22] , and have a high proliferation rate and a striking morphology. They are rounded, with a high nucleus-to-cytoplasm ratio 14,22,23 , and their nucleus is pleomorphic (i.e.…”

“…As the disease progresses, the proliferating cells infiltrate other tissues, which leads to the loss of integration and normal functioning of organs 24,25 . DN may reach an epizootic level, causing mass mortality in wild and commercial bivalve populations 14,22,24 .…”

“…It employs flow cytometry of hemocytes labeled with fluorescent dyes specifically binding DNA 19 . Flow cytometry is less sensitive to the initial stages of DN than hemocytology but allows an efficient examination of large samples 19,22 .…”