Investigating the Environmental Survival of Marteilia refringens, a Marine Protozoan Parasite of the Flat Oyster Ostrea edulis, Through an Environmental DNA and Microscopy-Based Approach

Abstract

Outside-host parasite survival is a key parameter to better understand disease mechanisms, especially for marine pathogens transmitted from one host to another through an environmental stage. For non-cultivable micro-parasites like Marteilia refringens, a protozoan parasite infecting the flat oyster Ostrea edulis, investigating this parameter requires innovative approaches. In the present study, we have developed an Environmental DNA (eDNA)-based method allowing detecting and quantifying up to 25 and 10 parasites DNA in seawater and sediment, respectively. This method was used in combination with light and transmission electron microscopy (TEM) to study experimentally parasite survival in seawater and flat oyster faeces after its release from naturally infected oysters. M. refringens DNA could be detected up to 20 days, in both seawater and oyster faeces with a more stable detection over time in the latter. Light and transmission microscopy confirm that parasites leaving flat oysters are sporangia. We also observed a membrane dissolution of the sporangia over time that could indicate the release of parasite spore. This study not only improves our understanding of M. refringens ecology but also highlights the interest to combine molecular and microscopical analysis to study non-cultivable micro-parasites life cycle outside their host.