Search

In the fifty years since the introduction of the Pacific oyster Crassostrea gigas and the first reports of the parasites Marteilia refringens and Bonamia ostreae in European waters, numerous research projects dedicated to the native European flat oyster Ostrea edulis have been conducted, notably in France. Most of these projects have been dedicated to developing controlled reproduction and hatchery technology for seed production, examining pathological aspects to understand and control diseases, and using genetics to develop resistant lines. While the long-term objective of most studies has been to revive the aquaculture production of O. edulis, a smaller number have addressed the ecology of local remnant beds and reefs in France. This article provides an overview of the major outcomes of all these projects, focusing on results obtained in France and prospects for future work there, taking into account the rising interest in increasing aquaculture production and ecological motivation to restore declining populations as part of the framework of the Native Oyster Restoration Alliance (NORA) and in line with UN Decade for Ecosystem Restoration. [ABSTRACT FROM AUTHOR]

The Pacific cupped oyster Crassostreagigas is one of the most 'globalized' marine invertebrates and its production is predominant in many parts of the world including Europe. However, it is threatened by mortality events associated with pathogenic microorganisms such as the virus OsHV‐1 and the bacteria Vibrio aestuarianus. C. gigas is also a host for protozoan parasites including haplosporidians. In contrast with Haplosporidium nelsoni previously detected in Europe, H. costale was considered exotic although its presence in French oysters was suggested in the 1980s based on ultrastructural examination. Here, a combination of light and transmission electron microscopy, PCR and sequencing allowed characterizing the presence of the parasite in the context of low mortality events which occurred in 2019 in France. Histological observation revealed the presence of uninucleated, plasmodial and spore stages within the connective tissues of some oysters. Ultrastructural features were similar to H. costale ones in particular the presence of axe‐shaped haplosporosomes in spore cytoplasms. Three fragments of the genome including partial small subunit rRNA gene, the ITS‐1, 5.8S and ITS‐2 array and part of the actin gene were successfully sequenced and grouped with H. costale homologous sequences. This is the first time that the presence of H. costale was confirmed in C. gigas in France. Furthermore, a TaqMan real‐time PCR assay was developed and validated [DSe = 92.6% (78.2–99.8) and DSp = 95.5% (92.3–98.6)] to enable the rapid and specific detection of the parasite. The application of the PCR assay on archived samples revealed that the parasite has been present in French oyster populations at least since 2008. Considering the little information available on this parasite, the newly developed TaqMan assay will be very helpful to investigate the temporal and geographic distribution and the life cycle of the parasite in France and more generally in C. gigas geographic range. [ABSTRACT FROM AUTHOR]

Abstract: In April 2010, the sampling of juvenile Pacific cupped oysters originating from France was undertaken from a farm located off the coast of the Marche region in Italy. Samples were sent to the national reference laboratory for mollusc diseases for bacteriological, histological, electron microscopical and molecular analysis. Classical and Real-Time PCR indicated the presence of the microvariant (OsHV-1 μvar) of Type 1 Ostreid Herpes virus (OsHV-1) despite the absence of clinical and pathological signs normally associated with the presence of this variant. Further molecular and sequencing analysis revealed the presence of OsHV-1 μvar and OsHV-1 DNA in one oyster indicating that simultaneous detection of both viruses is possible. This is the first report of the presence of OsHV-1 μvar in Italy and also the first time that evidence of possible co-infection of OsHV-1 and OsHV-1 μvar has been provided. [Copyright &y& Elsevier]

Paracartia grani originates from high latitudes and has progressively been recorded in the Mediterranean Sea since the 1980s. In Thau lagoon, while Acartia clausi and Acartia discaudata present maximum peaks of abundance from January to April and in November and December, both species being perennial over the year, P. grani and Paracartia latisetosa appear in May, develop during summer, in July and August, and slowly decrease in autumn before disappearing from the water column in December. The Acartiidae populations have changed over the years at our monitoring station. Indeed, P. latisetosa was last recorded in August 1983 while P. grani appeared in the lagoon in 1998. [ABSTRACT FROM AUTHOR]

Abstract: Ostreid herpesvirus 1 (OsHV-1) infections have been reported around the world and are associated with high mortalities of the Pacific oyster (Crassostrea gigas). In the summer 2008, abnormal mortality rates ranging from 80% to 100% were reported in France and affected only Pacific oysters. Analyses of oyster samples collected during mortality outbreaks demonstrated a significant detection of OsHV-1 (75% of analysed batches), which appeared stronger than previous years. DNA sequencing based on C and IA regions was carried out on 28 batches of OsHV-1 infected Pacific oysters collected in 2008. Polymorphisms were described in both the C and IA regions and characterized a genotype of OsHV-1 not already reported and termed OsHV-1 μVar. A microsatellite zone present in the C region showed several deletions. Additionally, 44 isolates collected in France and in the USA, from 1995 to 2007 were sequenced and compared to the 2008 sequences. The analyses of 76 sequences showed OsHV-1 μVar detection only in 2008 isolates. These data suggest that OsHV-1 μVar can be assumed as an emergent genotype. [Copyright &y& Elsevier]

• M. refringens, B. ostreae and B. exitiosa are regulated parasites of flat oysters. • First validated PCR detecting all regulated pathogens of flat oysters in one step. • Multiplex PCR performances similar or higher than currently recommended PCR assays. • Represents a substantial improvement in time, resources and accuracy. • Useful to monitors parasites prevalence or for demonstration of freedom. Aquaculture including shellfish production is an important food resource worldwide which is particularly vulnerable to infectious diseases. Marteilia refringens, Bonamia ostreae and Bonamia exitiosa are regulated protozoan parasites infecting flat oysters Ostrea edulis that are endemic in Europe. Although some PCR assays have been already developed for their detection, a formal validation to assess the performances of those tools is often lacking. In order to facilitate the diagnosis of flat oyster regulated diseases, we have developed and evaluated a new multiplex Taqman® PCR allowing the detection of both M. refringens and Bonamia sp. parasites in one step. First part of this work consisted in assessing analytical sensitivity and specificity of the new PCR assay. Then, diagnostic performances were assessed by testing a panel of field samples with the new real-time PCR and currently recommended conventional PCR methods for the detection of M. refringens and Bonamia sp. Samples were collected from the main flat oyster production sites in France (N = 386 for M. refringens and N = 349 for B. ostreae). In the absence of gold standard, diagnostic sensitivity and specificity of the new PCR were estimated through Bayesian latent class analysis (DSe 87,2% and DSp 98,4% for the detection M. refringens , DSe 77,5% and DSp 98,4% for the detection of Bonamia sp.). Those results suggest equivalent performances for the detection of Bonamia sp. and an improved sensitivity for the detection of M. refringens compared to commonly used conventional protocols. Finally, the new PCR was evaluated in the context of an inter-laboratory comparison study including 17 European laboratories. Results revealed a very good reproducibility with a global accordance (intra-laboratory precision) >96% and a global concordance (inter-laboratory precision) >93% for both targets, demonstrating that this new tool is easily transferable to different laboratory settings. This is the first assay designed to detect both Marteilia refringens and Bonamia sp. in a single step and it should allow reducing the number of analysis to monitor both diseases, and where relevant to demonstrate freedom from infection. [ABSTRACT FROM AUTHOR]

The protozoan parasite Bonamia ostreae has been associated with the decline of flat oyster Ostrea edulis populations in some European countries. Control of shellfish diseases mostly relies on prevention measures including transfer restrictions and stock management measures such as breeding programmes. These prevention and mitigation measures require a better understanding of interactions between host and pathogens. Previous in vitro studies allowed identifying apoptosis as a mechanism activated by the flat oyster in response to B. ostreae. However, these experiments also suggested that the parasite is able to regulate apoptosis in order to survive and multiply within hemocytes. By simplifying the conditions of infection, in vitro studies allow identifying most distinct features of the response of the host. In order to appreciate the relative importance of apoptosis in this response at the oyster scale, in vivo trials were carried out by injecting with parasites oysters from two French locations, Quiberon Bay (Brittany) and Diana Lagoon (Corsica). Apoptosis was investigated on pools of hemolymph from oysters collected at early and later times after injection using previously developed tools. Apoptotic cellular activities including intracytoplasmic calcium concentration, mitochondrial membrane potential and phosphatidyl serine externalization were analysed using flow cytometry. Moreover, the expression of flat oyster genes involved in both extrinsic and intrinsic pathways was measured using real time quantitative PCR., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Microcell parasites are small intracellular protozoans mostly detected in molluscs and can be associated with mortalities. In 2010 and 2011, strong increases in mortality events were reported in different wild beds of the wedge clam Donax trunculus Linnaeus, along the Atlantic coast of France and the presence of potential pathogens, including microcells, was investigated., Methods: Clams collected in different beds showing mortality were examined by histology. Based on histological observations, confirmatory analyses were carried out, including transmission electron microscopy (TEM) and molecular characterization., Results: Histological analyses revealed the presence of small protozoans similar to microcell parasites in different tissues of Donax trunculus, particularly in muscular and connective tissues. TEM examination confirmed the intracellular localization of the protozoans. Moreover, the lack of haplosporosomes and mitochondria suggested that the observed parasites belong to the genus Mikrocytos Farley, Wolf & Elston, 1988. Mikrocytos genus-specific PCR and in situ hybridization results supported the microscopic observations. Sequence fragments of the 18S rRNA gene shared 75-83% identity with the different Mikrocytos spp. described previously, including Mikrocytos mackini Farley, Wolf & Elston, 1988 and M. boweri Abbott, Meyer, Lowe, Kim & Johnson, 2014. Phylogenetic analyses confirmed that the microcell parasites observed in Donax trunculus in France belong to the genus Mikrocytos and suggest the existence of two distinct species., Conclusions: Based on morphological, ultrastructural, molecular data and host information, the two microcell parasites detected in Donax trunculus belong to the genus Mikrocytos and are distinct from previously described members of this genus. This is the first report of Mikrocytos spp. found in France and infecting the clam Donax trunculus. Mikrocytos veneroïdes n. sp. was detected in different wild beds and Mikrocytos donaxi n. sp. was detected only in Audierne Bay.

Since its molecular characterisation, Ostreid herpesvirus 1 (OsHV-1) has been regularly detected in Crassostrea gigas in France. Although its pathogenicity was demonstrated on larval stages, its involvement during mortality outbreaks at the juvenile stage was highly suspected but not evidenced. To investigate mortality outbreaks, the French National Network for Surveillance and Monitoring of Mollusc Health (REPAMO) carried out two surveys in juvenile C. gigas. The first survey lasted from 1998 to 2006 and was an epidemiological inquiry occurring when oyster farmers reported mortality outbreaks. The second survey, a longitudinal one, was set up in 1998 to complete the network observations on OsHV-1. Data analysis showed a specific pattern of mortality outbreaks associated with OsHV-1 detection. Ostreid herpesvirus 1 detection mainly appeared during the summer, suggesting the influence of the seawater temperature on its occurrence. It mostly presented a patchy distribution in the field in contrast to the nursery. Significant relationship between OsHV-1 detection and spat mortality was found, preferentially in sheltered and closed environments. The longitudinal survey confirmed most of the network observations. Although subsequent works particularly epidemiological surveys would be useful to confirm the causal link between the detection of OsHV-1 and the mortality outbreaks in juvenile C. gigas, the role of OsHV-1 in oyster mortality is progressing.

Viral gametocytic hypertrophy was reported for the first time in 2001 in Pacific oyster Crassostrea gigas in France. Since this date, the number of reported cases and the distribution area have increased every year; however, the cases are not associated with macroscopic signs or increased mortality rates. Both male and female gametes were hypertrophied and basophilic inclusions were observed in gamete nuclei. Transmission electron microscopy revealed the presence of viral particles in these intranuclear basophilic inclusions. These particles had characteristics similar to those of the Papillomaviridae and Polyoma viridae families: they were small, non-enveloped, icosahedral, and 44 to 56 nm in diameter. The viral particles were found in male, female and hermaphrodite oysters and no significant difference in viral infection was observed between those groups. The frequency of detection and the intensity of infection were low and no host defence reaction was recognised, suggesting that the viral particles had a weak impact on C. gigas. The viral particles described in the present study seem to be similar to these described in C. virginica in the USA and Canada and in C. gigas in Korea, but further studies are required to confirm their identity. The issue of a possible emergence of this infection is discussed.