What's New in Bivalve Mollusc Pathology?

24, Bull. Eur. Ass. Fish Pathol., 36(1) 2016

What’s new in bivalve mollusc pathology Overview o articles published in the last two years

A. Villalba

Centro de Investigacións Mariñas (CIMA), Consellería do Mar da Xunta de ǰȱřŜŜŘŖȱȱȱǰȱǲȱ ¢ȱǰȱȱȱ Life Sciences, University of Alcalá, 28871 Alcalá de Henares, Spain

Abstract This is an overview o articles on bivalve mollusc diseases published between the two last Inter- national Conerences on Diseases o Fish and Shellsh organised by the EAFP (September 2013 to September 2015). The need to limit the number o pages o this overview has involved selecting (and discarding) articles, prioritising by novelty and relevance, which entails an element o subjectivity.

Journal special issues Highly relevant was the publication o special will be key reerences in their respective elds issues ocused on this eld by three journals or years. (in chronological order): (1) Microcell parasites o molluscs (Diseases of Aquatic Organisms Immune response 110 (1-2), 2014), including recent research on Bivalve immune response captured much at- protistan parasites o molluscs in the genera tention in the reviewed period and numerous, Bonamia and Mikrocytos; (2) Molluscan Im- excellent articles were published, mostly on host munity (ȱǭȱęȱ ¢ 46 (1), transcriptomic/proteomic changes associated 2015), including reviews o many aspects o with pathogens/diseases (viruses, bacteria, pro- the immunology o molluscs as well as an tistan, stress, etc), showing the complexity o the announcement o the orthcoming eort to immune system o bivalve molluscs. Hopeully, sequence the genome o the eastern oyster, advances in this eld will be used to design Crassostrea virginica; and (3) Pathogens and strategies to ght diseases o economically rel- Disease Processes in Marine Molluscs (Journal evant molluscs, such as identiying molecular of Invertebrate Pathology 131, 2015), ocusing markers o disease resistance. The list o articles on new knowledge about the pathogens o is too long to address bivalve immunity in this commercially important marine molluscs, ex- overview; nevertheless, the increasing aen- panding the scope to include associated topics tion paid to apoptosis within bivalve immune important to understand disease processes. context deserves to be mentioned (reviewed by Articles included in these three special issues Romero et al., 2015); the apoptotic eect o UV

* Corresponding author’s e-mail: [email protected] Bull. Eur. Ass. Fish Pathol., 36(1) 2016, 25 radiation on haemocytes o the European at Regarding sampling procedures and diagnostic oyster Ostrea edulis suggests that apoptosis is tools, new primers or PCR diagnosis were an important mechanism against stress actors assayed (Lynch et al., 2013). Three procedures (Gervais et al., 2015). (qPCR, evaluation o mortality provoked by thermal challenge and qPCR during thermal Viruses challenge) were assessed to prevent mortality The inection o the Pacic oyster Crassostrea (Normand et al., 2014a). A centriugation pro- gigas with the Ostreid herpesvirus 1 (OsHV1) cedure was arranged to improve viral detection is one o the bivalve mollusc pathology items in water samples (Evans et al., 2014). A survey- standing out or the high number o articles ing protocol, using hydrodynamic dispersion published in the last two years, which reects models to determine epidemiological units, the economic relevance o this virus or the was designed (Pande et al., 2015). An in situ widespread production o C. gigas; this research hybridisation assay or detecting OsHV1 mRNA eort, on a wide thematic spectrum, has result- was designed (Corbeil et al., 2015). ed in useul knowledge to ght this disease. OsHV1 genetic variability within geographic The pathogenesis o inection with OsHV1-μvar and host ranges was reported rom Mexico (Gri- in oyster was also addressed. A proteomic jalva-Chon et al., 2013), South Korea (Hwang et approach showed cytoskeleton disorganisa- al., 2013), China (Bai et al., 2015), Portugal and tion, dysregulation o protein turnover, stress Spain (Batista et al., 2015) and The Netherlands signals, dysregulation o steroid metabolism, (Gienberger et al., 2015). Temporal variation and Warburg eect (Corporeau et al., 2014). o genetic prole was also detected in Spain Additionally, metabolic changes in the oyster (Andree et al., 2014) and Ireland (Morrisey et were detected (Tamayo et al., 2014). A protocol al., 2015). The widest perspective on genetic or experimental inection (Paul-Pont et al., variability was provided by Mineur et al. (2015). 2015) will allow urther characterisation.

A genome analysis o OsHV1 ocused on 40 Research was also ocused on the oyster immune ORFs allowed distinguishing virus variants response to OsHV1 and the search or oyster re- (Martenot et al., 2013) while genotyping a mi- sistance associated genes. The analysis o oyster crosatellite locus allowed dierentiating clinical transcriptome across mortality episodes in the OsHV1 specimens (Renault et al., 2014a). Viral eld, using microarray approach (Jouaux et al., transcriptome was also addressed; analysis 2013), and ater experimental challenge with Os- o the expression o 39 viral genes provided HV1-μvar, through Illumina RNA sequencing inormation on the virus cycle based on the ki- (He et al., 2015), showed oyster exhibit a strong netics o viral DNA replication and transcription and complex anti-viral response. Two MyD88 (Segarra et al., 2014a, 2014b) and dierences o genes o the Pacic oyster involved in the an- gene expression were ound between viruses ti-viral immune response were characterised inecting oyster spat and those inecting adults (Du et al., 2013) and candidate genes to be used (Segarra et al. 2014c). in marker-assisted selection or resistance were identied (Normand et al. 2014b). Induction o 26, Bull. Eur. Ass. Fish Pathol., 36(1) 2016 oyster anti-viral response by challenging them Husbandry and management procedures were with double strand RNA was demonstrated, tested to minimise problems due to this disease. involving TLR, MyD88, IRF, PKR genes and Bearing in mind all those host and environ- protecting oysters rom OsHV1-μvar but not mental risk actors, mitigation strategies can be rom Vibrio splendidus (Green and Montagnani, designed (Peon et al., 2015). Oyster mortality 2013); the anti-viral induction is inuenced by at nursery acilities can be prevented by treating temperature and host age (Green et al., 2014a) pumped water, namely aging water or 48 h and and is absent in a second challenge with double ltration through 5 μm, which supports the stranded RNA (Green et al., 2014b). Anti-her- hypothesis that OsHV1 is carried on particles pesvirus activity o cavortin, the major Pacic (Whiington et al., 2015a). Elevating growing oyster plasma protein, was demonstrated height in intertidal oyster culture areas aected (Green et al., 2014c). by this disease, in Woolooware Bay, Australia, was eective or adult oysters but not or spat Host actors, especially size but also age, gen- (Whiington et al., 2015b). Selective breeding otype (genetic selection) and ploidy (triploid/ or resistance appears as the most promising diploid) were shown to inuence oyster suscep- strategy (reviewed by Degremont et al., 2015a); tibility to OsHV1-μvar (Degremont, 2013); the oyster lines that had been selected or resistance percentage o somatic aneuploidy cells may be against summer mortality (with a amily-based associated with oyster susceptibility (Batista et design) were less susceptible to OsHV1-μvar al., 2014). Regarding the inuence o environ- than non-selected oysters (Degremont, 2013). mental actors on disease dynamics, seawater Furthermore, successul results were aained temperature above 16 C avours disease pro- through classic mass selection: 69% survival gression and oyster mortality (Degremont, 2013; o the 4th generation selected oysters vs. 7% o Peon et al., 2013, 2015; Renault et al., 2014b); non-selected ones (Degremont et al., 2015b). The seawater temperature also inuences how long heritability or oyster mortality ranged rom 0.49 the virus keeps inective in the water (Martenot to 0.60 (Degremont et al., 2015c), which supports et al., 2015). Availability o good quality ood the potential o the selective-breeding strategy. (diatoms) decreases mortality risk (Pernet et al., 2014a), while the dinoagellate Alexandrium Bacteria catenella aects OsHV1-oyster interaction and Most articles on bacteria and bivalve health reduces inection prevalence (Lassudrie et al., were ocused on particular species or groups al- 2015). Field studies during OsHV1-μvar out- though a ew o them involved wide taxonomic breaks associated with Pacic oyster mortality spectrum. The search or probiotic bacteria that were perormed in Italy (Domeneghei et al., can be used or bivalve mollusc rearing has con- 2014), New Zealand (Keeling et al., 2014) and tinued (Newaj-Fyzul et al., 2014; Escamilla-Mon- Australia (Paul-Pont et al., 2014), taking the tes et al., 2015). Bacteriological issues associated chance to evaluate risk actors in Ireland (Clegg with bivalve hatcheries in Chile were reviewed et al., 2014) and France (Pernet et al., 2014b). by de la Fuente et al. (2015). The eects o temperature, temperature stress and challenge with a virulent Vibrio sp. strain on the microbiota o Bull. Eur. Ass. Fish Pathol., 36(1) 2016, 27 the Pacic oyster C. gigas haemolymph were ad- and V. tubiashii (Mersni-Achour et al., 2014, dressed analysing the microbioma (Lokmer and 2015). Brown ring disease o Manila clams R. Wegner, 2015). Changes in bacterial microbiota phillipinarum was also subject o study, in par- were characterised through culture process o ticular the population structure, evolution and oysters ȱ£, C. gigas and C. taxonomy o Vibrio tapetis (Balboa and Romalde, sikamea (Trabal Fernndez et al. 2014). 2013; Balboa et al., 2014) and disease dynamics (Paillard et al., 2014). Regarding vibrios, articles describing new Vibrio species isolated rom bivalve molluscs were Mollusc nocardiosis seemed to spread in Europe published (Gonzlez Castillo et al., 2014; Lasa because Nocardia crassostreae was detected in et al. 2014; Prado et al., 2014b; Romalde et al., oysters O. edulis and mussels M. galloprovincialis 2014; Gonzlez Castillo et al., 2015). Hatchery rom Italy (Carella et al., 2014). A real-time PCR larval mortality o scallops Argopecten purpu- diagnostic assay was designed or N. crassotreae ratus (Rojas et al., 2015) and o razor clams (Carrasco et al., 2013a). Solen marginatus (Prado et al., 2014a) caused by Vibrio splendidus-like strains, o European Protista oysters O. edulis and Manila clams Ruditapes Perkinsosis, bonamiosis, marteiliosis, and philippinarum associated with Vibrio tubiashii mikrocytosis are protistan diseases o bivalve europaensis (Prado et al., 2015) and o Pacic molluscs that captured much aention in the oysters C. gigas caused by Vibrio coralliilitychus reviewed period, but also QPX and other patho- and V. tubiashii (Travers et al., 2014; Richards genic protists were the subject o research. Es- et al., 2015) were reported. The involvement o pecially relevant or ghting bivalve diseas- Vibrio spp. in C. gigas summer mortality events, es, selective breeding programmes continue in naturalised populations (Thieltges et al., 2013; providing promising, even successul results, Wendling and Wegner, 2013) and culture areas such as the cases o the inection o Sydney (Barbosa Solomieu et al., 2015), was addressed. rock oyster Saccostrea cucullata with Marteilia The invasive capacity o Crassostrea gigas in sydneyi (Dove et al., 2013a, b), the inection o Europe was ound to be linked with rapid evo- the European oyster O. edulis with Bonamia lution o resistance to local Vibrio spp. (Wendling ostreae (Lynch et al., 2014), and the inections o and Wegner, 2015). The dynamics and pathoge- the American oyster C. virginica with Perkinsus nicity o Vibrio communities in the haemolymph marinus and Haplosporidium nelsoni (Frank-La- o C. gigas (Wendling et al., 2014) and those o V. wale et al., 2014). Furthermore, identication o splendidus and V. aestuarianus isolates in mussels molecular markers o resistance (Ratos et al., (Romero et al., 2014) were studied. Regarding 2014; Nikapitiya et al., 2014) should contribute diagnosis, real-time PCR assays were designed to improve those programmes. or V. coralliilyticus (Wilson et al., 2013) and V. aestuarianus (McCleary and Henshilwood, Marteiliosis has reappeared in the bivalve 2015). Vibrios’ extracellular products and vir- disease scene due to mass mortalities o cockles ulence actors were also studied, namely in V. Cerastoderma edule caused by the new species tapetis and V. aestuarianus (Madec et al., 2014) Marteilia cochillia (Carrasco et al., 2013b; Villal- 28, Bull. Eur. Ass. Fish Pathol., 36(1) 2016 ba et al., 2014) and the detection o parasites viewed in a special issue o the journal Diseases o the genus Marteilia in Asia and America, of Aquatic Organisms. The scope o mikrocytids namely the new species M. granula inecting the broadened as divergent lines were identied Manila clam R. philippinarum in Japan (Itoh et al., and diverse invertebrate hosts were ound; 2014) and PCR positive cases or M. refringens in particular, the aetiological agents o two in oysters C. gigas and ǯȱ£ in Mexico emerging diseases o oysters C. gigas and crabs (Grijalva-Chon et al., 2015). New studies on Cancer pagurus rom the United Kingdom were the disease dynamics and parasite stages out ound to be two divergent new mikrocytid o mollusc hosts, involving mussels in the case species, Mikrocytos mimicus and Paramikrocytos o France (Boyer et al., 2013; Arzul et al., 2014) canceri, respectively (Hartikainen et al., 2014a). and oysters in Australia (Adlard and Nolan, Additionally, Mikrocytos mackini was ound in 2015), with the copepod Paracartia grani and the Pacic oysters C. gigas o Washington state, at polychaete Nephtys australiensis as putative in- lower prevalence and severity than previously termediate hosts, respectively, have contributed reported rom British Columbia, but it was not to a beer knowledge o marteiliosis. in native oysters Ostrea lurida (Elston et al., 2015). A real-time PCR diagnostic assay was Regarding perkinsosis, a comprehensive designed or M. mackini (Polinski et al., 2015). review o host-pathogen interaction stands out (Soudant et al., 2013); within this context, Regarding the inection o the hard clam Mer- putative virulence genes o Perkinsus marinus cenaria mercenaria with QPX, the genome o this were shown to be modulated by oyster mucous parasite was partially characterised and the (Pales-Espinosa et al., 2014). Experimental chal- inuence o temperature in its gene expression lenge o Manila clams R. philippinarum with was assessed (Garcia-Vedrenne et al., 2013). Perkinsus olseni (Waki and Yoshinaga, 2013) and mass mortality o Manila clams in Italy caused A group o protists rarely mentioned in the by this parasite (Preo et al., 2014) highlighted literature o bivalve diseases, Amoebozoa, its pathogenicity. The population genetics o P. entered the stage with the detection o a para- marinus was elucidated (Thompson et al., 2014a, sitic amoeba in the emale gonads o Crassostrea b), and proteomic approaches were used to gasar rom Brazil (Shnel et al., 2014). analyse both inter- and intra-species parasite variability (Fernndez-Boo et al., 2014, 2015). Pathogen DNA in environmental Various surveys have shown Perkinsus spp. in samples new places and new hosts in Brazil (da Silva Searching or pathogen DNA in environmental et al., 2014; Ferreira et al., 2015; Queiroga et al., samples allowed detecting pathogen prints in 2015), western Mediterranean area (Carrasco remote disease-ree locations as well as dis- et al., 2014; Valencia et al., 2014; Ramilo et al., closing unsuspected pathogen diversity (new 2015) and Australia (Dang et al., 2015). lineages, new branches in phylogenetic trees). That was the case o mikrocytids (Hartikainen As mentioned above, recent research on bona- et al., 2014a), haplosporidians (Hartikainen miosis and mikrocytosis was thoroughly re- et al., 2014b) and perkinsids (Chambouvet et Bull. Eur. Ass. Fish Pathol., 36(1) 2016, 29 al., 2014, 2015). This approach will be useul sia is spreading between clams M. arenaria as a to get knowledge on many aspects o patho- clonal transmissible cell derived rom a single gens aecting bivalves, such as pathogen lie original clam (Meger et al., 2015). cycle, phylogeny, geographic range, sanitary control and management and actors inuenc- Other items ing disease dynamics (reviewed by Bass et al., Unortunately, articles devoted to many other 2015). New inormation on technical procedures items, such as environmental stress, climate was published (Bonilla et al., 2015; Schul and change, health condition surveys, health man- Lance, 2015) agement, bibliometry, and other group o pathogens (algae, metazoans, etc), have not Cancer been covered to avoid overpassing the allo- The aetiology o disseminated neoplasia cated space. (leukaemia-like cancer) in bivalves has been a controversial issue or many years. Disease References transmissibility (inectious character), occa- Adlard RD and Nolan MJ (2015). Elucidating the lie cycle o Marteilia sydneyi, the aetiological sional (questioned) experimental transmission agent o QX disease in the Sydney rock with ultra-ltrated, homogenised tissues rom oyster (Saccostrea glomerata). International aected molluscs, sporadic observation o viral Journal for Parasitology 45, 419-426. particles in neoplastic cells, and high levels Andree KB, Carrasco N, Rodgers CJ, Roque A, o reverse transcriptase activity in aected Gairin I and Furones D (2014). The apparent individuals o various bivalve species have disappearance o oyster herpes virus type 1 (OsHV-1) rom the Ebro Delta, Spain. been arguments to support retroviral aetiology Bulletin of the European Association of Fish (reviewed by Carballal et al., 2015). Two im- Pathologists 34, 195-200. pacting contributions have changed the scope, Arriagada, Meger MJ, Muray A, Sherry a novel retrotransposon, called Steamer, was J, Reinisch C, Street C, Lipkin WI and characterised and its DNA copy number per Gol SP (2014). Activation o transcription genome was ound at enormously high levels and retrotransposition o a novel element Steamer in neoplastic hemocytes o the in neoplastic cells o the sotshell clam Mya mollusk Mya arenaria. Proceedings of the arenaria (Arriagada et al., 2014), which explains National Academy of Sciences 111, 14175- the reverse transcriptase activity in aected 14180. individuals. Additionally, using three types o Arzul I, Chollet B, Boyer S, Bonnet D, Gaillard DNA markers (sequences o the Steamer inte- J, Baldi Y, Robert M, Joly JP, Garcia C and gration sites, SNPs in mitochondrial genes and Bouchoucha M (2014). Contribution to the understanding o the cycle o the protozoan microsatellites in nuclear DNA), the genotype parasite Marteilia refringens. Parasitology o neoplastic cells was ound to be dierent 141, 227-240. rom that o normal cells o the host clams M. Bai C, Wang C, Xia J, Sun H, Zhang S and arenaria, whereas the neoplastic cells o clams Huang J (2015). Emerging and endemic rom dispersed locations in Canada and north types o Ostreid herpesvirus 1 were detected eastern USA all have nearly identical genotypes; in bivalves in China. Journal of Invertebrate these results indicate that disseminated neopla- Pathology 124, 98-106. 30, Bull. Eur. Ass. Fish Pathol., 36(1) 2016

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