Ostreid Herpesvirus 1 μVar Research Articles

Susceptibility of shellfish aquaculture species in the Chesapeake Bay and Maryland coastal bays to ostreid herpesvirus 1 microvariants.

Ostreid herpesvirus 1 (OsHV-1) and its microvariants (µVars) cause economically devastating mass mortalities of oysters and pose a threat to the shellfish aquaculture industry globally. OsHV-1 outbreaks can cause up to 100% mortality in the Pacific oyster Crassostrea gigas. However, OsHV-1 and its variants have a broad host range and can infect at least 7 bivalve species, including bay scallops Argopecten irradians and eastern oysters C. virginica. Determining the susceptibility of economically and ecologically important bivalve species to OsHV-1 is critical for improving biosecurity and disease management to protect the aquaculture industry. Surveys of eastern oysters were conducted in June to August 2021 in the Maryland portion of the Chesapeake Bay to determine the prevalence and viral load of OsHV-1 at 5 aquaculture farms. Using quantitative PCR, OsHV-1 was not detected at any sites. Experiments examined the susceptibility of single stocks of eastern oysters and hard clams Mercenaria mercenaria to the virus and their ability to horizontally transmit it using OsHV-1 µVar SD (San Diego, California) and OsHV-1 µVar FRA (Marennes-Olreon, France). Results showed that OsHV-1 µVars did not cause mortality or symptomatic infection in the single stocks of eastern oysters and hard clams used in these experiments using natural infection pathways. However, the eastern oyster stock, when injected with OsHV-1, did transmit the virus to naïve Pacific oysters. Further experimentation using additional stocks and lines and establishment of surveillance programs along the east and Gulf coasts of the USA are necessary to prepare for the potential spread and impact of OsHV-1 related disease.

Herpesvirus de los ostreidos tipo 1 en la almeja mano de león Nodipecten subnodosus de Baja California Sur, México

This study is the first report of the presence of Ostreid herpes virus 1 (OsHV-1) in the scallop Nodipecten subnodosus from Baja California Sur, Mexico; collected from 2010 to 2012.The retrospective study was conducted to evaluate whether OsHV-1 was implicated in the population decline of 2010. The virus was detected by PCR, and viral particles were quantified by qPCR. The OsHV-1 detection was conducted by PCR amplification of three different regions of the genome used to identify OsHV-1 variants. A phylogenetic analysis indicated the phylogenetic position of N. subnodusus OsHV-1 among OsHV-1 samples previously collected in different countries and bivalves. Phylogeny analyses indicated differences between OsHV-1 of N. subnodosus and OsHV-1 μVar. Similarity analyses showed differences between the OsHV-1 of Crassostrea gigas and the OsHV-1 of N. subnodosus from Mexico, however the sequence was identical to that of OsHV-1 from France 1993. This study contributes to better understand the genetic diversity of OsHV-1 and to identify potential host of the virus or species involved in the viral cycle.

Genomic Diversity of the Ostreid Herpesvirus Type 1 Across Time and Location and Among Host Species.

The mechanisms underlying virus emergence are rarely well understood, making the appearance of outbreaks largely unpredictable. This is particularly true for pathogens with low per-site mutation rates, such as DNA viruses, that do not exhibit a large amount of evolutionary change among genetic sequences sampled at different time points. However, whole-genome sequencing can reveal the accumulation of novel genetic variation between samples, promising to render most, if not all, microbial pathogens measurably evolving and suitable for analytical techniques derived from population genetic theory. Here, we aim to assess the measurability of evolution on epidemiological time scales of the Ostreid herpesvirus 1 (OsHV-1), a double stranded DNA virus of which a new variant, OsHV-1 μVar, emerged in France in 2008, spreading across Europe and causing dramatic economic and ecological damage. We performed phylogenetic analyses of heterochronous (n = 21) OsHV-1 genomes sampled worldwide. Results show sufficient temporal signal in the viral sequences to proceed with phylogenetic molecular clock analyses and they indicate that the genetic diversity seen in these OsHV-1 isolates has arisen within the past three decades. OsHV-1 samples from France and New Zealand did not cluster together suggesting a spatial structuration of the viral populations. The genome-wide study of simple and complex polymorphisms shows that specific genomic regions are deleted in several isolates or accumulate a high number of substitutions. These contrasting and non-random patterns of polymorphism suggest that some genomic regions are affected by strong selective pressures. Interestingly, we also found variant genotypes within all infected individuals. Altogether, these results provide baseline evidence that whole genome sequencing could be used to study population dynamic processes of OsHV-1, and more broadly herpesviruses.

The first detection of a novel OsHV-1 microvariant in San Diego, California, USA

The spread, emergence, and adaptation of pathogens causing marine disease has been problematic to fisheries and aquaculture industries for the last several decades creating the need for strategic management and biosecurity practices. The Pacific oyster (Crassostrea gigas), a highly productive species globally, has been a target of disease and mortality caused by a viral pathogen, the Ostreid herpesvirus 1 (OsHV-1) and its microvariants (OsHV-1 µvars). During routine surveillance to establish health history at a shellfish aquaculture nursery system in San Diego, California, the presence of OsHV-1 in Pacific oyster juveniles was detected. Quantification of OsHV-1 in tissues of oysters revealed OsHV-1 viral loads > 106 copies/mg. We characterized and identified the OsHV-1 variant by sequencing of ORFs 4 (C2/C6) and 43 (IA1/IA2), which demonstrated that this variant is a novel OsHV-1 microvariant: OsHV-1 µvar SD. A pilot transmission study indicates that OsHV-1 µvar SD is infectious with high viral loads ~ 7.57 × 106 copies/mg detected in dead individuals. The detection of OsHV-1 µvar SD in a large port mirrors previous studies conducted in Australia where aquaculture farms and feral populations near port locations may be at a higher risk of OsHV-1 emergence. Further research is needed to understand the impacts of OsHV-1 µvar SD, such as transmission studies focusing on potential vectors and characterization of virulence as compared to other OsHV-1 µvars. To increase biosecurity of the global aquaculture industry, active and passive surveillance may be necessary to reduce spread of pathogens and make appropriate management decisions.

The role of the mussel Mytilus spp. in the transmission of ostreid herpesvirus-1 microVar.

The Pacific oyster Crassostrea gigas contributes significantly to global aquaculture; however, C. gigas culture has been affected by ostreid herpesvirus-1 (OsHV-1) and variants. The dynamics of how the virus maintains itself at culture sites is unclear and the role of carriers, reservoirs or hosts is unknown. Both wild and cultured mussels Mytilus spp. (Mytilus edulis, Mytilus galloprovincialis and hybrids) are commonly found at C. gigas culture sites. The objective of this study was to investigate if Mytilus spp. can harbour the virus and if viral transmission can occur between mussels and oysters. Mytilus spp. living at oyster trestles, 400-500 m higher up the shore from the trestles and up to 26 km at non-culture sites were screened for OsHV-1 and variants by all the World Organization for Animal Health (OIE) recommended diagnostic methods including polymerase chain reaction (PCR), quantitative PCR (qPCR), histology, in situ hybridization and confirmation using direct sequencing. The particular primers that target OsHV-1 and variants, including OsHV-1 microVar (μVar), were used in the PCR and qPCR. OsHV-1 μVar was detected in wild Mytilus spp. at C. gigas culture sites and more significantly the virus was detected in mussels at non-culture sites. Cohabitation of exposed wild mussels and naïve C. gigas resulted in viral transmission after 14 days, under an elevated temperature regime. These results indicate that mussels can harbour OsHV-1 μVar; however, the impact of OsHV-1 μVar on Mytilus spp. requires further investigation.

Complete genome sequence of Ostreid herpesvirus type 1 µVar isolated during mortality events in the Pacific oyster Crassostrea gigas in France and Ireland

Infections with Ostreid herpesvirus 1 (OsHV-1) microvariants in young Pacific oysters are associated with massive mortality events and significant economic losses. Previous studies, focusing on few regions of the genome, have revealed the genomic diversity of these genotypes with respect to the reference type. We used a NGS process to sequence the whole genome of the OsHV-1 µVar in infected individuals, collected during mortality events in France and Ireland. The final genome length of OsHV-1 µVar was approximately 205kbp, shorter than the reference genotype and the overall genome organisation resembled herpes simplex viruses. 94.4% similarity was observed with the OsHV-1 reference genotype. Large indels, including five deletions and three insertions were found to induce the loss and the addition of several ORFs, summed with codon substitutions in 64% of genes shared with the reference type. This diversity raises the question of the exact origin and evolution of OsHV-1 µVar.

Ostreid herpesvirus type 1 genomic diversity in wild populations of Pacific oyster Crassostrea gigas from Italian coasts

Ostreid herpesvirus 1 (OsHV-1) is a significant pathogen affecting the young Pacific oyster Crassostrea gigas, worldwide. A new variant, OsHV-1 μVar, has been associated with recurrent mortality events in Europe since 2008. Epidemiological data collection is key for global risk assessment; however little is known about health status and genotypes present in European wild oyster beds. Most studies to date have involved only cultivated individuals during mortality events, and reported low genotype diversity. With this study, conducted along the Italian coasts, we investigated for the first time the presence of OsHV-1 in European natural oyster beds. Analysis of three genomic regions revealed the presence of at least nine different genotypes, including two variants close to the OsHV-1 reference, known since the early 1990s but with no European record reported since 2010, and highlights relevant genotype diversity in natural environment. Phylogenetic analysis distinguished two distinct clusters and geographical distribution of genotypes, with the exception of a variant very closely related to the μVar, which appeared the single genotype present in all the Adriatic sites. Interestingly, these wild symptom free populations could represent, in Europe, an accessible alternative to the import of OsHV-1-resistant oyster strains from the East Pacific, the native area of C. gigas, avoiding the high-risk of non-native marine species and new pathogen introductions.

Experimental infection of European flat oyster Ostrea edulis with ostreid herpesvirus 1 microvar (OsHV-1 μvar): Mortality, viral load and detection of viral transcripts by in situ hybridization

Ostreid herpesvirus 1 (OsHV-1) infections have been reported in several bivalve species. Mortality of Pacific oyster Crassostrea gigas spat has increased considerably in Europe since 2008 linked to the spread of a variant of OsHV-1 called μvar. In the present study we demonstrated that O. edulis juveniles can be infected by OsHV-1μvar when administered as an intramuscular injection. Mortality in the oysters injected with OsHV-1μvar was first detected 4 days after injection and reached 25% mortality at day 10. Moreover, the high viral load observed and the detection of viral transcripts by in situ hybridization in several tissues of dying oysters suggested that OsHV-1μvar was the cause of mortality in the O. edulis juveniles. This is therefore the first study to provide evidence about the pathogenicity of OsHV-1μvar in a species that does not belong to the Crassostrea genus. Additionally, we present a novel method to detect OsHV-1 transcripts in infected individuals’ using in situ hybridization.

Detection of undescribed ostreid herpesvirus 1 (OsHV-1) specimens from Pacific oyster, Crassostrea gigas

The ostreid herpesvirus 1 (OsHV-1) and variants were implicated in mass mortality affecting the young Pacific cupped oysters, Crassostrea gigas, in European countries and those around the world. From 2008 onwards, oyster mortality had greatly increased on the French coast and was associated with the detection of a new OsHV-1 variant, entitled OsHV-1 μVar. The OsHV-1 μVar is predominant in oysters; however, other OsHV-1 variants have been detected in samples collected during mortality periods or collected out of mortality periods in France, Ireland, Spain, Portugal, Italy, Mexico, United States, South Korea, Australia, and New Zealand. A retrospective study conducted on 1047 OsHV-1 specimens sampled mainly in France between 2009 and 2012, revealed 17 undescribed OsHV-1 variants found in 65 oyster samples. These specimens presented point mutations situated downstream and upstream from the microsatellite area in the C region (ORF 4/5) which were different from the OsHV-1 reference and the OsHV-1 μVar. In the present work, investigation was performed to further characterize these OsHV-1 specimens by sequencing two habitually targeted regions to study genetic polymorphism of the virus: ORF 41/42 and ORF 35–38. An OsHV-1 variant detected in six oyster samples, contained a nucleotide substitution in the C region which impacted the amino acid sequence and might modify the function of the unknown protein encoding by ORF 4. For the ORF 41/42 region, only two specimens presented a synonymous mutation in comparison with the OsHV-1 μVar. All specimens contained the same deletion with the OsHV-1 μVar in ORF 35–38. Then, a phylogenetic analysis based on the C region was performed to investigate the distribution of undescribed specimens among 21 OsHV-1 DNA sequences notified in GenBank and collected from different countries (France, Japan, New Zealand, China, Ireland, and United States) between 1995 and 2012. All analyzed samples and the OsHV-1 μVar were placed in the same group, excepted for a Japan specimen.Our results contribute to improve the description of the genetic diversity of the OsHV-1 and the C region (ORF 4/5) appears to be a better target than ORF 42/42 and 35–38 to distinguish variants between themselves.

Stability of Ostreid herpesvirus-1 (OsHV-1) and assessment of disinfection of seawater and oyster tissues using a bioassay

Microvariant genotypes of Ostried herpesvirus-1 (OsHV-1 μVar) have recently emerged as a cause of epizootic mortality in Pacific oysters (Crassostrea gigas) in many countries. Measures that can reduce the spread of the virus are required to decrease the incidence and distribution of the disease at the local and global scales. Disease management strategies and biosecurity plans require data describing the stability of OsHV-1 in the environment and the methods required for effective disinfection of the virus. A bioassay using intramuscular injection of 10month old oyster spat had a limit of detection of 3.6×103 copies of OsHV-1 genome per oyster. This was 10-fold more sensitive compared to immersion challenge of 5month old spat, even though the oysters were exposed to a greater volume of inoculum over a period of 2h. OsHV-1 remained infectious in seawater for 2days at 20°C and in wet or dry, non-living oyster tissues for at least 7days at 20°C. OsHV-1 was inactivated by: commercial multipurpose disinfectants used according to label directions (Virkon-S, Dupont; quaternary ammonium preparation, Livingston); sodium hydroxide (20g/L 10min), iodine (0.1% 5min) and formalin (10% v/v 30min); and physical measures including heating to 50°C for 5min and exposure to a high dose of ultraviolet light. Ineffective disinfectant treatments were: heating to 42°C for 5min, and alkaline detergent (2000ppm, 10min) (Pyroneg, Johnson Diversey). Sodium hypochlorite (50ppm available chlorine, 15min) inactivated OsHV-1 in relatively clean seawater, but this treatment was not effective after addition of protein, 10% v/v foetal bovine serum. A concentration of 200ppm available chlorine for 15min did not inactivate OsHV-1 in oyster tissue. This study provides information that will assist in modelling the disease risk posed by OsHV-1 and data that are necessary to devise effective biosecurity strategies to control the spread of OsHV-1 in a variety of situations. The potential role of fomites in the spread of the virus, and recurrence of disease in endemically infected areas were highlighted by the persistence of infectivity in non-living oyster tissues. Statement of relevanceData on the duration of infectivity and effective disinfection of OsHV-1 in filed relevant preparations is needed for improved disease control.

Resistance of Black-lip learl oyster, Pinctada margaritifera, to infection by Ostreid herpes virus 1μvar under experimental challenge may be mediated by humoral antiviral activity

Ostreid herpesvirus 1 (OsHV-1) has induced mass mortalities of the larvae and spat of Pacific oysters, Crassostrea gigas, in Europe and, more recently, in Oceania. The production of pearls from the Black-lip pearl oyster, Pinctada margaritifera, represents the second largest source of income to the economies of French Polynesia and many Pacific Island nations that could be severely compromised in the event of a disease outbreak. Coincidentally with the occurrence of OsHV-1 in the southern hemisphere, C. gigas imported from New Zealand and France into French Polynesia tested positive for OsHV-1. Although interspecies viral transmission has been demonstrated, the transmissibility of OsHV-1 to P. margaritifera is unknown. We investigated the susceptibility of juvenile P. margaritifera to OsHV-1 μvar that were injected with tissue homogenates sourced from either naturally infected or healthy C. gigas. The infection challenge lasted 14 days post-injection (dpi) with sampling at 0, 1, 2, 3, 5, 7 and 14 days. Mortality rate, viral prevalence, and cellular immune responses in experimental animals were determined. Tissues were screened by light microscopy and TEM. Pacific oysters were also challenged and used as a positive control to validate the efficiency of OsHV-1 μvar infection. Viral particles and features such as marginated chromatin and highly electron dense nuclei were observed in C. gigas but not in P. margaritifera. Mortality rates and hemocyte immune parameters, including phagocytosis and respiratory burst, were similar between challenged and control P. margaritifera. Herpesvirus-inhibiting activity was demonstrated in the acellular fraction of the hemolymph from P. margaritifera, suggesting that the humoral immunity is critical in the defence against herpesvirus in pearl oysters. Overall, these results suggest that under the conditions of the experimental challenge, P. margaritifera was not sensitive to OsHV-1 μvar and was not an effective host/carrier. The nature and spectrum of activity of the humoral antiviral activity is worthy of further investigation.

Genotype of mucopolysaccharidosis type II severe form and the efficacy of enzyme replacement therapy or hematopoietic stem cell transplantation on cognitive function

The Ostreid herpesvirus 1 (OsHV-1) is associated with the mortality of young oysters Crassostrea gigas in France. The virus could infect different marine bivalve species and OsHV-1 DNA was previously detected in crustacean, marine gastropod, and sediment. Nevertheless, the effect of the temperature on the stability of infectious viral particles in sea water remains unpublished. During a mortality event, dead oysters may release virus in the sea water such as free viral particles, viral particles included in cells or attached to a support like plankton, suspended particulate matters, and microalgae. Oysters placed around the infectious source may filtrate the virus and then be infected by OsHV-1. In the present study, we investigated the OsHV-1 μVar virulence in sea water at 16 °C and 25 °C. Filtered infected tissue homogenates were kept at 16 °C or 25 °C prior to injection challenge of uninfected spats. A real-time reverse transcription PCR targeting the OsHV-1 polymerase gene has been developed to confirm the death cause oysters collected during the experiment. The RNA detection of herpesvirus in their hosts is a sign of viral replication and indicates that the virus is certainly infectious. Results showed that the OsHV-1 μVar remained infectious and induced mortality after 33 h at 25 °C and 54 h at 16 °C in controlled conditions. The OsHV-1 μVar is thus able to persist in sea water and high temperatures seem to reduce its infectivity. However, the OsHV-1 μVar virulence may be modulated by biological, physical and chemical factors which are present in sea water.

Virulence of Ostreid herpesvirus 1 μVar in sea water at 16 °C and 25 °C

Abstract The Ostreid herpesvirus 1 (OsHV-1) is associated with the mortality of young oysters Crassostrea gigas in France. The virus could infect different marine bivalve species and OsHV-1 DNA was previously detected in crustacean, marine gastropod, and sediment. Nevertheless, the effect of the temperature on the stability of infectious viral particles in sea water remains unpublished. During a mortality event, dead oysters may release virus in the sea water such as free viral particles, viral particles included in cells or attached to a support like plankton, suspended particulate matters, and microalgae. Oysters placed around the infectious source may filtrate the virus and then be infected by OsHV-1. In the present study, we investigated the OsHV-1 μVar virulence in sea water at 16 °C and 25 °C. Filtered infected tissue homogenates were kept at 16 °C or 25 °C prior to injection challenge of uninfected spats. A real-time reverse transcription PCR targeting the OsHV-1 polymerase gene has been developed to confirm the death cause oysters collected during the experiment. The RNA detection of herpesvirus in their hosts is a sign of viral replication and indicates that the virus is certainly infectious. Results showed that the OsHV-1 μVar remained infectious and induced mortality after 33 h at 25 °C and 54 h at 16 °C in controlled conditions. The OsHV-1 μVar is thus able to persist in sea water and high temperatures seem to reduce its infectivity. However, the OsHV-1 μVar virulence may be modulated by biological, physical and chemical factors which are present in sea water.

Sequence variation in ostreid herpesvirus 1 microvar isolates detected in dying and asymptomatic Crassostrea angulata adults in the Iberian Peninsula: Insights into viral origin and spread

The Portuguese oyster Crassostrea angulata was the main oyster species produced in Europe until the 1970s when mass mortalities almost led to its disappearance. At that time, mortality events were associated with detection of irido-like viruses and no record of herpes-like virus exists. Nowadays, C. angulata is only present in relatively few locations in the Iberian Peninsula, unlike in Asia where production is widespread. In 2011 and 2013, we observed mortality outbreaks of C. angulata adults collected in the Ria Formosa (Portugal) with a cumulative mortality of 47 and 59%, respectively, a few days after the oysters were transferred to indoor conditions. Ostreid herpesvirus 1 (OsHV-1) DNA was detected by PCR in all dying oysters analyzed and a high viral load was observed by real time PCR. Sequence analysis of ORFs 4, 42, 43 and 88 as well as a non-coding region near ORF4 revealed that the OsHV-1 genotype was the microvar (μvar) variant. No nucleotide sequence variation was observed in the regions analyzed in any of the 18 isolates. However, polymorphism was observed in a non-coding region (NC1/NC2 region) located between ORFs 49 and 50. Two genotypes in the NC1/NC2 region differing by several nucleotides were identified during the 2 mortality outbreaks and each genotype was unique to each mortality event. In addition, a third genotype in the NC1/NC2 region of OsHV-1 μvar was observed in asymptomatic adults of C. angulata collected in Carrera River (Spain). OsHV-1 DNA was not detected by PCR in any of the oysters collected in Sado and Mira estuaries. The results obtained in the present study suggest that the NC1/NC2 region may be a highly polymorphic region and hence of particular interest for molecular epidemiological studies. In light of the results of the present and previous studies on the genetic diversity and detection of OsHV-1 in C. angulata, we hypothesize that the virus was introduced recently in Europe from Asia, possibly with the introduction of C. gigas in the 1970s or soon after.

Development and Assessment of a Sensitive and Cost-Effective Polymerase Chain Reaction to Detect Ostreid Herpesvirus 1 and Variants

Ostreid herpesvirus 1 (OsHV-1) and variants (OsHV-1 var, OsHV-1 µvar, Irish genotype) have had a significant impact on the Pacific oyster, Crassostrea gigas, industry worldwide and on the survival of larvae and juveniles of several other bivalve species in Europe. Diagnostic techniques used to screen for this pathogen include histology, polymerase chain reaction (PCR), and quantitative polymerase chain reaction (qPCR), which allows for quantification of the virus. Techniques used to confirm infection include PCR, in situ hybridization, and transmission electron microscopy, which have variable sensitivity and specificity. In this study, several new primers were designed to amplify the C region in the ORF4 gene of the virus. This region is variable and diagnostic among OsHV-1 and variants. To date, the routinely used C2/C6 primers, which are used to screen for OsHV-1 and variants, amplify a product of 709 bp whereas the new primers described in this study amplify products of 296 bp, 385 bp, and 400 bp. Several C. gigas samples, which had been exposed to herpes virus and variants were screened, and a sample of wild mussels (Mytilus spp.) was used as a negative control. The performance of the new PCR and primers was compared with the performance of the C2/C6 primers and qPCR. The effect of tissue storage and DNA extraction on PCR performance was also examined. The results of this study indicate that the new PCR and primers are more rapid, sensitive, and cost-effective compared with the C2/C6 PCR, and are as sensitive as qPCR. The results also highlight the impact that sample storage, tissue selection, DNA extraction method used, and subsequent storage of DNA have on PCR success or failure.

Identification and characterisation of an ostreid herpesvirus-1 microvariant (OsHV-1 µ-var) in Crassostrea gigas (Pacific oysters) in Australia

Between November 2010 and January 2011, triploid Crassostrea gigas (Pacific oysters) cultivated in the Georges River, New South Wales, experienced >95% mortality. Mortalities also occurred in wild diploid C. gigas in the Georges River and shortly thereafter in the adjacent Parramatta River estuary upstream from Sydney Harbour. Neighbouring Saccostrea glomerata (Sydney rock oysters) did not experience mortalities in either estuary. Surviving oysters were collected to investigate the cause of mortalities. Histologically all oysters displayed significant pathology, and molecular testing revealed a high prevalence of ostreid herpesvirus-1 (OsHV-1). Quantitative PCR indicated that many C. gigas were carrying a high viral load at the time of sampling, while the load in S. glomerata was significantly lower (p < 0.001). Subsequent in situ hybridisation experiments confirmed the presence of a herpesvirus in C. gigas but not S. glomerata tissues, suggesting that S. glomerata is not susceptible to infection with OsHV-1. Naïve sentinel triploid C. gigas placed in the Georges River estuary in January 2011 quickly became infected and experienced nearly 100% mortality within 2 wk of exposure, indicating the persistence of the virus in the environment. Phylogenetic analysis of sequences derived from the C2/C6 region of the virus revealed that the Australian strain of OsHV-1 belongs to the microvariant (µ-var) cluster, which has been associated with severe mortalities in C. gigas in other countries since 2008. Environmental data revealed that the Woolooware Bay outbreaks occurred during a time of considerable environmental disturbance, with increased water temperatures, heavy rainfall, a toxic phytoplankton bloom and the presence of a pathogenic Vibrio sp. all potentially contributing to oyster stress. This is the first confirmed report of OsHV-1 µ-var related C. gigas mortalities in Australia.

Detection of Ostreid Herpesvirus 1 from adult Pacific Oysters Crassostrea gigas Cultured in Korea

The presence of ostreid herpesvirus 1 (OsHV-1) and the percentage of viral DNA detected in Pacific oyster Crassostrea gigas adults were investigated monthly between May and November 2012 at three locations along the southern coast of Korea. Among 210 oysters examined by polymerase chain reaction (PCR) analysis, OsHV-1 DNA was detected in only one oyster collected in August. The low detection rate of OsHV-1 DNA was consistent with the lack of reported OsHV-1-associated disease in C. gigas cultured in Korea. The sequence of the present PCR product amplified with the C2/C6 primer pair was identical to that of OsHV-1 μVar except for one nucleotide, and the sequence amplified with Del36-37F2/Del36-37R showed a 605-bp deletion as in OsHV-1 μVar. Although these sequence data are insufficient to determine genotype, the results suggest that the herpesvirus detected was similar to OsHV-1 μVar. This is the first report on the presence of OsHV-1 in adult Pacific oysters cultured in Korea.

Variants of herpetic virus ostreid herpesvirus-1 (OsHV-1) in young Pacific oyster Crassostrea gigas.

Since the 1990s, high mortality rates of young Pacific oysters Crassostrea gigas have been regularly recorded in France and affect mainly spat (oysters less than 1yearold). Pathogens infecting marine bivalve mollusks have been studied particularly due to their economic and ecologic impacts on the farmed and wild oysters. A herpes virus belonging to the family Malacoherpesviridae and called Ostreid Herpesvirus-1 (OsHV-1) has been detected in infected oysters. Since the summer 2008, the mortality of spat has greatly increased on the French coast (with a mortality rate ranging from 40 to 80%) and may be linked to a variant of the OsHV-1, named OsHV-1 μVar. Few variants of the OsHV-1 and genotypes closed to the OsHV-1 μVar have been described in several areas in the world. However, the OsHV-1 μVar is predominant in analyzed samples and the reference genotype of the OsHV-1 has not been detected since 2008. The causes of the emergence of the OsHV-1 μVar and its rapid spreading in the world are currently unknown.

A previously undescribed ostreid herpes virus 1 (OsHV-1) genotype detected in the pacific oyster, Crassostrea gigas, in Ireland

Significant mortalities of the Pacific oyster, Crassostrea gigas, have been reported worldwide since the 1950s. The impact these re-occurring mortality events have had on the C. gigas industry has highlighted the necessity to determine the factors that may be causing these mortalities. This study investigated the possible role of ostreid herpes virus (OsHV-1) in C. gigas mortalities over 2 successive summers at 2 study areas in Ireland. A single sample of adult C. gigas, which had been experiencing mortalities at one of the sites was screened. Successive cohorts of C. gigas spat obtained from a hatchery outside Ireland was relayed to both sites in 2003 and in 2004. Spat were screened each year prior to relaying. Samples were collected every 2 weeks and mortality counts were recorded and observed at both sites. Polymerase chain reaction (PCR) analysis and subsequent sequencing indicated that a previously undocumented variant genotype of OsHV-1 was present in the single cohort of adult C. gigas and in seed and juveniles at both sites, in both years. Analysis suggests that the Irish OsHV-1 μvar variant genotype is closely related to OsHV-1 μvar, first described in France in 2008.

Investigation of mortality in Pacific oysters associated with Ostreid herpesvirus-1 μVar in the Republic of Ireland in 2009

High levels of mortality in Pacific oysters Crassostrea gigas in the Republic of Ireland were recorded during the summer of 2009. The new variant of Ostreid herpes 1 (OsHV-1μVar) which first emerged in France in 2008 was identified from affected stocks. Retrospective data was collected from 70 oyster farmers through an interviewer-administered questionnaire to investigate the distribution and determinants of the mortality. Based on farmer recall, data were recorded at the batch level for cumulative mortality during 2009, start dates and duration of the mortality event and the age of animals affected. Observable mortalities were recorded in 109 out of 346 batches at 47 sites; 104 of the 109 batches were located in bays where OsHV-1μVar had been detected. The records from bays where OsHV-1μVar had been detected were analysed to characterize the pattern of mortality and potential risk factors. Batch mortality averaged 37% (18–65% quartiles) but showed a bimodal distribution (half the batches had mortality less than 45%). Mortalities started at the end of May and continued until early August, peaking in early July. On average oysters died over a period of 18 days. Mortality varied considerably both between and within bays. Mortality started in recently introduced batches and occurred later in the summer in established oysters, which is consistent with the introduction of an infectious agent. Mortality was significantly lower in adults compared with other age groups, which supports observations from France. Three variables were significantly (P<0.05) associated, in both bivariate screening and a logistic regression, with high batch-level mortality (>40%): oysters (i) introduced as juveniles, (ii) during or since the winter of 2008/9 and (iii) which spent less than 8h out of water (in a tidal cycle) (compared with oysters introduced as adults before the winter of 2008/9 and spending more than 8h out of water). Twenty-one percent of triploid batches experienced “high” (>40%) mortality compared with 10% for diploid batches which was significant (P<0.05) in the initial bivariate screening but not in the final logistic regression model. Future studies should develop improved methods to assess oyster mortality and follow stocks over time to better determine the influence of management and environmental factors on mortality.