Ostreid Herpesvirus Research Articles

First investigation of Ostreid herpesvirus-1 and human enteric viruses in a major scallop production area in Brazil.

Bivalve mollusks may be affected by numerous infectious diseases, which cause high mortality rates and economic burdens for producers. Another challenge for bivalve aquaculture is the protection of farms from human contamination, such as sewage and stormwater discharges. Ilha Grande Bay (IGB), located in Rio de Janeiro state, is the largest Brazilian producer of scallops (Nodipecten nodosus). This region has recently suffered a mass mortality of mollusks, and several environmental contaminants have been reported in the area. To contribute to the elucidation of scallop collapse and better characterize the human impacts, this study assessed the circulation of mollusk (Ostreid herpesvirus-1 [OsHV-1]) and human (mastadenovirus [HAdV] and norovirus GII) viral pathogens in waters and animals produced at IGB. Neither water nor animals were positive for OsHV-1. However, of the 7 points analyzed, 5 points showed contamination by HAdV or norovirus. HAdV and norovirus were detected in 5.5 and 6.9% of the analyzed water samples, respectively, in concentrations ranging from 2.39 × 103 to 1 × 105 genome copies/L. One scallop sample was positive for norovirus (4.5%). These results demonstrate human contamination in the region, presenting a risk of consumer contamination, and a non-association between OsHV-1 and the mass mortality described in scallops.

Metavirome Insights into the Diversity and Potential Pathogenic Infection of Chlamys farreri in the Coastal Seas of the Republic of Korea

Chlamys farreri is primarily cultivated in Japan, China, and South Korea. Although mass mortality of scallops has been occurring recently, likely caused by high temperatures or infectious diseases, the underlying cause remains unclear. Little is known regarding the viral diseases affecting them. Therefore, we explored DNA virus diversity in the mid-gut gland of C. farreri and compared it with that of seawater. C. farreri was cultivated at depths below 5 m from the sea surface in the coastal waters of South Korea and sampled from May to August 2018. Different DNA viral communities were observed in both C. farreri and seawater. In C. farreri, prevalent groups included Mimiviridae (7%), Poxviridae (6%), and Phycodnaviridae (5%). Conversely, the dominant groups in seawater were Autographiviridae (20%), Kyanoviridae (12%), and Zobellviridae (10%). We identified C. farreri-specific viral communities and potentially infectious viruses, such as Ostreid herpesvirus 1 and Abalone herpesvirus Victoria/AUS/2009. Furthermore, C. farreri acts as a reservoir for various viruses, which impact microbial community dynamics and disease transmission in marine ecosystems. Understanding these viral communities is crucial to protecting and restoring coastal ecosystems by highlighting their role in the transmission of potential avian- and bivalve-specific viruses.

Environmental Conditions Associated with Four Index Cases of Pacific Oyster Mortality Syndrome (POMS) in Crassostrea gigas in Australia Between 2010 and 2024: Emergence or Introduction of Ostreid herpesvirus-1?

Warm water temperature is a risk factor for recurrent mass mortality in farmed Pacific oysters Crassostrea gigas caused by Ostreid herpesvirus-1, but there is little information on environmental conditions when the disease first appears in a region-the index case. Environmental conditions between four index cases in Australia (2010, 2013, 2016 and 2024) were compared to provide insight into possible origins of the virus. Each index case was preceded by unusually low rainfall and higher rates of temperature change that could increase oyster susceptibility through thermal flux stress. Water temperature alone did not explain the index cases, there being no consistency in sea surface, estuary or air temperatures between them. Tidal cycles and chlorophyll-a levels were unremarkable, harmful algae were present in all index cases and anthropogenic environmental contamination was unlikely. The lack of an interpretable change in the estuarine environment suggests the recent introduction of OsHV-1; however, viral emergence from a local reservoir cannot be excluded. Future events will be difficult to predict. Temperature flux and rainfall are likely important, but they are proxies for a range of undetermined factors and to identify these, it will be necessary to develop comprehensive protocols for data acquisition during future index cases.

Long-read transcriptomics of Ostreid herpesvirus 1 uncovers a conserved expression strategy for the capsid maturation module and pinpoints a mechanism for evasion of the ADAR-based antiviral defence

Abstract Ostreid herpesvirus 1 (OsHV-1), a member of the family Malacoherpesviridae (order Herpesvirales), is a major pathogen of bivalves. However, the molecular details of the malacoherpesvirus infection cycle and its overall similarity to the replication of mammalian herpesviruses (family Orthoherpesviridae) remain obscure. Here, to gain insights into the OsHV-1 biology, we performed long read sequencing of infected blood clams, Anadara broughtonii, which yielded over one million OsHV-1 long reads. This data enabled the annotation of the viral genome with 78 gene units and 274 transcripts, of which 67 were polycistronic mRNAs, 35 ncRNAs and 20 natural antisense transcripts (NATs). Transcriptomics and proteomics data indicate preferential transcription and independent translation of the capsid scaffold protein as an OsHV-1 capsid maturation protease isoform. The conservation of this transcriptional architecture across Herpesvirales likely indicates its functional importance and ancient origin. Moreover, we traced RNA editing events using short read sequencing and supported the presence of inosine nucleotides in native OsHV-1 RNA, consistent with the activity of ADAR1. Our data suggests that, whereas RNA hyper-editing is concentrated in specific regions of the OsHV-1 genome, single nucleotide editing is more dispersed along the OsHV-1 transcripts. In conclusion, we reveal the existence of a conserved pan-Herpesvirales transcriptomic architecture of the capsid maturation module and uncover a transcription-based viral counter defence mechanism, which presumably facilitates the evasion of the host ADAR antiviral system.

Reproductive aging weakens offspring survival and constrains the telomerase response to herpesvirus in Pacific oysters.

Telomere length (TL) is increasingly recognized as a molecular marker that reflects how reproductive aging affects intergenerational transmissions. Here, we investigated the effects of parental age on offspring survival and the regulation of TL by examining the telomere-elongating activity of telomerase in the Pacific oyster. We assessed the classical hallmarks of aging in parents at three age classes (young, middle-aged, and old) and crossbred them using a split-brood design to examine the consequences of the nine maternal-by-paternal age combinations on their offspring. Reproductive aging leads to increased larval mortality and accelerated telomere shortening in spats, rendering them more susceptible to infection by the Ostreid herpesvirus. Viral exposure stimulates telomerase activity, a response that we identified as adaptive, but weakened by parental aging. While telomerase lengthens a spat's telomere, paradoxically, longer individual TL predicts higher mortality in adults. The telomerase-telomere complex appeared as a conservative biomarker for distinguishing survivors and losers upon exposure to polymicrobial diseases.

Localization and Tissue Tropism of Ostreid Herpesvirus 1 in Blood Clam Anadara broughtonii.

OsHV-1 caused detrimental infections in a variety of bivalve species of major importance to aquaculture worldwide. Since 2012, there has been a notable increase in the frequency of mass mortality events of the blood clam associated with OsHV-1 infection. The pathological characteristics, tissue and cellular tropisms of OsHV-1 in A. broughtonii remain unknown. In this study, we sought to investigate the distribution of OsHV-1 in five different organs (mantle, hepatopancreas, gill, foot, and adductor muscle) of A. broughtonii by quantitative PCR, histopathology and in situ hybridization (ISH), to obtain insight into the progression of the viral infection. Our results indicated a continuous increase in viral loads with the progression of OsHV-1 infection, reaching a peak at 48 h or 72 h post-infection according to different tissues. Tissue damage and necrosis, as well as colocalized OsHV-1 ISH signals, were observed primarily in the connective tissues of various organs and gills. Additionally, minor tissue damage accompanied by relatively weak ISH signals was detected in the foot and adductor muscle, which were filled with muscle tissue. The predominant cell types labeled by ISH signals were infiltrated hemocytes, fibroblastic-like cells, and flat cells in the gill filaments. These results collectively illustrated the progressive alterations in pathological confusion and OsHV-1 distribution in A. broughtonii, which represent most of the possible responses of cells and tissues to the virus.

Pathogenic and genomic characterization of Ostreid herpesvirus 1 associated with mass mortalities of blood cockle (Anadara kagoshimensis)

Ostreid herpesvirus 1 (OsHV-1) is an infectious viral disease of bivalve mollusks with a wide geographic distribution across five continents. Since its initial identification in 1991, the disease has been identified as the cause of mortality in more than ten species of bivalve mollusks. The transmission of OsHV-1 is thought to be facilitated by water. In 2019 and 2021, mass mortalities of blood cockle (Anadara kagoshimensis) hatchery broodstock associated with OsHV-1 infection were reported. The presence of OsHV-1 DNA was confirmed in all hatchery samples with levels exceeding 106 copies per milligram of DNA. OsHV-1 was not detected in the wild populations from which the broodstock was collected, indicating that the virus was introduced into the hatchery with seawater from the open ocean. The results of the histopathological examination indicated that tissue lesions and associated hemocyte infiltration were most prevalent in the connective tissues of the digestive tract, mantle, gills and foot of the diseased A. kagoshimensis. In situ hybridization (ISH) signals of OsHV-1 DNA were frequently observed in infiltrated hemocytes, putative fibroblasts, and on occasion, in muscle cells with weak staining. Transmission electron microscopy analysis demonstrated the presence of herpes-like viral particles within the nucleus of infected cells. The genomes of the OsHV-1 variants from cases 2019 and 2021 were successfully resolved through direct high-throughput sequencing of infected tissues. A phylogenetic analysis of the complete genome sequences of 32 OsHV-1 variants revealed that the segregation of the virus was primarily structured by host species and geographic origin, with some exceptions. We propose that the sympatric distribution of different host species and the potential occurrence of cross-species transmission of OsHV-1 are responsible for the inconsistency between the phylogenetic segregation and their host species. This is the first report on the pathogenic and genomic characterization of OsHV-1 infecting A. kagoshimensis. The epidemiological and phylogenetic analyses offer new insights into the evolution of the virus and provide valuable information for the prevention and control of the disease.

Laboratory Replication of Ostreid Herpes Virus (OsHV-1) Using Pacific Oyster Tissue Explants.

Pacific oysters (Crassostrea or Magallana gigas) are one of the most economically important aquaculture species globally. Over the past two decades, ostreid herpesvirus (OsHV-1) has become a major pathogen of cultured Pacific oysters, resulting in widespread mortality with a global distribution. Experimental use of OsHV-1 is challenging for many reasons, including both complexity of host-pathogen dynamics and a lack of functioning model systems. The goal of this study was to improve the tools available for working with OsHV-1 in both whole animals and in tissue explants established from oysters maintained in controlled laboratory conditions. Tissue explants were taken from oysters originating from two different sources that have different levels of mortality in experimental OsHV-1 infections and were exposed to OsHV-1. A whole-animal infection experiment was run concurrently as a comparison. Quantitative PCR and electron microscopy were used to confirm that the explants were capable of replicating OsHV-1. Furthermore, the quantitative PCR results suggest that the source of the oysters was significant in determining the outcome of infection in the explants, supporting the validity of the explant model for OsHV-1 infection. This tissue explant approach for studying OsHV-1 allows for the control of confounding factors in the disease outcome that is not possible in whole-animal experiments, providing a new tool for the study of OsHV-1 in Pacific oysters.

Strong genotype-by-environment interaction across contrasted sites for summer mortality syndrome in the Pacific oyster Crassostrea gigas

Genotype-by-environment (GxE) interaction in aquaculture is usually estimated for continuous traits and based on data from a limited number of 2–3 rearing sites. Here we report the results of a GxE study for resistance to Pacific oyster mortality syndrome (POMS), a multi-factorial disease that severely impacts Pacific oyster production worldwide. The syndrome is largely associated with Ostreid Herpes Virus 1 (OsHV-1). Resistance to OsHV-1 in Crassostrea gigas has been shown to be heritable, meaning that selective breeding is a suitable strategy for reducing mortalities. However, limited information was available about GxE interaction or the possible need to consider it in selective breeding. Survival of two cohorts (C1 and C2), consisting of a total of 104 full-sib families, was evaluated during the summer of 2013 in 7 sites along the French Atlantic, Channel and Mediterranean coasts. Mean survivals in autumn 2013 were 12.6% ± 10.9 and 4.6% ± 6.4 for C1 and C2, respectively. Genetic parameters were computed by MCMC, which is suitable for binary data like survival. Heritability estimates ranged from 0.16 to 0.42 depending on site and cohort, with a mean of 0.24 [0.20; 0.27] when including all data. GxE interactions were estimated by the genetic correlations between pairs of sites. Genetic correlations were high (ρ > 0.80) for C1 between most tidal Atlantic and Channel sites, and intermediate between tidal sites and a Mediterranean lagoon site, while they were lower and more variable for C2 (0.21–0.77). Expected genetic gains were maximal when production site was the same as selection site. They were closed to this expected maximum when the selection site was different from the production site along the Atlantic or Channel coast. Limited GxE interaction along the French Atlantic coast is favorable to wide dissemination of genetically improved oysters along this coast. Limited potential improvement was shown in the Mediterranean site if selection was carried out elsewhere, confirming the specificity of this environment. Consequently, a specific strategy such as dedicated breeding should be used to achieve genetic progress for this site.

An amplicon panel for high-throughput and low-cost genotyping of Pacific oyster.

Maintaining genetic diversity in cultured shellfish can be challenging due to high variance in individual reproductive success, founder effects, and rapid genetic drift, but is important to retain adaptive potential and avoid inbreeding depression. To support broodstock management and selective breeding in cultured Pacific oysters (Crassostrea (Magallana) gigas), we developed an amplicon panel targeting 592 genomic regions and SNP variants with an average of 50 amplicons per chromosome. Target SNPs were selected based on elevated observed heterozygosity or differentiation in Pacific oyster populations in British Columbia, Canada. The use of the panel for parentage applications was evaluated using multiple generations of oysters from a breeding program on Vancouver Island, Canada (n = 181) and families selected for Ostreid herpesvirus-1 resistance from the Molluscan Broodstock Program in Oregon, USA (n = 136). Population characterization was evaluated using wild, naturalized, farmed, or hatchery oysters sampled throughout the Northern Hemisphere (n = 190). Technical replicates showed high genotype concordance (97.5%; n = 68 replicates). Parentage analysis found suspected pedigree and sample handling errors, demonstrating the panel's value for quality control in breeding programs. Suspected null alleles were identified and found to be largely population dependent, suggesting population-specific variation impacting target amplification. Null alleles were identified using existing data without the need for pedigree information, and once they were removed, assignment rates increased to 93.0% and 86.0% of possible assignments in the two breeding program datasets. A pipeline for analyzing the amplicon sequence data from sequencer output, amplitools, is also provided.

Investigations of the involvement of Vibrio species with Ostreid herpesvirus-1 in mass mortality events in the Pacific oyster Crassostrea gigas

Pacific oyster mortality syndrome (POMS) in Pacific oysters Crassostrea gigas is defined by mass mortality with Ostreid herpesvirus I (OsHV-1). In this study the association of Vibrio species with oysters before, during and after POMS events was investigated at population level to clarify their involvement with OsHV-1 in mass mortality events in the Georges and Hawkesbury River estuaries, Australia. In the POMS affected Georges River estuary, three patterns were observed concurrently at different sites: i) Vibrio counts and OsHV-1 DNA concentrations increased in adults and spat as mortalities started then decreased as mortalities stopped ii) bacterial counts increased in spat but not in adults associated with mortalities due to OsHV-1 iii) bacterial counts increased in the absence of mass mortality or high concentrations of OsHV-1 DNA. Although almost half of the 120 bacterial isolates were identified as an unknown Vibrio species belonging to the Splendidus clade with two predominant biotypes (A and H), there was no association between biotype and mortality. Concurrently in the Hawkesbury River estuary, neither mortality nor OsHV-1 were found, Vibrio counts fluctuated significantly and of 84 isolates identified, V. alginolyticus, V. parahaemolyticus and unidentified Vibrio sp. dominated; only five Splendidus clade isolates were identified. However, when POMS emerged in the Hawkesbury River estuary, the Splendidus-clade isolate became the dominant bacterial species. Mortality also occurred there without OsHV-1 and the dominant bacteria then were V. alginolyticus and V. harveyi, but the Splendidus-clade isolate was not detected. None of the bacterial species were specifically associated with POMS, although the Splendidus clade isolate increased in prevalence after POMS emerged in the Hawkesbury River estuary. V. aestuarianus was not detected in this study. While the findings suggest that bacterial proliferation follows infection with OsHV-1 in POMS, this was not a consistent feature. The number and diversity of bacteria within oyster tissues varied over time, between estuaries, between sites within an estuary and were associated with flooding, growing height, the age of oysters, mortality and OsHV-1.

Assessing selection potential for Pacific oyster (Crassostrea gigas) to a North American OsHV-1 μvar: Comparing two experimental assay methods

Ostreid Herpesvirus - 1 is a global threat to Pacific oyster aquaculture as catastrophic farm losses have occurred where OsHV-1 microvariants (μvars) become endemic. Genetic selection has proven effective for increasing survival to OsHV-1 exposure in multiple Pacific oyster breeding programs, but a novel μvar detected in San Diego Bay California has the potential to severely impact commercial aquaculture along the North American Pacific coast as no OsHV-1 μvar selected population is available commercially. Using a Molluscan Broodstock Program population descending from imports originating in the Miyagi prefecture (Japan), we evaluated the potential for genetic selection to improve survival against the San Diego Bay OsHV-1 μvar using a laboratory trial. Two experimental methods were evaluated, a common garden method where all 15 full-sibling families were pooled, and a petri dish method, where each family was confounded within replicate. No striking differences were found between experimental methods, and both experiments displayed high heritability (0.69 and 0.64) of the tested trait, a broad range of estimated breeding values, and a moderate prediction accuracy for the survival trait. A significant genotype-by-environment interaction occurred, highlighting variability in laboratory-based disease challenges, but the two experiments yielded similar results with minor to moderate re-ranking of mean family survival. Overall, the Miyagi population of Pacific oysters curated by the Molluscan Broodstock Program is a promising population to use for artificial selection to increase survival against San Diego Bay OsHV-1 μvar in attempts to obviate farm losses if the virus expands its currently isolated range.

Antiviral protection in the Pacific oyster Crassostrea (Magallana) gigas against OsHV-1 infection using UV-inactivated virus

The increase of the frequency and severity of marine diseases affecting farmed marine mollusks are currently threatening the sustainability of this aquaculture sector, with few available prophylactic or therapeutic solutions. Recent advances have shown that the innate immune system of invertebrates can develop memory mechanisms allowing for efficient protection against pathogens. These properties have been called innate immune memory, immune priming or trained immunity. Previous results demonstrated the possibility to elicit antiviral immune priming to protect Pacific oysters against the ostreid herpes virus 1 (OsHV-1), currently plaguing M. gigas production worldwide. Here, we demonstrate that UV-inactivated OsHV-1 is also a potent elicitor of immune priming. Previous exposure to the inactivated virus was able to efficiently protect oysters against OsHV-1, significantly increasing oyster survival. We demonstrate that this exposure blocked viral replication and was able to induce antiviral gene expression potentially involved in controlling the infection. Finally, we show that this phenomenon can persist for at least 3 months, suggesting the induction of innate immune memory mechanisms. This study unravels new ways to train the Pacific oyster immune system that could represent an opportunity to develop new prophylactic strategies to improve health and to sustain the development of marine mollusk aquaculture.

Temperature and microbe mediated impacts of the San Diego Bay ostreid herpesvirus (OsHV-1) microvariant on juvenile Pacific oysters

Abstract The ostreid herpesvirus (OsHV-1) was recently detected in San Diego Bay for the first time in farmed juvenile Pacific oysters (Crassostrea gigas). Due to the virus’ ability to cause mass mortality (50%–100%), it is important to determine the factors that promote infection as well as the consequences of infection. Here, we assess the role of temperature in controlling OsHV-1 induced mortality. Pacific oysters were exposed to the San Diego Bay microvariant of OsHV-1 at four different temperatures (15°C, 18°C, 21°C, and 24°C). While OsHV-1 was able to replicate in oyster tissues at all temperatures, it did not induce mortality at 15°C, only at the higher temperatures. Additionally, we examined oyster tissue-associated bacterial response to OsHV-1 infection. As shown previously, bacterial richness increased following OsHV-1 exposure and then decreased as the oysters became sick and died. Four bacterial taxa linked to the San Diego Bay microvariant infection, including Arcobacter, Vibrio, Amphritea, and Pseudoalteromonas, were the same as those shown for other microvariant infections in other studies from globally distributed oysters, suggesting a similar spectrum of co-infection irrespective of geography and microvariant type. The significant shift in the bacterial community following exposure suggests a weakening of the host defenses as a result of OsHV-1 infection, which potentially leads to adverse opportunistic bacterial infection.

Relationships between high temperatures and Pacific Oyster disease and mortality in southeast Tasmania, Australia

Warm ocean temperature extremes, including marine heatwaves, have profound impacts on natural marine systems and aquaculture industries across the globe. In Tasmania, Australia, one aquaculture industry that has been significantly impacted by warm temperatures is Pacific Oyster (Magallana gigas, previously named Crassostrea gigas) farming, due to recurring outbreaks of the virus Ostreid herpesvirus 1. Such viral outbreaks are understood to be driven by high seawater temperatures, but the temperature threshold or duration for triggering disease and mortalities remain unclear. This study investigates the relationship between in-situ farm temperatures and oyster disease and mortality on the southeast coast of Tasmania, Australia using daily observations from three oyster growing areas (Pipe Clay Lagoon, Upper Pittwater, and Lower Pittwater) over three seasons. It is found that a 12-day averaged daily mean temperature is an excellent measure of the occurrence of high mortality. Specifically, a 21-day mean of 23.7 °C resulted in a 70% likelihood of high mortality, which is defined here as oyster losses of >15%. On the other hand, for lower levels of disease and mortality, a 12-day average of daily mean temperature gave the strongest relationship. A 12-day mean of 19.7 °C led to 70% probability of some disease and low mortality. The analysis also found in-situ farm temperature generally correlates well with remotely sourced temperature observations, indicating their potential usability for operational management. This study demonstrates a statistical risk analysis framework for the oyster farming industry, helping to improve the understanding of the detrimental impact of high temperatures on Pacific Oysters.

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.

Rapid detection of Japanese eel endothelial cell-infecting virus (JEECV) and Anguillid herpesvirus (AngHV-1) in Japanese eel (Anguilla japonica) using a qLAMP assay

Two DNA viruses, Japanese eel endothelial cell-infecting virus (JEECV) and Anguillid herpesvirus 1 (AngHV-1), cause serious diseases in eels that can negatively affect the economy of eel aquaculture industries in South Korea. Efficient virus detection is crucial for disease management; however, the practical application of current methodologies is challenging, in part owing to extensive time and specialized equipment requirements. The aim of this study was to develop a rapid real-time quantitative loop-mediated isothermal amplification (qLAMP) technique for the detection of JEECV and AngHV-1. Six different primers were designed to target eight different regions of the large T-antigen gene for JEECV detection, and the DNA polymerase catalytic subunit gene for AngHV-1 detection. Both sets of primers worked optimally at 65 °C. The qLAMP assay exhibited a minimum detection limit of 2.39 × 104 copies/μL for JEECV and 2.37 × 104 copies/μL for AngHV-1. Moreover, the primers demonstrated excellent specificity, showing no reaction with two major viruses, namely ostreid herpesvirus-1 microvariant (OsHV-1 μvar) and red sea bream iridovirus (RSIV). In addition, JEECV primers did not react with AngHV-1, and vice versa. Based on the standard curve and coefficient of determination values, both viruses could be quantified using this method. The qLAMP assay was validated with 104 clinical samples, including 72 infected and 32 healthy eels, and the assay findings were compared with previously reported duplex qPCR results that reported high sensitivity and specificity. The qLAMP results concorded well with the previous results. Evaluation of the threshold time (Tt) of the qLAMP results revealed that the samples could be analyzed in less than 15 min. The clinical diagnostic sensitivity (dSe) and diagnostic specificity (dSp) were assessed based on the confusion matrix; dSe and dSp for JEECV were 100% and 94.1%, respectively, and dSe and dSp for AngHV-1 were 96.8% and 77.8%, respectively. The Kappa values, which were used to represent the level of agreement between qLAMP and duplex qPCR results, for JEECV and AngHV-1 were 0.956 and 0.755, respectively, indicating that the qLAMP assay is a promising detection method for those viruses. Thus, the qLAMP assay developed herein could be applied as a rapid and sensitive tool to diagnose diseases caused by JEECV and AngHV-1 in the eel aquaculture industry.

No trade-off between growth and OsHV-1 tolerance in Pacific oysters (Crassostrea gigas) farmed on the West Coast, USA.

An important pathogen of concern for Pacific oyster growers in the USA is ostreid herpesvirus 1 (OsHV-1). Currently, oyster stocks exist that are tolerant to OsHV-1; however, it is uncertain if a trade-off exists between their tolerance to OsHV-1 and their growth in different environments. To investigate any potential trade-offs, Pacific oyster families with varying levels of OsHV-1 tolerance were grown in a bay where OsHV-1 is endemic (Tomales Bay, CA) and in a bay where OsHV-1 is absent (Willapa Bay, WA). In Tomales Bay, we found that oysters from OsHV-1 tolerant families grew faster than oysters from OsHV-1 susceptible families, while in Willapa Bay, no statistically significant difference in growth was found between oyster families with different levels of OsHV-1 tolerance observed in Tomales Bay. These findings indicate that Pacific oysters bred to be tolerant to OsHV-1 would not be expected to have a longer time-to-market regardless of the presence of OsHV-1 in the growing environment.

Inactivation of two oyster pathogens by photocatalysis and monitoring of changes in the microbiota of seawater: A case study on Ostreid herpes virus 1 μVar and Vibrio harveyi.

The pollution of seawater by both biotic (bacteria, viruses) and abiotic contaminants (biocides, pharmaceutical residues) frequently leads to economic losses in aquaculture activities mostly mortality events caused by microbial infection. Advanced Oxidation Processes (AOPs) such as heterogeneous photocatalysis allow the removal of all organic contaminants present in water and therefore could reduce production losses in land-based farms. Oysters in land-based farms such as hatcheries and nurseries suffer from a large number of mortality events, resulting in significant losses. If photocatalysis has been widely studied for the decontamination, its application for disinfection is still overlooked, especially on seawater for viruses. We therefore studied seawater disinfection using the photocatalysis (UV365/TiO2) method in the context of Pacific oyster mortality syndrome (POMS). POMS has been defined as a polymicrobial disease involving an initial viral infection with Ostreid Herpes Virus 1, accompanied by multiple bacterial infections. We investigated the impact of treatment on Vibrio harveyi, a unique opportunistic pathogenic bacterium, and on a complex microbial community reflecting a natural POMS event. Viral inactivation was monitored using experimental infections to determine whether viral particles were still infectious after. Changes in the total bacterial community in seawater were studied by comparing UV365/TiO2 treatment with UV365-irradiated seawater and untreated seawater. In the case of OsHV-1, a 2-h photocatalytic treatment prevents POMS disease and oyster mortality. The same treatment also inactivates 80% of viable Vibrio harveyi culture (c.a. 1.5 log). Since OsHV-1 and Vibrio harveyi are effectively inactivated without long-term destabilization of the total bacterial microbiota in the seawater, photocatalysis appears to be a relevant alternative for disinfecting seawater in land-based oyster beds.

Ostreid herpesvirus 1 latent infection and reactivation in adult Pacific oysters, Crassostrea gigas

Ostreid herpesvirus 1 (OsHV-1) is one of the most economically important pathogens of Pacific oysters. Understanding the pathogenesis of this virus is critical to developing tools to control outbreaks on shellfish farms. OsHV-1 is genetically related to vertebrate herpesviruses, which have a lytic and a latent stage, with the latent stage capable of being reactivated to the lytic stage. Here, OsHV-1 latency in Pacific oysters was investigated in experimentally and naturally infected oysters. Lytic infection in one-year-old oysters injected with the Tomales Bay strain of OsHV-1 was detectable between 1 and 4 days post-injection (dpi) but was not detectable after 5 dpi. The injected oysters shed 1 × 102 to 1 × 104 DNA copies/ml into the water during the 4-day acute phase. Lytic shedding was not detectable in two-year-old oysters injected similarly with the same strain of OsHV-1; however, the OsHV-1 genome was detectable by qPCR in the adductor muscle, gill, mantle, and hemocytes within the first 3 dpi, after which it became undetectable. No OsHV-1 was detectable in the adductor muscle, gill, or mantle from experimentally infected oysters on days 15 and 21 post-injection or from oysters sampled 9 months after surviving an OsHV-1 mortality event; however, OsHV-1 DNA could be detected in hemocytes of both experimentally infected oysters at 21 dpi and naturally infected oysters using nested PCR. In addition, lytic viral gene transcription was detectable in hemocytes of experimentally infected oysters between 1 and 21 dpi and in hemocytes of naturally infected oysters. Furthermore, OsHV-1 reactivation from latency was induced in experimentally infected oysters at 21 dpi and in naturally infected oysters 12 months after an OsHV-1 outbreak.