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Richard J. Whittington - One of the best experts on this subject based on the ideXlab platform.
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detection of Ostreid Herpesvirus 1 in plankton and seawater samples at an estuary scale
Diseases of Aquatic Organisms, 2020Co-Authors: Ika Paulpont, Navneet K. Dhand, Ana Rubio, Richard J. WhittingtonAbstract:: Ostreid Herpesvirus-1 (OsHV-1) is known to associate with particles in seawater, leading to infection and disease in the Pacific oyster Crassostrea gigas. The estuarine environment is highly complex and changeable, and this needs to be considered when collecting environmental samples for pathogen detection. The aims of this study were to (1) compare different aspects of collecting natural seawater and plankton samples for detection of OsHV-1 DNA and (2) determine whether detection of OsHV-1 DNA in such environmental samples has merit for disease risk prediction. The results of one experiment suggest that sampling on the outgoing tide may improve the detection of OsHV-1 DNA in seawater and plankton tow samples (odds ratio 2.71). This statistical comparison was not possible in 2 other experiments. The method (plankton tow or beta bottle) and depth of collection (range: 250-1250 mm) had no effect on the likelihood of detection of OsHV-1. OsHV-1 DNA was found at low concentrations in plankton tow and seawater samples, and only when outbreaks of mortality associated with OsHV-1 were observed in nearby experimental or farmed populations of C. gigas. This suggests that single point in time environmental samples of seawater or plankton are not sufficient to rule out the presence of OsHV-1 in an estuary. The association of OsHV-1 with particles in seawater needs to be better understood in order to determine whether more selective and sensitive methods can be devised to detect it, before environmental samples can be reliably used in disease risk prediction.
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Different in vivo growth of Ostreid Herpesvirus 1 at 18 °C and 22 °C alters mortality of Pacific oysters (Crassostrea gigas)
Archives of Virology, 2019Co-Authors: Maximilian C De Kantzow, Richard J. Whittington, Paul M. HickAbstract:Seasonally recurrent outbreaks of mass mortality in Pacific oysters ( Crassostrea gigas ) caused by microvariant genotypes of Ostreid Herpesvirus 1 (OsHV-1) occur in Europe, New Zealand and Australia. The incubation period for OsHV-1 under experimental conditions is 48-72 hours and depends on water temperature, as does the mortality. An in vivo growth curve for OsHV-1 was determined by quantifying OsHV-1 DNA at 10 time points between 2 and 72 hours after exposure to OsHV-1. The peak replication rate was the same at 18 °C and 22 °C; however, there was a longer period of amplification leading to a higher peak concentration at 22 °C (2.34 × 10^7 copies/mg at 18 hours) compared to 18 °C (1.38 × 10^5 copies/mg at 12 hours). The peak viral concentration preceded mortality by 72 hours and 20 hours at 18 °C and 22 °C, respectively. Cumulative mortality to day 14 was 45.9% at 22 °C compared to 0.3% at 18 °C. The prevalence of OsHV-1 infection after 14 days at 18 °C was 33.3%. No mortality from OsHV-1 occurred when the water temperature in tanks of oysters challenged at 18 °C was increased to 22 °C for 14 days. The influence of water temperature prior to exposure to OsHV-1 and during the initial virus replication is an important determinant of the outcome of infection in C. gigas .
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different in vivo growth of Ostreid Herpesvirus 1 at 18 c and 22 c alters mortality of pacific oysters crassostrea gigas
Archives of Virology, 2019Co-Authors: Maximilian C De Kantzow, Richard J. Whittington, Paul HickAbstract:: Seasonally recurrent outbreaks of mass mortality in Pacific oysters (Crassostrea gigas) caused by microvariant genotypes of Ostreid Herpesvirus 1 (OsHV-1) occur in Europe, New Zealand and Australia. The incubation period for OsHV-1 under experimental conditions is 48-72 hours and depends on water temperature, as does the mortality. An in vivo growth curve for OsHV-1 was determined by quantifying OsHV-1 DNA at 10 time points between 2 and 72 hours after exposure to OsHV-1. The peak replication rate was the same at 18 °C and 22 °C; however, there was a longer period of amplification leading to a higher peak concentration at 22 °C (2.34 × 107 copies/mg at 18 hours) compared to 18 °C (1.38 × 105 copies/mg at 12 hours). The peak viral concentration preceded mortality by 72 hours and 20 hours at 18 °C and 22 °C, respectively. Cumulative mortality to day 14 was 45.9% at 22 °C compared to 0.3% at 18 °C. The prevalence of OsHV-1 infection after 14 days at 18 °C was 33.3%. No mortality from OsHV-1 occurred when the water temperature in tanks of oysters challenged at 18 °C was increased to 22 °C for 14 days. The influence of water temperature prior to exposure to OsHV-1 and during the initial virus replication is an important determinant of the outcome of infection in C. gigas.
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prior exposure to Ostreid Herpesvirus 1 oshv 1 at 18 c is associated with improved survival of juvenile pacific oysters crassostrea gigas following challenge at 22 c
Aquaculture, 2019Co-Authors: Maximilian C De Kantzow, Richard J. Whittington, Paul HickAbstract:Abstract The high mortality and economic loss in farmed Pacific oysters (Crassostrea gigas) caused by Ostreid Herpesvirus 1 (OsHV-1) has spurred research into strategies to mitigate the impact of the disease. Water temperature strongly influences the outcome of exposure with 14 °C not being permissive for an epidemic, infection occurring with limited mortality at 18 °C if there is a high viral dose, and full disease expression occurring at 22 °C. Observations in the field suggest improved survival to subsequent exposures in cohorts of oysters that have survived an OsHV-1 outbreak, but this assessment is complicated by the increasing age of the oysters, different viral exposures, different environmental conditions and the removal of naturally susceptible individuals. A laboratory infection model was used to evaluate the effect on survival of two temporally-spaced challenges with OsHV-1 under different water temperature regimes. Triploid Pacific oyster spat were recruited from commercial farm stocks at 6 months of age and exposed to OsHV-1 by intramuscular injection at 18 °C or 22 °C and then re-exposed at either 18 °C or 22 °C. Mortality did not occur in oysters exposed at 18 °C or when the temperature was increased to 22 °C at 14 days post challenge. Oysters challenged with OsHV-1 at 22 °C were protected if pre-exposed to OsHV-1 at 18 °C (Hazard ratio: 0.22, 95% CI: 0.06–0.80). The present study suggests that non-lethal exposure to OsHV-1 can reduce the mortality on subsequent exposure to the virus, but further experiments are required to investigate the duration of the protective response and its effect in different environments. Such protection may help describe the dynamics of seasonally recurrent disease epidemics and provide a novel approach to disease management.
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effect of emersion on the mortality of pacific oysters crassostrea gigas infected with Ostreid Herpesvirus 1 oshv 1
Aquaculture, 2019Co-Authors: Olivia Evans, Navneet K. Dhand, Richard J. Whittington, Erandi Pathirana, Paul HickAbstract:Abstract Microvariant genotypes of Ostreid Herpesvirus-1 (OsHV-1) have been responsible for mass mortalities in farmed Pacific oyster (Crassostrea gigas) populations in Europe, New Zealand and Australia since its first detection in France in 2008. Previous studies conducted in the Georges River estuary, New South Wales, Australia demonstrated a significant protective effect of increased emersion time for adult oysters, when oysters were grown 300 mm above the standard intertidal growing height used by local farmers. The aim of this study was to investigate the protective effect of tidal emersion on infection by OsHV-1 in a controlled laboratory environment. Adult C. gigas (24 months old 70–90 mm) and C. gigas spat (5 months old, 20–40 mm) were infected with OsHV-1 by intramuscular injection and housed with either twice daily emersion or constant immersion. Oysters were monitored daily for gaping of the valves, as an indicator of clinical disease. Contrary to observations in prior field studies, adult oysters subjected to emersion in this study had significantly higher mortalities (67.2%) compared to the constantly immersed adults (11.3%). No significant difference was observed between treatments for the spat. Real-time quantitative PCR confirmed that all mortality was associated with a high concentration of OsHV-1 DNA. Constant immersion appeared to have a protective effect on infected adult oysters, as many gaping oysters recovered. These results suggest that the beneficial effect of high growing height on adult oysters in the field is due to avoidance of infection with OsHV-1, rather than an effect on oyster physiology. Furthermore, it suggests that if infected oysters can be immersed in such a way that predators and secondary infections are avoided, then many may survive the infection.
Tristan Renault - One of the best experts on this subject based on the ideXlab platform.
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Autophagy in Pacific oyster, Crassostrea gigas
2020Co-Authors: Pierrick Moreau, Benjamin Morga, Delphine Tourbiez, Kevin Moreau, Marie-agnès Travers, David C. Rubinsztein, Tristan RenaultAbstract:Mass mortality outbreaks of Pacific oysters, Crassostrea gigas, are reported in different areas around the world affecting seriously the shellfish aquaculture sector. Two pathogens are associated with these mortality outbreaks, Ostreid Herpesvirus 1 (OsHV-1) and Vibrio aestuarianus. In this contexte, a better knowledge is needed in terms of oyster immunity. For this purpose, as an important degradation pathway autophagy was investigated in the Pacific oysters. An in silico research of genes involved in autophagy using the C. gigas genome allowed identification of homologs of ATG1 and ATG8/LC3(Microtubule-associated protein 1A/1B-light chain 3). Gene expression and protein detection were analysed using real-time PCR and western blot, respectively. ATG1 and ATG8 gene expression was upregulated during an experimental viral infection. Western blot analysis showed an increase of LC3 protein during the infection, suggesting an activation of autophagy. Ammonium chloride treatment was associated with increased oyster mortality whereas less mortality was reported after carbamazepine treatment in experimentally infected animals. Results suggest a protective role of autophagy against OsHV-1 and V. aestuarianus infection. This study contributes to better understand the innate immune system of Pacific oysters.
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First detection of Ostreid Herpesvirus 1 in wild Crassostrea gigas in Argentina
Journal of Invertebrate Pathology, 2019Co-Authors: Elena S. Barbieri, Nuria Vázquez, Carla Fiorito, Antonela Martelli, Andrés Wigdorovitz, Evangelina Schwindt, Cintia Débora Medina, Tristan Renault, Benjamin Morga, Viviana ParrenoAbstract:Abstract Ostreid Herpesvirus 1 (OsHV-1) is a DNA virus of the genus Ostreavirus (Malacoherpesviridae family, Herpesvirales order). Worldwide, OsHV-1 and its microvariants have been associated with increased mortality of Pacific oysters, Crassostrea gigas. Adult asymptomatic oysters also have shown a high prevalence of viral infection. As a consequence, surveillance is needed to better describe OsHV-1 diversity, pathogenicity, clinical signs, and geographical distribution. We examined Crassostrea gigas sampled in October 2017 from the inner zone of the Bahia Blanca Estuary, Argentina, and found that 8 of 30 specimens (26.7%) presented macroscopic lesions in mantle tissues. Histological analysis revealed abnormal presentation of mantle epithelial cells and connective tissues. Conventional and real-time PCR conducted on the oyster samples revealed 70% to be positive for presence of OsHV-1 DNA. The nucleotide sequence of the amplicon obtained from one sample using the primer pair IA1/IA2 (targeting ORF 42/43) was 99% identical to OsHV-1 reference as well as µVar strains B and A (KY271630, KY242785.1), sequenced from France and Ireland. This finding represents the first detection of OsHV-1 DNA in a wild population of C. gigas in Argentina in association with gross mantle lesions.
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insights on the association between somatic aneuploidy and Ostreid Herpesvirus 1 detection in the oysters crassostrea gigas c angulata and their f1 hybrids
Aquaculture Research, 2016Co-Authors: Tristan Renault, Jose I Navas, Frederico M Batista, Pierre Boudry, Monserrat Lopezsanmartin, Francisco Ruano, Vera G Fonseca, Alexandra LeitaoAbstract:Cytogenetic abnormalities associated with viral infections, including from viruses of the Herpesvirales order, have been reported in vertebrate species. Ostreid Herpesvirus 1 (OsHV-1) has been detected worldwide during mortality outbreaks of the Pacific oyster Crassostrea gigas. On the other hand, a high proportion of aneuploid cells in somatic tissues have been observed in C. gigas. In this study, we analysed the putative association between aneuploidy levels and the detection of OsHV-1 in gills of C. gigas, the Portuguese oyster C. angulata and their F1 hybrids cultured in Ria Formosa (Portugal). OsHV-1 was detected by PCR in 5.4% of the total of oysters analysed (n = 111) namely in 11.1%, 8.0% and 1.7% of C. gigas, C. angulata and F1 hybrid respectively. Sequencing analysis of a viral fragment amplified with the C2/C6 primer pair revealed a high similarity with the OsHV-1 reference type. Moreover, in situ hybridization confirmed the presence of OsHV-1 in gill tissue. Oysters where OsHV-1 was detected had a significantly higher mean percentage of aneuploid cells (25%) than the ones where the virus was not detected (18%). However, the overall low percentage of positive samples contrasted with the high mean percentage of aneuploidy observed, with 50% of the oysters analysed showing a percentage of aneuploid cells between 20% and 30%. We hypothesize that somatic aneuploidy may adversely affect oysters making them more prone to OsHV-1 infection, but the virus is unlikely to be the cause of somatic aneuploidy.
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in situ localization and tissue distribution of Ostreid Herpesvirus 1 proteins in infected pacific oyster crassostrea gigas
Journal of Invertebrate Pathology, 2016Co-Authors: Claire Martenot, Nicole Faury, Amelie Segarra, Laury Baillon, Maryline Houssin, Tristan RenaultAbstract:Abstract Immunohistochemistry (IHC) assays were conducted on paraffin sections from experimentally infected spat and unchallenged spat produced in hatchery to determine the tissue distribution of three viral proteins within the Pacific oyster, Crassostrea gigas. Polyclonal antibodies were produced from recombinant proteins corresponding to two putative membrane proteins and one putative apoptosis inhibitor encoded by ORF 25, 72, and 87, respectively. Results were then compared to those obtained by in situ hybridization performed on the same individuals, and showed a substantial agreement according to Landis and Koch numeric scale. Positive signals were mainly observed in connective tissue of gills, mantle, adductor muscle, heart, digestive gland, labial palps, and gonads of infected spat. Positive signals were also reported in digestive epithelia. However, few positive signals were also observed in healthy appearing oysters (unchallenged spat) and could be due to virus persistence after a primary infection. Cellular localization of staining seemed to be linked to the function of the viral protein targeted. A nucleus staining was preferentially observed with antibodies targeting the putative apoptosis inhibitor protein whereas a cytoplasmic localization was obtained using antibodies recognizing putative membrane proteins. The detection of viral proteins was often associated with histopathological changes previously reported during OsHV-1 infection by histology and transmission electron microscopy. Within the 6 h after viral suspension injection, positive signals were almost at the maximal level with the three antibodies and all studied organs appeared infected at 28 h post viral injection. Connective tissue appeared to be a privileged site for OsHV-1 replication even if positive signals were observed in the epithelium cells of different organs which may be interpreted as a hypothetical portal of entry or release for the virus. IHC constitutes a suited method for analyzing the early infection stages of OsHV-1 infection and a useful tool to investigate interactions between OsHV-1 and its host at a protein level.
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detection and distribution of Ostreid Herpesvirus 1 in experimentally infected pacific oyster spat
Journal of Invertebrate Pathology, 2016Co-Authors: Amelie Segarra, Nicole Faury, Laury Baillon, Delphine Tourbiez, Tristan RenaultAbstract:High mortality rates are reported in spat and larvae of Pacific oyster Crassostrea gigas and associated with Ostreid Herpesvirus 1 (OsHV-1) detection in France. Although the viral infection has been experimentally reproduced in oyster larvae and spat, little knowledge is currently available concerning the viral entry and its distribution in organs and tissues. This study compares OsHV-1 DNA and RNA detection and localization in experimentally infected oysters using two virus doses: a low dose that did not induce any mortality and a high dose inducing high mortality. Real time PCR demonstrated significant differences in terms of viral DNA amounts between the two virus doses. RNA transcripts were detected in oysters receiving the highest dose of viral suspension whereas no transcript was observed in oysters injected with the low dose. This study also allowed observing kinetics of viral DNA and RNA detection in different tissues of oyster spat. Finally, viral detection was significantly different in function of tissues (p 0.005), time (p 0.005) with an interaction between tissues and time (p 0.005) for each probe.
Paul Hick - One of the best experts on this subject based on the ideXlab platform.
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different in vivo growth of Ostreid Herpesvirus 1 at 18 c and 22 c alters mortality of pacific oysters crassostrea gigas
Archives of Virology, 2019Co-Authors: Maximilian C De Kantzow, Richard J. Whittington, Paul HickAbstract:: Seasonally recurrent outbreaks of mass mortality in Pacific oysters (Crassostrea gigas) caused by microvariant genotypes of Ostreid Herpesvirus 1 (OsHV-1) occur in Europe, New Zealand and Australia. The incubation period for OsHV-1 under experimental conditions is 48-72 hours and depends on water temperature, as does the mortality. An in vivo growth curve for OsHV-1 was determined by quantifying OsHV-1 DNA at 10 time points between 2 and 72 hours after exposure to OsHV-1. The peak replication rate was the same at 18 °C and 22 °C; however, there was a longer period of amplification leading to a higher peak concentration at 22 °C (2.34 × 107 copies/mg at 18 hours) compared to 18 °C (1.38 × 105 copies/mg at 12 hours). The peak viral concentration preceded mortality by 72 hours and 20 hours at 18 °C and 22 °C, respectively. Cumulative mortality to day 14 was 45.9% at 22 °C compared to 0.3% at 18 °C. The prevalence of OsHV-1 infection after 14 days at 18 °C was 33.3%. No mortality from OsHV-1 occurred when the water temperature in tanks of oysters challenged at 18 °C was increased to 22 °C for 14 days. The influence of water temperature prior to exposure to OsHV-1 and during the initial virus replication is an important determinant of the outcome of infection in C. gigas.
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prior exposure to Ostreid Herpesvirus 1 oshv 1 at 18 c is associated with improved survival of juvenile pacific oysters crassostrea gigas following challenge at 22 c
Aquaculture, 2019Co-Authors: Maximilian C De Kantzow, Richard J. Whittington, Paul HickAbstract:Abstract The high mortality and economic loss in farmed Pacific oysters (Crassostrea gigas) caused by Ostreid Herpesvirus 1 (OsHV-1) has spurred research into strategies to mitigate the impact of the disease. Water temperature strongly influences the outcome of exposure with 14 °C not being permissive for an epidemic, infection occurring with limited mortality at 18 °C if there is a high viral dose, and full disease expression occurring at 22 °C. Observations in the field suggest improved survival to subsequent exposures in cohorts of oysters that have survived an OsHV-1 outbreak, but this assessment is complicated by the increasing age of the oysters, different viral exposures, different environmental conditions and the removal of naturally susceptible individuals. A laboratory infection model was used to evaluate the effect on survival of two temporally-spaced challenges with OsHV-1 under different water temperature regimes. Triploid Pacific oyster spat were recruited from commercial farm stocks at 6 months of age and exposed to OsHV-1 by intramuscular injection at 18 °C or 22 °C and then re-exposed at either 18 °C or 22 °C. Mortality did not occur in oysters exposed at 18 °C or when the temperature was increased to 22 °C at 14 days post challenge. Oysters challenged with OsHV-1 at 22 °C were protected if pre-exposed to OsHV-1 at 18 °C (Hazard ratio: 0.22, 95% CI: 0.06–0.80). The present study suggests that non-lethal exposure to OsHV-1 can reduce the mortality on subsequent exposure to the virus, but further experiments are required to investigate the duration of the protective response and its effect in different environments. Such protection may help describe the dynamics of seasonally recurrent disease epidemics and provide a novel approach to disease management.
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effect of emersion on the mortality of pacific oysters crassostrea gigas infected with Ostreid Herpesvirus 1 oshv 1
Aquaculture, 2019Co-Authors: Olivia Evans, Navneet K. Dhand, Richard J. Whittington, Erandi Pathirana, Paul HickAbstract:Abstract Microvariant genotypes of Ostreid Herpesvirus-1 (OsHV-1) have been responsible for mass mortalities in farmed Pacific oyster (Crassostrea gigas) populations in Europe, New Zealand and Australia since its first detection in France in 2008. Previous studies conducted in the Georges River estuary, New South Wales, Australia demonstrated a significant protective effect of increased emersion time for adult oysters, when oysters were grown 300 mm above the standard intertidal growing height used by local farmers. The aim of this study was to investigate the protective effect of tidal emersion on infection by OsHV-1 in a controlled laboratory environment. Adult C. gigas (24 months old 70–90 mm) and C. gigas spat (5 months old, 20–40 mm) were infected with OsHV-1 by intramuscular injection and housed with either twice daily emersion or constant immersion. Oysters were monitored daily for gaping of the valves, as an indicator of clinical disease. Contrary to observations in prior field studies, adult oysters subjected to emersion in this study had significantly higher mortalities (67.2%) compared to the constantly immersed adults (11.3%). No significant difference was observed between treatments for the spat. Real-time quantitative PCR confirmed that all mortality was associated with a high concentration of OsHV-1 DNA. Constant immersion appeared to have a protective effect on infected adult oysters, as many gaping oysters recovered. These results suggest that the beneficial effect of high growing height on adult oysters in the field is due to avoidance of infection with OsHV-1, rather than an effect on oyster physiology. Furthermore, it suggests that if infected oysters can be immersed in such a way that predators and secondary infections are avoided, then many may survive the infection.
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Counting the dead to determine the source and transmission of the marine Herpesvirus OsHV-1 in Crassostrea gigas
Veterinary Research, 2018Co-Authors: Richard J. Whittington, Olivia Evans, Paul Hick, Ika Paul-pont, Navneet K. DhandAbstract:Marine Herpesviruses are responsible for epizootics in economically, ecologically and culturally significant taxa. The recent emergence of microvariants of Ostreid Herpesvirus 1 (OsHV-1) in Pacific oysters Crassostrea gigas has resulted in socioeconomic losses in Europe, New Zealand and Australia however, there is no information on their origin or mode of transmission. These factors need to be understood because they influence the way the disease may be prevented and controlled. Mortality data obtained from experimental populations of C. gigas during natural epizootics of OsHV-1 disease in Australia were analysed qualitatively. In addition we compared actual mortality data with those from a Reed–Frost model of direct transmission and analysed incubation periods using Sartwell’s method to test for the type of epizootic, point source or propagating. We concluded that outbreaks were initiated from an unknown environmental source which is unlikely to be farmed oysters in the same estuary. While direct oyster-to-oyster transmission may occur in larger oysters if they are in close proximity (
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both age and size influence susceptibility of pacific oysters crassostrea gigas to disease caused by Ostreid Herpesvirus 1 oshv 1 in replicated field and laboratory experiments
Aquaculture, 2018Co-Authors: Paul Hick, Olivia Evans, Navneet K. Dhand, Ana Rubio, Richard J. WhittingtonAbstract:Abstract New farm management practices are required to reduce the impact of disease caused by Ostreid Herpesvirus-1 (OsHV-1) on commercial production of Crassostrea gigas. Both the age and size of oysters are thought to be correlated with survival after exposure to OsHV-1. It is important to distinguish between the protective effects of these factors because the phenotypic plasticity of C. gigas enables a wide range of oyster sizes at any given age. The purpose of this study was to: (1) determine if the age and size of C. gigas have independent effects on susceptibility to disease caused by OsHV-1; (2) determine whether deliberate manipulation of the size of oyster spat (defined as oysters ≤ 12 months old) or adults (oysters > 12 months old) would alter their susceptibility to disease; and (3) assess whether size manipulation could be used as a feasible management strategy to protect C. gigas from mortality due to OsHV-1. Two experiments were conducted, with replicated field and laboratory challenges. In Experiment 1, a protective effect of larger size, independent of age (spat v. adults), was shown for oysters that had grown to be larger under standard commercial farming conditions. After adjusting for clustering due to site and basket, the hazard of death in small oysters was 1.9 times that of large spat and adult oysters. In a second experiment, the size of oysters was deliberately manipulated by restricting the growth of a half of the oysters (independent of starting size); immersion time was limited using a higher longline height compared to oysters held at the optimal intertidal growing height for 6 months, in a region free of OsHV-1. After controlling for the effects of variation in exposure due to location in the field, size group did not impact mortality for either age group but the hazard of death for oyster spat (8 months old) was 5.5 times that of adult oysters (17 months old). Laboratory trials for these prospectively differentiated oysters indicated an interaction between age and size whereby smaller spat (growth restricted) were relatively protected (hazard ratio 0.6) compared to those grown to a larger size. For adults the hazard of death was higher (HR = 2.3) for smaller oysters. Further investigation is required before size manipulation of oysters can be effectively utilised as a management strategy to protect C. gigas from OsHV-1 associated mortality. A thorough understanding of the physiological and metabolic condition of oysters, produced under different grow-out conditions when challenged with OsHV-1 is required.
Chongming Wang - One of the best experts on this subject based on the ideXlab platform.
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influence of temperature on the pathogenicity of Ostreid Herpesvirus 1 in ark clam scapharca broughtonii
Journal of Invertebrate Pathology, 2020Co-Authors: Hao Chen, Bowen Huang, Chongming WangAbstract:Abstract OsHV-1 is an epidemic pathogen of molluscs, and temperature has been recognized as a decisive environmental factor in its pathogenicity. In recent years, ark clam, Scapharca broughtonii, emerged as a host for OsHV-1. In the north of China, massive summer mortalities of ark clams infected with OsHV-1 have been continuously reported since 2012. However, the interaction between temperature and the pathogenicity of OsHV-1 was unknown in ark clams. In this study, the effect of temperature (10 °C to 18 °C stepped by 2 °C) on the occurrence of OsHV-1 disease in ark clams was analyzed. OsHV-1 infection led to gill erosion but not below the critical low temperature (between 12 °C and 14 °C). However, OsHV-1 persisted for more than 2 weeks at 12 °C post inoculation and replication was reactivated when the temperature was elevated to 18 °C. No significant reduction of OsHV-1 DNA load was found when the temperature descended to 12 °C from 18 °C, while the gill erosion remained unchanged. Ark clams failed to show the capability of effective clearance of OsHV-1 below the critical low temperature. Our results demonstrated that the pathogenicity of OsHV-1 was influenced significantly by temperature. Moreover, high temperature favored infection, which could provide more information to understand summer mortality of ark clams.
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long range pcr and high throughput sequencing of Ostreid Herpesvirus 1 indicate high genetic diversity and complex evolution process
Virology, 2019Co-Authors: Benjamin Morga, Umberto Rosani, Chen Li, Chongming WangAbstract:Abstract Ostreid Herpesvirus 1 (OsHV-1) is an important pathogen associated with mass mortalities of cultivated marine mollusks worldwide. Since no cell line allows OsHV-1 replication in vitro, it is difficult to isolate enough high-purity viral DNA for High-Throughput Sequencing (HTS). We developed an efficient approach for the enrichment of OsHV-1 DNA for HTS with long-range PCR. Twenty-three primer pairs were designed to cover 99.3% of the reference genome, and their performances were examined on ten OsHV-1 infected samples. Amplicon mixtures from six successfully amplified samples were sequenced with Illumina platform, and one of them (ZK0118) was also sequenced with the PacBio platform. PacBio reads were assembled into 2 scaffolds compared to 9−68 scaffolds assembled from the Illumina reads. Genomic comparison confirmed high genetic diversity among OsHV-1 variants. Phylogenetic analysis revealed that OsHV-1 evolution was mainly impacted by its host species rather than spatial segregation.
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dual transcriptomic analysis of Ostreid Herpesvirus 1 infected scapharca broughtonii with an emphasis on viral anti apoptosis activities and host oxidative bursts
Fish & Shellfish Immunology, 2018Co-Authors: Umberto Rosani, Chen Li, Qingchen Wang, Guiyang Li, Chongming WangAbstract:Abstract The ark shell, Scapharca (Anadara) broughtonii, is an economically important marine shellfish species in Northwestern Pacific. Mass mortalities of ark shell adults related to Ostreid Herpesvirus-1 (OsHV-1) infection have occurred frequently since 2012. However, due to the lack of transcriptomic resource of ark shells, the molecular mechanisms underpinning the virus-host interaction remains largely undetermined. In the present study, we resolved the dual transcriptome changes of OsHV-1 infected ark shell with Illumina sequencing. A total of 44 M sequence reads were generated, of which 67,119 reads were mapped to the OsHV-1 genome. De novo assembly of host reads resulted in 276,997 unigenes. 74,529 (26.90%), 47,653 (17.20%) and 19, 611 (7.07%) unigenes were annotated into GO, KOG and KEGG database, respectively. According to RSEM expression values, we identified 2998 differentially expressed genes (DEGs) between control and challenged groups, which included 2065 up-regulated unigenes and 933 down-regulated unigenes. Further analysis of functional pathways indicated that OsHV-1 could inhibit host cell apoptosis mainly by the up-regulation of inhibitor of apoptosis protein (IAP), and thus facilitating its successful replication. While host hemoglobins could induce oxidative burst by suppressing its peroxidase activity, and thus defense against OsHV-1 infection. Although we reported a narrow expression of the OsHV-1 genome compared to Crassostrea gigas infection, we highlighted several common viral genes highly expressed in the two hosts, suggesting an important functional role. This study offers insights into the pathogenesis mechanisms of OsHV-1 infection in bivalve mollusks of the Arcidae family.
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Ostreid Herpesvirus 1 infects specific hemocytes in ark clam scapharca broughtonii
Viruses, 2018Co-Authors: Chen Li, Chongming WangAbstract:: High levels of Ostreid Herpesvirus 1 (OsHV-1) were detected in hemocytes of OsHV-1 infected mollusks. Mollusk hemocytes are comprised of different cell types with morphological and functional heterogeneity. Granular cells are considered the main immunocompetent hemocytes. This study aimed to ascertain if OsHV-1 infects specific types of hemocytes in ark clams. Types of hemocytes were first characterized through microexamination and flow cytometry. In addition to a large group of red cells, there were three types of recognizable granular cells in ark clams. Type II granular cells were mostly found with OsHV-1 infection by transmission electron microscope (TEM) examination, and represented the hemocyte type that was susceptible to OsHV-1 infection. The subcellular location of OsHV-1 particles in apoptotic type II granular cells was further analyzed. Some OsHV-1 particles were free inside the apoptotic cells, which may contribute to OsHV-1 transmission among cells in the host, some particles were also found enclosed inside apoptotic bodies. Apoptosis is an important part of the host defense system, but might also be hijacked by OsHV-1 as a strategy to escape host immune attack. Following this investigation, a primary culture of type II granular cells with OsHV-1 infection would facilitate the research on the interaction between OsHV-1 and mollusk hosts.
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identification and characterization of Ostreid Herpesvirus 1 associated with massive mortalities of scapharca broughtonii broodstocks in china
Diseases of Aquatic Organisms, 2016Co-Authors: Chongming Wang, Tao Yu, Tianwen Zhang, Qingchen Wang, Jie HuangAbstract:Abstract In the early summer of 2012 and 2013, mass mortalities of blood ark shell (Scapharca [Anadara] broughtonii), broodstocks were reported in several hatcheries on the coast of northern China. Clinical signs including slow response, gaping valves and pale visceral mass were observed in diseased individuals. In response to these reported mortalities, 238 samples were collected from hatcheries at 6 sites. Microscopic changes including lysed connective tissue, dilation of the digestive tubules, eosinophilic inclusion bodies, nuclear chromatin margination and pyknosis were found in affected animals. Transmission electron microscopy (TEM) revealed herpes-like viral particles within the connective tissue of the mantle. Quantative PCR (qPCR) and nested PCR (nPCR) analysis using primers specific for Ostreid Herpesvirus 1 (OsHV-1) indicated significant higher prevalence of OsHV-1 DNA in cases associated with mass mortalities than those without mass mortalities (p = 0.0012 for qPCR, p < 0.0001 for nPCR). qPCR also indicated that samples associated with mass mortalities carried high viral DNA loads, while the loads in apparently healthy samples were significantly lower (t = 3.15, df = 92, p = 0.002). Sequence analysis of the C2/C6 region of nPCR products revealed 5 newly described variants, which were closely related to each other. Phylogenetic analysis of the 5 virus variants and 48 virus variants reported in previous studies identified 2 main phylogenetic groups, and the 5 virus variants identified here were allocated to a separate subclade. To our knowledge, this is the first report of mass mortalities of bivalve broodstocks associated with OsHV-1 infection.
Claire Martenot - One of the best experts on this subject based on the ideXlab platform.
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in situ localization and tissue distribution of Ostreid Herpesvirus 1 proteins in infected pacific oyster crassostrea gigas
Journal of Invertebrate Pathology, 2016Co-Authors: Claire Martenot, Nicole Faury, Amelie Segarra, Laury Baillon, Maryline Houssin, Tristan RenaultAbstract:Abstract Immunohistochemistry (IHC) assays were conducted on paraffin sections from experimentally infected spat and unchallenged spat produced in hatchery to determine the tissue distribution of three viral proteins within the Pacific oyster, Crassostrea gigas. Polyclonal antibodies were produced from recombinant proteins corresponding to two putative membrane proteins and one putative apoptosis inhibitor encoded by ORF 25, 72, and 87, respectively. Results were then compared to those obtained by in situ hybridization performed on the same individuals, and showed a substantial agreement according to Landis and Koch numeric scale. Positive signals were mainly observed in connective tissue of gills, mantle, adductor muscle, heart, digestive gland, labial palps, and gonads of infected spat. Positive signals were also reported in digestive epithelia. However, few positive signals were also observed in healthy appearing oysters (unchallenged spat) and could be due to virus persistence after a primary infection. Cellular localization of staining seemed to be linked to the function of the viral protein targeted. A nucleus staining was preferentially observed with antibodies targeting the putative apoptosis inhibitor protein whereas a cytoplasmic localization was obtained using antibodies recognizing putative membrane proteins. The detection of viral proteins was often associated with histopathological changes previously reported during OsHV-1 infection by histology and transmission electron microscopy. Within the 6 h after viral suspension injection, positive signals were almost at the maximal level with the three antibodies and all studied organs appeared infected at 28 h post viral injection. Connective tissue appeared to be a privileged site for OsHV-1 replication even if positive signals were observed in the epithelium cells of different organs which may be interpreted as a hypothetical portal of entry or release for the virus. IHC constitutes a suited method for analyzing the early infection stages of OsHV-1 infection and a useful tool to investigate interactions between OsHV-1 and its host at a protein level.
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detection of undescribed Ostreid Herpesvirus 1 oshv 1 specimens from pacific oyster crassostrea gigas
Journal of Invertebrate Pathology, 2015Co-Authors: Claire Martenot, Elise Oden, Emmanuelle Travaille, Suzanne Trancart, Ophelie Lethuillier, Sarah Fourour, Maryline HoussinAbstract:Abstract 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.
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virulence of Ostreid Herpesvirus 1 μvar in sea water at 16 c and 25 c
Aquaculture, 2015Co-Authors: Claire Martenot, Elise Oden, Emmanuelle Travaille, Lucie Denechere, Paul Hubert, Laetitia Metayer, Suzanne TrancartAbstract: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.
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genome exploration of six variants of the Ostreid Herpesvirus 1 and characterization of large deletion in oshv 1μvar specimens
Virus Research, 2013Co-Authors: Claire Martenot, Emmanuelle Travaille, Ophelie Lethuillier, Christophe Lelong, Maryline HoussinAbstract:Abstract The genetic polymorphism of the Ostreid Herpesvirus 1 (OsHV-1) has generally been investigated in three areas: ORFs 4/5, ORFs 42/43, and ORFs 35 to 38. The present study, however, focuses on 40 ORFs, representing 30% of the OsHV-1 genome, encoding four categories of putative proteins: 4 ORFs encoding putative inhibitor of apoptosis proteins; 17 ORFs encoding membrane proteins; 10 ORFs encoding secreted proteins; and 9 ORFs encoding RING finger proteins. The potential role of these proteins in major steps of the life cycle of the OsHV-1 motivated their selection. Seven specimens have been selected in accordance with their nucleotide variations in the C region (area located between the end of the ORF4 and the beginning of ORF 5): 3 OsHV-1 μVar specimens, 2 OsHV-1 μVar Δ9, one specimen of OsHV-1 μVar Δ15, and one OsHV-1 specimen (reference control) close to the reference genome to validate PCRs. The OsHV-1 μVar is mainly characterized by a deletion of 12 consecutive nucleotides followed by a deletion of one adenine in a microsatellite area located in the C region. A representation of nucleotide modifications between the different specimens was performed by building evolutionary trees with respect to the category of ORFs. This phylogenetic analysis revealed two groups: the first one corresponded to the reference control and the reference genome AY509253, and the second one included the 6 OsHV-1 variants. These results suggest that the two main groups come from the same common ancestor, and that the divergence between the reference OsHV-1 and its variants occurred quite far back in time. Moreover, consequences of nucleotide variations in the amino acid sequences, especially the change of the N glycoslyation sites, were investigated. Herein is the first report of four important deletions in these OsHV-1 μVar variants: a deletion of 1385 bp in ORF 11; a deletion of 599 bp in ORF 48; a deletion of 3549 bp in ORFs 61 to 64; and a deletion of 712 bp in ORF 114. The size of the deletions differed between OsHV-1 μVar specimens, OsHV-1 μVar Δ9 specimens, and the OsHV-1 μVar Δ15 specimen. These zones seem to correspond to special points of gene rearrangements for producing new proteins. Further investigation necessary proves to link such nucleotide modifications with consequences of protein functions in the OsHV-1 life cycle.
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Variants of herpetic virus Ostreid Herpesvirus-1 (OsHV-1) in young Pacific oyster Crassostrea gigas.
Virologie (Montrouge France), 2013Co-Authors: Claire MartenotAbstract: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 1 year old). 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.
