Genomes of anguillid herpesvirus 1 strains reveal evolutionary disparities and low genetic diversity in the genus cyprinivirus

Anguillid herpesvirus 1 (AngHV-1) is a pathogen of eels and a member of the genus Cyprinivirus in the family Alloherpesviridae. We have compared the biological and genomic features of different AngHV-1 strains, focusing on their growth kinetics in vitro and genetic content, diversity, and recombination. Comparisons based on three core genes conserved among alloherpesviruses revealed that AngHV-1 exhibits a slower rate of change and less positive selection than other cypriniviruses. We propose that this may be linked to major differences in host species and corresponding epidemiological circumstances. Efforts to derive evolutionary rate estimates for cypriniviruses under various theoretical models were ultimately unrewarding. We highlight the potential value of future collaborative efforts towards generating short-term evolutionary rate estimates based on known sequence sampling dates. Finally, we revealed that there is significantly less genetic diversity in core gene sequences within cyprinivirus species clades compared to species in the family Herpesviridae. This suggests that cyprinivirus species may have undergone much more vigorous purifying selection post species clade divergence. We discuss whether this may be linked to biological and anthropogenic factors or to sampling bias, and we propose that the comparison of short-term evolutionary rates between species may provide further insights into these differences

Susceptibility and permissivity of zebrafish (Danio rerio) larvae to cypriniviruses

No abstract available

Infection de l'anguille européenne par l'Anguillid herpesvirus 1: de la recherche fondamentale aux programmes de conservation

The European eel (Anguilla Anguilla) is a culturally and commercially important fish species and along with other members of the genus Anguilla, they are flagship species for aquatic conservation efforts. Anguillid eels have a fascinating and complex life cycle involving different life stages and extended migration through both marine and freshwater environments. The lifecycle of the European eel begins in the Atlantic Ocean. After hatching, leptocephali larvae drift thousands of kilometres across the ocean to reach continental coastal waters where they metamorphose into glass eels. These juvenile eels then migrate upstream towards freshwater habitats where they grow into yellow eels and remain there for several years. Finally, they fully mature into silver eels and migrate back to their place of birth, where they reproduce before dying. Within the last 50 years, the European eel population has dramatically decline and the species is now critically endangered. The reason for this steep decline in population is likely to be multifactorial, with one potential major factor being infectious diseases. In terms of viral diseases, Anguillid herpesvirus 1 (AngHV 1) is the causative agent of a haemorrhagic disease affecting freshwater eels (Anguilla spp.), and is one of the most frequently detected viruses from European eels, especially in fish farms. This herpesvirus is a member of the genus cyprinivirus of the family Alloherpesviridae. As a consequence of this decline, the European Commission has established the “Eel Recovery Plan” within member states with the aim of protecting and restoring the population of European eel. One of the main measures of this plan relies on restocking. This process involves the active transfer of eels to inland areas where eel populations have declined. Although this restocking is widely practiced in Europe, additional studies are still needed to evaluate and improve its effectiveness. The broad objective of the present thesis was to contribute to the conservation of the critically endangered European eel, through focusing on infectious diseases as one of the potential causative factors of its decline. Firstly, we investigated the sanitary status of wild caught glass eels entering estuaries and aimed to develop quarantine stations that would facilitate the implementation of therapeutic and prophylactic measures prior to their reintroduction into the wild. Our results showed that glass eels were negative for AngHV-1 before their reintroduction into Belgian rivers. We also established that under certain conditions, a two-week pre-release quarantine period could be implemented without major impact on important metrics such as eel survival rate, sanitary status, health and morphology. One of the major goals of this PhD project was to gain insights into AngHV-1 pathogenesis using recombinant viral strains combined with in vivo bioluminescent imaging (IVIS). As a prerequisite for the study of AngHV 1 pathogenesis, we sequenced the entire genome of seven strains of AngHV-1 from different geographic origins and compared their biological properties in vitro, which also provided insights into the evolution of this virus. Based on this, we selected 4 strains representative of the viral species, compared their properties in vivo and produced recombinants expressing luciferase and a copepod green fluorescent protein (here after named LucGFP). Characterization of these recombinants revealed that they are appropriate for the study of AngHV-1 with no detectable negative effect associated with the insertion of the transgene. As all strains expressed comparable properties in vivo, the UK LucGFP strain was selected for the next steps of the project. Using the UK LucGFP strain and the IVIS, we investigated major questions related to the pathogenesis of AngHV-1 infection in its natural host: These experiments led to the following observations: (i) Glass eels are not susceptible to AngHV-1 infection through the natural route tested. (ii) Inoculation of elvers led to few positive subjects expressing bioluminescence on their cephalic part, mainly gills and jaws, with no spreading of the infection between subjects according to time. (iii) Inoculation of yellow eels led to infection of all subjects expressing bioluminescence signal associated with strong clinical signs. Morbidity and cumulated mortality reached 100% and 20% respectively. (iv) The gills and the periodontal mucosa represented major portals of entry of the virus into naïve subjects. (v) Two modes of transmission of AngHV-1 were highlighted in this study, i.e. direct transmission through interspecific agonistic interactions (e.g. biting) between fish, and indirect transmission through environmental water. Our results indicate that AngHV 1 transmission via contaminated water requires a high density of subjects and virions, and is thus probably restricted to intensive aquaculture conditions. On the other hand, AngHV-1 infection mediated by biting appears highly efficient and is likely to be facilitated in environments with either high or low host density. We hypothesize that viral transmission mediated by interspecific agonistic interactions may partially explain the high prevalence of AngHV 1 in the wild despite the low infectivity of AngHV-1 in water, the low and declining host population density, the solitary behaviour of the host, and the strong ability of fish skin mucus to neutralize pathogens such as AngHV 1. In conclusion, this project contributed to recommendations for optimization of glass eel restocking practices in Belgium and highlighted major aspects of AngHV 1 pathogenesis. The findings create ample scope for future projects in relation to eel conservation, AngHV 1 evolution, host-pathogen dynamics in aquatic environments as well as setting the groundwork for AngHV 1 vaccine development

Fish and Amphibian Alloherpesviruses

peer reviewe

Fish and Amphibian Alloherpesviruses (Herpesviridae)

peer reviewe

A complete check-up of European eel after eight years of restocking in an upland river: Trends in growth, lipid content, sex ratio and health status

By combining field research and careful laboratory analysis of samples over the course of an eight-year study, we met the challenge of assessing the life history traits and health status of eels restocked in freshwater ecosystems. We found that restocked eels exhibited good growth performance; moreover, the stocks were femaledominated, showed a good Fulton's condition factor (K) and lipid stores and had high survival probability estimated using the best model of Jolly-Seber stock assessment method for open populations. A necropsy revealed the absence of internal lesions. A swimbladder examination revealed the absence of the parasite Anguillicola crassus. Polymerase chain reaction (PCR) analyses revealed an increase of Anguillid herpesvirus-1 (AngHV-1) prevalence throughout the study. Most positive subjects expressed viral loads compatible with a latent infection and correlated positively with K. All restocked eels were contaminated by at least one of the organic pollutant congeners studied, but the pollution loads corresponded to the lowest range of pollutant concentrations reported in the available literature for European eels and did not exceed the maximum residue and contaminant limits in food and feed of several national and international regulations. Pollutant loads were negatively correlated with K, lipid content and eel density for polychlorinated biphenyls PCB 138, 153 and 180 and K for pesticides p.p’- DDE, p.p’-DDD, p.p’-DDT and PBDE47. This study highlights the potential role played by upland aquaticecosystems in enhancing riverine silver eel production from the perspective of species conservation. To be successful, restocking must be accompanied by improved ecosystem quality and migration routes for eels in inland freshwaters. We also provide some recommendations for future research to improve the management of restocking programmes

In Vivo Imaging Sheds Light on the Susceptibility and Permissivity of <i>Carassius auratus</i> to Cyprinid Herpesvirus 2 According to Developmental Stage

Cyprinid herpesvirus 2 (CyHV-2) is a virus that causes mass mortality in economically important Carassius spp. However, there have been no comprehensive studies into host susceptibility or permissivity with respect to developmental stage, and the major portal of viral entry into the host is still unclear. To help bridge these knowledge gaps, we developed the first ever recombinant strain of CyHV-2 expressing bioluminescent and fluorescent reporter genes. Infection of Carassius auratus hosts with this recombinant by immersion facilitated the exploitation of various in vivo imaging techniques to establish the spatiotemporal aspects of CyHV-2 replication at larval, juvenile, and adult developmental stages. While less susceptible than later developmental stages, larvae were most permissive to CyHV-2 replication, leading to rapid systemic infection and high mortality. Permissivity to CyHV-2 decreased with advancing development, with adults being the least permissive and, thus, also exhibiting the least mortality. Across all developmental stages, the skin was the most susceptible and permissive organ to infection at the earliest sampling points post-infection, indicating that it represents the major portal of entry into these hosts. Collectively these findings provide important fundamental insights into CyHV-2 pathogenesis and epidemiology in Carassius auratus with high relevance to other related economically important virus-host models

Correction: He et al. In Vivo Imaging Sheds Light on the Susceptibility and Permissivity of <i>Carassius auratus</i> to Cyprinid Herpesvirus 2 According to Developmental Stage. <i>Viruses</i> 2023, <i>15</i>, 1746

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