Author: marina02

Molecular characterization of Proteocephalus (Cestoda) species parasitizing salmonids across Russian freshwater ecosystems

Introduction.Proteocephalus spp. are widely distributed cestode parasites infecting freshwater fishes. Recent taxonomic revisions within the genus have led to uncertainties regarding its species diversity. Under the current taxonomy, cestodes parasitizing salmonids in Russia have been classified as P. thymalli (infecting graylings, Thymallus spp.) and P. longicollis and P. exiguus (found in other salmonid fishes). This study aims to clarify the species composition of Proteocephalus parasitizing salmonids (Salmonidae) in Russian water bodies through molecular analysis of partial mitochondrial cox1 and nuclear 28S rRNA gene sequences.

Methodology. We examined cestodes collected from six fish species (Coregonus lavaretus, C. peled, C. migratorius, C. muksun, Salvelinus malma, and Thymallus baicalensis) across 11 water bodies spanning from the Kola Peninsula to Kamchatka. Notably, two of these fish species (C. lavaretus and S. malma) form complexes of sympatric morphs in the studied lakes. Gene fragments of the nuclear 28S rRNA (1,434 bp) and mitochondrial cox1 (1,548 bp) were amplified and sequenced. Phylogenetic analyses were conducted using maximum likelihood (ML) in IQ-TREE 2 and Bayesian inference (BI) in MrBayes v.3.2.1.

Results. Both maximum likelihood (ML) and Bayesian inference (BI) analyses of the 28S and cox1 gene sequences produced phylogenies with congruent topologies. The cestode sequences derived from salmonid hosts formed seven well-supported, species-level clades. Sequences from Coregonus spp. populations from Transbaikalia, the Kola Peninsula, and a broad geographic range from the Altai region to northern Yakutia clustered into three distinct clades. One clade consisted of samples from Lake Verkhnee Ondomozero (Kola Peninsula) and included GenBank reference sequences of P. fallax. The remaining two clades showed no matches to existing reference sequences, indicating they likely represent undescribed Proteocephalus taxa. Cestodes from C. migratorius formed a separate, monophyletic clade at the species level. Sequences from T. baicalensis cestodes grouped into a distinct clade identified as P. thymalli. Finally, cestodes from S. malma in Lake Kronotskoye (Kamchatka) and Lake Gytgyl (Chukotka) formed two non-sister clades.

Conclusions. Our findings reveal at least three regionally distributed Proteocephalus species parasitizing whitefish. For graylings we confirmed parasitism by the valid cestode species P. thymalli. The Baikal omul appears to host a previously undescribed Proteocephalus species. Furthermore, our data indicate that S. malma serves as a host for at least two putative Proteocephalus species.

This work was supported by the Russian Science Foundation no. 23-74-10101.

1. VLASENKO, PAVEL, ISEA SB RAS, Author
2. KOLMOGOROVA, TATIANA, ISEA SB RAS, Author
3. MARKEVICH, GREGORY, IPEE RAS, Author
4. KASHINSKAYA, ELENA, ISEA SB RAS, Author
5. SOLOVYEV, MIKHAIL, ISEA SB RAS, Presenter

Reassessment of Myxobolus toyamai morphology in koi carp using an enhanced Giemsa warming technique: A first report from Vietnam

Myxobolus toyamai, a well-established gill parasite of common carp, Cyprinus carpio, has been implicated in infections across multiple continents. Despite numerous attempts to classify this parasite based on the structural features of its polar capsules, existing staining techniques have proven inadequate, resulting in unclear structural definitions. This study represents the first report of M. toyamai infection in koi fish in Vietnam, along with the optimisation of a staining technique using Giemsa warming to accurately define its morphological characteristics. In addition, the study includes the analysis of the 18S rRNA gene and histopathological evaluations. The results show that the Giemsa warming staining method effectively clarifies the structure of the polar capsules, revealing distinct measurements that differ from those reported in previous studies. Furthermore, sequence analysis of the 18S rDNA gene demonstrates a high degree of similarity to M. toyamai, confirming its identification in the examined specimens.

1. HOAI, TRUONG, VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE, Presenter
2. Andrew, Shinn, INVE, Author

A retrospective histopathological study of disease in marine ornamental fish

This retrospective histopathological study reviews diseases of marine ornamental fish submitted by clinicians from public aquaria or from private practices to the Veterinary Pathology laboratory at the University of Camerino between 2012 and 2024. A total of 416 fish belonging to 158 different species (381 teleosts and 35 elasmobranchs) were examined. Hippocampus was the most commonly submitted genus (29 cases). Case materials included formalin-fixed fish or tissues. Tissues were routinely processed, sectioned at 5 microns, and stained with hematoxylin and eosin (HE). Select tissues were stained with the following special stains: Ziehl-Neelsen (ZN), Fite’s acid fast (AF), periodic acid–Schiff (PAS), Brown and Brenn (B&B), Giemsa and Gomori methenamine silver (GMS). Immunohistochemical staining or polymerase chain reaction testing was performed when attempting to identify infectious agents. Based on the morphological and etiological diagnoses obtained from case reports, the cases were classified into different disease groups as: inflammatory/infectious, inflammatory/non infectious, regressive (degenerative, metabolic), vascular and neoplastic. Inflammatory/infectious disease was the most frequently diagnosed pathological process (74%). The predominant etiological agents were bacteria, with a large prevalence of Gram-negative bacteria frequently causing sepsis, branchitis, enteritis and mycobacteria causing variably organized granulomas localized to one or more organs. Histological lesions consistent with megalocytivirus infection were pointed out in two cases of fish died during the quarantine period confirmed by real-time PCR analysis. The most commonly encountered parasitic diseases were cryptocaryoniasis and scuticociliatosis, both corresponding to 22% of the diagnosed parasitic diseases. In order of frequency, infectious diseases were followed by regressive/degenerative processes (17%) and noninfectious inflammatory processes (14%). Noteworthy was the presence of multiple pathological processes in the same case: in 12% of cases a condition of comorbidity was observed. In 52% of the cases in which the co-presence of more than one infection was observed, one of the etiological agents was Mycobacterium spp.

1. Magi, Gian Enrico, University of Camerino, Presenter
2. Pritelli, Antonella, Veterinary Practitioner, Author
3. Ciiulli, Sara, University of Bologna, Author
4. Bergamini, Chiara, University of Camerino, Author

First detection of the Fancy-Tailed Guppy Virus (FTGV) in Europe

Introduction: The Fancy-Tailed Guppy Virus (FTGV) is a pathogen of the guppy (Poecilia reticulata) and a new member of the emerging Amnoonviridae family of viruses, which are proving to be some of the most virulent and dangerous for fish. Due to its segmented genome, the virus may be prone to recombination, which can affect infectivity and complicate diagnostics. Until now, the virus has been detected in the USA in fish populations originating from Asia and South America, in both clinically affected and healthy fish. As the guppy is widely used as an ornamental pet and a biological model, health surveillance programmes are needed.

Methodology: Samples were collected from guppies presented for examination over one month after a disease outbreak leading to a large-scale die-off. These fish were collected from an undisclosed ornamental fish retailer and originated from an ornamental fish breeder in Asia. They did not display any clinical signs of infection. Additional samples were collected from clinically healthy, unrelated, laboratory-raised experimental populations with no history of health problems, originating from South America. Classical virological methods such as cell culture-based virus cultivation and a range of qPCR and RT-qPCR assays were employed in combination with metabarcoding of bacterial populations (Nanopore 16S sequencing) and immune response profiling to identify the agent causing the disease and mortality. Further targeted FTGV testing was performed to confirm the absence of the pathogen in fish from the laboratory-raised experimental populations.

Results: A total of five batches of fish from the retailer and five lab-raised populations were tested. Two populations originating from the disease outbreak were confirmed to be positive for FTGV, with high or moderate Ct values indicating persistent subclinical infection. The virus was re-isolated using E-11 cells. Partial genome sequencing and phylogenetic analysis revealed significant variation in the FTGV isolates. No clear indication of the involvement of pathogenic bacteria was detected. Evaluation of immune responses was not helpful due to the late subclinical phase of the infection. The laboratory-raised experimental populations were confirmed to the FTGV-free.

Conclusions: This study confirms the presence of FTGV in guppy populations associated with a major mortality event. It highlights the potential for persistent subclinical infection and significant genetic variability of the virus and emphasises the need for targeted surveillance and biosecurity measures in the ornamental fish trade.

1. ADAMEK, MIKOLAJ, UNIVERSITY OF VETERINARY MEDICINE HANNOVER, Presenter
2. Ramirez Stoll, Tamara, UNIVERSITY OF VETERINARY MEDICINE HANNOVER, Author
3. Debnath, Dipa, UNIVERSITY OF VETERINARY MEDICINE HANNOVER, Author
4. Krebs, Torben, UNIVERSITY OF VETERINARY MEDICINE HANNOVER, Author
5. Korytar, Tomas, Laboratory of Fish Immunology, Institute of Parasitology, Biology Centre CAS, Ceske Budejovice, Czechia, Author
6. Falco, Alberto, Fish Pathology Group, Institute of Aquaculture Torre de La Sal-Spanish National Research Council (IATS-CSIC), Cabanes, Castell&a, Author
7. Soto, Esteban, Department of Medicine and Epidemiology, School of Veterinary Medicine, University of California, Davis, California, USA, Author
8. Rakus, Krzystof, Department of Evolutionary Immunology, Institute of Zoology and Biomedical Research, Faculty of Biology, Jagiellonian University, Author
9. Herdegen-Radwan, Magdalena, Department of Behavioural Ecology, Institute of Environmental Biology, Faculty of Biology, Adam Mickiewicz University, Poznan, P, Author
10. Radwan, Jacek, Evolutionary Biology Group, Faculty of Biology, Adam Mickiewicz University, Poznan, Poland, Author
11. Jung-Schroers, Verena, UNIVERSITY OF VETERINARY MEDICINE HANNOVER, Author

Unveiling Perkinsus olsenii and P. chesapeaki across their French distribution using a touch of ecology and genetics

Marine parasites, particularly Perkinsus spp., pose significant threats to mollusc health, compromising biodiversity and generating ecological and economic consequences. Perkinsus olseniiand P. chesapeaki, two protozoans detected in clam populations in Europe and Asia, were likely introduced to the French coastline alongside their -suggested- host species, Ruditapes philippinarum and Mya arenaria, respectively. These parasites may have since expanded their host range to endemic French clams. In France, data concerning their distribution remains limited, thus necessitating a more in-depth investigation to facilitate a more comprehensive understanding of their role within host populations.

In the present study, we aim to assess the distribution of P. olsenii and P. chesapeaki along the French coastline, exploring their dynamics and genetic diversity across different sites, seasons and host species (Ruditapes philippinarum and R. decussatus).

Clams were collected from four sites: a Mediterranean site, (Thau lagoon) and three Atlantic sites (Saint-Brévin, Noirmoutier and Pen Bé-Croisic) at four distinct time points in 2023. DNA from ca. 600 clams was extracted for host identification and the detection of P. olsenii and/or P. cheasapeaki using PCR. Parasite diversity was studied by sequencing the ITS and actin regions. The prevalence of Perkinsus was not influenced by the season, but rather site-dependent, with infection ranging from 0% at Saint-Brévin to 94% at Thau lagoon. Infection rates were also host-dependant, R. decussatus showing significantly higher infection rates than R. philippinarum. Genetic analyses on Perkinsus sp. showed a low polymorphism along the French coast, suggesting an important founder effect.

These findings underscore the importance of considering various aspects including host species and genetic factors into the study of the Perkinsus parasites in clam populations. Furthermore, they highlight the need for continued monitoring and research in order to mitigate the potential ecological and economic risks associated with these parasites.

1. DURAND, ANTOINE, IFREMER, Author
2. CHOLLET, BRUNO, IFREMER, Author
3. ARZUL, ISABELLE, IFREMER, Presenter
4. GARCIA, CELINE, IFREMER, Author
5. RIQUET, FLORENTINE, IFREMER, Author

Cytopathology and phylogeny of Pinna nobilis Picornavirus (PnPV) from the Faro Lake (Southern Italy)

Introduction

Pinna nobilis Picornavirus (PnPv) is an RNA virus that proliferates in the immune cells of Pinna nobilis. As part of the LIFE PINNARCA project, this study was conducted to assess the health of the Pinna nobilis population in Lake Faro, Sicily, a protected site previously identified as a hot-spot for this species survival. The study detected the presence of PnPv in the Pinna nobilis population that was previously reported in Spain (Catalonia and Murcia) and Italy (Venice lagoon).

Methodology

Conservative sampling, such as mantle biopsies and haemolymph, were performed on six individuals from the Faro Lake (Sicily). Immune cells were analysed by transmission electron microscope (TEM), cytology and used for RNA-based qPCR. DNA and RNA extraction was performed from the mantle samples to detect Haplospodium pinnae parasites and PnPv, also determining the tropism of PnPV.

Results

Transmission Electron Microscopy detected the presence of the picornavirus in all specimens. Molecular diagnostic of Haplosporidium pinnae detected the parasite in three out of six animals, while acid-fast bacteria were found in two out of six individuals in cytological preparations. Phylogenetic analysis of PnPv confirmed that the virus belongs to the family Marnaviridae and probably descends from an algal host ancestor.

Conclusion

This discovery provided important information on the evolutionary relationships and taxonomy of picornaviruses, improving the understanding of the disease and bringing new insights into the cytopathology and phylogeny of PnPV. It remains necessary to develop a diagnostic test capable of simultaneously extracting DNA and RNA from hemolymph samples to understand disease pathogenesis. Future investigations should also check the infection presence in residual surviving P. nobilis populations in other geographic areas.

1. IARIA, CARMELO, UNIVERSITY OF MESSINA, Presenter
2. PIRO, MARIA GIOVANNA, UNIVERSITY OF MESSINA, Author
3. NATALE, SABRINA, UNIVERSITY OF MESSINA, Author
4. CAPPARUCCI, FABIANO, UNIVERSITY OF MESSINA, Author
5. VILLARI, GRAZIA, UNIVERSITY OF NAPLES FEDERICO II, Author
6. DE VICO, GIONATA, UNIVERSITY OF NAPLES FEDERICO II, Author
7. MARINO, FABIO, UNIVERSITY OF MESSINA, Author
8. CARELLA, FRANCESCA, UNIVERSITY OF NAPLES FEDERICO II, Author

Mining NCBI sequence read archive (SRA) data reveals the presence of Macrobrachium rosenbergii golda virus (MrGV) in giant river prawn larvae across Asia

Background: Macrobrachium rosenbergii golda virus (MrGV) was first characterised in Macrobrachium rosenbergii larvae in 2020, associated with mass mortalities in multiple Southern Bangladesh prawn hatcheries. MrGV has since been detected in two metatranscriptomic datasets from M. rosenbergii in China, however disease or poor condition was only noted in one animal. The major objective of this study was to further characterise the geographical spread of MrGV by mining the NCBI Sequence Read Archive (SRA) database.

Methods: Utilising a new database, Logan, generated from assembling the entire NCBI sequence read achieve (SRA), we mine the Macrobrachium SRA database up to December 2023 for MrGV to further characterise the geographical spread of the virus, phylogenetics associated with geographical location, and the species and life stages MrGV sequence data are present in.

Results: We discovered MrGV sequence data in M. rosenbergii SRAs from China, Thailand, and India. Presence and relative abundance of MrGV was mostly associated with the larval life stage of M. rosenbergii, with MrGV reads present in 61% of larval M. rosenbergii SRAs (n=150), and only 7% of adult SRAs (n=201). Using data from an NCBI BioProject that studied the transcriptomics of larvae from hatching to 25 days post-hatch, we also show that relative abundance or MrGV increased over time.

Conclusions: These results provide insights into the current prevalence of MrGV globally and suggest the life stages of prawn that should be screened to prevent spread of the virus. We also show that the Logan database allows NCBI SRAs to be searched more efficiently and at an affordable cost to detect the presence of known organisms in large sequencing datasets.

1. Hooper, Chantelle, The Centre for Environment, Fisheries and Aquaculture Science, Presenter
2. van Aerle, Ronny, The Centre for Environment, Fisheries and Aquaculture Science, Author
3. Ryder, David, The Centre for Environment, Fisheries and Aquaculture Science, Author
4. Coyle, Nicola, The Centre for Environment, Fisheries and Aquaculture Science, Author

Assessing PFOS Impact on Oxidative Stress and Tissue Damage in Ruditapes sp.

Introduction

In recent years, there has been an increasing attention on the issue of pollution by perfluoroalkyl compounds (PFAS), particularly perfluorooctanesulfonic acid (PFOS). This highly persistent and bioaccumulative compound, which is widely used in industrial and consumer products, has raised concerns due to its potential toxicity and consequent impact on marine organisms and the safety of seafood products. Marine bivalves, such as Ruditapes sp., are valuable bioindicators of environmental pollution, as they are directly exposed to contaminants in coastal waters and aquaculture areas. In these organisms, PFOS bioaccumulation can impair essential physiological functions by altering oxidative status, tissue structure, and immune response.

Methodology

We aimed to investigate the chronic effects of PFOS exposure on oxidative stress, tissue integrity, and immune responses in Ruditapes sp. specimens collected from local aquaculture farm. Initially, the median lethal concentration (LC??) of perfluorooctanesulfonate (PFOS) at 25.89 mg/L was determined. Subsequently, sixty clams were divided into a control group (T1) and three experimental groups (T2, T3, T4), which were exposed to increasing concentrations of PFOS (1, 10 and 20 mg/L, respectively) for 15 days at 22°C. At the end of the exposure period, a necropsy analysis was conducted, and tissues from the gills and digestive glands were sampled for histological and immunohistochemical analysis to assess pathological changes and immune system impairment. Indeed, molecular assays were conducted to quantify oxidative damage and defence mechanisms. Specifically, malondialdehyde (MDA) levels were measured as an indicator of lipid peroxidation, catalase activity was assessed to evaluate antioxidant responses, and nitrite levels were analysed to investigate nitrosative stress.

Results

The results showed a dose-dependent toxicity with a significant increase in nitrite formation, lipid peroxidation and catalase. Indeed, exposure to 20 mg/L of PFOS led to a substantial upregulation in the expression of the three damage markers associated with contaminant toxicity, in comparison to the lower and control groups. Histological examination revealed the presence of inflammatory elements, lipofuscin accumulation, and areas of vacuolization in the gills and deformation of the digestive gland tubules, while immunohistochemical analysis indicated an altered immune response.

Conclusion

Overall, this study highlights the susceptibility of Ruditapes sp. to PFOS contamination and underscores the potential ecological and public health risks associated with the presence of these persistent pollutants in aquaculture areas. Further research is essential to understand the long-term consequences of PFOS exposure on commercially significant bivalves.

1. NATALE, SABRINA, UNIVERSITÀ DI MESSINA, Presenter
2. PERITORE, ALESSIO FILIPPO, UNIVERSITÀ DI MESSINA, Author
3. CAPPARUCCI, FABIANO, UNIVERSITÀ DI MESSINA, Author
4. IARIA, CARMELO, UNIVERSITÀ DI MESSINA, Author
5. LANTERI, GIOVANNI, UNIVERSITÀ DI MESSINA, Author

Impact of oral immune priming on Manila clam (Ruditapes phillippinarum) microbiota

Growing evidence demonstrates that invertebrates display adaptive-like immune abilities, commonly known as “immune priming”: if invertebrates were previously exposed to the pathogen they will be better protected to subsequent re-infections. Immune priming has been demonstrated to date in a few species from different Phyla (Ctenophora, Cnidaria, Mollusca, Nematoda and Arthropoda). However, it has been barely studied in bivalve mollusks, and particularly in clams, despite its ecological significance and the high economic impact in worldwide aquaculture, such as Manila clam (Ruditapes phillippinarum) a Top Ten species for this industry.
Immune priming has been triggered by injection of the pathogen in the host in most bivalve’s reports, including Manila clam. However, bivalves are filter feeders and the oral route is their natural route of infection and, thus, the best way to obtain an accurate picture about how immune priming works. Here, we have established Manila clam as an experimental model organism for studying oral immune priming to mimic the bivalve’s natural route of infection. Manila clam juveniles orally exposed to a sub-lethal concentration of the bacterial pathogen Vibrio europaeus showed increased survival upon a subsequent challenge with a lethal dose of this bacterium. The impact of the immune priming on the Manila clam microbiota was also investigated for first time. Clam’s microbiota was characterized by full-length 16S rRNA metabarcoding and the bacterial pathogen, Vibrio spp and total bacteria quantified by qPCR. Different bacterial taxa and species overrepresented were identified in primed clams and the bacterial dynamics due to immune priming were established. Although the exact mechanism of oral immune priming in clams is unclear, we have demonstrated that host’s microbiota shifted as consequence of immune priming. Our results suggested that microbiota could play an important role in this phenomenon, and thus, immune priming could be influenced by the interaction of hosts/symbionts/pathogens.

1. Dubert, Javier, University of Santiago de Compostela, Presenter
2. Rodino-Janeiro, Bruno K., University of Santiago de Compostela, Author
3. Rodriguez, Sergio, University of Santiago de Compostla, Author
4. Dieguez, Ana L., University of Santiago de Compostela, Author
5. Toranzo, Alicia E., University of Santiago de Compostela, Author

The WAVES Consortium: Working towards adaptive and versatile environmental sustainability in mollusc aquaculture in Southeast Asia

Introduction: Mollusc aquaculture produces 20 million tonnes live biomass annually, with Asia hosting >95% of activity and bivalves dominating production. Non-fed species, such as oysters, mussels and benthic clams, offer a ‘low-carbon’ solution to high-quality nutritional security whilst conferring environmental benefits for biodiversity and seawater nutrient status. Molluscs are inexpensive and nutritionally rich, and sector expansion can enhance food security in Southeast Asia. Nevertheless, mollusc output as a proportion of aquatic animal aquaculture has declined to 20.3% from 30.2% since 2000, with producers facing many challenges including climate change and disease. Questions remain as to whether mollusc culture can develop and grow into a sustainable and resilient industry by surmounting seed supply constraints and changes in production resulting from climate change. The WAVES Consortium is a UKRI-funded project (2025-28) aiming to address sector challenges holistically to enhance system resilience and promote sustainable sector growth.

Methodology: A systems-thinking approach with stakeholders at its heart is core to achieving our ambition. We are: conducting systems mapping of current mollusc production in Vietnam, Malaysia and Indonesia to understand activities, livelihoods and climate change threats; creating a systems dynamic model to forecast plausible futures for mollusc aquaculture; generating data to support species diversification and hatchery development; and developing context-sensitive roadmaps for sustainable and resilient sector expansion.

Results: Our international, multidisciplinary consortium held its first stakeholder workshop focused on clam production in Hanoi, Vietnam. A systems map was co-produced that provided a visual representation of sector activities and threats, including those driven by climate change such as incidences of disease. The map was interrogated by participants to propose interventions that would mitigate against the identified challenges. Supplementing this new map with data derived from stakeholder interviews, surveys and peer-reviewed and other literature provides the platform for our next step, which is the creation of a systems dynamic model of the sector.

Conclusions: An holistic approach, working closely with stakeholders, is key to identifying pathways towards a resilient and sustainable expansion of the mollusc sector in Southeast Asia. Development of non-fed aquaculture, through improved productivity and enhanced natural resource use, is a key strategy to tackling climate change and marine environment degradation, whilst supporting livelihoods and providing increased food and nutritional security.

1. DESBOIS, ANDREW, INSTITUTE OF AQUACULTURE, UNIVERSITY OF STIRLING, Presenter