PhD students

Giulia Zarantonello

Title of the PhD project
Early warning for disease: improving aquaculture monitoring with real-time microbiome sequencing.

Supervisors
Argelia Cuenca, Lone Madsen & Thomas Nordahl Petersen.

Background of the project
The aquaculture microbiome balance is crucial for the health status of the system, such that dysbiosis has been reported when a stressor, such as a pathogen, is introduced. Current farmed fish diagnostics methods consist in infection event monitoring and imply histopathology, culture isolation, and targeted molecular diagnostics for the suspected pathogen. However, this approach comes with drawbacks: action is only taken after manifestation of clinical signs or increased mortality, and standard diagnostics are targeted towards known pathogens, which impairs new pathogen discovery, especially when the microorganisms are unculturable.

About the project
My PhD project aims to develop a rapid, untargeted NGS-mediated workflow for early detection of declining health conditions and microbial disease for farmed animals in aquaculture, by exploiting the microbiome as an indicator for the state of the system. First, I will implement a microbiome sequencing protocol with Oxford Nanopore Technology from various aquaculture-relevant samples, both environmental (eDNA) and host-associated. Then, I will test the protocol in stress-induced conditions (pathogen, organic waste) in experimental RAS facilities, to detect possible changes in the healthy microbiome associated with fish health. The protocol will then be translated to detect distress-correlated dysbiosis in industry samples. Finally, I will apply metagenomics sequencing for pathogen discovery for a salmonid skin disease, whose unculturable disease-causing agent is still unknown.

Perspectives
My project aims to exploit aquaculture-related microbiome sequencing to integrate current diagnostics with a non-lethal, fast and untargeted community surveillance method. If successful, the outcome of my PhD could represent the first steps towards the development of a novel indicator of fish health, such as microbiome risk scores for disease prediction. Early detection of distress could favour preventive strategies to minimize the impact on aquaculture production.

Alejandra Villamil Alonso

Title of the PhD project
Bacterial kidney disease in rainbow trout, with focus in infection kinetics and molecular tracing of Renibacterium salmoninarum in Denmark

Supervisors
Argelia Cuenca, Lone Madsen & Niccolò Vendramin

Background of the project
Bacterial kidney disease (BKD) is a systemic infection that affects wild and farmed salmonids, compromising aquaculture systems worldwide. The causative agent of BKD is Renibacterium salmoninarum, a Gram-positive intracellular bacterium characterized by chronic disease progression and able to spread both horizontally and vertically. Although efforts have been made to characterize the bacteria mechanisms of transmission, pathogenesis, and immune evasion, they remain poorly understood. Moreover, no knowledge is available on the introduction and molecular evolution of the pathogen in Denmark, which is of foremost importance for understanding the current and past movements of R. salmoninarum. 

About the project
My PhD project seeks to elucidate the interplay between host-pathogen-environment by first establishing a challenge model of R. salmoninarum in rainbow trout (Oncorhynchus mykiss), the predominant fish species farmed in Denmark, in recirculating aquaculture-like systems. Bacteria route and persistence inside the fish will be studied, as well as the influence of different environmental stressors such as water temperature and water quality in BKD development. I will also work on the development of improved diagnostics methods for R. salmoninarum, focusing on qPCR and targeted detection of eDNA on water systems. Eventually, I will study the origin and molecular evolution of R. salmoninarum in Denmark by genome sequencing of a collection of historical and new isolates originated in Danish farms for phylogeographic and molecular analyses.

Perspectives
With this project, we expect to gain knowledge on BKD transmission and progression in rainbow trout, as well as to build a deeper understanding on the virulence mechanisms of the pathogen and the subsequent host immune response. Together with the planned genomic studies, this research will facilitate the development of novel diagnostic methods and contribute to the identification of improved prevention and treatment measures for BKD.

Shana Fresnido Genavia

Title of the PhD project
Potential role of circular viral DNA in the shrimp immune system 

Supervisors
Niels Lorenzen and Dagoberto Andres Sepulveda Araneda, DTU Aqua & Morten Schiøtt, DTU Bioengineering

Background of the project
The immune system in animals is a multi-faceted defense mechanism against pathogens. In vertebrates, the immune system operates through both innate and adaptive mechanisms, the latter being capable of generating immunological memory. Traditionally, invertebrates, such as shrimp, have been understood to possess only innate immune systems, devoid of any capacity for immune memory. This paradigm is being challenged as more research suggests invertebrates demonstrating a form of immunological memory, albeit not antibody-based. Understanding this could have a broad impact on industries like aquaculture, which often grapple with viral diseases that threaten shrimp populations.

About the project
This PhD project aims to explore the role of circular viral DNA (cvDNA) in the immune response mechanism of shrimps against invading viruses. Utilizing Penaeus vannamei (Pacific whiteleg shrimp) as a model organism, the project will investigate whether cvDNA molecules are produced during viral infections and if these molecules serve as templates for RNAi-induced antiviral immune response. The project aims to address the biological aspects of cvDNA as well as its potential for conferring viral resistance and longevity of immunity. Methodologically, the study will employ a combination of molecular techniques, sequencing, and bioinformatics analysis to examine  cvDNA and its implications.

Perspectives
The implications of this research extend beyond shrimp aquaculture. If shrimp do possess a form of immune memory mediated by cvDNA, this could revolutionize our understanding of invertebrate immunity and even have repercussions for vertebrate immune systems. Moreover, it could open up new avenues for combating viral diseases in commercial aquaculture. The robust nature of cvDNA may offer innovative methods for studying virus-host interactions, both in contemporary and historical contexts. This has implications not only for animal husbandry but also for broader public health policies, especially considering the emerging evidence that vertebrates may utilize similar pathways.

Joyce Arguelles Hilario

Title of the PhD Project
Development of mucosal vaccine delivery strategies for farmed rainbow trout

Supervisors
Niels Lorenzen & Dagoberto Andres Sepulveda Araneda, DTU Aqua

Background of the project
Aquaculture is the most sustainable form of animal husbandry in terms of environmental impact, but infectious diseases cause significant losses, often requiring antibiotics. Lost production as well as use of antibiotics compromise both financial and environmental sustainability of aquaculture. The key to overcoming these problems is improved disease prophylaxis, with vaccination being one of the most effective tools. While injection vaccination is well established for larger-sized fish, early vaccination of smaller fish is less developed due to the lack of efficient vaccines delivered by mucosal routes. The mechanisms of immune activation and protection remain to be fully understood.

About the project
The project aims to develop and test formulation strategies for mucosal delivery of recombinant viral vaccines based on DNA and/or recombinant proteins in rainbow trout (Oncorhynchus mykiss), targeting diseases that infect early life stages. First, I will optimize the vaccine formulation by incorporating mucosal adjuvants and explore different delivery methods, including inactivated transformed bacterial cells. Next, I will test vaccination strategies, focusing on dose, delivery, and adjuvant aspects under laboratory conditions. Finally, I will evaluate the protective immunity in vaccinated fish through infection trials and identify immune response elements correlating with protection, providing insights into immune mechanisms and ensuring long-term vaccine efficacy.

Perspectives
Optimized mucosal vaccines that enhance uptake and stimulate both mucosal and systemic immune responses could address disease problems in small-sized fish. These vaccines would be easier to administer, reduce labor costs, and ultimately contribute to the overall health and welfare of fish population. This approach will not only improve aquaculture productivity and sustainability but also offer valuable insights into the mechanisms of fish mucosal immunity, thus, contributing to the broader field of immunology and vaccine development.

New PhD students

• Chiana Cialini
  Prophylactic measures against disease with Flavobacterium psychrophilum in rainbow trout; effect on pathogen as well as host, elucidated by e.g. microbiome studies

• Caitlin Yoo
  Economic evaluation of diseases in Danish aquaculture 

• Odunayo Esther Sholanke
  Development and testing of bioencapsulation of fish vaccines for oral delivery

Presentations of the PhD projects will follow.

Previous PhD students within the research area Fish and Shellfish Diseases

Juliane Sørensen
Mitigating the impact of PRV-3 infection in rainbow trout production by high-throughput diagnostic platforms 

Sofie Barsøe
Vaccination of sea bass against a lethal viral disease and characterization of protective immunity

Valentina Laura Donati
Bacteriophage-based control of Flavobacterium psychrophilum in rainbow trout. Studies on phage-treatment of rainbow trout at fry and eyed egg stages and effects on gut microbial communities

Anna Luiza Farias Alencar
Study of virulence markers in Viral Haemorrhagic Septicaemia Virus (VHSV)

Niccoló Vendramin
Piscine orthoreovirus. Distribution, characterization and experimental infections in salmonids