Shedding light on the pathogenic potential of Leishmania tarentolae parasites

Leishmania are protozoan parasites responsible for causing leishmaniasis, a spectrum of severe immunopathologies that affect both humans and animals worldwide. These diseases, which manifest in various forms, including cutaneous and visceral leishmaniasis, fall under the category of Neglected Tropical Diseases (NTDs), because they have historically received less attention and funding for research, prevention, and treatment compared to other infectious diseases. In contrast, Leishmania tarentolae parasites (Ltar), a close relative of human-infecting Leishmania species, are considered non-pathogenic to humans, as they primarily infect reptiles. However, recent studies indicate that cryptic transient infections in mammals may be possible, raising important questions about the true pathogenic potential of Ltar. Additionally, Ltar’s geographical distribution has recently been revisited, with evidence indicating that it is endemic to Southern Italy and likely other Mediterranean countries. 

Enter Estefanía Calvo Alvarez, a researcher at the Department of Pharmacological and Biomolecular Sciences, University of Milan in Italy. Her project, supported by ISIDORe funding, uses multimodal imaging services from Euro-BioImaging’s FiBI Node to analyse, for the first time, Ltar’s homing, tissue tropism, and innate inflammatory responses in both immunocompetent and immunosuppressed mice. Given that immunosuppression is a major risk factor for leishmaniasis, her ultimate goal is to test the hypothesis that, under immunosuppressed conditions, Ltar might exhibit its full pathogenic potential, an insight with potential implications for public health.

“I’m convinced that the pathogenic traits of Ltar have long been overlooked because we were not looking in the right direction. With the growing global population of immunocompromised individuals due to the prevalence of HIV/AIDS, the broad use of immunosuppressive therapies in autoimmune diseases, and advanced cancer treatments, it is crucial to thoroughly investigate whether Ltar can cause disease in these vulnerable subjects and potentially emerge as a pathogen of concern. To address these questions, I’ve been trying to understand its potential pathogenic features  with in vitro imaging and model systems, but there are limitations to this approach. That is why I needed access to in vivo imaging, which is not available at my home institute.”

Estefanía comes from an in vivo imaging background. She did her PhD in Spain using in vivo imaging approaches to study the infection biology of Leishmania parasites. During her post-doc at Institut Pasteur in Paris, she worked on a relative parasite species, Trypanosoma brucei, also using in vivo imaging systems. But when she joined the University of Milan, she moved away from in vivo techniques. Her current group specializes in immunology, but they do not work with animals. Estefanía knew she would have to work collaboratively with an in vivo imaging center to build her experiment. 

Making projects happen

Estefanía made contact with the in vivo imaging service at San Raffaele Hospital in Milan, part of Euro-BioImaging's MMMI node, in the context of another research project. This was her first introduction to Euro-BioImaging. Later on, while browsing the Euro-BioImaging website, she learned about ISIDORe funding - a perfect fit for her infectious disease-focused project. She immediately applied for funding to access imaging services through  ISIDORe.

Image: Estefanía in the sample prep room at the University of Milan.

Her project passed the first pre-selection phase and was assigned to Euro-BioImaging. Erika Cerutti, Euro-BioImaging's Scientific Project Manager at the Med-Hub in Torino supported her during the application process and put her into contact with the potential Node that could support her project. 

Following the project remotely

But the project took on a new scope. Based on the imaging technology required for her project , Euro-BioImaging recommended a facility at the University of Helsinki, part of the Finnish Biomedical Imaging Node. After the initial excitement came a big disappointment for Estefanía as she was not able to travel  to undertake the experiment. And no one in her group had the expertise to replace her. But what if the project could be carried out remotely, by colleagues at the Finnish Biomedical Imaging Node? 

“I would have really loved to be there. I know how to perform in vivo imaging and was looking forward to carrying out the experiment myself. Luckily, Marja Lohela of the University of Helsinki was willing to do the experiment on my behalf. I know it wasn’t easy for her, she had to juggle many things - she even hired a student to help out - but she did great.  We interacted a lot by email, telephone, or WhatsApp - she always kept me in the loop.” 

Marja Lohela is an intravital imaging specialist  working at the Biomedicum Imaging Unit, a core facility that is part of Euro-BioImaging’s FiBI Node. “We are used to working with visiting scientists and helping them with their imaging projects, but this was the first time we performed an entire experiment without the researcher present on site,” explains Marja. It was clear from the start that I could not manage a big in vivo experiment with a challenging timetable on my own, and being able to recruit a capable student was instrumental for the completion of Estefanía’s project.” 

“An in vivo imaging project conducted as a remote access service needs extremely careful planning beforehand,” explains Marja. In addition to the detailed experimental plans defined at the planning and application stages, Estefanía prepared diagrams of the progress of the experiment, with daily checklists for all procedures, and gave detailed instructions for handling the parasite cultures, sample collection, and other parts of the experiment that were outside of the facility’s imaging expertise. Constant communication with Estefanía was very important, because despite the most careful communication, one can never think of absolutely everything - and Biology always serves us with some surprises!” adds Marja.

Thank you, Euro-BioImaging, and in particular the FiBI Node, for your help! I would really recommend Euro-BioImaging to everyone, because it has really been a great experience.

• - Estefanía Calvo Alvarez
  

The strain of mice was chosen together with the FiBI Node. Marja and the student hired at University of Helsinki prepared the entire experiment including animals management and acquisition of the reagents. Estefanía prepared the parasites, and shipped the culture to Finland.

An efficient project process

“Shipping parasites is easy enough,” explains Estefanía. “They are considered non-pathogenic and they arrive live, overnight delivery.” 

Since Marja had never worked with Leishmania parasites before, she made a video to show Estefanía what the parasites looked like when they arrived. “It was really nice of her - just to make sure everything was OK,” says Estefanía.

“Marja ran the project really efficiently,” remarks Estefanía. “We started our planning and discussions in September 2024 and the project was over by November. The infection week was respected, infection and imaging took two weeks. I’ve received some of the data and we are performing data analysis at the moment.”

“It was very exciting to be involved in such an important and interesting project, getting to know something about an entirely new field of research. It was also a great pleasure to work with Estefanía, whose enthusiasm, clear scientific focus and careful planning really made it possible to conduct this project as a remote access service. This has also been a very useful learning experience for me about the management of such services, which will definitely benefit both me and the facility in the future,” says Marja (to learn Marja’s recommendations for running a remote user project, read the full story here).

“Erika and all of the team at Euro-BioImaging have been very kind and supportive, the team has really facilitated the project, making it possible. My initial project had to be modified, Euro-BioImaging had a lot of interactions with the ISIDORe team on my behalf,” comments Estefanía.

“Thank you, Euro-BioImaging, and in particular the FiBI Node, for your help!” exclaims Estefanía in conclusion. “I would really recommend Euro-BioImaging to everyone, because it has really been a great experience.”