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The development of UAVs and multispectral cameras has led to remote sensing applications with unprecedented spatial resolution. However, uncertainty remains on the radiometric calibration process for converting raw images to surface reflectance. Several calibration methods exist, but the advantages and disadvantages of each are not well understood. We performed an empirical analysis of five different methods for calibrating a 10-band multispectral camera, the MicaSense RedEdge MX Dual Camera System, by comparing multispectral images with spectrometer measurements taken in the field on the same day. Two datasets were collected, one in clear-sky and one in overcast conditions on the same field. We found that the empirical line method (ELM), using multiple radiometric reference targets imaged at mission altitude performed best in terms of bias and RMSE. However, two user-friendly commercial solutions relying on one single grey reference panel were only slightly less accurate and resulted in sufficiently accurate reflectance maps for most applications, particularly in clear-sky conditions. In overcast conditions, the increase in accuracy of more elaborate methods was higher. Incorporating measurements of an integrated downwelling light sensor (DLS2) did not improve the bias nor RMSE, even in overcast conditions. Ultimately, the choice of the calibration method depends on required accuracy, time constraints and flight conditions. When the more accurate ELM is not possible, commercial, user-friendly solutions like the ones offered by Agisoft Metashape and Pix4D can be good enough.

Visceral leishmaniasis (VL) is a lethal parasitic disease caused by the intracellular protozoan Leishmania and is transmitted through the bites of sand flies. In the vertebrate host, parasites infect cells of the liver, spleen and bone marrow (BM). Treatment options are scarce, as are novel leads in the R&D pipeline. Moreover, toxicity, resistance and post-treatment relapse are common and currently no effective test-of-cure exists. There is no human vaccine available and knowledge on protective immunity is limited. All of these obstruct disease elimination and constitute a major clinical concern. In this thesis, a specific cell type was uncovered as source of relapse, namely the long-term hematopoietic stem cell (LT-HSC) in the BM. An enormous number of parasites reside within this hospitable niche that is characterized by low oxidative burst levels and a higher tolerance to antileishmanial drugs. Infected LT-HSC express a unique transcriptional signature, termed StemLeish. Cross-species analyses revealed significant overlap with human VL and HIV co-infected blood transcriptomes. Using siRNA silencing, knockdown of major upregulated StemLeish genes Rgs1, Cxcr4, Ell2, Vav1 and Twistnb confirmed their prominence in regulating LT-HSC infection. Cxcr4 proved important for the observed decrease in oxidative burst, suggesting a central position of this gene in shaping the LT-HSC niche, corroborated by in vivo therapeutic exploration. In LT-HSC, amastigotes rapidly enter quiescence, a phenotype with a higher capacity to survive antileishmanial treatment, providing an informal link to relapse. Parasites that transitioned through quiescence displayed an increased cellular infectivity and high sand fly transmission capacity. Transcriptional profiling revealed a novel set of markers and potential drivers of quiescence. Lastly, the impact of VL on B cell lymphopoiesis and humoral immunity was investigated. Corroborating the documented VL clinical manifestation of polyclonal hypergammaglobulinemia, infection was found to increase B cell progenitors in the BM, and all analyzed B cell subtypes of the spleen. Using immunization against a fluorescent heterologous antigen, it was shown that VL infection does not impair humoral immune memory. Surprisingly, the B cell interacts with Leishmania, carrying amastigotes, extracellularly attached while remaining in a resting, non-replicative state. Collectively, the results in this thesis deliver unprecedented insights regarding post-treatment relapse and humoral immunity during VL infection, providing novel drug targets and biomarkers for both host-directed and parasite-directed therapeutics using differential genes from quiescent amastigotes residing in a relapse-prone niche. This data will have a direct impact on drug screening efforts tweaked to overcome relapse.

Human African trypanosomiasis is a neglected tropical disease caused by the tsetse fly-transmitted unicellular protozoa Trypanosoma brucei gambiense and T. b. rhodesiense. Inoculation of the parasite into the skin results in infection of the host. The first stage of the disease is characterized by non-specific flu-like symptoms due to parasite multiplication in the lymphatics and the bloodstream. Parasite invasion of the central nervous system causes severe neurological symptoms. A wide range of other trypanosome species are responsible for disease in wild and domestic animals causing a widespread socioeconomic problem. The therapies that are currently available are limited in number and face disadvantages of toxicity and drug resistance. Despite decades of research, effective vaccines to protect humans and animals from infection are still lacking. In addition, asymptomatic individuals who remain undiagnosed and hence untreated can still contribute to disease transmission. The development of novel treatment strategies will therefore be crucial to achieve disease elimination. The first aim of this thesis was to evaluate and characterize novel anti-trypanosomal compounds. The medicinal chemistry group of the UGent developed a series of nucleoside analogues based on structural elements of the natural nucleoside antibiotics tubercidin and cordycepin, which were previously shown to have high antiparasitic potency but accompanied by extreme host toxicity. We identified 2 promising nucleoside analogues with high activity against Stage-II disease. Mechanistic evaluation using a whole-genome RNAi library identified TbAT1, ADKIN and 4EIP as involved in their mode-of-action. Evaluation of the adenosine analogues for the treatment of animal trypanosomiasis was able to identify compounds showing a broad activity profile across multiple parasite species/strains. The second aim was to characterize the early immunological events that occur upon the bite of an infected tsetse fly to gather the scientific knowledge necessary to develop novel intervention strategies based on the host-parasite interplay. Neutrophils were previously identified as being rapidly recruited to the site of infection, exhibiting a surprising pro-parasitic response. Detailed in vitro and in vivo analysis of the role of the neutrophil showed that neutrophils become activated upon parasitic stimulation but without a negative impact om parasite replication. In naturally infected mouse models, neutrophils play a role in parasite retention at the site of infection and impact on systemic organ colonization. Analysis of parasite burdens in various tissues identified the lungs as a site of strong parasite proliferation pointing to a potential role as parasitic tissue reservoir. Parasite presence resulted in a large influx of immune cells and change in the transcriptional profile, however, without detrimental effect on lung function. Depletion of specific immune cell subsets including eosinophils, NK cells and IgM+ B cells may underly the increased susceptibility of patients to opportunistic infections.