Verification and monitoring of visceral leishmaniasis in hamsters caused by Leishmania infantum, using non-invasive approaches involving ultrasound imaging and blood gases

摘要

Hamsters are a suitable experimental model for visceral leishmaniasis (VL) because they mimic the features of the human disease. However, the infection after inoculation can only be verified after sacrifice of the animal or several months following infection, when obvious signs of the disease appear, compromising animal welfare in both cases. Unlike other studies, the present work used an inoculum of 5 x 10(8) promastigotes to induce Leishmania infantum infection, which are easier to produce than amastigotes, in in vitro culture. The infection in hamsters was detected using non-invasive methods such as ultrasound imaging (USG) and blood gases, in addition to alterations in hematological parameters and weight loss. USG imaging identified changes in the size and echogenicity of the spleen, liver, and kidney as early as week 9 (W9) after experimental inoculation. However, blood gases, specially lactate, was increased in response to the infection, with statistically significant differences between W9 and WO (before infection) (p 0.0001). The conventional hematological parameters showed progressive pancytopenia and weight loss of 15% and 10% in infected males and females respectively, at W9 versus WO (p 0.0001). Histological changes in the liver, kidney, and spleen correlated with changes detected by USG imaging and the number of parasites increased proportionately to the progression of infection, being higher at W24. In conclusion, USG imaging, lactate levels, hematocrit and hemoglobin parameters, along with weight loss allowed early detection of infection, which was then confirmed by the identification and quantification of parasites in the blood, liver, and spleen by qRT-PCR. In contrast, blood chemistry was not a useful tool in the early detection of VL infection because it did not correlate with alterations evident in other techniques. The use of non-invasive tools eliminates the need for animal sacrifice to confirm infection, thus reducing the number of animals required for a given study and eliminating the need to wait until the appearance of severe signs of infection, which affect animal welfare. These tools are therefore advantageous for use in preclinical studies, for studying pathogenesis as also for vaccine and drug development.