Leishmania development in their vectors is closely connected with bloodmeal digestion. This thesis focuses on factors affecting bloodmeal digestion, egg development and Leishmania infection within the sand fly gut. First, we compared the effect of mammalian (rabbit) and avian (chicken) blood on digestion and eggs development in Phlebotomus duboscqi. Sand flies fed on chickens had twice lower protein concentrations in the midgut and significantly lower trypsin activity compared to those fed on rabbits. The highest differences in the trypsin activity were observed during first 24 hours post bloodmeal. In addition, females fed on chickens had slower eggs development and their eggs were 10 % smaller compared to those fed on rabbits. In the second part of the thesis we tested the effect of mosquito hormone TMOF on the trypsin activity and eggs development of Lutzomyia longipalpis. Rabbit blood with TMOF (28 mg/ml) was presented to the females via a membrane feeding system. Sand flies fed on blood with TMOF had 15 - 35 % less trypsin activity than control females fed on only rabbit blood. In addition, females fed on blood with TMOF had developed 30 % less eggs and their eggs were 12 - 24 % smaller compared to control group. However the effect of TMOF we observed was lower than that described previously...

Sand flies serve as the vectors of leishmaniasis and their saliva was shown to affect the outcome of Leishmania infection by immunomodulation of the host. On the other hand, sand fly saliva contains a large scale of farmacologically active proteins that are strongly immunogenous for bitten hosts and specific anti-saliva immunity initiated by repeated sand fly feeding provides protection against Leishmania infection. Specific cell-mediated immunity was shown to be the core of the protectivity; however, our data suggests that the protective immunity has certain limitations. In mice bitten by sand flies for prolonged periods, we observed the desenzitization in term of abrogation of the protective immunity. Thus, we can speculate that the protective effect of immunity is linked solely with the short-term exposure. Nevertheless, our experiments showed that this aspect is also conditioned by the immediate infection after the protective short-term immunization. Taken together, it seems that these limitations may explain the circulation of leishmaniasis in endemic areas, even though humans and animals are frequently immunized by bites of uninfected sand flies. Repeated sand fly feeding on various hosts also promotes production of anti-saliva antibodies that reflect the intensity of exposure. We...

anglický Phlebotomine sand flies (Diptera: Psychodidae) are the only proven vectors of Leishmania parasites (Kinetoplastida: Trypanosomatidae). In Nematoceran Diptera, including sand flies, adults produce a type 1 peritrophic matrix (PM) which is secreted in response to the distension of the midgut caused by blood meal. The PM is an acellular envelope composed of chitin fibres and proteins, which protects the midgut epithelium against abrasion and pathogens and improves digestion. In hematophagous insects, the PM also plays a central role in heme detoxification. Female sand flies acquire Leishmania with a bloodmeal and the parasites undergo complicated development in their gut finished by the colonization of the stomodeal valve. The PM is one of the most important barriers in Leishmania development and its role in the vector competence of the S. schwetzi is the main topic of this master thesis. The PM's kinetic and morfology in the S. schwetzi is compared with other three sand fly species which differ in susceptibility to L. donovani. The key role of the PM in S. schwetzi vector competence is finally proved by disrupting the PM using the exogenous chitinase from Beauveria bassiana. Under these artificial conditions, the Leishmania parasites (L. donovani and L. major) are able to exit the PM,...

This thesis focuses on the development of protozoan parasites from Leishmania donovani complex in their insect vectors and summarizes results of five parts of the project I participated in during my Ph.D. studies. Sand flies of genera Phlebotomus and Lutzomyia are the only proven vectors of leishmaniasis, however, the role of alternative vectors, like ticks, fleas and biting midges is frequently discussed in the literature. In this work, we showed that Eurasian species of biting midge Culicoides nubeculosus does not support late stage infections of L. major and L. infantum. We also demonstrated that microscopical observation of Leishmania promastigotes in the digestive tract of bloodfeeding arthropods remains a crucial method for any conclusion about the vector competence of the suspected insect. In the second part of our study were compared the life-cycle parameters and vector competence of two Ethiopian P. orientalis colonies for L. donovani. Marked differences between colonies were found in life-cycle parametes, however, molecular analyses did not reveal any genetic differences. Experimental infections showed that both P. orientalis colonies are very susceptible to L. donovani infection and even the lowest infective dose tested (2 × 103 promastigotes/ml; corresponding to 1-2 promastigotes) was...

Phlebotomine sand flies (Diptera, Psychodidae) are small blood sucking insects which are only proven vectors of neglected tropical disease called leishmaniasis. Sand flies of the genus Phlebotomus act as vectors in the Old World and those of genus Lutzomyia are vectors in the New World. However, not all of them are capable of transmitting the disease. It is therefore crutial to conclusively determinate the species and have up-to-date knowledge of their natural occurence. Routine identification based on morphological characters is challenging due to intraspecific variability of these or their possible damage during the capture and preparation. In adition, correct assessment of distinctive cahracters, especially for females, is difficult and requires certain expertise. Thus, approaches of molecular taxonomy have been recently increasingly used for sand flies species determination. This thesis presents usage of DNA sequencing and MALDI-TOF protein profiling for routine determination of sand flies caught in southeastern Europe and adjacent regions, where we have lack of information of present sand flies species. Another part of this thesis focus on closer examination of relations between selected closely related species within species complexes. Key words: Phlebotomus, molacular identification, DNA...