Abstract
Phlebotomine sand flies (Diptera, Psychodidae) belonging to the Lutzomyia genus transmit zoonoses in the New World. Lutzomyia longipalpis is the main vector of Leishmania infantum, which is the causative agent of visceral leishmaniasis in Brazil. To identify key molecular aspects involved in the interaction between vector and pathogens and contribute to developing disease transmission controls, we investigated the sand fly innate immunity mediated by the Janus kinase/signal transducer and activator of transcription (Jak-STAT) pathway in response to L. infantum infection. We used two study models: L. longipalpis LL5 embryonic cells co-cultured with L. infantum and sand fly females artificially infected with the parasite. We used qPCR to follow the L. longipalpis gene expression of molecules involved in the Jak-STAT pathway. Also, we modulated the Jak-STAT mediated immune response to understand its role in Leishmania parasite infection. For that, we used RNAi to silence the pathway regulators, protein inhibitor of activated STATs (PIAS) in LL5 cells, and STAT in adult females. In addition, the pathway suppression effect on parasite development within the vector was assessed by light microscopy in late-phase infection. The silencing of the repressor PIAS in LL5 cells led to a moderate increase in a protein tyrosine phosphatase 61F (PTP61F) expression. It suggests a compensatory regulation between these two repressors. L. infantum co-culture with LL5 cells upregulated repressors PIAS, suppressor of cytokine signaling (SOCS), and PTP61F. It also downmodulated virus-induced RNA-1 (VIR-1), a pathway effector, indicating that the parasite could repress the Jak-STAT pathway in LL5 cells. In Leishmania-infected L. longipalpis females, STAT and the antimicrobial peptide attacin were downregulated on the third day post-infection, suggesting a correlation that favors the parasite survival at the end of blood digestion in the sand fly. The antibiotic treatment of infected females showed that the reduction of gut bacteria had little effect on the Jak-STAT pathway regulation. STAT gene silencing mediated by RNAi reduced the expression of inducible nitric oxide synthase (iNOS) and favored Leishmania growth in sand flies on the first day post-infection. These results indicate that STAT participated in the iNOS regulation with subsequent effect on parasite survival.
Highlights
Phlebotomine sand flies (Diptera, Psychodidae) are vectors of a group of parasitic diseases classified as cutaneous or visceral leishmaniases, which are transmitted by the bite of an infected female sand fly
We have previously identified L. longipalpis components of the sand fly innate immunity regulated by the Toll and immune deficiency (IMD) pathways that produce antimicrobial peptides (AMPs) such as attacin, cecropin, and defensins [6,7,8]
The attacin expression upregulation may be caused by another pathway such as Toll or IMD as indicated in our previous studies [6, 41] posing an additional microbial control. These results indicate that the L. longipalpis putative STAT is associated with the inducible nitric oxide synthase (iNOS) expression, while attacin and virusinduced RNA-1 (VIR-1) had no direct correlation with STAT silencing in sucrose-fed females
Summary
Phlebotomine sand flies (Diptera, Psychodidae) are vectors of a group of parasitic diseases classified as cutaneous or visceral leishmaniases, which are transmitted by the bite of an infected female sand fly. These diseases are caused by protozoan parasites belonging to the Leishmania genus, with over 20 species identified as infective to humans and endemic to 98 countries and territories distributed from temperate to tropical regions of the globe [1]. Lutzomyia is the most important genus in the American continent due to its wide distribution and diversity of species (reviewed in 2) This genus includes Lutzomyia longipalpis, the main vector of visceral leishmaniasis in Brazil, caused by Leishmania infantum We are interested in understanding how the L. longipalpis immune system responds to parasitic challenges, contributing to developing novel molecular-based tools to control the Leishmania cycle in the insect
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