Bioecological aspects of Phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae) in endemic area for visceral leishmaniasis in São Paulo state

Author(s)

João Augusto Franco Leonel

• Document type: Master's Dissertation
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina Veterinária e Zootecnia (FMVZ/SBD)
• Defense date: 2019-12-13

Examining board members

Trícia Maria Ferreira de Sousa Oliveira; Claudio Casanova; Fredy Galvis Ovallos

• Advisor: Trícia Maria Ferreira de Sousa Oliveira

Abstract

Nowadays, we have observed important changes in the transmission patterns of tegumentary (TL) and visceral leishmaniases (VL) on endemic areas of Brazil. In addition, the VL advances rapidly to new areas on the past considered indene. The Phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae) are small midges with great relevance in medical and veterinary sciences, as they are vectors of viruses, Bartonella bacilliformis and mainly Leishmania spp. The studies on Phlebotomine sandflies are fundamental to improve the comprehension of complex leishmaniasis epidemiology, revealing biological aspects of these vectors and its relationships with the parasite and its reservoirs. Molecular approaches have been included in these studies and have proven to be powerful tools for research about these dipterans. The aim of this work was to investigate the bioecological aspects of Phlebotomine sandflies on a TL and VL endemic area of São Paulo state, Brazil. Thus, an entomological survey was conducted to describe the fauna, its seasonality, rate of positive PCR by Leishmania spp. DNA and blood meals of sandflies in the study area. The insects were screened, identified and the thorax and abdomen of the females sampled submitted to DNA extraction and then PCR for the detection of Kinetoplast DNA (kDNA) of Leishmania spp. parasites. Next, DNA samples from engorged, partially engorged and gravid females were submitted to PCR amplification of the mitochondrial cytochrome B gene (CYT-B) of vertebrates. Amplified products in CYT-B reactions were purified and subjected to sequencing. The sequences found were compared in a database GenBank, to identify the vertebrate species involved in the blood meal of the female sandflies. By employing molecular tools, this work also analyzed the performance of some DNA extraction methods from individual samples of thorax and abdomen of females of these insects. In our study, twelve species of sandflies were found, among proven and suspected vectors in the TL and VL epidemiological cycles. No seasonal patterns were observed between climatic factors studied (temperature, relative humidity and rainfall) and the fauna of these insects, captured with CDC light traps. Regarding DNA extraction methods, commercial kits are a good option for this type of sample. However, in house methods based on phenol-chloroform/isoamyl alcohol or NaCl/alcohol showed better results. Molecular analyzes revealed one non-engorged Ev. (Ald.) carmelinoi female PCR positive by Leishmania spp. kDNA. Blood meal identification showed swine, humans, dogs, cattle, chickens and opossums as vertebrate hosts of sandflies in the study area. The implications of these findings are discussed in two scientific articles submitted for evaluation in journals of the area. (AU)

• FAPESP's process: 17/23436-0 - Hosts of Lutzomyia longipalpis in endemic area for visceral leishmaniasis
• Grantee: João Augusto Franco Leonel
• Support Opportunities: Scholarships in Brazil - Master