Phlebotomine Sandfly (Diptera: Psychodidae) Fauna and The Association Between Climatic Variables and The Abundance of Lutzomyia longipalpis sensu lato in an Intense Transmission Area for Visceral Leishmaniasis in Central Western Brazil.

The presence, abundance, and distribution of sandflies are strongly influenced by climate and environmental changes. This study aimed to describe the sandfly fauna in an intense transmission area for visceral leishmaniasis and to evaluate the association between the abundance of Lutzomyia longipalpis sensu lato (Lutz & Neiva 1912) (Diptera: Psychodidae) and climatic variables. Captures were carried out 2 yr (July 2017 to June 2019) with automatic light traps in 16 sites of the urban area of Campo Grande, Mato Grosso do Sul state. The temperature (°C), relative humidity (%), precipitation (mm3), and wind speed (km/h) were obtained by a public domain database. The Wilcoxon test compared the absolute frequencies of the species by sex. The association between climatic variables and the absolute frequency of Lu. longipalpis s.l. was assessed using the Spearman's correlation coefficient. A total of 1,572 sandflies into four species were captured. Lutzomyia longipalpis s.l. was the most abundant species and presented a significant correlation with the average temperature, humidity, and wind speed in different periods. Lutzomyia longipalpis s.l. was captured in all months, showing its plasticity in diverse weather conditions. We emphasize the importance of regular monitoring of vectors and human and canine cases, providing data for surveillance and control actions to continue to be carried out in the municipality.

Standardization of DNA extraction from sand flies: Application to genotyping by next generation sequencing.

Standardization of the methods for extraction of DNA from sand flies is essential for obtaining high efficiency during subsequent molecular analyses, such as the new sequencing methods. Information obtained using these methods may contribute substantially to taxonomic, evolutionary, and eco-epidemiological studies. The aim of the present study was to standardize and compare two methods for the extraction of genomic DNA from sand flies for obtaining DNA in sufficient quantities for next-generation sequencing. Sand flies were collected from the municipalities of Campo Grande, Camapuã, Corumbá and Miranda, state of Mato Grosso do Sul, Brazil. Three protocols using a silica column-based commercial kit (ReliaPrep™ Blood gDNA Miniprep System kit, Promega®), and three protocols based on the classical phenol-chloroform extraction method (Uliana et al., 1991), were compared with respect to the yield and quality of the extracted DNA. DNA was quantified using a Qubit 2.0 fluorometer. The presence of sand fly DNA was confirmed by PCR amplification of the IVS6 region (constitutive gene), followed by electrophoresis on a 1.5% agarose gel. A total of 144 male specimens were analyzed, 72 per method. Significant differences were observed between the two methods tested. Protocols 2 and 3 of phenol-chloroform extraction presented significantly better performance than all commercial kit extraction protocols tested. For phenol-chloroform extraction, protocol 3 presented significantly better performance than protocols 1 and 2. The IVS6 region was detected in 70 of 72 (97.22%) samples extracted with phenol, including all samples for protocols 2 and 3. This is the first study on the standardization of methods for the extraction of DNA from sand flies for application to next-generation sequencing, which is a promising tool for entomological and molecular studies of sand flies.