Leishmania amazonensis DNA in wild females ofLutzomyia cruzi (Diptera: Psychodidae) in the state of Mato Grosso do Sul, Brazil

Oliveira, Everton Falcão de; Casaril, Aline Etelvina; Mateus, Nathália Lopes Fontoura; Murat, Paula Guerra; Fernandes, Wagner; Oshiro, Elisa Teruya; Oliveira, Alessandra Gutierrez de; Galati, Eunice Aparecida Bianchi

doi:10.1590/0074-02760150317

Cited by 13 publications

(10 citation statements)

References 46 publications

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“…An ideal DNA extraction protocol should meet some criteria such as concentrated yield, elimination of PCR inhibitors, implementable as laboratory technique, low cost, reproducibility, and safety (Rantakokko-Javala and Javala, 2002). Researchers have used different extraction methods to extract insect DNA, such as phenolchloroform-isoamyl alcohol, detergents like Triton X-100 and Triton X-100/DDT, silica column solid phase extraction, and Chelex ® Molecular Biology Grade Resin (Bio-Rad Laboratories, Hercules, USA) (Cova et al, 2015;Oliveira et al, 2015;Paiva et al, 2007;Pinto et al, 2015;Santos et al, 2013).…”

Section: Discussionmentioning

confidence: 99%

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

Casaril

Oliveira

Alonso

et al. 2017

Experimental Parasitology

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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.

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Section: Discussionmentioning

confidence: 99%

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

Casaril

Oliveira

Alonso

et al. 2017

Experimental Parasitology

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“…amazonensis DNA was detected in wild-caught females of Lu . cruzi during project execution [ 28 ]. We therefore decided to include this parasite in the study.…”

Section: Discussionmentioning

confidence: 99%

“…The contents of the slide and the digestive tube were ground using a plastic pestle in 1.5-mL tubes with 300 μL of 5% Chelex resin solution (Bio-Rad, Hercules, CA, USA). DNA extraction was performed according to the protocol described by Oliveira [ 28 ].…”

Section: Methodsmentioning

confidence: 99%

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Experimental infection and transmission of Leishmania by Lutzomyia cruzi (Diptera: Psychodidae): Aspects of the ecology of parasite-vector interactions

et al. 2017

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Several parameters should be addressed before incriminating a vector for Leishmania transmission. Those may include its ability to become infected by the same Leishmania species found in humans, the degree of attractiveness for reservoirs and humans and capacity to sustain parasite infection under laboratory conditions. This study evaluated the vectorial capacity of Lutzomyia cruzi for Leishmania infantum and gathered information on its ability to harbor L. amazonensis. Laboratory-reared Lu. cruzi were infected experimentally by feeding them on dogs infected naturally with L. infantum and hamsters infected with L. amazonensis. Sand fly attractiveness to dogs and humans was determined using wild caught insects. The expected daily survival of infected Lu. cruzi, the duration of the gonotrophic cycle, and the extrinsic incubation period were also investigated for both parasites. Vector competence was investigated for both Leishmania species. The mean proportion of female sand flies that fed on hosts was 0.40. For L. infantum and L. amazonensis, Lu. cruzi had experimental infection rates of 10.55% and 41.56%, respectively. The extrinsic incubation period was 3 days for both Leishmania species, regardless of the host. Survival expectancy of females infected with L. infantum and L. amazonensis after completing the gonotrophic cycle was 1.32 and 0.43, respectively. There was no association between L. infantum infection and sand fly longevity, but L. amazonensis–infected flies had significantly greater survival probabilities. Furthermore, egg-laying was significantly detrimental to survival. Lu. cruzi was found to be highly attracted to both dogs and humans. After a bloodmeal on experimentally infected hosts, both parasites were able to survive and develop late-stage infections in Lu. cruzi. However, transmission was demonstrated only for L. amazonensis–infected sand flies. In conclusion, Lu. cruzi fulfilled several of the requirements of vectorial capacity for L. infantum transmission. Moreover, it was also permissive to L. amazonensis.

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“…Thereafter, collected females that did not have blood in their gut were dissected, and identified on the basis of their spermatheca characteristics. These females were then placed in 1.5-mL microtubes with isopropyl alcohol to determine the presence of Leishmania DNA, the results of which are described by Oliveira et al [21]. …”

Section: Methodsmentioning

confidence: 99%

Monthly Distribution of Phlebotomine Sand Flies, and Biotic and Abiotic Factors Related to Their Abundance, in an Urban Area to Which Visceral Leishmaniasis Is Endemic in Corumbá, Brazil

et al. 2016

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The monthly distribution and abundance of sand flies are influenced by both biotic and abiotic factors. The present study aimed to evaluate the seasonal distribution of sand flies and the relation between their abundance and environmental parameters, including vegetation and climate. This study was conducted over a 2-year period (April 2012 to March 2014). Monthly distribution was evaluated through the weekly deployment of CDC light traps in the peridomicile area of 5 residences in an urban area of the municipality of Corumbá in the State of Mato Grosso do Sul, Brazil. Meteorological data were obtained from the Mato Grosso do Sul Center for Weather, Climate, and Water Resources. The spectral indices were calculated based on spatial resolution images (GeoEye) and the percentage of vegetal coverage. Differences in the abundance of sand flies among the collection sites were assessed using the Kruskal-Wallis test, and the strength of correlations between environmental variables was determined by calculating Spearman’s correlation coefficients. Lutzomyia cruzi, Lu. forattinii, and Evandromyia corumbaensis were the most frequently found species. Although no significant association was found among these sand fly species and the tested environmental variables (vegetation and climate), high population peaks were found during the rainy season, whereas low peaks were observed in the dry season. The monthly distribution of sand flies was primarily determined by Lu. cruzi, which accounted for 93.94% of the specimens collected each month throughout the experimental period. The fact that sand flies were detected year-round indicates a continuous risk of infection to humans, demonstrating the need for targeted management and education programs.

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Leishmania amazonensis DNA in wild females ofLutzomyia cruzi (Diptera: Psychodidae) in the state of Mato Grosso do Sul, Brazil

Cited by 13 publications

References 46 publications

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

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

Experimental infection and transmission of Leishmania by Lutzomyia cruzi (Diptera: Psychodidae): Aspects of the ecology of parasite-vector interactions

Monthly Distribution of Phlebotomine Sand Flies, and Biotic and Abiotic Factors Related to Their Abundance, in an Urban Area to Which Visceral Leishmaniasis Is Endemic in Corumbá, Brazil

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