Rv treatment downregulates apoptotic pathway.

<p>Adult mosquitoes were reared until six days old under each dietary condition. The midguts of female mosquitoes were dissected and homogenized in TRIzol, and total RNA was extracted. These samples were used to perform qPCR for the genes: (<b>A</b>) 16S, (<b>B</b>) ARGONAUTE 2, (<b>C</b>) CASPASE 16, (<b>D</b>) AeDRONC, (<b>E</b>) AeIAP1 and (<b>F</b>) ATG8. Data show means and standard error of at least three independent experiments. Ctrl—0.05% ethanol plus 10% sucrose; Rv—100 μM Rv in 0.05% ethanol plus 10% sucrose; AbMix– 10% sucrose, 10 U/mL penicillin, 10 U/mL streptomycin and 15 U/mL gentamicin. *—p<0.05; **—p<0.01; ***—p<0.001; ****—p<0.0001, as calculated by Student’s t-test (F as calculated by one-way ANOVA with Tukey post test). (Error bars, s.e.m., n = 3 experiments).</p

Summary data of the effects of polyphenols on mosquito lifespan.

<p>Summary data of the effects of polyphenols on mosquito lifespan.</p

Rv triggers autophagy in mosquito midgut.

<p><b>(A, B</b>) Typical ultrastructural appearance of mosquito midgut fed on a control diet. (<b>C-F</b>) Midguts from insects fed on Rv. White stars indicate the formation of concentric membrane structures. Black arrows indicate preserved mitochondrial morphology and normal cristae aspects. N: nucleus; M: mitochondria; Mi: microvilli. Bars in panels A and C: 2 μm. Bars in panels B, D-F: 0.5 μm. (<b>G</b>) Midgut images obtained under a fluorescence microscope after incubation with LysoTracker Red (upper panels, DIC) (lower panels, fluorescence). (<b>H</b>) Densitometry of LysoTracker fluorescence images shown in panel G. (<b>I)</b> Images obtained as in panel G following the silencing of mosquito AMPK (DsAMPK) or an unrelated protein (DsMal). Insects were kept on the indicated Control (Ctrl), Rv or AICAR diets. <b>(J</b>) Densitometry of LysoTracker fluorescence images shown in the lower part of panels I. Rv—100 μM Rv; AICAR—1 mM AICAR. **—p<0.01; ****—p<0.0001 as calculated by one-way ANOVA with Tukey post test (I) and by Student’s t-test (J). Data in panels I, J represent means and s. e. m. (n = 3).</p

Polyphenols increase mosquito lifespan while still allowing dengue virus infection.

<p>(<b>A-D</b>) Effect of polyphenols on male (<b>A, C</b>) and female (<b>B, D</b>) lifespan. The 50% mean lifespan is indicated by the horizontal dotted line. (<b>E</b>) Dengue virus RNA from mosquitoes fed or not before infection with Rv. (<b>F</b>) Percentage of dengue-infected mosquitoes. Ctrl—Control, Epi—100 μM epi-gallo-catechin-gallate; Gen—100 μM genistein; Que—100 μM quercetin. (Error bars,s.e.m., n = 3 experiments with 200 mosquitoes in each cage, p<0.05 in each survivor curve compared with control as calculated by Mantel-Cox and Gehan-Breslow-Wilcoxon test).</p

Dietary polyphenols decrease lipid accumulation in mosquitoes through AMPK activation.

<p>(<b>A</b>) Densitometric analysis of triacylglycerol content measured by TLC. (<b>B</b>) Densitometric analysis of western blots for midgut p-AMPK. (<b>C</b>) Densitometric analysis of western blots for fat body p-AMPK. (<b>D</b>) Densitometric analysis of triacylglycerol levels as measured by TLC in mosquitoes fed as indicated. Actin was used as loading control in panels B and C. (HepG2) HepG2 cells are a positive control for pAMPK antibody. Insets show representative images of either TLCs or blots. Ctrl—control; Rv—100 μM Rv; Epi—100 μM epi-gallo-catechin-gallate; Gen—100 μM genistein; Que—100 μM quercetin; CC—100 μM Compound C; CC+Rv—100 μM Compound C and 100 μM Rv; AICAR—1 mM AICAR. *—p<0.05; **—p<0.01, as calculated by one-way ANOVA with Tukey post test. (Error bars, s.e.m., n = 3 experiments).</p