Cluster organization. The symbol (*, asterisk) indicates the miRNA cloned in this study. Exons are symbolized using broken boxes. Position of the mature miRNA within its stem-loop precursor is indicated using a darker line. Additional information is gathered in Supplementary Data (Table S2).

Expression profiling of Anopheles miRNAs across gender and body parts. (A) Representative examples of the autoradiographies of primer-extension assays performed on total RNA samples from Anopheles. (B) Profiling of the miRNA expression in Anopheles body parts using primer-extension assays. The intensities of the reverse transcriptase stops were quantified (see Materials and Methods section). The stop intensities obtained using total RNAs extracted from the female gut was set as the internal reference. F is used for female and M for male. As stated in Materials and Methods section, ratios are required to be outside of the estimated confidence interval [0.67; 1.33] to be deemed as significant.

Incidence of regular and infectious blood feeding on miRNA expression profiles. The intensity of the reverse transcriptase stops was quantified (see Materials and Methods section). (A) Effect of blood feeding on miRNA expression in leftovers samples. The female leftovers signal was set as the internal reference. (B) Effect of P. berghei presence on miRNA expression in leftovers samples. The blood-fed female leftovers signal was set as the internal reference. (C) Effect of blood feeding on miRNA expression in midguts samples. The female midguts signal was set as the internal reference. (D) Effect of P. berghei presence on miRNA expression in midguts samples. The blood-fed female midguts signal was set as the internal reference. (E) Representative examples of autoradiographies of the primer-extension assays. Total RNAs were extracted from leftovers or from midguts, 24–48 h after blood feeding on a regular mouse or after blood feeding (BF) on P. berghei-infected mouse.

Survival of P. berghei in drosha, Dicer1 and Argonaute1 depleted mosquitoes. Females were infected 4 days after injection, and midguts were dissected 10 days later and fixed. (A) Representative composite image between the green fluorescent channel and a differential interference contrast (DIC) image of dissected midguts showing parasite development in the dsRNA-injected mosquitoes. (B–D) Survival of P. berghei parasites in control ds-lacZ and in ds-drosha-, ds-Dicer1- or ds-Ago1-injected mosquitoes. Data were collected from three independent experiments. The mean of parasite number observed in each experiment was similar so that the data were pooled. Statistically significant differences between samples were evaluated by Mann–Whitney test and P-values are indicated on the graph. Black bars represent mean parasite numbers per group. Here, n represents the number of guts, m the average number of parasites per gut and me the median number of parasites per gut.