Background The olive fruit fly (Bactrocera oleae) is the most destructive pest of the olive cultivation worldwide causing significant production losses and olive fruit impoverishment, as its larvae feed exclusively on the olive fruit. Reproductive and sexual behavior, as well as host-plant recognition of the fly, are highly dependent on its chemosensory system. Therefore, exploring the role of genes that play a critical role in olfaction, could reveal potential molecular targets that determine species-specific features on chemical communication and could be used to impair sexual behavior. Results In this study we identified the gene that encodes the conserved olfactory co-receptor Orco (Odorant receptor co-receptor), which interacts with all divergent insect odorant receptors, and investigated how disruption of its expression affects chemoreception. We initially searched the expression profile of Bo-Orco in both sexes during sexual maturation, as well as pre- and post-mating communication by relative quantitative real time PCR (qRT-PCR) analysis suggesting that Bo-Orco was abundantly expressed in sexually mature adults. We further investigated the functional role of Bo-Orco in mating and oviposition behavior via transient gene silencing that was performed through in vivo dsRNA hemolymph injections in sexually mature flies 7 days after eclosion. Orco-knockdown phenotypes in both sexes showed reduced copulation rates in mating competitiveness tests, possibly through impaired olfactory-mediated detection of sex pheromone. In addition, oviposition was significantly inhibited in dsRNA-Orco injected females in a post-mating behavior test. Conclusions Our results demonstrate that Orco plays a crucial role in the reproductive behavior of the olive fruit fly, since pre- and post-mating processes were affected. This is the first report in the olive fruit fly that links the chemosensory pathway with the mating behavior and the reproductive potential at a molecular basis, rendering this gene a potential target for the improvement of the olive fruit fly population control techniques.

New biomarkers have to be developed in order to increase the performance of current antigen-based malaria rapid diagnosis. Antibody production often involves the use of laboratory animals and is time-consuming and costly, especially when the target is Plasmodium, whose variable antigen expression complicates the development of long-lived biomarkers. To circumvent these obstacles, we have applied the Systematic Evolution of Ligands by EXponential enrichment method to the rapid identification of DNA aptamers against Plasmodium falciparum-infected red blood cells (pRBCs). Five 70 b-long ssDNA sequences, and their shorter forms without the flanking PCR primer-binding regions, have been identified having a highly specific binding of pRBCs versus non-infected erythrocytes. Structural analysis revealed G-enriched sequences compatible with the formation of G-quadruplexes. The selected aptamers recognized intracellular epitopes with apparent Kds in the μM range in both fixed and non-fixed saponin-permeabilized pRBCs, improving >30-fold the pRBC detection in comparison with aptamers raised against Plasmodium lactate dehydrogenase, the gold standard antigen for current malaria diagnostic tests. In thin blood smears of clinical samples the aptamers reported in this work specifically bound all P. falciparum stages versus non-infected erythrocytes, and also detected early and late stages of the human malaria parasites Plasmodium vivax, Plasmodium ovale and Plasmodium malariae. The results are discussed in the context of their potential application in future malaria diagnostic devices.

Long non-coding RNA (lncRNA) research has emerged as an independent scientific field in recent years. Despite their association with critical cellular and metabolic processes in plenty of organisms, lncRNAs are still a largely unexplored area in mosquito research. We propose that they could serve as exceptional tools for pest management due to unique features they possess. These include low inter-species sequence conservation and high tissue specificity. In the present study, we investigated the role of ovary-specific lncRNAs in the reproductive ability of the Asian tiger mosquito, Aedes albopictus. Through the analysis of transcriptomic data, we identified several lncRNAs that were differentially expressed upon blood feeding; we called these genes Norma (NOn-coding RNA in Mosquito ovAries). We observed that silencing some of these Normas resulted in significant impact on mosquito fecundity and fertility. We further focused on Norma3 whose silencing resulted in 43% oviposition reduction, in smaller ovaries and 53% hatching reduction of the laid eggs, compared to anti-GFP controls. Moreover, a significant downregulation of 2 mucins withing a neighboring (∼100 Kb) mucin cluster was observed in smaller anti-Norma3 ovaries, indicating a potential mechanism of in-cis regulation between Norma3 and the mucins. Our work constitutes the first experimental proof-of-evidence connecting lncRNAs with mosquito reproduction and opens a novel path for pest management.

Long non-coding RNAs (lncRNAs) play critical regulatory roles in various cellular and metabolic processes in mosquitoes and all other organisms studied thus far. In particular, their involvement in essential processes such as reproduction makes them potential targets for the development of novel pest control approaches. However, their function in mosquito biology remains largely unexplored. To elucidate the role of lncRNAs in mosquitoes’ reproduction and vector competence for arboviruses, we have implemented a computational and experimental pipeline to mine, screen, and characterize lncRNAs related to these two biological processes. Through analysis of publicly available Zika virus (ZIKV) infection-regulated Aedes aegypti transcriptomes, at least six lncRNAs were identified as being significantly upregulated in response to infection in various mosquito tissues. The roles of these ZIKV-regulated lncRNAs (designated Zinc1, Zinc2, Zinc3, Zinc9, Zinc10 and Zinc22), were further investigated by dsRNA-mediated silencing studies. Our results show that silencing of Zinc1, Zinc2, and Zinc22 renders mosquitoes significantly less permissive to ZIKV infection, while silencing of Zinc22 also reduces fecundity, indicating a potential role for Zinc22 in trade-offs between vector competence and reproduction. We also found that silencing of Zinc9 significantly increases fecundity but has no effect on ZIKV infection, suggesting that Zinc9 may be a negative regulator of oviposition. Our work demonstrates that some lncRNAs play host factor roles by facilitating viral infection in mosquitoes. We also show that lncRNAs can influence both mosquito reproduction and permissiveness to virus infection, two biological systems with important roles in mosquito vectorial capacity.