Visual threats reduce blood-feeding and trigger escape responses in Aedes aegypti mosquitoes.

The diurnal mosquitoes Aedes aegypti are vectors of several arboviruses, including dengue, yellow fever, and Zika viruses. To find a host to feed on, they rely on the sophisticated integration of olfactory, visual, thermal, and gustatory cues reluctantly emitted by the hosts. If detected by their target, this latter may display defensive behaviors that mosquitoes need to be able to detect and escape. In humans, a typical response is a swat of the hand, which generates both mechanical and visual perturbations aimed at a mosquito. While the neuro-sensory mechanisms underlying the approach to the host have been the focus of numerous studies, the cues used by mosquitoes to detect and identify a potential threat remain largely understudied. In particular, the role of vision in mediating mosquitoes' ability to escape defensive hosts has yet to be analyzed. Here, we used programmable visual displays to generate expanding objects sharing characteristics with the visual component of an approaching hand and quantified the behavioral response of female mosquitoes. Results show that Ae. aegypti is capable of using visual information to decide whether to feed on an artificial host mimic. Stimulations delivered in a LED flight arena further reveal that landed females Ae. aegypti display a stereotypical escape strategy by taking off at an angle that is a function of the distance and direction of stimulus introduction. Altogether, this study demonstrates mosquitoes can use isolated visual cues to detect and avoid a potential threat.

Blood meal analysis of Anopheles vectors of simian malaria based on laboratory and field studies

Blood feeding and host-seeking behaviors of a mosquito play an imperative role in determining its vectorial capacity in transmitting pathogens. Unfortunately, limited information is available regarding blood feeding behavior of Anopheles species in Malaysia. Collection of resting Anopheles mosquitoes for blood meal analysis poses a great challenge especially for forest dwelling mosquitoes. Therefore, a laboratory-based study was conducted to evaluate the potential use of mosquitoes caught using human landing catch (HLC) for blood meal analysis, and subsequently to document blood feeding behavior of local Anopheles mosquitoes in Peninsular Malaysia. The laboratory-based experiment from this study revealed that mosquitoes caught using HLC had the potential to be used for blood meal analysis. Besides HLC, mosquitoes were also collected using manual aspirator and Mosquito Magnet. Overall, 47.4% of 321 field-caught Anopheles mosquitoes belonging to six species were positive for vertebrate host DNA in their blood meal. The most frequent blood meal source was human (45.9%) followed by wild boar (27.4%), dog (15.3%) and monkey (7.5%). Interestingly, only Anopheles cracens and Anopheles introlatus (Leucosphyrus Group) fed on monkey. This study further confirmed that members of the Leucosphyrus Group are the predominant vectors for knowlesi malaria transmission in Peninsular Malaysia mainly due to their simio-anthropophagic feeding behavior.

Dengue virus infection modifies mosquito blood-feeding behavior to increase transmission to the host

Mosquito blood-feeding behavior is a key determinant of the epidemiology of dengue viruses (DENV), the most-prevalent mosquito-borne viruses. However, despite its importance, how DENV infection influences mosquito blood-feeding and, consequently, transmission remains unclear. Here, we developed a high-resolution, video-based assay to observe the blood-feeding behavior of Aedes aegypti mosquitoes on mice. We then applied multivariate analysis on the high-throughput, unbiased data generated from the assay to ordinate behavioral parameters into complex behaviors. We showed that DENV infection increases mosquito attraction to the host and hinders its biting efficiency, the latter resulting in the infected mosquitoes biting more to reach similar blood repletion as uninfected mosquitoes. To examine how increased biting influences DENV transmission to the host, we established an in vivo transmission model with immuno-competent mice and demonstrated that successive short probes result in multiple transmissions. Finally, to determine how DENV-induced alterations of host-seeking and biting behaviors influence dengue epidemiology, we integrated the behavioral data within a mathematical model. We calculated that the number of infected hosts per infected mosquito, as determined by the reproduction rate, tripled when mosquito behavior was influenced by DENV infection. Taken together, this multidisciplinary study details how DENV infection modulates mosquito blood-feeding behavior to increase vector capacity, proportionally aggravating DENV epidemiology. By elucidating the contribution of mosquito behavioral alterations on DENV transmission to the host, these results will inform epidemiological modeling to tailor improved interventions against dengue.

Tyrosine-O-sulfation is a widespread affinity enhancer among thrombin interactors

Tyrosine-O-sulfation is a common post-translational modification (PTM) of proteins following the cellular secretory pathway. First described in human fibrinogen, tyrosine-O-sulfation has long been associated with the modulation of protein–protein interactions in several physiological processes. A number of relevant interactions for hemostasis are largely dictated by this PTM, many of which involving the serine proteinase thrombin (FIIa), a central player in the blood-clotting cascade. Tyrosine sulfation is not limited to endogenous FIIa ligands and has also been found in hirudin, a well-known and potent thrombin inhibitor from the medicinal leech, Hirudo medicinalis. The discovery of hirudin led to successful clinical application of analogs of leech-inspired molecules, but also unveiled several other natural thrombin-directed anticoagulant molecules, many of which undergo tyrosine-O-sulfation. The presence of this PTM has been shown to enhance the anticoagulant properties of these peptides from a range of blood-feeding organisms, including ticks, mosquitos and flies. Interestingly, some of these molecules display mechanisms of action that mimic those of thrombin's bona fide substrates.

The AMA1-RON complex drives Plasmodium sporozoite invasion in the mosquito and mammalian hosts

Plasmodium sporozoites that are transmitted by blood-feeding female Anopheles mosquitoes invade hepatocytes for an initial round of intracellular replication, leading to the release of merozoites that invade and multiply within red blood cells. Sporozoites and merozoites share a number of proteins that are expressed by both stages, including the Apical Membrane Antigen 1 (AMA1) and the Rhoptry Neck Proteins (RONs). Although AMA1 and RONs are essential for merozoite invasion of erythrocytes during asexual blood stage replication of the parasite, their function in sporozoites is still unclear. Here we show that AMA1 interacts with RONs in mature sporozoites. By using DiCre-mediated conditional gene deletion in P. berghei, we demonstrate that loss of AMA1, RON2 or RON4 in sporozoites impairs colonization of the mosquito salivary glands and invasion of mammalian hepatocytes, without affecting transcellular parasite migration. Our data establish that AMA1 and RONs facilitate host cell invasion across Plasmodium invasive stages, and suggest that sporozoites use the AMA1-RON complex to safely enter the mosquito salivary glands without causing cell damage, to ensure successful parasite transmission. These results open up the possibility of targeting the AMA1-RON complex for transmission-blocking antimalarial strategies.

Comparative Insecticidal Effects of Dry Ocimum Gratissium (Scent Leaves) and Rambo™ Paper on Mosquitoes in Jos, Nigeria

Insecticides are chemicals or biological substances that are used to kill or disable insects. Blood feeding mosquitoes are responsible for the intolerable biting nuisance and transmission of large number of diseases such as malaria, yellow fever, dengue fever, filarias is amongst others, causing serious health problems to humans and obstacles to socioeconomic development of developing nations like Nigeria. The insecticidal effect of scent leaves (Ocimum gratissum) and Rambo™ paper on mosquitoes was investigated. The study is aimed at comparing the insecticidal effects of Ocimum gratissimum and Rambo™ paper on mosquitoes in Jos. 100 mosquitoes were exposed to dried Ocimum gratissum and Rambo™ paper at different time intervals of 5, 10 and 12 min. Results obtained showed a time dependent insecticidal effect on mosquitoes, which was 54.2%, 54.0% and 55.6% total mortality of mosquitoes at respective time intervals on exposures to both Rambo™ paper insecticide and Osimum gratissum, indicating that there was a significant difference in the lethal effect of Rambo™ paper insecticide and scent leave on mosquitoes at (p<0.05). In comparing the lethal effect of Rambo™ paper insecticide and Ocimum grasstisimum on mosquitoes at differnt locations in Jos, at Dogon Karfe, after 10 min of treatment, Rambo™ paper had the highest lethal effect of 21 (84.0%) compared to scent leaves 6 (24.0%) and at Abattoir Jos, after 12 min of treatment, Rambo™ paper had the highest lethal effect of 17 (94.4%) compared to Ocimum grasstisimum 3 (16.7%). These comparisons were significant at p 0.05. This study provides evidence that Ocimum grasstisimum has a mosquitocidal effect. However, the Rambo™ paper gave a better mosquitocidal effect than Ocimum grasstisimum. There is a need to discover better additive or extract options that could give Ocimum grasstisimum a better effect as a natural product available in Africa towards the malaria eradication programme.

Screening for Odorant Receptor Genes Expressed in Aedes Aegypti Involved in Blood-Feeding and Oviposition Behaviors

Background: Aedes aegypti is one of the most important vector worldwide, and its survival and reproductive processes depend heavily on the olfactory system. In this study, the expression levels of all odorant receptor (OR) genes of Ae. aegypti were explored in different physiological periods to identify olfactory genes that may be associated with mosquito blood sucking and searching for oviposition sites.Methods: Four groups, consisting of Ae. aegypti males (M), pre-blood-feeding females (F), post-blood-feeding females (B) and post-oviposition females (O), were established. A total of 114 pairs of primer targeting all OR genes were designed based on the whole genome of Ae. aegypti. The expression of OR genes was evaluated by real-time fluorescence quantitative PCR for relative quantification and the comparison of differences between groups.Results: A total of 53 differentially expressed OR genes were identified between males and females in Ae. aegypti antennae. And 8, 5 and 13 differentially expressed OR genes were identified before versus after blood feeding, before versus after oviposition and post-blood-feeding versus post-oviposition, respectively. Meanwhile, 16 OR genes were significantly differentially expressed in multiple physiological periods of mosquitoes.Conclusions: A large number of ORs with significant intergroup differences and high expression levels were screened in this study, including OR75, OR88, OR110 and OR115 and so on. Some of these genes are reported for the first time, providing possible targets for the development of mosquito control pathways based on the olfactory system.

A transcriptomic atlas of Aedes aegypti reveals detailed functional organization of major body parts and gut regional specializations in sugar-fed and blood-fed adult females

Mosquito vectors transmit numerous pathogens, but large gaps remain in our understanding of their physiology. To facilitate future explorations of mosquito biology, with specific attention to the major vector Aedes aegypti, we have created Aegypti-Atlas (http://aegyptiatlas.buchonlab.com/), an online resource hosting RNAseq profiles of Ae. aegypti body parts (head, thorax, abdomen, gut, Malpighian tubules, and ovaries), gut regions (crop, proventriculus, anterior and posterior midgut, and hindgut), and a time course of blood meal digestion in the gut. Using Aegypti-Atlas, we provide new insights into the regionalization of gut function, blood feeding response, and immune defenses. We find that the anterior and posterior regions of the mosquito midgut possess clearly delineated digestive specializations which are preserved in the blood-fed state. Blood feeding initiates the sequential transcriptional induction and repression/depletion of multiple cohorts of peptidases throughout blood meal digestion. With respect to defense, immune signaling components, but not recognition or effector molecules, show enrichment in ovaries. Basal expression of antimicrobial peptides is dominated by two genes, holotricin and gambicin, that are expressed in the carcass and the digestive tissues, respectively, in a near mutually exclusive manner. In the midgut, gambicin and other immune effector genes are almost exclusively expressed in the anterior regions, while the posterior midgut exhibits the hallmarks of immune tolerance. Finally, in a cross-species comparison between the midguts of Ae. aegypti and Anopheles gambiae, we observe that regional digestive and immune specializations are closely conserved, indicating that our data may yield inferences that are broadly relevant to multiple mosquito vector species. We further demonstrate that the expression of orthologous genes is highly correlated, with the exception of a ‘species signature’ comprising a small number of highly/disparately expressed genes. With this work, we show the potential of Aegypti-Atlas to unlock a more complete understanding of mosquito biology.

Barcoded Asaia bacteria enable mosquito in vivo screens and identify novel systemic insecticides and inhibitors of malaria transmission

This work addresses the need for new chemical matter in product development for control of pest insects and vector-borne diseases. We present a barcoding strategy that enables phenotypic screens of blood-feeding insects against small molecules in microtiter plate-based arrays and apply this to discovery of novel systemic insecticides and compounds that block malaria parasite development in the mosquito vector. Encoding of the blood meals was achieved through recombinant DNA-tagged Asaia bacteria that successfully colonised Aedes and Anopheles mosquitoes. An arrayed screen of a collection of pesticides showed that chemical classes of avermectins, phenylpyrazoles, and neonicotinoids were enriched for compounds with systemic adulticide activity against Anopheles. Using a luminescent Plasmodium falciparum reporter strain, barcoded screens identified 48 drug-like transmission-blocking compounds from a 400-compound antimicrobial library. The approach significantly increases the throughput in phenotypic screening campaigns using adult insects and identifies novel candidate small molecules for disease control.