Project description:RNA-Seq was used for transcriptome sequencing of 11 samples covering various life stages and tissues from the yellow fever mosquito Ae. aegypti.
Project description:The impact of global climate change on the transmission dynamics of infectious diseases is the subject of extensive debate. The transmission of mosquito-borne viral diseases is particularly complex, with climatic variables directly affecting many parameters associated with the prevalence of disease vectors. While evidence shows that warmer temperatures often decrease the extrinsic incubation period of an arthropod-borne virus (arbovirus), exposure to cooler temperatures often predisposes disease vector mosquitoes to higher infection rates. RNA interference pathways are essential to antiviral immunity in the mosquito; however, few experiments have explored the effects of temperature on the RNAi machinery. Total small RNAs (miRNAs, siRNAs, piRNAs, etc.) were isolated and sequenced from the heads of sensor strain Aedes aegypti mosquitoes, or from the whole bodies of CHIKV-infected Aedes albopictus mosquitoes 8 hours post infection. Mosquitoes were grown at 18C or 28C in replicates of 1 (Ae. aegypti) or 3 (Ae. albopictus).
Project description:Aedes aegypti (L.) is the primary vector of many emerging arboviruses. Insecticide resistance among mosquito populations is a consequence of the application of insecticides for mosquito control. We used RNA-sequencing to compare transcriptomes between permethrin resistant and susceptible strains of Florida Ae. aegypti in response to Zika virus infection. A total of 2,459 transcripts were expressed at significantly different levels between resistant and susceptible Ae. aegypti. Gene ontology analysis placed these genes into 7 categories of biological processes. The 863 transcripts were expressed at significantly different levels between two strains (up/down regulated) more than 2-fold. Quantitative real-time PCR analysis validated Zika-infected response, and suggested a highly overexpressed P450, with AAEL014617 and AAEL006798 as potential candidates for the molecular mechanism of permethrin resistance in Ae. aegypti. Our findings indicated that most detoxification enzymes and immune system enzymes altered their gene expression between the two strains of Ae. aegypti in response to Zika virus infection. Understanding the interactions of arboviruses with resistant mosquito vectors at the molecular level allows for the possible development of new approaches in mitigating arbovirus transmission. This information sheds light on Zika-induced changes in the insecticide resistance of Ae. aegypti with implications for mosquito control strategies.
Project description:As a key vector for the major arthropod-borne viruses (arboviruses), such as dengue, Zika and chikungunya, control of Aedes aegypti represents a major challenge in public health. Bloodmeal acquisition is necessary for the reproduction of vector mosquitoes and pathogen transmission. Blood contains potentially toxic amounts of iron while it provides nutrients for mosquito offspring; disruption of iron homeostasis in the mosquito may therefore lead to a novel control strategies. We previously described a potential iron exporter in Ae. aegypti after a targeted functional screen of ZIP and ZnT family genes. In this study, we performed an RNAseq-based screen in an Ae. aegypti cell line cultured under iron-deficient and iron-excess conditions. A subset of differentially expressed genes were analyzed via a cytosolic iron-sensitive dual-luciferase reporter assay with several gene candidates potentially involved in iron transport. In vivo gene silencing resulted in significant reduction of fecundity (egg number) and fertility (hatch rate) for one gene, AAEL000471. Consistent with a role in iron homeostasis, ferritin expression in the midgut was reduced upon silencing of AAEL000471. Silencing of AAEL000471 also resulted in delayed/impaired excretion and digestion. Together these studies increase our understanding of gene regulatory networks controlling iron homeostasis in bloodfeeding mosquitoes, and suggest new targets for vector control based on the disruption of this critical process.
Project description:Zika virus (ZIKV) of the Flaviviridae family is a recently emerged mosquito-borne virus that has been implicated in the surge of the number of microcephaly instances in south America. The virus is transmitted mainly by the mosquito Aedes aegypti that also vectors dengue virus. Considering rather recent rapid spread of the virus and its declaration as a global health emergency by the World Health Organization, little is known about the interactions of the virus with the mosquito vector. In this study, we investigated the transcriptome profiles of whole Ae. aegypti mosquitoes in response to ZIKV infection at 2, 7 and 14 days post-infection using deep sequencing. Results showed a large number of transcripts were altered at each time point following infection, but 18 transcripts were commonly changed at the three time points. The outcomes provide a basic understanding of Ae. aegypti responses to ZIKV and help determining host factors involved in replication or anti-viral response against the virus.
Project description:RNA-Seq was used for transcriptome sequencing of 11 samples covering various life stages and tissues from the yellow fever mosquito Ae. aegypti. RNA was isolated from described samples for RNA-Seq. Samples included four embryonic time points from 0-12 hr, larvae, pupae, adult males, 0-1 day old ovaries, ovaries from 24 post blood fed females, and 24 hr post blood fed female carcass (ovaries extirpated). Raw data not available for GSM847403, GSM847404, GSM847405, and GSM847406
Project description:The dengue virus (DENV) cause frequent epidemics infecting ~390 million people annually in over 100 countries. There are no approved vaccines or antiviral drugs for treatment of infected patients. However, there is a novel approach to control transmission of DENV by the mosquito vectors, Aedes aegypti and Ae. albopictus, using Wolbachia symbiont. The wMelPop strain of Wolbachia suppresses DENV transmission and shortens the mosquito life span. However, the underlying mechanism is poorly understood. To clarify this mechanism, either naïve Ae. albopictus (C6/36) or wMelPop-C6/36 cells were infected with DENV2. Analysis of host transcript profiles by RNAseq revealed that the presence of wMelPop had profound effects on mosquito host cell transcription in response to DENV2 infection. The viral RNA evolved from wMelPop-C6/36 contained low frequency mutations (~25%) within the coding region of transmembrane domain-1 (TMD1) of E protein. Mutations with >97 % frequencies were distributed within other regions of E, NS5 RNA-dependent RNA polymerase (NS5POL) domain, the TMDs of NS2A, NS2B, and NS4B. Moreover, while DENV2-infected naïve C6/36 cells showed syncytia formation, DENV2-infected wMelPop-C6/36 cells did not. The Wolbachia-induced mutant DENV2 can readily infect and replicate in naïve C6/36 cells; whereas, in the mutant DENV2- infected BHK-21 or Vero cells, the virus replication was delayed. In LLC-MK2 cells, the mutant failed to produce plaques. Additionally, in BHK-21 cells, many mutations in the viral genome reverted to WT and compensatory mutations in NS3 gene appeared. Our results suggest that wMelPop impacts significantly the interactions of DENV2 with mosquito and mammalian host cells.
Project description:Background: The piRNA pathway has been shown in model organisms to be involved in silencing of transposons thereby providing genome stability. In D. melanogaster the majority of piRNAs map to these sequences. The medically important mosquito species Aedes aegypti has a large genome size, a high transposon load which includes Miniature Inverted repeat Transposable Elements (MITES) and an expansion of the piRNA biogenesis genes. Studies of transgenic lines of Ae. aegypti have indicated that introduced transposons are poorly remobilized and we sought to explore the basis of this. We wished to analyze the piRNA profile of Ae. aegypti and thereby determine if it be responsible for transposon silencing in this mosquito. Results: Estimated piRNA sequence diversity was comparable between Ae. aegypti and D. melanogaster, but surprisingly only 19% of mosquito piRNAs mapped to transposons compared to 51% for D. melanogaster. Ae. aegypti piRNA clusters made up a larger percentage of the total genome than those of D. melanogaster but did not contain significantly higher percentages of transposon derived sequences than other regions of the genome. Ae. aegypti contains a number of protein coding genes that may be sources of piRNA biogenesis with two, traffic jam and maelstrom, implicated in this process in model organisms. Several genes of viral origin were also targeted by piRNAs. Examination of six mosquito libraries that had previously been transformed with transposon derived sequence revealed that new piRNA sequences had been generated to the transformed sequences, suggesting that they may have stimulated a transposon inactivation mechanism. Conclusions: Ae. aegypti has a large piRNA complement that maps to transposons but primarily gene sequences, including many viral-derived sequences. This, together the more uniform distribution of piRNA clusters throughout its genome suggest that some aspects of the piRNA system differ between Ae. aegypti and D. melanogaster. 5 small RNA libraries were generated from total RNA of whole adult Aedes aegypti tissues, two of these libraries were sequenced twice (technical replicates). 1 small RNA library was generated from total RNA of a whole adult Drosophila melanogaster tissue.
Project description:We have generated transgenic Ae. aegypti that express a ZIKV-specific inverted repeat RNA intended to trigger the mosquito siRNA anti-viral immune pathway. To confirm that the transgene is expressed and processed, we performed small RNA sequencing on both Higgs White Eye (HWE) & anti-NS3/4A-ZIKV transgenic Ae. aegypti 24 hours post-bloodmeal
Project description:We compare the transcriptome of gnotobiotic Ae. aegypti generated by contaminating axenic (bacteria-free) larvae with bacterial isolates found in natural mosquito breeding sites. We focused on four bacterial isolates (Lysobacter, Flavobacterium, Paenibacillus and Enterobacteriaceae) and found that different gnotobiotic treatments resulted in massive transcriptomic changes throughout the mosquito development.