Insulin-like peptides in the mosquito Anopheles stephensi: identification and expression in response to diet and infection with Plasmodium falciparum.

Insulin-like peptides (ILPs) regulate a multitude of biological processes, including metabolism and immunity to infection, and share similar structural motifs across widely divergent taxa. Insulin/insulin-like growth factor signaling (IIS) pathway elements are similarly conserved. We have shown that IIS regulates reproduction, innate immunity, and lifespan in female Anopheles stephensi, a major mosquito vector of human malaria. To further explore IIS regulation of these processes, we identified genes encoding five ILPs in this species and characterized their expression in tissues. Antisera to ILP homologs in Anopheles gambiae were used to identify cellular sources in An. stephensi females by immunocytochemistry. We analyzed tissue-specific ILP transcript expression in young and older females, in response to different feeding regimens, and in response to infection with Plasmodiumfalciparum with quantitative reverse transcriptase-PCR assays. While some ILP transcript changes were evident in older females and in response to blood feeding, significant changes were particularly notable in response to hormonal concentrations of ingested human insulin and to P. falciparum infection. These changes suggest that ILP secretion and action may be similarly responsive in Plasmodium-infected females and potentially alter metabolism and innate immunity.

Correction: Activation of Akt signaling reduces the prevalence and intensity of malaria parasite infection and lifespan in Anopheles stephensi mosquitoes.

Malaria (Plasmodium spp.) kills nearly one million people annually and this number will likely increase as drug and insecticide resistance reduces the effectiveness of current control strategies. The most important human malaria parasite, Plasmodium falciparum, undergoes a complex developmental cycle in the mosquito that takes approximately two weeks and begins with the invasion of the mosquito midgut. Here, we demonstrate that increased Akt signaling in the mosquito midgut disrupts parasite development and concurrently reduces the duration that mosquitoes are infective to humans. Specifically, we found that increased Akt signaling in the midgut of heterozygous Anopheles stephensi reduced the number of infected mosquitoes by 60-99%. Of those mosquitoes that were infected, we observed a 75-99% reduction in parasite load. In homozygous mosquitoes with increased Akt signaling parasite infection was completely blocked. The increase in midgut-specific Akt signaling also led to an 18-20% reduction in the average mosquito lifespan. Thus, activation of Akt signaling reduced the number of infected mosquitoes, the number of malaria parasites per infected mosquito, and the duration of mosquito infectivity.

Activation of Akt signaling reduces the prevalence and intensity of malaria parasite infection and lifespan in Anopheles stephensi mosquitoes.

Malaria (Plasmodium spp.) kills nearly one million people annually and this number will likely increase as drug and insecticide resistance reduces the effectiveness of current control strategies. The most important human malaria parasite, Plasmodium falciparum, undergoes a complex developmental cycle in the mosquito that takes approximately two weeks and begins with the invasion of the mosquito midgut. Here, we demonstrate that increased Akt signaling in the mosquito midgut disrupts parasite development and concurrently reduces the duration that mosquitoes are infective to humans. Specifically, we found that increased Akt signaling in the midgut of heterozygous Anopheles stephensi reduced the number of infected mosquitoes by 60-99%. Of those mosquitoes that were infected, we observed a 75-99% reduction in parasite load. In homozygous mosquitoes with increased Akt signaling parasite infection was completely blocked. The increase in midgut-specific Akt signaling also led to an 18-20% reduction in the average mosquito lifespan. Thus, activation of Akt signaling reduced the number of infected mosquitoes, the number of malaria parasites per infected mosquito, and the duration of mosquito infectivity.

The role of NF-kappaB factor REL2 in the Aedes aegypti immune response.

Mosquitoes transmit numerous diseases that continue to be an enormous burden on public health worldwide. Transgenic mosquitoes impervious to vector-borne pathogens, in concert with vector control and drug and vaccine development, comprise an arsenal of means anticipated to defeat mosquito-spread diseases in the future. Mosquito transgenesis allows tissue-specific manipulation of their major immune pathways and enhances the ability to study mosquito-pathogen interactions. Here, we report the generation of two independent transgenic strains of Aedes aegypti overexpressing the NF-?B transcriptional factor REL2, a homologue of Drosophila Relish, which is shown to be under the control of the vitellogenin promoter in the mosquito fat body after a blood meal. We show that this REL2 overexpression in the fat body results in transcriptional activation of Defensins A, C, and D, and Cecropins A and N, as well as translation and secretion of Defensin A protein into the hemolymph. We also demonstrate that induction of REL2 results in the increased resistance of the mosquito to tested Gram-negative and Gram-positive bacteria. Importantly, induction of transgenic REL2 leads to the significant decrease in susceptibility of A. aegypti to Plasmodium gallinaceum infection. Consistently, RNAi knockdown of REL2 in wild-type mosquitoes results in a delay in Defensin A and Cecropin A expression in response to infection and in increased susceptibility to both bacteria and P. gallinaceum. Moreover, our transgenic assays demonstrate that the N-terminus of the mosquito REL2, which includes the His/Gln-rich and serine-rich regions, plays a role in its transactivation properties.