Beyond the eye: Kynurenine pathway impairment causes midgut homeostasis dysfunction and survival and reproductive costs in blood-feeding mosquitoes.

Insect ommochrome biosynthesis pathways metabolize tryptophan to generate eye-color pigments and wild-type alleles of pathway genes are useful phenotypic markers in transgenesis studies. Pleiotropic effects of mutations in some genes exert a load on both survival and reproductive success in blood-feeding species. Here, we investigated the challenges imposed on mosquitoes by the increase of tryptophan metabolites resulting from blood meal digestion and the impact of disruptions of the ommochrome biosynthesis pathway. Female mosquitoes with spontaneous and induced mutations in the orthologs of the genes encoding kynurenine hydroxylase in Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus exhibited impaired survival and reproductive phenotypes that varied in type and severity among the species. A compromised midgut permeability barrier function was also observed in An. stephensi. Surprisingly, mutant mosquitoes displayed an increase in microbiota compared to controls that was not accompanied by a general induction of immune genes. Antibiotic treatment rescued some deleterious traits implicating a role for the kynurenine pathway (KP) in midgut homeostasis. Supplemental xanthurenic acid, a KP end-product, rescued lethality and limited microbiota proliferation in Ae. aegypti. These data implicate the KP in the regulation of the host/microbiota interface. These pleiotropic effects on mosquito physiology are important in the development of genetic strategies targeting vector mosquitoes.

Aedes aegypti post-emergence transcriptome: Unveiling the molecular basis for the hematophagic and gonotrophic capacitation.

The adult females of Aedes aegypti mosquitoes are facultative hematophagous insects but they are unable to feed on blood right after pupae emergence. The maturation process that takes place during the first post-emergence days, hereafter named hematophagic and gonotrophic capacitation, comprises a set of molecular and physiological changes that prepare the females for the first gonotrophic cycle. Notwithstanding, the molecular bases underlying mosquito hematophagic and gonotrophic capacitation remain obscure. Here, we investigated the molecular and biochemical changes in adult Ae. aegypti along the first four days post-emergence, prior to a blood meal. We performed a RNA-Seq analysis of the head and body, comparing male and female gene expression time courses. A total of 811 and 203 genes were differentially expressed, respectively in the body and head, and both body parts showed early, mid, and late female-specific expression profiles. Female-specific up-regulation of genes involved in muscle development and the oxidative phosphorylation pathway were remarkable features observed in the head. Functional assessment of mitochondrial oxygen consumption in heads showed a gradual increase in respiratory capacity and ATP-linked respiration as a consequence of induced mitochondrial biogenesis and content over time. This pattern strongly suggests that boosting oxidative phosphorylation in heads is a required step towards blood sucking habit. Several salivary gland genes, proteases, and genes involved in DNA replication and repair, ribosome biogenesis, and juvenile hormone signaling were up-regulated specifically in the female body, which may reflect the gonotrophic capacitation. This comprehensive description of molecular and biochemical mechanisms of the hematophagic and gonotrophic capacitation in mosquitoes unravels potentially new targets for vector control.

Methodological description and preliminary results of a cohort study on the influence of the first 1,000 days of life on the children's future health.

The aim of this report is to describe general and methodological characteristics of a cohort study in southern Brazil (Coorte Brasil Sul), aimed at understanding the impact of the first 1,000 days of life on children's health. It is a cohort study involving all children born in 2009 and their families living in the municipality of Palhoça, State of Santa Catarina, Brazil. Face-to-face interviews with parents at home using a structured questionnaire and children's physical and clinical examinations at schools have been carried out. Cross-sectional analyzes, longitudinal comparisons and hierarquical regression analysis will allow understanding if the first 1,000 days of life can influence on 6-year-old children's health. The Coorte Brasil Sul is in its retrospective phase together with the children's physical data collection. Preliminary data (n=1270) related to nutritional status point to a high prevalence of overweight (16.4%) and obesity (15.5%). With the continuity of the study, it is expected to evaluate if the first phases of life can influence health during adolescence and in adult life, mainly in relation to chronic diseases.

Tyrosine Detoxification Is an Essential Trait in the Life History of Blood-Feeding Arthropods.

Blood-feeding arthropods are vectors of infectious diseases such as dengue, Zika, Chagas disease, and malaria [1], and vector control is essential to limiting disease spread. Because these arthropods ingest very large amounts of blood, a protein-rich meal, huge amounts of amino acids are produced during digestion. Previous work on Rhodnius prolixus, a vector of Chagas disease, showed that, among all amino acids, only tyrosine degradation enzymes were overexpressed in the midgut compared to other tissues [2]. Here we demonstrate that tyrosine detoxification is an essential trait in the life history of blood-sucking arthropods. We found that silencing Rhodnius tyrosine aminotransferase (TAT) and 4-hydroxyphenylpyruvate dioxygenase (HPPD), the first two enzymes of the phenylalanine/tyrosine degradation pathway, caused the death of insects after a blood meal. This was confirmed by using the HPPD inhibitor mesotrione, which selectively killed hematophagous arthropods but did not affect non-hematophagous insects. In addition, mosquitoes and kissing bugs died after feeding on mice that had previously received a therapeutic effective oral dose (1 mg/kg) of nitisinone, another HPPD inhibitor used in humans for the treatment of tyrosinemia type I [3]. These findings indicate that HPPD (and TAT) can be a target for the selective control of blood-sucking disease vector populations. Because HPPD inhibitors are extensively used as herbicides and in medicine, these compounds may provide an alternative less toxic to humans and more environmentally friendly than the conventional neurotoxic insecticides that are currently used, with the ability to affect only hematophagous arthropods.

Amino acids trigger down-regulation of superoxide via TORC pathway in the midgut of Rhodnius prolixus.

Sensing incoming nutrients is an important and critical event for intestinal cells to sustain life of the whole organism. The TORC is a major protein complex involved in monitoring the nutritional status and is activated by elevated amino acid concentrations. An important feature of haematophagy is that huge amounts of blood are ingested in a single meal, which results in the release of large quantities of amino acids, together with the haemoglobin prosthetic group, haem, which decomposes hydroperoxides and propagates oxygen-derived free radicals. Our previous studies demonstrated that reactive oxygen species (ROS) levels were diminished in the mitochondria and midgut of the Dengue fever mosquito, Aedes aegypti, immediately after a blood meal. We proposed that this mechanism serves to avoid oxidative damage that would otherwise be induced by haem following a blood meal. Studies also performed in mosquitoes have shown that blood or amino acids controls protein synthesis through TORC activation. It was already proposed, in different models, a link between ROS and TOR, however, little is known about TOR signalling in insect midgut nor about the involvement of ROS in this pathway. Here, we studied the effect of a blood meal on ROS production in the midgut of Rhodnius prolixus We observed that blood meal amino acids decreased ROS levels in the R. prolixus midgut immediately after feeding, via lowering mitochondrial superoxide production and involving the amino acid-sensing TORC pathway.

The antioxidant role of xanthurenic acid in the Aedes aegypti midgut during digestion of a blood meal.

In the midgut of the mosquito Aedes aegypti, a vector of dengue and yellow fever, an intense release of heme and iron takes place during the digestion of a blood meal. Here, we demonstrated via chromatography, light absorption and mass spectrometry that xanthurenic acid (XA), a product of the oxidative metabolism of tryptophan, is produced in the digestive apparatus after the ingestion of a blood meal and reaches milimolar levels after 24 h, the period of maximal digestive activity. XA formation does not occur in the White Eye (WE) strain, which lacks kynurenine hydroxylase and accumulates kynurenic acid. The formation of XA can be diminished by feeding the insect with 3,4-dimethoxy-N-[4-(3-nitrophenyl)thiazol-2-yl] benzenesulfonamide (Ro-61-8048), an inhibitor of XA biosynthesis. Moreover, XA inhibits the phospholipid oxidation induced by heme or iron. A major fraction of this antioxidant activity is due to the capacity of XA to bind both heme and iron, which occurs at a slightly alkaline pH (7.5-8.0), a condition found in the insect midgut. The midgut epithelial cells of the WE mosquito has a marked increase in occurrence of cell death, which is reversed to levels similar to the wild type mosquitoes by feeding the insects with blood supplemented with XA, confirming the protective role of this molecule. Collectively, these results suggest a new role for XA as a heme and iron chelator that provides protection as an antioxidant and may help these animals adapt to a blood feeding habit.

The major insect lipoprotein is a lipid source to mosquito stages of malaria parasite.

Once mosquito midgut barrier was crossed malaria parasite faces a extensive metabolic developmental program in order to ensure its transmission. In the hemolymph of the mosquito the dynamics of lipid metabolism is conducted by a major lipoprotein, lipophorin (Lp). It was recently shown that Lp is engaged in the mosquito immune response to parasite infection. However, it is not clear if Lp is uptaken by the parasite. Here, we show that oocysts are able to uptake mosquito Lp. The uptake of FITC-labeled Lp was demonstrated in midgut-associated oocysts. Alternatively, to confirm Lp incorporation by oocysts we have conducted another set of experiments with iodinated Lp ((125)I-Lp). Oocysts were able to incorporate (125)I-Lp and the process is both time and temperature dependent. This set of results indicated that no matter oocysts are attached to mosquito midgut wall they bear a lipid sequestering machinery from its surroundings. Phospholipid transfer to sporozoites was also demonstrated. In conclusion, these results demonstrate for the first time that malaria parasite undergoes lipid uptake while in the invertebrate host.