Kissing Bug Rhodnius Prolixus Research Articles

Aversive sexual learning in the kissing bug Rhodnius prolixus: Modulation of different sexual responses in males and females

Although sexual learning can be a key process in the reproductive success of animals, research focused on the experience-dependent modulation of courtship in insects is scarce. In kissing bugs, the behavioural steps implicated in mating have been exhaustively studied, but not the involvement of learning in them. Our objective was to determine whether the sexual behaviour of Rhodnius prolixus could be modulated by experience. During training, couples were submitted to eight assays, in which they received a vibration (negative reinforcement) when the male attempted to copulate the female. Immediately after, they were separated, not allowing the occurrence of copulation. We found that along training, males’ latency to perform a copulatory attempt increased, male’s copulatory attempts were less frequent, and females’ locomotor activity did not change. These results suggest that males, and not females, learned to avoid the vibration by reducing their intention to copulate. In post-training tests, conditioned males presented with new naïve females reverted to low copulatory attempt latencies, suggesting that the modulation was partner-specific. Besides, conditioned females increased their rejection frequencies to males’ copulatory attempts, suggesting that a second type of learning occurred in females. These results constitute the first evidence of sexual learning in hematophagous insects. Males and females seem to change their selectivity according to their previous sexual experience. We discuss the relevance that this plasticity might have in the fitness of this epidemiologically relevant insect.

The mlpt smORF gene is essential for digestive physiology and molting during nymphal stages in the kissing bug Rhodnius prolixus

Chagas disease affects around 8 million people globally, with Latin America bearing approximately 10,000 deaths each year. Combatting the disease relies heavily on vector control methods, necessitating the identification of new targets. Within insect genomes, genes harboring small open reading frames (smORFs - < 100 amino acids) present numerous potential candidates. In our investigation, we elucidate the pivotal role of the archetypal smORF-containing gene, mille-pattes/polished-rice/tarsalless (mlpt/pri/tal), in the post-embryonic development of the kissing bug Rhodnius prolixus. Injection of double-stranded RNA targeting mlpt (dsmlpt) during nymphal stages yields a spectrum of phenotypes hindering post-embryonic growth. Notably, fourth or fifth stage nymphs subjected to dsmlpt do not undergo molting. These dsmlpt nymphs display heightened mRNA levels of JHAMT-like and EPOX-like, enzymes putatively involved in the juvenile hormone (JH) pathway, alongside increased expression of the transcription factor Kr-h1, indicating changes in the hormonal control. Histological examination reveals structural alterations in the hindgut and external cuticle of dsmlpt nymphs compared to control (dsGFP) counterparts. Furthermore, significant changes in the vector's digestive physiology were observed, with elevated hemozoin and glucose levels in the posterior midgut of dsmlpt nymphs. Importantly, dsmlpt nymphs exhibit impaired metacyclogenesis of Trypanosoma cruzi, the causative agent of Chagas disease, underscoring the crucial role of proper gut organization in parasite differentiation. Thus, our findings constitute the first evidence of a smORF-containing gene's regulatory influence on vector physiology, parasitic cycle, and disease transmission.

The pharyngeal taste organ of a blood-feeding insect functions in food recognition

BackgroundObligate blood-feeding insects obtain the nutrients and water necessary to ensure survival from the vertebrate blood. The internal taste sensilla, situated in the pharynx, evaluate the suitability of the ingested food. Here, through multiple approaches, we characterized the pharyngeal organ (PO) of the hematophagous kissing bug Rhodnius prolixus to determine its role in food assessment. The PO, located antero-dorsally in the pharynx, comprises eight taste sensilla that become bathed with the incoming blood.ResultsWe showed that these taste sensilla house gustatory receptor neurons projecting their axons through the labral nerves to reach the subesophageal zone in the brain. We found that these neurons are electrically activated by relevant appetitive and aversive gustatory stimuli such as NaCl, ATP, and caffeine. Using RNA-Seq, we examined the expression of sensory-related gene families in the PO. We identified gustatory receptors, ionotropic receptors, transient receptor potential channels, pickpocket channels, opsins, takeouts, neuropeptide precursors, neuropeptide receptors, and biogenic amine receptors. RNA interference assays demonstrated that the salt-related pickpocket channel Rproppk014276 is required during feeding of an appetitive solution of NaCl and ATP.ConclusionsWe provide evidence of the role of the pharyngeal organ in food evaluation. This work shows a comprehensive characterization of a pharyngeal taste organ in a hematophagous insect.

Evidence that hematophagous triatomine bugs may eat plants in the wild

Blood feeding is a secondary adaptation in hematophagous bugs. Many proteins are secreted in the saliva that are devoted to coping with the host's defense and to process the blood meal. Digestive enzymes that are no longer required for a blood meal would be expected to be eventually lost. Yet, in many strictly hematophagous arthropods, α-amylase genes, which encode the enzymes that digest starch from plants, are still present and transcribed, including in the kissing bug Rhodnius prolixus (Hemiptera, Reduviidae) and its related species, which transmit the Chagas disease. We hypothesized that retaining α-amylase could be advantageous if the bugs occasionally consume plant tissues. We first checked that the α-amylase protein of Rhodnius robustus retains normal amylolytic activity. Then we surveyed hundreds of gut DNA extracts from the sylvatic R. robustus to detect traces of plants. We found plant DNA in 8% of the samples, mainly identified as Attalea palm trees, where R. robustus are usually found. We suggest that although of secondary importance in the blood-sucking bugs, α-amylase may be needed during occasional plant feeding and thus has been retained.

Heme protonation affects iron-NO binding in the NO transport protein nitrophorin

The nitrophorins (NPs) comprise an unusual group of heme proteins with stable ferric heme iron nitric oxide (Fe-NO) complexes. They are found in the salivary glands of the blood-sucking kissing bug Rhodnius prolixus, which uses the NPs to transport the highly reactive signaling molecule NO. Nuclear resonance vibrational spectroscopy (NRVS) of both isoform NP2 and a mutant NP2(Leu132Val) show, after addition of NO, a strong structured vibrational band at around 600 cm−1, which is due to modes with significant Fe-NO bending and stretching contribution. Based on a hybrid calculation method, which uses density functional theory and molecular mechanics, it is demonstrated that protonation of the heme carboxyl groups does influence both the vibrational properties of the Fe-NO entity and its electronic ground state. Moreover, heme protonation causes a significant increase of the gap between the highest occupied and lowest unoccupied molecular orbital by almost one order of magnitude leading to a stabilization of the Fe-NO bond.

ATP synthase affects lipid metabolism in the kissing bug Rhodnius prolixus beyond its role in energy metabolism

ATP synthase plays an essential role in mitochondrial metabolism, being responsible for the production of ATP in oxidative phosphorylation. However, recent results have shown that it may also be present in the cell membrane, involved in lipophorin binding to its receptors. Here, we used a functional genetics approach to investigate the roles of ATP synthase in lipid metabolism in the kissing bug Rhodnius prolixus. The genome of R. prolixus encodes five nucleotide-binding domain genes of the ATP synthase α and β family, including the α and β subunits of ATP synthase (RpATPSynα and RpATPSynβ), and the catalytic and non-catalytic subunits of the vacuolar ATPase (RpVha68 and RpVha55). These genes were expressed in all analyzed organsn highest in the ovaries, fat body and flight muscle. Feeding did not regulate the expression of ATP synthases in the posterior midgut or fat body. Furthermore, ATP synthase is present in the fat body's mitochondrial and membrane fractions. RpATPSynβ knockdown by RNAi impaired ovarian development and reduced egg-laying by approximately 85%. Furthermore, the lack of RpATPSynβ increased the amount of triacylglycerol in the fat body due to increased de novo fatty acid synthesis and reduced transfer of lipids to lipophorin. RpATPSynα knockdown had similar effects, with altered ovarian development, reduced oviposition, and triacylglycerol accumulation in the fat body. However, ATP synthases knockdown had only a slight effect on the amount of ATP in the fat body. These results support the hypothesis that ATP synthase has a direct role in lipid metabolism and lipophorin physiology, which are not directly due to changes in energy metabolism.

Using axenic and gnotobiotic insects to examine the role of different microbes on the development and reproduction of the kissing bug Rhodnius prolixus (Hemiptera: Reduviidae).

Kissing bugs (Hempitera: Reduviidae) are obligately and exclusively blood feeding insects. Vertebrate blood is thought to provide insufficient B vitamins to insects, which rely on symbiotic relationships with bacteria that provision these nutrients. Kissing bugs harbour environmentally acquired bacteria in their gut lumen, without which they are unable to develop to adulthood. Rhodococcus rhodnii was initially identified as the sole symbiont of Rhodnius prolixus, but modern studies of the kissing bug microbiome suggest that R. rhodnii is not always present or abundant in wild-caught individuals. We asked whether R. rhodnii or other bacteria alone could function as symbionts of R. prolixus. We produced insects with no bacteria (axenic) or with known microbiomes (gnotobiotic). Gnotobiotic insects harbouring R. rhodnii alone developed faster, had higher survival, and laid more eggs than those harbouring other bacterial monocultures, including other described symbionts of kissing bugs. R. rhodnii grew to high titre in the guts of R. prolixus while other tested species were found at much lower abundance. Rhodococcus species tested had nearly identical B vitamin biosynthesis genes, and dietary supplementation of B vitamins had a relatively minor effect on development and survival of gnotobiotic R. prolixus. Our results indicate that R. prolixus have a higher fitness when harbouring R. rhodnii than other bacteria tested, that this may be due to R. rhodnii existing at higher titres and providing more B vitamins to the host, and that symbiont B vitamin synthesis is probably a necessary but not sufficient function of gut bacteria in kissing bugs.

Deficiency of Acetyl-CoA Carboxylase Impairs Digestion, Lipid Synthesis, and Reproduction in the Kissing Bug Rhodnius prolixus.

Rhodnius prolixus is a hematophagous insect, vector of Chagas disease. After feeding, as blood is slowly digested, amino acids are used as substrates to fuel lipid synthesis, and adult females accumulate lipids in the fat body and produce eggs. In order to evaluate the importance of de novo fatty acid synthesis for this insect metabolism, we generated acetyl-CoA carboxylase (ACC) deficient insects. The knockdown (AccKD) females had delayed blood digestion and a shorter lifespan. Their fat bodies showed reduced de novo lipogenesis activity, did not accumulate triacylglycerol during the days after blood meal, and had smaller lipid droplets. At 10 days after feeding, there was a general decrease in the amounts of neutral lipids and phospholipids in the fat body. In the hemolymph, no difference was observed in lipid composition at 5 days after blood meal, but at day ten, there was an increase in hydrocarbon content and a decrease in phospholipids. Total protein concentration and amino acid composition were not affected. The AccKD females laid 60% fewer eggs than the control ones, and only 7% hatched (89% for control), although their total protein and triacylglycerol contents were not different. Scanning electron microscopy of the egg surface showed that chorion (eggshell) from the eggs laid by the AccKD insects had an altered ultrastructural pattern when compared to control ones. These results show that ACC has a central role in R. prolixus nutrient homeostasis, and its appropriate activity is important to digestion, lipid synthesis and storage, and reproductive success.

An updated catalog of lipocalins of the chagas disease vector Rhodnius prolixus (Hemiptera, Reduviidae)

The haematophagy process by arthropods has been one of the main targets of studies in the parasite-host interaction, and the kissing-bug Rhodnius prolixus, vector of the protozoan Trypanosoma cruzi, has been one of the main models for such studies. Still in the 1980s, it was identified that among the salivary proteins for disrupting vertebrate host homeostasis, lipocalins were among the most relevant proteins for this process. Since then, 30 lipocalins have been identified in the salivary glands of R. prolixus, that promotes vasodilatation, prevents inflammation, act as anticoagulants and inhibits platelet aggregation. The present work aims to identify new lipocalins from R. prolixus, combining transcriptome and genome data. Identified new genes were mapped and had their structure annotated. To infer an evolutionary relationship between lipocalins, and to support the predicted functions for each lipocalin, all amino acid sequences were used to construct phylogenetic trees. We identified a total of 29 new lipocalins, 3 new bioaminogenic-biding proteins (which act to inhibit platelet aggregation and vasodilation), 9 new inhibitors of platelet aggregation, 7 new apolipoproteins and 10 lipocalins with no putative function. In addition, we observed that several of the lipocalins are also expressed in different R. prolxius tissues, including gut, central nervous system, antennae, and reproductive organs. In addition to newly identified lipocalins and a mapping the new and old lipocalins in the genome of R. prolixus, our study also carried out a review on functional status and nomenclature of some of the already identified lipocalins. Our study reinforces that we are far from understanding the role of lipocalins in the physiology of R. prolixus, and that studies of this family are still of great relevance.

The Fate of Dietary Cholesterol in the Kissing Bug Rhodnius prolixus.

Insects are unable to synthesize cholesterol and depend on the presence of sterols in the diet for cell membrane composition and hormone production. Thus, cholesterol absorption, transport, and metabolism are potential targets for vector and pest control strategies. Here, we investigate the dietary cholesterol absorption and tissue distribution in the kissing bug Rhodnius prolixus using radiolabeled cholesterol. Both the anterior and posterior midguts absorbed cholesterol from the ingested blood, although the anterior midgut absorbed more. We also observed esterified cholesterol labeling in the epithelium, indicating that midgut cells can metabolize and store cholesterol. Only a small amount of labeled cholesterol was found in the hemolymph, where it was mainly in the free form and associated with lipophorin (Lp). The fat body transiently accumulated cholesterol, showing a labeled cholesterol peak on the fifth day after the blood meal. The ovaries also incorporated cholesterol, but cumulatively. The insects did not absorb almost half of the ingested labeled cholesterol, and radioactivity was present in the feces. After injection of 3H-cholesterol-labeled Lp into females, a half-life of 5.5 ± 0.7 h in the hemolymph was determined. Both the fat body and ovaries incorporated Lp-associated cholesterol, which was inhibited at low temperature, indicating the participation of active cholesterol transport. These results help describe an unexplored part of R. prolixus lipid metabolism.

"Urate and NOX5 Control Blood Digestion in the Hematophagous Insect Rhodnius prolixus".

Low levels of reactive oxygen species (ROS) are now recognized as essential players in cell signaling. Here, we studied the role of two conserved enzymes involved in redox regulation that play a critical role in the control of ROS in the digestive physiology of a blood-sucking insect, the kissing bug Rhodnius prolixus. RNAi-mediated silencing of RpNOX5 and RpXDH induced early mortality in adult females after a blood meal. Recently, a role for RpNOX5 in gut motility was reported, and here, we show that midgut peristalsis is also under the control of RpXDH. Together with impaired peristalsis, silencing either genes impaired egg production and hemoglobin digestion, and decreased hemolymph urate titers. Ultrastructurally, the silencing of RpNOX5 or RpXDH affected midgut cells, changing the cells of blood-fed insects to a phenotype resembling the cells of unfed insects, suggesting that these genes work together in the control of blood digestion. Injection of either allopurinol (an XDH inhibitor) or uricase recapitulated the gene silencing effects, suggesting that urate itself is involved in the control of blood digestion. The silencing of each of these genes influenced the expression of the other gene in a complex way both in the unfed state and after a blood meal, revealing signaling crosstalk between them that influences redox metabolism and nitrogen excretion and plays a central role in the control of digestive physiology.

Gene annotation of nuclear receptor superfamily genes in the kissing bug Rhodnius prolixus and the effects of 20-hydroxyecdysone on lipid metabolism.

The hormone 20-hydroxyecdysone is fundamental for regulating moulting and metamorphosis in immature insects, and it plays a role in physiological regulation in adult insects. This hormone acts by binding and activating a receptor, the ecdysone receptor, which is part of the nuclear receptor gene superfamily. Here, we analyse the genome of the kissing bug Rhodnius prolixus to annotate the nuclear receptor superfamily genes. The R. prolixus genome displays a possible duplication of the HNF4 gene. All the analysed insect organs express most nuclear receptor genes as shown by RT-PCR. The quantitative PCR analysis showed that the RpEcR and RpUSP genes are highly expressed in the testis, while the RpHNF4-1 and RpHNF4-2 genes are more active in the fat body and ovaries and in the anterior midgut, respectively. Feeding does not induce detectable changes in the expression of these genes in the fat body. However, the expression of the RpHNF4-2 gene is always higher than that of RpHNF4-1. Treating adult females with 20-hydroxyecdysone increased the amount of triacylglycerol stored in the fat bodies by increasing their lipogenic capacity. These results indicate that 20-hydroxyecdysone acts on the lipid metabolism of adult insects, although the underlying mechanism is not clear.

Impact of alkaloids in food consumption, metabolism and survival in a blood-sucking insect

The sense of taste provides information about the “good” or “bad” quality of a food source, which may be potentially nutritious or toxic. Most alkaloids taste bitter to humans, and because bitter taste is synonymous of noxious food, they are generally rejected. This response may be due to an innate low palatability or due to a malaise that occurs after food ingestion, which could even lead to death. We investigated in the kissing bug Rhodnius prolixus, whether alkaloids such as quinine, caffeine and theophylline, are merely distasteful, or if anti-appetitive responses are caused by a post-ingestion physiological effect, or both of these options. Although anti-appetitive responses were observed for the three alkaloids, only caffeine and theophylline affect metabolic and respiratory parameters that reflected an underlying physiological stress following their ingestion. Furthermore, caffeine caused the highest mortality. In contrast, quinine appears to be a merely unpalatable compound. The sense of taste helps insects to avoid making wrong feeding decisions, such as the intake of bitter/toxic foods, and thus avoid potentially harmful effects on health, a mechanism preserved in obligate hematophagous insects.

Expression of acyl-CoA-binding protein 5 fromRhodnius prolixusand its inhibition by RNA interference.

The acyl-CoA-binding proteins (ACBP) act by regulating the availability of acyl-CoA in the cytoplasm and must have essential functions in lipid metabolism. The genome of the kissing-bug Rhodnius prolixus encodes five proteins of this family, but little is known about them. In this study we investigated the expression and function of RpACBP-5. Feeding induced RpACBP-5 gene expression in the posterior midgut, and an increase of about four times was observed two days after the blood meal. However, the amount of protein, which was only detected in this organ, did not change during digestion. The RpACBP-5 gene was also highly expressed in pre-vitellogenic and vitellogenic oocytes. Recombinant RpACBP-5 was shown to bind to acyl-CoA of different lengths, and it exhibited nanomolar affinity to lauroyl-CoA in an isothermal titration assay, indicating that RpACBP-5 is a functional ACBP. RpACBP-5 knockdown by RNA interference did not affect digestion, egg laying and hatching, survival, or accumulation of triacylglycerol in the fat body and oocytes. Similarly, double knockdown of RpACBP-1 and RpACBP-5 did not alter egg laying and hatching, survival, accumulation of triacylglycerol in the fat body and oocytes, or the neutral lipid composition of the posterior midgut or hemolymph. These results show that RpACBP-5 is a functional ACBP but indicate that the lack of a detectable phenotype in the knockdown insects may be a consequence of functional overlap of the proteins of the ACBP family found in the insect.

Multiple Roles of the Polycistronic Gene Tarsal-less/Mille-Pattes/Polished-Rice During Embryogenesis of the Kissing Bug Rhodnius prolixus

Genes encoding small open-reading frames (smORFs) have been characterized as essential players of developmental processes. The smORF tarsaless/mille-pattes/polished-rice has been thoroughly investigated in holometabolous insects, such as the fruit fly Drosophila melanogaster and the red flour beetle Tribolium castaneum, while its function in hemimetabolous insects was only recently investigated. Thus, we analyzed the function of the tal/pri/mlpt ortholog in a hemimetabolous insect, the kissing bug Rhodnius prolixus (Rp). Sequence analysis shows that Rp-mlpt polycistronic mRNA encodes two small peptides (11 to 14 amino acids) containing a LDPTG motif. Interestingly, a new hemipteran-specific conserved peptide of approximately 80 amino acids was also identified by in silico analysis. In silico docking analysis supports the binding of the small Mlpt peptides to the transcription factor Shavenbaby. Rp-mlpt in situ hybridization and knockdown via RNA interference showed a conserved role of Rp-mlpt during embryogenesis, with a major role in the regulation of thoracic versus abdominal segmentation, leg development and head formation. Altogether, our study shows that mlpt segmentation role is conserved in the common ancestor of Paraneoptera and suggests that polycistronic genes might generate order specific smORFs.

The neuropeptide CCHamide2 regulates diuresis in the Chagas disease vector Rhodnius prolixus.

Given that hematophagous insects ingest large quantities of blood in a single meal, they must undergo a rapid post-prandial diuresis in order to maintain homeostasis. In the kissing bug Rhodnius prolixus (Hemiptera: Reduviidae), the coordinated activity of the Malpighian tubules and anterior midgut maintains water and ion balance during the post-prandial diuresis. Three to four hours after the meal, the diuretic process finishes, and the animal enters an antidiuretic state to ensure water conservation until the next blood intake. The diuretic and antidiuretic processes are tightly regulated by serotonin and neuropeptides in this insect. In the present work, we report that the neuropeptide precursor CCHamide2 is involved in the regulation of the post-prandial diuresis in R. prolixus Our results suggest a dual effect of RhoprCCHamide2 peptide, enhancing the serotonin-induced secretion by Malpighian tubules, and inhibiting serotonin-induced absorption across the anterior midgut. To our knowledge, this is the first report of a hormone presenting opposite effects in the two osmoregulatory organs (i.e. midgut and Malpighian tubules) in insects, probably reflecting the importance of a well-tuned diuretic process in hematophagous insects during different moments after the blood meal.

Nezara viridula (Hemiptera: Pentatomidae) transcriptomic analysis and neuropeptidomics

Stinkbugs (Hemiptera: Pentatomidae) are of major economic importance as pest of crops. Among the species composing the stinkbug complex, Nezara viridula is one of the most abundant in Brazil, Argentina and the Southern USA. However, this species has been poorly characterized at the genetic and physiological level. Here we sequenced and analyzed the complete transcriptome of N. viridula male and female adults. We identified neuropeptide precursor genes and G-protein coupled receptors for neuropeptides in this transcriptome. Mature neuropeptides were identified in N. viridula brain extracts by liquid chromatography-tandem mass spectrometry. We also analyzed the neuropeptide precursor complement in the genome sequence of Halyomorpha halys, another pentatomid of economic relevance. We compared the results in both pentatomids with the well-characterized neuropeptide repertoire from the kissing bug Rhodnius prolixus (Hemiptera: Reduviidae). We identified both group-specific features (which could be related to the different feeding habits) and similarities that could be characteristic of Heteroptera. This work contributes to a deeper knowledge of the genetic information of these pests, with a focus on neuroendocrine system characterization.

Learning Spatial Aversion Is Sensory-Specific in the Hematophagous Insect Rhodnius prolixus.

Even though innate behaviors are essential for assuring quick responses to expected stimuli, experience-dependent behavioral plasticity confers an advantage when unexpected conditions arise. As being rigidly responsive to too many stimuli can be biologically expensive, adapting preferences to time-dependent relevant environmental conditions provide a cheaper and wider behavioral reactivity. According to their specific life habits, animals prioritize different sensory modalities to maximize environment exploitation. Besides, when mediating learning processes, the salience of a stimulus usually plays a relevant role in determining the intensity of an association. Then, sensory prioritization might reflect an heterogeneity in the cognitive abilities of an individual. Here, we analyze in the kissing bug Rhodnius prolixus if stimuli from different sensory modalities generate different cognitive capacities under an operant aversive paradigm. In a 2-choice walking arena, by registering the spatial distribution of insects over an experimental arena, we evaluated firstly the innate responses of bugs confronted to mechanical (rough substrate), visual (green light), thermal (32°C heated plate), hygric (humidified substrate), gustatory (sodium chloride), and olfactory (isobutyric acid) stimuli. In further experimental series bugs were submitted to an aversive operant conditioning by pairing each stimulus with a negative reinforcement. Subsequent tests allowed us to analyze if the innate behaviors were modulated by such previous aversive experience. In our experimental setup mechanical and visual stimuli were neutral, the thermal cue was attractive, and the hygric, gustatory and olfactory ones were innately aversive. After the aversive conditioning, responses to the mechanical, the visual, the hygric and the gustatory stimuli were modulated while responses to the thermal and the olfactory stimuli remained rigid. We present evidences that the spatial learning capacities of R. prolixus are dependent on the sensory modality of the conditioned stimulus, regardless their innate valence (i.e., neutral, attractive, or aversive). These differences might be given by the biological relevance of the stimuli and/or by evolutionary aspects of the life traits of this hematophagous insect.

Orcokinins regulate the expression of neuropeptide precursor genes related to ecdysis in the hemimetabolous insect Rhodnius prolixus

Ecdysis is a vital process for insects, during which they shed the old cuticle in order to emerge as the following developmental stage. Given its relevance for survival and reproduction, ecdysis is tightly regulated by peptidic hormones that conform an interrelated neuromodulatory network. This network was studied in species that undergo a complete metamorphosis, but not in hemimetabola. In a recent work, we demonstrated that orcokinin neuropeptides are essential for ecdysis to occur in the kissing bug Rhodnius prolixus. Here we performed gene silencing, quantitative PCR and in vitro treatments in order to study the interrelationships between RhoprOKs and hormones such as ecdysis triggering hormone, corazonin, eclosion hormone, crustacean cardioactive peptide and ecdysone. Our results suggest that RhoprOKs directly or indirectly regulate the expression of other genes. Whereas RhoprOKA is centrally involved in the regulation of gene expression, RhoprOKB is implicated in processes related to midgut physiology. Therefore, we propose that the different transcripts encoded in RhoprOK gene could integrate signaling cues, in order to coordinate the nutritional state with development and ecdysis. Given the emerging data that point to OKs as important factors for survival and reproduction, they could be candidates in the search for new insect management strategies based on neuroendocrine targets.

Under-Expression of Chemosensory Genes in Domiciliary Bugs of the Chagas Disease Vector Triatoma brasiliensis.

BackgroundIn Latin America, the bloodsucking bugs Triatominae are vectors of Trypanosoma cruzi, the parasite that causes Chagas disease. Chemical elimination programs have been launched to control Chagas disease vectors. However, the disease persists because native vectors from sylvatic habitats are able to (re)colonize houses—a process called domiciliation. Triatoma brasiliensis is one example. Because the chemosensory system allows insects to interact with their environment and plays a key role in insect adaption, we conducted a descriptive and comparative study of the chemosensory transcriptome of T. brasiliensis samples from different ecotopes.Methodology/Principal FindingIn a reference transcriptome built using de novo assembly, we found transcripts encoding 27 odorant-binding proteins (OBPs), 17 chemosensory proteins (CSPs), 3 odorant receptors (ORs), 5 transient receptor potential channel (TRPs), 1 sensory neuron membrane protein (SNMPs), 25 takeout proteins, 72 cytochrome P450s, 5 gluthatione S-transferases, and 49 cuticular proteins. Using protein phylogenies, we showed that most of the OBPs and CSPs for T. brasiliensis had well supported orthologs in the kissing bug Rhodnius prolixus. We also showed a higher number of these genes within the bloodsucking bugs and more generally within all Hemipterans compared to the other species in the super-order Paraneoptera. Using both DESeq2 and EdgeR software, we performed differential expression analyses between samples of T. brasiliensis, taking into account their environment (sylvatic, peridomiciliary and domiciliary) and sex. We also searched clusters of co-expressed contigs using HTSCluster. Among differentially expressed (DE) contigs, most were under-expressed in the chemosensory organs of the domiciliary bugs compared to the other samples and in females compared to males. We clearly identified DE genes that play a role in the chemosensory system.Conclusion/SignificanceChemosensory genes could be good candidates for genes that contribute to adaptation or plastic rearrangement to an anthropogenic system. The domiciliary environment probably includes less diversity of xenobiotics and probably has more stable abiotic parameters than do sylvatic and peridomiciliary environments. This could explain why both detoxification and cuticle protein genes are less expressed in domiciliary bugs. Understanding the molecular basis for how vectors adapt to human dwellings may reveal new tools to control disease vectors; for example, by disrupting chemical communication.