RESUMEN
In triatomines, blood-feeding triggers many physiological processes including post embryonic development and reproduction. Different feeding habits, such as hematophagy, can shape gene functions to meet the challenges of each type of diet. The gut of blood-sucking insects faces particular challenges after feeding due to the quantity and the quality of the food ingested. A comparison of transcriptomic and proteomic data indicates that post transcriptional regulation of gene expression is crucial in the triatomine gut. It was proposed that eukaryotic translation initiation factor 3 subunit m (eIF3m) and eIF3e define 2 different eIF3 complexes with a distinct affinity for the different mRNAs, thus selecting the set of mRNAs to be translated and constituting a post transcriptional mode of regulation of gene expression. Because the eIF3m is mainly expressed in the gut, we evaluated its relevance in Rhodnius prolixus physiology through RNA interference-mediated gene silencing. The knockdown of eIF3m reduced the digestion rate, affecting the processes triggered by a blood meal. Its silencing inhibited molting and caused premature death in nymphs while impaired ovary development, oviposition and increased resistance to starvation in adult females. The survival of males after feeding (resistance to starvation) was not affected by eIF3m knockdown. The information regarding the eIF3m function in insects is scarce and the phenotypes observed in R. prolixus upon eIF3m silencing are different and more severe than those previously described in Drosophila melanogaster, indicating a pleiotropic role of this gene in triatomines.
RESUMEN
In ecdysozoan animals, moulting entails the production of a new exoskeleton and shedding of the old one during ecdysis. It is induced by a pulse of ecdysone that regulates the expression of different hormonal receptors and activates a peptide-mediated signalling cascade. In Holometabola, the peptidergic cascade regulating ecdysis has been well described. However, very little functional information regarding the neuroendocrine regulation of ecdysis is available for Hemimetabola, which display an incomplete metamorphosis. We use Rhodnius prolixus as a convenient experimental model to test two hypotheses: (1) the role of neuropeptides that regulate ecdysis in Holometabola is conserved in hemimetabolous insects; and (2) the neuropeptides regulating ecdysis play a role in the regulation of female reproduction during the adult stage. The RNA interference-mediated reduction of ecdysis triggering hormone (ETH) mRNA levels in fourth-instar nymphs resulted in lethality at the expected time of ecdysis. Unlike in holometabolous insects, knockdown of eth and orcokinin isoform A (oka) did not affect oviposition in adult females, pointing to a different endocrine regulation of ovary maturation. However, eth knockdown prevented egg hatching. The blockage of egg hatching appears to be a consequence of embryonic ecdysis failure. Most of the first-instar nymphs hatched from the eggs laid by females injected with dsRNA for eclosion hormone (dsEH), crustacean cardioactive peptide (dsCCAP) and dsOKA died at the expected time of ecdysis, indicating the crucial involvement of these genes in post-embryonic development. No phenotypes were observed upon corazonin (cz) knockdown in nymphs or adult females. The results are relevant for evolutionary entomology and could reveal targets for neuropeptide-based pest control tools.
Asunto(s)
Neuropéptidos , Rhodnius , Animales , Femenino , Metamorfosis Biológica , Muda/fisiología , Neuropéptidos/metabolismo , Reproducción , Rhodnius/genéticaRESUMEN
BACKGROUND: Blood-sucking insects incorporate many times their body weight of blood in a single meal. Because proteins are the major component of vertebrate blood, its digestion in the gut generates extremely high concentrations of free amino acids. Previous reports showed that the tyrosine degradation pathway plays an essential role in adapting these animals to blood feeding. Inhibition of 4-hydroxyphenylpyruvate dioxygenase (HPPD), the rate-limiting step of tyrosine degradation, results in the death of insects after a blood meal. Therefore, it has been suggested that compounds that block the catabolism of tyrosine could act selectively on blood-feeding insects. Here, we evaluated the toxicity against mosquitoes of three HPPD inhibitors currently used as herbicides and in human health. RESULTS: Of the compounds tested, nitisinone (NTBC) proved to be more potent than mesotrione (MES) and isoxaflutole (IFT) in Aedes aegypti. NTBC was lethal to Ae. aegypti in artificial feeding assays [median lethal dose (LD50 ): 4.53 µm] and in topical application (LD50 : 0.012 nmol/mosquito). NTBC was also lethal to Ae. aegypti populations that were resistant to neurotoxic insecticides, and to other mosquito species (Anopheles and Culex). CONCLUSION: HPPD inhibitors, particularly NTBC, represent promising new drugs for mosquito control. Because they affect only blood-feeding organisms, they represent a safer and more environmentally friendly alternative to conventional neurotoxic insecticides. © 2021 Society of Chemical Industry.
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4-Hidroxifenilpiruvato Dioxigenasa , Aedes , Culex , Insecticidas , 4-Hidroxifenilpiruvato Dioxigenasa/antagonistas & inhibidores , Animales , Humanos , Control de Mosquitos , Mosquitos VectoresRESUMEN
The use of chemical insecticides is the main control method for Blattella germanica worldwide. The prolonged and frequent use of insecticides produced the selection of insecticide-resistant individuals. The German cockroach is one of the most widespread urban pests in Argentina. In the last decades, resistance monitoring studies in this country demonstrated that there is a high prevalence of pyrethroid-resistant populations of B. germanica in the field. In this work, we studied the resistance mechanisms of a field-collected strain of B. germanica at toxicological, enzymatic, and molecular levels. A resistance ratio of 100 was obtained for the resistant strain when it was exposed to ß-cypermethrin. The pretreatment with specific synergists (piperonyl butoxide and triphenyl phosphate) led to a significant increase in the toxicity of the pyrethroid, suggesting an involvement of oxidases and esterases in the detoxification of this insecticide. Moreover, esterase and oxidase activities in the resistant strain were 1.5-fold and 2-fold higher respectively, compared to the susceptible individuals. On the other hand, the voltage-gated sodium channel gene of the resistant cockroaches did not show nucleotidic substitutions in the domain II which are associated to knockdown resistance in this species. These results suggest that the main mechanism of resistance of the studied cockroaches' strain is metabolic, mainly due to an increase in the activity of oxidase and esterase enzymes. The results of this work in addition to other reports found in literature show that the extended use of a single active principle for cockroach control promotes the development of resistance leading to control failure in the field. In contrast, integrated pest management strategies include the use of different control tools in addition to chemical insecticides, which delay the appearance of resistance increasing the efficacy of pest control.
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Blattellidae , Insecticidas , Piretrinas , Animales , Argentina , Resistencia a los Insecticidas/genética , Insecticidas/farmacología , Piretrinas/farmacologíaRESUMEN
Tsetse transmit African trypanosomiasis, which is a disease fatal to both humans and animals. A vaccine to protect against this disease does not exist so transmission control relies on eliminating tsetse populations. Although neurotoxic insecticides are the gold standard for insect control, they negatively impact the environment and reduce populations of insect pollinator species. Here we present a promising, environment-friendly alternative to current insecticides that targets the insect tyrosine metabolism pathway. A bloodmeal contains high levels of tyrosine, which is toxic to haematophagous insects if it is not degraded and eliminated. RNA interference (RNAi) of either the first two enzymes in the tyrosine degradation pathway (tyrosine aminotransferase (TAT) and 4-hydroxyphenylpyruvate dioxygenase (HPPD)) was lethal to tsetse. Furthermore, nitisinone (NTBC), an FDA-approved tyrosine catabolism inhibitor, killed tsetse regardless if the drug was orally or topically applied. However, oral administration of NTBC to bumblebees did not affect their survival. Using a novel mathematical model, we show that NTBC could reduce the transmission of African trypanosomiasis in sub-Saharan Africa, thus accelerating current disease elimination programmes.
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Ciclohexanonas/uso terapéutico , Reposicionamiento de Medicamentos , Control de Infecciones/métodos , Nitrobenzoatos/uso terapéutico , Tripanosomiasis Africana/prevención & control , 4-Hidroxifenilpiruvato Dioxigenasa/antagonistas & inhibidores , 4-Hidroxifenilpiruvato Dioxigenasa/metabolismo , Animales , Abejas/efectos de los fármacos , Femenino , Humanos , Insecticidas/uso terapéutico , Masculino , Metaboloma/efectos de los fármacos , Ratones , Modelos Teóricos , Enfermedades Desatendidas/prevención & control , Producción de Medicamentos sin Interés Comercial , Ratas , Ratas Wistar , Pruebas de Toxicidad , Tripanosomiasis Africana/transmisión , Moscas Tse-Tse/efectos de los fármacos , Moscas Tse-Tse/metabolismo , Tirosina/metabolismoRESUMEN
The Indian meal moth Plodia interpunctella is a pest of stored products worldwide. Plant-derived essential oils with insecticidal activity could be safe products to control this species. The scarce information about the mode of action of most plant-derived products limits their use for the control of insect pests. Here, we demonstrate that an essential oil distilled from Lippia turbinata ("poleo") has insecticidal activity on P. interpunctella larvae. Furthermore, we performed a comprehensive characterization of P. interpunctella neuroendocrine system, in comparison with other lepidopteran species.
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Lippia/química , Mariposas Nocturnas/efectos de los fármacos , Aceites de Plantas/toxicidad , Animales , Insecticidas , Larva/efectos de los fármacos , Mariposas Nocturnas/crecimiento & desarrollo , Mariposas Nocturnas/fisiología , Sistemas NeurosecretoresRESUMEN
BACKGROUND: Rhodnius prolixus has become a model for revealing the molecular bases of insect sensory biology due to the publication of its genome and its well-characterized behavioural repertoire. Gene expression modulation underlies behaviour-triggering processes at peripheral and central levels. Still, the regulation of sensory-related gene transcription in sensory organs is poorly understood. Here we study the genetic bases of plasticity in antennal sensory function, using R. prolixus as an insect model. RESULTS: Antennal expression of neuromodulatory genes such as those coding for neuropeptides, neurohormones and their receptors was characterized in fifth instar larvae and female and male adults by means of RNA-Sequencing (RNA-Seq). New nuclear receptor and takeout gene sequences were identified for this species, as well as those of enzymes involved in the biosynthesis and processing of neuropeptides and biogenic amines. CONCLUSIONS: We report a broad repertoire of neuromodulatory and neuroendocrine-related genes expressed in the antennae of R. prolixus and suggest that they may serve as the local basis for modulation of sensory neuron physiology. Diverse neuropeptide precursor genes showed consistent expression in the antennae of all stages studied. Future studies should characterize the role of these modulatory components acting over antennal sensory processes to assess the relative contribution of peripheral and central regulatory systems on the plastic expression of insect behaviour.
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Perfilación de la Expresión Génica/veterinaria , Proteínas de Insectos/genética , Rhodnius/crecimiento & desarrollo , Animales , Antenas de Artrópodos/química , Femenino , Regulación del Desarrollo de la Expresión Génica , Larva , Masculino , Neuropéptidos/genética , Neurotransmisores/genética , Filogenia , Receptores de Neuropéptido/genética , Receptores de Neurotransmisores/genética , Rhodnius/genética , Análisis de Secuencia de ARN/veterinariaRESUMEN
Cuticle tanning occurs in insects immediately after hatching or molting. During this process, the cuticle becomes dark and rigid due to melanin deposition and protein crosslinking. In insects, different from mammals, melanin is synthesized mainly from dopamine, which is produced from DOPA by the enzyme DOPA decarboxylase. In this work, we report that the silencing of the RpAadc-2 gene, which encodes the putative Rhodnius prolixus DOPA decarboxylase enzyme, resulted in a reduction in nymph survival, with a high percentage of treated insects dying during the ecdysis process or in the expected ecdysis period. Those treated insects that could complete ecdysis presented a decrease in cuticle pigmentation and hardness after molting. In adult females, the knockdown of AADC-2 resulted in a reduction in the hatching of eggs; the nymphs that managed to hatch failed to tan the cuticle and were unable to feed. Despite the failure in cuticle tanning, knockdown of the AADC-2 did not increase the susceptibility to topically applied deltamethrin, a pyrethroid insecticide. Additionally, our results showed that the melanin synthesis pathway did not play a major role in the detoxification of the excess (potentially toxic) tyrosine from the diet, an essential trait for hematophagous arthropod survival after a blood meal.
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Dopa-Decarboxilasa/fisiología , Proteínas de Insectos/fisiología , Integumento Común/fisiología , Rhodnius/enzimología , Rhodnius/crecimiento & desarrollo , Animales , Dopa-Decarboxilasa/farmacología , Femenino , Inactivación Metabólica , Proteínas de Insectos/farmacología , Insecticidas , Melaninas/metabolismo , Muda/fisiología , Nitrilos , Pigmentación/fisiología , Piretrinas , Reproducción , Rhodnius/genética , Tirosina/metabolismoRESUMEN
The phenylalanine/tyrosine degradation pathway is frequently described as a catabolic pathway that funnels aromatic amino acids into citric acid cycle intermediates. Previously, we demonstrated that the accumulation of tyrosine generated during the hydrolysis of blood meal proteins in Rhodnius prolixus is potentially toxic, a harmful outcome that is prevented by the action of the first two enzymes in the tyrosine degradation pathway. In this work, we further evaluated the relevance of all other enzymes involved in phenylalanine/tyrosine metabolism in the physiology of this insect. The knockdown of most of these enzymes produced a wide spectrum of distinct phenotypes associated with reproduction, development and nymph survival, demonstrating a highly pleiotropic role of tyrosine metabolism. The phenotypes obtained for two of these enzymes, homogentisate dioxygenase and fumarylacetoacetase, have never before been described in any arthropod. To our knowledge, this report is the first comprehensive gene-silencing analysis of an amino acid metabolism pathway in insects. Amino acid metabolism is exceptionally important in haematophagous arthropods due to their particular feeding behaviour.
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Monofenol Monooxigenasa/genética , Rhodnius/enzimología , Tirosina/metabolismo , Animales , Silenciador del Gen , Redes y Vías Metabólicas , Fenotipo , Rhodnius/genéticaRESUMEN
Vertebrate blood composition is heavily biased towards proteins, and hemoglobin, which is a hemeprotein, is by far the most abundant protein. Typically, hematophagous insects ingest blood volumes several times their weight before the blood meal. This barbarian feast offers an abundance of nutrients, but the degradation of blood proteins generates toxic concentrations of amino acids and heme, along with unparalleled microbiota growth. Despite this challenge, hematophagous arthropods have successfully developed mechanisms that bypass the toxicity of these molecules. While these adaptations allow hematophagous arthropods to tolerate their diet, they also constitute a unique mode of operation for cell signaling, immunity, and metabolism, the study of which may offer insights into the biology of disease vectors and may lead to novel vector-specific control methods.
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Vectores Artrópodos/metabolismo , Artrópodos/metabolismo , Hemoproteínas/metabolismo , Fenómenos Fisiológicos de la Nutrición/fisiología , Adaptación Fisiológica , Animales , Vectores Artrópodos/inmunología , Vectores Artrópodos/microbiología , Artrópodos/inmunología , Artrópodos/microbiología , Conducta Alimentaria/fisiología , Transducción de Señal/fisiologíaRESUMEN
To grow and develop insects must undergo ecdysis. During this process, the individual sheds the old cuticle to emerge as the following developmental stage. During ecdysis, different programed behaviors are regulated by neuropeptidergic pathways. In general, components of these pathways are better characterized in crustacean and holometabolous insects than in hemimetabola. In insects, the orkoninin gene produces two different neuropeptide precursors by alternative splicing: orcokinin A and orcokinin B. Although orcokinins are well conserved in insect species, their physiological role remains elusive. Here we describe a new splicing variant of the orcokinin gene in the hemimetabolous triatomine Rhodnius prolixus. We further analyze the expression pattern and the function of the alternatively spliced RhoprOK transcripts by means of immunohistochemistry and RNAi-mediated gene silencing. Our results indicate that orkoninis play an essential role in the peptidergic signaling pathway regulating ecdysis in the hemimetabolous insect Rhodnius prolixus.
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Muda , Neuropéptidos/metabolismo , Rhodnius/crecimiento & desarrollo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Femenino , Masculino , Datos de Secuencia Molecular , Interferencia de ARN , Rhodnius/metabolismoRESUMEN
Chagas' disease, affecting up to 6-7 million people worldwide, is transmitted to humans through the feces of triatomine kissing bugs. From these, Rhodnius prolixus, Triatoma dimidiata, Triatoma infestans and Triatoma pallidipennis are important vectors distributed throughout the Latin American subcontinent. Resistance to pyrethroids has been developed by some triatomine populations, especially T. infestans, obstructing their control. Given their role in the regulation of physiological processes, neuroendocrine-derived factors have been proposed as a source of molecular targets for new-generation insecticides. However, the involvement of neuropeptides in insecticide metabolism and resistance in insects has been poorly studied. In the present work, the sequences of 20 neuropeptide precursor genes in T. infestans, 16 in T. dimidiata, and 13 in T. pallidipennis detected in transcriptomic databases are reported, and a comparative analysis in triatomines is presented. A total of 59 neuropeptides were validated by liquid chromatography-tandem mass spectrometry in brain and nervous ganglia from T. infestans, revealing the existence of differential post-translational modifications, extended and truncated forms. The results suggest a high sequence conservation in some neuropeptide systems in triatomines, whereas remarkable differences occur in several others within the core domains. Comparisons of the basal expression levels for several neuropeptide precursor genes between pyrethroid sensitive and resistant population of T. infestans are also presented here, in order to introduce a proof of concept to test the involvement of neuropeptides in insecticide resistance. From the precursors tested, NVP and ITG peptides are significantly higher expressed in the resistant population. To our knowledge, this is the first report to associate differential neuropeptide expression with insecticide resistance. The information provided here contributes to creating conditions to widely extend functional and genetic studies involving neuropeptides in triatomines.
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Neuropéptidos/genética , Neuropéptidos/metabolismo , Triatoma/genética , Triatoma/metabolismo , Animales , Perfilación de la Expresión Génica , Resistencia a los Insecticidas/genética , Especificidad de la Especie , Triatoma/clasificaciónRESUMEN
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.
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4-Hidroxifenilpiruvato Dioxigenasa/genética , Silenciador del Gen , Proteínas de Insectos/genética , Rhodnius/genética , Tirosina Transaminasa/genética , Tirosina/metabolismo , 4-Hidroxifenilpiruvato Dioxigenasa/metabolismo , Animales , Femenino , Inactivación Metabólica , Proteínas de Insectos/metabolismo , Masculino , Ninfa/genética , Ninfa/crecimiento & desarrollo , Ninfa/metabolismo , Rhodnius/crecimiento & desarrollo , Rhodnius/metabolismo , Tirosina Transaminasa/metabolismoRESUMEN
The importance of Chagas disease motivated the scientific effort to obtain the complete genomic sequence of the vector species Rhodnius prolixus, this information is also relevant to the understanding of triatomine biology in general. The central nervous system is the key regulator of insect physiology and behavior. Neurohormones (neuropeptides and biogenic amines) are the chemical messengers involved in the regulation and integration of neuroendocrine signals. In insects, this signaling is mainly mediated by the interaction of neurohormone ligands with G protein coupled receptors (GPCRs). The recently sequenced R. prolixus genome provides us with the opportunity to analyze this important family of genes in triatomines, supplying relevant information for further functional studies. Next-generation sequencing methods offer an excellent opportunity for transcriptomic exploration in key organs and tissues in the presence of a reference genome as well as when a reference genome is not available. We undertook a genomic analysis to obtain a genome-wide inventory of opsines and the GPCRs for neurohormones in R. prolixus. Furthermore, we performed a transcriptomic analysis of R. prolixus central nervous system, focusing on neuropeptide precursor genes and neurohormone and opsines GPCRs. In addition, we mined the whole transcriptomes of Triatoma dimidiata, Triatoma infestans and Triatoma pallidipennis - three sanitary relevant triatomine species - to identify neuropeptide precursors and GPCRs genes. Our study reveals a high degree of sequence conservation in the molecular components of the neuroendocrine system of triatomines.
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Genoma de los Insectos , Neuropéptidos/metabolismo , Neurotransmisores/metabolismo , Opsinas/genética , Receptores Acoplados a Proteínas G/genética , Rhodnius/genética , Transcriptoma , Animales , Sistema Nervioso Central/metabolismo , Opsinas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Rhodnius/metabolismoRESUMEN
Metamorphosis, which depends upon a fine balance between two groups of lipid-soluble hormones such as juvenile hormones (JHs) and ecdysteroids, is an important feature in insect evolution. While it is clear that the onset of metamorphosis depends on the decrease of JH levels, the way in which these hormones exert their activities is not fully understood in Triatominae species. The discovery of a Drosophila melanogaster mutant resistant to the treatment with the JH analog methoprene, led finally to the description of the methoprene-tolerant gene in Tribolium castaneum (TcMet) as a putative JH receptor. Here we present the genomic and functional characterization of an ortholog of the methoprene-tolerant gene in the hemimetabolous insect Rhodnius prolixus (RpMet). The analysis of the R. prolixus gene showed that the exonic structure is different from that described for holometabolous species, although all the critical protein motifs are well conserved. Expression analysis showed the presence of RpMet mRNA in all the tested tissues: ovary, testis, rectum, Malpighian tubules and salivary glands. When juvenile individuals were treated with RpMet specific double strand RNA (dsRNA), we observed abnormal molting events that resulted in individuals with morphological alterations (adultoids). Similarly, treatment of newly emerged fed females with dsRNA resulted in an abnormal development of the ovaries, with eggs revealing anomalies in size and accumulation of yolk, as well as a decrease in the amount of heme-binding protein. Altogether, our results validate that RpMet is involved in the transduction of JH signaling, controlling metamorphosis and reproduction in R. prolixus.
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Proteínas Portadoras/metabolismo , Genes de Insecto/fisiología , Genómica/métodos , Hemoproteínas/metabolismo , Resistencia a los Insecticidas , Larva/metabolismo , Metopreno/farmacología , Rhodnius/genética , Secuencia de Aminoácidos , Animales , Femenino , Regulación del Desarrollo de la Expresión Génica , Técnicas de Silenciamiento del Gen , Proteínas de Unión al Hemo , Hormonas Juveniles/farmacología , Larva/efectos de los fármacos , Larva/crecimiento & desarrollo , Metamorfosis Biológica/efectos de los fármacos , Metamorfosis Biológica/genética , Datos de Secuencia Molecular , Oocitos/citología , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Ovario/citología , Ovario/efectos de los fármacos , Ovario/metabolismo , ARN Mensajero/metabolismo , Rhodnius/crecimiento & desarrollo , Rhodnius/metabolismo , Homología de Secuencia de Aminoácido , Transducción de SeñalRESUMEN
In insects, neuropeptides play a central role in the control of most physiological processes. The knowledge and characterization of new neuropeptide families, is of interest on the fields of Genetics, Genomics, Neurobiology, Endocrinology and Evolution. This knowledge also provides the tools for the design of peptidomimetics, pseudopeptides or small molecules, capable of disrupting the physiological processes regulated by the signaling molecules and their receptors. This is a promising target for a novel generation of insecticides. Using database searches, mass spectrometry and RACE-PCR, we identified a neuropeptide precursor transcript encoding a new family of insect neuropeptides in the hemipteran Rhodnius prolixus. We named this precursor Orcokinin B, because is originated by the alternative splicing of the Orcokinin gen. EST and genomic data suggests that Orcokinin B is expressed in the nervous system and gut from several insect species, with the exception of Drosophila sp. (Diptera) and Acyirthosiphon pisum (Hemiptera). Mass spectrometry and RT-PCR confirmed the expression of Orcokinin B in brain and anterior midgut of R. prolixus. Furthermore, we identified orthologues of this new family of peptides in genomic and EST databases from Arachnids and Crustaceans.
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Proteínas de Insectos/genética , Neuropéptidos/genética , Rhodnius/genética , Empalme Alternativo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Sistema Nervioso Central/citología , Sistema Nervioso Central/metabolismo , Tracto Gastrointestinal/citología , Tracto Gastrointestinal/metabolismo , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Datos de Secuencia Molecular , Neuropéptidos/química , Neuropéptidos/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Rhodnius/química , Rhodnius/metabolismo , Alineación de Secuencia , Espectrometría de Masas en TándemRESUMEN
The voltage-gated sodium channel is the target site of pyrethroid insecticides. Point mutations in the domain II region of the channel have been implicated in pyrethroid resistance of several insect species. We identified the sequence of domain II from the para sodium channel in Rhodnius prolixus, a vector of Chagas' disease. With this information, we cloned and sequenced the domain II of the sodium channel from the other main Chagas' disease vector: Triatoma infestans. We also identified the presence of a resistance-conferring mutation (L1014F) in a pyrethroid-resistant population of T. infestans from Argentina, and present a PCR-based method to detect this mutation in individuals from field populations. These findings have important implications for the implementation of strategies for resistance management, and for the rational design of campaigns for the control of Chagas' disease transmission.
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Insectos Vectores/genética , Resistencia a los Insecticidas/genética , Mutación Puntual , Canales de Sodio/genética , Triatoma/genética , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Secuencia de Bases , Enfermedad de Chagas/transmisión , Vectores de Enfermedades , Orden Génico , Insecticidas/farmacología , Datos de Secuencia Molecular , Piretrinas/farmacología , Alineación de SecuenciaRESUMEN
In hematophagous insects, blood intake triggers a prompt response mediated by neuropeptides, which regulates a variety of physiological processes. Here we report a quantitative proteomic analysis of the postfeeding response in the central nervous system of Rhodnius prolixus, a vector of Chagas disease. The concentration of neuropeptides NVP-like, ITG-like, kinin-precursor peptide, and neuropeptide-like precursor 1 (NPLP1) significantly changes in response to blood intake. We also performed a neuropeptidomic analysis of other feeding-related organs, namely salivary glands and gut. We identified NPLP1 in salivary glands and myosuppressin in midgut. This is the first report suggesting a role for NPLP1, involving the peptides processed from this precursor in the hormonal control of the production and/or release of saliva. Our results contribute to the understanding of the postprandial neuroendocrine response in hematophagous and provide important information for physiological and pharmacological studies aimed to the design of next-generation insecticides such as peptidomimetics.
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Conducta Alimentaria , Proteínas de Insectos/metabolismo , Neuropéptidos/metabolismo , Proteómica , Rhodnius/metabolismo , Secuencia de Aminoácidos , Animales , Cromatografía Liquida , Proteínas de Insectos/química , Datos de Secuencia Molecular , Neuropéptidos/química , Rhodnius/fisiología , Espectrometría de Masa por Ionización de Electrospray , Espectrometría de Masas en TándemRESUMEN
Haematophagous insects incorporate a large quantity of blood with each meal, producing a big quantity of urine in a few hours. The activity of the Malpighian tubules (MTs) is facilitated by the increase of the circulation of the haemolymph produced by the increase of the aorta contractions as well as, of the peristaltic waves of the anterior midgut. MTs of Triatoma infestans secrete an allatotropin-like peptide, which has a myostimulatory effect on the hindgut, inducing the mixing and voiding of the content during post-prandial diuresis. We are reporting now the activity of allatotropin (AT) as a cardioacceleratory and a myostimulatory peptide at the level of the anterior midgut. The peptide induced the increase of the rate of contractions of the anterior midgut and the aorta in a wide range of concentrations. The cardioacceleratory effect of AT was dependent on the feeding state of the insects and on the presence of serotonin. The response showed the existence of a differential behavior between sexes, inducing a higher increase on the frequency of contractions, as well as, the width of the aorta in males than in females. Finally, our results suggest that AT interacts with serotonin to facilitate the circulation of haemolymph during post-prandial diuresis.