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1.
PLoS One ; 16(1): e0243992, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33428654

RESUMEN

Insecticide resistance is a worldwide threat for vector control around the world, and Aedes aegypti, the main vector of several arboviruses, is a particular concern. To better understand the mechanisms of resistance, four isofemale strains originally from French Guiana were isolated and analysed using combined approaches. The activity of detoxification enzymes involved in insecticide resistance was assayed, and mutations located at positions 1016 and 1534 of the sodium voltage-gated channel gene, which have been associated with pyrethroid resistance in Aedes aegypti populations in Latin America, were monitored. Resistance to other insecticide families (organophosphates and carbamates) was evaluated. A large-scale proteomic analysis was performed to identify proteins involved in insecticide resistance. Our results revealed a metabolic resistance and resistance associated with a mutation of the sodium voltage-gated channel gene at position 1016. Metabolic resistance was mediated through an increase of esterase activity in most strains but also through the shifts in the abundance of several cytochrome P450 (CYP450s). Overall, resistance to deltamethrin was linked in the isofemale strains to resistance to other class of insecticides, suggesting that cross- and multiple resistance occur through selection of mechanisms of metabolic resistance. These results give some insights into resistance to deltamethrin and into multiple resistance phenomena in populations of Ae. aegypti.


Asunto(s)
Aedes/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Proteínas de Insectos/genética , Resistencia a los Insecticidas/genética , Canales de Sodio Activados por Voltaje/genética , Aedes/efectos de los fármacos , Aedes/genética , Animales , Esterasas/metabolismo , Femenino , Guyana Francesa , Técnicas de Silenciamiento del Gen , Genotipo , Inactivación Metabólica/genética , Proteínas de Insectos/antagonistas & inhibidores , Proteínas de Insectos/metabolismo , Insecticidas/farmacología , Mucosa Intestinal/metabolismo , Nitrilos/farmacología , Oligonucleótidos/metabolismo , Polimorfismo de Nucleótido Simple , Proteoma/análisis , Proteómica , Piretrinas/farmacología , Canales de Sodio Activados por Voltaje/química , Canales de Sodio Activados por Voltaje/metabolismo
2.
PLoS Pathog ; 17(1): e1009199, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33465145

RESUMEN

The insecticidal Cry11Aa and Cyt1Aa proteins are produced by Bacillus thuringiensis as crystal inclusions. They work synergistically inducing high toxicity against mosquito larvae. It was proposed that these crystal inclusions are rapidly solubilized and activated in the gut lumen, followed by pore formation in midgut cells killing the larvae. In addition, Cyt1Aa functions as a Cry11Aa binding receptor, inducing Cry11Aa oligomerization and membrane insertion. Here, we used fluorescent labeled crystals, protoxins or activated toxins for in vivo localization at nano-scale resolution. We show that after larvae were fed solubilized proteins, these proteins were not accumulated inside the gut and larvae were not killed. In contrast, if larvae were fed soluble non-toxic mutant proteins, these proteins were found inside the gut bound to gut-microvilli. Only feeding with crystal inclusions resulted in high larval mortality, suggesting that they have a role for an optimal intoxication process. At the macroscopic level, Cry11Aa completely degraded the gastric caeca structure and, in the presence of Cyt1Aa, this effect was observed at lower toxin-concentrations and at shorter periods. The labeled Cry11Aa crystal protein, after midgut processing, binds to the gastric caeca and posterior midgut regions, and also to anterior and medium regions where it is internalized in ordered "net like" structures, leading finally to cell break down. During synergism both Cry11Aa and Cyt1Aa toxins showed a dynamic layered array at the surface of apical microvilli, where Cry11Aa is localized in the lower layer closer to the cell cytoplasm, and Cyt1Aa is layered over Cry11Aa. This array depends on the pore formation activity of Cry11Aa, since the non-toxic mutant Cry11Aa-E97A, which is unable to oligomerize, inverted this array. Internalization of Cry11Aa was also observed during synergism. These data indicate that the mechanism of action of Cry11Aa is more complex than previously anticipated, and may involve additional steps besides pore-formation activity.


Asunto(s)
Aedes/efectos de los fármacos , Sinergismo Farmacológico , Endotoxinas/metabolismo , Tracto Gastrointestinal/efectos de los fármacos , Proteínas Hemolisinas/metabolismo , Insecticidas/metabolismo , Larva/efectos de los fármacos , Aedes/metabolismo , Animales , /toxicidad , Proteínas Bacterianas , Endotoxinas/genética , Endotoxinas/toxicidad , Tracto Gastrointestinal/metabolismo , Proteínas Hemolisinas/genética , Proteínas Hemolisinas/toxicidad , Insecticidas/toxicidad , Larva/metabolismo , Unión Proteica
3.
PLoS Pathog ; 16(12): e1009068, 2020 12.
Artículo en Inglés | MEDLINE | ID: mdl-33382858

RESUMEN

Originating from African forests, Zika virus (ZIKV) has now emerged worldwide in urbanized areas, mainly transmitted by Aedes aegypti mosquitoes. Although Aedes albopictus can transmit ZIKV experimentally and was suspected to be a ZIKV vector in Central Africa, the potential of this species to sustain virus transmission was yet to be uncovered until the end of 2019, when several autochthonous transmissions of the virus vectored by Ae. albopictus occurred in France. Aside from these few locally acquired ZIKV infections, most territories colonized by Ae. albopictus have been spared so far. The risk level of ZIKV emergence in these areas remains however an open question. To assess Ae. albopictus' vector potential for ZIKV and identify key virus outbreak predictors, we built a complete framework using the complementary combination of (i) dose-dependent experimental Ae. albopictus exposure to ZIKV followed by time-dependent assessment of infection and systemic infection rates, (ii) modeling of intra-human ZIKV viremia dynamics, and (iii) in silico epidemiological simulations using an Agent-Based Model. The highest risk of transmission occurred during the pre-symptomatic stage of the disease, at the peak of viremia. At this dose, mosquito infection probability was estimated to be 20%, and 21 days were required to reach the median systemic infection rates. Mosquito population origin, either temperate or tropical, had no impact on infection rates or intra-host virus dynamic. Despite these unfavorable characteristics for transmission, Ae. albopictus was still able to trigger and yield large outbreaks in a simulated environment in the presence of sufficiently high mosquito biting rates. Our results reveal a low but existing epidemic potential of Ae. albopictus for ZIKV, that might explain the absence of large scale ZIKV epidemics so far in territories occupied only by Ae. albopictus. They nevertheless support active surveillance and eradication programs in these territories to maintain the risk of emergence to a low level.


Asunto(s)
Mosquitos Vectores/metabolismo , Mosquitos Vectores/virología , Infección por el Virus Zika/transmisión , Aedes/metabolismo , Aedes/virología , Animales , Brotes de Enfermedades , Vectores de Enfermedades , Epidemias , Humanos , Modelos Teóricos , Saliva/virología , Carga Viral , Viremia/transmisión , Virus Zika/patogenicidad , Infección por el Virus Zika/epidemiología , Infección por el Virus Zika/virología
4.
PLoS Negl Trop Dis ; 14(9): e0008531, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32911504

RESUMEN

Pathogens may manipulate their human and mosquito hosts to enhance disease transmission. Dengue, caused by four viral serotypes, is the fastest-growing transmissible disease globally resulting in 50-100 million infections annually. Transmission of the disease relies on the interaction between humans and the vector Aedes aegypti and is largely dependent on the odor-mediated host seeking of female mosquitoes. In this study, we use activity monitors to demonstrate that dengue virus-1 affects the locomotion and odor-mediated behavior of Ae. aegypti, reflecting the progression of infection within the mosquito. Mosquitoes 4-6 days post-infection increase locomotion, but do not alter their odor-driven host-seeking response. In contrast, females 14-16 days post-infection are less active, yet more sensitive to human odors as assessed by behavioral and electrophysiological assays. Such an increase in physiological and behavioral sensitivity is reflected by the antennal-specific increase in abundance of neural signaling transcripts in 14 days post-infection females, as determined by transcriptome analysis. This suggests that the sensitivity of the mosquito peripheral olfactory system is altered by the dengue virus by enhancing the overall neural responsiveness of the antenna, rather than the selective regulation of chemosensory-related genes. Our study reveals that dengue virus-1 enhances vector-related behaviors in the early stages post-infection that aid in avoiding predation and increasing spatial exploration. On the other hand, at the later stages of infection, the virus enhances the host-seeking capacity of the vector, thereby increasing the risk of virus transmission. A potential mechanism is discussed.


Asunto(s)
Aedes/virología , Dengue , Conducta de Búsqueda de Hospedador , Aedes/genética , Aedes/metabolismo , Aedes/fisiología , Animales , Antenas de Artrópodos/fisiología , Conducta Animal , Virus del Dengue/fisiología , Femenino , Perfilación de la Expresión Génica , Humanos , Locomoción , Mosquitos Vectores/fisiología , Mosquitos Vectores/virología
5.
Ecotoxicol Environ Saf ; 204: 111034, 2020 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-32758695

RESUMEN

Trehalose is the major blood sugar in insects; it not only serves as an energy source but also plays important roles in physiological responses to adverse conditions. However, only a few studies have explored the effects of heavy metal exposure stress on trehalose metabolism in insects. Therefore, in this study, we examined the effects of cadmium stress on changes in trehalose metabolism in Aedes albopictus. Three concentrations of cadmium (0.005, 0.01, and 0.1 mg/L) were selected for evaluation of long-term stress in Ae. albopictus (from eggs to adults); Ae. albopictus in double-distilled water was used as the control group. The trehalose and glucose contents, trehalase activity, and trehalose metabolism-related gene expression were determined. The effects of long-term cadmium exposure on growth, development, and reproduction were also assessed. Trehalose contents were increased, whereas glucose contents and trehalase activity were decreased in Ae. albopictus following long-term exposure to low concentrations of cadmium compared with those in untreated individuals. Moreover, the expression of trehalose-6-phosphate synthase was upregulated, and that of trehalase was downregulated, indicating that Ae. albopictus may enhance trehalose synthesis to resist cadmium stress. Cadmium exposure also caused Ae. albopictus individuals to become smaller with a longer developmental duration, whereas both reproduction and hatching rates of the offspring were decreased compared with those in the control group. Our findings demonstrated that cadmium exposure affected the morphology, physiology, and biochemistry of Ae. albopictus. These findings also confirmed the role of trehalose in the response of Ae. albopictus to cadmium stress, providing insights into the effects of heavy metal stress on trehalose metabolism in an insect model.


Asunto(s)
Aedes/efectos de los fármacos , Cadmio/efectos adversos , Trehalosa/metabolismo , Contaminantes Químicos del Agua/efectos adversos , Aedes/crecimiento & desarrollo , Aedes/metabolismo , Animales , Femenino , Larva/efectos de los fármacos , Larva/crecimiento & desarrollo , Larva/metabolismo , Óvulo/efectos de los fármacos , Óvulo/crecimiento & desarrollo , Óvulo/metabolismo , Pupa/efectos de los fármacos , Pupa/crecimiento & desarrollo , Pupa/metabolismo
6.
PLoS One ; 15(7): e0229314, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32706777

RESUMEN

INTRODUCTION: Many arboviruses of public health significance are maintained in zoonotic cycles with complex transmission pathways. The presence of serum antibody against arboviruses in vertebrates provides evidence of their historical exposure but reveals nothing about the vector-reservoir relationship. Moreover, collecting blood or tissue samples from vertebrate hosts is ethically and logistically challenging. We developed a novel approach for screening the immune status of vertebrates against Ross River virus that allows us to implicate the vectors that form the transmission pathways for this commonly notified Australian arboviral disease. METHODS: A micro-plaque reduction neutralisation test (micro-PRNT) was developed and validated on koala (Phascolarctos cinereus) sera against a standard PRNT. The ability of the micro-PRNT to detect RRV antibodies in mosquito blood meals was then tested using two mosquito models. Laboratory-reared Aedes aegypti were fed, via a membrane, on sheep blood supplemented with RRV seropositive and seronegative human sera. Aedes notoscriptus were fed on RRV seropositive and seronegative human volunteers. Blood-fed mosquitoes were harvested at various time points after feeding and their blood meals analysed for the presence of RRV neutralising antibodies using the micro-PRNT. RESULTS: There was significant agreement of the plaque neutralisation resulting from the micro-PRNT and standard PRNT techniques (R2 = 0.65; P<0.0001) when applied to RRV antibody detection in koala sera. Sensitivity and specificity of the micro-PRNT assay were 88.2% and 96%, respectively, in comparison with the standard PRNT. Blood meals from mosquitoes fed on sheep blood supplemented with RRV antibodies, and on blood from RRV seropositive humans neutralised the virus by ≥50% until 48 hr post feeding. The vertebrate origin of the blood meal was also ascertained for the same samples, in parallel, using established molecular techniques. CONCLUSIONS: The small volumes of blood present in mosquito abdomens can be used to identify RRV antibodies and therefore host exposure to arbovirus infection. In tandem with the accurate identification of the mosquito, and diagnostics for the host origin of the blood meal, this technique has tremendous potential for exploring RRV transmission pathways. It can be adapted for similar studies on other mosquito borne zoonoses.


Asunto(s)
Aedes/metabolismo , Alimentación Animal/análisis , Anticuerpos Antivirales/análisis , Pruebas de Neutralización/métodos , Virus del Río Ross/inmunología , Aedes/virología , Infecciones por Alphavirus/diagnóstico , Infecciones por Alphavirus/transmisión , Infecciones por Alphavirus/veterinaria , Alimentación Animal/virología , Animales , Anticuerpos Antivirales/sangre , Vectores de Enfermedades , Femenino , Humanos , Phascolarctidae/virología , Sensibilidad y Especificidad
7.
PLoS Negl Trop Dis ; 14(5): e0008216, 2020 05.
Artículo en Inglés | MEDLINE | ID: mdl-32384079

RESUMEN

The extensive use of insecticides for vector control has led to the development of insecticide resistance in Aedes aegypti populations on a global scale, which has significantly compromised control actions. Insecticide resistance, and its underlying mechanisms, has been investigated in several countries, mostly in South American and Asian countries. In Africa, however, studies reporting insecticide resistance are rare and data on resistance mechanisms, notably knockdown resistance (kdr) mutations, is scarce. In this study, the recently described V410L kdr mutation is reported for the first time in old world Ae. aegypti populations, namely from Angola and Madeira island. Two additional kdr mutations, V1016I and F1534C, are also reported for the first time in populations from Angola and Cape Verde. Significant associations with the resistance phenotype were found for both V410L and V1016I individually as well as for tri-locus genotypes in the Angolan population. However, no association was found in Madeira island, probably due to the presence of a complex pattern of multiple insecticide resistance mechanisms in the local Ae. aegypti population. These results suggest that populations carrying the same kdr mutations may respond differently to the same insecticide, stressing the need for complementary studies when assessing the impact of kdr resistance mechanisms in the outcome of insecticide-based control strategies.


Asunto(s)
Aedes/efectos de los fármacos , Aedes/genética , Proteínas de Insectos/genética , Resistencia a los Insecticidas , Mosquitos Vectores/efectos de los fármacos , Mutación Missense , Aedes/metabolismo , Angola , Animales , Femenino , Genotipo , Proteínas de Insectos/metabolismo , Insecticidas/farmacología , Mosquitos Vectores/genética , Mosquitos Vectores/metabolismo , Portugal
8.
Biochim Biophys Acta Gene Regul Mech ; 1863(8): 194576, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32389826

RESUMEN

Juvenile hormones (JH) and ecdysone coordinately regulate metamorphosis in Aedes aegypti. We studied the function of an epigenetic regulator and multifunctional transactivator, CREB binding protein (CBP) in A. aegypti. RNAi-mediated knockdown of CBP in Ae. aegypti larvae resulted in suppression of JH primary response gene, Krüppel-homolog 1 (Kr-h1), and induction of primary ecdysone response gene, E93, resulting in multiple effects including early metamorphosis, larval-pupal intermediate formation, mortality and inhibition of compound eye development. RNA sequencing identified hundreds of genes, including JH and ecdysone response genes regulated by CBP. In the presence of JH, CBP upregulates Kr-h1 by acetylating core histones at the Kr-h1 promoter and facilitating the recruitment of JH receptor and other proteins. CBP suppresses metamorphosis regulators, EcR-A, USP-A, BR-C, and E93 through the upregulation of Kr-h1 and E75A. CBP regulates the expression of core eye specification genes including those involved in TGF-ß and EGFR signaling. These studies demonstrate that CBP is an essential player in JH and 20E action and regulates metamorphosis and compound eye development in Ae. aegypti.


Asunto(s)
Aedes/metabolismo , Proteína de Unión a CREB/metabolismo , Ojo/crecimiento & desarrollo , Metamorfosis Biológica/fisiología , Organogénesis/fisiología , Aedes/genética , Animales , Proteína de Unión a CREB/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila , Ecdisona/genética , Ecdisona/metabolismo , Ecdisona/farmacología , Femenino , Regulación del Desarrollo de la Expresión Génica , Histonas/metabolismo , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Hormonas Juveniles/metabolismo , Hormonas Juveniles/farmacología , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Larva , Organogénesis/efectos de los fármacos , Organogénesis/genética , Regiones Promotoras Genéticas , Pupa/crecimiento & desarrollo , Transducción de Señal , Factores de Transcripción/metabolismo , Fiebre Amarilla/genética
9.
PLoS Pathog ; 16(4): e1008433, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-32282862

RESUMEN

The insect bacterium Wolbachia pipientis is being introgressed into Aedes aegypti populations as an intervention against the transmission of medically important arboviruses. Here we compare Ae. aegypti mosquitoes infected with wMelCS or wAlbB to the widely used wMel Wolbachia strain on an Australian nuclear genetic background for their susceptibility to infection by dengue virus (DENV) genotypes spanning all four serotypes. All Wolbachia-infected mosquitoes were more resistant to intrathoracic DENV challenge than their wildtype counterparts. Blocking of DENV replication was greatest by wMelCS. Conversely, wAlbB-infected mosquitoes were more susceptible to whole body infection than wMel and wMelCS. We extended these findings via mosquito oral feeding experiments, using viremic blood from 36 acute, hospitalised dengue cases in Vietnam, additionally including wMel and wildtype mosquitoes on a Vietnamese nuclear genetic background. As above, wAlbB was less effective at blocking DENV replication in the abdomen compared to wMel and wMelCS. The transmission potential of all Wolbachia-infected mosquito lines (measured by the presence/absence of infectious DENV in mosquito saliva) after 14 days, was significantly reduced compared to their wildtype counterparts, and lowest for wMelCS and wAlbB. These data support the use of wAlbB and wMelCS strains for introgression field trials and the biocontrol of DENV transmission. Furthermore, despite observing significant differences in transmission potential between wildtype mosquitoes from Australia and Vietnam, no difference was observed between wMel-infected mosquitoes from each background suggesting that Wolbachia may override any underlying variation in DENV transmission potential.


Asunto(s)
Aedes/microbiología , Aedes/virología , Virus del Dengue/fisiología , Mosquitos Vectores/microbiología , Mosquitos Vectores/virología , Wolbachia/fisiología , Aedes/genética , Aedes/metabolismo , Animales , Femenino , Masculino , Mosquitos Vectores/genética , Mosquitos Vectores/metabolismo , Control Biológico de Vectores , Replicación Viral
10.
Sci Rep ; 10(1): 3300, 2020 02 24.
Artículo en Inglés | MEDLINE | ID: mdl-32094450

RESUMEN

Aedes aegypti is the primary vector for transmission of Dengue, Zika and chikungunya viruses. Previously it was shown that Dengue virus infection of the mosquito led to an in increased expression of the odorant binding protein 22 (AeOBP22) within the mosquito salivary gland and that siRNA mediated knockdown of AeOBP22 led to reduced mosquito feeding behaviors. Insect OBPs are implicated in the perception, storage and transport of chemosensory signaling molecules including air-borne odorants and pheromones. AeOBP22 is unusual as it is additionally expressed in multiple tissues, including the antenna, the male reproductive glands and is transferred to females during reproduction, indicating multiple roles in the mosquito life cycle. However, it is unclear what role it plays in these tissues and what ligands it interacts with. Here we present solution and X-ray crystallographic studies that indicate a potential role of AeOBP22 binding to fatty acids, and that the specificity for longer chain fatty acids is regulated by a conformational change in the C-terminal tail that leads to creation of an enlarged binding cavity that enhances binding affinity. This study sheds light onto the native ligands for AeOBP22 and provides insight into its potential functions in different tissues.


Asunto(s)
Aedes/metabolismo , Ácidos Grasos/metabolismo , Proteínas de Insectos/química , Proteínas de Insectos/metabolismo , Odorantes , Animales , Apoproteínas/química , Ácido Araquidónico/metabolismo , Cristalografía por Rayos X , Concentración de Iones de Hidrógeno , Cinética , Ligandos , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Unión Proteica , Conformación Proteica , Estabilidad Proteica , Soluciones , Homología Estructural de Proteína
11.
Sci Rep ; 10(1): 1755, 2020 02 04.
Artículo en Inglés | MEDLINE | ID: mdl-32020001

RESUMEN

Insect CAPA neuropeptides are homologs of mammalian neuromedin U and are known to influence ion and water balance by regulating the activity of the Malpighian 'renal' tubules (MTs). Several diuretic hormones are known to increase primary fluid and ion secretion by insect MTs and, in adult female mosquitoes, a calcitonin-related peptide (DH31) called mosquito natriuretic peptide, increases sodium secretion to compensate for the excess salt load acquired during blood-feeding. An endogenous mosquito anti-diuretic hormone was recently described, having potent inhibitory activity against select diuretic hormones, including DH31. Herein, we functionally deorphanized, both in vitro and in vivo, a mosquito anti-diuretic hormone receptor (AedaeADHr) with expression analysis indicating highest enrichment in the MTs where it is localized within principal cells. Characterization using a heterologous in vitro system demonstrated the receptor was highly sensitive to mosquito CAPA neuropeptides while in vivo, AedaeADHr knockdown abolished CAPA-induced anti-diuretic control of DH31-stimulated MTs. CAPA neuropeptides are produced within a pair of neurosecretory cells in each of the abdominal ganglia, whose axonal projections innervate the abdominal neurohaemal organs, where these neurohormones are released into circulation. Lastly, pharmacological inhibition of nitric oxide synthase (NOS) and protein kinase G (PKG) signaling eliminated anti-diuretic activity of CAPA, highlighting the role of the second messenger cGMP and NOS/PKG in this anti-diuretic signaling pathway.


Asunto(s)
Aedes/metabolismo , Fármacos Antidiuréticos/metabolismo , Proteínas de Insectos/metabolismo , Mosquitos Vectores/metabolismo , Neuropéptidos/metabolismo , Transducción de Señal/fisiología , Animales , GMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Vectores de Enfermedades , Humanos , Túbulos de Malpighi/metabolismo , Óxido Nítrico Sintasa/metabolismo , Sistemas de Mensajero Secundario/fisiología
12.
PLoS Negl Trop Dis ; 14(2): e0007948, 2020 02.
Artículo en Inglés | MEDLINE | ID: mdl-32012156

RESUMEN

Aedes cadherin (AaeCad, AAEL024535) has been characterized as a receptor for Bacillus thuringiensis subsp. israelensis (Bti) Cry11A toxins. However, its role in development is still unknown. In this study, we modified the cadherin gene using ZFN and TALEN. Even though we obtained heterozygous deletions, no homozygous mutants were viable. Because ZFN and TALEN have lower off-targets than CRISPR/Cas9, we conclude the cadherin gene is essential for Aedes development. In contrast, in lepidopteran insects loss of a homologous cadherin does not appear to be lethal, since homozygous mutants are viable. To analyze the role of AaeCad in vivo, we tagged this protein with EGFP using CRISPR-Cas9-mediated homologous recombination and obtained a homozygous AaeCad-EGFP line. Addition of Aedes Rad51 mRNA enhanced the rate of recombination. We then examined AaeCad protein expression in most tissues and protein dynamics during mosquito development. We observe that AaeCad is expressed in larval and adult midgut-specific manner and its expression pattern changed during the mosquito development. Confocal images showed AaeCad has high expression in larval caecae and posterior midgut, and also in adult midgut. Expression of AaeCad is observed primarily in the apical membranes of epithelial cells, and not in cell-cell junctions. The expression pattern observed suggests AaeCad does not appear to play a role in these junctions. However, we cannot exclude its role beyond cell-cell adhesion in the midgut. We also observed that Cry11A bound to the apical side of larval gastric caecae and posterior midgut cells exactly where AaeCad-EGFP was expressed. Their co-localization suggests that AaeCad is indeed a receptor for the Cry11A toxin. Using this mosquito line we also observed that low doses of Cry11A toxin caused the cells to slough off membranes, which likely represents a defense mechanism, to limit cell damage from Cry11A toxin pores formed in the cell membrane.


Asunto(s)
Aedes/metabolismo , Proteínas Bacterianas/metabolismo , Endotoxinas/metabolismo , Proteínas Hemolisinas/metabolismo , Proteínas de Insectos/metabolismo , Larva/crecimiento & desarrollo , Aedes/genética , Aedes/crecimiento & desarrollo , Animales , Proteínas Bacterianas/genética , Cadherinas/metabolismo , Sistema Digestivo/crecimiento & desarrollo , Sistema Digestivo/metabolismo , Endotoxinas/genética , Proteínas Hemolisinas/genética , Proteínas de Insectos/genética , Larva/genética , Larva/metabolismo , Unión Proteica
13.
Insect Biochem Mol Biol ; 119: 103325, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-31978586

RESUMEN

RNA activation (RNAa) is a newly emerging area of research in which dsRNA targeting promoter regions can induce the expression of the target gene. Although still in its infancy, it is already having significant impacts in several research areas in particular as cancer therapeutics. So far, the scope of RNAa has been limited to mammals and Caenorhabditis elegans with no indication of its prevalence in insects. In this study, we aimed to demonstrate the presence of RNAa in the insect dengue vector Aedes aegypti. Furthermore, we looked to uncover some details surrounding the involvement of host factors in order to present this as a new technique for insect research. The outcomes of this study provide new opportunities to further research into arthropod-borne diseases and insect biology in the same way as RNA interference.


Asunto(s)
Aedes/genética , Mosquitos Vectores/genética , ARN Bicatenario/genética , Activación Transcripcional , Aedes/metabolismo , Animales , Animales Modificados Genéticamente/genética , Animales Modificados Genéticamente/metabolismo , Mosquitos Vectores/metabolismo , ARN Bicatenario/metabolismo
14.
Nat Commun ; 11(1): 260, 2020 01 14.
Artículo en Inglés | MEDLINE | ID: mdl-31937766

RESUMEN

Transmission from an infected mosquito to a host is an essential process in the life cycle of mosquito-borne flaviviruses. Numerous studies have demonstrated that mosquito saliva facilitates viral transmission. Here we find that a saliva-specific protein, named Aedes aegypti venom allergen-1 (AaVA-1), promotes dengue and Zika virus transmission by activating autophagy in host immune cells of the monocyte lineage. The AG6 mice (ifnar1-/-ifngr1-/-) bitten by the virus-infected AaVA-1-deficient mosquitoes present a lower viremia and prolonged survival. AaVA-1 intracellularly interacts with a dominant negative binder of Beclin-1, known as leucine-rich pentatricopeptide repeat-containing protein (LRPPRC), and releases Beclin-1 from LRPPRC-mediated sequestration, thereby enabling the initialization of downstream autophagic signaling. A deficiency in Beclin-1 reduces viral infection in mice and abolishes AaVA-1-mediated enhancement of ZIKV transmission by mosquitoes. Our study provides a mechanistic insight into saliva-aided viral transmission and could offer a potential prophylactic target for reducing flavivirus transmission.


Asunto(s)
Aedes/metabolismo , Autofagia , Infecciones por Flavivirus/transmisión , Flavivirus/fisiología , Proteínas de Insectos/metabolismo , Mosquitos Vectores/metabolismo , Proteínas y Péptidos Salivales/metabolismo , Aedes/virología , Animales , Beclina-1/deficiencia , Beclina-1/metabolismo , Virus del Dengue/fisiología , Infecciones por Flavivirus/virología , Humanos , Proteínas de Insectos/deficiencia , Proteínas de Insectos/genética , Ratones , Mosquitos Vectores/virología , Proteínas de Neoplasias/metabolismo , Unión Proteica , Proteínas y Péptidos Salivales/deficiencia , Proteínas y Péptidos Salivales/genética , Células THP-1 , Replicación Viral , Virus Zika/fisiología
15.
Insect Biochem Mol Biol ; 118: 103311, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-31901476

RESUMEN

The initial signal that governs sex determination is highly variable among insects. A homolog of Nix, the male-determining factor in Aedes aegypti, was previously found in the Asian tiger mosquito Ae. albopictus. Here we show that the Ae. albopictus Nix (AalNix) is more complex in gene structure and splice isoforms than its Ae. aegypti homolog (AaeNix). AalNix shows a similar transcription profile compared to AaeNix. CRISPR/Cas9-mediated knockouts of AalNix in vivo and in the Ae. albopictus C6/36 cells lead to a shift of dsx and fru splicing towards the female isoforms. G0 knockout males showed feminization and deformities including feminized antennae, absence or partial absence of gonocoxites, gonostyli, testes and accessory glands, and the formation of ovaries. Despite ~70 MY of divergence, Nix functions as a conserved male-determining factor in the two most important arboviral vectors, namely Ae. aegypti and Ae. albopictus.


Asunto(s)
Aedes/genética , Proteínas de Insectos/genética , Aedes/crecimiento & desarrollo , Aedes/metabolismo , Animales , Secuencia de Bases , Femenino , Proteínas de Insectos/metabolismo , Masculino , Alineación de Secuencia , Procesos de Determinación del Sexo
16.
Parasit Vectors ; 13(1): 11, 2020 Jan 09.
Artículo en Inglés | MEDLINE | ID: mdl-31918753

RESUMEN

BACKGROUND: Dengue fever outbreaks tend to spread northward in China, and Jining is the northernmost region where local dengue fever cases have been detected. Therefore, it is important to investigate the density of Aedes albopictus and its resistance to deltamethrin. METHODS: The Breteau index (BI) and container index (CI) were calculated to assess the larval density of Ae. albopictus and human-baited double net trap (HDN) surveillance was performed in six subordinate counties (Rencheng, Yanzhou, Sishui, Liangshan, Zoucheng and Jiaxiang) of Jining City in 2017 and 2018. The resistance of Ae. albopictus adults to deltamethrin was evaluated using the World Health Organization (WHO) standard resistance bioassay. The mutations at Vgsc codons 1532 and 1534 were also analysed to determine the association between kdr mutations and phenotypic resistance in adult mosquitoes. RESULTS: The average BI, CI and biting rate at Jining were 45.30, 16.02 and 1.97 (female /man/hour) in 2017 and 15.95, 7.86 and 0.59 f/m/h in 2018, respectively. In August 26, 2017, when the first dengue fever case was diagnosed, the BI at Qianli village in Jiaxiang County was 107.27. The application of prevention and control measures by the government sharply decreased the BI to a value of 4.95 in September 3, 2017. The mortality of field-collected Ae. albopictus females from Jiaxiang was 41.98%. I1532T, F1534L and F1534S mutations were found in domain III of the Vgsc gene. This study provides the first demonstration that both I1532T and F1534S mutations are positively correlated with the deltamethrin-resistant phenotype. CONCLUSIONS: Mosquito density surveillance, resistance monitoring and risk assessment should be strengthened in areas at risk for dengue to ensure the sustainable control of Ae. albopictus and thus the prevention and control of dengue transmission.


Asunto(s)
Aedes/efectos de los fármacos , Dengue/transmisión , Resistencia a los Insecticidas , Insecticidas/farmacología , Aedes/genética , Aedes/metabolismo , Aedes/virología , Altitud , Animales , China , Dengue/virología , Virus del Dengue/genética , Virus del Dengue/aislamiento & purificación , Virus del Dengue/fisiología , Ecología , Femenino , Humanos , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Larva/efectos de los fármacos , Larva/genética , Larva/metabolismo , Larva/virología , Mosquitos Vectores/efectos de los fármacos , Mosquitos Vectores/genética , Mosquitos Vectores/metabolismo , Mosquitos Vectores/virología , Mutación , Nitrilos , Piretrinas
17.
Artículo en Inglés | MEDLINE | ID: mdl-31707089

RESUMEN

Compounds having insecticidal activity can be used to control Aedes aegypti mosquitoes, a major worldwide vector, and several plants have a source of such molecules. A principal component analysis (PCA) was carried out to determine the criterion to select larvicidal metabolites. The insecticidal activity of seven selected metabolites by PCA was validated by determining its lethal concentrations 50 (LC50) by probit analysis. Six of the seven evaluated molecules presented LC50 values <100 ppm. The effects of these six molecules on acetylcholinesterase and the respiratory chain complexes of the mitochondria of Ae. aegypti were evaluated. Four metabolites presenting the highest inhibition effects on these targets were mixed in 11 different combinations, and the percentage of mortality of each mixture on Ae. aegypti larvae were determined. Secondary metabolites such as geranyl acetate, α-humulene, ß-caryophyllene, geraniol, nerol, and n-octanol presented LC50 values under 100 ppm (44, 41, 48, 84, 87, and 98 ppm, respectively), whereas 1,8-cineole presented a LC50 value of 183 ppm. We found that, geranyl acetate, α-humulene, ß-caryophyllene, nerol, n-octanol, and geraniol inhibited at least one of the six targets with an efficiency between 25 and 41%. Overall, the evaluation of the different mixtures revealed a synergistic effect between geranyl acetate and geraniol, and an antagonistic effect between α-humulene and ß-caryophyllene compounds.


Asunto(s)
Aedes/metabolismo , Insecticidas/toxicidad , Mitocondrias/metabolismo , Control de Mosquitos/métodos , Metabolismo Secundario , Acetatos/toxicidad , Monoterpenos Acíclicos/toxicidad , Animales , Sesquiterpenos Monocíclicos/toxicidad , Oxidación-Reducción , Sesquiterpenos Policíclicos/toxicidad
18.
Curr Biol ; 29(24): 4241-4248.e5, 2019 12 16.
Artículo en Inglés | MEDLINE | ID: mdl-31761702

RESUMEN

Dengue has enormous health impacts globally. A novel approach to decrease dengue incidence involves the introduction of Wolbachia endosymbionts that block dengue virus transmission into populations of the primary vector mosquito, Aedes aegypti. The wMel Wolbachia strain has previously been trialed in open releases of Ae. aegypti; however, the wAlbB strain has been shown to maintain higher density than wMel at high larval rearing temperatures. Releases of Ae. aegypti mosquitoes carrying wAlbB were carried out in 6 diverse sites in greater Kuala Lumpur, Malaysia, with high endemic dengue transmission. The strain was successfully established and maintained at very high population frequency at some sites or persisted with additional releases following fluctuations at other sites. Based on passive case monitoring, reduced human dengue incidence was observed in the release sites when compared to control sites. The wAlbB strain of Wolbachia provides a promising option as a tool for dengue control, particularly in very hot climates.


Asunto(s)
Aedes/microbiología , Dengue/prevención & control , Control Biológico de Vectores/métodos , Wolbachia/metabolismo , Aedes/genética , Aedes/metabolismo , Animales , Virus del Dengue/metabolismo , Virus del Dengue/patogenicidad , Femenino , Humanos , Insectos Vectores , Malasia , Masculino , Mosquitos Vectores , Wolbachia/genética
19.
Curr Biol ; 29(22): 3946-3952.e5, 2019 11 18.
Artículo en Inglés | MEDLINE | ID: mdl-31679930

RESUMEN

Globally, diseases transmitted by arthropod vectors, such as mosquitoes, remain a major cause of morbidity and mortality [1]. The defense responses of mosquito and other arthropod vectors against parasites are important for understanding disease transmission dynamics and for the development of novel disease-control strategies. Consequently, the mechanisms by which mosquitoes resist parasitic infection (e.g., immune-mediated killing) have long been studied [2, 3]. However, the ability of mosquitoes to ameliorate the negative fitness consequences of infection through tolerance mechanisms (e.g., tissue repair) has been virtually ignored (but see [4, 5]). Ignoring parasite tolerance is especially taxing in vector biology because unlike resistance, which typically reduces vectorial capacity, tolerance is expected to increase vectorial capacity by reducing parasite-mediated mortality without killing parasites [6], contributing to the recurrent emergence of vector-borne diseases and its stabilization and exacerbation. Despite its importance, there is currently no evidence for the evolution of tolerance in natural mosquito populations. Here, we use a common-garden experimental framework to measure variation in resistance and tolerance to dog heartworm (Dirofilaria immitis) between eight natural Aedes albopictus mosquito populations representing areas of low and high transmission intensity. We find significant inter-population variation in tolerance and elevated tolerance where transmission intensity is high. Additionally, as expected, we find that increased tolerance is associated with higher vectorial capacity. Consequently, our results indicate that high transmission intensity can lead to the evolution of more competent disease vectors, which can feed back to impact disease risk.


Asunto(s)
Aedes/metabolismo , Aedes/parasitología , Dirofilaria immitis/patogenicidad , Aedes/fisiología , Animales , Vectores de Enfermedades , Interacciones Huésped-Parásitos/fisiología , Mosquitos Vectores/metabolismo , Mosquitos Vectores/parasitología , Parásitos , Enfermedades Parasitarias/inmunología , Enfermedades Parasitarias en Animales/inmunología
20.
Proc Natl Acad Sci U S A ; 116(43): 21501-21507, 2019 10 22.
Artículo en Inglés | MEDLINE | ID: mdl-31570611

RESUMEN

The yellow fever mosquito, Aedes aegypti, vectors human pathogens. Juvenile hormones (JH) control almost every aspect of an insect's life, and JH analogs are currently used to control mosquito larvae. Since RNA interference does not work efficiently during the larval stages of this insect, JH regulation of larval development and mode of action of JH analogs are not well studied. To overcome this limitation, we used a multiple single guide RNA-based CRISPR/Cas9 genome-editing method to knockout the methoprene-tolerant (Met) gene coding for a JH receptor. The Met knockout larvae exhibited a black larval phenotype during the L3 (third instar larvae) and L4 (fourth instar larvae) stages and died before pupation. However, Met knockout did not affect embryonic development or the L1 and L2 stages. Microscopy studies revealed the precocious synthesis of a dark pupal cuticle during the L3 and L4 stages. Gene expression analysis showed that Krüppel homolog 1, a key transcription factor in JH action, was down-regulated, but genes coding for proteins involved in melanization, pupal and adult cuticle synthesis, and blood meal digestion in adults were up-regulated in L4 Met mutants. These data suggest that, during the L3 and L4 stages, Met mediates JH suppression of pupal/adult genes involved in the synthesis and melanization of the cuticle and blood meal digestion. These results help to advance our knowledge of JH regulation of larval development and the mode of action of JH analogs in Ae. aegypti.


Asunto(s)
Aedes/genética , Proteínas Portadoras/genética , Proteínas de Insectos/genética , Hormonas Juveniles/metabolismo , Metopreno/metabolismo , Aedes/crecimiento & desarrollo , Aedes/metabolismo , Animales , Proteínas Portadoras/metabolismo , Regulación del Desarrollo de la Expresión Génica , Técnicas de Inactivación de Genes , Proteínas de Insectos/metabolismo , Larva/genética , Larva/crecimiento & desarrollo , Larva/metabolismo , Mosquitos Vectores/genética , Mosquitos Vectores/crecimiento & desarrollo , Mosquitos Vectores/metabolismo , Pupa/genética , Pupa/crecimiento & desarrollo , Pupa/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
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