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Aedes aegypti mosquitoes carrying self-spreading, virus-blocking Wolbachia bacteria are being deployed to suppress dengue transmission. However, there are challenges in applying this technology in extreme environments. We introduced two Wolbachia strains into Ae. aegypti from Saudi Arabia for a release program in the hot coastal city of Jeddah. Wolbachia reduced infection and dissemination of dengue virus (DENV2) in Saudi Arabian mosquitoes and showed complete maternal transmission and cytoplasmic incompatibility. Wolbachia reduced egg hatch under a range of environmental conditions, with the Wolbachia strains showing differential thermal stability. Wolbachia effects were similar across mosquito genetic backgrounds but we found evidence of local adaptation, with Saudi Arabian mosquitoes having lower egg viability but higher adult desiccation tolerance than Australian mosquitoes. Genetic background effects will influence Wolbachia invasion dynamics, reinforcing the need to use local genotypes for mosquito release programs, particularly in extreme environments like Jeddah. Our comprehensive characterization of Wolbachia strains provides a foundation for Wolbachia-based disease interventions in harsh climates.
Assuntos
Aedes , Dengue , Wolbachia , Animais , Arábia Saudita , Austrália , Ambientes ExtremosRESUMO
Excitotoxicity, a neuronal death process in neurological disorders such as stroke, is initiated by the overstimulation of ionotropic glutamate receptors. Although dysregulation of proteolytic signaling networks is critical for excitotoxicity, the identity of affected proteins and mechanisms by which they induce neuronal cell death remain unclear. To address this, we used quantitative N-terminomics to identify proteins modified by proteolysis in neurons undergoing excitotoxic cell death. We found that most proteolytically processed proteins in excitotoxic neurons are likely substrates of calpains, including key synaptic regulatory proteins such as CRMP2, doublecortin-like kinase I, Src tyrosine kinase and calmodulin-dependent protein kinase IIß (CaMKIIß). Critically, calpain-catalyzed proteolytic processing of these proteins generates stable truncated fragments with altered activities that potentially contribute to neuronal death by perturbing synaptic organization and function. Blocking calpain-mediated proteolysis of one of these proteins, Src, protected against neuronal loss in a rat model of neurotoxicity. Extrapolation of our N-terminomic results led to the discovery that CaMKIIα, an isoform of CaMKIIß, undergoes differential processing in mouse brains under physiological conditions and during ischemic stroke. In summary, by identifying the neuronal proteins undergoing proteolysis during excitotoxicity, our findings offer new insights into excitotoxic neuronal death mechanisms and reveal potential neuroprotective targets for neurological disorders.
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Morte Celular , Neurônios , Sinapses , Animais , Masculino , Camundongos , Ratos , Calpaína/metabolismo , Células Cultivadas , Inibidores de Cisteína Proteinase/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/metabolismo , Neurônios/patologia , Neurônios/fisiologia , Neuroproteção , Proteoma/análise , Ratos Wistar , Acidente Vascular Cerebral/patologia , Sinapses/patologia , Sinapses/fisiologiaRESUMO
[This corrects the article DOI: 10.1371/journal.ppat.1008103.].
RESUMO
With dengue virus (DENV) becoming endemic in tropical and subtropical regions worldwide, there is a pressing global demand for effective strategies to control the mosquitoes that spread this disease. Recent advances in genetic engineering technologies have made it possible to create mosquitoes with reduced vector competence, limiting their ability to acquire and transmit pathogens. Here we describe the development of Aedes aegypti mosquitoes synthetically engineered to impede vector competence to DENV. These mosquitoes express a gene encoding an engineered single-chain variable fragment derived from a broadly neutralizing DENV human monoclonal antibody and have significantly reduced viral infection, dissemination, and transmission rates for all four major antigenically distinct DENV serotypes. Importantly, this is the first engineered approach that targets all DENV serotypes, which is crucial for effective disease suppression. These results provide a compelling route for developing effective genetic-based DENV control strategies, which could be extended to curtail other arboviruses.
Assuntos
Aedes/genética , Anticorpos Antivirais/imunologia , Anticorpos Amplamente Neutralizantes/imunologia , Vírus da Dengue/imunologia , Aedes/virologia , Animais , Anticorpos Antivirais/biossíntese , Anticorpos Antivirais/genética , Anticorpos Amplamente Neutralizantes/biossíntese , Anticorpos Amplamente Neutralizantes/genética , Feminino , Humanos , Masculino , Engenharia de Proteínas , Anticorpos de Cadeia Única/genéticaRESUMO
Recent Zika virus (ZIKV) outbreaks have highlighted the necessity for development of novel vector control strategies to combat arboviral transmission, including genetic versions of the sterile insect technique, artificial infection with Wolbachia to reduce population size and/or vectoring competency, and gene drive-based methods. Here, we describe the development of mosquitoes synthetically engineered to impede vector competence to ZIKV. We demonstrate that a polycistronic cluster of engineered synthetic small RNAs targeting ZIKV is expressed and fully processed in Aedes aegypti, ensuring the formation of mature synthetic small RNAs in the midgut where ZIKV resides in the early stages of infection. Critically, we demonstrate that engineered Ae. aegypti mosquitoes harboring the anti-ZIKV transgene have significantly reduced viral infection, dissemination, and transmission rates of ZIKV. Taken together, these compelling results provide a promising path forward for development of effective genetic-based ZIKV control strategies, which could potentially be extended to curtail other arboviruses.
Assuntos
Mosquitos Vetores/genética , Infecção por Zika virus/genética , Zika virus/genética , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/virologia , Surtos de Doenças , Humanos , Mosquitos Vetores/virologia , Saliva/virologia , Carga Viral/genética , Wolbachia/patogenicidade , Wolbachia/virologia , Zika virus/patogenicidade , Infecção por Zika virus/transmissão , Infecção por Zika virus/virologiaRESUMO
The aim of this article is to summarise the virology content presented at the 9th Lorne Infection and Immunity Conference, Australia, in February 2019. The broad program included virology as a key theme, and the commentary herein highlights several key virology presentations at the meeting.
Assuntos
Virologia , Viroses/imunologia , Animais , Austrália , Quirópteros/virologia , Congressos como Assunto , Reservatórios de Doenças/virologia , Humanos , Viroses/prevenção & controle , Viroses/transmissão , Vírus/genética , Vírus/patogenicidadeRESUMO
BACKGROUND: Zika virus infection in new born is linked to congenital syndromes, especially microcephaly. Studies have shown that these neuropathies are the result of significant death of neuronal progenitor cells in the central nervous system of the embryo, targeted by the virus. Although cell death via apoptosis is well acknowledged, little is known about possible pathogenic cellular mechanisms triggering cell death in neurons. METHODS: We used in vitro embryonic mouse primary neuron cultures to study possible upstream cellular mechanisms of cell death. Neuronal networks were grown on microelectrode array and electrical activity was recorded at different times post Zika virus infection. In addition to this method, we used confocal microscopy and Q-PCR techniques to observe morphological and molecular changes after infection. RESULTS: Zika virus infection of mouse primary neurons triggers an early spiking excitation of neuron cultures, followed by dramatic loss of this activity. Using NMDA receptor antagonist, we show that this excitotoxicity mechanism, likely via glutamate, could also contribute to the observed nervous system defects in human embryos and could open new perspective regarding the causes of adult neuropathies. CONCLUSIONS: This model of excitotoxicity, in the context of neurotropic virus infection, highlights the significance of neuronal activity recording with microelectrode array and possibility of more than one lethal mechanism after Zika virus infection in the nervous system.
Assuntos
Potenciais de Ação/fisiologia , Morte Celular , Rede Nervosa/virologia , Neurônios/virologia , Infecção por Zika virus/virologia , Zika virus/fisiologia , Animais , Encéfalo/citologia , Encéfalo/virologia , Células Cultivadas , Ácido Glutâmico/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Modelos Neurológicos , Rede Nervosa/patologia , Neurônios/metabolismo , Neurônios/patologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Transdução de Sinais/genética , Transmissão Sináptica , Replicação Viral , Infecção por Zika virus/patologiaRESUMO
Although mosquitoes serve as vectors of many pathogens of public health importance, their response to viral infection is poorly understood. It also remains to be investigated whether viruses deploy some mechanism to be able to overcome this immune response. Here, we have used an RNA-Seq approach to identify differentially regulated genes in Culex quinquefasciatus cells following West Nile virus (WNV) infection, identifying 265 transcripts from various cellular pathways that were either upregulated or downregulated. Ubiquitin-proteasomal pathway genes, comprising 12% of total differentially regulated genes, were selected for further validation by real time RT-qPCR and functional analysis. It was found that treatment of infected cells with proteasomal inhibitor, MG-132, decreased WNV titers, indicating importance of this pathway during infection process. In infection models, the Culex ortholog of mammalian Cul4A/B (cullin RING ubiquitin ligase) was found to be upregulated in vitro as well as in vivo, especially in midguts of mosquitoes. Gene knockdown using dsRNA and overexpression studies indicated that Culex Cul4 acts as a pro-viral protein by degradation of CxSTAT via ubiquitin-proteasomal pathway. We also show that gene knockdown of Culex Cul4 leads to activation of the Jak-STAT pathway in mosquitoes leading to decrease viral replication in the body as well as saliva. Our results suggest a novel mechanism adopted by WNV to overcome mosquito immune response and increase viral replication.
Assuntos
Culex/virologia , Proteínas Culina/metabolismo , Indução Enzimática , Evasão da Resposta Imune , Proteínas de Insetos/metabolismo , Replicação Viral , Vírus do Nilo Ocidental/fisiologia , Aedes/imunologia , Aedes/metabolismo , Aedes/virologia , Animais , Linhagem Celular , Culex/imunologia , Culex/metabolismo , Proteínas Culina/antagonistas & inibidores , Proteínas Culina/genética , Vírus da Dengue/imunologia , Vírus da Dengue/fisiologia , Feminino , Trato Gastrointestinal/imunologia , Trato Gastrointestinal/metabolismo , Trato Gastrointestinal/virologia , Técnicas de Silenciamento de Genes , Proteínas de Insetos/antagonistas & inibidores , Proteínas de Insetos/genética , Janus Quinases/antagonistas & inibidores , Janus Quinases/genética , Janus Quinases/metabolismo , RNA/antagonistas & inibidores , RNA/metabolismo , Interferência de RNA , RNA Viral/antagonistas & inibidores , RNA Viral/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Fatores de Transcrição STAT/antagonistas & inibidores , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Transcriptoma , Vírus do Nilo Ocidental/imunologia , Vírus do Nilo Ocidental/isolamento & purificaçãoRESUMO
RNA viruses exhibit substantial structural, ecological and genomic diversity. However, genome size in RNA viruses is likely limited by a high mutation rate, resulting in the evolution of various mechanisms to increase complexity while minimising genome expansion. Here we conduct a large-scale analysis of the genome sequences of 99 animal rhabdoviruses, including 45 genomes which we determined de novo, to identify patterns of genome expansion and the evolution of genome complexity. All but seven of the rhabdoviruses clustered into 17 well-supported monophyletic groups, of which eight corresponded to established genera, seven were assigned as new genera, and two were taxonomically ambiguous. We show that the acquisition and loss of new genes appears to have been a central theme of rhabdovirus evolution, and has been associated with the appearance of alternative, overlapping and consecutive ORFs within the major structural protein genes, and the insertion and loss of additional ORFs in each gene junction in a clade-specific manner. Changes in the lengths of gene junctions accounted for as much as 48.5% of the variation in genome size from the smallest to the largest genome, and the frequency with which new ORFs were observed increased in the 3' to 5' direction along the genome. We also identify several new families of accessory genes encoded in these regions, and show that non-canonical expression strategies involving TURBS-like termination-reinitiation, ribosomal frame-shifts and leaky ribosomal scanning appear to be common. We conclude that rhabdoviruses have an unusual capacity for genomic plasticity that may be linked to their discontinuous transcription strategy from the negative-sense single-stranded RNA genome, and propose a model that accounts for the regular occurrence of genome expansion and contraction throughout the evolution of the Rhabdoviridae.
Assuntos
Evolução Molecular , Genoma Viral/fisiologia , Fases de Leitura Aberta/fisiologia , RNA Viral/genética , Rhabdoviridae/genética , Sequência de Bases , Dados de Sequência MolecularRESUMO
BACKGROUND: Mosquitoes are responsible for transmission of viruses, including dengue, West Nile and chikungunya viruses. Female mosquitoes are infected when they blood-feed on vertebrates, a required step for oogenesis. During this process, mosquitoes encounter high iron loads. Since iron is an essential nutrient for most organisms, including pathogens, one of the defense mechanisms for the host includes sequestration of iron away from the invading pathogen. Here, we determine whether iron availability affects viral replication in mosquitoes. METHODS: To elucidate effect of iron availability on mosquito cells during infection, Culex cells were treated with either ferric ammonium citrate (FAC) or the iron chelator, deferoxamine (DFX). Real time RT-PCR was performed using ferritin (heavy chain) and NRAMP as a measure of iron homeostasis in cells. To determine iron requirement for viral replication, Culex cells were knocked down for NRAMP using dsRNA. Finally, the results were validated in Culex mosquito-infection model, by treating infected mosquitoes with DFX to reduce iron levels. RESULTS: Our results show that infection of Culex cells led to induction in levels of ferritin (heavy chain) and NRAMP mRNAs in time-dependent manner. Results also showed that treatment of cells with FAC, reduced expression of NRAMP (iron transporter) and increase levels of ferritin (heavy chain). Interestingly, increasing iron levels increased viral titers; while reducing intracellular iron levels, either by NRAMP knock-down or using DFX, reduced viral titers. The results from Culex mosquito infection showed that mosquitoes treated with DFX had reduced viral titers compared with untreated controls in midgut as well as carcass 8 days pi. Saliva from mosquitoes treated with DFX also showed reduced viral titers compared with untreated controls, indicating low viral transmission capacity. CONCLUSIONS: Our results indicate that iron is required for viral replication in mosquito cells. Mosquitoes respond to viral infection, by inducing expression of heavy chain ferritin, which sequesters available iron, reducing its availability to virus infected cells. The data indicates that heavy chain ferritin may be part of an immune mechanism of mosquitoes in response to viral infections.
Assuntos
Culex/virologia , Ferro/metabolismo , Mosquitos Vetores/virologia , Oligoelementos/metabolismo , Replicação Viral , Vírus do Nilo Ocidental/fisiologia , Animais , Linhagem Celular , Culex/imunologia , Ferritinas/metabolismo , Imunidade Inata , Mosquitos Vetores/imunologia , Ligação ProteicaRESUMO
UNLABELLED: Wongabel virus (WONV) is an arthropod-borne rhabdovirus that infects birds. It is one of the growing array of rhabdoviruses with complex genomes that encode multiple accessory proteins of unknown function. In addition to the five canonical rhabdovirus structural protein genes (N, P, M, G, and L), the 13.2-kb negative-sense single-stranded RNA (ssRNA) WONV genome contains five uncharacterized accessory genes, one overlapping the N gene (Nx or U4), three located between the P and M genes (U1 to U3), and a fifth one overlapping the G gene (Gx or U5). Here we show that WONV U3 is expressed during infection in insect and mammalian cells and is required for efficient viral replication. A yeast two-hybrid screen against a mosquito cell cDNA library identified that WONV U3 interacts with the 83-amino-acid (aa) C-terminal domain of SNF5, a component of the SWI/SNF chromatin remodeling complex. The interaction was confirmed by affinity chromatography, and nuclear colocalization was established by confocal microscopy. Gene expression studies showed that SNF5 transcripts are upregulated during infection of mosquito cells with WONV, as well as West Nile virus (Flaviviridae) and bovine ephemeral fever virus (Rhabdoviridae), and that SNF5 knockdown results in increased WONV replication. WONV U3 also inhibits SNF5-regulated expression of the cytokine gene CSF1. The data suggest that WONV U3 targets the SWI/SNF complex to block the host response to infection. IMPORTANCE: The rhabdoviruses comprise a large family of RNA viruses infecting plants, vertebrates, and invertebrates. In addition to the major structural proteins (N, P, M, G, and L), many rhabdoviruses encode a diverse array of accessory proteins of largely unknown function. Understanding the role of these proteins may reveal much about host-pathogen interactions in infected cells. Here we examine accessory protein U3 of Wongabel virus, an arthropod-borne rhabdovirus that infects birds. We show that U3 enters the nucleus and interacts with SNF5, a component of the chromatin remodeling complex that is upregulated in response to infection and restricts viral replication. We also show that U3 inhibits SNF5-regulated expression of the cytokine colony-stimulating factor 1 (CSF1), suggesting that it targets the chromatin remodeling complex to block the host response to infection. This study appears to provide the first evidence of a virus targeting SNF5 to inhibit host gene expression.
Assuntos
Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona/metabolismo , Interações Hospedeiro-Patógeno , Rhabdoviridae/imunologia , Rhabdoviridae/fisiologia , Fatores de Transcrição/metabolismo , Proteínas Virais/metabolismo , Animais , Linhagem Celular , Núcleo Celular/química , Cromatografia de Afinidade , Insetos , Mamíferos , Microscopia Confocal , Técnicas do Sistema de Duplo-HíbridoRESUMO
We undertook a quantitative trait locus (QTL) analysis in mice to identify modifier genes that might influence the severity of human iron disorders. We identified a strong QTL on mouse chromosome 9 that differentially affected macrophage iron burden in C57BL/10J and SWR/J mice. A C57BL/10J missense allele of an evolutionarily conserved gene, Mon1a, cosegregated with the QTL in congenic mouse lines. We present evidence that Mon1a is involved in trafficking of ferroportin, the major mammalian iron exporter, to the surface of iron-recycling macrophages. Differences in amounts of surface ferroportin correlate with differences in cellular iron content. Mon1a is also important for trafficking of cell-surface and secreted molecules unrelated to iron metabolism, suggesting that it has a fundamental role in the mammalian secretory apparatus.
Assuntos
Proteínas de Transporte/genética , Ferro/metabolismo , Macrófagos/metabolismo , Locos de Características Quantitativas , Animais , Proteínas de Transporte/metabolismo , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Cromossomos de Mamíferos , Cruzamentos Genéticos , Feminino , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos , Transporte Proteico , RNA Interferente PequenoRESUMO
Mitochondrial iron is essential for the biosynthesis of heme and iron-sulfur ([Fe-S]) clusters in mammalian cells. In developing erythrocytes, iron is imported into the mitochondria by MFRN1 (mitoferrin-1, SLC25A37). Although loss of MFRN1 in zebrafish and mice leads to profound anemia, mutant animals showed no overt signs of porphyria, suggesting that mitochondrial iron deficiency does not result in an accumulation of protoporphyrins. Here, we developed a gene trap model to provide in vitro and in vivo evidence that iron regulatory protein-1 (IRP1) inhibits protoporphyrin accumulation. Mfrn1(+/gt);Irp1(-/-) erythroid cells exhibit a significant increase in protoporphyrin levels. IRP1 attenuates protoporphyrin biosynthesis by binding to the 5'-iron response element (IRE) of alas2 mRNA, inhibiting its translation. Ectopic expression of alas2 harboring a mutant IRE, preventing IRP1 binding, in Mfrn1(gt/gt) cells mimics Irp1 deficiency. Together, our data support a model whereby impaired mitochondrial [Fe-S] cluster biogenesis in Mfrn1(gt/gt) cells results in elevated IRP1 RNA-binding that attenuates ALAS2 mRNA translation and protoporphyrin accumulation.
Assuntos
5-Aminolevulinato Sintetase/metabolismo , Regulação da Expressão Gênica , Proteína 1 Reguladora do Ferro/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Porfirias/metabolismo , Animais , Blastocisto/citologia , Diferenciação Celular , Linhagem Celular Tumoral , Feminino , Genótipo , Células HEK293 , Heme/química , Humanos , Ferro/química , Proteínas Ferro-Enxofre/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Biossíntese de Proteínas , Protoporfirinas/metabolismo , Peixe-ZebraRESUMO
Although West Nile virus (WNV) and other arthropod-borne viruses are a major public health problem, the mechanisms of antiviral immunity in mosquitoes are poorly understood. Dicer-2, responsible for the RNAi-mediated response through the C-terminal RNase-III domain, also contains an N-terminal DExD/H-box helicase domain similar to mammalian RIG-I/MDA5 which, in Drosophila, was found to be required for activation of an antiviral gene, Vago. Here we show that the Culex orthologue of Vago (CxVago) is up-regulated in response to WNV infection in a Dicer-2-dependent manner. Further, our data show that CxVago is a secreted peptide that restricts WNV infection by activation of the Jak-STAT pathway. Thus, Vago appears to function as an IFN-like antiviral cytokine in mosquitoes.
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Culex/imunologia , Citocinas/imunologia , Imunidade Inata/fisiologia , Proteínas de Insetos/imunologia , Janus Quinases/imunologia , Fatores de Transcrição STAT/imunologia , Vírus do Nilo Ocidental/imunologia , Animais , Chlorocebus aethiops , Cricetinae , Culex/metabolismo , Culex/virologia , Citocinas/metabolismo , RNA Helicases DEAD-box/imunologia , RNA Helicases DEAD-box/metabolismo , Drosophila melanogaster , Humanos , Proteínas de Insetos/metabolismo , Janus Quinases/metabolismo , RNA Viral/imunologia , RNA Viral/metabolismo , Ribonuclease III/imunologia , Ribonuclease III/metabolismo , Fatores de Transcrição STAT/metabolismo , Células Vero , Febre do Nilo Ocidental/imunologia , Febre do Nilo Ocidental/metabolismo , Febre do Nilo Ocidental/transmissão , Vírus do Nilo Ocidental/metabolismoRESUMO
Mosquito-borne diseases such as malaria, dengue, Zika, and chikungunya cause significant morbidity and mortality globally, resulting in over 600,000 deaths from malaria and around 36,000 deaths from dengue each year, with millions of people infected annually, leading to substantial economic losses. The existing mosquito control measures, such as long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS), helped to reduce the infections. However, mosquito-borne diseases are still among the deadliest diseases, forcing us to improve the existing control methods and look for alternative methods simultaneously. Advanced monitoring techniques, including remote sensing, and geographic information systems (GIS) have significantly enhanced the efficiency and effectiveness of mosquito control measures. Mosquitoes' behavioural traits, such as locomotion, blood-feeding, and fertility are the key determinants of disease transmission and epidemiology. Technological advancements, such as high-resolution cameras, infrared imaging, and artificial intelligence (AI) driven object detection models, including groundbreaking convolutional neural networks, have provided efficient and precise options to monitor various mosquito behaviours, including locomotion, oviposition, fertility, and host-seeking. However, they are not commonly employed in mosquito-based research. This review highlights the novel and significant advancements in behaviour-monitoring tools, mostly from the last decade, due to cutting-edge video monitoring technology and artificial intelligence. These advancements can offer enhanced accuracy, efficiency, and the ability to quickly process large volumes of data, enabling detailed behavioural analysis over extended periods and large sample sizes, unlike traditional manual methods prone to human error and labour-intensive. The use of behaviour-assaying techniques can support or replace existing monitoring techniques and directly contribute to improving control measures by providing more accurate and real-time data on mosquito activity patterns and responses to interventions. This enhanced understanding can help establish the role of behavioural changes in improving epidemiological models, making them more precise and dynamic. As a result, mosquito management strategies can become more adaptive and responsive, leading to more effective and targeted interventions. Ultimately, this will reduce disease transmission and significantly improve public health outcomes.
RESUMO
Understanding the flight behaviour of dengue-infected mosquitoes can play a vital role in various contexts, including modelling disease risks and developing effective interventions against dengue. Studies on the locomotor activity of dengue-infected mosquitoes have often faced challenges in terms of methodology. Some studies used small tubes, which impacted the natural movement of the mosquitoes, while others that used cages did not capture the three-dimensional flights, despite mosquitoes naturally flying in three dimensions. In this study, we utilised Mask RCNN (Region-based Convolutional Neural Network) along with cubic spline interpolation to comprehensively track the three-dimensional flight behaviour of dengue-infected Aedes aegypti mosquitoes. This analysis considered a number of parameters as characteristics of mosquito flight, including flight duration, number of flights, Euclidean distance, flight speed, and the volume (space) covered during flights. The accuracy achieved for mosquito detection and tracking was 98.34% for flying mosquitoes and 100% for resting mosquitoes. Notably, the interpolated data accounted for only 0.31%, underscoring the reliability of the results. Flight traits results revealed that exposure to the dengue virus significantly increases the flight duration (p-value 0.0135 × 10-3) and volume (space) covered during flights (p-value 0.029) whilst decreasing the total number of flights compared to uninfected mosquitoes. The study did not observe any evident impact on the Euclidean distance (p-value 0.064) and speed (p-value 0.064) of Aedes aegypti. These results highlight the intricate relationship between dengue infection and the flight behaviour of Aedes aegypti, providing valuable insights into the virus transmission dynamics. This study focused on dengue-infected Aedes aegypti mosquitoes; future research can explore the impact of other arboviruses on mosquito flight behaviour.
Assuntos
Aedes , Vírus da Dengue , Dengue , Animais , Reprodutibilidade dos Testes , Mosquitos VetoresRESUMO
Mosquitoes of the Culex genus are responsible for a large burden of zoonotic virus transmission globally. Collectively, they play a significant role in the transmission of medically significant diseases such as Japanese encephalitis virus and West Nile virus. Climate change, global trade, habitat transformation and increased urbanisation are leading to the establishment of Culex mosquitoes in new geographical regions. These novel mosquito incursions are intensifying concerns about the emergence of Culex-transmitted diseases and outbreaks in previously unaffected areas. New mosquito control methods are currently being developed and deployed globally. Understanding the complex interaction between pathogens and mosquitoes is essential for developing new control strategies for Culex species mosquitoes. This article reviews the role of Culex mosquitos as vectors of zoonotic disease, discussing the transmission of viruses across different species, and the potential use of Wolbachia technologies to control disease spread. By leveraging the insights gained from recent successful field trials of Wolbachia against Aedes-borne diseases, we comprehensively discuss the feasibility of using this technique to control Culex mosquitoes and the potential for the development of next generational Wolbachia-based control methods.
Assuntos
Culex , Controle de Mosquitos , Mosquitos Vetores , Wolbachia , Wolbachia/fisiologia , Animais , Culex/microbiologia , Culex/virologia , Mosquitos Vetores/microbiologia , Mosquitos Vetores/virologia , Humanos , Controle de Mosquitos/métodosRESUMO
Mon1a was originally identified as a modifier gene of vesicular traffic, as a mutant Mon1a allele resulted in increased localization of cell surface proteins, whereas reduced levels of Mon1a showed decreased secretory activity. Here we show that Mon1a affects different steps in the secretory pathway including endoplasmic reticulum-to-Golgi traffic. siRNA-dependent reduction of Mon1a levels resulted in a delay in the reformation of the Golgi apparatus after Brefeldin A treatment. Endoglycosidase H treatment of ts045VSVG-GFP confirmed that knockdown of Mon1a delayed endoplasmic reticulum-to-Golgi trafficking. Reductions in Mon1a also resulted in delayed trafficking from Golgi to the plasma membrane. Immunoprecipitation and mass spectrometry analysis showed that Mon1a associates with dynein intermediate chain. Reductions in Mon1a or dynein altered steady state Golgi morphology. Reductions in Mon1a delayed formation of ERGIC-53-positive vesicles, whereas reductions in dynein did not affect vesicle formation. These data provide strong evidence for a role for Mon1a in anterograde trafficking through the secretory apparatus.
Assuntos
Proteínas de Transporte/metabolismo , Retículo Endoplasmático/metabolismo , Complexo de Golgi/metabolismo , Vesículas Secretórias/metabolismo , Animais , Brefeldina A/farmacologia , Células COS , Proteínas de Transporte/genética , Chlorocebus aethiops , Dineínas/genética , Dineínas/metabolismo , Retículo Endoplasmático/genética , Glicosídeo Hidrolases/farmacologia , Complexo de Golgi/genética , Células HeLa , Humanos , Lectinas de Ligação a Manose/genética , Lectinas de Ligação a Manose/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Células NIH 3T3 , Inibidores da Síntese de Proteínas/farmacologia , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/fisiologia , Vesículas Secretórias/genéticaRESUMO
Mosquito-borne diseases cause a huge burden on public health worldwide. The viruses that cause these diseases impact the behavioural traits of mosquitoes, including locomotion and feeding. Understanding these traits can help in improving existing epidemiological models and developing effective mosquito traps. However, it is difficult to understand the flight behaviour of mosquitoes due to their small sizes, complicated poses, and seemingly random moving patterns. Currently, no open-source tool is available that can detect and track resting or flying mosquitoes. Our work presented in this paper provides a detection and trajectory estimation method using the Mask RCNN algorithm and spline interpolation, which can efficiently detect mosquitoes and track their trajectories with higher accuracy. The method does not require special equipment and works excellently even with low-resolution videos. Considering the mosquito size, the proposed method's detection performance is validated using a tracker error and a custom metric that considers the mean distance between positions (estimated and ground truth), pooled standard deviation, and average accuracy. The results showed that the proposed method could successfully detect and track the flying (≈ 96% accuracy) as well as resting (100% accuracy) mosquitoes. The performance can be impacted in the case of occlusions and background clutters. Overall, this research serves as an efficient open-source tool to facilitate further examination of mosquito behavioural traits.
Assuntos
Aedes , Animais , Algoritmos , Redes Neurais de Computação , Mosquitos VetoresRESUMO
BACKGROUND: Mosquito-borne diseases exert a huge impact on both animal and human populations, posing substantial health risks. The behavioural and fitness traits of mosquitoes, such as locomotion and fecundity, are crucial factors that influence the spread of diseases. In existing egg-counting tools, each image requires separate processing with adjustments to various parameters such as intensity threshold and egg area size. Furthermore, accuracy decreases significantly when dealing with clustered or overlapping eggs. To overcome these issues, we have developed EggCountAI, a Mask Region-based Convolutional Neural Network (RCNN)-based free automatic egg-counting tool for Aedes aegypti mosquitoes. METHODS: The study design involves developing EggCountAI for counting mosquito eggs and comparing its performance with two commonly employed tools-ICount and MECVision-using 10 microscopic and 10 macroscopic images of eggs laid by females on a paper strip. The results were validated through manual egg counting on the strips using ImageJ software. Two different models were trained on macroscopic and microscopic images to enhance egg detection accuracy, achieving mean average precision, mean average recall, and F1-scores of 0.92, 0.90, and 0.91 for the microscopic model, and 0.91, 0.90, and 0.90 for the macroscopic model, respectively. EggCountAI automatically counts eggs in a folder containing egg strip images, offering adaptable filtration for handling impurities of varying sizes. RESULTS: The results obtained from EggCountAI highlight its remarkable performance, achieving overall accuracy of 98.88% for micro images and 96.06% for macro images. EggCountAI significantly outperformed ICount and MECVision, with ICount achieving 81.71% accuracy for micro images and 82.22% for macro images, while MECVision achieved 68.01% accuracy for micro images and 51.71% for macro images. EggCountAI also excelled in other statistical parameters, with mean absolute error of 1.90 eggs for micro, 74.30 eggs for macro, and a strong correlation and R-squared value (0.99) for both micro and macro. The superior performance of EggCountAI was most evident when handling overlapping or clustered eggs. CONCLUSION: Accurate detection and counting of mosquito eggs enables the identification of preferred egg-laying sites and facilitates optimal placement of oviposition traps, enhancing targeted vector control efforts and disease transmission prevention. In future research, the tool holds the potential to extend its application to monitor mosquito feeding preferences.