The corpora allata-corpora cardiaca (CA-CC) is an endocrine gland complex that regulates mosquito development and reproduction through the synthesis of juvenile hormone (JH). Epoxidase (Epox) is a key enzyme in the production of JH. We recently utilized CRISPR/Cas9 to establish an epoxidase-deficient (epox-/-) Aedes aegypti line. The CA from epox-/- mutants do not synthesize epoxidated JH III but methyl farneosate (MF), a weak agonist of the JH receptor, and therefore have reduced JH signalling. Illumina sequencing was used to examine the differences in gene expression between the CA-CC from wild type (WT) and epox-/- adult female mosquitoes. From 18,034 identified genes, 317 were significantly differentially expressed. These genes are involved in many biological processes, including the regulation of cell proliferation and apoptosis, energy metabolism, and nutritional uptake. In addition, the same CA-CC samples were also used to examine the microRNA (miRNA) profiles of epox-/- and WT mosquitoes. A total of 197 miRNAs were detected, 24 of which were differentially regulated in epox-/- mutants. miRNA binding sites for these particular miRNAs were identified using an in silico approach; they target a total of 101 differentially expressed genes. Our results suggest that a lack of epoxidase, besides affecting JH synthesis, results in the diminishing of JH signalling that have significant effects on Ae. aegypti CA-CC transcriptome profiles, as well as its miRNA repertoire.
- MeSH
- Aedes * genetika metabolismus MeSH
- corpora allata metabolismus MeSH
- exprese genu MeSH
- juvenilní hormony metabolismus MeSH
- mikro RNA * genetika metabolismus MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Juvenile hormone (JH) controls the development and reproduction of insects. Therefore, a tight regulation of the expression of JH biosynthetic enzymes is critical. microRNAs (miRNAs) play significant roles in the post-transcriptional regulation of gene expression by interacting with complementary sequences in target genes. Previously, we reported that several miRNAs were differentially expressed during three developmental stages of Aedes aegypti mosquitoes with different JH levels (no JH, high JH, and low JH). One of these miRNAs was aae-miR-34-5p. In this study, we identified the presence of potential target sequences of aae-miR-34-5p in the transcripts of some genes encoding JH biosynthetic enzymes. We analysed the developmental expression patterns of aae-miR-34-5p and the predicted target genes involved in JH biogenesis. Increases in miRNA abundance were followed, with a delay, by decreases in transcript levels of target genes. Application of an inhibitor and a mimic of aae-miR-34-5p led respectively to increased and decreased levels of thiolase transcripts, which is one of the early genes of JH biosynthesis. Female adult mosquitoes injected with an aae-miR-34-5p inhibitor exhibited significantly increased transcript levels of three genes encoding JH biosynthetic enzymes, acetoacetyl-CoA thiolase (thiolase), farnesyl diphosphate phosphatase, and farnesal dehydrogenase. Overall, our results suggest a potential role of miRNAs in JH production by directly targeting genes involved in its biosynthesis.
- MeSH
- Aedes virologie MeSH
- dengue epidemiologie přenos MeSH
- horečka chikungunya epidemiologie přenos MeSH
- infekce virem zika epidemiologie přenos MeSH
- komáří přenašeči * virologie MeSH
- lidé MeSH
- virové nemoci epidemiologie přenos MeSH
- virus La Crosse MeSH
- žlutá zimnice epidemiologie přenos MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
Methyl farnesoate (MF) plays hormonal regulatory roles in crustaceans. An epoxidated form of MF, known as juvenile hormone (JH), controls metamorphosis and stimulates reproduction in insects. To address the evolutionary significance of MF epoxidation, we generated mosquitoes completely lacking either of the two enzymes that catalyze the last steps of MF/JH biosynthesis and epoxidation, respectively: the JH acid methyltransferase (JHAMT) and the P450 epoxidase CYP15 (EPOX). jhamt-/- larvae lacking both MF and JH died at the onset of metamorphosis. Strikingly, epox-/- mutants, which synthesized MF but no JH, completed the entire life cycle. While epox-/- adults were fertile, the reproductive performance of both sexes was dramatically reduced. Our results suggest that although MF can substitute for the absence of JH in mosquitoes, it is with a significant fitness cost. We propose that MF can fulfill most roles of JH, but its epoxidation to JH was a key innovation providing insects with a reproductive advantage.
- MeSH
- Aedes enzymologie genetika MeSH
- biologická proměna MeSH
- genetická zdatnost * MeSH
- juvenilní hormony biosyntéza MeSH
- molekulární evoluce * MeSH
- nenasycené mastné kyseliny metabolismus MeSH
- rozmnožování MeSH
- seskviterpeny metabolismus MeSH
- sexuální chování zvířat MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Defective viral genomes (DVGs) are truncated and/or rearranged viral genomes produced during virus replication. Described in many RNA virus families, some of them have interfering activity on their parental virus and/or strong immunostimulatory potential, and are being considered in antiviral approaches. Chikungunya virus (CHIKV) is an alphavirus transmitted by Aedes spp. that infected millions of humans in the last 15 years. Here, we describe the DVGs arising during CHIKV infection in vitro in mammalian and mosquito cells, and in vivo in experimentally infected Aedes aegypti mosquitoes. We combined experimental and computational approaches to select DVG candidates most likely to have inhibitory activity and showed that, indeed, they strongly interfere with CHIKV replication both in mammalian and mosquito cells. We further demonstrated that some DVGs present broad-spectrum activity, inhibiting several CHIKV strains and other alphaviruses. Finally, we showed that pre-treating Aedes aegypti with DVGs prevented viral dissemination in vivo.
- MeSH
- Aedes virologie MeSH
- antivirové látky farmakologie MeSH
- defektní viry genetika MeSH
- genom virový * MeSH
- horečka chikungunya imunologie přenos virologie MeSH
- komáří přenašeči virologie MeSH
- lidé MeSH
- replikace viru * MeSH
- virus chikungunya genetika růst a vývoj izolace a purifikace MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
- MeSH
- Aedes MeSH
- chronická hepatitida C epidemiologie prevence a kontrola MeSH
- COVID-19 epidemiologie farmakoterapie MeSH
- infekce přenášené vektorem MeSH
- klíšťová encefalitida epidemiologie MeSH
- klostridiové infekce epidemiologie MeSH
- kongresy jako téma MeSH
- kontrola infekčních nemocí * MeSH
- lidé MeSH
- listeriové infekce epidemiologie MeSH
- nemoci novorozenců MeSH
- očkovací programy organizace a řízení statistika a číselné údaje MeSH
- pemfigus epidemiologie MeSH
- pneumokokové infekce prevence a kontrola MeSH
- pokrytí očkováním MeSH
- sérologické testování na COVID-19 statistika a číselné údaje MeSH
- tularemie epidemiologie MeSH
- Check Tag
- lidé MeSH
ETHNOPHARMACOLOGICAL RELEVANCE: The use of Hazomalania voyronii, popularly known as hazomalana, to repel mosquitoes and resist against insect attacks is handed down from generation to generation in Madagascar. In the present study, we investigated the ability of the essential oils (EOs) obtained from the stem wood, fresh and dry bark of H. voyronii to keep important mosquito vectors (Aedes aegypti and Culex quinquefasciatus) away, as well as their toxicity on three insect species of agricultural and public health importance (Cx. quinquefasciatus, Musca domestica and Spodoptera littoralis). MATERIALS AND METHODS: Hydrodistillation was used to obtain EOs from stem wood, fresh and dry bark. The chemical compositions were achieved by gas chromatography-mass spectrometry (GC-MS). Toxicity assays using stem wood and bark EOs were performed on larvae of Cx. quinquefasciatus and S. littoralis, and adults of M. domestica by WHO and topical application methods, respectively. Mosquito repellent activity of the most effective EO, i.e. the bark one, was determined on human volunteers by arm-in-cage tests, and results were compared with that of the commercial repellent N,N-ddiethyl-m-toluamide (DEET). RESULTS: The H. voyronii EOs were characterized by oxygenated monoterpenes with perilla aldehyde (30.9-47.9%) and 1,8-cineole (19.7-33.2%) as the main constituents. The fresh and dry bark EOs were the most active on Cx. quinquefasciatus and S. littoralis larvae, respectively, with LC50/LD50 of 65.5 mg L-1, and 50.5 μg larva-1; the EOs from wood and fresh bark displayed the highest toxicity on M. domestica (LD50 values 60.8 and 65.8 μg adult-1, respectively). Repellence assay revealed an almost complete protection (>80%) from both mosquito species for 30 min when pure fresh bark EO was applied on the volunteers' arm, while DEET 10% repelled >80% of the mosquitoes up to 120 min from application. CONCLUSION: The traditional use of the bark EO to repel insects has been demonstrated although an extended-release formulation based on H. voyronii EOs is needed to increase the repellent effect over time. A wide spectrum of insecticidal activity has been provided as well, suggesting a possible use of H. voyronii EOs in the fabrication of green repellents and insecticides useful to control mosquito vectors and agricultural pests.
- MeSH
- Aedes účinky léků růst a vývoj MeSH
- časové faktory MeSH
- Culex účinky léků embryologie MeSH
- diethyltoluamid farmakologie MeSH
- dřevo * chemie MeSH
- Hernandiaceae * chemie MeSH
- kůra rostlin * chemie MeSH
- larva účinky léků růst a vývoj MeSH
- lidé MeSH
- moskyti - kontrola * MeSH
- moucha domácí účinky léků růst a vývoj MeSH
- oleje prchavé izolace a purifikace farmakologie MeSH
- oleje rostlin izolace a purifikace farmakologie MeSH
- repelenty proti hmyzu izolace a purifikace farmakologie MeSH
- Spodoptera účinky léků embryologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
Symbiotic bacteria affect competence for pathogen transmission in insect vectors, including mosquitoes. However, knowledge on mosquito-microbiome-pathogen interactions remains limited, largely due to methodological reasons. The current, cost-effective practice of sample pooling used in mosquito surveillance and epidemiology prevents correlation of individual traits (i.e., microbiome profile) and infection status. Moreover, many mosquito studies employ laboratory-reared colonies that do not necessarily reflect the natural microbiome composition and variation in wild populations. As a consequence, epidemiological and microbiome studies in mosquitoes are to some extent uncoupled, and the interactions among pathogens, microbiomes, and natural mosquito populations remain poorly understood. This study focuses on the effect the pooling practice poses on mosquito microbiome profiles, and tests different approaches to find an optimized low-cost methodology for extensive sampling while allowing for accurate, individual-level microbiome studies. We tested the effect of pooling by comparing wild-caught, individually processed mosquitoes with pooled samples. With individual mosquitoes, we also tested two methodological aspects that directly affect the cost and feasibility of broad-scale molecular studies: sample preservation and tissue dissection. Pooling affected both alpha- and beta-diversity measures of the microbiome, highlighting the importance of using individual samples when possible. Both RNA and DNA yields were higher when using inexpensive reagents such as NAP (nucleic acid preservation) buffer or absolute ethanol, without freezing for short-term storage. Microbiome alpha- and beta-diversity did not show overall significant differences between the tested treatments compared to the controls (freshly extracted samples or dissected guts). However, the use of standardized protocols is highly recommended to avoid methodological bias in the data.
- MeSH
- Aedes mikrobiologie MeSH
- Bacteria klasifikace genetika MeSH
- Culex mikrobiologie MeSH
- komáří přenašeči mikrobiologie MeSH
- mikrobiologické techniky MeSH
- mikrobiota * MeSH
- ochrana biologická MeSH
- střevní mikroflóra MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Insects rely on the innate immune system for defense against pathogens, some aspects of which are under hormonal control. Here we provide direct experimental evidence showing that the juvenile hormone-binding protein (mJHBP) of Aedes aegypti is required for the regulation of innate immune responses and the development of mosquito blood cells (hemocytes). Using an mJHBP-deficient mosquito line generated by means of CRISPR-Cas9 gene editing technology we uncovered a mutant phenotype characterized by immunosuppression at the humoral and cellular levels, which profoundly affected susceptibility to bacterial infection. Bacteria-challenged mosquitoes exhibited significantly higher levels of septicemia and mortality relative to the wild type (WT) strain, delayed expression of antimicrobial peptides (AMPs), severe developmental dysregulation of embryonic and larval hemocytes (reduction in the total number of hemocytes) and increased differentiation of the granulocyte lineage. Interestingly, injection of recombinant wild type mJHBP protein into adult females three-days before infection was sufficient to restore normal immune function. Similarly, injection of mJHBP into fourth-instar larvae fully restored normal larval/pupal hemocyte populations in emerging adults. More importantly, the recovery of normal immuno-activation and hemocyte development requires the capability of mJHBP to bind JH III. These results strongly suggest that JH III functions in mosquito immunity and hemocyte development in a manner that is perhaps independent of canonical JH signaling, given the lack of developmental and reproductive abnormalities. Because of the prominent role of hemocytes as regulators of mosquito immunity, this novel discovery may have broader implications for the understanding of vector endocrinology, hemocyte development, vector competence and disease transmission.
- MeSH
- Aedes genetika růst a vývoj imunologie mikrobiologie MeSH
- hemocyty imunologie mikrobiologie MeSH
- hmyzí proteiny genetika imunologie MeSH
- juvenilní hormony imunologie MeSH
- larva genetika růst a vývoj imunologie mikrobiologie MeSH
- přirozená imunita MeSH
- Serratia marcescens fyziologie MeSH
- transportní proteiny genetika imunologie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Research Support, N.I.H., Extramural MeSH
- Research Support, N.I.H., Intramural MeSH
Virové infekční nemoci přenášené hmyzím vektorem (komár, klíště) jsou celosvětově na vzestupu. Severní Amerika zaznamenala v roce 2016 například více než ctyřicet tisíc případů Lymské boreliózy, oproti 12 000 případů v roce 1995. Od roku 1999 do současnosti bylo na území USA zachyceno 38 000 nemocných West Nile virem, více než patnáct set osob onemocnění podlehlo. Onemocnění dengue, stejně jako chicungunya se začaly pravidelně vyskytovat mezi obyvatelstvem západní polokoule. Hmyzí vektor nerozlišuje mezi civilní a vojenskou populací. A jsou to právě vojáci, kdo velmi často žijí, pracují, trénují, operují ve venkovním prostředí dnes a denně na celém světě, vystaveni riziku expozice hmyzím vektorem, přenosu infekční nemoci, potencionálně i letální. Jen některá z těchto onemocnění jsou preventabilní očkováním, eventuelně lékovou profylaxí (př. malárie, pomineme-li rezistentní kmeny). Základním prostředkem proti tomuto typu virových nemocí proto stále zůstávají osobní ochranné pomůcky – u cestovatelů v první linii repelenty, u vojáků chemicky specielně ošetřené uniformy.
Viral infectious diseases transmitted by insect vectors such as mosquito or tick are growing world-wide. For example, more than forty thousand cases of Lyme Disease were detected in North America during 2016 year, compared to 12 000 cases in 1995. Since 1999 up today we have found 38 000 US patients infected by West Nile Virus. More than one thousand five hundred patients died. Dengue fever, as well as Chicungunya have begun to regularly occur between the population in the Western Hemisphere. Insect vector does not distinguish between civilian and military populations. And there are exactly soldiers, who often live, work, train, operate in the outdoors every day around the world, at risk of exposure of insect vector, infectious disease, potentially lethal. Just some of these diseases are preventable by vaccines. Anyway a personal protective equipment is still the basic protection for humans.
- Klíčová slova
- Aedes aegypti,
- MeSH
- Aedes patogenita virologie MeSH
- arbovirové infekce * prevence a kontrola přenos virologie MeSH
- dezinfekce MeSH
- diethyltoluamid MeSH
- komáří přenašeči patogenita virologie MeSH
- kontrola infekčních nemocí metody MeSH
- lidé MeSH
- odívání MeSH
- ozbrojené síly MeSH
- permethrin MeSH
- vakcinace MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- přehledy MeSH
