Odorant receptors for floral- and plant-derived volatiles in the yellow fever mosquito, Aedes aegypti (Diptera: Culicidae).

Adult mosquitoes require regular sugar meals, including nectar, to survive in natural habitats. Both males and females locate potential sugar sources using sensory proteins called odorant receptors (ORs) activated by plant volatiles to orient toward flowers or honeydew. The yellow fever mosquito, Aedes aegypti (Linnaeus, 1762), possesses a large gene family of ORs, many of which are likely to detect floral odors. In this study, we have uncovered ligand-receptor pairings for a suite of Aedes aegypti ORs using a panel of environmentally relevant, plant-derived volatile chemicals and a heterologous expression system. Our results support the hypothesis that these odors mediate sensory responses to floral odors in the mosquito's central nervous system, thereby influencing appetitive or aversive behaviors. Further, these ORs are well conserved in other mosquitoes, suggesting they function similarly in diverse species. This information can be used to assess mosquito foraging behavior and develop novel control strategies, especially those that incorporate mosquito bait-and-kill technologies.

Odorant receptors for floral- and plant-derived volatiles in the yellow fever mosquito, Aedes aegypti (Diptera: Culicidae).

Adult mosquitoes require regular sugar meals, usually floral nectar, to survive and flourish in natural habitats. Both males and females locate potential sugar sources using sensory proteins called odorant receptors activated by plant volatiles that facilitate orientation toward flowers or honeydew. The Yellow Fever mosquito, Aedes aegypti (Linnaeus, 1762), possesses a large repertoire of odorant receptors, many of which are likely to support floral odor detection and nectar-seeking. In this study, we have employed a heterologous expression system and the two-electrode voltage clamping technique to identify environmentally relevant chemical compounds that activate specific odorant receptors. Importantly, we have uncovered ligand-receptor pairings for a suite of Aedes aegypti odorant receptors likely to mediate appetitive or aversive behavioral responses, thus shaping a critical aspect of the life history of a medically important mosquito. Moreover, the high degree of conservation of these receptors in other disease-transmitting species suggests common mechanisms of floral odor detection. This knowledge can be used to further investigate mosquito foraging behavior to either enhance existing, or develop novel, control strategies, especially those that incorporate mosquito bait-and-kill or attractive toxic sugar bait technologies.

The evolution of borneol repellency in culicine mosquitoes.

Insects have developed remarkable adaptations to effectively interact with plant secondary metabolites and utilize them as cues to identify suitable hosts. Consequently, humans have used aromatic plants for centuries to repel mosquitoes. The repellent effects of plant volatile compounds are mediated through olfactory structures present in the antennae, and maxillary palps of mosquitoes. Mosquito maxillary palps contain capitate-peg sensilla, which house three olfactory sensory neurons, of which two are mainly tuned to either carbon dioxide or octenol - two animal host odorants. However, the third neuron, which expresses the OR49 receptor, has remained without a known ecologically-relevant odorant since its initial discovery. In this study, we used odorant mixtures and terpenoid-rich Cannabis essential oils to investigate the activation of OR49. Our results demonstrate that two monoterpenoids, borneol and camphor, selectively activate OR49, and OR9-expressing neurons, as well as the MD3 glomerulus in the antennal lobe. We confirm that borneol repels female mosquitoes, and knocking out the gene encoding the OR49 receptor suppresses the response of the corresponding olfactory sensory neuron. Importantly, this molecular mechanism of action is conserved across culicine mosquito species, underscoring its significance in their olfactory systems.

Carboxylic acids that drive mosquito attraction to humans activate ionotropic receptors.

The mosquito, Aedes aegypti, is highly anthropophilic and transmits debilitating arboviruses within human populations and between humans and non-human primates. Female mosquitoes are attracted to sources of blood by responding to odor plumes that are emitted by their preferred hosts. Acidic volatile compounds, including carboxylic acids, represent particularly salient odors driving this attraction. Importantly, carboxylic acids are major constituents of human sweat and volatiles generated by skin microbes. As such, they are likely to impact human host preference, a dominant factor in disease transmission cycles. A more complete understanding of mosquito host attraction will necessitate the elucidation of molecular mechanisms of volatile odor detection that function in peripheral sensory neurons. Recent studies have shown that members of the variant ionotropic glutamate receptor gene family are necessary for physiological and behavioral responses to acidic volatiles in Aedes. In this study, we have identified a subfamily of variant ionotropic receptors that share sequence homology across several important vector species and are likely to be activated by carboxylic acids. Moreover, we demonstrate that selected members of this subfamily are activated by short-chain carboxylic acids in a heterologous cell expression system. Our results are consistent with the hypothesis that members of this receptor class underlie acidic volatile sensitivity in vector mosquitoes and provide a frame of reference for future development of novel mosquito attractant and repellent technologies.

Recurring germline mosaicism in a family due to reversion of an inherited derivative chromosome 8 from an 8;21 translocation with interstitial telomeric sequences.

BACKGROUND: Mosaicism for chromosomal structural abnormalities, other than marker or ring chromosomes, is rarely inherited. METHODS: We performed cytogenetics studies and breakpoint analyses on a family with transmission of mosaicism for a derivative chromosome 8 (der(8)), resulting from an unbalanced translocation between the long arms of chromosomes 8 and 21 over three generations. RESULTS: The proband and his maternal half-sister had mosaicism for a der(8) cell line leading to trisomy of the distal 21q, and both had Down syndrome phenotypic features. Mosaicism for a cell line with the der(8) and a normal cell line was also detected in a maternal half-cousin. The der(8) was inherited from the maternal grandmother who had four abnormal cell lines containing the der(8), in addition to a normal cell line. One maternal half-aunt had the der(8) and an isodicentric chromosome 21 (idic(21)). Sequencing studies revealed microhomologies at the junctures of the der(8) and idic(21) in the half-aunt, suggesting a replicative mechanism in the rearrangement formation. Furthermore, interstitial telomeric sequences (ITS) were identified in the juncture between chromosomes 8 and 21 in the der(8). CONCLUSION: Mosaicism in the proband, his half-sister and half-cousin resulting from loss of chromosome 21 material from the der(8) appears to be a postzygotic event due to the genomic instability of ITS and associated with selective growth advantage of normal cells. The reversion of the inherited der(8) to a normal chromosome 8 in this family resembles revertant mosaicism of point mutations. We propose that ITS could mediate recurring revertant mosaicism for some constitutional chromosomal structural abnormalities.

Functional conservation of Anopheline linalool receptors through 100 million years of evolution.

Insects rely on olfactory receptors to detect and respond to diverse environmental chemical cues. Detection of semiochemicals by these receptors modulates insect behavior and has a direct impact on species fitness. Volatile organic compounds (VOCs) are released by animals and plants and can provide contextual cues that a blood meal host or nectar source is present. One such VOC is linalool, an enantiomeric monoterpene, that is emitted from plants and bacteria species. This compound exists in nature as one of two possible stereoisomers, (R)-(-)-linalool or (S)-(+)-linalool. In this study, we use a heterologous expression system to demonstrate differential responsiveness of a pair of Anopheline odorant receptors (Ors) to enantiomers of linalool. The mosquitoes Anopheles gambiae and Anopheles stephensi encode single copies of Or29 and Or53, which are expressed in the labella of An. gambiae. (S)-(+)-linalool activates Or29 orthologs with a higher potency than (R)-(-)-linalool, while the converse is observed for Or53 orthologs. The conservation of these receptors across a broad range of Anopheline species suggests they may function in the discrimination of linalool stereoisomers, thereby influencing the chemical ecology of mosquitoes. One potential application of this knowledge would be in the design of novel attractants or repellents to be used in integrated pest management practices.

Multi-Year Study of the Chemical and Sensory Effects of Microwave-Assisted Extraction of Musts and Stems in Cabernet Sauvignon, Merlot and Syrah Wines from the Central Coast of California.

Microwave technology (MW) was applied to musts and stems over three consecutive vintages in Cabernet Sauvignon, Merlot and Syrah wines from California (USA). Stems were added to musts at a rate of 50 and 100% (50% Stems and 100% Stems), either as untreated or after MW (50% MW Stems and 100% MW Stems). Stem additions lowered ethanol (up to 1.15% v/v reduction), but increased pH (up to 0.16 units) and the tannin content of the wines. In 2016, tannins increased by 103% (100% Stems), and 124% (100% MW Stems). In 2017, tannins increased by 39% in stem-added Merlot wines and by 63% (100% Stems) and 85% (100% MW Stems) in Syrah wines. In 2018, tannins in Syrah wines increased by 250% (100% MW Stems) and by 743% (100% Stems). Wines made with 50% Stems exhibited intermediate tannin contents. Must MW increased flavonols (up to 278% in Syrah wines), monoglucosylated, acylated and anthocyanin-derived pigments. Stem additions reduced wine color and polymeric pigment formation in Syrah. Must MW decreased the perception of coarseness and herbaceous flavors in Merlot, whereas stem additions increased herbaceous aromas in Syrah. Despite higher tannin contents in stem-added wines, no concomitant increases in astringency were observed.

Identification and functional characterization of olfactory indolergic receptors in Musca domestica.

In mosquitoes, indolic compounds are detected by a group of olfactory indolergic Odorant Receptors (indolORs). The ancient origin of indole and 3-methylindole as chemical signals suggest that they may be detected by insects outside the Culicidae clade. To test this hypothesis, we have identified potential indolOR genes in brachyceran flies based on sequence homology. Because of the crucial roles of indolic compounds in oviposition and foraging, we have focused our attention on the housefly Musca domestica. Using a heterologous expression system, we have identified indolOR transcript expression in the female antennae, and have characterized MdomOR30a and MdomOR49b as 3-methylindole and indole receptors, respectively. We have identified a set of 92 putative indolOR genes encoded in the genomes of Culicoidea, Psychodidae and brachycera, described their phylogenetic relationships, and exon/intron structures. Further characterization of indolORs will impact our understanding of insect chemical ecology and will provide targets for the development of novel odor-based tools that can be integrated into existing vector surveillance and control programs.

Carboxylic acid responses by a conserved odorant receptor in culicine vector mosquitoes.

Many mosquito behaviours that are critical for survival and reproduction depend upon timely responses to chemical cues. Of interest are the effects of volatile organic compounds like carboxylic acids (CAs) that are released by potential blood meal hosts. Short chain CAs are among the primary attractants for host-seeking females and influence host selection in vector species. Although the behavioural relevance of CA's has been established, less is known about the molecular receptive events that evoke responses to specific compounds, with the Ir family of chemoreceptors being broadly implicated in their detection. In this study, we demonstrate that Or orthologs from two vector species, Aedes aegypti (L.) and Aedes albopictus (Skuse), are selectively activated by straight chain carboxylic acids and that these responses are attenuated by the commercial insect repellant N,N-Diethyl-meta-toluamide. Our results suggest that multiple chemoreceptors, representing diverse families, are able to mediate molecular responses to CAs and may therefore underlie important behaviours that directly impact disease-transmission cycles.

Identification of Mosquito Bloodmeals Collected in Diverse Habitats in Malaysian Borneo Using COI Barcoding.

Land cover and land use change (LCLUC) acts as a catalyst for spillover of arthropod-borne pathogens into novel hosts by shifting host and vector diversity, abundance, and distribution, ultimately reshaping host-vector interactions. Identification of bloodmeals from wild-caught mosquitoes provides insight into host utilization of particular species in particular land cover types, and hence their potential role in pathogen maintenance and spillover. Here, we collected 134 blood-engorged mosquitoes comprising 10 taxa across 9 land cover types in Sarawak, Malaysian Borneo, a region experiencing intense LCLUC and concomitant spillover of arthropod-borne pathogens. Host sources of blood were successfully identified for 116 (87%) mosquitoes using cytochrome oxidase subunit I (COI) barcoding. A diverse range of hosts were identified, including reptiles, amphibians, birds, and mammals. Sixteen engorged Aedes albopictus, a major vector of dengue virus, were collected from seven land cover types and found to feed exclusively on humans (73%) and boar (27%). Culex tritaeniohynchus (n = 2), Cx. gelidus (n = 3), and Cx. quiquefasciatus (n = 3), vectors of Japanese encephalitis virus, fed on humans and pigs in the rural built-up land cover, creating potential transmission networks between these species. Our data support the use of COI barcoding to characterize mosquito-host networks in a biodiversity hotspot.

An odorant receptor from Anopheles gambiae that demonstrates enantioselectivity to the plant volatile, linalool.

Insects express chemical receptors within sensory neurons that are activated by specific cues in the environment, thereby influencing the acquisition of critical resources. A significant gap in our current understanding of insect chemical ecology is defining the molecular mechanisms that underlie sensitivity to plant-emitted volatiles. Linalool is a commonly-occurring monoterpene that has various effects on insect behavior, either acting as an attractant or a repellent, and existing in nature as one of two possible stereoisomers, (R)-(-)-linalool and (S)-(+)-linalool. In this study, we have used a cell-based functional assay to identify linalool and structurally-related compounds as ligands of Odorant receptor 29, a labellum-expressed receptor in the malaria vector mosquito, Anopheles gambiae (AgamOr29). While (R)-(-)-linalool activates AgamOr29, a mixture of the (R) and (S) stereoisomers activates the receptor with higher potency, implying enantiomeric selectivity. Orthologs of Or29 are present in the genomes of Anophelines within the Cellia subgenus. The conservation of this receptor across Anopheline lineages suggests that this ecologically important compound might serve as an attraction cue for nectar-seeking mosquitoes. Moreover, the characterization of a mosquito terpene receptor could serve as a foundation for future ligand-receptor studies of plant volatiles and for the discovery of compounds that can be integrated into push-pull vector control strategies.

Chemical consequences of extended maceration and post-fermentation additions of grape pomace in Pinot noir and Zinfandel wines from the Central Coast of California (USA).

Pinot noir and Zinfandel wines were produced with traditional maceration (Control), and extended maceration for one (1 month-EM), and six months (6 months-EM). Addition of an extra amount of pomace during EM was also evaluated through a treatment referred to as Double pomace. The application of EM and Double pomace decreased the chromatic as well as the anthocyanin and anthocyanin-derived pigment composition of the wines. Large polymeric pigments were favored in EM and Double pomace wines, but the total polymeric pigment content was not improved by any of the winemaking treatments. The 6 months-EM wines showed a 13-fold (Pinot noir) and a 1.6-fold increase (Zinfandel) in tannins, respectively, relative to Control wines. However, the 1 month-EM and Double pomace treatments did not affect tannin levels, suggesting that in these wines, the extraction of tannins during EM may depend upon desorption of previously extracted tannins during extended contact time.

Nanoparticles of poorly water-soluble drugs prepared by supercritical fluid extraction of emulsions.

PURPOSE: The aim of the study was to develop and evaluate a new method for the production of micro- and nanoparticles of poorly soluble drugs for drug delivery applications. METHODS: Fine particles of model compounds cholesterol acetate (CA), griseofulvin (GF), and megestrol acetate (MA) were produced by extraction of the internal phase of oil-in-water emulsions using supercritical carbon dioxide. The particles were obtained both in a batch or a continuous manner in the form of aqueous nanosuspensions. Precipitation of CA nanoparticles was used for conducting a mechanistic study on particle size control and scale-up. GF and MA nanoparticles were produced in several batches to compare their dissolution behavior with that of micronized materials. The physical analysis of the particles produced was performed using dynamic light scattering (particle size), scanning electron microscopy (morphology), powder X-ray diffraction (crystallinity), gas chromatography (residual solvent), and a dissolution apparatus. RESULTS: Particles with mean volume diameter ranging between 100 and 1000 nm were consistently produced. The emulsion droplet size, drug solution concentration, and organic solvent content in the emulsion were the major parameters responsible for particle size control. Efficient and fast extraction, down to low parts-per-million levels, was achieved with supercritical CO2. The GF and MA nanoparticles produced were crystalline in nature and exhibited a 5- to 10-fold increase in the dissolution rate compared with that of micronized powders. Theoretical calculations indicated that this dissolution was governed mainly by the surface kinetic coefficient and the specific surface area of the particles produced. It was observed that the necessary condition for a reliable and scalable process was the sufficient emulsion stability during the extraction time. CONCLUSION: The method developed offers a viable alternative to both the milling and constructive nanoparticle formation processes. Although preparation of a stable emulsion can be a challenge for some drug molecules, the new technique significantly shortens the processing time and overcomes the current limitations of the conventional precipitation techniques in terms of large waste streams, product purity, and process scale-up.