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2024
Marois Eric
Using the CRISPR / Cas9 system for genome editing in Anopheles mosquitoes Online
2024, visited: 08.01.2024.
Abstract | Links | BibTeX | Tags: Anopheles, CRISPR/Cas9, Genome editing, M3i, marois, mosquitoes
@online{Marois2024,
title = {Using the CRISPR / Cas9 system for genome editing in Anopheles mosquitoes},
author = { Eric Marois},
url = {https://hal.science/hal-04380430/document},
doi = {HAL Id: hal-04380430},
year = {2024},
date = {2024-01-08},
urldate = {2024-01-08},
journal = {HAL science},
abstract = {The advent of the CRISPR / Cas9 technology permits the targeted editing of mosquito genomes, ranging from site-directed mutagenesis of genes of interest yielding knockout mutations (which arise by insertion / deletion of a few nucleotides) to site-specific insertion of exogenous DNA sequences such as fluorescence markers or even large gene drive cassettes, themselves encoding the components of the CRISPR / Cas9 system. To obtain these heritable targeted changes, genome editing requires the delivery of Cas9 protein and its guide RNA(s) to the developing germ tissue of an embryo. Different species require adaptation of this basic principle to accommodate for their specific biology. Here, we describe a technical pipeline based on delivering the CRISPR/Cas9 components in the form of injected plasmid or as transgenes, resulting in highly efficient gene editing in Anopheles malaria vector mosquitoes. We have reliably employed these methods to mutagenize > 20 different loci of interest in Anopheles coluzzii to date. },
keywords = {Anopheles, CRISPR/Cas9, Genome editing, M3i, marois, mosquitoes},
pubstate = {published},
tppubtype = {online}
}
The advent of the CRISPR / Cas9 technology permits the targeted editing of mosquito genomes, ranging from site-directed mutagenesis of genes of interest yielding knockout mutations (which arise by insertion / deletion of a few nucleotides) to site-specific insertion of exogenous DNA sequences such as fluorescence markers or even large gene drive cassettes, themselves encoding the components of the CRISPR / Cas9 system. To obtain these heritable targeted changes, genome editing requires the delivery of Cas9 protein and its guide RNA(s) to the developing germ tissue of an embryo. Different species require adaptation of this basic principle to accommodate for their specific biology. Here, we describe a technical pipeline based on delivering the CRISPR/Cas9 components in the form of injected plasmid or as transgenes, resulting in highly efficient gene editing in Anopheles malaria vector mosquitoes. We have reliably employed these methods to mutagenize > 20 different loci of interest in Anopheles coluzzii to date.
2023
SC Weng, I Antoshechkin, E Marois, O Akbari
Efficient Sex Separation by Exploiting Differential Alternative Splicing of a Dominant Marker in Aedes aegypti Journal Article
In: PLoS Genetics, 2023.
Abstract | Links | BibTeX | Tags: Aedes aegypti, Introns, larvae, M3i, marois, mosquitoes, pupae, rna sequencing, transcriptome analyses
@article{nokey,
title = {Efficient Sex Separation by Exploiting Differential Alternative Splicing of a Dominant Marker in Aedes aegypti},
author = {Weng SC and Antoshechkin I and Marois E and Akbari O},
url = {https://doi.org/10.1371/journal.pgen.1011065},
doi = {10.1371/journal.pgen.1011065},
year = {2023},
date = {2023-11-27},
urldate = {2023-11-27},
journal = {PLoS Genetics},
abstract = {Only female mosquitoes consume blood giving them the opportunity to transmit deadly human pathogens. Therefore, it is critical to remove females before conducting releases for genetic biocontrol interventions. Here we describe a robust sex-sorting approach termed SEPARATOR (Sexing Element Produced by Alternative RNA-splicing of A Transgenic Observable Reporter) that exploits sex-specific alternative splicing of an innocuous reporter to ensure exclusive dominant male-specific expression. Using SEPARATOR, we demonstrate reliable sex selection from early larval and pupal stages in Aedes aegypti, and use a Complex Object Parametric Analyzer and Sorter (COPAS) to demonstrate scalable high-throughput sex-selection of first instar larvae. Additionally, we use this approach to sequence the transcriptomes of early larval males and females and find several genes that are sex-specifically expressed. SEPARATOR can simplify mass production of males for release programs and is designed to be cross-species portable and should be instrumental for genetic biocontrol interventions.},
keywords = {Aedes aegypti, Introns, larvae, M3i, marois, mosquitoes, pupae, rna sequencing, transcriptome analyses},
pubstate = {published},
tppubtype = {article}
}
Only female mosquitoes consume blood giving them the opportunity to transmit deadly human pathogens. Therefore, it is critical to remove females before conducting releases for genetic biocontrol interventions. Here we describe a robust sex-sorting approach termed SEPARATOR (Sexing Element Produced by Alternative RNA-splicing of A Transgenic Observable Reporter) that exploits sex-specific alternative splicing of an innocuous reporter to ensure exclusive dominant male-specific expression. Using SEPARATOR, we demonstrate reliable sex selection from early larval and pupal stages in Aedes aegypti, and use a Complex Object Parametric Analyzer and Sorter (COPAS) to demonstrate scalable high-throughput sex-selection of first instar larvae. Additionally, we use this approach to sequence the transcriptomes of early larval males and females and find several genes that are sex-specifically expressed. SEPARATOR can simplify mass production of males for release programs and is designed to be cross-species portable and should be instrumental for genetic biocontrol interventions.
PELLETIER Julien, DAWIT Mengistu, GHANINIA Majid, MAROIS Eric, IGNELL Rickard
A mosquito-specific antennal protein is critical for the attraction to human odor in the malaria vector Anopheles gambiae Journal Article
In: Insect Biochemistry and Molecular Biology, vol. 159, iss. August 2023, 2023.
Abstract | Links | BibTeX | Tags: antenna, chemoreceptor, M3i, marois, mosquitoes, olfaction
@article{IGNELL2023,
title = {A mosquito-specific antennal protein is critical for the attraction to human odor in the malaria vector Anopheles gambiae},
author = {Julien PELLETIER AND Mengistu DAWIT AND Majid GHANINIA AND Eric MAROIS AND Rickard IGNELL},
editor = { },
url = {https://doi.org/10.1016/j.ibmb.2023.103988},
doi = {j.ibmb.2023.103988},
year = {2023},
date = {2023-07-11},
urldate = {2023-07-11},
journal = {Insect Biochemistry and Molecular Biology},
volume = {159},
issue = {August 2023},
abstract = {Mosquitoes rely mainly on the sense of smell to decipher their environment and locate suitable food sources, hosts for blood feeding and oviposition sites. The molecular bases of olfaction involve multigenic families of olfactory proteins that have evolved to interact with a narrow set of odorants that are critical for survival. Understanding the complex interplay between diversified repertoires of olfactory proteins and ecologically-relevant odorant signals, which elicit important behaviors, is fundamental for the design of novel control strategies targeting the sense of smell of disease vector mosquitoes. Previously, large multigene families of odorant receptor and ionotropic receptor proteins, as well as a subset of odorant-binding proteins have been shown to mediate the selectivity and sensitivity of the mosquito olfactory system. In this study, we identify a mosquito-specific antennal protein (MSAP) gene as a novel molecular actor of odorant reception. MSAP is highly conserved across mosquito species and is transcribed at an extremely high level in female antennae. In order to understand its role in the mosquito olfactory system, we generated knockout mutant lines in Anopheles gambiae, and performed comparative analysis of behavioral and physiological responses to human-associated odorants. We found that MSAP promotes female mosquito attraction to human odor and enhances the sensitivity of the antennae to a variety of odorants. These findings suggest that MSAP is an important component of the mosquito olfactory system, which until now has gone completely unnoticed.},
keywords = {antenna, chemoreceptor, M3i, marois, mosquitoes, olfaction},
pubstate = {published},
tppubtype = {article}
}
Mosquitoes rely mainly on the sense of smell to decipher their environment and locate suitable food sources, hosts for blood feeding and oviposition sites. The molecular bases of olfaction involve multigenic families of olfactory proteins that have evolved to interact with a narrow set of odorants that are critical for survival. Understanding the complex interplay between diversified repertoires of olfactory proteins and ecologically-relevant odorant signals, which elicit important behaviors, is fundamental for the design of novel control strategies targeting the sense of smell of disease vector mosquitoes. Previously, large multigene families of odorant receptor and ionotropic receptor proteins, as well as a subset of odorant-binding proteins have been shown to mediate the selectivity and sensitivity of the mosquito olfactory system. In this study, we identify a mosquito-specific antennal protein (MSAP) gene as a novel molecular actor of odorant reception. MSAP is highly conserved across mosquito species and is transcribed at an extremely high level in female antennae. In order to understand its role in the mosquito olfactory system, we generated knockout mutant lines in Anopheles gambiae, and performed comparative analysis of behavioral and physiological responses to human-associated odorants. We found that MSAP promotes female mosquito attraction to human odor and enhances the sensitivity of the antennae to a variety of odorants. These findings suggest that MSAP is an important component of the mosquito olfactory system, which until now has gone completely unnoticed.
C Lutrat, M Burckbuchler, RP Olmo, R Beugnon, A Fontaine, OS Akbari, R Argiles-Herrero, T Baldet, J Bouyer, E Marois
Combining two Genetic Sexing Strains allows sorting of non-transgenic males for Aedes genetic control Journal Article
In: Commun Biol., vol. 6, iss. 1, pp. 646, 2023.
Abstract | Links | BibTeX | Tags: Aedes, M3i, marois, mosquitoes, Olmo, vectoring
@article{Lutrat2023,
title = {Combining two Genetic Sexing Strains allows sorting of non-transgenic males for Aedes genetic control},
author = {Lutrat C and Burckbuchler M and Olmo RP and Beugnon R and Fontaine A and Akbari OS and Argiles-Herrero R and Baldet T and Bouyer J and Marois E},
url = {https://www.nature.com/articles/s42003-023-05030-7},
doi = {10.1038/s42003-023-05030-7},
year = {2023},
date = {2023-06-16},
urldate = {2023-06-16},
journal = {Commun Biol.},
volume = {6},
issue = {1},
pages = {646},
abstract = {Chemical control of disease vectoring mosquitoes Aedes albopictus and Aedes aegypti is costly, unsustainable, and increasingly ineffective due to the spread of insecticide resistance. The Sterile Insect Technique is a valuable alternative but is limited by slow, error-prone, and wasteful sex-separation methods. Here, we present four Genetic Sexing Strains (two for each Aedes species) based on fluorescence markers linked to the m and M sex loci, allowing for the isolation of transgenic males. Furthermore, we demonstrate how combining these sexing strains enables the production of non-transgenic males. In a mass-rearing facility, 100,000 first instar male larvae could be sorted in under 1.5 h with an estimated 0.01–0.1% female contamination on a single machine. Cost-efficiency analyses revealed that using these strains could result in important savings while setting up and running a mass-rearing facility. Altogether, these Genetic Sexing Strains should enable a major upscaling in control programmes against these important vectors.},
keywords = {Aedes, M3i, marois, mosquitoes, Olmo, vectoring},
pubstate = {published},
tppubtype = {article}
}
Chemical control of disease vectoring mosquitoes Aedes albopictus and Aedes aegypti is costly, unsustainable, and increasingly ineffective due to the spread of insecticide resistance. The Sterile Insect Technique is a valuable alternative but is limited by slow, error-prone, and wasteful sex-separation methods. Here, we present four Genetic Sexing Strains (two for each Aedes species) based on fluorescence markers linked to the m and M sex loci, allowing for the isolation of transgenic males. Furthermore, we demonstrate how combining these sexing strains enables the production of non-transgenic males. In a mass-rearing facility, 100,000 first instar male larvae could be sorted in under 1.5 h with an estimated 0.01–0.1% female contamination on a single machine. Cost-efficiency analyses revealed that using these strains could result in important savings while setting up and running a mass-rearing facility. Altogether, these Genetic Sexing Strains should enable a major upscaling in control programmes against these important vectors.
Klug Dennis, Gautier Amandine, Calvo Eric, Marois Eric, Blandin Stéphanie A.
The salivary protein Saglin facilitates efficient midgut colonization of Anopheles mosquitoes by malaria parasites Journal Article
In: Plos Pathogens, vol. 19, iss. 3, no. 3, 2023.
Abstract | Links | BibTeX | Tags: blandin, BLOOD, M3i, malarial parasites, marois, mosquitoes, Oocysts, Parasitic Diseases, Plasmodium, salivary glands, sporozoites
@article{Klug2023,
title = {The salivary protein Saglin facilitates efficient midgut colonization of Anopheles mosquitoes by malaria parasites},
author = {Dennis Klug and Amandine Gautier and Eric Calvo and Eric Marois and Stéphanie A. Blandin},
url = {https://doi.org/10.1371/journal.ppat.1010538},
doi = {10.1371/journal.ppat.1010538},
year = {2023},
date = {2023-03-02},
urldate = {2023-03-02},
booktitle = {Plos Pathogens},
journal = {Plos Pathogens},
volume = {19},
number = {3},
issue = {3},
abstract = {Female mosquitoes rely on blood feeding to acquire sufficient nutrients for egg development. Because of the importance of this process mosquitoes evolved salivary proteins with a broad range of functions acting as blood thinners, anti-coagulants and immunosuppressants. The effect of these proteins on the blood at the bite site directly influences the size of the blood bolus a female takes up in a given time frame. Both, time of feeding and bolus size, are important parameters for fecundity and survival. Recent studies have shown that a significant proportion of salivated proteins is re-ingested during feeding and becomes part of the blood meal. Here we investigated the salivary protein Saglin which has been previously suggested as putative receptor mediating malaria parasite entry into the salivary gland. By engineering a loss-of-function mutant in An. coluzzi we could show that the absence of Saglin impairs the development of parasite stages in the blood meal of the rodent malaria parasite P. berghei and the human malaria parasite P. falciparum lowering the parasite burden of subsequent stages and preventing efficient transmission at low infection densities. Furthermore, we could show that Saglin is present in the blood meal after feeding possibly indicating a previously overlooked parasite-vector interaction.},
keywords = {blandin, BLOOD, M3i, malarial parasites, marois, mosquitoes, Oocysts, Parasitic Diseases, Plasmodium, salivary glands, sporozoites},
pubstate = {published},
tppubtype = {article}
}
Female mosquitoes rely on blood feeding to acquire sufficient nutrients for egg development. Because of the importance of this process mosquitoes evolved salivary proteins with a broad range of functions acting as blood thinners, anti-coagulants and immunosuppressants. The effect of these proteins on the blood at the bite site directly influences the size of the blood bolus a female takes up in a given time frame. Both, time of feeding and bolus size, are important parameters for fecundity and survival. Recent studies have shown that a significant proportion of salivated proteins is re-ingested during feeding and becomes part of the blood meal. Here we investigated the salivary protein Saglin which has been previously suggested as putative receptor mediating malaria parasite entry into the salivary gland. By engineering a loss-of-function mutant in An. coluzzi we could show that the absence of Saglin impairs the development of parasite stages in the blood meal of the rodent malaria parasite P. berghei and the human malaria parasite P. falciparum lowering the parasite burden of subsequent stages and preventing efficient transmission at low infection densities. Furthermore, we could show that Saglin is present in the blood meal after feeding possibly indicating a previously overlooked parasite-vector interaction.