scholarly journals Efficient population modification gene-drive rescue system in the malaria mosquito Anopheles stephensi

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Adriana Adolfi ◽  
Valentino M. Gantz ◽  
Nijole Jasinskiene ◽  
Hsu-Feng Lee ◽  
Kristy Hwang ◽  
...  
Keyword(s):  
Anopheles Stephensi ◽  
Pathogen Transmission ◽  
Gene Drive ◽  
Initial Release ◽  
Genetic Technologies ◽  
Rescue System ◽  
Vector Borne ◽  
Kynurenine Hydroxylase ◽  
Drive Systems ◽  
Advanced Development

Abstract Cas9/gRNA-mediated gene-drive systems have advanced development of genetic technologies for controlling vector-borne pathogen transmission. These technologies include population suppression approaches, genetic analogs of insecticidal techniques that reduce the number of insect vectors, and population modification (replacement/alteration) approaches, which interfere with competence to transmit pathogens. Here, we develop a recoded gene-drive rescue system for population modification of the malaria vector, Anopheles stephensi, that relieves the load in females caused by integration of the drive into the kynurenine hydroxylase gene by rescuing its function. Non-functional resistant alleles are eliminated via a dominantly-acting maternal effect combined with slower-acting standard negative selection, and rare functional resistant alleles do not prevent drive invasion. Small cage trials show that single releases of gene-drive males robustly result in efficient population modification with ≥95% of mosquitoes carrying the drive within 5-11 generations over a range of initial release ratios.

scholarly journals Efficient population modification gene-drive rescue system in the malaria mosquito Anopheles stephensi

2020 ◽  
Author(s):  
Adriana Adolfi ◽  
Valentino M. Gantz ◽  
Nijole Jasinskiene ◽  
Hsu-Feng Lee ◽  
Kristy Hwang ◽  
...  
Keyword(s):  
Anopheles Stephensi ◽  
Pathogen Transmission ◽  
Gene Drive ◽  
Initial Release ◽  
Genetic Technologies ◽  
Rescue System ◽  
Vector Borne ◽  
Kynurenine Hydroxylase ◽  
Drive Systems ◽  
Population Suppression

ABSTRACTThe development of Cas9/gRNA-mediated gene-drive systems has bolstered the advancement of genetic technologies for controlling vector-borne pathogen transmission. These include population suppression approaches, genetic analogs of insecticidal techniques that reduce the number of vector insects, and population modification (replacement/alteration) approaches, which interfere with competence to transmit pathogens. We developed a recoded gene-drive rescue system for population modification in the malaria vector, Anopheles stephensi, that relieves the load in females caused by integration of the drive into the kynurenine hydroxylase gene by rescuing its function. Non-functional resistant alleles are eliminated via a dominantly-acting maternal effect combined with slower-acting standard negative selection, and a functional resistant allele does not prevent drive invasion. Small cage trials show that single releases of gene-drive males robustly result in efficient population modification with ≥95% of mosquitoes carrying the drive within 5-11 generations over a range of initial release ratios.

scholarly journals Population dynamics of underdominance gene drive systems in continuous space

2018 ◽  
Author(s):  
Jackson Champer ◽  
Joanna Zhao ◽  
Joanna Zhao ◽  
Samuel E. Champer ◽  
Jingxian Liu ◽  
...  
Keyword(s):  
Decision Making ◽  
Population Dynamics ◽  
Local Population ◽  
Spatial Context ◽  
Decision Making Process ◽  
Gene Drive ◽  
Continuous Space ◽  
Vector Borne Diseases ◽  
Vector Borne ◽  
Drive Systems

ABSTRACTUnderdominance gene drive systems promise a mechanism for rapidly spreading payload alleles through a local population while otherwise remaining confined, unable to spread into neighboring populations due to their frequency-dependent dynamics. Such systems could provide a new tool in the fight against vector-borne diseases by disseminating transgenic payloads through vector populations. If local confinement can indeed be achieved, the decision-making process for the release of such constructs would likely be considerably simpler compared to other gene drive mechanisms such as CRISPR homing drives. So far, the confinement ability of underdominance systems has only been demonstrated in models of panmictic populations linked by migration. How such systems would behave in realistic populations where individuals move over continuous space remains largely unknown. Here, we study several underdominance systems in continuous-space population models and show that their dynamics are drastically altered from those in panmictic populations. Specifically, we find that all underdominance systems we studied can fail to persist in such environments, even after successful local establishment. At the same time, we find that a two-locus two-toxin-antitoxin system can still successfully invade neighboring populations in many scenarios even under weak migration. This suggests that the parameter space for underdominance systems to both establish in a given region and remain confined to that region would likely be highly limited. Overall, these results indicate that spatial context must be considered when assessing strategies for the deployment of underdominance systems.

scholarly journals Digital droplet PCR and IDAA for the detection of CRISPR indel edits in the malaria species Anopheles stephensi

BioTechniques ◽  
2020 ◽  
Vol 68 (4) ◽  
pp. 172-179 ◽  
Author(s):  
Rebeca Carballar-Lejarazú ◽  
Adam Kelsey ◽  
Thai Binh Pham ◽  
Eric P Bennett ◽  
Anthony A James
Keyword(s):  
Anopheles Stephensi ◽  
Field Testing ◽  
Nonhomologous End Joining ◽  
Mosquito Vector ◽  
Gene Drive ◽  
End Joining ◽  
Indel Detection ◽  
Vector Mosquito ◽  
Droplet Pcr ◽  
Drive Systems

CRISPR/Cas9 technology is a powerful tool for the design of gene-drive systems to control and/or modify mosquito vector populations; however, CRISPR/Cas9-mediated nonhomologous end joining mutations can have an important impact on generating alleles resistant to the drive and thus on drive efficiency. We demonstrate and compare the insertions or deletions (indels) detection capabilities of two techniques in the malaria vector mosquito Anopheles stephensi: Indel Detection by Amplicon Analysis (IDAA™) and Droplet Digital™ PCR (ddPCR™). Both techniques showed accuracy and reproducibility for indel frequencies across mosquito samples containing different ratios of indels of various sizes. Moreover, these techniques have advantages that make them potentially better suited for high-throughput nonhomologous end joining analysis in cage trials and contained field testing of gene-drive mosquitoes.

scholarly journals Design and analysis of CRISPR-based underdominance toxin-antidote gene drives

2019 ◽  
Author(s):  
Jackson Champer ◽  
Samuel E. Champer ◽  
Isabel Kim ◽  
Andrew G. Clark ◽  
Philipp W. Messer
Keyword(s):  
Critical Frequency ◽  
Essential Gene ◽  
Geographic Location ◽  
Early Embryo ◽  
Gene Drive ◽  
Wide Range ◽  
Vector Borne ◽  
Drive Systems ◽  
Invasion Threshold ◽  
Gene Drives

ABSTRACTCRISPR gene drive systems offer a mechanism for transmitting a desirable transgene throughout a population for purposes ranging from vector-borne disease control to invasive species suppression. In this simulation study, we assess the performance of several CRISPR-based underdominance gene drive constructs employing toxin-antidote principles. These drives disrupt the wild-type version of an essential gene using a CRISPR nuclease (the toxin) while simultaneously carrying a recoded version of the gene (the antidote). Drives of this nature allow for releases that could be potentially confined to a desired geographic location. This is because such drives have a nonzero invasion threshold frequency, referring to the critical frequency required for the drive to spread through the population. We model drives which target essential genes that are either haplosufficient or haplolethal, using nuclease promoters with expression restricted to the germline, promoters that additionally result in cleavage activity in the early embryo from maternal deposition, and promoters that have ubiquitous somatic expression. We also study several possible drive architectures, considering both “same-site” and “distant-site” systems, as well as several reciprocally targeting drives. Together, these drive variants provide a wide range of invasion threshold frequencies and options for both population modification and suppression. Our results suggest that CRISPR toxin-antidote underdominance drive systems could allow for the design of highly flexible and potentially confinable gene drive strategies.

scholarly journals MGDrivE 2: A simulation framework for gene drive systems incorporating seasonality and epidemiological dynamics

2020 ◽  
Author(s):  
Sean L. Wu ◽  
Jared B. Bennett ◽  
Héctor M. Sánchez C. ◽  
Andrew J. Dolgert ◽  
Tomás M. León ◽  
...  
Keyword(s):  
Population Dynamics ◽  
Disease Transmission ◽  
Field Trials ◽  
Pathogen Transmission ◽  
Environmental Data ◽  
List Type ◽  
Gene Drive ◽  
Simulation Framework ◽  
Implementation Framework ◽  
Drive Systems

AbstractInterest in gene drive technology has continued to grow as promising new drive systems have been developed in the lab and discussions are moving towards implementing field trials. The prospect of field trials requires models that incorporate a significant degree of ecological detail, including parameters that change over time in response to environmental data such as temperature and rainfall, leading to seasonal patterns in mosquito population density. Epidemiological outcomes are also of growing importance, as: i) the suitability of a gene drive construct for release will depend on its expected impact on disease transmission, and ii) initial field trials are expected to have a measured entomological outcome and a modeled epidemiological outcome.We present MGDrivE 2 (Mosquito Gene Drive Explorer 2): an extension of and development from the MGDrivE 1 simulation framework that investigates the population dynamics of a variety of gene drive architectures and their spread through spatially-explicit mosquito populations. Key strengths and improvements of the MGDrivE 2 framework are: i) the ability of parameters to vary with time and induce seasonal population dynamics, ii) an epidemiological module accommodating reciprocal pathogen transmission between humans and mosquitoes, and iii) an implementation framework based on stochastic Petri nets that enables efficient model formulation and flexible implementation.Example MGDrivE 2 simulations are presented to demonstrate the application of the framework to a CRISPR-based homing gene drive system intended to drive a disease-refractory gene into a population, incorporating time-varying temperature and rainfall data, and predict impact on human disease incidence and prevalence. Further documentation and use examples are provided in vignettes at the project’s CRAN repository.MGDrivE 2 is an open-source R package freely available on CRAN. We intend the package to provide a flexible tool capable of modeling gene drive constructs as they move closer to field application and to infer their expected impact on disease transmission.

scholarly journals Host Associations Of Biting Midges (Diptera: Ceratopogonidae: Culicoides) Near Sentinel Chicken Surveillance Locations In Florida, USA

2019 ◽  
Vol 35 (3) ◽  
pp. 200-206 ◽  
Author(s):  
Kristin E. Sloyer ◽  
Carolina Acevedo ◽  
Alfred E. Runkel ◽  
Nathan D. Burkett-Cadena
Keyword(s):  
Mosquito Control ◽  
Pathogen Transmission ◽  
Host Use ◽  
Culicoides Species ◽  
Biting Midges ◽  
Total Blood ◽  
Avian Blood ◽  
Vector Borne ◽  
Arbovirus Surveillance ◽  
Blood Meals

ABSTRACT Quantifying host use is important for understanding transmission of vector-borne pathogens. Despite the importance of biting midges (Diptera: Ceratopogonidae) in pathogen transmission, the vector–host relationships of most Culicoides species are poorly documented, even in locations where active arbovirus surveillance is conducted. Polymerase chain reaction–based blood-meal analysis was performed on 663 blood-engorged Culicoides females collected by 7 Florida mosquito control districts at 24 sentinel chicken arbovirus surveillance sites in 2017. A total of 638 blood meals were successfully analyzed to determine host species source, representing 11 Culicoides species. The most commonly bitten host was domestic chicken (Gallus gallus) (presumably sentinel chickens), constituting 565 of 638 (88.6%) the total blood meals. Other common hosts included humans (5.8%), white-tailed deer (Odoocoileus virginianus) (2.5%), and brown basilisk (Basiliscus vittatus) (1.6%). Significant differences in distribution of mammal and avian blood meals were found for a number of Culicoides species, and these patterns did not vary across locations. These results indicate that sentinel chickens are exposed to bites by Culicoides, potentially exposing them to Culicoides-borne pathogens. The findings that vertebrate host use was consistent across locations suggests that each Culicoides species has affinity for one or more specific animal groups, and does not feed randomly upon available animals.

scholarly journals Optimized CRISPR tools and site-directed transgenesis in Culex quinquefasciatus mosquitoes for gene drive development

2021 ◽  
Author(s):  
Xuechun Feng ◽  
Víctor López Del Amo ◽  
Enzo Mameli ◽  
Megan Lee ◽  
Alena L. Bishop ◽  
...  
Keyword(s):  
Culex Quinquefasciatus ◽  
Technology Development ◽  
Companion Animals ◽  
Fruit Fly ◽  
Valuable Insight ◽  
Gene Drive ◽  
Future Technology ◽  
Drive Systems ◽  
Global Vector ◽  
Human And Animal Diseases

ABSTRACTCulex mosquitoes are a global vector for multiple human and animal diseases, including West Nile virus, lymphatic filariasis, and avian malaria, posing a constant threat to public health, livestock, companion animals, and endangered birds. While rising insecticide resistance has threatened the control of Culex mosquitoes, advances in CRISPR genome-editing tools have fostered the development of alternative genetic strategies such as gene drive systems to fight disease vectors. However, though gene-drive technology has quickly progressed in other mosquitoes, advances have been lacking in Culex. Here, we developed a Culex-specific Cas9/gRNA expression toolkit and used site-directed homology-based transgenesis to generate and validate a Culex quinquefasciatus Cas9-expressing line. We showed that gRNA scaffold variants improve transgenesis efficiency in both Culex and Drosophila and boost gene-drive performance in the fruit fly. These findings support future technology development to control Culex mosquitoes and provide valuable insight for improving these tools in other species.

scholarly journals A CRISPR homing gene drive targeting a haplolethal gene removes resistance alleles and successfully spreads through a cage population

2020 ◽  
Vol 117 (39) ◽  
pp. 24377-24383 ◽  
Author(s):  
Jackson Champer ◽  
Emily Yang ◽  
Esther Lee ◽  
Jingxian Liu ◽  
Andrew G. Clark ◽  
...  
Keyword(s):  
Gene Drive ◽  
End Joining ◽  
Large Cage ◽  
Vector Borne Diseases ◽  
Guide Rnas ◽  
Genetic Modifications ◽  
New Strategy ◽  
Cage Population ◽  
Vector Borne ◽  
Gene Drives

Engineered gene drives are being explored as a new strategy in the fight against vector-borne diseases due to their potential for rapidly spreading genetic modifications through a population. However, CRISPR-based homing gene drives proposed for this purpose have faced a major obstacle in the formation of resistance alleles that prevent Cas9 cleavage. Here, we present a homing drive in Drosophila melanogaster that reduces the prevalence of resistance alleles below detectable levels by targeting a haplolethal gene with two guide RNAs (gRNAs) while also providing a rescue allele. Resistance alleles that form by end-joining repair typically disrupt the haplolethal target gene and are thus removed from the population because individuals that carry them are nonviable. We demonstrate that our drive is highly efficient, with 91% of the progeny of drive heterozygotes inheriting the drive allele and with no functional resistance alleles observed in the remainder. In a large cage experiment, the drive allele successfully spread to all individuals within a few generations. These results show that a haplolethal homing drive can provide an effective tool for targeted genetic modification of entire populations.

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