Biology subjects

Bennett, J. B.

Publications and source records attributed to Bennett, J. B..

14 recordsLinked to original sources

bioRxiv2026

Split suppression gene drives reveal a design tradeoff between inheritance and reproductive load in Aedes aegypti

Mosquito-borne pathogens cause approximately 700,000 deaths annually, highlighting the need for effective and sustainable vector control strategies. CRISPR-based gene drives offer a potential approach for suppressing disease-vector populations, but autonomous drives may spread beyond intended populations. Split gene drives, in which the Cas9 and guide RNA components are genetically separated, provide an inherently confinable alternative because biased inheritance depends on the co-occurrence of both components. Here, we develop the first split suppression gene drives in Ae. aegypti, targeting the female reproductive genes doublesex (dsx) and yellow-g (yg). The yg-targeting drive exhibited moderate super-Mendelian inheritance, reaching 73.4% transmission in the highest-performing cross, whereas the dsx-targeting drive exhibited little detectable inheritance bias. Conversely, disruption of dsx imposed substantially stronger reproductive fitness costs, including complete sterility in homozygous females, whereas yg disruption produced a less severe fertility phenotype. Mathematical modeling parameterized with these experimental data predicted that three of four split-drive configurations could achieve population suppression at lower release intensities or shorter release durations than pgSIT across portions of the modeled parameter space. Together, these results reveal that inheritance efficiency and reproductive load can impose competing constraints on split suppression drive performance: stronger suppression-associated fitness effects do not necessarily translate into improved drive performance when accompanied by limited drive propagation. Our findings establish the feasibility of split suppression gene drives in Ae. aegypti and demonstrate that effective suppression drive design requires balancing inheritance efficiency with reproductive fitness costs rather than maximizing either independently.

bioengineering↗

bioRxiv2025

Spatial close-kin mark-recapture methods to estimate dispersal parameters and barrier strength for mosquitoes

Close-kin mark-recapture (CKMR) methods have recently been used to infer demographic parameters for several aquatic and terrestrial species. For mosquitoes, the spatial distribution of close-kin pairs has been used to estimate mean dispersal distance, of relevance to vector-borne disease transmission and genetic biocontrol strategies. Close-kin methods have advantages over traditional mark-release-recapture (MRR) methods as the mark is genetic, removing the need for physical marking and recapturing that may interfere with movement behavior. Here, we extend CKMR methods to accommodate spatial structure alongside life history for mosquitoes and comparable insects. We derive kinship probabilities for parent-offspring and full-sibling pairs in a spatial context, where an individual in each pair may be a larva or adult. Using the dengue vector Aedes aegypti as a case study, we use an individual-based model of mosquito life history to test the effectiveness of this approach at estimating parameters such as mean dispersal distance, daily staying probability, and the strength of a barrier to movement. Considering a simulated population of 9,025 adult mosquitoes arranged on a 19-by-19 grid, we find the CKMR approach provides unbiased and precise estimates of mean dispersal distance given a total of 2,500 adult females sampled over a three-month period using 25 traps evenly spread throughout the landscape. The CKMR approach is also able to estimate parameters of more complex dispersal kernels, such as the daily staying probability of a zero-inflated exponential kernel, or the strength of a barrier to movement, provided the magnitude of these parameters is greater than 0.5. These results suggest that CKMR provides an insightful characterization of mosquito dispersal that is complementary to conventional MRR methods. Author summaryClose-kin mark-recapture (CKMR) is a genetic analogue of mark-release-recapture (MRR) in which the frequency of marked individuals in a sample is used to infer demographic parameters such as census population size and mean dispersal distance. These methods have been widely applied to aquatic species; however their application to mosquitoes is yet to be rigorously explored. Previous theoretical work demonstrated the potential for CKMR to infer parameters such as population size and mortality rate for randomly-mixing mosquito populations, and close-kin-based methods have been used to infer movement patterns for Aedes aegypti mosquitoes in Singapore and Malaysia. Here, we use simulations to explore the potential for formal CKMR methods to characterize mosquito dispersal patterns. We find that formal CKMR methods are able to accurately estimate mean dispersal distance, and to estimate additional parameters, such as the strength of a landscape barrier and the probability that a mosquito remains within its population node each day. CKMR and other close-kin-based methods provide insights into mosquito dispersal complementary to commonly-used alternatives such as MRR, as they capture displacement across several generations and are not compromised by the marking process.

ecology↗

bioRxiv2022

Close-kin mark-recapture methods to estimate demographic parameters of mosquitoes

Close-kin mark-recapture (CKMR) methods have recently been used to infer demographic parameters such as census population size and survival for fish of interest to fisheries and conservation. These methods have advantages over traditional mark-recapture methods as the mark is genetic, removing the need for physical marking and recapturing that may interfere with parameter estimation. For mosquitoes, the spatial distribution of close-kin pairs has been used to estimate mean dispersal distance, of relevance to vector-borne disease transmission and novel biocontrol strategies. Here, we extend CKMR methods to the life history of mosquitoes and comparable insects. We derive kinship probabilities for mother-offspring, father-offspring, full-sibling and half-sibling pairs, where an individual in each pair may be a larva, pupa or adult. A pseudo-likelihood approach is used to combine the marginal probabilities of all kinship pairs. To test the effectiveness of this approach at estimating mosquito demographic parameters, we develop an individual-based model of mosquito life history incorporating egg, larva, pupa and adult life stages. The simulation labels each individual with a unique identification number, enabling close-kin relationships to be inferred for sampled individuals. Using the dengue vector Aedes aegypti as a case study, we find the CKMR approach provides unbiased estimates of adult census population size, adult and larval mortality rates, and larval life stage duration for logistically feasible sampling schemes. Considering a simulated population of 3,000 adult mosquitoes, estimation of adult parameters is accurate when ca. 40 adult females are sampled biweekly over a three month period. Estimation of larval parameters is accurate when adult sampling is supplemented with ca. 120 larvae sampled biweekly over the same period. The methods are also effective at detecting intervention-induced increases in adult mortality and decreases in population size. As the cost of genome sequencing declines, CKMR holds great promise for characterizing the demography of mosquitoes and comparable insects of epidemiological and agricultural significance. Author summaryClose-kin mark-recapture (CKMR) methods are a genetic analogue of traditional mark-recapture methods in which the frequency of marked individuals in a sample is used to infer demographic parameters such as census population size and mean dispersal distance. In CKMR, the mark is a close-kin relationship between individuals (parents and offspring, siblings, etc.). While CKMR methods have mostly been applied to aquatic species to date, opportunities exist to apply them to insects and other terrestrial species. Here, we explore the application of CKMR to mosquitoes, with Aedes aegypti, a primary vector of dengue, chikungunya and yellow fever, as a case study. By analyzing simulated Ae. aegypti populations, we find the CKMR approach provides unbiased estimates of adult census population size, adult and larval mortality rates, and larval life stage duration, and may be informative of intervention impact. Optimal sampling schemes are compatible with Ae. aegypti ecology and field studies. This study represents the first theoretical exploration of the application of CKMR to an insect species, and demonstrates its potential for characterizing the demography of insects of epidemiological and agricultural importance.

ecology↗

bioRxiv2021

Genetic conversion of a split-drive into a full-drive element

Gene-drive systems offer an important new avenue for spreading beneficial traits into wild populations. Their core components, Cas9 and guide RNA (gRNA), can either be linked within a single cassette (full gene drive, fGD) or provided in two separate elements (split gene drive, sGD) wherein the gRNA-bearing element drives in the presence of an independent static source of Cas9. We previously designed a system engineered to turn split into full gene drives. Here, we provide experimental proof-of-principle for such a convertible system inserted at the spo11 locus, which is recoded to restore gene function. In multigenerational cage studies, the reconstituted spo11 fGD cassette initially drives with slower kinetics than the unlinked sGD element (using the same Mendelian vasa-Cas9 source), but eventually reaches a similar level of final introgression. Different kinetic behaviors may result from transient fitness costs associated with individuals co-inheriting Cas9 and gRNA transgenes during the drive process.

genetics↗

bioRxiv2021

Eliminating Aedes aegypti from its southern margin in Australia: insights from genomic data and simulation modeling

A rare example of a successful long-term elimination of the mosquito Aedes aegypti is in Brisbane, Queensland, where the legislatively-enforced removal of rainwater tanks drove its disappearance by the mid-1950s. However, a decade-long drought led to the mass installation of rainwater tanks throughout the region, re-introducing critical breeding sites for the mosquitos persistence in this subtropical region. With Ae. aegypti re-invading towns just 150 km north of Brisbane, we examined the potential for their sustained elimination. Through genomic analyses, we estimated historical expansion and current isolation between neighboring populations as close as 15 kilometers. The estimated recent migration rate, entomological and meteorological data were used to calibrate the simulations of elimination campaigns in the two southernmost populations. Our simulations indicate that Ae. aegypti could be eliminated with moderate release numbers of incompatible Wolbachia-infected (IIT) males (sorted with an error rate [≤]10-6) if non-compliant rainwater tanks are removed first. With this combined campaign, highly effective suppression (>99%) was predicted in both towns, and complete elimination was predicted in 35% of simulations in one town. Without tank removal, however, IIT led to a moderate suppression (61-93%) even with a 40:1 ratio of released IIT males to local males. Moreover, with a ratio of >20:1, Wolbachia establishment was predicted when the sorting error was >10-7. Our conservative estimates of intervention outcomes inform the planning of Ae. aegypti elimination in the region, and offer insight into the effective combinations of conventional and novel control tools, particularly for vulnerable mosquito populations at range margins. SignificanceAfter decades of range stagnation in Australia, the Aedes aegypti mosquito is expanding southward, approaching the most-densely-populated areas of Queensland. Using population genomics and simulation modeling of elimination campaigns, we show that Australias southernmost populations of this disease vector are genetically isolated and could be eliminated with moderate releases of incompatible Wolbachia-infected males if major larval breeding sites (non-compliant rainwater tanks) are removed first. The risk of Wolbachia establishment for this approach is low, and so is the risk of quick mosquito re-invasion. Our conservative estimates of intervention outcomes inform the planning of Ae. aegypti elimination in the region, and offer new insight into the benefits of combining conventional and novel control tools, particularly for mosquito populations at range margins.

systems biology↗

bioRxiv2021

Eliminating Mosquitoes with Precision Guided Sterile Males

The mosquito Aedes aegypti is the principal vector for arboviruses including dengue/yellow fever, chikungunya, and zika, infecting hundreds of millions of people annually. Unfortunately, traditional control methodologies are insufficient, so innovative control methods are needed. To complement existing measures, here we develop a molecular genetic control system termed precision guided sterile insect technique (pgSIT) in Aedes aegypti. PgSIT uses a simple CRISPR-based approach to generate sterile males that are deployable at any life stage. Supported by mathematical models, we empirically demonstrate that released pgSIT males can compete, suppress, and eliminate mosquitoes in multigenerational population cages. This platform technology could be used in the field, and adapted to many vectors, for controlling wild populations to curtail disease in a safe, confinable, and reversible manner.

genetics↗

bioRxiv2020

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

O_LIInterest 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. C_LIO_LIWe 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. C_LIO_LIExample 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 projects CRAN repository. C_LIO_LIMGDrivE 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. C_LI

bioinformatics↗

bioRxiv2020

Inherently confinable split-drive systems in Drosophila

CRISPR-based gene drive systems, which copy themselves based on gene conversion mediated by the homology directed repair (HDR) pathway, have potential to revolutionize vector control. However, mutant alleles generated by the competing non-homologous end-joining (NHEJ) pathway that are rendered resistant to Cas9 cleavage can interrupt the spread of genedrive elements. We hypothesized that drives targeting genes essential for viability or reproduction also carrying recoded sequences to restore endogenous gene functionality should benefit from dominantly-acting maternal clearance of NHEJ alleles, combined with recessive Mendelian processes. Here, we test split gene-drive (sGD) systems in Drosophila melanogaster that were inserted into essential genes required for viability (rab5, rab11, prosalpha2) or fertility (spo11). In single generation crosses, sGDs copy with variable efficiencies and display sex-biased transmission. In multi-generational cage trials, sGD follow distinct drive trajectories reflecting their differential tendencies to induce target chromosome damage or lethal/sterile mosaic phenotypes, leading to inherently confineable drive outcomes.

genetics↗

bioRxiv2020

A home and rescue gene drive forces its inheritance stably persisting in populations

Homing based gene drives, engineered using CRISPR/Cas9, have been proposed to spread desirable genes into target populations. However, spread of such drives can be hindered by the accumulation of resistance alleles. To overcome this significant obstacle, we engineer an inherently confinable population modification Home-and-Rescue (HomeR) drive in Drosophila melanogaster that, by creative design, limits the accumulation of such alleles. We demonstrate that HomeR can achieve nearly [~]100% transmission enabling it to spread and persist at genotypic fixation in several multi-generational population cage experiments, underscoring its long term stability and drive potential. Finally, we conduct mathematical modeling determining HomeR can outperform contemporary gene drive architectures for population modification over wide ranges of fitness and transmission rates. Given its straightforward design, HomeR could be universally adapted to a wide range of species.

genetics↗

bioRxiv2020

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

The 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.

genetics↗

bioRxiv2019

Split-gene drive system provides flexible application for safe laboratory investigation and potential field deployment

CRISPR-based gene drives spread through populations bypassing the dictates of Mendelian genetics, offering a population-engineering tool for tackling vector-borne diseases, managing crop pests, and helping island conservation efforts; unfortunately, current technologies raise safety concerns for unintended gene propagation. Herein, we address this by splitting the two drive components, Cas9 and gRNAs, into separate alleles to form a novel trans-complementing split-gene-drive (tGD) and demonstrate its ability to promote super-Mendelian inheritance of the separate transgenes. This bi-component nature allows for individual transgene optimization and increases safety by restricting escape concerns to experimentation windows. We employ the tGD and a small- molecule-controlled version to investigate the biology of component inheritance and use our system to study the maternal effects on CRISPR inheritance, impaired homology on efficiency, and resistant allele formation. Lastly, mathematical modeling of tGD spread in a population shows potential advantages for improving current gene-drive technologies for field population modification.

genetics↗

bioRxiv2019

Confinement and reversibility of threshold-dependent gene drive systems in spatially-explicit Aedes aegypti populations

BackgroundThe discovery of CRISPR-based gene editing and its application to homing-based gene drive systems has been greeted with excitement, for its potential to control mosquito-borne diseases on a wide scale, and concern, for the invasiveness and potential irreversibility of a release. Gene drive systems that display threshold-dependent behavior could potentially be used during the trial phase of this technology, or when localized control is otherwise desired, as simple models predict them to spread into partially isolated populations in a confineable manner, and to be reversible through releases of wild-type organisms. Here, we model hypothetical releases of two recently-engineered threshold-dependent gene drive systems - reciprocal chromosomal translocations and a form of toxin-antidote-based underdominance known as UDMEL - to explore their ability to be confined and remediated. ResultsWe simulate releases of Aedes aegypti, the mosquito vector of dengue, Zika and other arboviruses, in Yorkeys Knob, a suburb of Cairns, Australia, where previous biological control interventions have been undertaken on this species. We monitor spread to the neighboring suburb of Trinity Park to assess confinement. Results suggest that translocations could be introduced on a suburban scale, and remediated through releases of non-disease-transmitting male mosquitoes with release sizes on the scale of what has been previously implemented. UDMEL requires fewer releases to introduce, but more releases to remediate, including of females capable of disease transmission. Both systems are expected to be confineable to the release site; however, spillover of translocations into neighboring populations is less likely. ConclusionsOur analysis supports the use of translocations as a threshold-dependent drive system capable of spreading disease-refractory genes into Ae. aegypti populations in a confineable and reversible manner. It also highlights increased release requirements when incorporating life history and population structure into models. As the technology nears implementation, further ecological work will be essential to enhance model predictions in preparation for field trials.

ecology↗

bioRxiv2019

Microfluidic Rapid and Autonomous Analytical Device (microRAAD) to Detect HIV from Whole Blood Samples

Early Human Immunodeficiency Virus (HIV) testing is critical to preventing transmission and providing treatment to HIV-positive individuals, yet an estimated 30% of HIV-positive individuals do not know their status because of barriers to early diagnosis. Readily accessible, highly sensitive, and rapid diagnostic tests would enable patients prompt treatment with anti-retroviral therapies and reduce transmission. However, existing HIV diagnostic technologies either do not detect early stages of infection or require multiple days of laboratory processing, delaying notification of patients status.\n\nMolecular techniques that amplify HIV RNA can detect the earliest stages of infection, within 8-10 days after transmission. However, most of these molecular assays require cold-chain storage of reagents, significant sample preparation, and extensive laboratory infrastructure. To achieve early detection, we developed a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay with a limit of detection of 10 HIV-1 RNA copies visualized by eye using a lateral flow immunoassay. To demonstrate automated sample-to-answer detection of HIV, we incorporate dried amplification reagents and wax valves in low-cost substrates with resistive heating elements and circuitry. By combining controlled heating with papers capillary flow, our assembled device automatically isolates viral particles from human blood samples, amplifies HIV-1 RNA, and transports products to a detection zone. We determine that as few as 105 HIV-1 viral particles can be separated from whole blood, amplified, and visually detected within 90 minutes of sample addition into our Microfluidic Rapid and Autonomous Analysis Device (microRAAD). The low-cost and automated attributes of microRAAD demonstrate its utility as a point-of-care testing platform.

bioengineering↗

bioRxiv2018

MGDrivE: A modular simulation framework for the spread of gene drives through spatially-explicit mosquito populations

Malaria, dengue, Zika, and other mosquito-borne diseases continue to pose a major global health burden through much of the world, despite the widespread distribution of insecticide-based tools and antimalarial drugs. The advent of CRISPR/Cas9-based gene editing and its demonstrated ability to streamline the development of gene drive systems has reignited interest in the application of this technology to the control of mosquitoes and the diseases they transmit. The versatility of this technology has also enabled a wide range of gene drive architectures to be realized, creating a need for their population-level and spatial dynamics to be explored. To this end, we present MGDrivE (Mosquito Gene Drive Explorer): a simulation framework designed to investigate the population dynamics of a variety of gene drive architectures and their spread through spatially-explicit mosquito populations. A key strength of the MGDrivE framework is its modularity: a) a genetic inheritance module accommodates the dynamics of gene drive systems displaying user-defined inheritance patterns, b) a population dynamic module accommodates the life history of a variety of mosquito disease vectors and insect agricultural pest species, and c) a landscape module accommodates the distribution of insect metapopulations connected by migration in space. Example MGDrivE simulations are presented to demonstrate the application of the framework to CRISPR/Cas9-based homing gene drive for: a) driving a disease-refractory gene into a population (i.e. population replacement), and b) disrupting a gene required for female fertility (i.e. population suppression), incorporating homing-resistant alleles in both cases. We compare MGDrivE with other genetic simulation packages, and conclude with a discussion of future directions in gene drive modeling.

genetics↗