Biology subjects

Marshall, J. M.

Publications and source records attributed to Marshall, J. M..

At least 19 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

A target product profile for a rapid diagnostic test to monitor mosquito gene drive presence and frequency

Malaria remains a major global health challenge, with over 263 million cases and nearly 600,000 deaths reported in 2023, the majority in sub-Saharan Africa. While conventional interventions such as insecticide-treated nets, indoor residual spraying and antimalarial drugs have reduced transmission, progress has stalled due to the limitations of these interventions and the emergence of resistance. Gene drive-modified mosquitoes represent a promising, potentially transformative vector control strategy, capable of spreading malaria-refractory traits or suppressing mosquito populations. Successful field deployment will depend upon monitoring systems to track the presence and frequency of gene drive constructs as they spread and persist. Current molecular surveillance techniques, though effective, are resource-intensive and reliant on laboratory infrastructure and technical competencies. Here, we make the case for a near-universal and low-cost rapid diagnostic test (RDT) designed to detect gene drive mosquitoes in the field, to complement existing surveillance infrastructure. Two use cases are outlined: i) to detect the presence of the drive construct in a new population, and ii) to provide an estimate of drive frequency prior to more accurate laboratory-based measurements. We provide a target product profile for the RDT outlining minimally essential and ideal characteristics, including test procedures, sensitivity, specificity, usability by a range of stakeholders in field settings, and compatibility with pooled testing of mosquito samples. An RDT for gene drive construct detection would support community access and participation in monitoring, enhance regulatory oversight, and promote transparency in field trials, thereby facilitating responsible deployment of gene drive-based malaria interventions.

genetics↗

bioRxiv2025

Y-linked editors for invasive rodent control

Invasive rodents are major contributors to biodiversity loss, particularly on islands where native species have evolved in their absence, rendering them vulnerable to predation. Genetic biocontrol offers a promising suite of species-specific technologies that may contribute to rodent suppression and elimination. Here, we evaluate the potential of Y-linked genome editors (YLEs), which contain a nuclease that disrupts a female-essential fertility or viability gene, to suppress or eliminate a mouse population on a small, isolated island. We benchmark YLEs against other self-limiting genetic biocontrol tools, such as sterile male releases, and self-sustaining tools, such as population suppression gene drives and Y-linked X chromosome-shredders. We use the MouseGD simulation framework to model the inheritance patterns of these systems in the context of rodent life history, calculating elimination probabilities, times to elimination and other outcomes for a range of construct designs, fitness costs and release schemes. We find that YLEs are more efficient than other self-limiting tools, and are capable of achieving rodent elimination within a short timeframe for modest release sizes. For a mouse population size of 10,000, elimination is predicted within five years for releases of 350 males per month, and within ten years for releases of 150 males per month. This scale of production is well within existing capabilities, potentially enabling suppression to encompass a larger spatial scale. We found that elimination could be achieved for YLEs targeting both haploinsufficient and haplosufficient target genes; but for a more restrictive parameter space for the haplosufficient case. Gene drives were predicted to achieve suppression and elimination for smaller release sizes; but also to spread to non-target populations. In contrast, YLEs do not bias inheritance and hence display minimal spillover. Altogether, these characteristics present YLEs as a promising, ecologically-manageable biocontrol tool for elimination of invasive rodents and conservation of native species.

ecology↗

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↗

bioRxiv2025

Mosquito Dispersal in Context

Mosquito dispersal plays an important role in mosquito ecology and mosquito-borne pathogen transmission. While reaction-diffusion and patch-based models with simple flux assumptions for emigration have played a predominant role in modeling mosquito dispersal, mosquito behavioral ecology - in particular, the process of searching for resources - is usually ignored by diffusion-based models. We thus set out to analyze mosquito movement using highly mimetic models, to see what we could learn from a different approach. Here, we present two behavioral state microsimulation models, in which mosquitoes are in behavioral states and search for required resources, which are distributed on a landscape. Models of this sort are laborious and challenging to work with, so we developed ramp.micro, an R package to build, solve, analyze, and visualize behavioral state microsimulation models for mosquitoes. Mosquito population dispersal is an emergent feature in these complex systems, so we developed new methods to describe and understand it. We show that population structure might not be revealed by a crude analysis of observed mosquito movement patterns, unless it can somehow account for sequences of flights for mosquitoes in behavioral states trying to accomplish a goal. We also show that even when resources are distributed randomly and uniformly, mosquito populations tend to form highly spatially structured communities. We also show that some heterogeneity in mosquito population densities is attributed to features of a network defined by searching and the spatial distribution of resources. These models highlight the importance of understanding mosquito behavioral states and resource availability in understanding mosquito population dispersal, and they point to the importance of local context. Motivated by these dynamics, spatial models for mosquito ecology and mosquito-borne pathogen transmission would benefit from considering resource availability as a factor affecting mosquito movement and dispersal. Author summaryWe develop highly realistic population dynamic models for mosquito behavioral ecology where mosquitoes move around on landscapes made up of resources located at points in space. The mosquitoes are in behavioral states - blood feeding, egg laying, or sugar feeding - and they end each search at a point where the resource they need can be found. Even with simple assumptions, mosquito populations can be highly structured. Because mosquitoes tend to leave areas that lack a resource, and they tend to stay in areas that have all required resources, the scarcest resources end up determining the structure of mosquito populations.

ecology↗

bioRxiv2024

Finding divergent sequences of homomorphic sex chromosomes via diploidized nanopore-based assembly from a single male

Although homomorphic sex chromosomes can have non-recombining regions with elevated sequence divergence between its complements, such divergence signals can be difficult to detect bioinformatically. If found in genomes of e.g. insect pests, these sequences could be targeted by the engineered genetic sexing and control systems. Here, we report an approach that can leverage long-read nanopore sequencing of a single XY male to identify divergent regions of homomorphic sex chromosomes. Long-read data are used for de novo genome assembly that is diploidized in a way that maximizes sex-specific differences between its haploid complements. We show that the correct assembly phasing is supported by the mapping of nanopore reads from the males haploid Y-bearing sperm cells. The approach revealed a highly divergent region (HDR) near the centromere of the homomorphic sex chromosome of Aedes aegypti, the most important arboviral vector, for which there is a great interest in creating new genetic control tools. HDR is located [~]5Mb downstream of the known male-determining locus on chromosome 1 and is significantly enriched for ovary-biased genes. While recombination in HDR ceased relatively recently ([~]1.4 MYA), HDR gametologs have divergent exons and introns of protein coding genes, and most lncRNA genes became X-specific. Megabases of previously invisible sex-linked sequences provide new putative targets for engineering the genetic systems to control this deadly mosquito. Broadly, our approach expands the toolbox for studying cryptic structure of sex chromosomes.

genomics↗

bioRxiv2023

MGDrivE 3: A decoupled vector-human framework for epidemiological simulation of mosquito genetic control tools and their surveillance

Novel mosquito genetic control tools, such as CRISPR-based gene drives, hold great promise in reducing the global burden of vector-borne diseases. As these technologies advance through the research and development pipeline, there is a growing need for modeling frameworks incorporating increasing levels of entomological and epidemiological detail in order to address questions regarding logistics and biosafety. Epidemiological predictions are becoming increasingly relevant to the development of target product profiles and the design of field trials and interventions, while entomological surveillance is becoming increasingly important to regulation and biosafety. We present MGDrivE 3 (Mosquito Gene Drive Explorer 3), a new version of a previously-developed framework, MGDrivE 2, that investigates the spatial population dynamics of mosquito genetic control systems and their epidemiological implications. The new framework incorporates three major developments: i) a decoupled sampling algorithm allowing the vector portion of the MGDrivE framework to be paired with a more detailed epidemiological framework, ii) a version of the Imperial College London malaria transmission model, which incorporates age structure, various forms of immunity, and human and vector interventions, and iii) a surveillance module that tracks mosquitoes captured by traps throughout the simulation. Example MGDrivE 3 simulations are presented demonstrating the application of the framework to a CRISPR-based homing gene drive linked to dual disease-refractory genes and their potential to interrupt local malaria transmission. Simulations are also presented demonstrating surveillance of such a system by a network of mosquito traps. MGDrivE 3 is freely available as an open-source R package on CRAN (https://cran.r-project.org/package=MGDrivE2) (version 2.1.0), and extensive examples and vignettes are provided. We intend the software to aid in understanding of human health impacts and biosafety of mosquito genetic control tools, and continue to iterate per feedback from the genetic control community. Author summaryVector-borne diseases such as malaria cause massive morbidity and mortality throughout much of the world. Currently-available control measures, such as insecticide-based tools and antimalarial drugs, have limited impact and are waning in effectiveness, hence there is a need for novel tools to complement existing ones. Mosquito genetic control tools, such as gene drive systems and genetic versions of the sterile insect technique, offer a range of promising options, the development of which has greatly expanded since the advent of CRISPR-based gene-editing. Recently, we proposed MGDrivE 2 (Mosquito Gene Drive Explorer 2), which incorporates epidemiology into simulations of the dynamics of these systems in spatially-structured mosquito populations; however, that framework relied on simple model representations of vector-borne diseases. Here, we present MGDrivE 3, which decouples the vector portion of the model from the human portion, allowing the mosquito genetic control framework to be paired with more-detailed epidemiological frameworks. As an example, we implement the human transmission dynamics of the Imperial College London malaria model. We also incorporate a network of mosquito traps for surveillance. As genetic control technology edges closer towards field implementation, more detailed predictions of its epidemiological and biosafety implications are needed. We propose MGDrivE 3 to fulfill this role.

ecology↗

bioRxiv2023

Cost-effectiveness of Precision Guided SIT for Control of Anopheles gambiae in the Upper River Region, The Gambia.

Precision-guided sterile insect technique (pgSIT) is an extremely promising vector control intervention that can reduce and potentially eliminate the unacceptable malaria burden, particularly in sub-Saharan Africa. The deployment of pgSIT shows the greatest promise and does not have a near peer competitor in the fight to eradicate malaria. Here we explore the cost effectiveness of using this approach in Africa using mathematical modeling and economical analysis. Overall, we find that pgSIT represents a cost-effective and promising approach to A. gambiae control in The Gambia, with the potential to deliver significant economic and social benefits. SummaryPrecision-guided sterile insect technique (pgSIT) is an extremely promising vector control intervention that can reduce and potentially eliminate the unacceptable malaria burden, particularly in sub-Saharan Africa. pgSIT is a safe, innovative, and highly targeted approach to mosquito control that combines the principles of the sterile insect technique (SIT) with advanced state-of-the-art technologies of genetic engineering (Akbari et al. 2023; M. Li et al. 2021; Kandul, Liu, and Akbari 2021; Kandul et al. 2019, 2022). The technique involves inundative releases of genetically sterile male mosquitoes into the environment to mate with their wild counterparts, sterilizing them in the process. After multiple releases, this method can suppress, and even eradicate pest populations without the use of chemical pesticides or other less specific agents. The use of pgSIT has the potential to be a lasting, safe, cost-effective, sustainable, and environmentally friendly method to suppress a target species, acting as a chemical-free, species-specific insecticide. Before this work is considered for field application, however, we need to have a robust, data-driven modeling framework that will accurately predict the outcome of pgSIT release scenarios for malaria control, and we need an assessment of the costs and health benefits of implementing this technology in a region in Africa. This cost assessment evaluates the economic feasibility of both capacity building and establishing the infrastructure for a pgSIT facility in The Gambia to control the deadly malaria mosquito vector Anopheles gambiae. We focused on the Upper River region (URR) for three key reasons: (1) this area is known to have the highest per capita malaria rates in The Gambia (2) this region has more comprehensive historical and current data on malaria incidence and prevalence, malaria-associated healthcare and prevention costs, and human demographic data and (3) its location could feasibly demonstrate that a single pgSIT production facility can support phased suppression on a regional scale and then upon local extinction be repurposed to support vector suppression on a country or continent-wide scale. The pgSIT treatment of the URR ([~]2069 km2) is predicted to prevent approximately 230 deaths and about 48,000 sick days per year. This estimate is based on a model for localized extinction of A. gambiae, reaching full epidemiological impact by the third year. There are multiple ways to calculate the value of this intervention monetarily, from the value of statistical life (VSL) to quality adjusted life years (QALY), which ranges from 367 million to 880 million USD saved in the first ten years of the facility being active. Other metrics such as willingness-to-pay (WTP), estimates the willingness of locals to contribute to malaria prevention financially, and estimates based on gross domestic product (GDP) growth predict this model to save either 53 million or 551 million USD, respectively, in the first ten years of intervention. This model assumes localized extinction of A. gambiae by the second year of intervention with repeated releases to maintain extinction despite seasonal mosquito migration from beyond the treatment area. Localized extinction, however, is expected to have a year-to-year suppression effect making it easier to suppress mosquito populations with reduced sterile male releases in subsequent years. It is, therefore, likely an underestimate of the costs and benefits of pgSIT sterile male releases. In later years, the release of sterile males from this facility could be redirected to new areas to expand the suppression region. Additionally, this facility would have a significant off-season where the facility is not producing sterile male A. gambiae. This off-season could, therefore, produce sterile males to suppress A. gambiae populations in other regions with seasonal malaria during the off-season in The Gambiae. It could even be used to mass produce pgSIT sterile males for other mosquito species, such as the dengue vector, Aedes aegypti, which have eggs that can be stored for many months and consequently can be stockpiled to aid in the suppression of dengue outbreaks, which are also common in the area. Initially, however, the off-season can be used to build local capacity for genetically engineered (GE) mosquitos, which would consist of training staff, optimizing procedures, and troubleshooting any issues that arose during the higher production phases. The costs for this approach will vary as there are some unknowns and variability in the expected efficiencies of the facility, equipment, and procedures. In particular, mosquito survival rates and fecundity may vary more widely at scale and selection of the rearing equipment and protocols, and the mosquito sorting technology will dictate production levels and procedures. This variability is factored into the cost, and, therefore, a range of technologies and costs are considered. The expected start-up costs range from 6 to 11.5 million USD, which includes all necessary development, field trials, equipment, facility construction, staffing, and other establishment costs. The major upfront costs vary by facility size and capital equipment, much of which is dependent on mosquito rearing efficiencies. The annual estimated operational costs range from approximately 315,000 USD, depending on equipment selection and the size of the facility. Annually, this intervention costs less than 1 USD per person to suppress mosquitoes in the URR and prevent malaria transmission, which is about 6% of a malaria prevention WTP based on current interventions or 0.3-11% when using the VSL or QALY metrics, with most variation from the VSL calculation method. This intervention costs 15 to 124 USD (2022) to save one life-year and prevents malaria infections at 13 to 113 USD per case prevented, making this method competitive with many current interventions. WTP demonstrates that this facility could be made locally sustainable long term by local funding. Overall, the estimates of the capital and operational costs associated with the production of pgSIT sterile males, and the construction and management of the production facility indicate the cost savings associated with the annual decrease in morbidity and mortality (value of life) resulting from the use of pgSIT are significantly higher than the implementation costs. These estimates suggest that pgSIT represents a cost-effective and promising approach to A. gambiae control in The Gambia, with the potential to deliver significant economic and social benefits.

genetics↗

bioRxiv2023

Eliminating Malaria Vectors with Precision Guided Sterile Males

Controlling the principal African malaria vector, the mosquito Anopheles gambiae, is considered essential to curtail malaria transmission. However existing vector control technologies rely on insecticides, which are becoming increasingly ineffective. Sterile insect technique (SIT) is a powerful suppression approach that has successfully eradicated a number of insect pests, yet the A. gambiae toolkit lacks the requisite technologies for its implementation. SIT relies on iterative mass-releases of non-biting, non-driving, sterile males which seek out and mate with monandrous wild females. Once mated, females are permanently sterilized due to mating-induced refractoriness, which results in population suppression of the subsequent generation. However, sterilization by traditional methods renders males unfit, making the creation of precise genetic sterilization methods imperative. Here we develop precision guided Sterile Insect Technique (pgSIT) in the mosquito A. gambiae for inducible, programmed male-sterilization and female-elimination for wide scale use in SIT campaigns. Using a binary CRISPR strategy, we cross separate engineered Cas9 and gRNA strains to disrupt male-fertility and female-essential genes, yielding >99.5% male-sterility and >99.9% female-lethality in hybrid progeny. We demonstrate that these genetically sterilized males have good longevity, are able to induce population suppression in cage trials, and are predicted to eliminate wild A. gambiae populations using mathematical models, making them ideal candidates for release. This work provides a valuable addition to the malaria genetic biocontrol toolkit, for the first time enabling scalable SIT-like confinable suppression in the species.

bioengineering↗

bioRxiv2023

MGSurvE: A framework to optimize trap placement for genetic surveillance of mosquito population

Genetic surveillance of mosquito populations is becoming increasingly relevant as genetics-based mosquito control strategies advance from laboratory to field testing. Especially applicable are mosquito gene drive projects, the potential scale of which leads monitoring to be a significant cost driver. For these projects, monitoring will be required to detect unintended spread of gene drive mosquitoes beyond field sites, and the emergence of alternative alleles, such as drive-resistant alleles or non-functional effector genes, within intervention sites. This entails the need to distribute mosquito traps efficiently such that an allele of interest is detected as quickly as possible - ideally when remediation is still viable. Additionally, insecticide-based tools such as bednets are compromised by insecticide-resistance alleles for which there is also a need to detect as quickly as possible. To this end, we present MGSurvE (Mosquito Gene SurveillancE): a computational framework that optimizes trap placement for genetic surveillance of mosquito populations such that the time to detection of an allele of interest is minimized. A key strength of MGSurvE is that it allows important biological features of mosquitoes and the landscapes they inhabit to be accounted for, namely: i) resources required by mosquitoes (e.g., food sources and aquatic breeding sites) can be explicitly distributed through a landscape, ii) movement of mosquitoes may depend on their sex, the current state of their gonotrophic cycle (if female) and resource attractiveness, and iii) traps may differ in their attractiveness profile. Example MGSurvE analyses are presented to demonstrate optimal trap placement for: i) an Aedes aegypti population in a suburban landscape in Queensland, Australia, and ii)an Anopheles gambiae population on the island of Sao Tome, Sao Tome and Principe. Further documentation and use examples are provided in projects documentation. MGSurvE is freely available as an open-source Python package on pypi (https://pypi.org/project/MGSurvE/). It is intended as a resource for both field and computational researchers interested in mosquito gene surveillance. Author summaryMosquito-borne diseases such as malaria and dengue fever continue to pose a major health burden throughout much of the world. The impact of currently-available tools, such as insecticides and antimalarial drugs, is stagnating, and gene drive-modified mosquitoes are considered a novel tool that could contribute to continuing reductions in disease transmission. Gene drive approaches are unique in the field of vector control in that they involve transgenes that could potentially spread on a wide scale, and consequently, surveillance is expected to be a major cost driver for the technology. This is needed to monitor for unintended spread of intact drive alleles, and the emergence of alternative alleles such as homing-resistance alleles and non-functional effector genes. Additionally, surveillance of insecticide-resistance alleles is of interest to support the impact of insecticide-based tools such as bednets. Here, we present MGSurvE, a computational framework that optimizes trap placement for genetic surveillance of mosquito populations in order to minimize the time to detection for an allele of interest. MGSurvE has been tailored to various features of mosquito ecology, and is intended as a resource for researchers to optimize the efficiency of limited surveillance resources.

bioinformatics↗

bioRxiv2023

Targeting Sex Determination to Suppress Mosquito Populations

Each year, hundreds of millions of people are infected with arboviruses such as dengue, yellow fever, chikungunya, and Zika, which are all primarily spread by the notorious mosquito Aedes aegypti. Traditional control measures have proven insufficient, necessitating innovations. In response, here we generate a next generation CRISPR-based precision-guided sterile insect technique (pgSIT) for Aedes aegypti that disrupts genes essential for sex determination and fertility, producing predominantly sterile males that can be deployed at any life stage. Using mathematical models and empirical testing, we demonstrate that released pgSIT males can effectively compete with, suppress, and eliminate caged mosquito populations. This versatile species-specific platform has the potential for field deployment to effectively control wild populations of disease vectors.

bioengineering↗

bioRxiv2023

Mechanical transmission of Dengue Virus by Aedes aegypti may influence disease transmission dynamics during outbreaks

The escalating number of dengue virus (DENV) outbreaks and their worldwide spread pose a major threat to global public health. DENV transmission dynamics significantly influence outbreak duration and magnitude. Conventional DENV transmission requires an incubation period between mosquitoes biting infected humans and the mosquitoes becoming infectious. However, the possibility of immediate, mechanical transmission of DENV without viral replication in the mosquito has received little attention despite its potential importance. Here, we show that Aedes aegypti mosquitoes can mechanically transmit DENV to susceptible mice immediately after biting infected mice without the need for an incubation period. By incorporating parameters from our experiments into a newly developed mathematical model, we found a significant impact on DENV outbreak characteristics. Mechanical transmission may amplify existing disease transmission routes and influence outbreak dynamics. Our findings have implications for vector control strategies that target mosquito lifespan and suggest the possibility of similar mechanical transmission routes in other disease-carrying mosquitoes.

microbiology↗

bioRxiv2022

wMel replacement of dengue-competent mosquitoes is robust to near-term climate change

Rising temperatures and increasing temperature variability are impacting the range and prevalence of mosquito-borne disease. A promising biocontrol technology replaces wild mosquitoes with those carrying the virus-blocking Wolbachia bacterium. Laboratory and field observations show that the most widely used strain, wMel, is adversely affected by heat stress. Here, we examine whether and how climate warming may impact wMel-based replacement. We integrate empirical data on the temperature sensitivity of wMel bacteria into a mechanistic model of population dynamics for the dengue vector Aedes aegypti and use CMIP5 climate projections and historical temperature records from Cairns, Australia to simulate vector control interventions. We show that higher mean temperatures are predicted to lower wMel infection frequency and that extended heatwaves have the potential to reverse the public health benefits of this intervention. Sensitivity analysis probing the thermal limits of wMel replacement reveal that, under existing projections, operational adaptations would be required for heatwaves lasting longer than two weeks. We conclude that this technology is expected to be robust to both the increased mean temperatures and heatwaves associated with near-term climate change in temperate regions. However, more rapid warming or tropical and inland regions that presently feature hotter baselines may challenge these tested limits, requiring further research.

ecology↗

bioRxiv2022

A confinable female-lethal population suppression system in the malaria vector, Anopheles gambiae

Malaria is among the worlds deadliest diseases, predominantly affecting sub-Saharan Africa, and killing over half a million people annually. Controlling the principal vector, the mosquito Anopheles gambiae, as well as other anophelines, is among the most effective methods to control disease spread. Here we develop an innovative genetic population suppression system termed Ifegenia (Inherited Female Elimination by Genetically Encoded Nucleases to Interrupt Alleles) in this deadly vector. In this bicomponent CRISPR-based approach, we disrupt a female-essential gene, femaleless (fle), demonstrating complete genetic sexing via heritable daughter gynecide. Moreover, we show that Ifegenia males remain reproductively viable, and can load both fle mutations and CRISPR machinery to induce fle mutations in subsequent generations, resulting in sustained population suppression. Through modeling, we demonstrate that iterative releases of non-biting Ifegenia males can act as an effective, confinable, controllable, and safe population suppression and elimination system.

bioengineering↗

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↗