Scholarly Literature
This is an aggregation of scholarly literature from peer-reviewed journals, bioRxiv , medRxiv and arXiv preprint servers focused on natural and engineered selfish genetic elements and genetic biocontrol. New citations are added as they appear. This content is intended for anyone interested in the scholarly literature on gene drive and genetic biocontrol.
Gene drive strategies of pest control in agricultural systems: challenges and opportunities
17836M. Legros, J. M. Marshall, S. Macfadyen, K. R. Hayes, A. Sheppard and L. G. Barrett, Evolutionary Applications, 2021-07-26 14:04:32.
Abstract Recent advances in gene editing technologies have opened new avenues for genetic pest control strategies, in particular around the use of gene drives to suppress or modify pest populations. Significant uncertainty, however, surrounds the applicability of these strategies to novel target species, their efficacy in natural populations, and their eventual safety and acceptability as control methods. In this article we identify issues associated with the potential use of gene drives in agricultural systems, to control pests and diseases that impose a significant cost to agriculture around the world. We first review the need for innovative approaches, and provide an overview of the most relevant biological and ecological traits of agricultural pests that could impact the outcome of gene drive approaches. We then describe the specific challenges associated with using gene drives in agricultural systems, as well as the opportunities that these environments may offer, focusing in particular on the advantages of high-threshold gene drives. Overall we aim to provide a comprehensive view of the potential opportunities and the remaining uncertainties around the use of gene drives in agricultural systems.
Resistance to a CRISPR-based gene drive at an evolutionarily conserved site is revealed by mimicking genotype fixation
17940S. Fuchs, W. Garrood, A. Beber, A. Hammond, R. Galizi, M. Gribble, G. Morselli, T.-Y. Hui, K. Willis, N. Kranjc, A. Burt, T. Nolan and A. Crisanti, bioRxiv, 2021-07-26 13:48:52.
CRISPR-based homing gene drives can be designed to disrupt essential genes whilst biasing their own inheritance, leading to suppression of mosquito populations in the laboratory. This class of gene drives relies on CRISPR-Cas9 cleavage of a target sequence and copying (‘homing’) therein of the gene drive element from the homologous chromosome. However, target site mutations that are resistant to cleavage yet maintain the function of the essential gene are expected to be strongly selected for. Targeting functionally constrained regions where mutations are not easily tolerated should lower the probability of resistance. Evolutionary conservation at the sequence level is often a reliable indicator of functional constraint, though the actual level of underlying constraint between one conserved sequence and another can vary widely. Here we generated a novel gene drive in the malaria vector Anopheles gambiae , targeting an ultra-conserved target site in a haplosufficient essential gene (AGAP029113) required during mosquito development, which fulfils many of the criteria for the target of a population suppression gene drive. We then designed a selection regime to experimentally assess the likelihood of generation and subsequent selection of gene drive resistant mutations at its target site. We simulated, in a caged population, a scenario where the gene drive was approaching fixation, where selection for resistance is expected to be strongest. Continuous sampling of the target locus revealed that a single, restorative, in-frame nucleotide substitution was selected. Our findings show that ultra-conservation alone need not be predictive of a site that is refractory to target site resistance. Our strategy to evaluate resistance in vivo could help to validate candidate gene drive targets for their resilience to resistance and help to improve predictions of the invasion dynamics of gene drives in field populations. <h4>Author summary</h4> Gene drives have the potential to be applied as novel control strategy of disease-transmitting mosquitoes, by spreading genetic traits that suppress or modify the target population. Many gene drive elements work by recognising and cutting a specific target sequence in the mosquito genome and copying themselves into that target sequence allowing the gene drive to increase in frequency in the population. Like other mosquito control interventions, efficacy will greatly depend on minimising the development of resistance to the gene drive mechanism - most likely via a change in the target sequence that prevents further cutting. One strategy to reduce resistance is to target sequences that are highly conserved, which implies that changes cannot easily be tolerated. We developed a strategy that simulates high selection pressure, under which resistance is most likely to emerge, and therefore provides a stringent test of its propensity to arise. Unlike previous results with another gene drive, we recovered a resistant allele within a few generations of gene drive exposure and at high frequency. Our results show that conserved sequences can vary hugely in ability to tolerate mutations and highlights the need to functionally validate future candidate gene drive target sites for their robustness to resistance.
Risk management recommendations for environmental releases of gene drive modified insects
17829Y. Devos, J. D. Mumford, M. B. Bonsall, D. C. M. Glandorf and H. D. Quemada, Biotechnology Advances, 2021-07-25 13:02:06.
The ability to engineer gene drives (genetic elements that bias their own inheritance) has sparked enthusiasm and concerns. Engineered gene drives could potentially be used to address long-standing challenges in the control of insect disease vectors, agricultural pests and invasive species, or help to rescue endangered species. However, risk concerns and uncertainty associated with potential environmental release of gene drive modified insects (GDMIs) have led some stakeholders to call for a global moratorium on such releases or the application of other strict precautionary measures to mitigate perceived risk assessment and risk management challenges. Instead, we provide recommendations that may help to improve the relevance of risk assessment and risk management frameworks for environmental releases of GDMIs. These recommendations include: (1) developing additional and more practical risk assessment guidance to ensure appropriate levels of safety; (2) making policy goals and regulatory decision-making criteria operational for use in risk assessment so that what constitutes harm is clearly defined; (3) ensuring a more dynamic interplay between risk assessment and risk management to manage uncertainty through closely interlinked pre-release modelling and post-release monitoring; (4) considering potential risks against potential benefits, and comparing them with those of alternative actions to account for a wider (management) context; and (5) implementing a modular, phased approach to authorisations for incremental acceptance and management of risks and uncertainty. Along with providing stakeholder engagement opportunities in the risk analysis process, the recommendations proposed may enable risk managers to make choices that are more proportionate and adaptive to potential risks, uncertainty and benefits of GDMI applications, and socially robust.
The Aedes aegypti (Diptera: Culicidae) hsp83 Gene Promoter Drives Strong Ubiquitous DsRed and ZsGreen Marker Expression in Transgenic Mosquitoes
18145S. H. Webster and M. J. Scott, Journal of Medical Entomology, 2021-07-24 17:29:43.
Transgenic strains of the mosquito disease vector Aedes aegypti (L.) are being developed for population suppression or modification. Transgenic mosquitoes are identified using fluorescent protein genes. Here we describe DsRed and ZsGreen marker genes driven by the constitutive Ae. aegypti heat shock protein 83 (hsp83) promoter in transgenic mosquitoes. Transgenic larvae and pupae show strong full body expression of the red and green fluorescent proteins. This greatly assists in screening for transgenic individuals while making new or maintaining already established lines. Transient marker gene expression after embryo microinjection was readily visible in developing larvae allowing the separation of individuals that are more likely to produce transgenic offspring. The strongly expressed marker genes developed in this study should facilitate the detection of transgenic Ae. aegypti larvae or pupae in the field.
Sex separation of Aedes spp. mosquitoes for sterile insect technique application: a review
17905B. M. Moran-Aceves, C. F. Marina, A. Dor, P. Liedo and J. Toledo, Entomologia Experimentalis Et Applicata, 10. 2021-07-24 15:04:30.
Separation of the sexes is necessary for the application of the sterile insect technique (SIT) in mosquitoes due to the hematophagous habits and disease vector activity of the females. In this review we analyze the history, current status, and future perspectives for the development of genetic sexing strains (GSS) of Aedes mosquitoes (Diptera: Culicidae). Various genetic control methods for mosquitoes are reviewed, as are their need for sex-separation methods. We focus on areas of opportunity where GSS developed with classical genetic methods can be used. Regulatory restrictions and social acceptance of various control methods are analyzed. We conclude that the development of GSS by classical methods represents the most viable option for separation of the sexes and the application of large-scale SIT programs within an area-wide integrated vector management (AW-IVM) approach.
Wolbachia as translational science: controlling mosquito-borne pathogens
17799E. P. Caragata, H. L. C. Dutra, P. H. F. Sucupira, A. G. A. Ferreira and L. A. Moreira, Trends in Parasitology, 2021-07-22 20:54:55.
In this review we examine how exploiting the Wolbachia?mosquito relationship has become an increasingly popular strategy for controlling arbovirus transmission. Field deployments of Wolbachia-infected mosquitoes have led to significant decreases in dengue virus incidence via high levels of mosquito population suppression and replacement, emphasizing the success of Wolbachia approaches. Here, we examine how improved knowledge of Wolbachia?host interactions has provided key insight into the mechanisms of the essential phenotypes of pathogen blocking and cytoplasmic incompatibility. And we discuss recent studies demonstrating that extrinsic factors, such as ambient temperature, can modulate Wolbachia density and maternal transmission. Finally, we assess the prospects of using Wolbachia to control other vectors and agricultural pest species.
CRISPR/Cas-9 mediated knock-in by homology dependent repair in the West Nile Virus vector Culex quinquefasciatus Say
17811D.-K. Purusothaman, L. Shackleford, M. A. E. Anderson, T. Harvey-Samuel and L. Alphey, Scientific Reports, 11:14964. 2021-07-22 14:14:08.
Culex quinquefasciatus Say is a mosquito distributed in both tropical and subtropical regions of the world. It is a night-active, opportunistic blood-feeder and vectors many animal and human diseases, including West Nile Virus and avian malaria. Current vector control methods (e.g. physical/chemical) are increasingly ineffective; use of insecticides also imposes hazards to both human and ecosystem health. Advances in genome editing have allowed the development of genetic insect control methods, which are species-specific and, theoretically, highly effective. CRISPR/Cas9 is a bacteria-derived programmable gene editing tool that is functional in a range of species. We describe the first successful germline gene knock-in by homology dependent repair in C. quinquefasciatus. Using CRISPR/Cas9, we integrated an sgRNA expression cassette and marker gene encoding a fluorescent protein fluorophore (Hr5/IE1-DsRed, Cq7SK-sgRNA) into the kynurenine 3-monooxygenase (kmo) gene. We achieved a minimum transformation rate of 2.8%, similar to rates in other mosquito species. Precise knock-in at the intended locus was confirmed. Insertion homozygotes displayed a white eye phenotype in early-mid larvae and a recessive lethal phenotype by pupation. This work provides an efficient method for engineering C. quinquefasciatus, providing a new tool for developing genetic control tools for this vector.
Using Moderate Transgene Expression to Improve the Genetic Sexing System of the Australian Sheep Blow Fly Lucilia cuprina
17753Y. Yan, M. E. Williamson and M. J. Scott, Insects, 11. 2021-07-19 14:36:16.
The sterile insect technique (SIT) is a promising strategy to control the Australian sheep blow fly Lucilia cuprina, a major pest of sheep. We have previously developed a transgenic embryonic sexing system (TESS) for this pest to facilitate the potential SIT application. TESS carry two transgenes, a tetracycline transactivator (tTA) driver and a tTA-activated pro-apoptotic effector. TESS females die at the embryonic stage unless tetracycline is supplied in the diet. However, undesired female sterility was observed in some TESS strains without tetracycline due to expression of tTA in ovaries. Here we investigate if TESS that combine transgenes with relatively low/moderate expression/activity improves the fertility of TESS females. tTA driver lines were evaluated for tTA expression by quantitative real time PCR and/or by crossing with a tTA-activated RFPex effector line. Fertility and lethality tests showed that a TESS strain containing a driver line with moderate tTA expression and an effector line showing moderate pro-apoptotic activity could recover the fertility of parental females and eliminated all female offspring at the embryonic stage. Consequently, such a strain could be further evaluated for an SIT program for L. cuprina, and such a “moderate strategy” could be considered for the TESS development in other pest species.
Combating mosquito-borne diseases using genetic control technologies
17735G.-H. Wang, S. Gamez, R. R. Raban, J. M. Marshall, L. Alphey, M. Li, J. L. Rasgon and O. S. Akbari, Nature Communications, 12:4388. 2021-07-19 13:06:45.
Mosquito-borne diseases, such as dengue and malaria, pose significant global health burdens. Unfortunately, current control methods based on insecticides and environmental maintenance have fallen short of eliminating the disease burden. Scalable, deployable, genetic-based solutions are sought to reduce the transmission risk of these diseases. Pathogen-blocking Wolbachia bacteria, or genome engineering-based mosquito control strategies including gene drives have been developed to address these problems, both requiring the release of modified mosquitoes into the environment. Here, we review the latest developments, notable similarities, and critical distinctions between these promising technologies and discuss their future applications for mosquito-borne disease control.
The Possible Role of Microorganisms in Mosquito Mass Rearing
17741L. Chersoni, A. Checcucci, M. Malfacini, A. Puggioli, F. Balestrino, M. Carrieri, I. Piunti, M. L. Dindo, P. Mattarelli and R. Bellini, Insects, 12. 2021-07-15 13:10:37.
In Europe, one of the most significant mosquitoes of public health importance is Aedes albopictus (Skuse), an allochthonous species of Asian origin. One of the most promising control methods against Aedes albopictus is the sterile insect technique (SIT), which consists of mass rearing the target species, separation of males from females, and male exposure to sterilizing ionizing radiation. Once released in the environment, the sterile males are expected to search for wild females to mate with. If mating occurs, no offspring is produced. The quality of sterile males is a crucial aspect in SIT programs in order to optimize effectiveness and limit production costs. The integration of probiotic microorganisms in larval and adult mosquito diets could enhance the quality parameters of the released sterile males. In this review, we attempt to give the most representative picture of the present knowledge on the relationships between gut microbiota of mosquitoes and the natural or artificial larval diet. Furthermore, the possible use of probiotic microorganisms for mosquito larvae rearing is explored. Based on the limited amount of data found in the literature, we hypothesize that a better understanding of the interaction between mosquitoes and their microbiota may bring significant improvements in mosquito mass rearing for SIT purposes.
Sterile Insect Technique (SIT) and Its Applications
17733K. Bourtzis and M. J. B. Vreysen, Insects, 12. 2021-07-13 15:57:10.
Although most insect species have a beneficial role in the ecosystems, some of them represent major plant pests and disease vectors for livestock and humans. During the last six–seven decades, the sterile insect technique (SIT) has been used as part of area-wide integrated pest management strategies to suppress, contain, locally eradicate or prevent the (re)invasion of insect pest populations and disease vectors worldwide. This Special Issue on “Sterile insect technique (SIT) and its applications”, which consists of 27 manuscripts (7 reviews and 20 original research articles), provides an update on the research and development efforts in this area. The manuscripts report on all the different components of the SIT package including mass-rearing, development of genetic sexing strains, irradiation, quality control as well as field trials.
Autocatalytic-protection for an unknown locus CRISPR-Cas countermeasure for undesired mutagenic chain reactions
17969E. Schonfeld, E. Schonfeld and D. Schonfeld, Journal of Theoretical Biology, 528:110831. 2021-07-11 13:41:33.
The mutagenic chain reaction (MCR) is a genetic tool to use a CRISPR–Cas construct to introduce a homing endonuclease, allowing gene drive to influence whole populations in a minimal number of generations (Esvelt et al., 2014, Gantz and Bier, 2015, Gantz and Bier, 2016). The question arises: if an active genetic terror event is released into a population, could we prevent the total spread of the undesired allele (Gantz, et al., 2015, Webber et al., 2015)? Thus far, effective protection methods require knowledge of the terror locus (Grunwald et al., 2019). Here we introduce a novel approach, an autocatalytic-Protection for an Unknown Locus (a-PUL), whose aim is to spread through a population and arrest and decrease an active terror event’s spread without any prior knowledge of the terror-modified locus, thus allowing later natural selection and ERACR drives to restore the normal locus (Hammond et al., 2017). a-PUL, using a mutagenic chain reaction, includes (i) a segment encoding a non-Cas9 endonuclease capable of homology-directed repair suggested as Type II endonuclease Cpf1 (Cas12a), (ii) a ubiquitously-expressed gene encoding a gRNA (gRNA1) with a U4AU4 3′-overhang specific to Cpf1 and with crRNA specific to some desired genomic sequence of non-coding DNA, (iii) a ubiquitously-expressed gene encoding two gRNAs (gRNA2/gRNA3) both with tracrRNA specific to Cas9 and crRNA specific to two distinct sites of the Cas9 locus, and (iv) homology arms flanking the Cpf1/gRNA1/gRNA2/gRNA3 cassette that are identical to the region surrounding the target cut directed by gRNA1 (Khan, 2016, Zetsche et al., 2015). We demonstrate the proof-of-concept and efficacy of our protection construct through a Graphical Markov model and computer simulation.
Earth system interventions as technologies of the Anthropocene
17730J. L. Reynolds, Environmental Innovation and Societal Transitions, 40:132-146. 2021-07-10 15:16:46.
Earth system interventions (ESIs)—intentional large-scale interventions in Earth systems—are not entirely new. However, in response to threats to sustainability, particularly from climate change and biodiversity loss, some scientists and others are researching, developing, and using new, largely technological ESIs. These include carbon dioxide removal, solar geoengineering, in situ genetically modified organisms, gene drive organisms, de-extinction, and high-tech ecosystem restoration. Some emerging ESIs appear to be effective and feasible, both technically and economically, and may be necessary to achieve important sustainability and human welfare objectives. They also pose serious environmental risks. This paper identifies more than a dozen social, political, and ethical challenges that are common across many of the emerging technological ESIs. Governance could mitigate and manage these issues, and it would be made more effective and robust by understanding and treating new ESIs as such and as a potentially transformative set of innovations in human-Earth system relations.
Improvement of the Mass-Rearing Protocols for the South American Fruit Fly for Application of the Sterile Insect Technique
17764T. Mastrangelo, A. Kovaleski, B. Maset, M. D. Costa, C. Barros, L. A. Lopes and C. Caceres, Insects, 12. 2021-07-09 14:59:26.
The existing rearing protocols for Anastrepha fraterculus must be reviewed to make economically viable the production of sterile flies for their area-wide application. Additionally, evidence of sexual incompatibility between a long-term mass-reared Brazilian strain and wild populations has been found. To address these issues, this study aimed to refine rearing protocols and to assess the suitability of an A. fraterculus strain for the mass production of sterile flies. A series of bioassays were carried out to evaluate incubation times for eggs in a bubbling bath and to assess the temporal variation of egg production from ovipositing cages at different adult densities. A novel larval diet containing carrageenan was also evaluated. Egg incubation times higher than 48 h in water at 25 °C showed reduced larval and pupal yields. Based on egg production and hatchability, the density of 0.3 flies/cm2 can be recommended for adult cages. The diet with carrageenan was suitable for mass production at egg-seeding densities between 1.0 and 1.5 mL of eggs/kg of diet, providing higher insect yields than a corn-based diet from Embrapa. Even after two years of being reared under the new rearing protocols, no sexual isolation was found between the bisexual strain and wild flies.
Comparative response to post-production process of two Anastrepha ludens strains: Application in the sterile insect technique
17725J. Arredondo, J. F. Aguirre-Medina, J. S. Meza, J. Cancino and F. Diaz-Fleischer, Journal of Applied Entomology, 11. 2021-07-07 14:54:23.
The new desiccation-resistant (DR) strain of Anastrepha ludens Loew differs in its life-history traits from the non-selected strain (NS). Given the innate resistance of DR flies to stressors, it is necessary to determine the packing and shipment conditions for their use in the sterile insect technique (SIT). First, we used pupae to evaluate the interaction of hypoxia period (12, 24 and 36 hr) and temperature (15.3 (+/- 0.9), 19.5 (+/- 1.0), 25.5 (+/- 0.6) and 30.1 (+/- 0.5) C-o). Second, adults were exposed to densities of 1.0, 1.3 and 1.5 flies/cm(2) and chilling periods of 0, 3 and 5 hr. We observed a negative relationship of emergence and flight ability with hypoxia period and temperature in both strains especially when flies were exposed to 36 hr of hypoxia and 30celcius. Interestingly, knock-down time decreased as male density increased, but when chilling time increased, recovery time also increased. Both chilling time and male density had negative effects on male weight. Chilling time reduced male survival when exposed to desiccation, while both chilling time and male density decreased male survival under starvation conditions. Negative effects on number of matings were observed only on the first day after chilling but not on the following days. We found that DR flies survive approximately twice as long as flies from the NS strain when exposed to starvation or desiccation stressors. We discuss our results on the basis of the potential use of DR flies in SIT programmes.
Distinct spermiogenic phenotypes underlie sperm elimination in the Segregation Distorter meiotic drive system
17832M. Herbette, X. L. Wei, C. H. Chang, A. M. Larracuente, B. Loppin and R. Dubruille, PLOS Genetics, 17:26. 2021-07-06 13:14:02.
Here we show that SD/SD+ males of different genotypes but with similarly strong degrees of distortion have distinct spermiogenic phenotypes. In some genotypes, SD+ spermatids fail to fully incorporate protamines after the removal of histones, and degenerate during the individualization stage of spermiogenesis. In contrast, in other SD/SD+ genotypes, protamine incorporation appears less disturbed, yet spermatid nuclei are abnormally compacted, and mature sperm nuclei are eventually released in the seminal vesicle. Our analyses of different SD+ chromosomes suggest that the severity of the spermiogenic defects associates with the copy number of the Rsp satellite. We propose that when Rsp copy number is very high (> 2000), spermatid nuclear compaction defects reach a threshold that triggers a checkpoint controlling sperm chromatin quality to eliminate abnormal spermatids during individualization. Author summary In diploid organisms, both alleles of the same gene have an equal chance to be transmitted to the progeny. However, in many species including mammals, insects and plants, selfish genetic elements perturb gametogenesis in a way that favors their own transmission to the detriment of the homologous chromosome that does not carry them. In Drosophila melanogaster, Segregation Distorter (Sd) is a well-characterized selfish locus that induces, by still-unclear mechanisms, the elimination of sperm cells which contain the homologous second chromosome when this carries a large heterochromatic block of repetitive DNA called Rsp. Here, we show that in Sd males, the replacement of histones by sperm specific protamine-like proteins is perturbed in the differentiating Rsp sperm cells, which are then eliminated before their release in the seminal vesicle. However, in some genetic backgrounds, many spermatids Rsp tend to escape this elimination and defective sperm nuclei are found in the seminal vesicle. We show that these phenotypes are partly linked to the number of repeats in the Rsp block and can be modulated by suppressors present in the genetic background. Our work thus helps to understand how selfish loci exploit gametogenesis to favor their own transmission and highlights the essential role of heterochromatin in spermiogenesis progression.
Effect of the timing of pupal irradiation on the quality and sterility of oriental fruit flies (Diptera: Tephritidae) for use in Sterile Insect Technique
17781T. J. Fezza, P. A. Follett and T. E. Shelly, Applied Entomology and Zoology, 8. 2021-07-05 17:37:25.
The Sterile Insect Technique (SIT) is a target-specific, biologically based method used to control pestiferous tephritids entailing the release of mass-reared, sterilized males of the target species to achieve sterile male x wild female matings. As documented for several tephritid species, sterilizing irradiation may have adverse effects on various biological parameters, including life span, flight ability, and mating competitiveness. To minimize these impacts and ensure sterility, released flies must be irradiated at a precise dose at a specific and uniform age. The objective of the present study was to determine the impact of pupal age at the time of irradiation on flight ability, male survival, fertility, and mating competitiveness for the oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae) irradiated at 100 Gy. We concluded that flies irradiated as pupae 4 days before emergence were significantly compromised with respect to flight ability and male survival when compared to flies irradiated 1 or 2 days before emergence. Additionally, we determined that crosses between males irradiated 1 day before emergence with non-irradiated females yielded significantly more pupae than the same cross with males irradiated at 2, 3, and 4 days before emergence. Although rare, the occurrence of pupae and emerged adults suggested low levels of fertility for flies irradiated as pupae 1 day before emergence, but complete sterility at 2, 3, and 4 days before emergence. These findings suggest that SIT programs can potentially irradiate pupae at 2 or 3 days before emergence without compromising the quality of the released flies, which would permit the distribution of flies over greater distances.
Gene drive that results in addiction to a temperature sensitive version of an essential gene triggers population collapse in Drosophila
17609G. Oberhofer, B. Hay and T. Ivy, bioRxiv, 2021.07.03.451005. 2021-07-04 14:38:01.
One strategy for population suppression seeks to use gene drive to spread genes that confer conditional lethality or sterility, providing a way of combining population modification with suppression. Stimuli of potential interest could be introduced by humans, such as an otherwise benign virus or chemical, or occur naturally on a seasonal basis, such as a change in temperature. Cleave and Rescue (ClvR) selfish genetic elements use Cas9 and gRNAs to disrupt endogenous versions of an essential gene, while also including a Rescue version of the essential gene resistant to disruption. ClvR spreads by creating loss-of-function alleles of the essential gene that select against those lacking it, resulting in populations in which the Rescue provides the only source of essential gene function. In consequence, if function of the Rescue, a kind of Trojan horse now omnipresent in a population, is condition-dependent, so too will be the survival of that population. To test this idea we created a ClvR in Drosophila in which Rescue activity of an essential gene, dribble, requires splicing of a temperature-sensitive intein (TS-ClvRdbe). This element spreads to transgene fixation at 23° C, but when populations now dependent on TS-ClvRdbe are shifted to 29° C death and sterility result in a rapid population crash. These results show that conditional population elimination can be achieved. A similar logic, in which Rescue activity is conditional, could also be used in HEG-based drive, and to bring about suppression and/or killing of specific individuals in response to other stimuli.Competing Interest StatementThe authors have filed patent applications on ClvR and related 336 technologies (U.S. Application No. 15/970,728 and No. 16/673,823 ; provisional patent No. 337 CIT-8511-P )
Remating in Ceratitis capitata sterile males: Implications in sterile insect technique programmes
17827M. Catala-Oltra, E. Llacer, O. Dembilio, I. Pla, A. Urbaneja and M. Perez-Hedo, Journal of Applied Entomology, 8. 2021-07-02 14:57:45.
Sterile insect technique (SIT) is used, among other biological control tools, as a sustainable measure for the management of Ceratitis capitata Wiedemann (Diptera: Tephritidae) in many agricultural regions where this pest can trigger severe economic impacts. The tendency of wild females to remate multiple times has been deeply studied; it has been a common point of controversy when evaluating SIT programmes. Nevertheless, the remating potential of the released sterile males remains unknown. Here, under laboratory conditions, the remating capability of mass-reared sterile males was determined. Wild-type virgin females were offered to sterile males (Vienna-8 strain), which had the opportunity to mate up to four consecutive times. The remating assays were carried out at 24 hr, 48 hr, 4 days and 7 days after the first mating. At the end of each tested time period, males were divided according to their mating response, mated or unmated, and subsequently reused for the next round of mating assays. The frequency of successful remating in each tested time period was obtained. Insemination was confirmed by determining the sperm transfer in mated female spermathecae by quantitative real-time PCR. Our results demonstrate that 73% of the mass-reared sterile males were able to remate 24 hr after the first mating, 55% of which remated again the day after. Close to 25% of the V8 sterile males tended to copulate in all of the four mating opportunities. The qPCR analysis of the spermathecae contents verified an effective transfer of V8 sperm to wild females with every mating; 99% of copulations resulted in sperm transfer. These findings shed light on the remating potential of V8 sterile males, an aspect until now underestimated in many SIT programmes.
Preventing COVID-59
17604Y. Erlich and D. Douek, github, 2021-06-30 14:30:52.
SARS-CoV-2 is the third betacoronavirus to enter the human population in the past 20 years, revealing a concerning pattern. Clearly, preventing a future pandemic from such viruses is a critical priority. Previous studies have shown that shRNAs can be powerful suppressors of RNA viruses in transgenic animals and substantially reduce transmission. Thus, we propose the introduction of anti-betacoronavirus shRNAs using CRISPR/CAS9 gene drive into the horseshoe bat population, the natural reservoir of those viruses, to combat this pandemic threat at its source. Importantly, our approach is not expected to create any harm to bats and can benefit other animals in the ecosystem that contract betacoronaviruses from bats. We map the ethical and the technical aspects and suggest guidelines for moving forward with this proposal.
INSIDER: alignment-free detection of foreign DNA sequences
17657A. P. Tay, B. Hosking, C. Hosking, D. C. Bauer and L. O. W. Wilson, Computational and Structural Biotechnology Journal, 19:3810-3816. 2021-06-29 19:39:26.
External DNA sequences can be inserted into an organism’s genome either through natural processes such as gene transfer, or through targeted genome engineering strategies. Being able to robustly identify such foreign DNA is a crucial capability for health and biosecurity applications, such as anti-microbial resistance (AMR) detection or monitoring gene drives. This capability does not exist for poorly characterised host genomes or with limited information about the integrated sequence. To address this, we developed the INserted Sequence Information DEtectoR (INSIDER). INSIDER analyses whole genome sequencing data and identifies segments of potentially foreign origin by their significant shift in k-mer signatures. We demonstrate the power of INSIDER to separate integrated DNA sequences from normal genomic sequences on a synthetic dataset simulating the insertion of a CRISPR-Cas gene drive into wild-type yeast. As a proof-of-concept, we use INSIDER to detect the exact AMR plasmid in whole genome sequencing data from a Citrobacter freundii patient isolate. INSIDER streamlines the process of identifying integrated DNA in poorly characterised wild species or when the insert is of unknown origin, thus enhancing the monitoring of emerging biosecurity threats.
A wAlbB Wolbachia< transinfection displays stable phenotypic effects across divergent Aedes aegypti mosquito backgrounds
17592P. A. Ross, X. Gu, K. L. Robinson, Q. Yang, E. Cottingham, Y. Zhang, H. L. Yeap, X. Xu, N. M. Endersby-Harshman and A. A. Hoffmann, bioRxiv, 2021.06.25.450002. 2021-06-26 14:00:37.
Aedes mosquitoes harboring intracellular Wolbachia bacteria are being released in arbovirus and mosquito control programs. With releases taking place around the world, understanding the contribution of host variation to Wolbachia phenotype is crucial. We generated a Wolbachia transinfection (wAlbBQ) in Aedes aegypti and performed backcrossing to introduce the infection into Australian or Malaysian nuclear backgrounds. Whole Wolbachia genome sequencing shows that the wAlbBQ transinfection is near-identical to the reference wAlbB genome, suggesting few changes since the infection was first introduced to Ae. aegypti over 15 years ago. However, these sequences were distinct from other available wAlbB genome sequences, highlighting the potential diversity of wAlbB in natural Ae. albopictus populations. Phenotypic comparisons demonstrate effects of wAlbB infection on egg hatch and nuclear background on fecundity and body size, but no interactions between wAlbB infection and nuclear background for any trait. The wAlbB infection was stable at high temperatures and showed perfect maternal transmission and cytoplasmic incompatibility regardless of host background. Our results demonstrate the stability of wAlbB across host backgrounds and point to its long-term effectiveness for controlling arbovirus transmission and mosquito populations.Competing Interest StatementThe authors have declared no competing interest.
A genetically encoded anti-CRISPR protein constrains gene drive spread and prevents population suppression
17565C. Taxiarchi, A. Beaghton, N. I. Don, K. Kyrou, M. Gribble, D. Shittu, S. P. Collins, C. L. Beisel, R. Galizi and A. Crisanti, Nature Communications, 12:3977. 2021-06-25 15:00:05.
CRISPR-based gene drives offer promising means to reduce the burden of pests and vector-borne diseases. These techniques consist of releasing genetically modified organisms carrying CRISPR-Cas nucleases designed to bias their inheritance and rapidly propagate desired modifications. Gene drives can be intended to reduce reproductive capacity of harmful insects or spread anti-pathogen effectors through wild populations, even when these confer fitness disadvantages. Technologies capable of halting the spread of gene drives may prove highly valuable in controlling, counteracting, and even reverting their effect on individual organisms as well as entire populations. Here we show engineering and testing of a genetic approach, based on the germline expression of a phage-derived anti-CRISPR protein (AcrIIA4), able to inactivate CRISPR-based gene drives and restore their inheritance to Mendelian rates in the malaria vector Anopheles gambiae. Modeling predictions and cage testing show that a single release of male mosquitoes carrying the AcrIIA4 protein can block the spread of a highly effective suppressive gene drive preventing population collapse of caged malaria mosquitoes.
A conditional female lethal system for genetic suppression of the global fruit crop pest Drosophila suzukii
F. Li, A. Yamamoto, E. J. Belikoff, A. Berger, E. H. Griffith and M. J. Scott, Pest Management Science, 2021-06-24 14:50:37.Abstract BACKGROUND Drosophila suzukii (Matsumura, 1931, Diptera: Drosophilidae) is a global pest of soft-skinned fruits such as blueberries, cherries and raspberries. Also known as spotted-wing drosophila, D. suzukii is native to Asia but is now widely distributed in the Americas and Europe and presents a serious challenge for growers. Genetic control strategies offer an environmentally-friendly approach for control of D. suzukii. RESULTS In this study we have developed transgenic strains of D. suzukii that carry dominant conditional female lethal transgenes. When raised in the absence of tetracycline, female D. suzukii die. We show that repeated releases of an excess of transgenic males can suppress D. suzukii populations in laboratory cage trials. CONCLUSION Our data suggests that the transgenic strain could provide an effective approach for control of this invasive pest of soft-skinned fruits. This article is protected by copyright. All rights reserved.
A transgenic female killing system for the genetic control of Drosophila suzukii
17506M. F. Schetelig, J. Schwirz and Y. Yan, Scientific Reports, 11:12938. 2021-06-24 14:13:24.
The spotted wing Drosophila (Drosophila suzukii) is an invasive pest of soft-skinned fruit crops. It is rapidly transmitted in Europe and North America, causing widespread agricultural losses. Genetic control strategies such as the sterile insect technique (SIT) have been proposed as environment-friendly and species-restricted approaches for this pest. However, females are inefficient agents in SIT programs. Here we report a conditional female-killing (FK) strategy based on the tetracycline-off system. We assembled sixteen genetic constructs for testing in vitro and in vivo. Twenty-four independent transgenic strains of D. suzukii were generated and tested for female-specific lethality. The strongest FK effect in the absence of tetracycline was achieved by the construct containing D. suzukii nullo promoter for early gene expression, D. suzukii pro-apoptotic gene hidAla4 for lethality, and the transformer gene intron from the Mediterranean fruit fly Ceratitis capitata for female-specific splicing. One strain carrying this construct eliminated 100% of the female offspring during embryogenesis and produced only males. However, homozygous females from these FK strains were not viable on a tetracycline-supplemented diet, possibly due to the basal expression of hidAla4. Potential improvements to the gene constructs and the use of such FK strains in an SIT program are discussed.
Potential use of gene drive modified insects against disease vectors, agricultural pests and invasive species poses new challenges for risk assessment
17551Y. Devos, J. D. Mumford, M. B. Bonsall, A. M. Camargo, L. G. Firbank, D. C. M. Glandorf, F. Nogué, K. Paraskevopoulos and E. A. Wimmer, Critical Reviews in Biotechnology, 2021-06-24 13:49:18.
Potential future application of engineered gene drives (GDs), which bias their own inheritance and can spread genetic modifications in wild target populations, has sparked both enthusiasm and concern. Engineered GDs in insects could potentially be used to address long-standing challenges in control of disease vectors, agricultural pests and invasive species, or help to rescue endangered species, and thus provide important public benefits. However, there are concerns that the deliberate environmental release of GD modified insects may pose different or new harms to animal and human health and the wider environment, and raise novel challenges for risk assessment. Risk assessors, risk managers, developers, potential applicants and other stakeholders at many levels are currently discussing whether there is a need to develop new or additional risk assessment guidance for the environmental release of GD modified organisms, including insects. Developing new or additional guidance that is useful and practical is a challenge, especially at an international level, as risk assessors, risk managers and many other stakeholders have different, often contrasting, opinions and perspectives toward the environmental release of GD modified organisms, and on the adequacy of current risk assessment frameworks for such organisms. Here, we offer recommendations to overcome some of the challenges associated with the potential future development of new or additional risk assessment guidance for GD modified insects and provide considerations on areas where further risk assessment guidance may be required.
Patterns and Mechanisms of Sex Ratio Distortion in the Collaborative Cross Mouse Mapping Population
17511B. A. Haines, F. Barradale and B. L. Dumont, bioRxiv, 2021-06-23 14:26:09.
In species with single-locus chromosome-based mechanisms of sex determination, the laws of segregation predict an equal ratio of females to males at birth. Here, we show that departures from this Mendelian expectation are commonplace in the 8-way recombinant inbred Collaborative Cross (CC) mouse population. More than one-third of CC strains exhibit significant sex ratio distortion (SRD) at wean, with twice as many male-biased than female-biased strains. We show that these pervasive sex biases persist across multiple breeding environments, are stable over time, are not fully mediated by maternal effects, and are not explained by sex-biased neonatal mortality. SRD exhibits a heritable component, but QTL mapping analyses and targeted investigations of sex determination genes fail to nominate any large effect loci. These findings, combined with the reported absence of sex ratio biases in the CC founder strains, suggest that SRD manifests from multilocus combinations of alleles only uncovered in recombined CC genomes. We speculate that the genetic shuffling of eight diverse parental genomes during the early CC breeding generations led to the decoupling of sex-linked drivers from their co-evolved suppressors, unleashing complex, multiallelic systems of sex chromosome drive. Consistent with this interpretation, we show that several CC strains exhibit copy number imbalances at co-evolved X- and Y-linked ampliconic genes that have been previously implicated in germline genetic conflict and SRD in house mice. Overall, our findings reveal the pervasiveness of SRD in the CC population and nominate the CC as a powerful resource for investigating sex chromosome genetic conflict in action.
Selective inheritance of target genes from only one parent of sexually reproduced F1 progeny in Arabidopsis
17567T. Zhang, M. Mudgett, R. Rambabu, B. Abramson, X. Dai, T. P. Michael and Y. Zhao, Nature Communications, 12:3854. 2021-06-22 15:05:16.
Sexual reproduction constrains progeny to inherit allelic genes from both parents. Selective acquisition of target genes from only one parent in the F1 generation of plants has many potential applications including the elimination of undesired alleles and acceleration of trait stacking. CRISPR/Cas9-based gene drives can generate biased transmission of a preferred allele and convert heterozygotes to homozygotes in insects and mice, but similar strategies have not been implementable in plants because of a lack of efficient homology-directed repair (HDR). Here, we place a gene drive, which consists of cassettes that produce Cas9, guide RNAs (gRNA), and fluorescent markers, into the CRYPTOCHROME 1 (CRY1) gene through CRISPR/Cas9-mediated HDR, resulting in cry1drive lines. After crossing the cry1drive/cry1drive lines to wild type, we observe F1 plants which have DNA at the CRY1 locus from only the cry1drive/cry1drive parent. Moreover, a non-autonomous trans-acting gene drive, in which the gene drive unit and the target gene are located on different chromosomes, converts a heterozygous mutation in the target gene to homozygous. Our results demonstrate that homozygous F1 plants can be obtained through zygotic conversion using a CRISPR/Cas9-based gene drive.
Haldane’s duel: intragenomic conflict, selfish Y chromosomes and speciation
17509S. W. Roy, Trends in Genetics, 2021-06-21 14:14:05.
Haldane?s rule, which states that the heterogametic sex (XY or ZW females) fares more poorly in interspecific hybrids, is generally attributed to absence of one of the two species' X/Z chromosomes. However, Haldane?s rule is also observed in mouse placentas despite paternal X silencing. This pattern could reflect Y chromosomes having evolved to promote growth due to maternal?paternal conflict. If so, balanced sex investment arises from a complex intra- and intergenomic duel.
Sustainable Food Production: The Contribution of Genome Editing in Livestock
17590A. Menchaca, Sustainability, 13. 2021-06-21 13:52:55.
This article is focused on the scope and perspectives for the application of this technology, which includes improving production traits, enhancing animal welfare through adaptation and resilience, conferring resistance to infectious diseases, and suppressing pests and invasive species that threaten livestock. The main advantages and concerns that should be overcome by science, policy and people are discussed with the aim that this technology can make a real contribution to our collective future. This review is part of the special issue “Genome Editing in Animal Systems to Support Sustainable Farming and Pest Control”.
Evaluating Flight Performance of Mass-Reared and Irradiated Navel Orangeworm (Lepidoptera: Pyralidae) for Sterile Insect Technique
17545J. Reger, J. A. Wenger, G. Brar, C. Burks and H. Wilson, Journal of Economic Entomology, 2021-06-21 13:28:59.
Navel orangeworm (Pyralidae: Amyelois transitella) is a key pest of almonds and pistachios in California. Moths directly infest nuts which leads to reduced crop yield and quality, and infestation can predispose nuts to fungal pathogens that produce aflatoxins. While several integrated pest management strategies have been developed for A. transitella, studies have recently been initiated to explore the use of sterile insect technique (SIT) as an additional control tool. Mass-rearing, sterilization, and transportation methods originally developed for Pectinophora gossypiella (Lepidoptera: Gelechiidae) are currently being used for production of A. transitella in a mass-rearing facility, but the impacts of these processes on performance of A. transitella remain unclear. In this study, computerized flight mills were used to evaluate multiple flight parameters of mass-reared and irradiated A. transitella males and females relative to non-irradiated mass-reared moths and two strains of locally reared moths which were neither mass-reared nor irradiated. Mass-reared non-irradiated females performed similarly to both strains of locally reared females, flying a mean 9.4–11.8 km per night, whereas mass-reared and irradiated males and mass-reared non-irradiated males all flew shorter distances, in the range of 3.0–6.7 km per night. All of the mass-reared moths compared to locally reared moths had significantly more non-fliers that did not engage in more than two minutes of continuous flight. Findings from this study suggest that mass-rearing conditions reduce A. transitella flight capacity, while irradiation interacts with moths in a sex-specific manner.
Population modification strategies for malaria vector control are uniquely resilient to observed levels of gene drive resistance alleles
17502G. C. Lanzaro, H. M. Sánchez C, T. C. Collier, J. M. Marshall and A. A. James, BioEssays, 2021-06-20 13:46:45.
Cas9/guide RNA (gRNA)-based gene drive systems are expected to play a transformative role in malaria elimination efforts., whether through population modification, in which the drive system contains parasite-refractory genes, or population suppression, in which the drive system induces a severe fitness load resulting in population decline or extinction. DNA sequence polymorphisms representing alternate alleles at gRNA target sites may confer a drive-resistant phenotype in individuals carrying them. Modeling predicts that, for observed levels of SGV at potential target sites and observed rates of de novo DRA formation, population modification strategies are uniquely resilient to DRAs. We conclude that gene drives can succeed when fitness costs incurred by drive-carrying mosquitoes are low enough to prevent strong positive selection for DRAs produced de novo or as part of the SGV and that population modification strategies are less prone to failure due to drive resistance.
Wolbachia-mediated sterility suppresses Aedes aegypti populations in the urban tropics
19443Project Wolbachia-Singapore Consortium, medRxiv, 2021-06-17 21:07:26.
Incompatible insect technique (IIT) via releases of male Wolbachiainfected mosquitoes is a promising tool for dengue control. In a three-year trial in Singaporean high-rise housing estates, we demonstrated that Wolbachia-based IIT dramatically reduces both wildtype Aedes aegypti populations [reductions of 92.7% (95% CI: 84.7%–95.8%) and 98.3% (97.7%–99.8%)] and dengue incidence [reductions of 71% (43%-87%) to 88% (57%-99%)] in the targeted areas. The study highlights the need to ensure adequate vertical distribution of released males in high-rise buildings, address immigration of wildtype females from neighboring areas, and prevent and mitigate stable establishment of Wolbachia in field mosquito populations. Our results demonstrate the potential of Wolbachia-based IIT (supplemented with irradiation, in Singapore’s context) for strengthening dengue control in tropical cities, where dengue burden is the greatest.
Stable high-density and maternally inherited Wolbachia infections in Anopheles moucheti and Anopheles demeilloni mosquitoes
17504T. Walker, S. Quek, C. L. Jeffries, J. Bandibabone, V. Dhokiya, R. Bamou, M. Kristan, L. A. Messenger, A. Gidley, E. A. Hornett, E. R. Anderson, C. Cansado-Utrilla, S. Hegde, C. Bantuzeko, J. C. Stevenson, N. F. Lobo, S. C. Wagstaff, C. A. Nkondjio, S. R., Current Biology, 31:2310. 2021-06-14 13:55:55.
Wolbachia, a widespread bacterium that can reduce pathogen transmission in mosquitoes, has recently been reported to be present in Anopheles (An.) species. In wild populations of the An. gambiae complex, the primary vectors of Plasmodium malaria in Sub-Saharan Africa, Wolbachia DNA sequences at low density and infection frequencies have been detected. As the majority of studies have used highly sensitive nested PCR as the only method of detection, more robust evidence is required to determine whether Wolbachia strains are established as endosymbionts in Anopheles species. Here, we describe high-density Wolbachia infections in geographically diverse populations of An. moucheti and An. demeilloni. Fluorescent in situ hybridization localized a heavy infection in the ovaries of An. moucheti, and maternal transmission was observed. Genome sequencing of both Wolbachia strains obtained genome depths and coverages comparable to those of other known infections. Notably, homologs of cytoplasmic incompatibility factor (cif) genes were present, indicating that these strains possess the capacity to induce the cytoplasmic incompatibility phenotype, which allows Wolbachia to spread through host populations. These strains should be further investigated as candidates for use in Wolbachia biocontrol strategies in Anopheles aiming to reduce the transmission of malaria.
Temperature-Inducible Precision Guided Sterile Insect Technique
17447N. P. Kandul, J. Liu and O. S. Akbari, bioRxiv, 2021.06.14.448312. 2021-06-14 13:13:08.
Releases of sterile males are the gold standard for many insect population control programs, and precise sex sorting to remove females prior to male releases is essential to the success of these operations. To advance traditional methods for scaling the generation of sterile males, we previously described a CRISPR-mediated precision-guided sterile insect technique (pgSIT), in which Cas9 and gRNA strains are genetically crossed to generate sterile males for release. While effective at generating F1 sterile males, pgSIT requires a genetic cross between the two parental strains which requires maintenance and sexing of two strains in a factory. Therefore, to further advance pgSIT by removing this crossing step, here we describe a next-generation Temperature-Inducible pgSIT (TI-pgSIT) technology and demonstrate its proof-of-concept in Drosophila melanogaster. Importantly, we were able to develop a true-breeding strain for TI-pgSIT that eliminates the requirement for sex sorting, a feature that may help further automate production at scale.
Area-Wide Integrated Management of a Glossina palpalis gambiensis Population from the Niayes Area Of Senegal: A review of operational research in support of a phased conditional approach
25657M. J. B. Vreysen, M. T. Seck, B. Sall, A. G. Mbaye, M. Bassene, A. G. Fall, M. Lo and J. Bouyer, AREA-WIDE INTEGRATED PEST MANAGEMENT:, 2021-06-12 13:45:08.
In 2005, the Government of Senegal initiated a project entitled "Projet de lutte contre les glossines dans les Niayes" (Tsetse control project in the Niayes) with the aim of creating a zone free of Glossina palpalis gambiensis in that area. The project received technical and financial support from the International Atomic Energy Agency (IAEA), the Food and Agriculture Organization of the United Nations (FAO), the Centre de Cooperation Internationale en Recherche Agronomique pour le Developpement (CIRAD) and the US Department of State through the Peaceful Uses Initiative (PUI). It was implemented in the context of the Pan African Tsetse and Trypanosomosis Eradication Campaign (PATTEC) following a phased conditional approach (PCA) that entails implementation in distinct phases, in which support to the next phase is conditional upon completion of all (or at least the majority of) activities in the previous phase. In the case of the tsetse project in Senegal, the PCA consisted of 4 phases: (1) commitment of all stakeholders and training, (2) baseline data collection, feasibility studies and strategy development, (3) preparatory pre-operational activities and (4) operational activities. This paper provides an overview of the main activities that were carried out within each phase, with emphasis on the operational research carried out in phases 2 and 3, that was instrumental in guiding the project's decision-making. Activities of phase 2 focused on the collection of entomological, veterinary, socio-economic and environmental baseline data, and a population genetics study that proved the isolated character of the G. p. gambiensis population of the Niayes. These data enabled the tsetse-infested area to be delimited to 1000 km(2), the impact of animal trypanosomosis on the farmers' welfare to be quantified (annual benefits of 2 million Euro in the tsetse-infested zone), and the formulation of an area-wide integrated pest management (AW-IPM) strategy that included a sterile insect (SIT) component to eradicate the isolated tsetse populations from the Niayes. In view of the extreme fragmentation of the remaining favourable habitat of the Niayes and the high human population density (peri-urban area), which excluded the possibility of using the Sequential Aerosol Technique, the IPM strategy that was selected comprised the suppression of the tsetse population with insecticide-impregnated traps/targets and the use of "pour-on" for cattle, followed by the release of sterile males to eliminate the remaining relic pockets. During phase 3, the pre-operational phase, a series of activities were carried out that were needed to implement the operational phase. These included the establishment of a colony of tsetse originating from the target area in Senegal, competitiveness studies between the sterile flies and those from the target area, development of transport methods for long-distance shipments of sterile male pupae, competitiveness of the sterile male flies after release in the target area, development of aerial release methods (including a new chilled adult release system) and development of a Maxent-based distribution model to guide the suppression, sterile male releases and monitoring of the eradication campaign. To be able to properly manage the eradication campaign in different phases, the entire target area was divided into 3 operational blocks. This paper demonstrates how, during the operational phase, scientific principles continued to guide the implementation process. The results to date are encouraging, i.e. the deployment of 269 insecticide-impregnated Vavoua traps in favourable habitat of Block 1 reduced the apparent density of the G. p. gambiensis population significantly (from 0.42 (SD 0.39) to 0.04 (SD 0.11) flies/trap/day). This was followed by the aerial release of sterile males that reduced the apparent density to zero after six months of releases. The last wild fly was trapped on August 9, 2012 in Block 1. In Block 2, during the suppression, the apparent fly density dropped from 1.24 (SD 1.23) to 0.005 (SD 0.017) flies/trap/day. Sterile male releases were initiated in February 2014 and expanded to cover the entire Block 2 in January 2015. The apparent fly density has so far been reduced to < 0.001 fly per trap per day until the end of 2018 and releases are still ongoing. The results of the campaign are discussed with respect to the "adaptive management approach" used, which was deemed critical for the success of the campaign.
Using Wolbachia to Eliminate Dengue: Will the Virus Fight Back?
18270M. Edenborough Kathryn, A. Flores Heather, P. Simmons Cameron, E. Fraser Johanna and C. Pierson Ted, Journal of Virology, 95:e02203-20. 2021-06-10 14:06:11.
Recent fieldtrials havedemonstratedthatdengue incidence can besubstantially reduced by introgressing strains of the endosymbiotic bacterium Wolbachia into Aedes aegypti mosquito populations. This strategy relies on Wolbachia reducing the susceptibility of Ae. aegypti to disseminated infection by positive-sense RNA viruses like dengue. However, RNA viruses are well known to adapt to antiviral pressures. Here, we review the viral infection stages where selection for Wolbachia-resistant virus variants could occur. We also consider the genetic constraints imposed on viruses that alternate between vertebrate and invertebrate hosts, and the likely selection pressures to which dengue virus might adapt in order to be effectively transmitted by Ae. aegypti that carry Wolbachia. While there are hurdles to dengue viruses developing resistance to Wolbachia, we suggest that long-term surveillance for resistant viruses should be an integral component of Wolbachia-introgression biocontrol programs.
Efficacy of Wolbachia-Infected Mosquito Deployments for the Control of Dengue
17300A. Utarini, C. Indriani, R. A. Ahmad, W. Tantowijoyo, E. Arguni, M. R. Ansari, E. Supriyati, D. S. Wardana, Y. Meitika, I. Ernesia, I. Nurhayati, E. Prabowo, B. Andari, B. R. Green, L. Hodgson, Z. Cutcher, E. Rancès, P. A. Ryan, S. L. O’Neill, S. M. Dufau, New England Journal of Medicine, 384:2177-2186. 2021-06-10 11:05:41.
BACKGROUND Aedes aegypti mosquitoes infected with the wMel strain of Wolbachia pipientis are less susceptible than wild-type A. aegypti to dengue virus infection. METHODS We conducted a cluster-randomized trial involving releases of wMel-infected A. aegypti mosquitoes for the control of dengue in Yogyakarta, Indonesia. We randomly assigned 12 geographic clusters to receive deployments of wMel-infected A. aegypti (intervention clusters) and 12 clusters to receive no deployments (control clusters). All clusters practiced local mosquito-control measures as usual. A test-negative design was used to assess the efficacy of the intervention. Patients with acute undifferentiated fever who presented to local primary care clinics and were 3 to 45 years of age were recruited. Laboratory testing was used to identify participants who had virologically confirmed dengue (VCD) and those who were test-negative controls. The primary end point was symptomatic VCD of any severity caused by any dengue virus serotype. RESULTS After successful introgression of wMel into the intervention clusters, 8144 participants were enrolled; 3721 lived in intervention clusters, and 4423 lived in control clusters. In the intention-to-treat analysis, VCD occurred in 67 of 2905 participants (2.3%) in the intervention clusters and in 318 of 3401 (9.4%) in the control clusters (aggregate odds ratio for VCD, 0.23; 95% confidence interval [CI], 0.15 to 0.35; P=0.004). The protective efficacy of the intervention was 77.1% (95% CI, 65.3 to 84.9) and was similar against the four dengue virus serotypes. The incidence of hospitalization for VCD was lower among participants who lived in intervention clusters (13 of 2905 participants [0.4%]) than among those who lived in control clusters (102 of 3401 [3.0%]) (protective efficacy, 86.2%; 95% CI, 66.2 to 94.3).
Fine-scale estimation of key life-history parameters of malaria vectors: implications for next-generation vector control technologies
17329A. L. Morris, A. Ghani and N. Ferguson, Parasites and Vectors, 14:311. 2021-06-08 14:48:41.
Mosquito control has the potential to significantly reduce malaria burden on a region, but to influence public health policy must also show cost-effectiveness. Gaps in our knowledge of mosquito population dynamics mean that mathematical modelling of vector control interventions have typically made simplifying assumptions about key aspects of mosquito ecology. Often, these assumptions can distort the predicted efficacy of vector control, particularly next-generation tools such as gene drive, which are highly sensitive to local mosquito dynamics.
Vector control: Discovery of Wolbachia in malaria vectors
17554P. A. Ross and A. A. Hoffmann, Current Biology, 31:R738-R740. 2021-06-07 14:28:04.
Wolbachia bacteria are being widely released for suppression of dengue transmitted by Aedes mosquitoes. Walker, Quek, Jeffries and colleagues present robust evidence for natural Wolbachia infections in malaria-vectoring Anopheles mosquitoes, paving the way for new Wolbachia-based intervention
Sequence of the supernumerary B chromosome of maize provides insight into its drive mechanism and evolution
17202N. Blavet, H. Yang, H. Su, P. Solanský, R. N. Douglas, M. Karafiátová, L. Šimková, J. Zhang, Y. Liu, J. Hou, X. Shi, C. Chen, M. El-Walid, M. E. McCaw, P. S. Albert, Z. Gao, C. Zhao, G. Ben-Zvi, L. Glick, G. Kol, J. Shi, J. Vrána, H. Šimková, J. C. Lamb,, Proceedings of the National Academy of Sciences, 118:e2104254118. 2021-06-05 15:32:03.
B chromosomes are nonvital chromosomes found in thousands of plants and animals that persist through various drive mechanisms. The drive mechanism of the maize B chromosome consists of mitotic nondisjunction at the second pollen division to produce two unequal sperm and then the sperm with the B chromosomes preferentially fertilizes the egg in double fertilization. A high-quality sequence of the maize B chromosome together with genetic analysis reveals the cis factor for nondisjunction is a B chromosome-specific repeat interspersed in and around the centromere. The gene and transposable element content of the B chromosome and relaxed purifying selection of transposed protein-encoding genes suggest that the chromosome has been present in the evolutionary lineage for millions of years.B chromosomes are enigmatic elements in thousands of plant and animal genomes that persist in populations despite being nonessential. They circumvent the laws of Mendelian inheritance but the molecular mechanisms underlying this behavior remain unknown. Here we present the sequence, annotation, and analysis of the maize B chromosome providing insight into its drive mechanism. The sequence assembly reveals detailed locations of the elements involved with the cis and trans functions of its drive mechanism, consisting of nondisjunction at the second pollen mitosis and preferential fertilization of the egg by the B-containing sperm. We identified 758 protein-coding genes in 125.9 Mb of B chromosome sequence, of which at least 88 are expressed. Our results demonstrate that transposable elements in the B chromosome are shared with the standard A chromosome set but multiple lines of evidence fail to detect a syntenic genic region in the A chromosomes, suggesting a distant origin. The current gene content is a result of continuous transfer from the A chromosomal complement over an extended evolutionary time with subsequent degradation but with selection for maintenance of this nonvital chromosome.Raw data used for sequence assembly of B73 line possessing B chromosome(s) and additional RNA-seq data used for B chromosome annotation are available in NCBI-SRA as BioProject PRJNA633287. Sequence reads for B-deficiency-carrying lines of maize are available in NCBI-SRA as BioProject PRJNA634743. Input-seq data are available in NCBI-GEO under accession GSE152074. The final B chromosome sequence and its annotation are available at MaizeGDB (https://www.maizegdb.org/) under the name Zm-B73_B-CHROMOSOME-MBSC-1.0 with the identifier Zm00044a.
Nondisjunction and unequal spindle organization accompany the drive of Aegilops speltoides B chromosomes
17169D. Wu, A. Ruban, J. Fuchs, J. Macas, P. Novák, M. Vaio, Y. Zhou and A. Houben, New Phytologist, 223:1340-1352. 2021-06-04 19:45:39.
B chromosomes (Bs) are supernumerary chromosomes, which are often preferentially inherited. When transmission rates of chromosomes are higher than 0.5, not obeying the Mendelian law of equal segregation, the resulting transmission advantage is collectively referred to as ?chromosome drive?. Here we analysed the drive mechanism of Aegilops speltoides Bs. The repeat AesTR-183 of A. speltoides Bs, which also can be detected on the Bs of Aegilops mutica and rye, was used to track Bs during pollen development. Nondisjunction of CENH3-positive, tubulin interacting B sister chromatids and an asymmetric spindle during first pollen grain mitosis are key for the accumulation process. A quantitative flow cytometric approach revealed that, independent of the number of Bs present in the mother plant, Bs accumulate in the generative nuclei to > 93%. Nine out of 11 tested (peri)centromeric repeats were shared by A and B chromosomes. Our findings provide new insights into the process of chromosome drive. Quantitative flow cytometry is a useful and reliable method to study the drive frequency of Bs. Nondisjunction and unequal spindle organization accompany during first pollen mitosis the drive of A. speltoides Bs. The prerequisites for the drive process seems to be common in Poaceae.
What is wrong in extinguishing a species? Charting the Ethical Challenges of using Gene-Drive Technologies to eradicate A. gambiae vector populations
17161M. Annoni and T. Pievani, Biolaw Journal-Rivista Di Biodiritto, 2021-05-31 19:04:09.
This article analyses three ethical arguments against the use of gene-drive technologies to control for, and possibly extinguish, a particular species of vector mosquitoes (Anopheles gambiae) causing the malaria infection. We conclude that none of these arguments is truly persuasive in the specific case and, therefore, that using gene-drive technologies to suppress or eradicate the population of Anopheles gambiae could be ethically justifiable provided certain cautions referring to ecological consequences, evolutionary effects and social engagement of local communities.
Analysis of off-target effects in CRISPR-based gene drives in the human malaria mosquito
17141W. T. Garrood, N. Kranjc, K. Petri, D. Y. Kim, J. A. Guo, A. M. Hammond, I. Morianou, V. Pattanayak, J. K. Joung, A. Crisanti and A. Simoni, Proceedings of the National Academy of Sciences, 118:e2004838117. 2021-05-31 13:24:14.
CRISPR-Cas9 nuclease-based gene drives have been developed toward the aim of control of the human malaria vector Anopheles gambiae. Gene drives are based on an active source of Cas9 nuclease in the germline that promotes super-Mendelian inheritance of the transgene by homology-directed repair (“homing”). Understanding whether CRISPR-induced off-target mutations are generated in Anopheles mosquitoes is an important aspect of risk assessment before any potential field release of this technology. We compared the frequencies and the propensity of off-target events to occur in four different gene-drive strains, including a deliberately promiscuous set-up, using a nongermline restricted promoter for SpCas9 and a guide RNA with many closely related sites (two or more mismatches) across the mosquito genome. Under this scenario we observed off-target mutations at frequencies no greater than 1.42%. We witnessed no evidence that CRISPR-induced off-target mutations were able to accumulate (or drive) in a mosquito population, despite multiple generations’ exposure to the CRISPR-Cas9 nuclease construct. Furthermore, judicious design of the guide RNA used for homing of the CRISPR construct, combined with tight temporal constriction of Cas9 expression to the germline, rendered off-target mutations undetectable. The findings of this study represent an important milestone for the understanding and managing of CRISPR-Cas9 specificity in mosquitoes, and demonstrates that CRISPR off-target editing in the context of a mosquito gene drive can be reduced to minimal levels.All raw amplicon sequencing files have been deposited in the National Center for Biotechnology Information (NCBI) BioProject (accession code PRJNA665154).
Experimental demonstration of tethered gene drive systems for confined population modification or suppression
17153M. Metzloff, E. Yang, S. Dhole, A. G. Clark, P. W. Messer and J. Champer, bioRxiv, 2021.05.29.446308. 2021-05-30 10:50:23.
Tethered drive systems, in which a locally confined gene drive provides the CRISPR nuclease needed for a homing drive, could provide a solution to this problem, offering the power of a homing drive and confinement of the supporting drive. Here, we demonstrate the engineering of a tethered drive system in Drosophila, using a TARE drive to support modification and suppression homing drives. Each drive was able to bias inheritance in its favor, and the TARE drive was shown to spread only when released above a threshold frequency in experimental cage populations. After the TARE drive had established in the population, it facilitated the spread of a subsequently released split homing modification drive (to all individuals in the cage) and of a homing suppression drive (to its equilibrium frequency). Our results show that the tethered drive strategy is a viable and easily engineered option for providing confinement of homing drives to target populations. Competing Interest StatementThe authors have declared no competing interest.
First evidence of deviation from Mendelian proportions in a conservation programme
17556C. E. Grueber, K. A. Farquharson, B. R. Wright, G. P. Wallis, C. J. Hogg and K. Belov, Molecular Ecology, 13. 2021-05-29 14:38:25.
Classic Mendelian inheritance is the bedrock of population genetics and underpins pedigree-based management of animal populations. However, assumptions of Mendelian inheritance might not be upheld in conservation breeding programmes if early viability selection occurs, even when efforts are made to equalise genetic contributions of breeders. To test this possibility, we investigated deviations from Mendelian proportions in a captive metapopulation of the endangered Tasmanian devil. This marsupial population is ideal for addressing evolutionary questions in conservation due to its large size, range of enclosure types (varying in environmental conditions), good genomic resources (which aid interpretation), and the species' biology. Devil mothers give birth to more offspring than they can nurse in the pouch, providing the potential for intense viability selection amongst embryos. We used data from 140 known sire-dam-offspring triads to isolate within-family selection from population-level mechanisms (such as mate choice or inbreeding), and compared observed offspring genotypes at 123 targeted SNPs to neutral (i.e., Mendelian) expectations. We found lower offspring heterozygosity than expected, and subtle patterns that varied across a gradient of management intensity from zoo-like enclosures to semi-wild environments for some loci. Meiotic drive or maternal-foetal incompatibilities are consistent with our results, although we cannot statistically confirm these mechanisms. We found some evidence that maternal genotype affects annual litter size, suggesting that family-level patterns are driven by differential offspring mortality before birth or during early development. Our results show that deviations from Mendelian inheritance can occur in conservation programmes, despite best-practice management to prevent selection.
Improving mosquito control strategies with population genomics
18781T. L. Schmidt, N. M. Endersby-Harshman and A. A. Hoffmann, Trends in Parasitology, 37:907-921. 2021-05-29 12:41:38.
Mosquito control strategies increasingly apply knowledge from population genomics research. This review highlights recent applications to three research domains: mosquito invasions, insecticide resistance evolution, and rear and release programs. Current research trends follow developments in reference assemblies, either as improvements to existing assemblies (particularly Aedes) or assemblies for new taxa (particularly Anopheles). With improved assemblies, studies of invasive and rear and release target populations are better able to incorporate adaptive as well as demographic hypotheses. New reference assemblies are aiding comparisons of insecticide resistance across sister taxa while helping resolve taxon boundaries amidst frequent introgression. Anopheles gene drive deployments and improved Aedes genome assemblies should lead to a convergence in research aims for Anopheles and Aedes in the coming years.
A Novel Genetic Sexing Strain of Anastrepha Ludens for Cost-Effective Sterile Insect Technique Applications: Improved Genetic Stability and Rearing Efficiency
17125E. Ramírez-Santos, P. Rendon, G. Gouvi, A. Zacharopoulou, K. Bourtzis, C. Cáceres and K. Bloem, Insects, 12. 2021-05-27 19:00:07.
Anastrepha ludens (Loew) is one of the most destructive insect pests damaging several fruits of economic importance. The sterile insect technique (SIT) is used under an area-wide integrated pest management approach, to suppress these pest populations. Mass rearing facilities were initially established to produce sterile males of bi-sexual strains in support of SIT. The first genetic sexing strain (GSS) for A. ludens, Tapachula-7, based on pupal color dimorphism, was a key development since the release of males-only significantly increases the SIT efficiency. In this study, we document the development of a novel pupal color-based GSS. Twelve radiation-induced translocation lines were assessed as potential GSS in terms of recombination rates and rearing efficiency at a small scale. The best one, GUA10, was cytogenetically characterized: it was shown to carry a single translocation between the Y chromosome and chromosome 2, which is known to carry the black pupae marker. This GSS was further evaluated at medium and large scales regarding its genetic stability, productivity and quality versus Tapachula-7. GUA10 presented better genetic stability, fecundity, fertility, production efficiency, flying ability, and male mating, clear indicators that GUA10 GSS can significantly improve the efficacy and cost-effectiveness of SIT applications against this pest species.
Suppression of female fertility in Aedes aegypti with a CRISPR-targeted male-sterile mutation
17112J. Chen, J. Luo, Y. Wang, A. S. Gurav, M. Li, O. S. Akbari and C. Montell, Proceedings of the National Academy of Sciences, 118:e2105075118. 2021-05-26 18:33:06.
We introduced and characterized the impact on female fertility of an Ae. aegypti mutation that disrupts a gene that is specifically expressed in testes. We used CRISPR/Cas9 to generate a null mutation in the Ae. aegypti β2-tubulin (B2t) gene, which eliminates male fertility. When we allowed wild-type females to first mate with B2t mutant males, most of the females did not produce progeny even after being subsequently exposed to wild-type males. We also introduced B2t mutant and wild-type males simultaneously with wild-type females and found that a larger number of B2t mutant males relative to the wild-type males was effective in significantly suppressing female fertility. These results raise the possibility of employing B2t sterile males to improve the efficacy of SIT in suppressing populations of Ae. aegypti through repeated releases and thereby reduce the transmission of viruses by these invasive mosquitoes.All study data are included in the main text.
The origin of island populations of the African malaria mosquito, Anopheles coluzzii
17287M. Campos, M. Hanemaaijer, H. Gripkey, T. C. Collier, Y. S. Lee, A. J. Cornel, J. Pinto, D. Ayala, H. Rompao and G. C. Lanzaro, Communications Biology, 4:9. 2021-05-26 10:26:11.
Anopheles coluzzii is a major malaria vector throughout its distribution in west-central Africa. Here we present a whole-genome study of 142 specimens from nine countries in continental Africa and three islands in the Gulf of Guinea. This sample set covers a large part of this species' geographic range. Our population genomic analyses included a description of the structure of mainland populations, island populations, and connectivity between them. Three genetic clusters are identified among mainland populations and genetic distances (F-ST) fits an isolation-by-distance model. Genomic analyses are applied to estimate the demographic history and ancestry for each island. Taken together with the unique biogeography and history of human occupation for each island, they present a coherent explanation underlying levels of genetic isolation between mainland and island populations. We discuss the relationship of our findings to the suitability of Sao Tome and Principe islands as candidate sites for potential field trials of genetic-based malaria control strategies. Campos, Lanzaro and colleagues use whole-genome sequencing and population genomic analyses to infer connectivity between mainland and island mosquito populations in West Africa. The unique biogeographic history for each island population is reported, and the findings highlight potential candidate sites for genetic-based malaria control strategies.
