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 Drives as Interventions into Nature: the Coproduction of Ontology and Morality in the Gene Drive Debate
25114K. Boersma, B. Bovenkerk and D. Ludwig, NanoEthics, 17:4. 2023-04-22 10:10:41.
Gene drives are potentially ontologically and morally disruptive technologies. The potential to shape evolutionary processes and to eradicate (e.g. malaria-transmitting or invasive) populations raises ontological questions about evolution, nature, and wilderness. The transformative promises and perils of gene drives also raise pressing ethical and political concerns. The aim of this article is to arrive at a better understanding of the gene drive debate by analysing how ontological and moral assumptions are coproduced in this debate. Combining philosophical analysis with a critical reading of the gene drive literature and an ethnographic study of two leading research groups, the article explores the hypothesis that the development of and debate about gene drives are characterized by a particular intervention-oriented mode of coproduction. Based on the results of this exploration, we highlight the need for a broadening of the perspective on gene drives in which empirical, moral, and ontological concerns are addressed explicitly in their interplay rather than in (disciplinary) isolation from each other.
The Role of Symbiont-Targeted Strategies in the Management of Pentatomidae and Tephritidae Pests under an Integrated Vision
24996E. Gonella and A. Alma, Agronomy, 13. 2023-04-21 15:11:32.
The interaction between insects and gut bacterial symbionts is, nowadays, regarded as an important element in the implementation of pest management, in consideration of the urgent need for sustainable alternatives to insecticide use. In this framework, a major tool is symbiotic control, with the main ready-to-use application represented by the interruption of obligate symbioses. Two insect families, namely Pentatomidae in the Hemiptera order and Tephritidae in Diptera, have been indicated as outstanding targets for symbiont-oriented control tactics. An important advantage of interrupting obligate symbioses is the target shift from insect to bacteria, which avoids insecticide use; however, the compatibility between this approach and other pest/disease management strategies is crucial to design low-impact pest control programs. Here we present the state of knowledge regarding the integration of symbiont manipulation in sustainable plant protection plans. Research assessing the potential for multitarget applications is reported, as well as studies on the impact of symbiont interruption on nontarget species. Besides symbiont-targeted pest control, another relevant outcome of symbiont manipulation is the restoration of microbial perturbation in mass-reared insects used in pest control programs, which is a required step to allow the success of other tactics, such as the Sterile Insect Technique. Despite the potential contribution that symbiont-targeted strategies may offer to integrated pest management, we point out that operational caveats may emerge in symbiont-oriented control in relation to the target extension on the label directions and to the number of required treatments. Future work is needed to increase the target range and the number of tested formulations exploiting the interruption of bacterial symbioses. This will also require assessment of the effect of different products on beneficial organisms, including biological control agents. Finally, the authorization of formulates for symbiotic control should be taken into consideration by the regulatory bodies, to really promote new readily available control options. © 2023 by the authors.
Standardization of sterile insect technique (SIT) for melon fruit fly
25144M. Math and Y. K. Kotikal, Journal of Entomological Research, 47:16-20. 2023-04-21 14:26:32.
Sterile technique was standardized for melon fly, Zeugodacus cucurbitae Coquillett under laboratory conditions. The Adult emergence of B. cucurbitae was significantly highest (96.12%) without radiation to the pupae. Among different radiation doses, 30 Gy resulted in significantly the highest adult emergence (64.12%). Significantly the highest deformed adults were observed at 90 Gy (10.61%) while, it was lowest without radiation (1.33%). When mature pupae were exposed to 30, 50, 70 and 90 Gy the male longevity was 32.37, 26.55, 24.57 and 20.63 days, respectively, as against 34.98 days for untreated males. At 90 Gy mating competitive values recorded was 5.66 and 8.29 with a treated male: Untreated male: Untreated female cross ratio of 2:1:1 and 1:2:1, respectively. Significantly the highest per cent sterility of Z. cucurbitae was recorded at 90 Gy with a cross ratio of 2:1:1 and 1:1:2 (68.36 and 55.28%, respectively). © 2023, Journal of Entomological Research. All Rights Reserved.
Targeting Sex Determination to Suppress Mosquito Populations
24988L. Ming, P. K. Nikolay, S. Ruichen, Y. Ting, D. B. Elena, J. B. Daniel, A. Igor, M. S. C. Hector, Z. Yinpeng, A. D. Nicolas, M. L. YuMin, P. S. Matthew, M. Craig, M. M. John and S. A. Omar, bioRxiv, 2023.04.18.537404. 2023-04-20 14:45:03.
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 control wild populations, safely curtailing disease transmission.Competing Interest StatementO.S.A. is a founder of Agragene, Inc. with equity interest. O.S.A., M.L., and N.P.K are founders of Synvect with equity interest. The terms of this arrangement have been reviewed and approved by the University of California, San Diego in accordance with its conflict of interest policies. All remaining authors declare no competing interests.
Evaluating the Effect of Irradiation on the Densities of Two RNA Viruses in Glossina morsitans morsitans
25137C. K. Mirieri, A. M. M. Abd-Alla, V. I. D. Ros and M. M. van Oers, Insects, 14. 2023-04-20 14:11:24.
Tsetse flies are cyclic vectors of Trypanosoma parasites, which cause debilitating diseases in humans and animals. To decrease the disease burden, the number of flies is reduced using the sterile insect technique (SIT), where male flies are sterilized through irradiation and released into the field. This procedure requires the mass rearing of high-quality male flies able to compete with wild male flies for mating with wild females. Recently, two RNA viruses, an iflavirus and a negevirus, were discovered in mass-reared Glossina morsitans morsitans and named GmmIV and GmmNegeV, respectively. The aim of this study was to evaluate whether the densities of these viruses in tsetse flies are affected by the irradiation treatment. Therefore, we exposed tsetse pupae to various doses (0–150 Gy) of ionizing radiation, either in air (normoxia) or without air (hypoxia), for which oxygen was displaced by nitrogen. Pupae and/or emerging flies were collected immediately afterwards, and at three days post irradiation, virus densities were quantified through RT-qPCR. Generally, the results show that irradiation exposure had no significant impact on the densities of GmmIV and GmmNegeV, suggesting that the viruses are relatively radiation-resistant, even at higher doses. However, sampling over a longer period after irradiation would be needed to verify that densities of these insect viruses are not changed by the sterilisation treatment. © 2023 by the authors.
Defining transformation events for gene drive in species complexes
25001J. B. Connolly, IOBC-WPRS Bulletin, 163:8-20. 2023-04-13 15:11:40.
Engineered gene drives (EGDs) that allow the super-Mendelian inheritance of genetic traits could one day be used to reduce the vectorial capacity of Anopheles species that transmit human malaria in Africa. Many Anopheles species belong to complexes of closely related sibling species that can produce fertile interspecific hybrid females. In cases where the genomic target locus of the EGD is conserved amongst sibling species from the same complex as the released target species, it would therefore be plausible that the EGD could be vertically transmitted from the target species to sibling species by interspecific mating. To differentiate genetically modified organisms, the term ‘transformation event’ is used, based on the specific genomic location of the transgenic construct, as a result of random genomic integration. In contrast, an EGD is generated via its precise and reproducible insertion in its specific genomic target locus. These considerations pose two key questions for the use of EGD in species complexes: (i) what does the definition of “transformation event” mean in the context of vertical gene drive transfer of the EGD to sibling species in species complexes and (ii) does de novo transformation of an EGD into a sibling species constitute the same transformation event as introgression by backcrossing into a sibling species of an EGD that had been originally transformed in the target species? While definitions of the term transformation event that have been provided by national and intergovernmental organisations are somewhat ambiguous, they do provide scope for broad interpretation of vertical gene drive transfer of a specific EGD to different sibling species of the target species as the same transformation event. There also appears to be some consensus that definitions of transformation event support the notion that de novo transformations of an EGD in sibling species constitute the same transformation events as introgression by backcrossing into sibling species of an EGD that had been originally inserted in the target species.
Dengue Exposure and Wolbachia wMel Strain Affects the Fertility of Quiescent Eggs of Aedes aegypti
25102M. T. Petersen, D. Couto-Lima, G. A. Garcia, M. G. Pavan, M. R. David and R. Maciel-de-Freitas, Viruses, 15. 2023-04-12 10:20:54.
(1) Background: The deployment of the bacterium Wolbachia to reduce arbovirus transmission is ongoing in several countries worldwide. When Wolbachia-carrying Aedes aegypti are released and established in the field, females may feed on dengue-infected hosts. The effects of simultaneous exposure on life-history traits of Ae. aegypti to Wolbachia wMel strain and dengue-1 virus DENV-1 remain unclear. (2) Methods: We monitored 4 groups (mosquitoes with either DENV-1 or Wolbachia, coinfected with DENV-1 and Wolbachia, as well as negative controls) to estimate Ae. aegypti survival, oviposition success, fecundity, collapsing and fertility of quiescent eggs for 12 weeks. (3) Results: Neither DENV-1 nor Wolbachia had a significant impact on mosquito survival nor on mosquito fecundity, although the last parameter showed a tendency to decrease with ageing. There was a significant decrease in oviposition success in individuals carrying Wolbachia. Wolbachia infection and storage time significantly increased egg collapse parameter on the egg viability assay, while DENV-1 had a slight protective effect on the first four weeks of storage. (4) Conclusions: Despite limitations, our results contribute to better understanding of the tripartite interaction of virus, bacteria and mosquito that may take place in field conditions and aid in guaranteeing the Wolbachia strategy success.
Spatial Distribution and Long-Term Persistence of Wolbachia-Infected Aedes aegypti in the Mentari Court, Malaysia
25100Y. L. Cheong, W. A. Nazni, H. L. Lee, A. NoorAfizah, I. C. MohdKhairuddin, G. M. R. Kamarul, N. M. N. Nizam, M. A. K. Arif, Z. M. NurZatilAqmar, S. M. Irwan, K. Khadijah, Y. M. Paid, O. Topek, A. H. Hasnor, R. AbuBakar, B. Singh Gill, K. Fadzilah, A. Tahi, Insects, 14. 2023-04-11 10:16:44.
Dengue is endemic in Malaysia, and vector control strategies are vital to reduce dengue transmission. The Wolbachia strain wAlbB carried by both sexes of Ae. aegypti was released in Mentari Court, a high-rise residential site, in October 2017 and stopped after 20 weeks. Wolbachia frequencies are still being monitored at multiple traps across this site, providing an opportunity to examine the spatiotemporal distribution of Wolbachia and mosquito density with respect to year, residential block, and floor, using spatial interpolation in ArcGIS, GLMs, and contingency analyses. In just 12 weeks, Wolbachia-infected mosquitoes were established right across the Mentari Court site with an overall infection frequency of >90%. To date, the Wolbachia frequency of Ae. aegypti has remained high in all areas across the site despite releases finishing four years ago. Nevertheless, the Wolbachia invaded more rapidly in some residential blocks than others, and also showed a relatively higher frequency on the eighth floor. The Ae. aegypti index tended to differ somewhat between residential blocks, whilst the Ae. albopictus index was relatively higher at the top and bottom floors of buildings. In Mentari Court, only a short release period was required to infiltrate Wolbachia completely and stably into the natural population. The results inform future releases in comparable sites in a dengue control programme.
DIPA-CRISPR gene editing in the yellow fever mosquito <em>Aedes aegypti</em> (Diptera: Culicidae)
24960S. Yu, T. Momoyo, O. Manabu, K. Hirotaka and D. Takaaki, bioRxiv, 2023.04.07.535996. 2023-04-07 10:28:04.
Current methods for gene editing in insects rely on embryonic microinjection, which can be challenging for non-specialist laboratories. Recently, an alternative method known as direct parental CRISPR (DIPA-CRISPR) was developed. This method involves injecting commercial Cas9 protein and single-guide RNA into adult females, which can efficiently introduce mutations into developing oocytes. However, its versatility has not been fully explored, particularly in insects that have the most derived, polytrophic meroistic ovaries. In this study, we successfully applied DIPA-CRISPR to the yellow fever mosquito Aedes aegypti, which has polytrophic meroistic ovaries. Following adult injection of Cas9 ribonucleoproteins (Cas9 RNPs) targeting the kynurenine 3-monooxygenase gene, we recovered gene-edited G0 individuals. Injection at 24 h after blood-feeding resulted in the highest gene editing efficiency (3.5%), confirming that a key parameter of DIPA-CRISPR is the stage in which the adult females are injected. Together with our previous study, we demonstrated that DIPA-CRISPR is applicable to all three types of insect ovaries (i.e., panoistic, telotrophic, and polytrophic), which indicates that DIPA-CRISPR is a generalizable approach for insect gene editing.Competing Interest StatementThe authors have declared no competing interest.
A bacterium against the tiger: further evidence of the potential of non-inundative releases of males with manipulated Wolbachia infection in reducing fertility of Aedes albopictus field populations in Italy
24966B. Caputo, R. Moretti, C. Virgillito, M. Manica, E. Lampazzi, G. Lombardi, P. Serini, V. Pichler, N. W. Beebe, A. Della Torre and M. Calvitti, Pest Management Science, 2023-04-06 10:42:30.
BACKGROUND: Incompatible Insect Technique (IIT) is a population suppression approach based on the release of males with manipulated Wolbachia infection inducing egg inviability in wild females. We here present results of multiple field releases of incompatible ARwP males carried out in 2019 in a 2.7-ha green area within urban Rome (Italy) to assess the effect on Ae. albopictus egg viability. Data are compared with results obtained in 2018, when the approach was tested for the first time in Europe. RESULTS: An average of 4,674 ARwP males were released weekly for 7 weeks, resulting in a mean ARwP/wild male ratio of 1.1:1 (versus 0.7:1 in 2018). Egg-viability dynamics in ovitraps significantly varied between Treated and Control Sites, with an estimated overall reduction of 35% (versus 15% in 2018). The estimated proportion of females classified as mated with ARwP males was 41.8% and the viability rate of eggs laid by these females (9.5%) was on average significantly lower than that of females only mated with wild males (87.8%); however, high variability in fertility was observed. Values of ARwP male competitiveness were 0.36 and 0.73 based on the overall viability rate of eggs in ovitraps and on female fertility, respectively, i.e. well above the conventional 0.2 threshold for an effective suppressive impact in the field. CONCLUSIONS: Results further support the potential of IIT as a tool to contribute to Ae. albopictus control in the urban context, stressing the need of larger field trials to evaluate the cost-efficacy of the approach in temperate regions.
Modelling the effect of migration on the localisation and spread of a gene drive
24955C. Benjamin James and F.-L. Alexandre Jules Hen, bioRxiv, 2023.04.02.535303. 2023-04-04 14:22:19.
Gene drives have the potential to address pressing ecological issues. Through the super-Mendelian inheritance of a gene drive, a trait can be spread through a population even in spite of a fitness cost. This ability to spread is both its greatest quality and detractor. We may not want a gene drive to spread universally. If a gene drive were designed to cause the collapse of a pest population, it may inadvertently cause the collapse of the entire species. Migration is the mechanism through which a gene drive can spread to distant populations. Understanding its effect on the progression of a gene drive is crucial to our ability to control a gene drive. While migration can spread the gene drive to other populations, equally it can bring in other alleles to the population that may disrupt the progression of the gene drive. Through our deterministic migration gene drive model we can assess the conditions in which a gene drive is likely to spread to unintended populations, and if a gene drive is likely to be displaced by incoming alleles.Competing Interest StatementThe authors have declared no competing interest.
The Effect of the Sterile Insect Technique on Vibrational Communication: The Case of Bagrada hilaris (Hemiptera: Pentatomidae)
24951C. Peccerillo, C. E. Mainardi, R. Nieri, J. M. Fouani, A. Cemmi, M. Cristofaro, G. Anfora and V. Mazzoni, Insects, 14. 2023-04-02 13:09:28.
The painted bug, Bagrada hilaris, is an agricultural pest in its original areas (Africa, South Asia, and the Middle East), and it has recently been recorded as an invasive species in southwestern part of the US, Chile, Mexico, and two islands in the Mediterranean basin. Its polyphagous diet causes severe damage to economically important crops. The control of this pest is primarily achieved by means of synthetic pesticides, which are often expensive, ineffective, and harmful to the ecosystem. Recent physiological bioassays to assess its potential control through the sterile insect technique demonstrated that mating between untreated females and males irradiated at doses of 64 and 100 Gy, respectively, resulted in 90% and 100% sterility of the eggs produced by the females. In this study, the mating abilities of virgin males irradiated at 60 and 100 Gy with virgin females were measured through a study of short-range courtship mediated by vibrational communication. The results indicate that males irradiated at 100 Gy emit signals with lower peak frequencies, mate significantly less than unirradiated males do, and do not surpass the early stages of courtship. Conversely, males irradiated at 60 Gy present vibrational signal frequencies that are comparable to those of the control and successfully mated males. Our findings suggest that B. hilaris individuals irradiated at 60 Gy are good candidates for the control of this species, given that they retain sexual competitiveness regardless of their sterility, through an area-wide program that incorporates the sterile insect technique.
Gamma-radiation of Glossina palpalis gambiensis revisited: effect on fertility and mating competitiveness
24993S. Pagabeleguem, O. Koughuindida, E. W. Salou, G. Gimonneau, A. I. Toé, B. A. Kaboré, K. S. M. Dera, H. Maïga, A. M. G. Belem, G. M. S. Sanou/Ouédraogo, M. J. Vreysen and J. Bouyer, Parasite, 30:8. 2023-03-31 14:58:29.
African animal trypanosomoses are vector-borne diseases that cause enormous livestock losses in sub-Saharan Africa, with drastic socio-economic impacts. Vector control in the context of an area-wide integrated pest management program with a sterile insect technique component requires the production of high-quality sterile male tsetse flies. In our study, we evaluated the effect of irradiation on the fecundity of Glossina palpalis gambiensis to identify the optimal dose that will induce maximum sterility while maintaining biological performance as much as possible. In addition, male mating performance was evaluated in semi-field cages. The irradiation doses used were 90, 100, 110, 120, 130, 140, and 150 Gy, and untreated males were used as the control. The results showed that pupal production and emergence rates were higher in batches of females that had mated with fertile males than in those that had mated with irradiated males with any experimental dose. A dose of 120 Gy administered to male flies induced 97-99% sterility after mating with virgin females. For the semi-field cage experiments, males irradiated with 120 Gy showed good sexual competitiveness as compared to fertile males and those irradiated with 140 Gy, considering the level of filling of spermatheca and the number of pairs formed. The optimal radiation dose of 120 Gy found in this study is slightly different from the traditional dose of 110 Gy that has been used in several eradication programmes in the past. The potential reasons for this difference are discussed, and an argument is made for the inclusion of reliable dosimetry systems in these types of studies.
Improved Quality Management of the Indian Meal Moth, Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae) for Enhanced Efficacy of the Sterile Insect Technique
25139M. M. Hasan, M. A. Hossain and C. G. Athanassiou, Insects, 14. 2023-03-31 14:15:48.
The sterile insect technique (SIT) is considered an environmentally friendly, autocidal control tactic to manage insect pests. This work dealt with the improvement of quality management of the Indian meal moth Plodia interpunctella (Hübner) for enhanced efficacy of the SIT. The results indicated that egg hatching of irradiated mature eggs of P. interpunctella was higher than that of younger eggs, indicating that mature eggs were significantly more tolerant than younger eggs. Moreover, our data revealed that a dose of 500 Gy completely prevented pupal formation in irradiated young and mature larvae. Crosses between irradiated and non-irradiated adults resulted in considerable variations in fecundity. The mating competitiveness index (CI) value was higher for a ratio of 5:1:1 (sterile male, fertile male, and fertile female, respectively) as compared with the ratio 1:1:1 for the irradiated individuals of all life stages. Low temperature (5 °C) maintenance of irradiated pupae significantly affected adult emergence. Using cylinders to assess flight ability, we found that the flight performance of adults that were developed from cold treated irradiated pupae was influenced by cylinder diameter, cylinder height and the number of hours the insects were in the cylinders. The percentage of DNA damage of the reproductive organs of adults developed from cold treated pupae that were irradiated with 100 and 150 Gy varied significantly. The results of this study should be used to implement pilot-scale field tests aiming at a sterile- to-fertile male ratio of 5 to 1. © 2023 by the authors.
Additive Effect of Releasing Sterile Insects Plus Biocontrol Agents against Fruit Fly Pests (Diptera: Tephritidae) under Confined Conditions
24928P. Montoya, E. Flores-Sarmiento, P. López, A. Ayala and J. Cancino, Insects, 14. 2023-03-30 13:54:53.
Pest control models integrating the use of the sterile insect technique (SIT) and augmentative biological control (ABC) have postulated that it is possible to obtain a synergistic effect from the joint use of these technologies. This synergistic effect is attributed to the simultaneous attack on two different biological stages of the pest (immature and adult flies), which would produce higher suppression on the pest populations. Here we evaluated the effect of the joint application of sterile males of A. ludens of the genetic sexing strain Tap-7 along with two parasitoid species at the field cage level. The parasitoids D. longicaudata and C. haywardi were used separately to determine their effect on the suppression of the fly populations. Our results showed that egg hatching percentage was different between treatments, with the highest percentage in the control treatment and a gradual reduction in the treatments with only parasitoids or only sterile males. The greatest induction of sterility (i.e., the lowest egg hatching percentage) occurred with the joint use of ABC and SIT, demonstrating that the earlier parasitism caused by each parasitoid species was important reaching high levels of sterility. Gross fertility rate decreased up to 15 and 6 times when sterile flies were combined with D. longicaudata and C. haywardi, respectively. The higher parasitism by D. longicaudata was determinant in the decrease of this parameter and had a stronger effect when combined with the SIT. We conclude that the joint use of ABC and SIT on the A. ludens population had a direct additive effect, but a synergistic effect was observed in the parameters of population dynamics throughout the periodic releases of both types of insects. This effect can be of crucial importance in the suppression or eradication of fruit fly populations, with the added advantage of the low ecological impact that characterizes both techniques.
One strain may hide another: Cryptic male-killing Wolbachia
24924E. A. Hornett and G. D. D. Hurst, PLOS Biology, 21:e3002076. 2023-03-30 13:44:51.
While heritable symbionts are common in insects, strains that act as male-killers are considered rare. This Primer explores a new study in PLOS Biology which reveals a novel male-killer hidden by co-infection and host resistance, highlighting the complexity of host-microbial interactions in natural systems.
Differences in gene expression in field populations of Wolbachia-infected Aedes aegypti mosquitoes with varying release histories in northern Australia
B. Wimalasiri-Yapa, B. Huang, P. A. Ross, A. A. Hoffmann, S. A. Ritchie, F. D. Frentiu, D. Warrilow and A. F. van den Hurk, PLoS Negl Trop Dis, 17:e0011222. 2023-03-29 08:14:14.Aedes aegypti is the principal mosquito vector of dengue, yellow fever, Zika and chikungunya viruses. The wMel strain of the endosymbiotic bacteria Wolbachia pipientis was introduced into the vector as a novel biocontrol strategy to stop transmission of these viruses. Mosquitoes with Wolbachia have been released in the field in Northern Queensland, Australia since 2011, at various locations and over several years, with populations remaining stably infected. Wolbachia infection is known to alter gene expression in its mosquito host, but whether (and how) this changes over the long-term in the context of field releases remains unknown. We sampled mosquitoes from Wolbachia-infected populations with three different release histories along a time gradient and performed RNA-seq to investigate gene expression changes in the insect host. We observed a significant impact on gene expression in Wolbachia-infected mosquitoes versus uninfected controls. Fewer genes had significantly upregulated expression in mosquitoes from the older releases (512 and 486 from the 2011 and 2013/14 release years, respectively) versus the more recent releases (1154 from the 2017 release year). Nonetheless, a fundamental signature of Wolbachia infection on host gene expression was observed across all releases, comprising upregulation of immunity (e.g. leucine-rich repeats, CLIPs) and metabolism (e.g. lipid metabolism, iron transport) genes. There was limited downregulation of gene expression in mosquitoes from the older releases (84 and 71 genes from the 2011 and 2013/14 release years, respectively), but significantly more in the most recent release (509 from the 2017 release year). Our findings indicate that at > 8 years post-introgression into field populations, Wolbachia continues to profoundly impact expression of host genes, such as those involved in insect immune response and metabolism. If Wolbachia-mediated virus blocking is underpinned by these differential gene expression changes, our results suggest it may remain stable long-term.
The Promise and Challenge of Genetic Biocontrol Approaches for Malaria Elimination
24901S. James and M. Santos, Tropical Medicine and Infectious Disease, 2023-03-29 07:50:09.
Malaria remains an ongoing public health challenge, with over 600,000 deaths in 2021, of which approximately 96% occurred in Africa. Despite concerted efforts, the goal of global malaria elimination has stalled in recent years. This has resulted in widespread calls for new control methods. Genetic biocontrol approaches, including those focused on gene-drive-modified mosquitoes (GDMMs), aim to prevent malaria transmission by either reducing the population size of malaria transmitting mosquitoes or making the mosquitoes less competent to transmit the malaria parasite. The development of both strategies has advanced considerably in recent years, with successful field trials of several biocontrol methods employing live mosquito products and demonstration of the efficacy of GDMMs in insectary-based studies. Live mosquito biocontrol products aim to achieve area-wide control with characteristics that differ substantially from current insecticide-based vector control methods, resulting in some different considerations for approval and implementation. The successful field application of current biocontrol technologies against other pests provides evidence for the promise of these approaches and insights into the development pathway for new malaria control agents. The status of technical development as well as current thinking on the implementation requirements for genetic biocontrol approaches are reviewed, and remaining challenges for public health application in malaria prevention are discussed.
Paternity Analyses for the Planning of SIT Projects against the Red Palm Weevil
25135S. Belvedere, S. Arnone, M. Cristofaro, A. La Marca and A. De Biase, Insects, 14. 2023-03-28 14:05:49.
The red palm weevil Rhynchophorus ferrugineus is an invasive pest from southeastern Asia and Melanesia that has spread widely across the Middle East and the Mediterranean Basin over the last 30 years. Its endophagous larvae cause huge amounts of damage to several palm tree species from the Arecaceae family. Many of these palms are economically important for agricultural and ornamental purposes. Therefore, a lot of attention has recently been focused on studying this species with the aim of identifying sustainable and effective eradication strategies. Sterile insect techniques are biological control strategies that are currently being investigated for their potential to eradicate this pest in selected invasion areas. Mating system features (e.g., polyandry and related features) can affect the success and suitability of these approaches. The main goal of this research was to assess the performance of a previously developed microsatellite panel in terms of the paternity assignment of progeny from laboratory mating experiments. Using a simulation approach, we evaluated the reliability of the microsatellite markers in the paternity tests both in complex laboratory experiment scenarios and on the progeny of wild-caught gravid females to help future studies on the RPW mating system. As a case study of the simulation results, we performed two double-mating experiments, genotyped the progeny and estimated the P2 values to compare to the expected progeny genotypes according to the crossing scheme of each experiment. The results of our simulations on laboratory experiments showed that it was possible to carry out paternity assignments for all progeny with reliable statistical confidence using our 13 microsatellites set. On the contrary the low genetic variability measured in red palm weevil populations in invaded areas made the resolution power of our loci too low to carry out paternity analyses on natural populations. Results of laboratory crossing were completely congruent with the expectations from the Mendelian laws. © 2023 by the authors.
The Wolbachia WalE1 effector alters Drosophila endocytosis
24855M. Martin and I. L. G. Newton, bioRxiv, 2023-03-27 15:30:37.
The most common intracellular bacterial infection is Wolbachia pipientis , a microbe that manipulates host reproduction and is used in control of insect vectors. Phenotypes induced by Wolbachia have been studied for decades and range from sperm-egg incompatibility to male killing. How Wolbachia alters host biology is less well understood. Previously, we characterized the first Wolbachia effector â€" WalE1, which encodes a synuclein domain at the N terminus. Purified WalE1 sediments with and bundles actin and when heterologously expressed in flies, increases Wolbachia titer in the developing oocyte. In this work, we first identify the native expression WalE1 by Wolbachia infecting both fly cells and whole animals. WalE1 appears as aggregates, separate from Wolbachia cells. We next show that WalE1 co-immunoprecipitates with the host protein Past1 and that WalE1 manipulates host endocytosis. Yeast expressing WalE1 show deficiency in uptake of FM4-64 dye, and flies harboring mutations in Past1 or overexpressing WalE1 are sensitive to AgNO (3) , a hallmark of endocytosis defects. Finally, we also show that Past1 null flies harbor more Wolbachia overall and in late egg chambers. Our results identify interactions between a Wolbachia secreted effector and a host protein and point to yet another important host cell process impinged upon by Wolbachia .
Modelling Emergence of Wolbachia Toxin-Antidote Protein Functions with an Evolutionary Algorithm
24926J. Beckmann, J. Gillespie and D. Tauritz, bioRxiv, 2023-03-25 13:50:46.
Evolutionary algorithms (EAs) simulate Darwinian evolution and adeptly mimic natural evolution. Most EA applications in biology encode high levels of abstraction in top-down ecological population models. In contrast, our research merges protein alignment algorithms from bioinformatics into codon based EAs that simulate molecular protein string evolution from the bottom up. We apply our EA to reconcile a problem in the field of Wolbachia induced cytoplasmic incompatibility (CI). Wolbachia is a microbial endosymbiont that lives inside insect cells. CI is conditional insect sterility that operates as a toxin antidote (TA) system. Although, CI exhibits complex phenotypes not fully explained under a single discrete model. We instantiate in-silico genes that control CI, CI factors ( cifs ), as strings within the EA chromosome. We monitor the evolution of their enzymatic activity, binding, and cellular localization by applying selective pressure on their primary amino acid strings. Our model helps rationalize why two distinct mechanisms of CI induction might coexist in nature. We find that nuclear localization signals (NLS) and Type IV secretion system signals (T4SS) are of low complexity and evolve fast, whereas binding interactions have intermediate complexity, and enzymatic activity is the most complex. Our model predicts that as ancestral TA systems evolve into eukaryotic CI systems, the placement of NLS or T4SS signals can stochastically vary, imparting effects that might impact CI induction mechanics. Our model highlights how preconditions, genetic diversity, and sequence length can bias evolution of cifs towards one mechanism or another.
A male-killing Wolbachia endosymbiont is concealed by another endosymbiont and a nuclear suppressor
24920K. M. Richardson, P. A. Ross, B. S. Cooper, W. R. Conner, T. Schmidt and A. A. Hoffmann, PLoS Biol, 21:e3001879. 2023-03-22 13:30:17.
Bacteria that live inside the cells of insect hosts (endosymbionts) can alter the reproduction of their hosts, including the killing of male offspring (male killing, MK). MK has only been described in a few insects, but this may reflect challenges in detecting MK rather than its rarity. Here, we identify MK Wolbachia at a low frequency (around 4%) in natural populations of Drosophila pseudotakahashii. MK Wolbachia had a stable density and maternal transmission during laboratory culture, but the MK phenotype which manifested mainly at the larval stage was lost rapidly. MK Wolbachia occurred alongside a second Wolbachia strain expressing a different reproductive manipulation, cytoplasmic incompatibility (CI). A genomic analysis highlighted Wolbachia regions diverged between the 2 strains involving 17 genes, and homologs of the wmk and cif genes implicated in MK and CI were identified in the Wolbachia assembly. Doubly infected males induced CI with uninfected females but not females singly infected with CI-causing Wolbachia. A rapidly spreading dominant nuclear suppressor genetic element affecting MK was identified through backcrossing and subsequent analysis with ddRAD SNPs of the D. pseudotakahashii genome. These findings highlight the complexity of nuclear and microbial components affecting MK endosymbiont detection and dynamics in populations and the challenges of making connections between endosymbionts and the host phenotypes affected by them.
Vector competence of sterile male Glossina fuscipes fuscipes for Trypanosoma brucei brucei: implications for the implementation of the sterile insect technique in a sleeping sickness focus in Chad
24883M. H. Mahamat, A. Ségard, J.-B. Rayaisse, R. Argiles-Herrero, A. G. Parker, P. Solano, A. M. M. Abd-Alla, J. Bouyer and S. Ravel, Parasites and Vectors, 16:111. 2023-03-22 07:59:24.
Human African trypanosomiasis (HAT) is a neglected tropical disease caused by Trypanosoma brucei gambiense transmitted by tsetse flies in sub‑Saharan West Africa. In southern Chad the most active and persistent focus is the Mandoul focus, with 98% of the reported human cases, and where African animal trypanosomosis (AAT) is also present. Recently, a control project to eliminate tsetse flies (Glossina fuscipes fuscipes) in this focus using the sterile insect technique (SIT) was initiated. However, the release of large numbers of sterile males of G. f. fuscipes might result in a potential temporary increase in transmission of trypanosomes since male tsetse flies are also able to transmit the parasite. The objective of this work was therefore to experimentally assess the vector competence of sterile males treated with isometamidium for Trypanosoma brucei brucei.
Identification and functional analysis of Cochliomyia hominivorax U6 gene promoters
27884R. Novas, T. Basika, M. E. Williamson, P. Fresia, A. Menchaca and M. J. Scott, Insect Molecular Biology, 2023-03-21 08:32:32.
The New World screwworm, Cochliomyia hominivorax, is an obligate parasite, which is a major pest of livestock. While the sterile insect technique was used very successfully to eradicate C. hominivorax from North and Central America, more cost-effective genetic methods will likely be needed in South America. The recent development of CRISPR/Cas9-based genetic approaches, such as homing gene drive, could provide a very efficient means for the suppression of C. hominivorax populations. One component of a drive system is the guide RNA(s) driven by a U6 gene promoter. Here, we have developed an in vivo assay to evaluate the activity of the promoters from seven C. hominivorax U6 genes. Embryos from the related blowfly Lucilia cuprina were injected with plasmid DNA containing a U6-promoter-guide RNA construct and a source of Cas9, either protein or plasmid DNA. Activity was assessed by the number of site-specific mutations in the targeted gene in hatched larvae. One promoter, Chom U6_b, showed the highest activity. These U6 gene promoters could be used to build CRISPR/Cas9-based genetic systems for the control of C. hominivorax.
Convergent Aedes and Drosophila CidB interactomes suggest cytoplasmic incompatibility targets are conserved
24875S. O. Oladipupo, J. D. Carroll and J. F. Beckmann, Insect Biochem Mol Biol, 103931. 2023-03-20 08:18:16.
Wolbachia-mediated cytoplasmic incompatibility (CI) is a conditional embryonic lethality induced when Wolbachia-modified sperm fertilizes an uninfected egg. The Wolbachia proteins, CidA and CidB control CI. CidA is a rescue factor that reverses lethality. CidA binds to CidB. CidB contains a deubiquitinating enzyme and induces CI. Precisely how CidB induces CI and what it targets are unknown. Likewise, how CidA prevents sterilization by CidB is not clear. To identify CidB substrates in mosquitos we conducted pull-down assays using recombinant CidA and CidB mixed with Aedes aegypti lysates to identify the protein interactomes of CidB and the CidB/CidA protein complex. Our data allow us to cross compare CidB interactomes across taxa for Aedes and Drosophila. Our data replicate several convergent interactions, suggesting that CI targets conserved substrates across insects. Our data support a hypothesis that CidA rescues CI by tethering CidB away from its substrates. Specifically, we identify ten convergent candidate substrates including P32 (protamine-histone exchange factor), karyopherin alpha, ubiquitin-conjugating enzyme, and bicoid stabilizing factor. Future analysis on how these candidates contribute to CI will clarify mechanisms.
Enhancing the scalability of Wolbachia-based vector-borne disease management: time and temperature limits for storage and transport of Wolbachia-infected Aedes aegypti eggs for field releases
24877M. J. Allman, Y. H. Lin, D. A. Joubert, J. Addley-Cook, M. C. Mejía-Torres, C. P. Simmons, H. A. Flores and J. E. Fraser, Parasit Vectors, 16:108. 2023-03-18 08:18:24.
BACKGROUND: Introgression of the bacterial endosymbiont Wolbachia into Aedes aegypti populations is a biocontrol approach being used to reduce arbovirus transmission. This requires mass release of Wolbachia-infected mosquitoes. While releases have been conducted using a variety of techniques, egg releases, using water-soluble capsules containing mosquito eggs and larval food, offer an attractive method due to its potential to reduce onsite resource requirements. However, optimisation of this approach is required to ensure there is no detrimental impact on mosquito fitness and to promote successful Wolbachia introgression. METHODS: We determined the impact of storage time and temperature on wild-type (WT) and Wolbachia-infected (wMel or wAlbB strains) Ae. aegypti eggs. Eggs were stored inside capsules over 8 weeks at 18 °C or 22 °C and hatch rate, emergence rate and Wolbachia density were determined. We next examined egg quality and Wolbachia density after exposing eggs to 4-40 °C to determine how eggs may be impacted if exposed to extreme temperatures during shipment. RESULTS: Encapsulating eggs for 8 weeks did not negatively impact egg viability or resulting adult emergence and Wolbachia density compared to controls. When eggs were exposed to temperatures within 4-36 °C for 48 h, their viability and resulting adult Wolbachia density were maintained; however, both were significantly reduced when exposed to 40 °C. CONCLUSIONS: We describe the time and temperature limits for maintaining viability of Wolbachia-infected Ae. aegypti eggs when encapsulated or exposed to extreme temperatures. These findings could improve the efficiency of mass releases by providing transport and storage constraints to ensure only high-quality material is utilised during field releases.
Functional analysis of Wolbachia Cid effectors unravels cooperative interactions to target host chromatin during replication
24869K. Terretaz, B. Horard, M. Weill, B. Loppin and F. Landmann, PLOS Pathogens, 19:e1011211. 2023-03-16 08:02:08.
Wolbachia are common bacteria among terrestrial arthropods. These endosymbionts transmitted through the female germline manipulate their host reproduction through several mechanisms whose most prevalent form called Cytoplasmic Incompatibility -CI- is a conditional sterility syndrome eventually favoring the infected progeny. Upon fertilization, the sperm derived from an infected male is only compatible with an egg harboring a compatible Wolbachia strain, this sperm leading otherwise to embryonic death. The Wolbachia Cif factors CidA and CidB responsible for CI and its neutralization function as a Toxin-Antitoxin system in the mosquito host Culex pipiens. However, the mechanism of CidB toxicity and its neutralization by the CidA antitoxin remain unexplored. Using transfected insect cell lines to perform a structure-function analysis of these effectors, we show that both CidA and CidB are chromatin interactors and CidA anchors CidB to the chromatin in a cell-cycle dependent-manner. In absence of CidA, the CidB toxin localizes to its own chromatin microenvironment and acts by preventing S-phase completion, independently of its deubiquitylase -DUB- domain. Experiments with transgenic Drosophila show that CidB DUB domain is required together with CidA during spermatogenesis to stabilize the CidA-CidB complex. Our study defines CidB functional regions and paves the way to elucidate the mechanism of its toxicity.
The sex pheromone heptacosane enhances the mating competitiveness of sterile Aedes aegypti males
24861L.-M. Wang, N. Li, M. Zhang, Q. Tang, H.-Z. Lu, Q.-Y. Zhou, J.-X. Niu, L. Xiao, Z.-Y. Peng, C. Zhang, M. Liu, D.-Q. Wang and S.-Q. Deng, Parasites and Vectors, 16:102. 2023-03-15 15:04:09.
Aedes aegypti is a vector that transmits various viral diseases, including dengue and Zika. The radiation-based sterile insect technique (SIT) has a limited effect on mosquito control because of the difficulty in irradiating males without reducing their mating competitiveness. In this study, the insect sex pheromone heptacosane was applied to Ae. aegypti males to investigate whether it could enhance the mating competitiveness of irradiated males. Heptacosane was smeared on the abdomens of Ae. aegypti males that were allowed to mate with untreated virgin females. The insemination rate was used to assess the attractiveness of heptacosane‑treated males to females. The pupae were irradiated with different doses of X‑rays and γ‑rays, and the emergence, survival time, egg number, and hatch rate were detected to find the optimal dose of X‑ray and γ‑ray radiation. The males irradiated at the optimal dose were smeared with heptacosane, released in different ratios with untreated males, and mated with females. The effect of heptacosane on the mating competitiveness of irradiated mosquitoes was then evaluated by the hatch rate, induced sterility, and mating competitiveness index. Results Applying heptacosane to Ae. aegypti males significantly increased the insemination rate of females by 20%. Pupal radiation did not affect egg number but significantly reduced survival time and hatch rate. The emergence of the pupae was not affected by X‑ray radiation but was affected by γ‑ray radiation. Pupae exposed to 60 Gy X‑rays and 40 Gy γ‑rays were selected for subsequent experiments. After 60 Gy X‑ray irradiation or 40 Gy γ‑ray irradiation, the average hatch rate was less than 0.1%, and the average survival time was more than 15 days. Moreover, at the same release ratio, the hatch rate of the irradiated group perfumed with heptacosane was lower than that of the group without heptacosane. Conversely, the male sterility and male mating competitiveness index were significantly increased due to the use of heptacosane.
RNA interference is essential to modulating the pathogenesis of mosquito-borne viruses in the yellow fever mosquito Aedes aegypti
24948G. H. Samuel, T. Pohlenz, Y. Dong, N. Coskun, Z. N. Adelman, G. Dimopoulos and K. M. Myles, Proceedings of the National Academy of Sciences, 120:e2213701120. 2023-03-14 10:39:54.
While it has long been known that the transmission of mosquito-borne viruses depends on the establishment of persistent and nonlethal infections in the invertebrate host, specific roles for the insects? antiviral immune pathways in modulating the pathogenesis of viral infections is the subject of speculation and debate. Here, we show that a loss-of-function mutation in the Aedes aegypti Dicer-2 (Dcr-2) gene renders the insect acutely susceptible to a disease phenotype upon infection with pathogens in multiple virus families associated with important human diseases. Additional interrogation of the disease phenotype demonstrated that the virus-induced pathology is controlled through a canonical RNA interference (RNAi) pathway, which functions as a resistance mechanism. These results suggest comparatively modest contributions of proposed tolerance mechanisms to the fitness of A. aegypti infected with these pathogens. Similarly, the production of virus-derived piwi-interacting RNAs (vpiRNAs) was not sufficient to prevent the pathology associated with viral infections in Dcr-2 null mutants, also suggesting a less critical, or potentially secondary, role for vpiRNAs in antiviral immunity. These findings have important implications for understanding the ecological and evolutionary interactions occurring between A. aegypti and the pathogens they transmit to human and animal hosts.
Distinct Wolbachia localization patterns in oocytes of diverse host species reveal multiple strategies of maternal transmission
24853Y. A. Radousky, M. T. J. Hague, S. Fowler, E. Paneru, A. Codina, C. Rugamas, G. Hartzog, B. S. Cooper and W. Sullivan, Genetics, 2023-03-13 15:24:48.
A broad array of endosymbionts radiate through host populations via vertical transmission, yet much remains unknown concerning the cellular basis, diversity and routes underlying this transmission strategy. Here we address these issues, by examining the cellular distributions of Wolbachia strains that diverged up to 50 million years ago in the oocytes of 18 divergent Drosophila species. This analysis revealed three Wolbachia distribution patterns: 1) a tight clustering at the posterior pole plasm (the site of germline formation); 2) a concentration at the posterior pole plasm, but with a significant bacteria population distributed throughout the oocyte; 3) and a distribution throughout the oocyte, with none or very few located at the posterior pole plasm. Examination of this latter class indicates Wolbachia accesses the posterior pole plasm during the interval between late oogenesis and the blastoderm formation. We also find that one Wolbachia strain in this class concentrates in the posterior somatic follicle cells that encompass the pole plasm of the developing oocyte. In contrast, strains in which Wolbachia concentrate at the posterior pole plasm generally exhibit no or few Wolbachia in the follicle cells associated with the pole plasm. Taken together, these studies suggest that for some Drosophila species, Wolbachia invade the germline from neighboring somatic follicle cells. Phylogenomic analysis indicates that closely related Wolbachia strains tend to exhibit similar patterns of posterior localization, suggesting that specific localization strategies are a function of Wolbachia-associated factors. Previous studies revealed that endosymbionts rely on one of two distinct routes of vertical transmission: continuous maintenance in the germline (germline-to-germline) or a more circuitous route via the soma (germline-to-soma-to-germline). Here we provide compelling evidence that Wolbachia strains infecting Drosophila species maintain the diverse arrays of cellular mechanisms necessary for both of these distinct transmission routes. This characteristic may account for its ability to infect and spread globally through a vast range of host insect species.
Assessing the hybridization potential between a hypothetical gene drive-modified Drosophila suzukii strain and non-target Drosophila species
24990J. Romeis, S. Wolf, J. Collatz, J. Enkerli and F. Widmer, IOBC-WPRS Bulletin, 163:108. 2023-03-13 14:49:23.
Genetically engineered gene drives (GD) are a potentially powerful tool to control pest insects by population suppression or even elimination. Before living GD modified insects can be released into the environment, they must pass an environmental risk assessment (ERA). A key question to be addressed is the potential acquisition of functional GD elements by nontarget species as this may lead to the loss of those species and to a disruption of the ecosystem services they provide. The main route for gene flow is through hybridization between the GD insect strain and closely related species that co-occur in the area of release. Using the invasive Drosophila suzukii as a case study, we demonstrate how the potential for hybridization can be assessed.
A male-killing gene encoded by a symbiotic virus of Drosophila
24838D. Kageyama, T. Harumoto, K. Nagamine, A. Fujiwara, T. N. Sugimoto, A. Jouraku, M. Tamura, T. K. Katoh and M. Watada, Nature Communications, 14:1357. 2023-03-13 08:49:35.
In most eukaryotes, biparentally inherited nuclear genomes and maternally inherited cytoplasmic genomes have different evolutionary interests. Strongly female-biased sex ratios that are repeatedly observed in various arthropods often result from the male-specific lethality (male-killing) induced by maternally inherited symbiotic bacteria such as Spiroplasma and Wolbachia. However, despite some plausible case reports wherein viruses are raised as male-killers, it is not well understood how viruses, having much smaller genomes than bacteria, are capable of inducing male-killing. Here we show that a maternally inherited double-stranded RNA (dsRNA) virus belonging to the family Partitiviridae (designated DbMKPV1) induces male-killing in Drosophila. DbMKPV1 localizes in the cytoplasm and possesses only four genes, i.e., one gene in each of the four genomic segments (dsRNA1−dsRNA4), in contrast to ca. 1000 or more genes possessed by Spiroplasma or Wolbachia. We also show that a protein (designated PVMKp1; 330 amino acids in size), encoded by a gene on the dsRNA4 segment, is necessary and sufficient for inducing male-killing. Our results imply that male-killing genes can be easily acquired by symbiotic viruses through reassortment and that symbiotic viruses are hidden players in arthropod evolution. We anticipate that host-manipulating genes possessed by symbiotic viruses can be utilized for controlling arthropods.
First report of natural Wolbachia infections in mosquitoes from Cuba
24933A. Ruiz, G. Gutiérrez-Bugallo, R. Rodríguez-Roche, L. Pérez, R. González-Broche, L. A. Piedra, L. C. Martínez, Z. Menéndez, A. Vega-Rúa and J. A. Bisset, Acta Tropica, 242:106891. 2023-03-11 08:20:20.
Mosquitoes are extensively responsible for the transmission of pathogens. Novel strategies using Wolbachia could transform that scenario, since these bacteria manipulate mosquito reproduction, and can confer a pathogen transmission-blocking phenotype in culicids. Here, we screened the Wolbachia surface protein region by PCR in eight Cuban mosquito species. We confirmed the natural infections by sequencing and assessed the phylogenetic relationships among the Wolbachia strains detected. We identified four Wolbachia hosts: Aedes albopictus, Culex quinquefasciatus, Mansonia titillans, and Aedes mediovittatus (first report worldwide). Knowledge of Wolbachia strains and their natural hosts is essential for future operationalization of this vector control strategy in Cuba.
CRISPR-based genetic control strategies for insect pests
24978Y. Yan, R. A. Aumann, I. Hacker and M. F. Schetelig, Journal of Integrative Agriculture, 22:651-668. 2023-03-11 07:53:08.
Genetic control strategies such as the sterile insect technique have successfully fought insect pests worldwide. The CRISPR (clustered regularly interspaced short palindromic repeats) technology, together with high-quality genomic resources obtained in more and more species, greatly facilitates the development of novel genetic control insect strains that can be used in area-wide and species-specific pest control programs. Here, we review the research progress towards state-of-art CRISPR-based genetic control strategies, including gene drive, sex ratio distortion, CRISPR-engineered genetic sexing strains, and precision-guided sterile insect technique. These strategies' working mechanisms, potential resistance development mechanisms, and regulations are illustrated and discussed. In addition, recent developments such as stacked and conditional systems are introduced. We envision that the advances in genetic technology will continue to be one of the driving forces for developing the next generation of pest control strategies.
Modeling Sustained Transmission of Wolbachia among Anopheles Mosquitoes: Implications for Malaria Control in Haiti
24916D. Florez, A. J. Young, K. J. Bernabé, J. M. Hyman and Z. Qu, Trop Med Infect Dis, 8. 2023-03-09 12:09:17.
Wolbachia infection in Anopheles albimanus mosquitoes can render mosquitoes less capable of spreading malaria. We developed and analyzed a mechanistic compartmental ordinary differential equation model to evaluate the effectiveness of Wolbachia-based vector control strategies among wild Anopheles mosquitoes in Haiti. The model tracks the mosquito life stages, including egg, larva, and adult (male and female). It also accounts for critical biological effects, such as the maternal transmission of Wolbachia through infected females and cytoplasmic incompatibility, which effectively sterilizes uninfected females when they mate with infected males. We derive and interpret dimensionless numbers, including the basic reproductive number and next-generation numbers. The proposed system presents a backward bifurcation, which indicates a threshold infection that needs to be exceeded to establish a stable Wolbachia infection. The sensitivity analysis ranks the relative importance of the epidemiological parameters at baseline. We simulate different intervention scenarios, including prerelease mitigation using larviciding and thermal fogging before the release, multiple releases of infected populations, and different release times of the year. Our simulations show that the most efficient approach to establishing Wolbachia is to release all the infected mosquitoes immediately after the prerelease mitigation process. Moreover, the model predicts that it is more efficient to release during the dry season than the wet season.
A mosquito population suppression model with a saturated Wolbachia release strategy in seasonal succession
24814Z. Zhang, L. Chang, Q. Huang, R. Yan and B. Zheng, J Math Biol, 86:51. 2023-03-06 08:55:06.
Releasing Wolbachia-infected male mosquitoes to suppress wild female mosquitoes through cytoplasmic incompatibility has shown great promise in controlling and preventing mosquito-borne diseases. To make the release logistically and economically feasible, we propose a saturated release strategy, which is only implemented during the epidemic season of mosquito-borne diseases. Under this assumption, the model becomes a seasonally switching ordinary differential equation model. The seasonal switch brings rich dynamics, including the existence of a unique periodic solution or exactly two periodic solutions, which are proved by using the qualitative property of the Poincaré map. Sufficient conditions are also obtained for determining the stability of the periodic solutions.
Analysing inhibition of dengue virus in Wolbachia-infected mosquito cells following the removal of Wolbachia
24822M. Hussain, K. Etebari and S. Asgari, Virology, 581:48-55. 2023-03-03 10:30:05.
Wolbachia pipientis is known to block replication of positive sense RNA viruses. Previously, we created an Aedes aegypti Aag2 cell line (Aag2.wAlbB) transinfected with the wAlbB strain of Wolbachia and a matching tetracycline-cured Aag2.tet cell line. While dengue virus (DENV) was blocked in Aag2.wAlbB cells, we found significant inhibition of DENV in Aag2.tet cells. RNA-Seq analysis of the cells confirmed removal of Wolbachia and lack of expression of Wolbachia genes that could have been due to lateral gene transfer in Aag2.tet cells. However, we noticed a substantial increase in the abundance of phasi charoen-like virus (PCLV) in Aag2.tet cells. When RNAi was used to reduce the PCLV levels, DENV replication was significantly increased. Further, we found significant changes in the expression of antiviral and proviral genes in Aag2.tet cells. Overall, the results reveal an antagonistic interaction between DENV and PCLV and how PCLV-induced changes could contribute to DENV inhibition.
Genomic and cytogenetic analysis of the Ceratitis capitata temperature-sensitive lethal region
25470G. Sollazzo, G. Gouvi, K. Nikolouli, R. A. Aumann, H. Djambazian, M. A. Whitehead, P. Berube, S.-H. Chen, G. Tsiamis, A. C. Darby, J. Ragoussis, M. F. Schetelig and K. Bourtzis, G3-Genes Genomes Genetics, 13. 2023-03-03 08:40:56.
Genetic sexing strains (GSS) are an important tool in support of sterile insect technique (SIT) applications against insect pests and disease vectors. The yet unknown temperature-sensitive lethal (tsl) gene and the recently identified white pupae (wp) gene have been used as selectable markers in the most successful GSS developed so far, the Ceratitis capitata (medfly) VIENNA 8 GSS. The molecular identification of the tsl gene may open the way for its use as a marker for the development of GSS in other insect pests and disease vectors of SIT importance. Prior studies have already shown that the tsl gene is located on the right arm of chromosome 5, between the wp and Zw loci (tsl genomic region). In the present study, we used genomic, transcriptomic, bioinformatic, and cytogenetic approaches to characterize and analyze this genomic region in wild-type and tsl mutant medfly strains. Our results suggested the presence of 561 genes, with 322 of them carrying SNPs and/or insertion–deletion (indel) mutations in the tsl genomic region. Furthermore, comparative transcriptomic analysis indicated the presence of 32 differentially expressed genes, and bioinformatic analysis revealed the presence of 33 orthologs with a described heat-sensitive phenotype of Drosophila melanogaster in this region. These data can be used in functional genetic studies to identify the tsl gene(s) and the causal mutation(s) responsible for the temperature-sensitive lethal phenotype in medfly, and potentially additional genes causing a similar phenotype.
Effects of Wolbachia on Transposable Element Expression Vary Between Drosophila melanogaster Host Genotypes
24871A. T. Eugénio, M. S. P. Marialva and P. Beldade, Genome Biology Evolution, 15. 2023-03-03 08:07:31.
Transposable elements (TEs) are repetitive DNA sequences capable of changing position in host genomes, thereby causing mutations. TE insertions typically have deleterious effects but they can also be beneficial. Increasing evidence of the contribution of TEs to adaptive evolution further raises interest in understanding what factors impact TE activity. Based on previous studies associating the bacterial endosymbiont Wolbachia with changes in the abundance of piRNAs, a mechanism for TE repression, and to transposition of specific TEs, we hypothesized that Wolbachia infection would interfere with TE activity. We tested this hypothesis by studying the expression of 14 TEs in a panel of 25 Drosophila melanogaster host genotypes, naturally infected with Wolbachia and annotated for TE insertions. The host genotypes differed significantly in Wolbachia titers inside individual flies, with broad-sense heritability around 20%, and in the number of TE insertions, which depended greatly on TE identity. By removing Wolbachia from the target host genotypes, we generated a panel of 25 pairs of Wolbachia-positive and Wolbachia-negative lines in which we quantified transcription levels for our target TEs. We found variation in TE expression that was dependent on Wolbachia status, TE identity, and host genotype. Comparing between pairs of Wolbachia-positive and Wolbachia-negative flies, we found that Wolbachia removal affected TE expression in 21.1% of the TE-genotype combinations tested, with up to 2.3 times differences in the median level of transcript. Our data show that Wolbachia can impact TE activity in host genomes, underscoring the importance this endosymbiont can have in the generation of genetic novelty in hosts.
Genome evolution of dengue virus serotype 1 under selection by Wolbachia pipientis in Aedes aegypti mosquitoes
27978D. Thi Hue Kien, K. Edenborough, D. da Silva Goncalves, T. Thuy Vi, E. Casagrande, H. Thi Le Duyen, V. Thi Long, L. Thi Dui, V. Thi Tuyet Nhu, N. Thi Giang, H. Thi Xuan Trang, E. Lee, I. a. Donovan-Banfield, H. Thi Thuy Van, N. Minh Nguyet, N. Thanh Phong, Virus Evolution, 9:vead016. 2023-03-03 07:47:19.
The introgression of antiviral strains of Wolbachia into Aedes aegypti mosquito populations is a public health intervention for the control of dengue. Plausibly, dengue virus (DENV) could evolve to bypass the antiviral effects of Wolbachia and undermine this approach. Here, we established a serial-passage system to investigate the evolution of DENV in Ae. aegypti mosquitoes infected with the wMel strain of Wolbachia. Using this system, we report on virus genetic outcomes after twenty passages of serotype 1 of DENV (DENV-1). An amino acid substitution, E203K, in the DENV-1 envelope protein was more frequently detected in the consensus sequence of virus populations passaged in wMel-infected Ae. aegypti than wild-type counterparts. Positive selection at residue 203 was reproducible; it occurred in passaged virus populations from independent DENV-1-infected patients and also in a second, independent experimental system. In wild-type mosquitoes and human cells, the 203K variant was rapidly replaced by the progenitor sequence. These findings provide proof of concept that wMel-associated selection of virus populations can occur in experimental conditions. Field-based studies are needed to explore whether wMel imparts selective pressure on DENV evolution in locations where wMel is established.
Wolbachia pipientis infections in populations of Aedes albopictus in the city of València (Spain): implications for mosquito control
24830R. Bueno-Marí, R. Domínguez-Santos, M. Trelis, E. Garrote-Sánchez, M. Cholvi, F. Quero de Lera, M. Khoubbane, A. Marcilla and R. Gi, Revista Española de Salud Pública, 97. 2023-03-02 16:20:52.
OBJECTIVE: The presence of Aedes albopictus, of high sanitary and social impact, was first reported in Valencia (Eastern Spain) in 2015. Innovative tools for its control include the use of the endosymbiotic bacterium Wolbachia pipientis. The release of mosquito males infected with the wPip strain, has proven very promising for large-scale Incompatible Insect Technique (IIT) applications. Before this strategy can be implemented in Valencia, it is important to know whether the natural local mosquito populations are Wolbachia-infected and, if so, identifying the infecting strains/supergroups, these being the objectives of the present work. METHODS: Eggs were collected from the 19 districts of the València city between May and October 2019. A total of 50 lab-reared adult Ae. albopictus individuals were processed and analyzed for Wolbachia detection and molecular characterization. These actions took place within the framework of a collaboration established with the Department of Health and Consumer Affairs of the city council of Valencia. Fisher's exact test was used to detect the statistical significance of the differences between groups. RESULTS: Our study revealed that 94% of the analyzed samples were naturally infected with Wolbachia. Both wAlbA and wAlbB supergroups were identified, with most samples (72% of the infected ones) carrying co-infections. CONCLUSIONS: These data provide the first characterization of the Wolbachia presence in natural populations of Ae. albopictus in the Mediterranean area of Spain. This information is relevant to evaluate the potential use of Wolbachia strains in order to achieve the suppression of the Asian tiger mosquito populations through massive release of artificially-infected males.
Hybrid incompatibilities in the anopheles gambiae species complex
24968A. Kriezis, Imperial College London, 2023-03-01 10:47:20.
Malaria is an infectious disease caused by parasites of the genus Plasmodium which is responsible for approximately 400,000 deaths annually, primarily in sub-Saharan Africa. Malaria is transmitted by mosquitoes belonging to the Anopheles gambiae species complex. While progress has been made to reduce the incidence of malaria, the emergence of insecticide resistance necessitates the development of novel vector control strategies. Gene drive technologies have seen significant advances in recent years, providing hope for their implementation in the near future. While gene flow has been identified between sibling species of the An. gambiae species complex, they are reproductively isolated by both pre- and post-zygotic isolation mechanisms. Interspecific crosses between most member species produce sterile hybrid males, in accordance with Haldane’s rule of speciation. The aim of this project was to support the development of gene drive technologies by investigating hybrid incompatibilities between two of the most significant vector species, Anopheles gambiae and Anopheles arabiensis. The potential for the introgression of genomic regions from one species into the genetic background of the other was investigated to help inform models regarding the spread of gene drives between sibling species. In addition, the identification of genetic elements involved in hybrid male sterility could provide potential targets for vector control strategies. Large autosomal regions were found to introgress and persist in interspecific genomes without a detectable fertility cost. In addition, the introduction of distinct autosomal regions of conspecific DNA into otherwise heterospecific genomes of hybrid males was found to overcome hybrid incompatibilities and partially restore fertility. While no specific genetic factors involved in hybrid incompatibilities could be identified, the results indicate that such factors are present at least on the X chromosome. Furthermore, the evidence suggests that asynapsis between interspecific homologous autosomes during gametogenesis plays a role in the manifestation of hybrid male sterility.
Satellite Rearing of Aedes Mosquito Eggs: Synchronized Empirical Test of a Novel Mass Rearing Model
24981K. L. Dobson, K. Blore, J. A. Henke, K. Y. Hung, T. Morgan, T. Posey, S. Sun, O. Sypes, N. P. Tremblay and S. L. Dobson, J Am Mosq Control Assoc, 39:12-17. 2023-03-01 09:41:35.
Mosquito suppression strategies based on "rear and release" of male mosquitoes are attracting renewed interest from governments, municipalities, and private businesses. These include irradiation-based sterile insect technique, Wolbachia-based technologies, and genetic modification. Each of these approaches requires the mass rearing and release of adult male mosquitoes, which typically is accomplished via a rearing facility near the release site. Although some release programs have relied on centralized rearing and shipment of adult males, adult male mosquitoes are relatively fragile, and their fitness can be diminished by temperature fluctuations, humidity, nutritional deficiencies, and other stresses that occur during shipment. Furthermore, expensive, expedited shipment is typically used to maximize the amount of adult lifetime in the field following the release. In contrast, Aedes aegypti and Ae. albopictus eggs can be desiccated and stored for long periods. They are small, and many millions of eggs can be shipped without specialized environmental conditions and using less expensive means. Here we examine a model in which mosquito eggs are centrally produced and then mailed to satellite rearing facilities. As a control, a replicate set of eggs was reared at the factory of origin. At each of the rearing sites, cloud-based software was used to track and compare rearing at the different locations. The results demonstrate similar rearing outcomes (i.e., egg hatch, immature development, and number of adult males) at each of the different sites for both species. We discuss the outcome in relation to downstream applications and potential future studies.
Ovarian Transcriptional Response to Wolbachia Infection in D. melanogaster in the Context of Between-Genotype Variation in Gene Expression
24807S. I. Frantz, C. M. Small, W. A. Cresko and N. D. Singh, G3-Genes Genomes Genetics, 2023-03-01 08:37:16.
Wolbachia is a maternally-transmitted endosymbiotic bacteria that infects a wide variety of arthropod and nematode hosts. The effects of Wolbachia on host biology are far-reaching and include changes in host gene expression. However, previous work on the host transcriptional response has generally been investigated in the context of a single host genotype. Thus, the relative effect of Wolbachia infection versus host genotype on gene expression is unknown. Here we explicitly test the relative roles of Wolbachia infection and host genotype on host gene expression by comparing the ovarian transcriptomes of four strains of D. melanogaster infected and uninfected with Wolbachia. Our data suggest that infection explains a small amount of transcriptional variation, particularly in comparison to variation in gene expression among strains. However, infection specifically affects genes related to cell cycle, translation, and metabolism. We also find enrichment of cell division and recombination processes among genes with infection-associated differential expression. Broadly, the transcriptomic changes identified in this study provide novel understanding of the relative magnitude of the effect of Wolbachia infection on gene expression in the context of host genetic variation, and also point to genes that are consistently differentially expressed in response to infection among multiple genotypes.
Alleviating the burden of malaria with gene drive technologies? A biocentric analysis of the moral permissibility of modifying malaria mosquitoes
24800N. de Graeff, K. R. Jongsma and A. L. Bredenoord, Journal of Medical Ethics, 2023-02-28 08:33:28.
Gene drive technologies (GDTs) have been proposed as a potential new way to alleviate the burden of malaria, yet have also raised ethical questions. A central ethical question regarding GDTs relates to whether it is morally permissible to intentionally modify or eradicate mosquitoes in this way and how the inherent worth of humans and non-human organisms should be factored into determining this. Existing analyses of this matter have thus far generally relied on anthropocentric and zoocentric perspectives and rejected an individualist biocentric outlook in which all living organisms are taken to matter morally for their own sake. In this paper, we reconsider the implications of taking a biocentric approach and highlight nuances that may not be evident at first glance. First, we shortly discuss biocentric perspectives in general, and then outline Paul Taylor's biocentric theory of respect for nature. Second, we explore how conflicting claims towards different organisms should be prioritised from this perspective and subsequently apply this to the context of malaria control using GDTs. Our ethical analysis shows that this context invokes the principle of self-defence, which could override the pro tanto concerns that a biocentrist would have against modifying malaria mosquitoes in this way if certain conditions are met. At the same time, the case study of GDTs underlines the relevance of previously posed questions and criticism regarding the internal consistency of Taylor's egalitarian biocentrism.
Engagement on risk assessment for gene drive mosquitoes by EFSA and Target Malaria
24794S. Hartley, A. Kokotovich, Y. Devos and J. Mumford, Environmental Science and Policy, 142:183-193. 2023-02-27 11:12:07.
As engineered gene drive technologies continue to advance, many actors are actively considering how environmental risk assessments (RAs) for gene drive organisms should be conducted, and how stakeholder engagement opportunities should be provided. There is, however, a lack of clarity concerning what constitutes engagement on gene drive RA and, furthermore, what forms of engagement already exist around gene drive RA. To address this gap, we reflect on the actions of a risk assessor (the European Food Safety Authority, EFSA) and a gene drive developer (Target Malaria) to understand: 1) the RA-related decisions that each are making concerning gene drive technology for mosquitoes and other harmful insects, 2) the existing role of engagement in those decisions, and 3) the implications for our understandings of engagement and RA. We found, first, that both EFSA and Target Malaria have already made many RA-related decisions, even though any preparation and evaluation of a formal RA for gene drive mosquitoes remains far off. This finding supports the idea that gene drive RA involves multiple processes and decisions in different forms across the entire technology and regulatory development process. Second, we found that both EFSA and Target Malaria have already integrated engagement into their respective RA-related decisions in different ways, reflecting their different roles. We conclude by considering how EFSA and Target Malaria could improve their existing RA-related engagement by explicitly considering disciplinary diversity and worldview diversity in their related decision making.
A toxin-antidote CRISPR gene drive system for regional population modification
24826J. Champer, E. Lee, E. Yang, C. Liu, A. G. Clark and P. W. Messer, Nature Communications, 11:1082. 2023-02-27 10:39:13.
Engineered gene drives based on a homing mechanism could rapidly spread genetic alterations through a population. However, such drives face a major obstacle in the form of resistance against the drive. In addition, they are expected to be highly invasive. Here, we introduce the Toxin-Antidote Recessive Embryo (TARE) drive. It functions by disrupting a target gene, forming recessive lethal alleles, while rescuing drive-carrying individuals with a recoded version of the target. Modeling shows that such drives will have threshold-dependent invasion dynamics, spreading only when introduced above a fitness-dependent frequency. We demonstrate a TARE drive in Drosophila with 88-95% transmission by female heterozygotes. This drive was able to spread through a large cage population in just six generations following introduction at 24% frequency without any apparent evolution of resistance. Our results suggest that TARE drives constitute promising candidates for the development of effective, flexible, and regionally confinable drives for population modification.
Simulations Reveal High Efficiency and Confinement of a Population Suppression CRISPR Toxin-Antidote Gene Drive
24791Y. Zhu and J. Champer, ACS Synthetic Biolog, 2023-02-24 09:56:44.
Though engineered gene drives hold great promise for spreading through and suppressing populations of disease vectors or invasive species, complications such as resistance alleles and spatial population structure can prevent their success. Additionally, most forms of suppression drives, such as homing drives or driving Y chromosomes, will generally spread uncontrollably between populations with even small levels of migration. The previously proposed CRISPR-based toxin-antidote system called toxin-antidote dominant embryo (TADE) suppression drive could potentially address the issues of confinement and resistance. However, it is a relatively weak form of drive compared to homing drives, which might make it particularly vulnerable to spatial population structure. In this study, we investigate TADE suppression drive using individual-based simulations in a continuous spatial landscape. We find that the drive is actually more confined than in simple models without space, even in its most efficient form with low cleavage rate in embryos from maternally deposited Cas9. Furthermore, the drive performed well in continuous space scenarios if the initial release requirements were met, suppressing the population in a timely manner without being severely affected by chasing, a phenomenon in which wild-type individuals avoid the drive by recolonizing empty areas. At higher embryo cut rates, the drive loses its ability to spread, but a single, widespread release can often still induce rapid population collapse. Thus, if TADE suppression gene drives can be successfully constructed, they may play an important role in control of disease vectors and invasive species when stringent confinement to target populations is desired.
Development of transgenic corn planthopper Peregrinus maidis that express the tetracycline transactivator
27707A. A. Patil, W. Klobasa, D. Espinoza-Rivera, O. Baars, M. D. Lorenzen and M. J. Scott, Insect Molecular Biology, 32:363-375. 2023-02-24 09:39:46.
Abstract The corn planthopper, Peregrinus maidis, is a vector of several maize viruses and is consequently a significant agricultural pest in many tropical and subtropical regions. As P. maidis has developed resistance to insecticides, the aim of this study was to develop transgenic P. maidis strains that could be used for future genetic biocontrol programs. To facilitate the identification of transgenic P. maidis, we isolated and characterized the promoters for the P. maidis ubiquitin-like and profilin genes. Transient expression assays with P. maidis embryos showed that both promoters were active. Transgenic lines were established using piggyBac vectors and fluorescent protein marker genes. The lines carried an auto-regulated tetracycline transactivator (tTA) gene, which has been widely used to establish conditional lethal strains in other insect species. The transgenic lines showed low levels of tTA expression but were viable on diet with or without doxycycline, which inhibits the binding of tTA to DNA. We discuss possible modifications to the tTA overexpression system that could lead to the successful development of conditional lethal strains. To our knowledge, this is the first report of a transgenic Hemiptera. The approach we have taken could potentially be applied to other Hemiptera and, for P. maidis, the technology will facilitate future functional genomics studies.
Next-generation CRISPR gene-drive systems using Cas12a nuclease
24783S. Sanz Juste, E. M. Okamoto, X. Feng and V. L. Del Amo, bioRxiv, 2023.02.20.529271. 2023-02-20 09:40:53.
One method for reducing the impact of vector-borne diseases is through the use of CRISPR-based gene drives, which manipulate insect populations due to their ability to rapidly propagate desired genetic traits into a target population. However, all current gene drives employ a Cas9 nuclease that is constitutively active, impeding our control over their propagation abilities and limiting the generation of novel gene drive arrangements. Yet, other nucleases such as the temperature-sensitive Cas12a have not been explored for gene drive designs. To address this, we herein present a proof-of-concept gene-drive system driven by Cas12a that can be regulated via temperature modulation. Furthermore, we combined Cas9 and Cas12a to build double gene drives capable of simultaneously spreading two independent engineered alleles. The development of Cas12a-mediated gene drives provides an innovative option for designing next-generation vector control strategies to combat disease vectors and agricultural pests.Competing Interest StatementAll authors declare no competing interests.
