Scholarly Literature

This is a database of scholarly literature that concentrates currently on natural and engineered selfish genetic elements (gene drives).  The latest are shown here.
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Rapid, scalable, combinatorial genome engineering by marker-less enrichment and recombination of genetically engineered loci in yeast

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M. Abdullah, B. M. Greco, J. M. Laurent, R. K. Garge, D. R. Boutz, M. Vandeloo, E. M. Marcotte and A. H. Kachroo,  Cell Rep Methods,  3:100464. 2023-05-10 12:35:58.
A major challenge to rationally building multi-gene processes in yeast arises due to the combinatorics of combining all of the individual edits into the same strain. Here, we present a precise and multi-site genome editing approach that combines all edits without selection markers using CRISPR-Cas9. We demonstrate a highly efficient gene drive that selectively eliminates specific loci by integrating CRISPR-Cas9-mediated double-strand break (DSB) generation and homology-directed recombination with yeast sexual assortment. The method enables marker-less enrichment and recombination of genetically engineered loci (MERGE). We show that MERGE converts single heterologous loci to homozygous loci at ∼100% efficiency, independent of chromosomal location. Furthermore, MERGE is equally efficient at converting and combining multiple loci, thus identifying compatible genotypes. Finally, we establish MERGE proficiency by engineering a fungal carotenoid biosynthesis pathway and most of the human α-proteasome core into yeast. Therefore, MERGE lays the foundation for scalable, combinatorial genome editing in yeast.

Combined actions of bacteriophage-encoded genes in Wolbachia-induced male lethality

25420
H. Arai, H. Anbutsu, Y. Nishikawa, M. Kogawa, K. Ishii, M. Hosokawa, S. R. Lin, M. Ueda, M. Nakai, Y. Kunimi, T. Harumoto, D. Kageyama, H. Takeyama and M. N. Inoue,  iScience,  26:106842. 2023-05-10 08:02:36.
Some Wolbachia endosymbionts induce male killing, whereby male offspring of infected females are killed during development; however, the origin and diversity of the underlying mechanisms remain unclear. In this study, we identified a 76 kbp prophage region specific to male-killing Wolbachia hosted by the moth Homona magnanima. The prophage encoded a homolog of the male-killing gene oscar in Ostrinia moths and the wmk gene that induces various toxicities in Drosophila melanogaster. Upon overexpressing these genes in D. melanogaster, wmk-1 and wmk-3 killed all males and most females, whereas Hm-oscar, wmk-2, and wmk-4 had no impact on insect survival. Strikingly, co-expression of tandemly arrayed wmk-3 and wmk-4 killed 90% of males and restored 70% of females, suggesting their conjugated functions for male-specific lethality. While the male-killing gene in the native host remains unknown, our findings highlight the role of bacteriophages in male-killing evolution and differences in male-killing mechanisms among insects.

Editorial: Genetic control of insect pest species—achievements, challenges, and perspectives

25423
I. Häcker, D. Bartsch, A. Choo and F. Marec,  Frontiers in Bioengineering and Biotechnology,  11. 2023-05-05 08:12:07.
Genetic control is a type ofbiological control and a promising approach to regulate insect pest populations in a species-specific manner. It is based on targeting the reproductive capacity of the target pest species to reduce population size to non-critical levels. The best known and also very successful genetic control strategy is the Sterile Insect Technique (SIT), which entails the continuous mass-release ofirradiation-sterilized males ofa given species to produce infertile matings in the field, leading to the decline in the target population over time. To date, SIT is only available for a few species, as its transfer to new target species is challenging and time consuming. Key aspects of this classical SIT and challenges in applying it to new pest species include mass rearing of target species, mass removal of female insects prior to irradiation and release, the sterilization procedure, and the biological quality control of the sterile insects produced. Besides this classical SIT strategy, current research efforts are also focused on the development of genetic control approaches based on transgenic, symbiont-mediated, or gene-drive strategies. Modern genetic technologies offer new solutions for the improvement of existing genetic control strategies and insect strains, for faster and easier transfer of existing strategies to new target species, and also for the development of new genetic control approaches. Publications within this Research Topic address pressing questions and challenges related to the genetic control of insect pests.

The boundary problem: Defining and delineating the community in field trials with gene drive organisms

25098
N. de Graeff, I. Pirson, R. van der Graaf, A. L. Bredenoord and K. R. Jongsma,  Bioethics,  2023-05-03 10:11:03.
Despite widespread and worldwide efforts to eradicate vector-borne diseases such as malaria, these diseases continue to have an enormous negative impact on public health. For this reason, scientists are working on novel control strategies, such as gene drive technologies (GDTs). As GDT research advances, researchers are contemplating the potential next step of conducting field trials. An important point of discussion regarding these field trials relates to who should be informed, consulted, and involved in decision-making about their design and launch. It is generally argued that community members have a particularly strong claim to be engaged, and yet, disagreement and lack of clarity exist about how this "community" should be defined and delineated. In this paper, we shed light on this "boundary problem": the problem of determining how boundaries of inclusion and exclusion in (GDT) community engagement should be drawn. As our analysis demonstrates, the process of defining and delineating a community is itself normative. First, we explicate why it is important to define and delineate the community. Second, we demonstrate that different definitions of community are used and intermingled in the debate on GDTs, and argue in favor of distinguishing geographical, affected, cultural, and political communities. Finally, we propose initial guidance for deciding who should (not) be engaged in decision-making about GDT field trials, by arguing that the definition and delineation of the community should depend on the rationale for engagement and that the characteristics of the community itself can guide the effective design of community engagement strategies.

Mathematical modeling of the performance of wild and transgenic mosquitoes in malaria transmission

25861
A. P. Wyse, A. J. B. dos Santos, J. D. Azevedo, A. C. de Meneses and V. M. D. Santos,  Plos One,  18:23. 2023-04-28 08:20:19.
A mathematical model that simulates malaria transmission under the influence of transgenic mosquitoes refractory to malaria is presented in this paper. The zygosity of transgenic mosquitoes is taken into account and, consequently, the total population of mosquitoes is comprised of wild type and heterozygous and homozygous transgenic mosquitoes. These three mosquito varieties interact by mating and competition, and the genetic characteristics of their resulting offspring are in accordance with Mendelian genetics or the mutagenic chain reaction (MCR) technique. Although the incorporation of transgenic mosquitoes into the ecosystem reduces the incidence of malaria, the model also takes into account the importance of completing treatment in individuals with confirmed infection and the imminent risk of increased environmental temperature.

B chromosomes reveal a female meiotic drive suppression system in Drosophila melanogaster

25112
S. L. Hanlon and R. S. Hawley,  Current Biology,  2023-04-27 10:03:45.
Selfish genetic elements use a myriad of mechanisms to drive their inheritance and ensure their survival into the next generation, often at a fitness cost to its host.(1)(,)(2) Although the catalog of selfish genetic elements is rapidly growing, our understanding of host drive suppression systems that counteract self-seeking behavior is lacking. Here, we demonstrate that the biased transmission of the non-essential, non-driving B chromosomes in Drosophila melanogaster can be achieved in a specific genetic background. Combining a null mutant of matrimony, a gene that encodes a female-specific meiotic regulator of Polo kinase,(3)(,)(4) with the TM3 balancer chromosome creates a driving genotype that is permissive for the biased transmission of the B chromosomes. This drive is female-specific, and both genetic components are necessary, but not individually sufficient, for permitting a strong drive of the B chromosomes. Examination of metaphase I oocytes reveals that B chromosome localization within the DNA mass is mostly abnormal when drive is the strongest, indicating a failure of the mechanism(s) responsible for the proper distribution of B chromosomes. We propose that some proteins important for proper chromosome segregation during meiosis, like Matrimony, may have an essential role as part of a meiotic drive suppression system that modulates chromosome segregation to prevent genetic elements from exploiting the inherent asymmetry of female meiosis.

Wolbachia-based strategies for control of agricultural pests

25104
J. T. Gong, T. P. Li, M. K. Wang and X. Y. Hong,  Curr Opin Insect Sci,  101039:10.1016/j.cois.2023.101039. 2023-04-25 10:26:15.
Wolbachia-based incompatible insect technique (IIT) and pathogen blocking technique (PBT) have been shown to be effective at protecting humans from mosquito-borne diseases in the past decades. Population suppression based on IIT and population replacement based on PBT have become major field application strategies that have continuously been improved by the translational research on Wolbachia-transinfected mosquitoes. Similarly, Wolbachia-based approaches have been proposed for the protection of plants from agricultural pests and their associated diseases. However, a bottleneck in Wolbachia-based strategies for the control of agricultural pests is the need for methods to establish Wolbachia-transinfected insect lines. As a first step in this direction, we compare field control strategies for mosquitos with the potential strategies for agricultural pests based on Wolbachia. Our results show that there is a critical need for establishing productive insect lines and accumulating field test data.

Anti-CRISPR Anopheles mosquitoes inhibit gene drive spread under challenging behavioural conditions in large cages

25086
A. Simoni, R. D'Amato, C. Taxiarchi, M. Galardini, A. Trusso, R. Minuz, S. Gilli, A. Somerville, D. Shittu, A. Khalil, R. Galizi and R. Muller,  Research Square,  2023-04-24 06:46:30.
CRISPR-based gene drives have the potential to spread within a population and are considered as promising vector control tools. A doublesex-targeting gene drive was shown effective to suppress laboratory populations in both small and large cages, and it is considered for field application. Challenges related to the field-use of gene drives and the evolving regulatory framework demand for systems able to modulate or revert the action of gene drives, as part of post-release risk-mitigation plans. We developed an improved AcrIIA4-based anti-drive strain and showed inhibition of gene drive spread, in complex feeding and reproductive behavioural conditions. A stochastic model predicted the experimentally-observed genotypes dynamics in overlapping generations in medium- and large-sized cages and further demonstrated the effectiveness of anti-drive in different release and fitness scenarios. This study provides a further validation for the use of anti-drive system in controlling the spread of gene drive in Anopheles under complex behavioural conditions.

Applications of Anti-CRISPR Proteins in Genome Editing and Biotechnology

C. Kraus and E. J. Sontheimer,  Journal of Molecular Biology,  2023-04-24 06:39:49.
In the ten years since the discovery of the first anti-CRISPR (Acr) proteins, the number of validated Acrs has expanded rapidly, as has our understanding of the diverse mechanisms they employ to suppress natural CRISPR-Cas immunity. Many, though not all, function via direct, specific interaction with Cas protein effectors. The abilities of Acr proteins to modulate the activities and properties of CRISPR-Cas effectors have been exploited for an ever-increasing spectrum of biotechnological uses, most of which involve the establishment of control over genome editing systems. This control can be used to minimize off-target editing, restrict editing based on spatial, temporal, or conditional cues, limit the spread of gene drive systems, and select for genome-edited bacteriophages. Anti-CRISPRs have also been developed to overcome bacterial immunity, facilitate viral vector production, control synthetic gene circuits, and other purposes. The impressive and ever-growing diversity of Acr inhibitory mechanisms will continue to allow the tailored applications of Acrs.

A rapidly spreading deleterious aphid endosymbiont that uses horizontal as well as vertical transmission

25082
X. Gu, P. A. Ross, A. Gill, Q. Yang, E. Ansermin, S. Sharma, S. Soleimannejad, K. Sharma, A. Callahan, C. Brown, P. A. Umina, T. N. Kristensen and A. A. Hoffmann,  Proceedings of the National Academy of Sciences,  120:e2217278120. 2023-04-24 06:34:04.
Endosymbiotic bacteria that live inside the cells of insects are typically only transmitted maternally and can spread by increasing host fitness and/or modifying reproduction in sexual hosts. Transinfections of Wolbachia endosymbionts are now being used to introduce useful phenotypes into sexual host populations, but there has been limited progress on applications using other endosymbionts and in asexual populations. Here, we develop a unique pathway to application in aphids by transferring the endosymbiont Rickettsiella viridis to the major crop pest Myzus persicae. Rickettsiella infection greatly reduced aphid fecundity, decreased heat tolerance, and modified aphid body color, from light to dark green. Despite inducing host fitness costs, Rickettsiella spread rapidly through caged aphid populations via plant-mediated horizontal transmission. The phenotypic effects of Rickettsiella were sensitive to temperature, with spread only occurring at 19 °C and not 25 °C. Body color modification was also lost at high temperatures despite Rickettsiella maintaining a high density. Rickettsiella shows the potential to spread through natural M. persicae populations by horizontal transmission and subsequent vertical transmission. Establishment of Rickettsiella in natural populations could reduce crop damage by modifying population age structure, reducing population growth and providing context-dependent effects on host fitness. Our results highlight the importance of plant-mediated horizontal transmission and interactions with temperature as drivers of endosymbiont spread in asexual insect populations.

Gene Drives as Interventions into Nature: the Coproduction of Ontology and Morality in the Gene Drive Debate

25114
K. 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

24996
E. 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

25144
M. 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

24988
L. 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

25137
C. 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

25001
J. 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

25102
M. 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

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Y. 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)

24960
S. 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

24966
B. 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.

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