Briefing Document: "Recombinant venom proteins in insect seminal fluid reduce female lifespan"

Source: Article "Recombinant venom proteins in insect seminal fluid reduce female lifespan" by Samuel J. Beach & Maciej Maselko, published in Nature Communications (DOI: 10.1038/s41467-024-54863-1).

Date: Current date (based on when the briefing document is created).

Prepared for: [Intended Audience - e.g., Pest Management Stakeholders, Research Community, Regulatory Bodies]

Executive Summary:

This research presents a novel genetic biocontrol technology termed the "toxic male technique" (TMT). The study demonstrates, using Drosophila melanogaster as a model, that genetically engineering males to express insecticidal venom proteins within their seminal fluid, which are then transferred to females during mating, significantly reduces the lifespan of the mated females. This intragenerational approach contrasts with existing mating-based genetic biocontrol technologies that primarily focus on reducing offspring viability or skewing sex ratios. Agent-based modeling of Aedes aegypti suggests that TMT has the potential to be significantly more effective in rapidly reducing blood-feeding rates, a key factor in disease transmission, compared to leading intergenerational biocontrol methods like female-specific Release of Insects carrying a Dominant Lethal gene (fsRIDL). TMT offers a promising avenue for rapidly controlling outbreaks of disease vectors and agricultural pests with potentially reduced reliance on chemical insecticides and lower risks of transgene persistence compared to gene drive technologies.

Main Themes and Important Ideas/Facts:

• The Need for Alternative Pest Management Tools:
• The emergence of insecticide resistance is a major challenge.
• Current genetic biocontrol technologies (GBTs) have limitations, particularly a generational lag in their impact, leaving mated females capable of causing harm (e.g., disease transmission) in the interim.
• The study highlights the significant human and economic costs associated with insect pests and the negative impacts of widespread insecticide use on non-target species and the environment.
• "The emergence of insecticide resistance has increased the need for alternative pest management tools."
• Introducing Intragenerational Genetic Biocontrol: The Toxic Male Technique (TMT):
• TMT aims to directly reduce the lifespan of mated females by expressing toxic proteins in the male reproductive tract (male accessory glands - MAG) that are transferred during mating.
• This is presented as the "first example of an intragenerational GBT."
• "Here we describe the toxic male technique (TMT), which is—to the best of our knowledge—the first example of an intragenerational GBT. By genetically engineering pest species to express toxic, low molecular weight compounds within the MAG, the survival of mated females can be reduced (Fig. 1)."
• Proof of Concept in Drosophila melanogaster:
• Seven insect-specific venom proteins were selected based on criteria including invertebrate specificity, low mammalian toxicity, and low molecular weight.
• MAG-specific expression was achieved using the antr-GAL4 driver in the UAS/GAL4 system.
• Mating with males expressing Γ-CNTX-Pn1a (from a spider) or δ-AITX-Avd2a (from a sea anemone) significantly reduced the median lifespan of wild-type females by 37-64% compared to controls.
• Increasing the ratio of TMT males to females (3:1) resulted in an even greater reduction in female lifespan (50% and 64% respectively).
• "The median lifespans of wild-type females mated to two of the TMT strains are reduced by 37–64% after initial exposure compared to control females mated to wild type males."
• Functionality and Specificity of Recombinant Venoms:
• Functionality of the venom proteins was confirmed through ubiquitous expression assays, where functional venoms resulted in non-viable offspring.
• The antr-GAL4 driver showed strong and specific expression in the male accessory glands with minimal off-target expression.
• Single-pair courtship assays indicated that TMT males were able to court females as effectively as wild-type males, suggesting no significant impact on mating behavior.
• However, competitive mating assays revealed reduced competitiveness of the transgenic males likely due to the GAL4-UAS genetic background.
• Modeling the Impact on Aedes aegypti Populations:
• Agent-based models simulating Aedes aegypti release programs compared TMT to SIT and fsRIDL.
• The model predicted that TMT could reduce blood-feeding rates by a further 40-60% during release periods compared to fsRIDL.
• TMT achieved faster population reduction (PR50 and PR95) compared to fsRIDL in the simulations.
• The median number of gonotrophic cycles (related to disease transmission potential) was significantly lower in the TMT simulations.
• "Agent-based models of Aedes aegypti predict that TMT could reduce rates of blood feeding by a further 40 – 60% during release periods compared to leading biocontrol technologies like fsRIDL."
• Impact of Mating Lethality and Male Sterility in Models:
• Simulations exploring reduced mating lethality showed that fertile TMT males required ≥80% mating lethality to outperform fsRIDL in reducing blood-feeding.
• Sterile TMT males (combined with a sterility mechanism) only required ≥40% mating lethality to outperform fsRIDL.
• This highlights the potential benefit of combining TMT with male sterilization to prevent transgene escape.
• Sensitivity Analysis of Model Parameters:
• Sensitivity analysis revealed that polyandry (multiple mating by females) and release ratio of TMT males had significant positive effects on TMT's relative performance.
• Density-dependent mortality had a negative effect, although this could be mitigated by increased release ratios.
• Higher rates of polyandry are predicted to enhance the effectiveness of TMT.
• Potential and Future Directions of TMT:
• TMT offers a rapid response to pest outbreaks, making it a potential alternative to chemical pesticides as a first-line response.
• The taxonomic selectivity of venom proteins allows for targeted pest control.
• Co-expression of multiple toxins could help prevent resistance.
• Future research needs to focus on identifying optimal venom proteins (considering molecular weight and toxicity), improving expression levels, and engineering stable, conditional expression systems for mass rearing.
• Determining the mating competitiveness of TMT males in target species under wild-type genetic backgrounds is crucial.
• Further studies are needed to understand the epidemiological effects and the impact on non-target organisms.
• "TMT is inherently self-limiting due to strong selection pressure against venom expression, which maybe advantageous from a regulatory perspective as, unlike GBT such as gene drives, there is little risk of substantial transgene introgression into non-target populations, and TMT transgenes are predicted to be rapidly lost from all populations in the absence of ongoing releases."

Quotes:

• "Here, we demonstrate intra-generational genetic biocontrol, wherein mating with engineered males reduces female lifespan."
• "The toxic male technique (TMT) involves the heterologous expres...