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ARS Home Midwest Area Peoria, Illinois National Center for Agricultural Utilization Research Crop Bioprotection Research Research





- Ramirez, Jose
- Muturi, Ephantus
- 2023
- 2022
- 2021
- Publications
- Product Quality and New Uses (306)
- Veterinary, Medical and Urban Entomology (104)

Research Project: Environmentally-Friendly, Microbial and Plant-Based Agents for Mosquito Control **Location:
                            Crop Bioprotection Research** 2023 Annual Report **Objectives**
                                

Objective 1: Enable the commercial production of microorganisms and their bioactive metabolites to control mosquitoes and the viruses they carry.
Goal 1.1: Evaluate larval-specific fungal/bacterial entomopathogens for mosquito control.
Goal 1.2: Characterize and select microbial isolates with potential for bioactive factor production.
Objective 2: Enable the commercial production of bioactive compounds from plants to control mosquitoes.
Goal 2.1: Identify essential oils with adulticidal activity against mosquitoes and their potential application as ingredients of attractive toxic sugar bait. 
Goal 2.2: Develop essential oil emulsions that are effective against mosquito larvae. **Approach**
                                

Mosquito control is a fundamental component of mosquito-borne disease prevention and outbreak control. The conventional approach to mosquito control relies heavily on synthetic chemical insecticides, but there is an urgent need for alternative vector control tools to tackle the rising problem of insecticide resistance and limit pesticide-related environmental hazards. Biopesticides are pest management agents based on living organisms or natural products and have a proven potential as ecofriendly alternatives to synthetic chemical insecticides. To date, only a limited number of biopesticides have been commercialized for use in mosquito control. Thus, the discovery of new biopesticide agents is one of the key priorities of vector biology research. This project will apply technologies allied with the fields of medical entomology, molecular biology, microbiology, chemical ecology and natural products chemistry to discover new microbial- and plant-based biopesticide agents to be developed and commercialized for mosquito control. Plant-based compounds that are highly effective against mosquitoes will be identified and developed into water-soluble and environmentally stable formulations for effective delivery to the target mosquitoes. The potential to harness bioactive compounds from plants as active ingredients for attractive toxic sugar-baits for mosquito control will also be explored. The project will focus primarily on plant essential oils because of their proven potential for pest and vector management. Additionally, we will explore and identify new entomopathogenic fungi and bacteria that kill different life stages of the mosquito. The bioactive compounds contributing to entomopathogenic activity of these fungi/bacteria will be isolated, characterized and examined for mosquitocidal and anti-arboviral activity. Successful completion of this project will lead to new discoveries that have great potential to propel the development and eventual commercialization of novel plant- and microbial-based agents for mosquito control. **Progress Report**
                                

We made significant progress in Objective 1, evaluating five bacterial strains for larvicidal activity against the yellow fever mosquito, Aedes aegypti. These bacteria were selected from the ARS Culture Collection (NRRL) because of their potential to be pathogenic to insects (i.e., isolated from an insect host). These bacterial strains encompassed five bacterial genera and represented a range of toxicity against mosquito larvae. Some of these bacteria also produce molecules with anti-bacterial properties. Our studies are allowing us to identify bacterial strains with potential to be lethal to mosquito larvae and to serve as leads for bioprospecting studies on new antimicrobial compounds. These studies also tested different entomopathogenic fungi for their ability to stimulate the production of mosquito antimicrobial peptides in the yellow fever mosquito, thus allowing us to further dissect the mechanism of mosquito resistance to fungal biocontrol agents. In a separate project, a Chromobacterium crude dry formulation was tested against two species of ticks (Ixodes scapularis and Amblyoma americanum) with preliminary results indicating significant mortality of I. scapularis following exposure. 
Substantial progress was also made in testing culture conditions to stimulate production of secondary metabolites by three species of entomopathogenic fungi. Crude secondary metabolite testing suggests that organic additives (i.e., chitin) influence production of secondary metabolites with potent larvicidal activity. Additionally, preliminary assays indicate that select fractions of crude secondary metabolites have anti-mycobacterial properties. Further experiments are currently ongoing to identify the chemical properties of this potential antimicrobial compound.
For Objective 2, the bioactivity of brassica seed meal and their major chemical constituents were tested for toxicity against mosquito larvae. This study indicated that different brassica seed meals have varying degrees of toxicity and can readily be developed into plant-based bioinsecticides against mosquitoes. Additional assays evaluated the toxicity of crude pennycress oil against mosquito larvae. **Accomplishments**
                                

1. Identification of mosquito-derived antimicrobial proteins responsible for resistance to fungal-based biocontrol agents. Microbial biopesticides represent ecofriendly and sustainable mosquito control strategies. Their efficacy depends on the virulence of the microbial agent and on mosquito-derived factors that protect it from microbial infections. In this regard, mosquito resistance to microbes (including fungi used for mosquito control) depend on a range of antimicrobial proteins. ARS researchers in Peoria, Illinois, identified several of these antimicrobial proteins that the mosquito uses to resist fungal infection and tested the antimicrobial proteins in-vivo activity in the yellow fever mosquito (Aedes aegypti) when exposed to diverse fungal entomopathogens. The results showed that removal of a single antimicrobial protein in the mosquito did not affect its ability to fight fungal infections. However, removal of four antimicrobial proteins significantly affected the mosquito survival to fungal infections. This study indicated that these proteins are acting in unison, rather than individually, to arrest the proliferation of the fungal agents used to kill the mosquito. This study provides a new understanding of the mechanisms that confer resistance to insect-killing fungi in an important mosquito vector. Furthermore, this new information allows us to improve fungal-based mosquito control strategies by selecting fungal strains that are better at overcoming the action of these mosquito proteins. 2. Determined mustard seed meals are effective mosquito control biopesticides. Plant-derived biopesticides are attractive alternatives to synthetic pesticides because they are biodegradable and categorized as low-risk pesticide to non-target organisms such as mammals, fish and amphibians. Recently, plants from the mustard family Brassicaceae have been found to be sources of bioactive compounds with potential insecticidal activity. ARS researchers in Peoria, Illinois, evaluated the larvicidal activity of four defatted Brassica seed meals and their individual chemical constituents against the yellow fever mosquito Aedes aegypti. This study indicated that Brassica seed meals were more toxic to mosquito larvae than individual chemical compounds derived from Brassicaceae seeds. Specifically, garden cress (Lepidium sativum) seed meal was the most lethal to mosquito larvae after a 24h exposure period. This is the first study documenting the insecticidal activity of brassica seed meal, a byproduct of oil seed extraction, against mosquito larvae. This study shows that brassica seed meal may signify a potential new ecofriendly larvicide for mosquito control. **Review Publications**
                            *Flor-Weiler, L., Behle, R.W., Berhow, M.A., McCormick, S.P., Vaughn, S.F., Muturi, E.J., Hay, W.T. 2023. Bioactivity of brassica seed meals and its compounds as ecofriendly larvicides against mosquitoes. Scientific Reports. 13. Article 3936. https://doi.org/10.1038/s41598-023-30563-6.*
                                *Ramirez, J.L., Hampton, K.J., Rosales, A.M., Muturi, E.J. 2023. Multiple mosquito AMPs are needed to potentiate their antifungal effect against entomopathogenic fungi. Frontiers in Microbiology. 13. Article 1062383. https://doi.org/10.3389/fmicb.2022.1062383.*


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