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Investigating the mode of action of fluensulfone on the plant parasitic nematode, Globodera pallida

Investigating the mode of action of fluensulfone on the plant parasitic nematode, Globodera pallida
Investigating the mode of action of fluensulfone on the plant parasitic nematode, Globodera pallida
Plant parasitic nematodes (PPNs) are a major agricultural pest that infest crops and affect food production worldwide. Current measures that exist for controlling PPN infestation are limited, with the majority of chemical control agents being withdrawn from use due to their off-target toxicity and impact on the environment. New nematicides are therefore required for safe and improved PPN management strategies. Fluensulfone (FLS) is a novel nematicide with a distinct profile of effects on PPN, suggesting a unique mode of action. Here, I investigate the effects of fluensulfone on G. pallida at two critical life cycle stages; the egg- and cyst-enclosed J2, and the free-living, non-feeding, infective J2. Investigation of the ability of fluensulfone to inhibit G. pallida hatching, relative to the efficacy of other distinct classes of nematicides, reinforced that fluensulfone has a distinct mode of action among existing nematicides and demonstrated that it is a potent inhibitor of hatching. At concentrations as low as 1μM (0.29 ppm), fluensulfone exhibited complete inhibition of G. pallida root diffusate-induced hatching. This effect was reversible at concentrations ≤5μM (1.46 ppm), partially reversible at concentrations ≤50μM (14.6 ppm) and irreversible at the maximum concentration tested of 500μM (146 ppm). Despite hatching inhibition being reversible at low concentrations, J2s that emerged under these conditions exhibited impaired motility. This suggests that fluensulfone impacts on the viability of the unhatched J2. The irreversibility of hatch inhibition at concentrations >50µM coincided with a granulated appearance of the egg-encased J2 and suggests a nematicidal action of fluensulfone on the unhatched J2. To inform mechanistic understanding of the nematastatic and nematicidal action of fluensulfone, I combined inhibitor studies to highlight the role played by distinct metabolic fluxes and provide evidence that lipid utilization is selectively disrupted. This was reinforced using Coherent anti-Stokes Raman scattering (CARS) spectroscopy to image lipid stores. I optimized approaches to extract major lipids from the PPN and probed for levels of key lipid. Interestingly, this failed to identify significant changes in the gross levels of lipids in G. pallida juveniles treated with fluensulfone. I discuss this mismatch. The effects of fluensulfone on lipid utilization were accompanied by morphological changes, manifest by the appearance of enlarged, lipid-bound organelles. These highlight that fluensulfone impacts lipid and/or membrane trafficking processes, accumulating vacuole-like structures. The gross features of these effects appear to be consistent with degradative and cell death pathways. In particular, the appearance of these structures show striking similarities to the accumulation of degradative water and lipid filled vacuoles associated with the cell death process of methuosis. Taken together, the data presented here present an increased understanding of the mode of action of fluensulfone. They support a model in which fluensulfone brings about a slowly developing metabolic insult that is nematicidal. This involves a disruption in lipid homeostasis and the nematicidal action of fluensulfone may be explained by the accumulation of toxic metabolic intermediates.
University of Southampton Press
Feist, Emily
17090654-ab15-4e64-aaa1-2e0f199365e7
Feist, Emily
17090654-ab15-4e64-aaa1-2e0f199365e7
Holden-Dye, Linda
8032bf60-5db6-40cb-b71c-ddda9d212c8e
O'connor, Vincent
8021b06c-01a0-4925-9dde-a61c8fe278ca

Feist, Emily (2020) Investigating the mode of action of fluensulfone on the plant parasitic nematode, Globodera pallida. University of Southampton, Doctoral Thesis, 291pp.

Record type: Thesis (Doctoral)

Abstract

Plant parasitic nematodes (PPNs) are a major agricultural pest that infest crops and affect food production worldwide. Current measures that exist for controlling PPN infestation are limited, with the majority of chemical control agents being withdrawn from use due to their off-target toxicity and impact on the environment. New nematicides are therefore required for safe and improved PPN management strategies. Fluensulfone (FLS) is a novel nematicide with a distinct profile of effects on PPN, suggesting a unique mode of action. Here, I investigate the effects of fluensulfone on G. pallida at two critical life cycle stages; the egg- and cyst-enclosed J2, and the free-living, non-feeding, infective J2. Investigation of the ability of fluensulfone to inhibit G. pallida hatching, relative to the efficacy of other distinct classes of nematicides, reinforced that fluensulfone has a distinct mode of action among existing nematicides and demonstrated that it is a potent inhibitor of hatching. At concentrations as low as 1μM (0.29 ppm), fluensulfone exhibited complete inhibition of G. pallida root diffusate-induced hatching. This effect was reversible at concentrations ≤5μM (1.46 ppm), partially reversible at concentrations ≤50μM (14.6 ppm) and irreversible at the maximum concentration tested of 500μM (146 ppm). Despite hatching inhibition being reversible at low concentrations, J2s that emerged under these conditions exhibited impaired motility. This suggests that fluensulfone impacts on the viability of the unhatched J2. The irreversibility of hatch inhibition at concentrations >50µM coincided with a granulated appearance of the egg-encased J2 and suggests a nematicidal action of fluensulfone on the unhatched J2. To inform mechanistic understanding of the nematastatic and nematicidal action of fluensulfone, I combined inhibitor studies to highlight the role played by distinct metabolic fluxes and provide evidence that lipid utilization is selectively disrupted. This was reinforced using Coherent anti-Stokes Raman scattering (CARS) spectroscopy to image lipid stores. I optimized approaches to extract major lipids from the PPN and probed for levels of key lipid. Interestingly, this failed to identify significant changes in the gross levels of lipids in G. pallida juveniles treated with fluensulfone. I discuss this mismatch. The effects of fluensulfone on lipid utilization were accompanied by morphological changes, manifest by the appearance of enlarged, lipid-bound organelles. These highlight that fluensulfone impacts lipid and/or membrane trafficking processes, accumulating vacuole-like structures. The gross features of these effects appear to be consistent with degradative and cell death pathways. In particular, the appearance of these structures show striking similarities to the accumulation of degradative water and lipid filled vacuoles associated with the cell death process of methuosis. Taken together, the data presented here present an increased understanding of the mode of action of fluensulfone. They support a model in which fluensulfone brings about a slowly developing metabolic insult that is nematicidal. This involves a disruption in lipid homeostasis and the nematicidal action of fluensulfone may be explained by the accumulation of toxic metabolic intermediates.

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Published date: 31 December 2020

Identifiers

Local EPrints ID: 450181
URI: http://eprints.soton.ac.uk/id/eprint/450181
PURE UUID: 68c9d870-328b-465b-9941-3110a775c816
ORCID for Linda Holden-Dye: ORCID iD orcid.org/0000-0002-9704-1217

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Date deposited: 15 Jul 2021 16:35
Last modified: 16 Jul 2021 01:34

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Contributors

Author: Emily Feist
Thesis advisor: Linda Holden-Dye ORCID iD
Thesis advisor: Vincent O'connor

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