Plant parasitic nematodes infest crops and present a threat to food security worldwide. Currently available chemical controls e.g. methyl bromide, organophosphates and carbamates have an unacceptable level of toxicity to non-target organisms and are being withdrawn from use. Fluensulfone is a new nematicide of the fluoroalkenyl thioether group that has significantly reduced environmental impact with low toxicity to non-target insects and mammals. Here, we show that the model genetic organism Caenorhabditis elegans is susceptible to the irreversible nematicidal effects of fluensulfone. Whilst the dose required is higher than that which has nematicidal activity against Meloidogyne spp. the profile of effects on motility, egg-hatching and survival is similar to that reported for plant parasitic nematodes. C. elegans thus provides a tractable experimental paradigm to analyse the effects of fluensulfone on nematode behaviour. We find that fluensulfone has pleiotropic actions and inhibits development, egg-laying, egg-hatching, feeding and locomotion. In the case of feeding and locomotion, an early excitation precedes the gross inhibition. The profile of these effects is notably distinct from other classes of anthelmintic and nematicide: the inhibition of motility caused by fluensulfone is not accompanied by the hypercontraction which is characteristic of organophosphates and carbamates and C. elegans mutants that are resistant to the carbamate aldicarb and the macrocyclic lactone ivermectin retain susceptibility to fluensulfone. These data indicate fluensulfone's mode of action is distinct from currently available nematicides and it therefore presents a promising new chemical entity for crop protection.

Genetic and chemical biology screens of C. elegans have been of enormous benefit in providing fundamental insight into neural function and neuroactive drugs. Recently the exploitation of microfluidic devices has added greater power to this experimental approach providing more discrete and higher throughput phenotypic analysis of neural systems. Here we make a significant addition to this repertoire through the design of a semi-automated microfluidic device, NeuroChip, which has been optimised for selecting worms based on the electrophysiological features of the pharyngeal neural network. We demonstrate this device has the capability to sort mutant from wild-type worms based on high definition extracellular electrophysiological recordings. NeuroChip resolves discrete differences in excitatory, inhibitory and neuromodulatory components of the neural network from individual animals. Worms may be fed into the device consecutively from a reservoir and recovered unharmed. It combines microfluidics with integrated electrode recording for sequential trapping, restraining, recording, releasing and recovering of C. elegans. Thus mutant worms may be selected, recovered and propagated enabling mutagenesis screens based on an electrophysiological phenotype. Drugs may be rapidly applied during the recording thus permitting compound screening. For toxicology, this analysis can provide a precise description of sub-lethal effects on neural function. The chamber has been modified to accommodate L2 larval stages showing applicability for small size nematodes including parasitic species which otherwise are not tractable to this experimental approach. We also combine NeuroChip with optogenetics for targeted interrogation of the function of the neural circuit. NeuroChip thus adds a new tool for exploitation of C. elegans and has applications in neurogenetics, drug discovery and neurotoxicology.

The nematicidal action of fluensulfone follows a time-course which progresses from an early impact on motility through to an accumulating metabolic impairment, an inability to access lipid stores and death.

No abstract

Plant parasitic nematodes (PPNs) infest the roots of crops and cause global losses with a severe economic impact on food production. Current chemical control agents are being removed from use due to environmental and toxicity concerns and there is a need for new approaches to crop protection. A key feature of parasitic behaviour for the majority of PPNs is a hollow stomastyle or odontostyle required for interaction with the host plant and feeding. This lance-like microscopic structure, often called a stylet, protrudes from the mouth of the worm and thrusts in a rhythmic manner to stab the host root. Studying stylet activity presents technical challenges and as a consequence the underlying biology is poorly understood. We have addressed this by designing a microfluidic chip which traps the PPN Globodera pallida and permits the recording of an electrophysiological signal concomitant with stylet thrusting. The PDMS chip incorporates a precisely designed aperture to trap the nematode securely around a mid-point of its body. It is fabricated using a novel combination of conventional photolithography and two photon polymerization. The chip incorporates valves for rapid application of test compounds and integral electrodes to facilitate acquisition of electrical signals. We show that stylet thrusting can be induced by controlled application of 5-HT (serotonin) to the worm. Each thrust and retraction produces an electrical waveform that characterises the physiological activity associated with the worm's behaviour. The ability to reproducibly record the stylet activity of PPNs provides a new platform for nematicide screening that specifically focuses on a behaviour that is integral to the parasite host interaction. This is the first report of a microfluidic chip capable of electrophysiological recording from nematodes other than Caenorhabditis elegans. The unique approach is optimised for trapping and recording from smaller worms or worms with distinct anterior body shapes and may be applied to other species of economic or medical importance.