Prof Howard Atkinson & Prof P E Urwin
We study fundamental aspects of plant parasitic nematodes and their interactions with plants. This knowledge is then used to underpin the development of new control strategies. Plant parasitic nematodes represent one of the major biotic causes of yield loss in crops estimated to be $125b annually. Potato cyst nematode costs the UK potato industry alone an estimated £43m/year. Control often depends on nematicides that are harmful to the environment and potentially to human health. We have progressed a range of new control options that have advanced from gene discovery through protein engineering to field trials and a full biosafety assessment.
Plant response to parasitism using microarrays: Many plant genes are differentially regulated inside the specialised feeding structure that nematodes induce. Cyst nematodes (e.g Globodera sp.) induce a syncytium in which the cytoplasm of neighbouring plant cells merges. The syncytium is different from the abnormally large so-called giant plant cells induced by root knot nematodes (Meloidogyne sp.). We have used microarrays to compare transcript abundance for plant genes in sections of root harbouring the syncytium of a female cyst nematode (Heterodera schachtii) with similar uninfected material. We found a similar number of gene expression changes in this and a parallel study using a root-knot nematode (Meloidogyne incognita). We are now carrying out further characterisation of plant genes that respond or either or both cyst and root knot nematodes.
RNA interference: RNAi was first described in the free living nematode Caenorhabditis elegans: It is a powerful and rapid method of inhibiting the function of a specific gene. We established RNAi for plant parasitic nematodes to facilitate functional analysis. Octopamine was used to induce dsRNA uptake and different genes were targeted to establish the general utility of the approach. We are using the approach to define gene products that are essential for parasitism. In addition the delivery of dsRNA in planta is being optimised to explore the potential for novel resistance.
Molecular characterisation of plant parasitic nematodes: We established that C. elegans has homologous digestive genes to those of plant parasitic nematodes. We have used C. elegans genomics to underpin a functional genomic study of digestive physiology of Globodera. Genes of Globodera encoding several digestive enzymes have been cloned using sequence information from conserved regions of predicted C. elegans digestive genes. A number of potential the digestive enzymes have also been identified and characterised using libraries isolated from small amounts of Globodera digestive tissue. We are studying the neurobiology of cyst nematodes in comparison to the architecture and function of the C. elegans chemoreception system. In another project that underpins pesticide discovery we aim to define the genes that are involved in the metabolism of xenobiotic chemicals by initially using C. elegans microarray technology to give leads to homologous genes in plant parasitic nematodes.
Molecular ecology: One postgraduate project is discerning the seasonal changes in gene expression of the plant parasitic nematode Longidorus elongatus. Our molecular expertise with Arabidopsis and microarray technology has led to collaboration with ecologists to define projects that are not centred on nematodes. We work with other groups to define the range margins of plants in the environment. This work currently focuses on the non-human commensal, Arabidopsis lyrata petraea and its restricted range in relation to thermal stress. The work forms part of a large collaborative project between groups at Leeds and Sheffield.
Strategic biotechnological applications of the work.
An anti-feedant approach: Cysteine proteinases are important digestive enzymes in many nematodes, but not humans. Cysteine proteinase activity is inhibited in female cyst nematodes after incubation with a cystatin. A gene encoding a rice cystatin, Oc-I, was engineered to have an enhanced inhibitory activity. Expression of the engineered variant in Arabidopsis plants was the first transgenic technology shown to work against both root-knot and cyst nematodes. The work has culminated in successful field and containment trials of transgenic potatoes expressing a cystatin. The best transgenic lines were shown to have commercially useful resistance. Full resistance to G. pallida was observed in the UK field by stacking natural and transgenic resistance. Transgenic potato plants with cystatin expression restricted largely to roots have been field trialled with similar resistance levels to those achieved with constitutive expression. By using specific promoters the genes can be programmed to work only in the plant tissues attacked by nematodes. This provides a safe and environmentally friendly method of control. Resistance has also been demonstrated against a range of nematode species and extended from potato to rice and banana.
Peptide technology: Our most recent strategic work has focused on the use of peptides that disrupt chemoreception in nematodes. We have isolated two different peptides that either inhibit acetylcholinesterase or bind to nematode nicotinic receptors. Both peptides inhibited chemoreception at c. 1 nM and probably undergo retrograde transport along certain chemoreceptive neurones. The strategic development of this approach provided a prototype potato plant that expressed the peptide that inhibits acetylcholinesterase. Protection levels of >80% have been achieved against nematodes. Other peptide mimetics are being sought and the efficacy of the technology is being evaluated further.
International collaborations: We aim to develop the technology for world agribusiness in partnership with biotechnology companies. We also donate the technology freely to subsistence farmers through not-for-profit organisations including the Department for International Development and USAID. We have collaborative links with India, China, Uganda, Hawaii and Brazil.