An effective basis for control of potato cyst-nematodes and assessment of the impact on soil health

Lead Research Organisation: University of Leeds
Department Name: Inst of Integrative & Comparative Biolog

Abstract

Plant parasitic nematodes cause losses to global agriculture of 100b dollars annually. Potato cyst nematode costs the UK potato industry an estimated 43 m pounds sterling/year. Control often depends on nematicides that are both harmful to the environment and possibly human health but often essential for economic potato cropping in the UK. We have developed novel resistance that controls many nematodes including potato cyst-nematodes. This antifeedant approach has been shown to be effective, completely safe in food and benign to the environment. It has been highlighted uniquely by two government reports for its importance to UK agriculture. This work will develop the approach and achieve 90 per cent resistance by stacking antifeedants with peptides that prevent nematode attraction to host roots. The number and diversity of other nematode species will be used as an indication of soil health. We will determine whether or not using the peptides has any effect on non-target nematode species. We will develop methods that will allow nematode diversity to be assessed in a large number of field samples.This will add a further appropriate basis to those we have already deployed. We wish to assess any environmental impact of our technology on soil organisms in the field and allow its comparison with the pesticide use it would replace.

Technical Summary

Plant parasitic nematodes cause losses to global agriculture of 100b Dollars annually. Potato cyst nematode costs the UK potato industry an estimated 43m pounds sterling/year. Control often depends on nematicides that are both harmful to the environment and possibly human health but often essential for economic potato cropping in the UK. We have developed genetically modified, nematode resistant (GMNR) potatoes that have been shown to have no substantial adverse effect on either invertebrate associates of potato above ground or on soil communities. The approach relies on a cysteine proteinase inhibitor already in food, which prevents nematodes from utilising their dietary protein. Toxicological studies establish that a margin of exposure is more than 2000 fold when root specific promoters are used to deliver the cystatin to where nematodes feed while providing low expression of a safe novel protein in food. Work by others has established that the protein is not an allergen. The proposed work will stack that approach with novel peptides that disrupt nematode chemosensory-mediated invasion of host roots. This will involve restricted expression of peptide at sites of invasion and in the rhizosphere. All the components of the stacked defence will be delivered behind promoters that optimise expression at site of invasion and feeding where it is required and limit expression elsewhere i.e. in tubers. New GMNR lines harbouring stacked defences will be challenged in containment and later, the best lines will be tested in the field. In this work we will extend our completed, comprehensive environmental impact studies from our GMNR-cystatin plants to those that disrupt nematode orientation to roots. Non-target nematodes form a dominant group of soil meiofauna, occur in all soil types, with both high abundance and biodiversity. Nematodes play an important role in soil functioning, e.g. in food webs and have been judged as the most promising indicators of soil quality. We will establish base lines that can be applied later to GMNR and other plants in a range of field soils. A recent, promising approach for nematode species identification and comparison between different populations is the use of molecular barcodes derived from PCR amplification, sequencing and analysis of the small subunit ribosomal RNA gene. We will use this approach in parallel with morphological recognition of changes in nematode abundance or prevalence in our soils. The aim of the work is to establish the potential of a fully biosafe GM replacement for current UK use of environmentally damaging pesticides. One novel variant of this technology depends on nematode control via GM allelopathic plants without production of tubers from GM potato plants.
 
Description Potato cyst nematode (PCN), a microscopic worm that attacks potato plants, costs the UK potato industry £43 million/year. Control of PCN relies on chemicals that are often essential for economic cropping in the UK. EU legislation has already resulted in the recent loss of two nematicides in response to environmental concerns. There is an intention to amend Directive 91/414/EEC regulating pesticide use. The UK Potato Council considers this will cause loss from the market of the three remaining nematicides. There is severe doubt that future pesticides with appropriate environmental safety can reach the high control levels that are required.

We have developed a novel resistance strategy involving peptide repellents that prevent attraction of nematodes to host plant roots and so reduce the level of nematode infestation. The main aim of this project was to evaluate the potential of the new technology to provide environmentally safe resistance to PCN in the field.

We first determined how the peptide exerted its effects on the nematode. The location and invasion of host roots by PCN requires them to respond to specific chemicals from the plant. These chemicals are recognised by sense organs that send signals through the nervous system of the nematode. The peptide repellent does not kill the nematodes but blocks the transmission of the signal so that the nematode no longer responds. Using a peptide with a fluorescent tag we showed that it is taken up along the nematode sense neurons and once it reaches connections with other neurons it prevents the nematodes being attracted to potato roots.

We produced potato plants that secreted the repellent from their roots. In some of the plants we controlled the expression of the repellent so that it was only produced in the tips of the roots - the specific region attacked by PCN. We measured the amount of repellent secreted by the roots to identify the best plant lines and tested the resistance of those lines to PCN. The trials were first carried out in a glasshouse and then the plants were evaluated in the field under consent from DEFRA. A number of the potato lines displayed good levels of resistance to PCN in the field with the best line causing a 77% reduction in nematodes. The plants that produced the repellent only in the root tips were more resistant than those where it was produced in all parts of the plant. Limiting the expression in this way will add to the biosafety of the approach.

We assessed the biosafety of the repellent and found that it did not harm a range of soil invertebrates or microorganisms. It is important to show that new transgenic plants do not affect the environment any more than do conventional agricultural practices. The number and diversity of other nematode species in the soil can be used as an indication of soil health. We developed methods that allowed nematode diversity to be assessed rapidly in a large number of field samples without the need for specialist skills in identification. Using these methods we demonstrated that the new technology does not damage soil nematode communities. The transgenic plants had no greater effect on the soil than resulted from cropping oil seed rape rather than potato. One important finding was that field trials are required to collect the necessary data for environmental impact assessments. Containment glasshouse experiments using field soil did not closely simulate effects in the field.

We investigated approaches to increase the level of PCN resistance achieved. We introduced two separate technologies into the same potato plant; the repellent and an anti-feedant. Some plant lines were resistant to PCN but more need to be tested to identify those that have better resistance than the repellent alone. We also showed that when certain brassicas were grown next to potato plants, the multiplication of PCN on the potatoes was reduced. We are continuing to explore the potential of these and other plants to enhance nematode control.
Exploitation Route This project was focussed on developing strategic applications of previous BBSRC-funded fundamental research. The outputs have a clear utility for the implementation of transgenic resistance strategies against plant parasitic nematodes both via the private sector and through public sector involvement in technology transfer to subsistence farmers in developing world countries. The technology for nematode resistant plants developed and evaluated within this project has attracted interest from the private and public sector both in the UK and abroad.

This technology has been donated, together with that already provided by our laboratory, to the developing world through the BBSRC/DFID SARID initiative. We now have transgenic plantain lines expressing some of the constructs first developed in this BBSRC project. The lines display nematode resistance in containment trials and are now undergoing field trial in Uganda. The constructs are also being used within our collaboration on nematode-resistant East African Highland banana as part of a USAID ABSPII project, also based in Uganda.

Plant transformation constructs from this project have been used to develop nematode-resistant transgenic egg-plants in India as part of a RCUK-funded project on sustainable agriculture.
Sectors Agriculture, Food and Drink

 
Description This project was focussed on developing strategic applications of previous BBSRC-funded fundamental research. The outputs have a clear utility for the implementation of transgenic resistance strategies against plant parasitic nematodes both via the private sector and through public sector involvement in technology transfer to subsistence farmers in developing world countries. The technology for nematode resistant plants developed and evaluated within this project has attracted interest from the private and public sector both in the UK and abroad. This technology has been donated, together with that already provided by our laboratory, to the developing world through the BBSRC/DFID SARID initiative. We now have transgenic plantain lines expressing some of the constructs first developed in this BBSRC project. The lines display nematode resistance in containment trials and are now undergoing field trial in Uganda. The constructs are also being used within our collaboration on nematode-resistant East African Highland banana as part of a USAID ABSPII project, also based in Uganda. Plant transformation constructs from this project have been used to develop nematode-resistant transgenic egg-plants in India as part of a RCUK-funded project on sustainable agriculture. The work to investigate the potential of brassica plants for control of nematodes was continued in a PhD studentship in collaboration with the Potato Council. A new award under the HAPI initiative is extending this work in collaboration with academic partners and industry.
First Year Of Impact 2011
Sector Agriculture, Food and Drink
Impact Types Cultural,Societal,Economic,Policy & public services

 
Description BBSRC/NERC HAPI Initiative
Amount £676,030 (GBP)
Funding ID BB/M017672/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 10/2015 
End 09/2020
 
Description Sinochem research funding
Amount £500,000 (GBP)
Organisation Sinochem Group 
Sector Private
Country China
Start 09/2012 
End 08/2015
 
Description Cafe Scientifique (Halifax) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact The talk was followed by a question and answer session and there was general discussion about the pros/cons of GM technology and, in particular, its benefits for the developing world.

Members of the audience reported that their views on GM crops had been changed after participating in the event
Year(s) Of Engagement Activity 2013
 
Description Cafe Scientifique (Hull) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact The talk was followed by a question and answer session and there was general discussion about the pros/cons of GM technology and, in particular, its benefits for the developing world.

Members of the audience reported that their views on GM crops had been changed after participating in the event
Year(s) Of Engagement Activity 2009
 
Description Cafe Scientifique (Leeds) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact A presentation entitled "GM crops: real benefits, real concerns?" was followed by questions from the audience and general discussion

A number of audience members reported a change in opinion after participating in the event
Year(s) Of Engagement Activity 2010
 
Description College visit (York) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Activity raised awareness of a career in science and research

A number of pupils reported that they only realised what a scientific career could offer as a result of my talk
Year(s) Of Engagement Activity 2009,2011
 
Description Frontier Potato Group Presentation 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Talk sparked questions and discussion afterwards: possibilities of commercial direction.

The audience were aware of GM technology that could provide a solution to nematode pathogens
Year(s) Of Engagement Activity 2012,2015
 
Description GM article in Sunday Times Magazine 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact P.U. and H.A. were interviewed for an article in the Sunday Times magazine focusing on developments in and applications of GM crops. The work undertaken in this grant was included in the article. An article was published in the Sunday Times Magazine on June 27th 2010.

no actual impacts realised to date
Year(s) Of Engagement Activity 2010
 
Description Interview for BBC news 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact PU was interviewed by BBC news to provide comment about GM field trials

None reported
Year(s) Of Engagement Activity 2009
 
Description KWS 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Information provided to company regarding new biotechnology

Company were able to make economic decisions
Year(s) Of Engagement Activity 2010
 
Description Parliamentary and Scientific Committee 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Policymakers/politicians
Results and Impact HJA gave a presentation to a meeting of the Parliamentary and Scientific Committee on the topic of "Can GM help to feed 9 billion people by 2050?"
The talk sparked questions and discussion amongst the attendees.

Impacts will be long term in influencing decision making concerning the support for GM crops
Year(s) Of Engagement Activity 2009
 
Description Potato Conference 2010 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact HJA led a workshop on potato cyst nematodes at the Potato Growers Research Association: 21st Annual Potato Conference. The focus of the workshop was "How will consumers affect potato production in the future?" This stimulated interesting discussion between growers/industry/researchers.

The workshop led to useful sharing of ideas and experiences. Direct dialogue with growers informs future research directions for nematode control
Year(s) Of Engagement Activity 2010
 
Description US Embassy Food Security 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact Discussion of Food security issues with particular emphasis on its potential for the developing world

None reported
Year(s) Of Engagement Activity 2009
 
Description University Open Days 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact The work undertaken by the research team in developing nematode resistant crops was demonstrated to visitors. Visitors were engaged in discussions about the work and GM technology in general. Postgraduate students, postdocs and technicians associated with the grants all took part in either preparing or demonstrating the events.

Prospective students encouraged to apply for courses with an applied biology focus
Year(s) Of Engagement Activity 2009,2010,2011,2012,2013
 
Description Worksop College (Nottingham) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Generate understanding an critical thinking about new biotechnological advances

All girl college: students reported a greater interest in continuing in the science field
Year(s) Of Engagement Activity Pre-2006
 
Description Yorkshire Show 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact All members of the research team participated in preparation and delivery of an interactive display at The Great Yorkshire Show. The display explained the problem of plant parasitic nematodes in agriculture and introduced the research being done in our group to both the general public and members of the agricultural community. Members of the group engaged in numerous discussions about our research with a wide audience.

Our exhibits at the Great Yorkshire Show have resulted in an interview on Radio York and also interviews with a number of freelance journalists who provide articles for publications such as the Farmers Guardian.
Year(s) Of Engagement Activity 2012
 
Description cafe scientifique (York) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Talk sparked a lively discussion about the development and use of GM crops

A number of audience members reported that their views on GM crops had changed after hearing the presentation and ensuing discussion
Year(s) Of Engagement Activity 2009