The molecular and genetic basis of aphid virulence

This project aims to advance our understanding of the genes involved in molecular interactions between aphids and their host plants. This is important because aphids are at the top of the global league table of insect pests that damage food and non-food crops, causing losses of hundreds of millions of dollars every single year. In addition to sucking sap, they are major transmitters of viruses, and necessitate extensive use of costly pesticides. Some of these chemicals will be withdrawn under EU legislation and others are no longer effective due to aphids becoming resistant, adding further urgency to develop new robust but sustainable routes to crop protection. The route we propose centres on genetics rather than chemicals. Most aphid species have a defined range of host plants on which they can successfully feed. Within those aphid-host species pairs, some genetic variants on both sides can dramatically affect the outcome: aphid success is known as virulence, and successful plant defence is known as resistance. Although some specific resistance to aphids most likely operates in a "gene-for-gene" manner similar to better known mechanisms acting against diseases, we do not yet understand the details of the interactions between the aphid virulence proteins, collectively known as effectors, and plant resistance proteins. This knowledge gap hampers progress toward crop genetic improvement. Because of huge advances in gene sequencing technologies and genomic databases for both pea aphid (Acyrthosiphon pisum) and one of its hosts, barrel medic (Medicago truncatula), there is for the first time the opportunity to study genes in both species as a route to defining those responsible for success of the aphid or resistance of the host.
Our recent work has generated vital knowledge and resources that make the proposed project feasible. First, we have made a series of novel aphid hybrids by mating virulent and non-virulent parents. Several of these hybrids exhibited virulence patterns not seen in either parent, which suggests that there are complex patterns of inheritance of virulence genes. Second, our pilot studies show that there are major differences between gene expression patterns and protein profiles between virulent and non-virulent aphids. Together, these findings enable an efficient route to narrow down the effectors responsible, and can greatly aid our understanding of how resistance to aphids works. Our aim is to define the major genes that determine aphid success or host resistance. Such knowledge can potentially lead to new routes to generating durable resistance in a wide array of crop plants. We will test how closely the mechanisms uncovered in the pea aphid-medic system are also relevant to other major UK pests such as peach-potato aphid (Myzus persicae), which attacks a very wide range of crops and for which little or no resistance is known.
The project has four major activities. First, we will generate further hybrid aphid populations that differ in virulence. These aphid sets will allow us to discover the major common differences across multiple virulent and non-virulent aphids. We will measure amounts of expressed gene products, both as RNA and as protein, and simultaneously look across the entire pea aphid genome for DNA sequence variants that may affect protein function. Second, we will test the top candidate genes and proteins by inserting them into medic plants to see if they specifically mimic host infestation symptoms and/or affect aphid virulence. Third, we will learn more about host resistance processes by characterising a medic gene that gives particularly strong resistance to some pea aphid types. Finally, we will compare pea aphid effectors with the their equivalents in peach-potato aphid, and will test whether pea aphid effectors inserted into host plants have the potential to affect resistance to other species, especially peach-potato aphid.

Although aphids are major global crop pests, the mechanisms by which they successfully infest plants remain unknown, and very few aphid resistance genes have been defined in crops. We predict that salivary secretions include effector proteins and other molecules that during probing and feeding will result in suppression of host defences by virulent genotypes, or will trigger R-gene dependent resistance against avirulent genotypes. We will exploit the advantages of the pea aphid-Medicago truncatula model system to attempt to make a step change in the understanding of genetics and genomics of aphid virulence. Specifically, we have demonstrated Mendelian inheritance of at least one major determinant of virulence, and have shown that both the transcriptomes and the proteomes of virulent and avirulent aphids differ substantially in candidate secreted effector proteins. We will develop further segregating F1 and F2 aphid populations for assessment of virulence phenotypes. Whole transcriptome analysis of bulked segregant aphid clones by RNAseq and parallel proteomics will narrow down candidate effector genes, through detection of sequence polymorphisms, and comparison of RNA and protein level between virulent and avirulent groups. The top aphid candidate genes will be evaluated by delivery into Medicago leaf tissue, looking for effects on aphid performance and on plant response phenotypes. We will initiate translational studies to other aphid species of major UK importance, specifically testing the extent of conservation of effector gene functions between the specialist pea aphid and the generalist Myzus persicae. We will advance our understanding of cognate host resistance genes by characterising the function of RAP1, a major aphid resistance locus. Together, the outputs of this project will define the potential for genetic rather than agrochemical routes to crop pest control.

Economic importance of the problem we are addressing:
Aphids infest many crops worldwide, causing extensive direct damage and transmitting many economically important plant viruses. Economic losses due to such pests in the UK alone are in the order of £100 million a year. As highlighted by the Horticultural Development Company (HDC), there is a need for new chemistries with improved environmental profiles, and for non-chemical approaches to protect field crops from pests and diseases.
Global problems:
Insect infestations are expected to increase due to the combined impacts of climate change, pressure to reduce pesticide use, and intensification of agriculture. Up to 50% of crop losses in developing nations result from plant pests and pathogens according CABI. With the pressing need for sustainably increased food production worldwide, there is unprecedented demand to develop new strategies to protect crops from pests and diseases.
Scientific impacts:
Our combined advances in understanding of inheritance of aphid virulence and effector functions open up new opportunities for scientific advances. This potential is greatly augmented by availability of pea aphid and Medicago genomic sequence and gene expression resources, providing opportunities to increase our understanding of the molecular fundamentals of gene-specific defences. Wider benefits from deep understanding of the pea aphid - Medicago system can emanate from our testing of translation into other aphid species.

In addition to the several academic beneficiaries described above, the proposed research is therefore expected to benefit i) agri-food and related industries, ii) research staff, iii) the general public.
The proposed project will benefit agri-food and related industries by enabling development of novel effective control strategies against crop pests. We have had contact with key crop groups through AHDB representatives for horticulture, potato and cereal sectors, and with BBRO, the UK sugarbeet research organisation.
The project aims to identify aphid proteins that are essential for virulence and/or trigger immunity. Such proteins are potential targets for the development of novel control strategies in economically important crops based on, for example, RNAi or novel chemical compounds. In addition, this project will characterise plant proteins involved in resistance.
Career benefits:
Research staff involved in the project will benefit in terms of career development. The research draws on a vast array of molecular biology and biochemistry techniques that represent transferable skills in the biological sciences. With results of the research expected to be of high impact and of great interest to the research community, this project will likely generate multiple publications in peer-reviewed journals as well as lead to invitations to national and international meetings. In addition, opportunities will be provided to young talented undergraduate and postgraduate students to receive training within the research groups involved in this project, aiming to inspire and engage potential future academics in biological sciences research.
Environmental and health benefits:
A reduction in the use of insecticides will benefit the general public. Currently, aphid control strategies, and those of other plant pests, rely on extensive use of such chemicals, some of which pose a threat to human health and the environment. By instead using genetics to work towards robust resistant crops, use of chemicals is potentially reduced. Such benefits are within the framework of sustainable agriculture, contributing to protection of natural environments and reducing the environmental impact of agricultural practices.