Nanopore technologies for Septoria Surveillance

Lead Research Organisation: National Institute of Agricultural Botany
Department Name: Genetics and Breeding

Abstract

Septoria tritici blotch (STB) of wheat is the most economically important disease affecting wheat production in Northern Europe. STB is controlled using a combination of agronomy, genetic resistance and fungicides however, selection and subsequent adaptation of the underlying pathogen populations can lead to breakdowns in host resistance and resistance to chemical controls. As the pathogen adapts we will need to understand the composition of the pest population (or pathotypes) to be able to describe specific differences between members of the same species and how these differences affect the range of wheat varieties affected by STB. An effective and cost-efficient solution for STB will directly depend on our understanding of the underlying pathogen (Zymoseptoria tritici).

The use of advanced sequencing technologies has the potential to quickly and efficiently screen many individual pathogen isolates, providing a wealth of information at a low cost. This type of approach has already been demonstrated to be a powerful tool in yellow rust of wheat. However, the cost of purchasing machinery required to conduct sequencing is often prohibitive and runs can take between 24-48 hours to complete. There are also requirements for high-powered computing facilities and experienced operators to conduct the relevant analyses to arrange sequencing data in to a practical format.

NanoPath will employ the MinION sequencer from Oxford Nanopore Technologies as a surveillance tool to improve the speed and ease at which researchers can identify and survey new pests in the field. This will first assess the utility of the MinION and help to develop a model and computational platform to apply nanopore sequencing to a wider range of crop pathogens. The MinION device is portable and able to generate sequencing data following a relatively easy-to-use process. We also propose to implement a data analysis pipeline that will make use of a number of open-source primary data software tools optimised for the discovery and characterisation of genetic variants this crop pathogen.

Technical Summary

Septoria tritici blotch (STB) of wheat, caused by the plant pathogen Zymoseptoria tritici, is the most economically important disease affecting wheat production in Northern Europe. STB is controlled using a combination of agronomy, genetic resistance and fungicides however, selection and subsequent adaptation in pathogen populations can lead to erosion of host resistance. These adaptations are associated to the occurrence of new pathotypes. Understanding the composition of Z. tritici populations and occurrence of different pathotypes is important when attempting to manage the disease effectively, especially when changes in virulence are apparent. Recently, changes in virulence in Z. tritici isolates have been reported in the UK. There is a large significant increase in the severity of disease symptoms when plants with partial resistance were inoculated with the new Zt isolate resulting in a higher frequency of dead leaves and a greater number of lesions than compared to the historic isolate. As suggested by experiments we conducted the new isolates are able to reproduce asexually having eroded the resistance in the host. NanoPath will employ the MinION sequencer from Oxford Nanopore Technologies as a surveillance tool to improve the speed and ease at which researchers can identify new Z. tritici isolates in the field. We will first assess the utility of the MinION for STB pathotyping and help develop a model and computational platform to apply nanopore sequencing to a wider range of crop pathogens. We also propose to implement a data analysis pipeline that will make use of a number of open-source primary data software tools optimised for the discovery and characterisation of genetic variants in Z. tritici. Concurrently, Illumina short read chemistry (HiSeq 4000) will be used to sequence the same isolates (10-20x coverage) as a strategy to validate the quality of reads produced on the MinION.

Planned Impact

The delivery of the NanoPath's objectives is focused on current diagnostics and surveillance needs for which suitable tools are lacking. In this way achieving impact is an integral part of the proposal. The recent advances in sequencing technologies have opened up new opportunities for the implementation of targeted approaches for the characterisation and surveillance of pathogens in general. The Programme's objectives will directly contribute solutions in areas of research relevant to the BBSRC scientific priority in food security and integrate novel methods in genomics with emerging data analysis protocols.

Academic, Economic and Commercial Impacts
The development of the NanoPath platform will generate new opportunities for collaborative work with R&D groups in industry and academic institutions. The characterisation of the genetic diversity in Septoria, for instance, will enable and support the design of novel approaches to control this crop disease which is one of the most important in Northern Europe in terms of economical losses.

Similarly the infrastructure and technology advances described in the objectives will have a direct effect in key UK R&D areas such small biotechnology industries and academic labs with limited access to compute power. The use of the MinION device coupled with the development of bioinformatics analysis pipelines that can be executed in cloud-based systems will be accessible to a great number of researchers with limited requirement for laboratories or computing facilities.

The transformative effect of the availability of large datasets in the development of technologies and services is undoubtedly one of the main drivers supporting the biotechnology industry. One example is the effect that genomics assisted methods will have the understanding of the genetic diversity in crop pests. The availability of pre-competitive data generated by NanoPath will have a direct impact on research in the crop pathogens area.

Societal impacts
The implementation of NanoPath will directly impact the agricultural sector with the potential to indirectly support improved yields by increasing productivity.

It is widely recognised that the shortage of expertise and skill in biomathematics and informatics in the UK and across the world is a major risks for a future development of data-driven science. In this context and in a modest measure we hope to support the training of talented computational biologists by demonstrating the value the MinION (or more general nanopore technologies) to implement effective surveillance strategies.

Policy: BBSRC, research councils and UK
NanoPath will contribute to reinforce the UK's reputation of leaders in the agri-food sector by supporting the use of cutting-edge technologies to monitor plant crops. The availability of reliable monitoring and surveillance tools will be one of the key pillars to support the most modern crop disease controls.

Publications

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Wood T A (Article) Applying new technology to pathogen surveillance in NIAB Landmark Magazine - Quarterly publication distributed to network membership (>700 subscriptions)

 
Description We have generated genome assemblies for more than 10 Septoria strains. These assemblies are the basis for the downstream analysis which will be focused on the identification of genetic/genomic markers for pathogenicity. This project also supported the development of novel plant disease surveillance approaches using inexpensive portable sequencing technologies.
Exploitation Route The sequences will be made available and a scientific manuscript will be accessible to the wider research community.
Sectors Agriculture

Food and Drink

Digital/Communication/Information Technologies (including Software)

 
Description The development of effective plant disease surveillance strategies is a fundamental objective of this project. The findings of this project are currently informing the work with collaborators in the UK to implement a service to support advanced, effective and accurate disease surveillance.
First Year Of Impact 2019
Sector Agriculture, Food and Drink
Impact Types Economic

 
Description BSPP Summer undergraduate internship
Amount £3,800 (GBP)
Organisation The British Society of Plant Pathology 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2021 
End 09/2021
 
Title RNAseq data set from compatible/incompatible interaction w/ Zymoseptoria tritici on contrasting Wheat cultivars Cougar and Gallant 
Description Transcriptomics was used to investigate differences in gene expression in virulent/avirulent Zymoseptoria tritici interactions on susceptible and resistant cultivars of UK wheat 
Type Of Material Technology assay or reagent 
Year Produced 2020 
Provided To Others? No  
Impact Enhanced knowledge and insight into the genetic basis of virulence in 'Cougar-type' Zymoseptoria tritici 
 
Title Re-sequenced genomic data for 44 Zymoseptoria tritici isolates 
Description 44 Zymoseptoria isolates have been re-sequenced providing extensive genomic data for conducting phylogenetic and comparative genetic studies. A pangenome has also been developed using the data set. 
Type Of Material Technology assay or reagent 
Year Produced 2018 
Provided To Others? No  
Impact Comparative genetic analysis between members of the panel have demonstrated that it is not possible to distinguish between contrasting pathotypes based on genetic profile alone, particularly with only a small number of isolates. This led to us developing additional transcriptomics datasets and mapping resources in order to elucidate the factors contributing to virulence in the host. 
 
Description Proyecto Omicas / Omics project 
Organisation Pontifical Xavierian University
Country Colombia 
Sector Academic/University 
PI Contribution Participation as an associated collaborator. The research program optimization Multiscale In-silico Agricultural Crops Sustainable (omics) is the winner of the second round of Colombia Science in focus Food which seeks, through seven projects, develop and implement strategies science and technology to improve agricultural varieties with the aim of contributing to food security and sustainable production worldwide .
Collaborator Contribution Omic is made up of researchers, undergraduate and graduate students , and support staff from 16 institutions, including institutions are higher education (IES) accredited, national and international, IES national non - accredited, international research centers in agriculture and domestic enterprises in the industrial sector, with the Pontificia Universidad Javeriana Cali anchor entity .
Impact This is a multidisciplinary project that involves collaborators working on genetics, genomics, bioinformatics, agriculture, phenotyping and social sciences. The Colombian government is also directly engaging with the collaborators.
Start Year 2019
 
Description Invited Speaker at the 10-year celebration of the Centre of Excellence in Genomics (ICRISAT, India) 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact 10-year anniversary of the Centre of Excellence in Genomics. This event congregated researchers and industry stakeholders from across the globe working in crop genomics.
Year(s) Of Engagement Activity 2017
URL http://www.icrisat.org/celebrating-a-decade-of-genomic-advances-in-agriculture/?utm_content=buffer55...
 
Description Key Note Speaker at VIII Argentinian Bioinformatics and Computational Biology Congress 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Key note speaker - meeting was followed by a number projects to apply for GCRF funding to work with researchers in Latin America
Year(s) Of Engagement Activity 2017
URL http://ufq.unq.edu.ar/8cab2c/
 
Description Myanmar - Green Fruits Dedicated Workshop 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact The Workshop, dedicated to Green Fruits in Green Horticulture, was organized in the context of (the ASEAN Institute for Green Economy, AIGE Component on Increasing and Improving Green and Blue Productivity and Sustainability The Dedicated Workshop was forward looking, taking into account fast emerging and converging regional and international trends on the imperative need for green and clean economy, articulated and adopted in an increasing number of national, regional and international for.
Year(s) Of Engagement Activity 2018
URL http://geggmyanmar.org/green-fruits-dedicated-workshop-report/
 
Description NIAB Director's day 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact I presented at the NIAB Director day - the focus of the presentation was on describing the application of new technologies to agriculture. I engaged with a diverse audience - explaining the advantages that new technologies could bring to agronomical practices.
Year(s) Of Engagement Activity 2017
 
Description Talk on pathogen genomics at NIAB Science Day 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Professional Practitioners
Results and Impact The nanopath project was presented in a talk discussing how genomics is being utilized to enhance pathogen surveillance, monitoring and detection. The talk was attended by approx. 80 delegates from NIAB/NIAB-EMR. The talked raised a number of questions from the audience and provided an excellent opportunity to demonstrate how the technology provides accurate and rapid pathotyping capability for crop pathogens such as Septoria.
Year(s) Of Engagement Activity 2017
 
Description Talk providing overview of the project 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact A talk was delivered to postgraduate students and professional researchers during a phytopathology symposium at Wageningen University. The talked sparked some good discussions on the practical implementation of pathogenomics for pathogen surveillance, specifically using nanopore sequencing.
Year(s) Of Engagement Activity 2018
 
Description Talk providing overview of the project 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact A talk providing an overview of the Panacea project was delivered to post-graduate researchers at NaCRRI in Uganda, highlighting the aims of the project, the new resources that will be generated and the importance of engaging with local researchers and experts to leverage impact from them. The talk generated some fruitful discussions on the relevance of pathogenomics approaches for providing improved motoring of crop diseases and how this type of strategy could be extended to other countries growing phaseolus bean and additional pathogens to create even wider benefits from the technologies.
Year(s) Of Engagement Activity 2018
 
Description Talk to staff members at NIAB-EMR 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Professional Practitioners
Results and Impact Talk on crop pathogen diagnostics and monitoring given to research staff members at NIAB-EMR, Kent. Staff were briefed on the Nanopath project and how nanopore sequencing is being used to screen for genetic differences in Zymoseptoria tritici
Year(s) Of Engagement Activity 2017