MARPLE Diagnostics

MARPLE Diagnostics provides real-time, point-of-care plant disease diagnostics and surveillance for complex fungal pathogens.

Working with obligate biotrophic pathogens that can’t be separated from their host and with genomes that are > 10,000 larger than for instance the Ebola virus we have developed a simple, mobile diagnostics platform.

Furthermore, with the whole system able to fit in a suitcase we have shown (read more here) it can be run in any lab irrespective of existing infrastructure, an office or even in the back of a car directly in the field

Welcome to the future of plant disease diagnostics and surveillance

Project Partners

Diane Saunders

John Innes Centre

Dave Hodson

CIMMYT

Tadessa Daba

Ethiopian Institute of Agricultural Research

Tesfaye Disasa

Ethiopian Institute of Agricultural Research

Rapid Mobile Diagnostics and Surveillance

Fast: From field sample to strain level identification within 48 hours
Mobile: Entire lab set up and pipeline can be transported in a single hard case
Resilient: Pipeline from field to sample can run on inconsistent power and uses reagents that can be kept at room temperature.
Simple: Easy-to-follow procedure with no prior expertise required.

Complex Fungal pathogens

The MARPLE platform was built initially for wheat yellow rust, which is an obligate biotrophic pathogen that can’t be separated from its wheat host and has a genome > 10,000 larger than for instance the Ebola virus. To address this, we developed a strategy to sequence specific regions of the pathogens genome that can be used to define individual strains and monitor key genes for mutations such as fungicide target genes.

Impact

This new, innovative, data-driven approach provides near real-time data on the rapidly shifting nature of pathogen population structure and dispersal that can feed directly into early-warning systems and control recommendations.

The Pipeline

Sample Collection

Crops showing fungal disease symptoms are identified in the field and small (< 1 cm²) leaf samples collected. These samples are processed using the MARPLE pipeline to confirm the causal pathogen and define the individual strain, whilst also monitoring important genes for mutations such as fungicide target genes.

DNA Extraction

Simple DNA preparation method: DNA is extracted from pathogen-infected leaf samples using a simple method that is independent of any complex, additional equipment. Following mechanical disruption of the leaf tissue, DNA is cleaned using a simple bead-based method to provide high-quality DNA ready for PCR.

MiniPCR

Mobile PCR amplification: Regions of the pathogens genome that are informative for defining individual strains and target genes of interest are amplified using the polymerase chain reaction (PCR). Currently, a total of 242 genes are PCR amplified across four pools.

Library Preparation and MinION Run

No cold-chain storage: Fragments of DNA generated from the PCR step are prepared for sequencing using a simple kit developed by Oxford Nanopore that can be shipped at room temperature to enable easy transfer between locations.

Hand-held sequencing device: Once prepared for sequencing DNA is loaded onto the MinION sequencer and the genetic code of these DNA fragments revealed.

Data Analysis

Offline data analysis: The genetic code for the regions of the pathogen’s genome that can be used to define strains is compared between samples using an analysis pipeline that works independently of internet connectivity.

Comparative analysis with data from known strains helps define the individual strain of the pathogen in the sample and thereby its likely threat to different crop varieties. Analysis of additional genes such as fungicide target genes can also give a real-time indication of likely breakdowns in their utility.

Access training materials

You can now access the training materials for the MARPLE pipeline. A full guide can now be found below

Download MARPLE training guide