Combination of next generation sequencing and metagenomic analysis in the diagnostics of severe hop stunting

Client ID: Agencija za raziskovalno dejavnost RS (J4-4153)
Project type: Basic Research and Application Project
Project duration: 2011 - 2014


In 2006, hop growers in the Savinja valley reported the appearance of hop plants showing stunted vigor. Such symptoms were visible in various cultivars, showing the disease agent’s ability to affect different hop genotypes. Infected plants reached only up to 50% of normal height. Symptoms were even more severe the next year and affected plants eventually declined. It was confirmed that the disease was spreading around the first observed point of infection, suggesting agrotechnical practice as the most likely cause of transmission of the disease to healthy plants, although disease spread by means of insect vectors could not be ruled out. Infected planting material exchange between hop farmers has been confirmed as a source of new outbreaks over longer distances.

The speed of spread and severity of this disease require quick action. In the proposed project, we aim to accomplish several tasks, which will subsequently allow prompt identification of the pathogen, by developing a method for its routine detection, comparing the transcriptome response of diseased and healthy plants and establishing guidelines for proper disease management.

In order to identify the pathogen, we propose to use a combination metagenomic approach of sequence comparison and next generation sequencing technology (NGS), which will allow us to identify as yet unknown pathogens present in stunted hop plants. There have been several successful studies utilizing metagenomics coupled with NGS for the diagnostics of human, animal and plant diseases, which leads us to believe that this approach will allow prompt novel pathogen detection or show a synergistic infection of several pathogens. On the basis of the retrieved sequence, we will develop a diagnostic method for routine detection of the pathogen. We will then study the epidemiological characteristics of the disease, aimed at clarifying whether weed plant species present in hop gardens are possible reservoirs of the pathogen or contribute its spread, and whether the pathogen can spread in a hop garden by means of mechanical injuries to plants.

In addition to identifying the pathogen(s), we propose to study differential gene expression in transcriptomes of three experimental groups. To understand the molecular mechanisms underlying host-pathogen interactions, it is necessary to identify the gene transcripts differentially expressed in infected and healthy plants. This study will be carried out using an RNA-seq analysis of the three described experimental groups and will include de novo assembly of the hop transcriptome. We anticipate that the proposed combination of metagenomic analysis of NGS data and RNA-seq studies will result in successful identification of the novel hop pathogen or explain the synergistic infection of several pathogens, while providing a detailed insight into the transcriptome of infected hop plants.