Tracking the dissemination of Erwinia amylovora in the Eurasian continent using a PCR targeted on the duplication of a single CRISPR spacer

Mirjam Kurz,Mila Grahovac,Tinatin Doolotkeldieva,Nataliya Drenova,Mery Dafny-Yelin,Theo H M Smits,Fabio Rezzonico,Milan Ivanović,Alexander L Lagonenko,Simon Carnal,Galiya Zharmukhamedova,Richard A Gottsberger,Orly Mairesse

doi:10.1186/s42483-021-00096-9

Abstract

Fire blight is the most devastating disease affecting pome fruit production globally. The pathogen is native to North America and was imported to western Europe in the 1950s, progressively spreading over the continent in the ensuing decades. Previous phylogenetic studies have revealed the extreme genetic homogeneity of the pathogen outside its center of origin, which makes epidemiological studies difficult. These are generally only possible using hypervariable regions of the genome such as those represented by CRISPRs (Clustered Regularly Interspaced Short Palindromic Repeats), which are, however, not practical to sequence due to their size and variability. Here, we present a simple PCR assay targeting the duplication of a single CRISPR spacer in Erwinia amylovora that was found to be an important marker to discriminate between two main European populations of the pathogen. We implemented the assay on a total of 582 isolates to follow the spread of fire blight across the continent over several decades and, wherever possible, within single countries. The results obtained point to the occurrence of two major separate introduction events for E. amylovora in Europe that occurred approximately 20 years apart, and confirmed the existence of two principal distribution areas located in Northeastern Europe and the Eastern Mediterranean Basin from which the pathogen moved on to colonize the Eurasian continent.

Highlights

Fire blight, caused by the enterobacterium Erwinia amylovora, was first described on apple trees in the Hudson Valley (State of New York) in the 1790’s
E. amylovora contains two seemingly poorly active Clustered regularly interspaced short palindromic repeats (CRISPR) repeat regions (CRR1 and CRR2) with their associated cas gene cluster, alongside with a third region (CRR4) that is apparently a relic of a second lost CRISPR/Cas system, which is present in other pathoadapted Erwinia species like Erwinia pyrifoliae, Erwinia tasmaniensis and Erwinia piriflorinigrans (Rezzonico et al 2011; Smits et al 2011, 2013)
CRR1 genotypes in E. amylovora genomes Analysis of the genomes of E. amylovora strains from Europe, the Mediterranean area, Asia, North America and Oceania revealed the presence of 16 distinct genotypes in CRR1 (Fig. 1)

Summary

Introduction

Fire blight, caused by the enterobacterium Erwinia amylovora, was first described on apple trees in the Hudson Valley (State of New York) in the 1790’s This severe disease, which affects plants belonging to the family Rosaceae, is thought to have originated in North America, moved to New Zealand, where it was first detected in 1910, and arrived in the United Kingdom in 1958, before (2021) 3:18 effect, resulting from a restricted number of dissemination events of so-called CRISPR group I strains radiating from the East Coast of the United States (Rezzonico et al 2011; McGhee and Sundin 2012) or possibly even introduced from New Zealand (Bühlmann 2015). Strains that display recent incorporation of new spacers at the 5’end or show deletions/ duplications in the internal region of the array cannot be ancestral (Additional file 1: Figure S1b)

Methods

Results

Conclusion