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

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

Fire blight cases in Almaty Region of Kazakhstan in the proximity of wild apple distribution area

Fire blight caused by Erwinia amylovora reached Kazakhstan in 2008. Here, the disease poses a threat not only to agricultural production of apples and pears, but also to the forests of wild Malus sieversii, the progenitor of most domesticated apple varieties worldwide. In the period 2019–2021, the spread of fire blight in the growth area of wild apples was limited by the weather conditions. In 2022, late spring and early summer were characterized by increased rainfall and moderate temperatures favorable for the disease. The goal of this study was to monitor the distribution of fire blight in private households and small orchards in the zones adjacent to wild apple distribution areas. A total of 91 samples with fire blight-compatible symptoms were collected from cultural apples (68), wild apple (10), pear (5), hawthorn (7), and quince (1) in south-eastern and eastern Kazakhstan, resulting in 21 isolates (one from pear, one from quince, and 19 from apple) of E. amylovora. All isolates belonged to the archetypal CRISPR genotype A. Considering the relative proximity of the infections to the forests of wild M. sieversii, additional measures for fire blight control and prevention will have to be implemented, including state monitoring of the wild apple forests for disease symptoms and awareness campaigns for specially protected natural territories that safeguard M. sieversii, as well as for local pomaceous-fruit growing communities

Assessment of fire blight introduction in the wild apple forests of Kazakhstan

Fire blight disease of apples continues its worldwide spread, having reached Kazakhstan in 2010. It is a great threat to the wild apple forests of Malus sieversii. The introduction of fire blight is already showing a considerable impact on cultural apple growing and demands radical efforts for conservation of the wild apple forests. A number of studies have been conducted to examine the presence of fire blight distribution within apples in agricultural areas; however, there has been no large-scale monitoring of wild apple tree populations. Here we present the results of three years of monitoring wild apples in three protected areas of Kazakhstan, looking for the presence of fire blight (Erwinia amylovora). A visual inspection showed no signs of fire blight on the trees of M. sieversii in three consecutive years. These findings were confirmed by lateral flow immunochromatography, and conventional and real-time polymerase chain reaction tests of the asymptomatic samples. The findings of this study will be used to produce recommendations for state authorities to prevent fire blight in wild apple forests of Kazakhstan