Molecular monitoring reveals two distinct Erwinia amylovora genotypes threatening cultivated orchards and wild Malus sieversii reservoirs in southern Kazakhstan

Fire blight, caused by Erwinia amylovora, poses a serious threat to both cultivated and wild apple populations in Kazakhstan, including Malus sieversii, a key progenitor of the domesticated apple. In 2024, systematic monitoring was carried out in the Almaty and Turkestan regions under meteorological conditions favorable for fire blight development. A total of 131 samples was collected from symptomatic and asymptomatic trees of apple, pear, quince, hawthorn, walnut, and wild M. sieversii. Most samples (77.9%) originated from cultivated apple trees, while 9.2% were collected from wild populations, including from the Sayram-Ugam State National Natural Park, where wild and cultivated apple trees coexist. Molecular screening of plant wash samples confirmed E. amylovora in 31 cases, none of which were collected from M. sieversii trees. Pure bacterial cultures were successfully isolated from 24 samples, while for seven additional qPCR-positive samples pure cultures could not be obtained. Diagnostic confirmation relied on real-time PCR targeting the chromosomal amsC gene and immunochromatographic assays. Genotyping was successfully conducted for 26 positive samples, including 24 isolates and two samples of nucleic acids extracted directly from plant washes. Our results revealed the coexistence of two archetypal E. amylovora CRISPR genotypes in Kazakhstan. Genotype A, previously reported lineage in the Almaty region, was predominantly identified in the southeast, while genotype D, in contrast, was detected as major genotype in southern Kazakhstan. The simultaneous detection of these genotypes suggests a diversification of the E. amylovora population after its first introduction in the country, that could be originating from adaptation to local conditions or from a novel introduction event. These findings underscore the importance of continued molecular surveillance to inform quarantine measures and guide strategies for the conservation of wild apple genetic resources.