Rasprostranjenost, domaćini i karakterizacija populacije Erwinia amylovora u Crnoj Gori / Distribution, plant hosts and characterization of Erwinia amylovora population in Montenegro

Erwinia amylovora (Burrill) Winslow is the causal agent of fireblight, one of the most destructive diseases of fruits and ornamental plants and one of the most damaging diseases of cultivated plants. In Montenegro, fireblight symptoms were observed for the first time in 1993 on pear trees in the municipalities of Berane and Andrijevica, in northern part of the country. The occurrence this bacterium was experimentally confirmed in 2003 on apple samples from vicinity of Nikšić. Studies of bacterium E. amylovora in Montenegro were carried out in this research. The study comprised of field and laboratory work, conducted in the period 2012‒2015. During fieldwork, the fireblight simptoms were observed on susceptible species, host range and distribution of bacterium in the municipalities was determined. More than 200 plant samples with fireblight symptoms were collected, from which the bacterium was successfully isolated. Widespread of the bacterium was confirmed in whole country, particularly in fruit growing regions in northeastern and western continental part. Presence of E. amylovora was confirmed on five hosts, including four fruit species: quince (Cydonia oblonga), pear (Pyrus communis), apple (Malus domestica), medlar (Mespilus germanica) and one species from spontaneous flora ‒ hawthorn (Crataegus sp.). Monitoring effects of meteorological factors on fireblight symptom occurrence in different climatic regions of Montenegro revealed their high correlation. Quince, pear and apple orchards, as well as single heavily infected trees in northeastern (localities Bijelo Polje and Berane) and western (locality Nikšić) parts of the country, have been confirmed as hotspots from which the bacterium spreads to other areas and new hosts. Presented data points to the fact that in the last few years E. amylovora spread in continental part of the country where it caused significant damages. Therefore, this bacterium presents is a serious threat to successful cultivation of pome fruits, especially because these areas are becoming larger each year in Montenegro. Collected samples plants from different hosts and localities yielded 60 bacterial isolates in laboratory studies. Studied isolates showed pronounced uniformity in pathogenic, cultural, morphological and biochemical‒physiological characteristics and it is confirmed that they belong to the species E. amylovora. Serological analysis (ELISA and IF tests) confirmed homogeneity in antigenic structure within 27 selected strains of E. amylovora originating from quince, pear, apple and hawthorn from different localities in Montenegro. This research carried out the first molecular techniques in studying of E. amylovora in Montenegro. Several molecular techniques were applied: Nested PCR (using two primer pairs AJ75/ AJ76, PEANT1/PEANT2), rep‒PCR (using REP, ERIC and BOX primers) and RAPD PCR (using two primers CUGEA3 and CUGEA5). Molecular analyzes enabled identification and studying of genetic structure of 18 strains of E. amylovora originating from quince, pear, apple and hawthorn, from different localities. The results showed genetic diversion of studied strains from pear compared to all strains originating from other host plants, regardless locality. This is the first study of genetic variability of E. amylovora in Montenegro.