Identification and Differentiation of Egyptian Isolates of Erwinia amylovora using 16S rRNA Gene and RAPD-PCR

Abstract

Erwinia amylovora the causative of fire blight disease on pear in Egypt was identified with some classical tests and molecular methods. Random Amplified Polymorphic DNA (RAPD) technique was used to identify the genetic variability among 6 Egyptian isolates of E. amylovora isolated from different governorates (Behera, Gharbiya, Nubariya, Menufiya, Kaluobiya and Ismailiya) and compared with one reference isolate. Profiles were obtained by amplification of DNA from these isolates with six random primers.The RAPD band patterns resulting from these primers were analyzed using UPGMA method to construct a similarity matrix and generate a dendogram indicating the relationships between the examinedisolates. The results obtained indicated that primers P4d, P5e and P6f were chosen because they reliably and reproducibly detected polymorphism among the selected isolates. Also, no variation in bacterial genome was observed among the examined isolates. In addition, 16SrRNA gene sequence was used to identify E. amylovora isolates using universal primers (8F and U1492R). The results showed that sequence of the isolates collected from different locations was matched with the sequence found in the Genbank database. The sequence of the fragment (16S rRNA gene) showed percentage homology with strain DSM30165 of Genbank strains ranged from 89–97%. The selected isolates were found to have theclosest homology to E. amylovora.