Pathogenicity and Genetic Diversity among the Strains of Clavibacter michiganensis subsp. insidiosus Causing Alfalfa Wilt Disease, Hamedan Province

Alfalfa (Medicago sativa) is among a few of the most important forage crops for the growth of which Hamedan province benefits from a suitable climate. Alfalfa bacterial wilt disease induced by Clavibacter michiganensis subsp. insidiosus cause substantial economic crop losses. This disease has not been so far investigated extensively in Hamedan province. In the course of this survey affected plant samples were collected from alfalfa growing areas in Hamedan province, and causative bacterial agents isolated. Pathogenicity of the strains was confirmed on plants and their phenotypic features characterized as based on standard bacteriological methods. Bacterial strains induced symptoms on tested alfalfa plants after a passage of 40 days. Pathogenic strains were identified as C. m. subsp. insidiosus. Effect of the strains on shoot and root length, wet and dry weight of the plant parts were evaluated in a randomized design in four replicates. The obtained data were analyzed through SAS software. Results indicated that all the tested strains reduced the above-mentioned traits showing significant differences in this regard. To determine of the genetic diversity among the tested strains Opa-03, Opa-15, Opb-10, Opb-15, Opb-18, Opb-20, Ope-01, Ope-02, Ope-03, Ope-04, Ope-07, Ope-14, Ope-16 and Ope-19 primers were picked up and clustering was performed. Results revealed four clusters comparised of: AH7, AH8, AH9 and AH6 strains (first cluster), Ah15 and AH2 strains (second cluster), AH10, AH11, AH12 and AH14 strains (third), and AH1 and AH3 (fourth cluster). EcoR1 digestion of the amplified fragment using CIRS-1 and CIRS-2 primers revealed two groups of strains. All the obtained data revealed genetic diversity among the C. m. subsp. insidiosus strains collected from Hamedan province.