Occurrence, identification and phylogenetic analysis of Fusarium proliferatum on bean seed (Phaseolus vulgaris L.) in Serbia

Abstract

During the routine quality control analysis on bean seeds in 2015, Fusarium fungal infection was observed on an average of 17% of the bean seed. The objective of this paper was isolation and identification of Fusarium sp. based on the pathogen's morphological and molecular characteristics. Morphological identification of Fusarium isolates was performed on PDA and CLA. DNA of 14 Fusarium sp. isolates was extracted directly from the mycelium (~ 100 mg wet weight), with a Dneasy Plant Mini Kit (Qiagen, Hilden, Germany). Following DNA extraction, the translation elongation factor 1-alpha region was amplified by PCR with the primer pair EF1 and EF2. An amplicon of 700 bp was amplified in all tested isolates. Identification of one isolate was performed by sequencing the translation elongation factor EF-1a gene. Completed morphological and molecular characteristics of isolates, as well as the results of sequencing confirmed that Fusarium proliferatum was the causal agent of bean seed rot.

Tokom rutinske kontrole kvaliteta semena pasulja (Phaseolus vulgaris L.) u 2015. godini ustanovljeno je prisustvo Fusarium sp. u visokom procentu, u proseku 17%. Cilj ovog rada bio je izolacija i identifikacija prouzrokovača truleži semena pasulja, na osnovu morfoloških i molekularnih karakteristika patogena. Odgajivačke karakteristike 14 odabranih izolata ocenenjene su na PDA i CLA podlogama. Ekstrakcija DNK 14 odabranih izolata obavljena je direktno iz 100 mg sveže micelije, korišćenjem Dneasy Plant Mini Kit (Qiagen, Hilden, Nemačka). Amplifikacija DNK obavljena je pomoću PCR korišćenjem para prajmera EF1 i EF2. U svim proučavanim izolatima formirani su amplikoni veličine 700 bp. Identifikacija jednog odabranog izolata izvršena je sekvenciranjem translacionog faktora EF-1 a gena. Na osnovu dobijenih rezultata ustanovljeno je da je prouzrokovač truleži semena pasulja Fusarium proliferatum.