Bean bacterial wilt is one of the serious and damaging diseases of beans, which has been reported in recent years from Iran. The disease characterized by interveinal necrosis, foliar blight and wilting of the bean plants due to clogging of the xylems by the causal agent, Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff).The strains of Cff are seed-borne pathogens and posse the potential to spread the disease to the next season’s crop in new area. Since the bean seed is produced and traded across the country, sampling and detection of cff contaminated seed is necessary to reduce the risk of pathogen spread through seed lots. To trace Cff in bean seeds, discolored and wrinkled bean seeds were collected from various regions of Iran. Five seeds from each sample were soaked for 12 hours in 10 ml buffer at room temperature with gentle agitation by shaking at 100 rpm. A loop full of each suspension was streaked on Nutrient agar and semi selective C- medium containting 10g peton, 1g Casein, 0/25g MgSo 4 .7H 2 O, 18g Agar, 10g Licl and 5g maltose. After incubating at 28? for two days, the yellow colonies with different morphologies were picked and recultured. In the preliminary examination, the genomic diversity analysis of the obtained isolates was carried out with the use of BOX IR and ERIC1R/2F fingerprinting methods. Both the ERIC and BOX PCR amplification data revealed the highly genetic heterogeneity of the isolates, regardless of their regional origin. Based on the corresponding dendrogram, the isolates clustered in five separate groups I (13 isolates), II (14 isolates), III (16 isolates), IV (six isolates) and V (four isolates) with the exception of strain BS54 which showed unique distinct pattern. To achieve a genus level identification of the members of each fingerprint group, nearly full – length 16S rRNA region of the chosen isolates were amplified using the prokaryotic specific primer pair 27F/1492R. Sequencing the 1500bp fragment of the isolates, alignment of the sequences and their comparison with the available 16S rRNA sequences of the prokaryotes in GenBank database showed that the isolates of groups I, II and III are related to Siccibacter sp., Enterobacter sp. and Kosakonia sp. (%99 similarity) which are known collectively as epiphytic bacteria. The members of distinct groups IV and V have strong homomology (%99.72) to the plant pathogenic bacteria Xanthomonas axonopodis pv. phaseoli and) and Curtobacterium flaccumfaciens pv. flaccumfaciens , respectively. The BS54 isolate was different from those groups and identified as Pantoea sp. (with %98 similarity to Pantoea agglomerans ). Further detailed morphological, physiological, biochemical and pathological tests results ensured that the yellow pigmented, Gram positive,oxidase negative isolates of group IV with sensitivity reaction on tobacco leaves are capable to induce typical wilt symptoms in bean plants. Cff specific primer pair (Cff-FOR2/Cff-REV4) also amplified the predicted 306 bp DNA fragment from all Cff isolates of group IV tested, which confirmed their taxonomic affiliation to C . flaccumfaciens pv . flaccumfaciens . The Cff strains, isolated mainly from the seeds taken from Isfahan and Markazi provinces, were found homogenous with respect to their biochemical and molecular characteristics. Considering the long term survival of Cff (at least 13 months in present research) in bean seeds and the importance of high capacity spread of the bacterium through bean seeds, it is necessary to develop an easy, sensitive and culture independent method for rapid detection of Cff in bean seeds. So far, only one PCR protocol using primer pair Cff-FOR2/Cff-REV4 with yield of 360bp fragment has been described for identification of Cff strains, which is too small for sequencing to validate the result. In order to develop a new PCR method for detection of Cff on bean seeds, two primer pairs (Cff1F/Cff1R and Cff2F/Cff2R) designed according to the DNA sequence of the 1500bp16S rRNA fragment of the reference isolate Cff-JSM05. The results for optimization and evaluation of specific detection of Cff by these primers demonstrated that Cff1F/Cff1R and Cff2F/Cff2R are able to amplify fragments 911bp and 603bp in Cff isolates from pure culture and artificially or naturally infested seeds. The results indicated that the Cff specific primers described in present study could be useful for the specific and rapid detection of Cff strains in bean seed lots. Key words Bean Bacterial wilt, Curtobacterium flaccumfaciens pv. flaccumfaciens , ERIC and BOX IR fingerprinting, specific primer, PCR detection