Bovine digital dermatitis (BDD) is a contagious infectious disease which causes lameness in dairy cows. It has a multifactorial etiology which is not yet fully understood but Treponema spp. seem to play a significant role in development of BDD lesions. This study evaluated the presence of Treponema phylotypes commonly associated with BDD (T. medium/T. vincentii, T. phagedenis and T. putidum/T. denticola), in four farms different areas in Iran. Single biopsies were taken from 113 Holstein cows with active BDD lesions (scored according to size) on the farms and polymerase chain reaction assays used to detect 16S rRNA nucleotide fragments of three BDD Treponema phylotype groups:‘‘T.medium/T.vincentii’’,‘‘T.phagedenis’’ and ‘‘T.putidum/T.denticola’’ (now T. pedis). Over 95.00% of samples were positive for at least one of phylotypes, with 89.00%, 91.00 %, and 66.00% of samples were positive for T.putidum/T.denticola,T.phagedenis and T. medium/T. vincentii, respectively. Out of the 113 samples, 60.00% were positive for all three phylotypes, the detection of T. putidum/T. denticola was positively associated with detection of both T. phagedenis and T. medium/T. vincentii. No association between lesion size and phylotypes identified was found but there were significant differences between farms in the proportion of each phylotypes identified. Further research is required to establish the factors influencing the proportions of individual phylotypes, especially at the farm level

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