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Although physicians currently rely on clinical signs along with non\u2010specific biomarkers of infection, such as erythrocyte sedimentation rate and C\u2010reactive protein, to diagnose and monitor DFU, there is no specific and sensitive measure available to monitor or prognosticate the success of foot salvage therapy (FST). To address this we performed a prospective, observational microbiome analysis to test the hypotheses that: (i) the initial microbiomes of healed versus non\u2010healed DFU are distinct; (ii) the microbial load, diversity and presence of pathogenic organism of the DFU change in response to antibiotics treatment; and (iii) the changes in the DFU microbiome during treatment are prognostic of clinical outcome. To test this, microbiome analyses were performed on 23 DFU patients undergoing FST, in which wound samples were collected at zero, four, and eight weeks following wound debridement and antibiotics treatment. Bacterial abundance was determined using quantitative polymerase chain reaction (qPCR). Eleven patients healed their DFU, while FDT failed to heal DFU in the other 12 patients. Microbiome results demonstrated that healing DFUs had a larger abundance Actinomycetales and Staphylococcaceae (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.05), while DFUs that did not heal had a higher abundance of Bacteroidales and Streptococcaceae (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.05). FST marked increases Actinomycetales in DFU, and this increase is significantly greater in patients that healed (<jats:italic>p<\/jats:italic>\u2009&lt;\u20090.05). Future studies to confirm the differential microbiomes, and that increasing Actinomycetales is prognostic of successful FST are warranted. Statement of Clinical Significance: Tracking changes in the prevalence of pathogens in diabetic foot ulcers may be a clinical tool for monitoring treatment response to foot salvage therapy and prognosticating the need for further surgical intervention. The initial wound sample microbiome may provide important prognostic information on the eventual clinical outcome of foot salvage therapy. It may serve as an important clinical tool for patient counseling and making surgical decision of pursuing foot salvage versus amputation. \u00a9 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. 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