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Background: The purpose of this cohort study was to assess the incidence of positive cultures in section's osseous slice biopsy (SOB) taken at the level of major limb amputation. In case of positive cultures we sought whether the microorganisms present in SOB could take origin from the primary infection site necessitating the amputation. The impact of diabetes on culture results was also investigated. Methods: This prospective cohort study, which aimed to confirm the results of the pilot study, analysed patients who underwent major limb amputation between 2012 and 2013 in three Lithuanian hospitals. SOBs at the amputation site (surgical bone biopsies) and percutaneous bone biopsies of the distal site were performed simultaneously during limb amputation. Tissue cultures were analysed by microbiologists, and species along with antibiograms were reported. Histopathological assessment and bacterial typing were also evaluated. A positive culture was defined as the identification of at least 1 bacteria not belonging to the skin flora, at least 2 bacteria belonging to the skin flora with the same antibiotic susceptibility profiles or the same bacteria belonging to the skin flora in two different sites. Fisher's exact test and Student's test were used to compare the populations and the microbiological results. The statistical significance level was set at P< 0.05. Results: Sixty-nine patients (35 males/34 females), mean age 68.7 (S= 13.6) years, including 21 (30.4 %) with diabetes underwent the major limb amputation. Forty-five amputations (65.2 %) were done above the knee. In total, 207 SOBs and 207 percutaneous distal site biopsies were studied. SOB cultures were positive in 11 (15.9 %) cases. In 5 (45.5 %) cases the same microorganisms were identified in both SOB and distal biopsy cultures. No association between culture results and presence of diabetes was identified. Conclusions: Our results suggest that, independently of the diabetes status, foot infection may silently spread along the bone and can achieve the site of major limb amputation. Additional investigations aiming to confirm this hypothesis and to evaluate a prognostic value are in progress. [ABSTRACT FROM AUTHOR]