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Introduction: The intestinal microbiota is vital to human health, and has a profound influence on several biological processes including inflammation and pathogen resistance. Antibiotic intake greatly impacts bacterial diversity, can increase antibiotic resistance and impair the equilibrium between bacterial species. The key to grasping post-antibiotic effects on the gut microbiota rests on the implementation of a suitable procedure to isolate microbial DNA and a meticulous consideration of experimental sequencing artefacts. Methods: We herein report the bacterial community dynamics of a cohort of 128 surgical oncology patients before and after the intravenous administration of cefuroxime, an antibiotic routinely used in surgical antibioprophylaxis with proven efficiency against both gram-positive and gram-negative bacteria. In our study, we analyzed patient fecal samples collected through rectal examination before and 7 days post cefuroxime treatment by employing a high-throughput sequencing assay which targets the V3-V4 region of the 16S rRNA gene. A first challenge in applying the study design was to extract an appropriate amount of DNA characteristic to the sampled microbiota, which implied the use of both mechanical (ceramic beads) and chemical (proteinase K, lysozyme and lysostaphin) lysis. Results: Gut microbiota richness and composition was significantly different between the two groups, but most differences were determined by additional perioperative procedures, rather than antibioprophylaxis. Intestinal microbiota composition was not significantly changed one week post cefuroxime treatment when compared to pre-treatment condition for patients without mechanical bowel preparation, but some loss in taxonomic variety could be observed. Discussion: Taken together, cefuroxime does not promote short-term dysbiosis in surgical patients without any additional perioperative procedures. [ABSTRACT FROM AUTHOR]