A mild traumatic brain injury (mTBI) can increase the risk of neurodegenerative-related disease, and serious long-term outcomes are often overlooked. In forensic science, the accurate identification of mTBIs can directly affect the application of evidence in practice cases. Recent research has revealed that the oral cavity and fecal microbiota play a fundamental role in deeply interconnecting the gut and brain injury. Therefore, we investigated the relationship between the temporal changes of the oral cavity and fecal bacterial communities with damage identification and post-injury time estimation after mTBI. In this study, we analyzed the oral cavity and fecal bacterial communities in mTBI rats under 12 different post-injury times (sham, 0 h, 2 h, 6 h, 12 h, 24 h, 2 d, 3 d, 5 d, 7 d, 10 d, and 14 d post-injury) using 16S rRNA sequencing technology. The sequence results revealed bacteria belonging to 36 phyla, 82 classes, 211 orders, 360 families, 751 genera, and 1398 species. Compared to the sham group, the relative abundance of the bacterial communities varied markedly in the post-injury groups. Importantly, our data demonstrated that Fusobacteria, Prevotellaceae, Ruminococcaceae, and Lactobacillaceae might be the potential candidates for mTBI identification, and 2 h post-injury was a critical time point to explore the temporal changes of mTBI injury-time estimation. The results also provide new ideas for mTBI treatment in the clinic.