An abnormal endometrial microbiota has been associated with implantation failure; therefore, it may be important to evaluate it in order to improve reproductive outcomes in infertile patients. The main objective of our study was to compare the endometrial microbiome of patients with recurrent implantation failure (RIF) and control patients undergoing assisted reproduction treatment (ART). A prospective cohort study including forty-five patients with their own or donated gametes. The endometrial microbiome was analysed by massive sequencing of the bacterial 16S rRNA gene. Different bacterial communities were detected in RIF and control patients. Lactobacillus stands out as the most frequent genus, with 92.27% in RIF patients and 97.96% in control patients, and significant differences were reported between the two groups (p = 0.002). No significant differences were found regarding alpha diversity index. In beta diversity analysis, a significant trend was observed in the separation of the bacterial community between established groups (p < 0.07). Relative abundance analysis identified genera Prevotella (p < 0.001), Streptococcus (p < 0.001), Bifidobacterium (p = 0.002), Lactobacillus (p = 0.002) and Dialister (p = 0.003). Our results demonstrated the existence of an endometrial microbiota characteristic of RIF patients and showed that there might be a relationship between population of the endometrial microbiome and embryo implantation failure, providing us the possibility to improve clinical results in this patients.