Backed by advancements in technologies like microbial sequencing, many studies indicate that the vaginal microbiome is a key marker of female reproductive health. However, further studies are still needed to investigate the correlation between vaginal microbiota (VMB) and outcomes of assisted reproductive technology (ART). Therefore, this study compared the VMB of two types of infertile women undergoing in vitro fertilization (IVF) with normal control women during the implantation window period and investigated the effects of VMB characteristics on IVF outcomes. Vaginal swabs from IVF patients (n = 85) were collected before embryo transfer (ET), and vaginal swabs from normal control women (n = 37) were collected during the 6-8 days of ovulation detection. Results showed differences in the vaginal community structure between infertile women with polycystic ovary syndrome (PCOS) and with tubal factor infertility (TFI) undergoing IVF treatment. The results revealed a higher relative abundance of Lactobacillus iners in the non-pregnant group compared with the pregnant group. The results also demonstrated the abundance of Pseudomonas spp. in both non-pregnant groups of infertile women. The findings suggested that the VMB composition affects the IVF outcomes and that the pre-ET high abundance of L. iners may potentially indicate an IVF failure. The abundance of Pseudomonas spp. in the vagina may be an adverse factor for ART pregnancy.