An optimal female reproductive tract (FRT) microbiome represents a cornerstone for successful reproductive and sexual health. Although understanding the link between the microbiota and our health is the focus of a growing number of research programs and research has advanced in the last decade, the FRT microbiome remains relatively poorly defined. This is despite it being one of the most critical factors impacting fertility and the health of the future generation. Important gaps in our knowledge remain about women-specific microbial niches, such as the lower FRT. The vaginal microbiome of healthy women is almost exclusively dominated by Lactobacillus species, making these species synonymous with vaginal health. Largely thanks to their capacity to produce high levels of lactic acid, lactobacilli are thought to inhibit most invading bacterial and viral pathogens. Nonetheless, the dogma that all Lactobacillus species are key beneficial species in the vaginal niche has been questioned, based on the association with disease for some of the major players in the niche, for example L. iners, particularly in non-European or Caucasian cohorts. Further, the ability of other (mostly anaerobic) species besides lactobacilli, to maintain similar homeostasis in the vaginal niche raises the question of what we define as optimal or healthy vaginal microbiota. Answering this question is essential since a non-optimal or dysbiotic vaginal microbiome has been linked to the acquisition of sexually transmitted infections (STIs), infertility, and adverse pregnancy outcomes. To confront these healthcare challenges we should be able to distinguish between an optimal and non-optimal microbiome. With the advance of sequence-based technologies, as well as other omics methodologies, including metaproteomics, metabolomics and culturomics, we are now able to begin to correlate the presence, as well as the activities, of specific organisms and microbial communities with health or disease. However, we are only beginning to understand the complexity of interactions between bacterial community members with each other and the human host. A better understanding of these relationships within this specialized environmental niche and in relation to female sexual and reproductive health is critical for the development of novel diagnostics and therapeutics to create and maintain an optimal FRT microbiome.