Previously established diagnostic approaches for the diagnosis of bacterial vaginosis (BV), such as the Amsel criteria or the Nugent scoring system, do not always correspond to modern trends in understanding the etiology and pathogenesis of polymicrobial conditions. Inter-examiner variability and interpretation of data complicate the wet mount microscopy method. Gram staining of smears does not always provide reliable information regarding bacterial taxa, biofilms, or vaginal dysbiosis. Therefore, the introduction of molecular techniques into clinical practice is extremely relevant. Molecular approaches allow not only the diagnosis of BV but also provide an assessment of microbial composition, which is especially important in the differential diagnosis of vaginal infections. The current review represents an expert opinion on BV diagnosis and is based on extensive experience in the field of vaginal infection diagnosis and treatment.