A new study published in the Journal of Medical Microbiology aimed to profile and compare pelvic and cervical microbiota from patients with pelvic inflammatory disease (PID) and to test the ascending infection hypothesis, which states that microbes in the vagina or cervix spread to the upper genital tract and cause PID. This study recruited 38 PID patients and 19 control patients who received salpingectomy or salpingo-oophorectomy. Both pelvic and cervical samples were collected and culture diagnosis and next-generation sequencing (NGS)-based 16S rRNA profiling were carried out. The findings revealed that in the PID group, half the samples were dominated by a single organism while the other half exhibited a polymicrobial infection. Moreover, the pelvic and cervical microbiota were similar in terms of both taxonomic richness and evenness. Pelvic microbiota was dominated by Acinetobacter, Escherichia, Sneathia and Streptococcus, whereas cervical microbiota was dominated by Lactobacillus and bGardnerella. Meanwhile, only six pelvic samples were positive by culture diagnosis, three of which matched the NGS-based results. The PID samples were further divided into 23 hydrosalpinx and 15 pyosalpinx samples according to the pelvic mass collected during surgery; the former displayed a richer bacterial composition. From the results, it was inferred that the NGS-based method provides a more informative profile of pelvic microbiota than the culture method. Since pelvic and cervical microbiota from the same patient were inconsistent with each other, it was stated that therapeutic decisions based on cervical microbiota may lead to antimicrobial misuse.