Prospective study of vaginal samples for the detection of Gardnerella vaginalis and G. swidsinskii/leopoldii using Gardnerella agar in CO2 and anaerobic atmospheres.

Gardnerella vaginalis is a key bacterium in the etiology of bacterial vaginosis (BV) and contributes to biofilm formation. Gardnerella is a facultative anaerobic bacterium, meaning it can grow both in the presence of oxygen and under anaerobic conditions. We conducted a prospective study in 2025, including 160 consecutive vaginal samples from women submitted for pathogenic bacteria. Clinical information on the referral form indicated in 87 women (54.4%), from which colpitis in 64 cases (73.6%). There were 43 pregnant women (26.9%), with an average age of 30.7 years. Each vaginal sample was cultured on Gardnerella agar and incubated in both CO2 and anaerobic atmospheres. The culture plates were examined after 24 and 48 h of incubation, independently of other results. In cases where suspicious colonies with beta-hemolysis were observed, bacterial identification was performed using MALDI-TOF mass spectrometry (Bruker). Gram-stained smears were evaluated according to the Nugent score. The bacteria Gardnerella vaginalis and Gardnerella leopoldii/swidsinskii were detected in 46 (28.8%) samples. The sensitivity of G. vaginalis and G. leopoldii/swidsinskii cultivation on Gardnerella agar in a CO2 atmosphere was 67.4% compared to anaerobic cultivation. Anaerobic cultivation on Gardnerella agar proved to be a superior method for the isolation of G. vaginalis and G. leopoldii/swidsinskii compared to cultivation in a CO2 atmosphere. The diagnosis of BV in clinical practice relies on standard Amsel's criteria, gram-stained vaginal smears evaluated using the Nugent scoring system or Ison-Hay criteria, and the isolation of G. vaginalis and G. leopoldii/swidsinskii.IMPORTANCEBacterial vaginosis (BV) is the most common vaginal infection among women of reproductive age and is associated with significant adverse outcomes, including preterm birth, spontaneous abortion, and increased susceptibility to sexually transmitted infections. Accurate and timely microbiological diagnosis is therefore clinically important. Our study demonstrates that anaerobic incubation on selective Gardnerella agar improves the detection of Gardnerella vaginalis and Gardnerella leopoldii/swidsinskii compared with incubation in a CO2-enriched atmosphere. Under anaerobic conditions, colonies were larger, hemolysis was more pronounced, and overall sensitivity was higher. Although BV diagnosis in clinical practice primarily relies on Amsel's criteria and Nugent scoring, optimized culture conditions provide valuable complementary microbiological evidence. These findings contribute to the optimization of laboratory procedures and may enhance the reliability of routine diagnostic workflows in women with suspected BV.