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We previously reported on the effects of a lactobacilli probiotic (SynForU-HerCare; two capsules/day of 9.5 log CFU/capsule) in improving symptoms of vaginal irritation, discharge and burning in pregnant women with vaginal candidiasis upon administration for 8 weeks, accompanied by improved emotional and social quality of life parameters. Thus, the present study aimed to analyse vaginal microbiota and inflammatory changes in hope to better understand the improved clinical symptoms as observed previously. Patients in the probiotic group showed a decreased abundance of Candida glabrata after 8 weeks ( p = 0.009) in the lower vaginal region, while patients in the placebo group did not show any changes over time. In the higher vaginal and cervicovaginal regions, patients in the placebo group showed a decreased abundance of Candida albicans only within 4 weeks ( p < 0.05) but no changes in abundance of C. glabrata over time, while patients in the probiotic group showed a continuous decreased abundance of C. albicans and C. glabrata over 8 weeks ( p < 0.05). Patients in the placebo group also had a decreased abundance of Lactobacillus crispatus over 4 weeks ( p = 0.023) in the lower vaginal region and a decreased abundance of L. jensenii over 8 weeks in the cervicovaginal region ( p = 0.001). Meanwhile, patients in the probiotic group had an increased abundance of L. crispatus in the lower vaginal region after 8 weeks ( p = 0.012) and Lactobacillus jensenii over 4 weeks in the cervicovaginal region ( p < 0.001). Inflammation may have occurred in both low and high vaginal regions, predominantly observed by the increased concentration of pro-inflammatory cytokine TNF-alpha in patients from the placebo group ( p < 0.05), while the administration of probiotics has shortened the period of inflammation as observed from the reduced need for anti-inflammatory cytokine IL-4 and IL-10 over time ( p < 0.05). Taken together, our present new data further support previous findings that probiotic SynForU-HerCare had a beneficial effect against vaginal candidiasis in pregnant women via modulation of the vaginal microbiota and microenvironment.