A Probiotic Intervention With Bifidobacterium longum NCC3001 on Perinatal Mood Outcomes (PROMOTE Study): Protocol for a Decentralized Randomized Controlled Trial

**Background:** Perinatal mood disturbances (PMD), including depression and anxiety, affect a substantial proportion of women, with up to 70% experiencing 'baby blues' postpartum and approximately 24% of women in their third trimester and 15% of postpartum women experiencing anxiety. These symptoms can negatively impact breastfeeding practices and infant development. Many women avoid pharmacological treatments during pregnancy and lactation, creating a need for alternative interventions. The probiotic Bifidobacterium longum (BL) NCC3001 has shown potential in reducing anxiety-like behavior in preclinical models and low mood in human adults with irritable bowel syndrome, but had not been tested in the perinatal period. The COVID-19 pandemic created additional barriers to conventional clinical trial conduct, particularly for vulnerable populations like pregnant women.

**Methods:** This was a double-blind, placebo-controlled, randomized, 3-parallel-arm study aiming to recruit 180 women in Singapore. Participants were randomized 1:1:1 to: (1) prepartum + postpartum probiotic (n=60), (2) prepartum placebo + postpartum probiotic (n=60), or (3) prepartum + postpartum placebo (n=60). The intervention was BL NCC3001 at 1.0 × 10^10 CFU/day in a maltodextrin carrier, consumed as a drink once daily. The placebo was a maltodextrin-based powder with similar sensory characteristics. The intervention period spanned from 28-32 weeks' gestation until 12 weeks postpartum. Mood outcomes were measured using STAI and EPDS questionnaires at baseline (28-32 weeks' gestation), 36 weeks' gestation, 9 days postpartum, and 4, 8, and 12 weeks postpartum. Secondary outcomes included Hospital Anxiety and Depression Scale scores, Parenting Stress Index, Pittsburgh Sleep Quality Index, Gastrointestinal Symptom Rating Scale, breastfeeding practices, salivary cortisol, and gut microbiota composition. The study was conducted entirely remotely using video calls, electronic questionnaires via Medidata Patient Cloud, courier delivery of study products, and self-collected biological samples. Ethics approval was granted by the A*STAR institutional research board (2020-065).

**Key Results:** Recruitment occurred between November 7, 2020, and August 20, 2021. A total of 520 women registered interest, of whom 184 (35.4%) were eligible and randomized. Of these, 5 (2.7%) withdrew after randomization, leaving 179 (97.3%) who completed the study. This exceeded the target of at least 48 participants per group (144 total) due to the much lower than anticipated dropout rate (2.7% vs 20%). Social media was the most effective recruitment strategy, bringing in 46.9% (244/520) of prospective participants, followed by parenting-specific websites (22.3%, 116/520), and public transport posters (12.3%, 64/520). Nationwide recruitment was achieved, with participants from across Singapore. Data processing is ongoing, and no outcome results are yet reported.

**Clinical Implications:** This study demonstrates the feasibility of conducting a fully decentralized randomized controlled trial in a vulnerable population (pregnant women) during a global pandemic. The high retention rate (97.3%) and successful nationwide recruitment suggest that remote trial designs can be effective when the population has high digital literacy and access to technology. The study will provide novel evidence on whether BL NCC3001 can reduce symptoms of perinatal mood disturbances, potentially offering a non-pharmacological intervention for women who wish to avoid medications during pregnancy and lactation. The results may inform the design of future decentralized trials and contribute to understanding the role of probiotics in perinatal mental health.