Article Text

Preventive effect of prenatal maternal oral probiotic supplementation on neonatal jaundice (POPS Study): A protocol for the randomised double-blind placebo-controlled clinical trial
  1. Bekalu Kassie Alemu1,2,
  2. May Wing Lee1,
  3. Maran Bo Wah Leung1,
  4. Wing Fong Lee1,
  5. Yao Wang1,3,
  6. Chi Chiu wang1,4,
  7. So Ling Lau1
  1. 1Department of Obstetrics and Gynaecology, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, Hong Kong SAR
  2. 2Department of Midwifery, College of Medicine and Health Science, Debre Markos University, Debre Markos, Ethiopia
  3. 3Institute of Health Sciences, The Chinese University, Hong Kong, Hong Kong SAR
  4. 4School of Biomedical Sciences, Joint Laboratory for Reproductive Medicine, The Chinese University, Hong Kong, Hong Kong SAR
  1. Correspondence to Dr So Ling Lau; solinglau{at}


Introduction Neonatal jaundice is a common and life-threatening health problem in neonates due to overaccumulation of circulating unconjugated bilirubin. Gut flora has a potential influence on bilirubin metabolism. The infant gut microbiome is commonly copied from the maternal gut. During pregnancy, due to changes in dietary habits, hormones and body weight, maternal gut dysbiosis is common, which can be stabilised by probiotics supplementation. However, whether probiotic supplements can reach the baby through the mother and reduce the incidence of neonatal jaundice has not been studied yet. Therefore, we aim to evaluate the effect of prenatal maternal probiotic supplementation on the incidence of neonatal jaundice.

Methods and analysis This is a randomised double-blind placebo-controlled clinical trial among 94 pregnant women (47 in each group) in a tertiary hospital in Hong Kong. Voluntary eligible participants will be recruited between 28 and 35 weeks of gestation. Computer-generated randomisation and allocation to either the intervention or control group will be carried out. Participants will take either one sachet of Vivomixx (450 billion colony-forming units per sachet) or a placebo per day until 1 week post partum. Neither the study participants nor researchers will know the randomisation and allocation. The intervention will be initiated at 36 weeks of gestation. Neonatal bilirubin level will be measured to determine the primary outcome (hyperbilirubinaemia) while the metagenomic microbiome profile of breast milk and maternal and infant stool samples as well as pregnancy outcomes will be secondary outcomes. Binary logistic and linear regressions will be carried out to assess the association of the microbiome data with different clinical outcomes.

Ethics and dissemination Ethics approval is obtained from the Joint CUHK-NTEC Clinical Research Ethics Committee, Hong Kong (CREC Ref: 2023.100-T). Findings will be published in peer-reviewed journals and presented at international conferences.

Trial registration number NCT06087874.

  • Clinical Trial
  • Fetal medicine
  • Maternal medicine
  • Prenatal diagnosis

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  • This randomised, double-blind, placebo-controlled superiority trial for reaching the baby through the mother to tackle neonatal jaundice is the first of its kind.

  • Using multistrain probiotics (Vivomixx with eight strains of beneficial bacteria) known for its safety during pregnancy is one of the strengths of this trial.

  • Biological samples from the mother and baby (stool and breast milk) will be collected to triangulate the effect of the intervention on clinical outcomes with microbiome data.

  • The participant with her baby will be appointed for the seventh-day follow-up to collect samples and measure the baby’s transcutaneous bilirubin, and this may increase the dropout.


Neonatal jaundice is a yellowish discolouration of the skin and sclera of neonates in the first month of life due to elevated levels of bilirubin in the blood (hyperbilirubinaemia).1–3 Hyperbilirubinaemia is a total serum bilirubin (TSB) level measurement of more than 5 mg/dL or >85.5 µmol/L in 24 hours and 12 mg/dL or 205 µmol/L after a day.4 Neonatal jaundice can be physiologic or pathologic in its type. The former usually appears after 24 hours of birth and goes off within 2 weeks of life while the latter arises within a day with a TSB level of more than 15 mg/dL and lasts beyond 14 days.2 5 Physiological jaundice is associated with different factors such as ethnicity, geographical location, birth season, maternal smoking and pharmacotherapy.6 It is also associated with low birth weight,7–9 prematurity and sepsis.8 Pathological jaundice is mostly caused by glucose-6-phosphate dehydrogenase deficiency,8 10 ABO incompatibility and rhesus haemolysis.11 Neonatal jaundice needs comprehensive management to avoid severe complications, such as brain damage. However, due to different barriers, evidence-based management of neonatal jaundice is still challenging.12 Even though there are certain managements such as phototherapy, they have various impacts on the vision, hearing and alertness of the newborn. It also affects the circadian rhythm of babies and causes dehydration, hypocalcaemia and renal damage.13

Neonatal jaundice is occurring in varied incidence (about 60%–90%) worldwide.6 The incidence in North America is about 34%.14 It is more prevalent among infants of Southeast and Far East Asian mothers and may be up to threefold higher than among whites.6 15 Among the three main regions in China, the Central South Region has extreme hyperbilirubinaemia. A study in Hong Kong showed about 50%–70% of normal-term infants have the problem.16 Though the bulk of neonatal jaundice causes is unknown, some of the associated factors include breast milk jaundice, breastfeeding jaundice and glucose-6-phosphate dehydrogenase deficiency as well as surgical disease.17 It is also hypothesised due to elevated enterohepatic recirculation of unconjugated bilirubin, which is mediated by β-glucuronidase enzyme and bad bacteria in the gut.18 19 Neonates’ gut flora, which is mainly copied from the mother in the uterus, during childbirth, and through breast feeding has a critical role in bilirubin metabolism by antagonising the activity of the β-glucuronidase enzyme and converting conjugated bilirubin into stercobilinogen (to be excreted via faeces) and urobilinogen (via urine).20 21 In addition to converting conjugated bilirubin into excretable form, good bacteria in the gut keep the neonate’s gut healthy and increase defecation frequency. Even though the gut microbiome cannot have an impact on the pathological causes of hyperbilirubinaemia such as haemolysis, it may facilitate the excretion of conjugated bacteria.22 23 Thus, it reduces enterohepatic recirculation. On the contrary, the recirculation of unconjugated bilirubin is facilitated during gut microbiome imbalance (dysbiosis).24

Preventing gut dysbiosis is very crucial and could be one of the management modalities of neonatal jaundice.25 The mechanism to modulate the infant’s gut is important. Modulating maternal gut microbiome could end up with beneficial bacteria enriched in breast milk. Our systematic review and meta-analysis proved this innovative approach.26 The way for this modulation of breast milk is the enteromammary pathway/gut-breast axis, where intestinal dendritic cells and CD18+ cells play an important role in helping beneficial bacteria translocate from the maternal gut to the mammary system.27–29 The infant gut microbiota is the collection of germs colonising the newborn’s intestine. It is thought to be established by breastfeeding and exposure during birth through the birth canal.30 31 Bioactive components of human breast milk, oligosaccharides, which are non-digestible carbohydrates forming the third largest solid component in human milk particularly are significantly important to shaping infant gut microbiota by nurturing the beneficial bacteria.32 Antimicrobial agents in breast milk also inactivate pathogens individually, additively and synergistically.33 34 In the case of gut dysbiosis, pathogenic micro-organisms will be more abundant and play a role in enhancing the beta-glucuronidase enzyme, which indirectly promotes deconjugation and bilirubin reuptake into the enterohepatic circulation.21

Supplementing pregnant women with products containing beneficial bacteria (probiotics) can maintain gut health.21 35 36 Probiotics are supplements composed of live beneficial microorganisms to improve microbial balance, lower intestinal pH, decrease colonisation and invasion by pathogenic organisms, and modify the host immune response, mainly in the gastrointestinal tract. They consist of Saccharomyces boulardii, Streptococcus thermophilus, Lactobacillus and Bifidobacterium species.37 Administration of probiotics directly to infants is evidenced to prevent and treat neonatal problems including jaundice through maintaining gut health, increasing stool frequency, decreasing beta-glucuronidase enzyme activity, compete with pathogens for food and receptors in the gut.36 38 39 From lines of literature, we found that probiotic species can modulate the physiology of bilirubin metabolism.21 Such bacteria include but are not limited to Bifidobacterium, Lactobacillus, Streptococcus and Saccharomyces species.


As compared with non-jaundiced babies, those affected by jaundice had significantly lower levels of Bifidobacterium39 (B. adolescentis, B. bifidum and B. longum),40 41 which imply Bifidobacterium has a bilirubin reduction effect.40 42 An animal study suggested that faecal beta-glucuronidase enzyme was significantly reduced in animals fed diets containing Bifidobacterium (B. longum) supplements.43 Another study conducted among 53 healthy volunteers confirmed that Bifidobacterium (B. breve, B. lactis and B. bifidum) has a suppressive effect on beta-glucuronidase activity.44 45 B. infantis also has an impact on maintaining mucosal and immune systems, reducing sepsis and increasing defecation frequency.46 47


Probiotic Lactobacillus rhamnosus GG has a significant effect on inhibition of serum bilirubin level and increasing defecation frequency to enhance bilirubin excretion.48L. rhamnosus and L. acidophilus are species that could reduce hospitalisation of neonates with hyperbilirubinaemia.45 Lactobacillus bulgaricus as one of the bifid triple viable has a therapeutic effect on neonatal jaundice.39 Lactobacillus plantarum has also a significant effect on the reduction of bilirubin and liver enzymes.41

Saccharomyces boulardii

S. boulardii, in an animal model study, affects preventing bacterial translocation and improvement of intestinal barrier function in obstructive jaundice.49


S. thermophilus is also one of the bifid triple viable probiotic components and it is effective in treating neonatal jaundice.39

Since their action on mucosal adhesion, immune system development and inhibition of pathogens is much better in combination than as a single species, multistrain probiotic products are chosen for better effect.50 51 Therefore, for this project, Vivomixx, a food supplement that is produced according to food Good Manufacturing Practices and a widely studied product on pregnant women with various outcomes of interest could be used. We have selected Vivomixx from available products in the market like ‘PGut pregnancy probiotics’52 and ‘Prenatal-Probiotics’53 because of two main reasons. First, Vivomixx is a well-applied probiotic on pregnant women with a null adverse event.54–61 Second, it contains a majority of probiotic species identified to affect neonatal jaundice/ bilirubin metabolism.56

This is the first study that aims to investigate the preventive effect of maternal probiotic supplementation on the incidence of neonatal jaundice and bilirubin level, pregnancy outcomes, breast milk, as well as maternal and infant stool microbiome profile.

Methods and analysis

Study design, setting and period

This is a randomised double-blind placebo-controlled parallel group superiority clinical trial in the Prince of Wales Hospital (PWH), Hong Kong from January 2024 to June 2025. This trial will be with pregnant women randomly assigned to either the probiotic group (Vivomixx with maltose) or the placebo group (maltose only). The trial protocol is reported as per the Standard Protocol Items: Recommendations for Interventional Trials guidelines.62


Pregnant women who will attend their antenatal care and plan to deliver in PWH will be invited to participate in this study.

Inclusion criteria

  • Pregnant women from 18 to 45 years old.

  • Gestational age between 28 and 35 weeks.

  • Normal singleton pregnancy.

  • Not on standing dose of antibiotic treatment during enrolment and initiation of the intervention.

  • Plan to exclusively breastfeed.

Exclusion criteria

  • Pregnant women with any fetal abnormality.

  • Couple with glucose 6-phosphate dehydrogenase enzyme deficiency.

  • Couple with known rhesus or haemolytic disease history.

  • Any known risk of developing pathological jaundice.

  • Plan to give birth at other hospitals than PWH.

  • Women with any contraindications for breast feeding.

  • Non-exclusively breastfeeding babies.

Recruitment and randomisation

Pregnant women will be recruited between 28 and 35 weeks in the outpatient clinics in a tertiary-level hospital (figure 1). After confirming the eligibility of pregnant women, the investigator will explain the study by using a printed information sheet that contains the title of the study, purpose, expected length of time for participation, description of all the procedures, confidentiality, voluntary participation, the right to refuse the intervention, withdraw from participation in the clinical trial at any time, contact information and all information will be kept securely. Sufficient time is allowed for their consideration, and all their questions will be answered. Pregnant women will voluntarily sign the written informed consent form (online supplemental file 1). Computer-generated randomisation and allocation will be made by a third party other than the investigators and clinicians to allocate the pregnant women to either probiotic or placebo groups in (1:1) ratio. Neither the study participants nor researchers will know the randomisation and allocation. The intervention will be initiated at 36 weeks of gestation for both study groups. To date, the recruitment has not started yet.

Figure 1

Flow diagram of participant recruitment and follow-up in the POPS trial using Consolidated Standards of Reporting Trials. DOL, days of life; N, total screened for eligibility; n, number of participants in each stage of enrolment, intervention and follow-up, POPS, Perinatal Oral Probiotics Supplementation.76


The probiotic group pregnant women will take one sachet of probiotic (a maltose containing Vivomixx) product orally daily from 36 weeks of gestation up to the seventh day of postpartum while pregnant women in the control group will take one sachet placebo (only maltose) orally daily for the same duration. The best time to take the product is in the morning before breakfast. Each participant in both groups will be advised to dilute a sachet of product provided with a cap of cold water. Detailed information will be provided to each participant with an information sheet prepared in local language. Both active Vivomixx and placebo are similar in colour and taste with the same white sachet package. The probiotic product in this trial is a widely studied supplement among pregnant women for other research outcomes including vaginal microbiota and cytokine secretion, bacterial vaginosis, breast milk beneficial microbiota and cytokines, glycaemic control, neonatal gastrointestinal functional symptoms and gut microbiota as well as immune response.54–59 Maximum gut microbiome colonisation is achieved in 2–3 weeks of supplementation.63

Investigators have trial identified about 12 different microbiome species from the literature that have direct or indirect bilirubin reduction effects (table 1). There are commercially available probiotic products that contain many of these species. Vivomixx is one of the accessible well-studied products for its safety during pregnancy. It is a non-GMO, gluten-free, high-potency microbiotic food supplement, containing eight strains of live bacteria (450 billion bacteria per sachet) including S. thermophilus DSM24731/NCIMB 30438, Bifidobacterium breve DSM24732/NCIMB 30441, Bifidobacterium longum DSM24736/NCIMB 30435, Bifidobacterium infantis DSM24737/NCIMB 30436, Lactobacillus acidophilus DSM24735/NCIMB 30442, Lactobacillus plantarum DSM24730/NCIMB 30437, Lactobacillus paracasei DSM24733/NCIMB 30439, Lactobacillus delbrueckii ssp. bulgaricus DSM24734/NCIMB 30440.64 From lines of affidavits, we have cross-evaluated and found that seven of the strains affect the bilirubin metabolism. It can be stored at room temperature for up to week without having a major effect on potency. Otherwise, it is safe at 4°C–8℃.65 66 We prepared an icebag that can keep the product at a low temperature during transportation to home and each participant will store the product in a refrigerator at home. It will be stored, distributed and administered as per the product instruction by clinicians.

Table 1

Summary of probiotic species that have direct or indirect effect on neonatal jaundice from different literatures

Outcome measures

Primary outcome

Hyperbilirubinaemia will be the primary outcome of this study. For all participants’ babies, sternal transcutaneous bilirubin (TcB) level will be measured within 2 days and on the seventh day of life by Dräger Meter JM-105, which is a non-invasive device. When TcB exceeds or is within 3 mg/dL of the phototherapy treatment threshold or if the TcB is 15 mg/dL, TSB will be measured as a confirmatory test.67 Hyperbilirubinaemia will be declared when TSB is >5 mg/dL/24 hours or >12 mg/dL after 1 day. Since our intervention will be initiated during pregnancy, all outcomes of each type of jaundice including jaundice diagnosed within 24 hours will be compared between the two groups by considering the intrauterine exposure to probiotics. The TcB measurement device is studied in different settings including a study conducted in Hong Kong in 2016 at Baby-friendly Hospital68 and Tuen Mun Hospital,69 and in a tertiary hospital in Malaysia.70 It is a very quick, effective and non-invasive method of measurement. It is also a reliable method with a better correlation coefficient with TSB measurement.71–73 The bilirubin level will be plotted on the standard neonatal bilirubin level nomogram chart to decide if the infant has considerable jaundice. A measurement of TcB level above the 95th percentile on the nomogram will be regarded as a high risk for subsequent hyperbilirubinaemia67 and will be investigated with TSB and followed.74 There is also a phototherapy and exchange transfusion threshold determining nomogram based on gestational age and age in hours.67 This threshold-determining nomogram will be used and the need for phototherapy and exchange transfusion will also be assessed as outcomes. By taking in utero exposure into consideration, all newborns irrespective of their health status and admission status to the special care baby unit will be included in the study.

Secondary outcomes

Secondary outcomes involve (1) Pregnancy outcomes will be collected by a questionnaire which contains different outcome measure variables including fetal well-being, preterm labour, pre-eclampsia, diabetes mellitus, infection/sepsis, induction of labour, mode of delivery, GA at birth, birth weight, Apgar score and …etc. (2) Milk and stool microbiome: Breast milk samples will be collected within the second and on the seventh day of giving birth. It will be kept at 4°C during transportation and stored at 80°C freezer. The maternal stool will be collected before initiation of the intervention (at 36 weeks of gestation) and after 3 weeks of intervention. Infant stool will be collected simultaneously with breast milk samples (figure 2). Both maternal and infant stool samples will be collected using a tube containing DNA stabiliser and then aliquoted and frozen at 80°C. Both samples will be processed based on the commercially available kit protocol. Metagenomic sequencing will be used to characterise the microbiome in each sample. For analytical measurement, the number of participants from whom provided probiotics were detected (relative risk) and the mean concentration of provided probiotic species in the breast milk and infant stool will be measured. Maternal stool sample microbiome profiles before and after intervention in both probiotic and placebo groups will be compared. (3) Safety for the mother and fetus/ infant: An independent safety monitoring committee and all research team members will monitor each participant for any adverse events, in particular, gastrointestinal symptoms and possible allergies. Mothers and infants will be followed regularly for possible side effects such as allergic reactions, stomach upset, diarrhoea, flatulence (passing gas) and bloating.37 Apart from discomfort causing symptoms such as bloating other severe side effects are uncommon. However, in any case, such a moderate to severe side effects, such as decreased/absent/fetal movement, fever, uncommon vaginal fluid leakage, severe headache, decreased abdominal girth or overdistended abdomen, the intervention will be stopped, the participant will be linked to the Accident and Emergency Department. If any adverse events occur in infants, infants will be linked to the paediatrics department.

Figure 2

Timeline of the POPS study from recruitment to end of follow-up. POPS, Perinatal Oral Probiotics Supplementation

Follow-up, compliance and dietary intake data

Pregnant women will be reminded to collect preintervention and postintervention stool samples, start the intervention at 36 weeks of gestation and be advised to report when labour starts or any discomfort. For better retention of participants, a reminder message will be sent to each participant using WhatsApp in the local language at four different time points. The pregnant women will be reminded to record daily on the diary form regarding the dose and time they take the supplement, supplemented with any use of prescribed antibiotics or adverse events such as any diarrhoea, hospitalisation, etc. In addition, they will also be reminded to keep their usual diet only and not to take any other probiotic products. The unused supplements report will be collected. Maternal dietary intake and diversity will be evaluated using standard individual dietary diversity measurement guidelines.75 76

Data and sample collection

For this randomised controlled clinical trial, data will be collected at least at three time points.

  • Recruitment: At this stage, participants’ characteristics such as age, marital status, educational level and other sociodemographic characteristics, obstetrics and gynaecological characteristics, medical and surgical factors, reproductive history-related factors including family size, behavioural-related factors (smoking or alcohol use), allergies, use of supplements/vitamins/probiotics, prior lactation history, use of standing dose of antibiotics, plan for the place of birth, current pregnancy condition (gestational age, type of pregnancy (singleton), presentation, fetal heartbeat, placental location, fetal weight, any supplement/ vitamin/ probiotic in the previous 1 month. Preintervention maternal stool samples will be collected from women in both groups. The intervention will be initiated at 36 weeks of gestation and after 3 weeks of intervention, postintervention data will be collected.

  • Within 2 days of life and before discharge:

    Having sufficient rest after delivery, mothers will be contacted until before discharge home for data collection at their most convenient time. The time for data collection will be marked. Data including gestational age at delivery, date and time of delivery, mode of delivery, any antibiotic use; infant birth weight, Apgar score, urination status of the infant and time of initiation of breast feeding will be documented. Breast milk and infant stool samples will be collected, and the volume of samples collected, and storage temperature for both samples. Moreover, the neonate’s TcB level will be measured and documented.

  • Follow-up on the seventh day of infant life: Breast milk and infant stool samples will be collected. Both the volume and storage temperature of samples will also be recorded. Infant body weight and bilirubin level will be measured. Breastfeeding status will be recorded. The diary record for supplements will be returned and the final sachet count report will be recorded.

In general, the data collection will be exclusively by the research staff appointed for this research using a pretested questionnaire. After recruitment, every mother will be counselled at each contact for better compliance with the intervention.

Data handling and quality assurance

All the data collected through this study will be handled by the principal investigator and researchers and kept confidential. Data quality will be checked regularly by all investigators separately and a trial managing committee.

Sample size

The sample size in this protocol is calculated using the G*power V. statistical software.77 Due to the lack of previous studies on pregnant women targeting bilirubin level reduction in neonates, we used a previous study conducted to assess the effect of a probiotic on neonatal hyperbilirubinaemia by providing the lactobacillus RGG directly to neonates in Turkey to calculate our study sample size (PMID: 30968632).78 The effect size was calculated using mean, SD and sample size, which are the most common parameters to measure effect size. In the paper conducted in Turkey, the mean and SD of bilirubin level and the sample size were mentioned, and we took the third-day bilirubin measurement records. The effect size was computed as 0.663. Finally, the sample size is calculated using the following assumptions: Two tailed, power 80, effect size 0.663, α=0.05, 1:1 allocation ratio. This yields about 74 (37 in each arm), and then we considered a 20% drop-out using the formula N=(n/1−DO). The final sample size, therefore, computed 94 (47 in each group).48 We have also calculated the sample size using a reduction of the incidence of neonatal jaundice in controls (p=46.6%) and in the probiotics group (p=14.6%) and yield (n=62; 31 in each group), which is smaller than the first sample size. To calculate this sample size, we used additional parameters (α=0.05, power=0.8, enrolment ratio 1:1) and calculated using an online platform.79

Statistical analysis

This study will use the Consolidated Standards of Reporting Trials guidelines.80 An intention-to-treat approach will be used to assess the cause-and-effect relation to maintain the effect of randomisation. Baseline characteristics will be compared between allocated treatment groups using frequencies and descriptive statistics. Pearson’s χ2 test and Fisher’s exact test will be run to assess the difference of different factors between the two intervention groups (online supplemental table). To assess the exclusive effect of the intervention on the occurrence of neonatal jaundice, a binary logistic regression model will be fitted. Additionally, to examine the effect of the intervention on the mean bilirubin level of neonates, mean beneficial bacteria level in breast milk, and in maternal and infant stool, linear regression will be employed. The microbial DNA will be extracted from different specimens, including breast milk, and maternal and infant stool samples using commercial kits. Quality control and metagenomics sequencing will be employed and analysed by bioinformatics experts to obtain microbiome information including taxonomy and function.

Study status

Participant recruitment has not started yet.

Patient and public involvement

The public representative was involved in the Joint CUHK-NTEC Clinical Research Ethics Committee, whose membership includes lay persons from the community, apart from clinical and nursing staffs. Results of the trial will be disseminated to the community including study participants through presentations in workshops and posters as well as flyers that will be posted in the outpatient clinics for ease of access. Plan for participant involvement in participant recruitment is not considered in this study. This trial has subsequent follow-up schedules. In each visit, participants will be asked for any adverse effects. A diary form is also prepared to mark down the dose taken and any side effects.

Ethics and dissemination

All the procedures in this clinical trial will be conducted as per the ethical principles of the Declaration of Helsinki of the World Medical Association and the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) Guideline for Good Clinical Practice (GCP) for medical research involving human subjects. Ethical approval was obtained from the CUHK-NTEC (The Chinese University of Hong Kong and New Territories East Cluster) Clinical Research Ethics Committee (CREC) Hong Kong (with CREC Reference number 2023.100-T). Participants will be identified and screened with eligibility criteria by reviewing their medical records and questioning them for these criteria. For those who fulfilled the inclusion criteria, adequate information will be provided by using the printed information sheet. Written informed consent will be taken from each participant after their approval. Codes instead of names will be used and all data will be kept confidential. These codes will be used throughout each communication among the research team and for data analysis.

In focus on increasing visibility, using any recognised benefits of the interventions resulting from this study and further evaluation, enhancing academic collaborations, linking relevant research areas in this field and identifying further areas where research could be effectively managed, this study result will be disseminated through presentations in academic seminars, workshops, and national and international symposiums. It will be submitted to The Chinese University of Hong Kong for archival and possible dissemination. The aggregated findings of this trial will be published in peer-reviewed academic journals for better visibility. A lay summary of the results and links to publications will be made available on databases.


Emerging evidence increasingly indicates that the microbiome established in early life, even before birth, plays a pivotal role in shaping human health and disease outcomes in later years.81 Neonatal jaundice, a prevalent problem in newborns, is attributed to dysfunction in bilirubin metabolism, production and excretion.82 Despite treatments such as phototherapy and exchange transfusion are commonly employed, however, preventive approaches for neonatal jaundice are limited. Gut flora is instrumental in converting conjugated bilirubin into stercobilinogen and urobilinogen, facilitating its excretion as stercobilin in faeces and urobilin in urine.20 Interestingly, the neonatal gut microbiome is primarily established by the maternal gut and milk microbiome during pregnancy and lactation periods.83 84 Therefore, remodelling the maternal microbiome provides us with a potential opportunity to shape the gut microbiome in newborns.

Probiotics have been proven to be an effective intervention to shape the maternal-offspring microbiome. Of note, lines of evidence suggest maternal probiotic supplementation not only positively influences the breast milk and infant’s gut microbiome but also enhances maternal health during pregnancy.26 Importantly, no adverse effects from these supplements during pregnancy have been reported to date. Previous studies showed that directly administering probiotics to neonates enables them to treat and prevent neonatal jaundice.39 85 However, direct neonatal administration of probiotics is not always practical. Therefore, we questioned if breast milk can be modulated by probiotic supplementation during pregnancy thereby reshaping the infant’s gut microbiome and reducing neonatal complications such as jaundice.

To this end, we hypothesised perinatal probiotic supplementation in pregnancies may serve as a feasible approach to preventing neonatal jaundice and designed this randomised controlled trial to evaluate its effect. Our study will revolutionise current neonatal care practices, offering a non-invasive, safe, inexpensive and potentially highly effective preventive strategy for neonatal jaundice. It also underscores the critical role of the maternal microbiome in infant health, paving the way for new approaches in prenatal care.

Ethics statements

Patient consent for publication


The Hong Kong Obstetrical & Gynaecological Trust Fund granted funding for the team to handle this study. The Clinical Research Pharmacy in the Prince of Wales Hospital, Hong Kong has signed agreement with this study team to store the investigational food products (Vivomixx and placebo) and dispense by keeping the cold chain of the product. The authors would like to acknowledge Mendes, the innovative microbiotic company in Europe, for supplying the investigational food product (Vivomixx and placebo). The trial team would like to sincerely acknowledge the Department of Obstetrics and Gynaecology, at the Chinese University of Hong Kong for supporting the trial in logistics.


Supplementary materials

  • Supplementary Data

    This web only file has been produced by the BMJ Publishing Group from an electronic file supplied by the author(s) and has not been edited for content.


  • Contributors SLL is the principal investigator of this trial. CCW and BKA conceived the idea. BKA developed the research question and study design and prepared the protocol for ethical approval submission under the supervision of CCW and YW. WFL and BKA prepared an information sheet, consent form and other logistics before submission to the ethics committee for approval. MBWL and BKA optimised the laboratory sample handling, storage and processing protocol. BKA, CCW and YW established collaboration with the innovation microbiotic company for free supply of the investigational food product. MWL, SLL, CCW, YW, BKA and WFL obtained the funding. CCW is the senior trial supervisor who coordinated the operational delivery of the study protocol. WFL provides support on recruitment. YW and BKA handle the data and analysis. BKA drafted the manuscript and all authors listed provided critical review and approved the manuscript for submission.

  • Funding The Hong Kong Obstetrical & Gynaecological Trust Fund supported the trial (grant number: N/A). Department of Obstetrics and Gynaecology, Faculty of Medicine, the Chinese University of Hong Kong provided logistics for the laboratory sample storage, processing and analysis.

  • Competing interests None declared.

  • Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.