Trial Review

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Trial registered on ANZCTR


Registration number
ACTRN12618000339257
Ethics application status
Approved
Date submitted
2/03/2018
Date registered
6/03/2018
Date last updated
1/06/2020
Date data sharing statement initially provided
3/07/2019
Type of registration
Prospectively registered

Titles & IDs
Public title
ECOBABe - Early Colonisation with Bacteria After Birth
Scientific title
Maternal bacteria to correct abnormal gut microbiota in babies born by C-section
Secondary ID [1] 293779 0
nil
Universal Trial Number (UTN)
U1111-1206-7066
Trial acronym
ECOBABe
Linked study record

Health condition
Health condition(s) or problem(s) studied:
Abnormal gut microbiota in infants born by C-section 305894 0
Condition category
Condition code
Metabolic and Endocrine 305088 305088 0 0
Metabolic disorders
Oral and Gastrointestinal 306032 306032 0 0
Other diseases of the mouth, teeth, oesophagus, digestive system including liver and colon

Intervention/exposure
Study type
Interventional
Description of intervention(s) / exposure
Phase 1. The purposes of this phase are to ensure that the gauze incubated in the maternal vagina will contain vaginal bacteria, and to determine the viability of these bacteria.
We will recruit 3 pregnant women aged 18 years and above, who are carrying a singleton fetus and are scheduled for an elective C-section (CS) at 34 to 41 weeks of gestation. Administration of maternal vaginal bacteria to the offspring is not part of Phase 1.
The woman (or the research midwife) will insert a sterile gauze into her vagina for approximately 30 minutes prior to the CS. After removal the gauze will be vertically divided in two equal sized pieces:
• piece one will be placed in a sterile syringe with 5 ml of sterile water to obtain a vaginal-water supernatant
• piece two will be kept at 4ºC in a sterile container
Both the vaginal-water supernatant (obtained from piece one) and the gauze itself (piece two) will undergo: (i) bacterial extraction using the QIAmp DNA extraction kit; and (ii) bacterial characterization by 16S amplicon sequencing. In addition, the vaginal-saline supernatant (obtained from piece one) will undergo: (iii) viability of bacteria using a culture-independent method that directly counts the numbers of live, dead and damaged cells by fluorescent cell sorting (i.e. FACS).

Phase 2. The aim of this phase of the trial is to determine whether babies receiving the maternal vaginal microbiota will have different gut microbiota profile compared to babies receiving placebo, with treated babies more closely resembling the bacterial profile of vaginal-born babies.
We aim to recruit 45 pregnant women and 45 offspring: 30 women carrying singletons (n=30 babies) and scheduled for an elective CS; and 15 women carrying singletons (n=15 babies) and having a vaginal birth (VB). All included women will give birth at 37 weeks of gestation or later. Briefly, approximately 30 minutes before CS, a sterile gauze will be inserted into the maternal vagina by the woman herself or the research midwife. After removal, the gauze will be placed in a sterile syringe with 5 ml of sterile water to obtain a vaginal-water supernatant, which will be kept at 4ºC in a sterile container. Shortly after birth, babies will be randomised to receive either 3 ml of the vaginal-water supernatant or placebo (3 ml of sterile water) orally by syringe. Parents will be blinded to their infant's exposure to maternal vaginal microbiota. The vaginal birth controls will receive neither vaginal-water supernatant nor placebo.
Intervention code [1] 299885 0
Treatment: Other
Comparator / control treatment
The case group (15 CS-born babies who will receive the vaginal-water supernatant) will be compared to two control groups:
- control group 1: 15 CS-born babies who will receive placebo (3 ml of sterile water orally by syringe); and
- control group 2: 15 vaginal-born babies who will receive neither the vaginal-water supernatant nor placebo.
Control group
Placebo

Outcomes
Primary outcome [1] 304267 0
Our primary outcome is a difference in the community structure of the gut microbiota between CS babies who received the active treatment (maternal vaginal microbiota) compared to those who received placebo (sterile water) at 1 month of age. This will be assessed using Bray-Curtis dissimilarity and stastically tested by Permutational Multivariate Analysis of Variance (PERMANOVA).
Timepoint [1] 304267 0
1 month of age
Secondary outcome [1] 341426 0
Difference in overall gut community structure between CS groups. This will be assessed using Bray-Curtis dissimilarity and stastically tested by Permutational Multivariate Analysis of Variance (PERMANOVA).
Timepoint [1] 341426 0
24 hours and 3 months
Secondary outcome [2] 341960 0
Infant anthropometry (weight SDS and length SDS)
Timepoint [2] 341960 0
At birth, at 1 month, and at 3 months (secondary timepoint) of age
Secondary outcome [3] 343845 0
Infant body composition (assessed using whole-body dual-energy X-ray absorptiometry -DXA-)
Timepoint [3] 343845 0
At 3 months (secondary timepoint) of age.
Secondary outcome [4] 383493 0
Similarity of CS seeded and placebo gut profiles to vaginally born cohort. This will be assessed using Bray-Curtis dissimilarity and statistically tested by Permutational Multivariate Analysis of Variance (PERMANOVA).
Timepoint [4] 383493 0
At 24 hours, at 1 month, and at 3 months of age.
Secondary outcome [5] 383494 0
Degree of maternal vaginal strain transfer in CS-born babies. Strain-level identification will be assessed by SNP haplotype based profiling using StrainPhlAn software. The resulting SNP haplotypes will be used to identify the proportion of shared and unrelated strains between mother-infant pairs, comparing these proportions across treatment groups.
Timepoint [5] 383494 0
At 24 hours, at 1 month, and at 3 months of age.

Eligibility
Key inclusion criteria
Maternal inclusion criteria:
- greater than or equal to 18 years of age
- delivery at greater than or equal to 37 weeks of gestation

Offspring (CS and vaginal) inclusion criteria:
- APGAR score greater or equal to 7 at 5 minutes after birth
Minimum age
0 Years
Maximum age
No limit
Sex
Both males and females
Can healthy volunteers participate?
No
Key exclusion criteria
Maternal exclusion criteria:
- emergency c-section
- multiple pregnancy
- carrying a fetus with chromosomal/single gene defects or syndromes
- type 1 diabetes, type 2 diabetes, or gestational diabetes
- use of probiotic supplements or antibiotics in the last two weeks of pregnancy
- a history suggestive of chorioamnionitis
- premature rupture of membranes (PROM) and prolonged PROM
- intrapartum fever >38°C
- history of genital herpes (CS group only)
- a previous group B streptococcus (GBS)-infected baby
- GBS bacteriuria of any count or a re-occurring GBS infection during the current pregnancy
- any transmissible viral, bacterial, or protozoan pathogens

Offspring exclusion criteria:
- birth before 37 weeks of gestation
- congenital abnormalities detected at birth
- perinatal asphyxia (5-minute Apgar score <7)
- respiratory distress requiring support/oxygen therapy with subsequent admission to NICU
- respiratory distress requiring support/oxygen therapy without NICU admission, but without approval of attending clinician for study inclusion

Study design
Purpose of the study
Treatment
Allocation to intervention
Randomised controlled trial
Procedure for enrolling a subject and allocating the treatment (allocation concealment procedures)
Randomisation sequences will be computer generated. Participants will be randomised in a 1:1 ratio to either treatment or placebo group, using block randomisation with variable block sizes of 2 and 4. Parents will be blinded to infants' treatment, To maintain the integrity of the trial evaluation, analysis of the primary outcome (i.e. microbial investigations) will be performed on encoded data, so that the researcher(s) who will perform these analyses will be blinded to group allocation.
Methods used to generate the sequence in which subjects will be randomised (sequence generation)
Computer-generated randomisation sequences
Masking / blinding
Blinded (masking used)
Who is / are masked / blinded?
The people receiving the treatment/s


The people analysing the results/data
Intervention assignment
Parallel
Other design features
Phase
Not Applicable
Type of endpoint/s
Efficacy
Statistical methods / analysis
Metagenomic sequencing data will be processed using bioBakery workflows using docker images available at http://huttenhower.sph.harvard.edu/biobakery_workflows. Briefly, sequence data quality control, including removal of any human reads, will be conducted using KneadData. Taxonomic and functional profiles of the microbiota will be generated using MetaPhlAn v2.6 (Segata et al., 2012) and HUMAnN2 (Fransoza et al., 2018) respectively. Additionally, strain-level taxonomic profiling will be achieved by SNP haplotype based profiling using StrainPhlAn software (Truong et al., 2017). The resulting SNP haplotypes will be used to identify the proportion of shared and unrelated strains between mother-infant pairs, comparing these proportions across treatment groups.
Permutational multivariate analysis of variance (PERMANOVA) will be used to identify any significant differences in the overall microbial community structure between treatment groups. Additionally, Multivariate Association with Linear Models (MaAsLin2) will be used to identify any significant associations between specific microbial taxa and treatment groups. Both taxonomic and functional profiles will be compared. Microbiota stability across time points will be assessed using Bray Curtis dissimilarity. Microbial alpha diversity will be assessed using Shannon’s diversity index and gene counts.
Treatment effects on clinical outcomes will be assessed on the principle of intention to treat, using data collected from all randomised participants. Generalised linear models will be used, adjusting for the baseline outcome value (where appropriate) and sex, and maybe gestational age. Model-adjusted estimates and the differences among the three groups will be calculated and tested. Planned subgroup analyses by sex will be conducted on primary and secondary outcomes to evaluate the consistency of possible treatment effects in males and females. Data analyses will be performed in SAS v.9.4, SPSS v26, and/or Minitab v16. Statistical tests will be two-tailed and significance maintained at 5% level.

References:
Segata N, Waldron L, Ballarini A, Narasimhan V, Jousson O, Huttenhower C. Metagenomic microbial community profiling using unique clade-specific marker genes. Nature methods. 2012;9(8):811-814.
Franzosa EA, McIver LJ, Rahnavard G, et al. Species-level functional profiling of metagenomes and metatranscriptomes. Nature methods. 2018;15(11):962-968.
Truong DT, Tett A, Pasolli E, Huttenhower C, Segata N. Microbial strain-level population structure and genetic diversity from metagenomes. Genome research. 2017;27(4):626-638.

Recruitment
Recruitment status
Stopped early
Data analysis
No data analysis planned
Reason for early stopping/withdrawal
Other reasons/comments
Other reasons
Recruitment has stopped because of the NZ government imposed national lockdown (i.e. mandatory physical isolation) as a result of the Covid-19 outbreak.
Date of first participant enrolment
Anticipated
3/09/2018
Actual
3/12/2018
Date of last participant enrolment
Anticipated
31/03/2020
Actual
16/03/2020
Date of last data collection
Anticipated
30/06/2020
Actual
Sample size
Target
48
Accrual to date
Final
50
Recruitment outside Australia
Country [1] 9441 0
New Zealand
State/province [1] 9441 0
Auckland

Funding & Sponsors
Funding source category [1] 298238 0
Government body
Name [1] 298238 0
Health Research Council (HRC) of New Zealand
Country [1] 298238 0
New Zealand
Primary sponsor type
University
Name
University of Auckland
Address
Liggins Institute
University of Auckland
85 Park Road
Grafton 1023
Auckland
New Zealand
Country
New Zealand
Secondary sponsor category [1] 297525 0
None
Name [1] 297525 0
n/a
Address [1] 297525 0
n/a
Country [1] 297525 0

Ethics approval
Ethics application status
Approved
Ethics committee name [1] 299247 0
Health and Disability Ethics Committees (HDEC)
Ethics committee address [1] 299247 0
Ethics committee country [1] 299247 0
New Zealand
Date submitted for ethics approval [1] 299247 0
29/03/2018
Approval date [1] 299247 0
31/05/2018
Ethics approval number [1] 299247 0
18/NTA/49

Summary
Brief summary
Trial website
Trial related presentations / publications
Public notes

Contacts
Principal investigator
Name 79786 0
Prof Wayne Cutfield
Address 79786 0
Liggins Institute
University of Auckland
85 Park Road
Grafton
Auckland 1023
New Zealand
Country 79786 0
New Zealand
Phone 79786 0
+64 9 3737599 Ext 84476
Fax 79786 0
Email 79786 0
[email protected]
Contact person for public queries
Name 79787 0
Wayne Cutfield
Address 79787 0
Liggins Institute
University of Auckland
85 Park Road
Grafton
Auckland 1023
New Zealand
Country 79787 0
New Zealand
Phone 79787 0
+64 9 3737599 Ext 84476
Fax 79787 0
Email 79787 0
[email protected]
Contact person for scientific queries
Name 79788 0
Wayne Cutfield
Address 79788 0
Liggins Institute
University of Auckland
85 Park Road
Grafton
Auckland 1023
New Zealand
Country 79788 0
New Zealand
Phone 79788 0
+64 9 3737599 Ext 84476
Fax 79788 0
Email 79788 0
[email protected]

Data sharing statement
Will individual participant data (IPD) for this trial be available (including data dictionaries)?
No
No/undecided IPD sharing reason/comment
The clinical data cannot be made available in a public repository due to strict conditions of the ethics approval of our study. We plan to make the study protocol publicly available through its publication in an open access journal. Further, anonymized and unidentifiable data, as well as study documents and materials could be made available to other investigators upon request to the principal investigator.


What supporting documents are/will be available?

Doc. No.TypeCitationLinkEmailOther DetailsAttachment
2740Study protocol  [email protected]
2741Statistical analysis plan  [email protected]
2742Informed consent form  [email protected]
2743Ethical approval  [email protected]
2744Analytic code  [email protected]



Results publications and other study-related documents

Documents added manually
No documents have been uploaded by study researchers.

Documents added automatically
SourceTitleYear of PublicationDOI
EmbaseOral administration of maternal vaginal microbes at birth to restore gut microbiome development in infants born by caesarean section: A pilot randomised placebo-controlled trial.2021https://dx.doi.org/10.1016/j.ebiom.2021.103443
Dimensions AIAn update on the current understanding of the infant skin microbiome and research challenges2023https://doi.org/10.1016/j.mib.2023.102364
N.B. These documents automatically identified may not have been verified by the study sponsor.