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THE EFFECT OF BALANCED ENERGY-PROTEIN SUPPLEMENTATION DURING PREGNANCY AND LACTATION ON BIRTH OUTCOMES AND INFANT GROWTH IN RURAL BURKINA FASO : STUDY PROTOCOL FOR A RANDOMIZED For peerCONTROLLED review TRIAL only Journal: BMJ Open

Manuscript ID bmjopen-2020-038393

Article Type: Protocol

Date Submitted by the 11-Mar-2020 Author:

Complete List of Authors: Vanslambrouck, Katrien; University of Ghent, Food Technology, Safety and Health, Faculty of Bioscience Engineering de Kok, Brenda; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering Toe, Laeticia Celine; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering; Institut de Recherche en Sciences de la Sante De Cock, Nathalie; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering

Ouedraogo, Moctar; AfricSanté http://bmjopen.bmj.com/ Ganaba, Rasmané; AfricSanté Lachat, Carl; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering Huybregts, Lieven; IFPRI, PHN division Kolsteren, Patrick; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering

Nutrition < TROPICAL MEDICINE, Community child health < Keywords: PAEDIATRICS, Protocols & guidelines < HEALTH SERVICES on September 29, 2021 by guest. Protected copyright. ADMINISTRATION & MANAGEMENT

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 8 9 I, the Submitting Author has the right to grant and does grant on behalf of all authors of the Work (as defined 10 in the below author licence), an exclusive licence and/or a non-exclusive licence for contributions from authors 11 who are: i) UK Crown employees; ii) where BMJ has agreed a CC-BY licence shall apply, and/or iii) in accordance 12 with the terms applicable for US Federal Government officers or employees acting as part of their official 13 duties; on a worldwide, perpetual, irrevocable, royalty-free basis to BMJ Publishing Group Ltd (“BMJ”) its 14 licensees and where the relevant Journal is co-owned by BMJ to the co-owners of the Journal, to publish the 15 Work in this journal and any other BMJ products and to exploit all rights, as set out in our licence. 16 17 The Submitting Author accepts and understands that any supply made under these terms is made by BMJ to 18 the Submitting Author Forunless you peer are acting as review an employee on behalf only of your employer or a postgraduate 19 student of an affiliated institution which is paying any applicable article publishing charge (“APC”) for Open 20 Access articles. Where the Submitting Author wishes to make the Work available on an Open Access basis (and 21 intends to pay the relevant APC), the terms of reuse of such Open Access shall be governed by a Creative 22 Commons licence – details of these licences and which Creative Commons licence will apply to this Work are set 23 out in our licence referred to above. 24 25 Other than as permitted in any relevant BMJ Author’s Self Archiving Policies, I confirm this Work has not been 26 accepted for publication elsewhere, is not being considered for publication elsewhere and does not duplicate 27 material already published. I confirm all authors consent to publication of this Work and authorise the granting 28 of this licence. 29 30 31 32 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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Word count : http://bmjopen.bmj.com/ 37 38 3.763 words 39 40 41 Correspondence : 42 [email protected] 43 44 45 Other email addresses : on September 29, 2021 by guest. Protected copyright. 46 [email protected] - [email protected] - [email protected] 47 48 [email protected] – [email protected] - [email protected] – [email protected] - 49 50 [email protected] 51 52 53 54 55 56 57 58 59 60

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1 2 3 [Main Text File] 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 Abstract 7 8 Introduction: 9 10 Adequate nutrition during pregnancy is crucial to both mother and child. Maternal malnutrition can 11 be the cause of stillbirth or lead to poor birth outcomes such as preterm delivery and small-for- 12 13 gestational-age new-borns. There is a possible positive effect of providing pregnant women a balanced 14 15 energy and protein (BEP) food supplement but more evidence is needed. The MISAME III project is 16 aiming to improve birth outcomes and infant growth by testing a BEP supplement during pregnancy 17 18 and lactation in rural ForBurkina Faso. peer This paper review presents the protocol. only 19 20 Methods and analysis: 21 22 MISAME III is an individually randomized efficacy trial implemented in six rural health center 23 catchments areas in the district of Houndé, Burkina Faso. Eligible pregnant women living in these areas 24 25 and in the reproductive age range (between 15-40 years old) will be enrolled. Women will be randomly 26 27 assigned to one of the four study groups: (i) prenatal intervention only, (ii) postnatal intervention only, 28 (iii) prenatal and postnatal intervention or (iv) no prenatal or postnatal intervention. The intervention 29 30 group will receive the BEP supplement and iron/folic acid (IFA) tablets while the control group will only 31 32 receive the IFA tablets following the national health protocol. Consumption will be supervised by 33 trained village women on a daily basis through home visits. The primary outcomes are small-for- 34 35 gestational age at birth and length-for-age z-score at six months of age. Secondary outcomes will be 36 37 measured at birth and during the first six months of the infants’ life. http://bmjopen.bmj.com/ 38 Ethics and dissemination: 39 40 MISAME III has been reviewed and approved by the University Hospital of Ghent University and the 41 42 ethics committee of Centre Muraz, Burkina Faso. Informed consent will be obtained from participants. 43 44 Results of the various studies will be published in relevant journals and shared with other researchers on September 29, 2021 by guest. Protected copyright. 45 and public health institutions. 46 47 Trial registration number on Clinical Trial: NCT03533712. 48 49 Protocol v1 50 51 6/03/2020 52 53 54 55 56 57 58 59 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 Strengths and limitations of this study 8 - This trial will help to fill in the gap on how to improve birth outcomes in pregnant women in 9 10 lower-and middle-income countries. 11 12 - Inclusion of formative research before the start of the efficacy trial will give us a better 13 14 understanding on what product type to choose and the factors that determine compliance. 15 - Daily consumption of the BEP supplement and/or IFA tablets will be done under supervision 16 17 during pregnancy and lactation. 18 For peer review only 19 - This study will be done in a rural district of Burkina Faso, generalizability of results might 20 therefore be limited. 21 22 - Blinding the participants and all staff members will not be possible since the supplements are 23 24 identifiable. 25 26 27 28 29 Keywords: Maternal nutrition, Dietary supplements, Pregnancy outcomes, Children’s growth, Small 30 31 for Gestational Age, Length for Age, Burkina Faso 32 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 8 9 Introduction 10 11 12 Pregnancy is a challenging period in the life of many women in lower- and middle-income countries 13 (LMIC). Maternal mortality remains high and many neonates suffer from premature delivery and/or 14 15 intra-uterine growth retardation both in length and in weight accumulation (1). An indicator to 16 17 measure neonates growth is small-for-gestational age (SGA). SGA is defined as a birthweight below the 18 10th percentile of a standardFor optimal peer reference review population for aonly given gestational age and sex (2). SGA 19 20 is often caused by growth-restriction in the womb and has been associated with neonatal and post- 21 22 neonatal mortality (2). It has also been linked to an increase in morbidity risks later in life, especially 23 non-communicable diseases (3). SGA affected 23.3 million term children in LMIC in 2012 (4). Adequate 24 25 nutrition during pregnancy is crucial for optimal maternal and newborn health (5, 6) and maternal 26 27 malnutrition has been associated with fetal growth restriction (7). An adequate dietary balance is 28 necessary to ensure sufficient energy intake for adequate growth of the fetus (8). Unfortunately, 29 30 maternal undernutrition remains a big problem in regions across Sub-Saharan Africa and Asia (9, 10). 31 32 33 Several types of food supplements have been developed and evaluated over the past years. A positive 34 effect of multiple-micronutrient supplements (MMS) during pregnancy on birth outcomes has been 35 36 found in previous studies (11). Keats et al. concluded in their review that MMS during pregnancy gave 37 http://bmjopen.bmj.com/ 38 a probable reduction in SGA and preterm births and can thus be used as a future guidance (11). 39 40 According to a multi-country randomized controlled trial done in LMIC, a positive effect of lipid-based 41 nutrient supplements on fetal growth-related birth outcomes can be seen starting before conception 42 43 or starting during the first trimester (12). Moreover, the latest evidence indicates a possible positive 44 45 effect of providing pregnant women a balanced energy and protein (BEP) food supplement (5, 13-15). on September 29, 2021 by guest. Protected copyright. 46 In line with that evidence, the 2016 WHO antenatal care guidelines state that pregnant women in 47 48 undernourished populations should receive - depending on the context - BEP supplements to reduce 49 50 the risk of stillbirth and SGA (6). Researchers, however, still highlight the limited amount of evidence 51 and a need to evaluate the effect of balanced supplements on birth outcomes, such as SGA (5, 13). 52 53 Randomized trials of high quality and big sample sizes, especially in undernourished pregnant women 54 55 are thus needed (13). Following this recommendation, a compositional guidance for a ready-to-use 56 food supplement for pregnant women was developed by the Bill and Melinda Gates Foundation 57 58 (BMGF) in 2016 (16). 59 60

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1 2 3 The MISAME III (MIcronutriments pour la SAnté de la Mère et de l’Enfant) study hypothesizes that (i) 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 providing women with a BEP supplement during pregnancy will decrease the incidence of SGA 6 7 compared to the control group and that (ii) providing them with a BEP supplement during the postnatal 8 period will increase growth in length by the age of six months compared to the control group. 9 10 11 Methods 12 13 14 MISAME III study design 15 16 17 The MISAME III project is an individually, randomized 2x2 factorial efficacy trial aiming to improve birth 18 For peer review only 19 outcomes and infant growth in rural Burkina Faso by testing a BEP supplement during pregnancy and 20 lactation (figure 1). At inclusion, pregnant women will individually and randomly be allocated to a pre- 21 22 natal intervention or control group and a postnatal intervention or control group. The intervention 23 24 group will receive daily BEP supplement to be consumed under supervision for the duration of 25 pregnancy/lactation. Both intervention and control groups will receive the standard IFA tablet through 26 27 the regular antenatal care program. In addition to the main trial, we propose a number of sub-studies 28 29 to test specific hypotheses in a subsample of pregnant/lactating women and children. MISAME III 30 started off with a formative research to identify the preferred product type for the provision of a 31 32 fortified BEP supplement for the RCT. 33 34 35 Study setting 36 37 http://bmjopen.bmj.com/ The study will be conducted in the district of Houndé in Burkina Faso, a land locked country situated 38 39 in West-Africa. Burkina Faso has an infant mortality rate of 53 per 1.000 livebirths (17) with an 40 41 estimated prevalence of LBW at 14% in 2013 (18). The prevalence of SGA has been estimated to be 42 43 between 32.2% and 41.6% in the district of Houndé (19). The Demographic and Health Survey (DHS) 44 of 2010 found that 16% of women had a body mass index (BMI) below 18.5 kg/m² , which indicates 45 on September 29, 2021 by guest. Protected copyright. 46 that chronic energy deficiencies are present (20). The highest prevalence can be found in the Eastern 47 48 region where 31% of women have a BMI lower than 18.5 kg/m² (21). And in particular, adolescent 49 Burkinabe girls between the age of 15 and 19 years have a low BMI with a prevalence of 23% (22). 50 51 Micronutrient deficiencies also still remain a problem in both infants and women of reproductive age 52 53 in the country (22, 23). 54 55 The climate of the country is Sudano-Sahelian, with a dry season from October to March/April and a 56 57 rainy season from May until September/October. The diet is essentially cereal-based (24) with maize 58 as staple food (25). 59 60

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1 2 3 MISAME III will be conducted in the same health district where two previous MISAME studies were 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 organized (19, 26). The study villages are concentrated around six health centers which are within an 6 7 accessible range from the district hospital. A list of all study sites can be found on 8 www.misame3.ugent. 9 10 11 Study population and recruitment 12 13 14 Women living in the study villages and between the age of 15 and 40 years will be identified through 15 a census. The villages have been selected based on their accessibility and distance to the nearest health 16 17 centre, number of facility-based deliveries per year and their agricultural model as some households 18 For peer review only 19 tend to reside on their fields during the harvest season. Trained village women (femmes 20 accompagnantes: FA), selected in collaboration with the community leaders, will visit the households 21 22 every five weeks to ask about women’s menstruation. In case of amenorrhea, women will be sent to 23 24 the nearest health center for a pregnancy test and a first antenatal consultation by our project midwife 25 when tested positive. An ultrasound examination will be done soon after inclusion by the project 26 27 medical doctor to assess gestational age. A baseline interview will also be done by the project 28 29 interviewers to assess the household members characteristics, household properties, water sanitation 30 31 and hygiene (WASH) and household food security. 32 33 Inclusion criteria: 34 35  Women between 15-40 years old at study inclusion; 36

 Confirmed pregnancy by a pregnancy test; http://bmjopen.bmj.com/ 37 38  Women who signed the informed consent form. 39 40 Exclusion criteria: 41 42  Pregnancies > 20 weeks of gestational age; 43 44  Women planning on leaving the area during their pregnancy; 45 on September 29, 2021 by guest. Protected copyright. 46  Women planning on delivering outside the study area; 47  Women who are allergic to peanuts; 48 49 50 FA will be informed by the project midwife when a participant has been included. FA will visit pregnant 51 women on a daily basis to hand out the BEP supplement and/or IFA tablet and to supervise 52 53 consumption. During the postnatal period, FA will hand out the supplements and IFA tablets to the 54 55 intervention group on a daily basis until six weeks after birth. From that moment, they will receive a 56 week-worth of BEP supplements. The postnatal control group will get the IFA tablets on a daily basis 57 58 during the first six weeks after birth, and participants will be visited once a week afterwards (without 59 60 any supplementation) to minimalize the effect of the home visits. The FA will inform women on the

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1 2 3 supplement’s function, the importance of antenatal visits during pregnancy, maintaining a healthy 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 diversified diet, the importance of delivering at a health facility, the importance of exclusive 6 7 breastfeeding and the introduction of complementary foods at the age of six months. Throughout the 8 study, the FA will be supervised by project interviewers. Supervision visits will be conducted by using 9 10 Lot Quality Assurance Sampling (LQAS) schemes and empty sachets counts to ensure that study 11 12 participant are visited according to the project protocol. 13 14 Manufacturing of twelve fortified BEP supplements and the formative study 15 16 17 Twelve fortified BEP supplements were pre-tested before the start of the RCT during a formative 18 For peer review only 19 research phase. Several food manufacturing companies were invited to produce ready-to-use BEP 20 supplements following the compositional guidelines proposed during an expert meeting hosted by Bill 21 22 and Melinda Gates Foundation (BMGF) (27) in September 2016 (table 1). The BEP supplements had to 23 24 be (i) ready to consume, (ii) not needing for a cold chain and (iii) microbiologically stable. 25 26 Table 1 The compositional guidelines for macro- and micronutrients 27 28 Nutrition component Target per serving 29 30 Total energy: 250-500 kcal per serving; 31 Fat content: 10-60% of energy intake; 32 33 Protein content: 16 g (14-18 g) with a digestible indispensable amino 34 acid score (DIAAS) of ≥ 0.9; 35 Carbohydrates: between 45g and 32g per 100g (added saccharose 36 between 20g and 10g per 100g); http://bmjopen.bmj.com/ 37 Trans fats: <1% of energy intake; 38 Fatty Acid (optional): min of 1.3g of n-3 or 300mg 39 docosahexaenoic acid (DHA) + eicosapentaenoic acid 40 41 (EPA) (of which 200mg DHA) to achieve a healthy n-6: 42 ratio of the supplement of 5:1; 43 Micronutrients: vitamin A, D, E, K, B1 (thiamin), B2 (riboflavin), B3 44 (niacin), B6 (pyridoxine), B9 (folate), B12 and C; 45 minerals: iron, zinc, iodine, calcium, phosphorous, on September 29, 2021 by guest. Protected copyright. 46 copper, and selenium; 47 Optionally: pantothenic acid, manganese, potassium, biotin and 48 49 choline will be included; 50 The final composition of the product will be 51 determined by the selected product as the 52 manufacturing process will influence the 53 macronutrient composition. 54 55 Seven out of 12 were characterized as sweet and five as savoury. Products have been produced in 56 57 different forms including a biscuit, pillow, wafer, bar, paste, instant drink, and soup 58 59 60

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1 2 3 In a first screening step of the formative research, the two most preferred BEP were identified by using 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 a combined evaluation approach consisting of a single meal test, sensory evaluation, and focus group 6 7 discussions in a convenience sample of 40 pregnant women. In a next step, we compared the 8 acceptability of the two pre-selected BEP supplements using a 10-week home-feeding study with 80 9 10 pregnant women to select to most preferred product for the RCT. We refer to both papers for detailed 11 12 information (manuscripts under review). 13 14 Study intervention 15 16 17 At inclusion, pregnant women will be randomly allocated to four different study groups; (i) prenatal 18 For peer review only 19 intervention only, (ii) postnatal intervention only, (iii) prenatal and postnatal intervention or (iv) no 20 prenatal and no postnatal intervention. Prenatal and postnatal supplementation will start right after 21 22 inclusion and birth, respectively. The prenatal intervention group will receive the fortified BEP and IFA, 23 24 the control group IFA tablets alone. The IFA tablets contain 65mg iron and 0.4mg folic acid. The 25 postnatal intervention group will receive the BEP supplement during six months together with the IFA 26 27 tablets for six weeks following the national protocol of Burkina Faso, the control group will receive IFA 28 29 alone for those six weeks. 30 31 32 Allocation/randomization 33 34 A stratified health center permuted block randomization schedule will be used to allocate women to 35 36 the intervention and control group. The double random sequence will be generated before the start http://bmjopen.bmj.com/ 37 of the study using Stata 15.1 (Statacorp, Texas) by an external research analyst. A time coded Stata 38 39 log-file is available upon request after the completion of the study to assess the randomization 40 41 sequence generation. The allocation group will be coded with two letters (pre- and postnatal study 42 group) and placed in a sequentially numbered sealed envelope by project employees not in direct 43 44 contact with participants. The trial midwife will draw a sealed envelope at inclusion and allocate the 45 on September 29, 2021 by guest. Protected copyright. 46 participant to the group defined in the envelop. Blinding of participants and community-based project 47 workers will not be possible since the supplements are identifiable. Blinding of outcome assessors will 48 49 be ensured to a maximum extent measuring primary and secondary outcomes. 50 51 52 Outcomes 53 54 55 Primary outcomes of the RCT 56 57 The trial has two primary study outcomes that is used to assess the impact of the prenatal and the 58 59 postnatal intervention respectively : 60

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1 2 3  th st

Incidence of SGA , defined as birth weight <10 centile Intergrowth 21 reference (28); BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 5  Length-for-age z-score (LAZ) calculated using the WHO 2006 growth reference at six 6 months of age (29). 7 8 Secondary outcomes of the RCT 9 10 11 The trial has a list of secondary outcomes which can be found in table 2. 12 Birthweight measurements will be defined using the Intergrowth 21st reference (28) and child 13 anthropometry using the Child Growth Standards developed by the WHO (29). 14 15 Table 2 Secondary outcomes of the RCT on maternal, newborn and child level 16 17 18 For peer review only 19 Maternal outcomes Newborn Child 20 21 22 Total and trimester specific Birth weight (measured within 72 Weight-for-Age Z-score at six 23 prenatal weight gain and hours after birth) months of age (WAZ) (and 9 and 24 gestational weight change 12 months on a subsample) 25 26 27 28 Probable and possible maternal Birth length (measured within 72 Weight-for-Length Z-score at six 29 post-natal depression at 2 and 6 hours after birth) months of age (WLZ) (and 9 and 30 months of child age 12 months on a subsample) 31 32 Maternal anemia at the third Ponderal or Rohrer’s index at birth Stunting at six months of age 33 antenatal consultation 34 35 36 Women’s mean and minimum Gestational age at delivery Wasting at six months of age 37 dietary diversity score (measured http://bmjopen.bmj.com/ 38 biweekly) 39 40 Large-for-gestational age Underweight at six months of age 41 42 43 Chest circumference (measured Duration of exclusive 44 within 72 hours after birth) breastfeeding during the first six 45 months of age on September 29, 2021 by guest. Protected copyright. 46 47 48 49 Head circumference (measured Incidence of child wasting over 50 within 72 hours after birth) first six months of life 51 52 53 Arm circumference (measured Weight gain over first six months 54 within 72 hours after birth) of life 55 56 Incidence of preterm birth Child mortality (between birth 57 and six months of age) 58 59 60

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1 2 3 Fetal loss Monthly change in LAZ over first BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 six months of life 5 6 Stillbirths Monthly change in WHZ over first 7 six months of life 8 9 Monthly change in WAZ over first 10 six months of life 11 12 Monthly change in head 13 circumference over first six 14 months of life 15 16 17 Child morbidity symptoms over 18 For peer review onlyfirst six months of life 19 20 Anemia at six months of age 21 22 23 Hemoglobin concentration at six 24 months of age. 25 26 27 28 Outcomes of the sub studies 29 30 Sub-study 1: Impact of the intervention on child and maternal body composition. 31 32 33 To compare body composition between intervention and control, body composition will be 34 35 determined in mother-child dyads by deuterium dilution, and analysis of saliva by a Fourier 36 Transform Infrared reader (Agilent FTIR 4500 series). 37 http://bmjopen.bmj.com/ 38 39 Sub-study 2: Impact of the intervention on dietary intake. 40 41 A dietary assessment study will be conducted by using 24-hour recalls in a subsample of 42 43 women. This will enable to determine if the food supplement actually supplements the diet or 44 45 whether there is a degree of dietary substitution. on September 29, 2021 by guest. Protected copyright. 46 47 Sub-study 3: Impact of the intervention on breastmilk. 48 49 50 Breastmilk samples will be taken in the four study groups to compare the composition and to 51 analyze the interaction between the supplementation periods. 52 53 54 Sample size 55 56 57 With an SGA prevalence of 32% and an anticipated decrease of 7%, a sample of 652 subjects per arm 58 is required with α = 0.05 and β = 0.2 (26). To accommodate for possible losses, the subjects per arm 59 60 was increased to 888 (total subjects of 1776). Possible losses are based on previous MISAME studies

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1 2 3 where the number was ~26 % due to a combination of abortions, still births, multifetal pregnancies, 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 out-migrants, maternal deaths and incomplete data (19, 26). For the analysis of an effect of the 6 7 postnatal intervention on length-for-age at six months of age, the minimally detectable effect depends 8 on the presence or absence of an interaction effect between the pre- and postnatal intervention. In 9 10 the absence of a statistically significant interaction between pre- and postnatal intervention, the 11 12 sample size of 588 children per postnatal study arm allows to detect a difference of 0.18 Z-score 13 (SD=1.1), based on a cross-sectional survey conducted in the Gourcy health district in Burkina Faso 14 15 (30), between study arms with α=0.05 and β=0.20. This implies that if ~1400 singleton live births are 16 17 available, we allow for a maximum lost to follow-up of 16%. In the presence of a statistically significant 18 interaction between For pre- and peer postnatal intervention, review a total only sample size of 1176 represents 294 19 20 children per factorial combination of the prenatal and postnatal study group (four groups in total). A 21 22 subgroup size of 294 allows to detect a difference in LAZ at six months of child age of 0.28 assuming a 23 SD of 1.1, α=0.025 (Bonferroni correction for two primary endpoints analyses) and β=0.20. 24 25 26 Data collection 27 28 29 Anthropometric and clinical procedures 30 31 32 At enrolment, anthropometric measurements from all women will be taken. Gestational age will be 33 determined during an ultrasound consultation by measuring crown-rump length (7-13 weeks), or by 34 35 calculating the mean of three to four measurements: biparietal diameter, head circumference, 36 37 abdominal circumference, and femur length (12-26 weeks) (31). http://bmjopen.bmj.com/ 38 39 During pregnancy, clinical follow up will consist of antenatal visits following the national guidelines. 40 41 42 At birth, anthropometric measurements of all neonates will be assessed in duplicate within the first 72 43 44 hours of life (in practice the aim is to measure within the 24 hours of life). After birth, mother and child on September 29, 2021 by guest. Protected copyright. 45 will visit the health care centers monthly for a follow up on clinical, anthropometric and child morbidity 46 47 measures (signs including fever, vomiting, diarrhea, cough, difficulty breathing and running nose). A 48 49 subsample will be visited at home by the project interviewers to collect data at month 9 and 12. 50 51 Hemoglobin concentrations will be measured in women at enrolment and during the 3th antenatal care 52 53 visit. This will be done at six months of age in children. 54 55 56 Baseline questionnaires 57 58 Pre- and postnatal maternal depression will be assessed using the standardized EPDS questionnaire 59 60 consisting of 10 questions (32). Project midwives will be trained for this and the questionnaire will be

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1 2 3 asked at inclusion and at month two and 6 after birth. Socio-economic and demographic information 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 from all participants will be collected once included. Trained project interviewers will ask questions on 6 7 household members characteristics, household properties, WASH environment and household food 8 security (33). The women’s dietary score will be measured in all participating women by the FA during 9 10 the home visits. This will be done twice a week per participant using the Women’s dietary diversity 11 12 score with 11 food groups (34). 13 14 Table 3 shows the overview of the time schedule and measurements of the trial. 15 16 17 Quality of all study data will be insured by a thorough training of all field staff. Procedures to handle 18 For peer review only 19 tools for data collection (questionnaires, anthropometric and clinical measurement material and 20 laboratory procedures) will be practiced during a training and pre-tested on the field during a dry-run 21 22 of ± three months. Anthropometry standardizations of the field staff will be repeated throughout the 23 24 trial on a regular basis. All anthropometric measurements will be taken in duplicate, newborns will be 25 measured within the 72 hours after birth (preferably within the 24 hours) and all weighing scales and 26 27 HemoCue 201+ devices will regularly be controlled on preciseness to ensure data quality. A WhatsApp 28 29 group will be set up where problems can be communicated and solved quickly. 30 31 All data collection forms of the trial can be found on www.misame3.ugent. 32 33 34 Women will be designated as lost to follow up if they move away or withdraw. Reasons for 35 36 discontinuation will be recorded. 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 Table 3 Participant timeline schedule of enrolment, interventions, assessment and visits. 4 5 Enrolment Allocation Post-allocation 6 7 8 Pregnancy and birth After birth 9 10 Start 5- ANC 1 Household Ultra- ANC 2, Birth Month Month Month Month Month Month Month Month 11 12 weeklyFor peervisit sound review3 and 4 1 only2 3 4 5 6 9 12 13 visits 14 15

16 Enrolment http://bmjopen.bmj.com/ 17 18 Census X 19 20 Pregnancy X 21 identification 22 23 Pregnancy X

24 confirmatory test on September 29, 2021 by guest. Protected copyright. 25 26 Informed consent and X 27 allocation to study 28 group 29 30 Study groups 31 32 Prenatal BEP + IFA X X 33 34 Prenatal IFA X X 35 36 Postnatal BEP + IFA* X X 37 38 Postnatal IFA X X 39 40 41 42 13 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from

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1 2 3 4 Assessments 5 6 Mothers 7 8 Baseline questionnaire X 9 10 Gestational age X 11 determination 12 For peer review only 13 Skinfold measurements X 14 15 Weight (kg) and arm X X X X X X X X X X X

16 http://bmjopen.bmj.com/ circumference (mm) 17 **/*** 18 19 20 Height (cm) X 21 22 Hemoglobin (g/dL) X X 23 (ANC3)

24 on September 29, 2021 by guest. Protected copyright. 25 Women’s dietary X 26 diversity score 27 (biweekly) 28 29 Maternal depression X X X 30 31 Infants 32 33 Birth weight (kg) X 34 35 Birth length (cm) X 36 37 Head circumference X X X X X X X 38 (mm) 39 40 41 42 14 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from

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1 2 3 4 Chest circumference X 5 (mm) 6 7 Arm circumference X X X X 8 (mm) 9 10 Morbidity X X X X X X 11 12 Mortality For peer reviewX XonlyX X X X X 13 14 Weight (kg) and height X X X X X X X X 15 (cm)

16 http://bmjopen.bmj.com/ 17 Breastfeeding practices X X X X X X 18 19 20 Hemoglobin (g/dL) X 21 * The IFA tablets will be given during the first 6 weeks after birth in the postnatal intervention group, according to the national health protocol. 22 ** Only maternal weight will be taken at birth. 23 *** In subsample during month 9 and 12

24 on September 29, 2021 by guest. Protected copyright. 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 15 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 17 of 28 BMJ Open

1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 Data management and analysis 5 6 7 FA will use smartphones with a CAPI programmed in CSPRO (version 7.3.1) to collect data during 8 9 household visits. The study data collected by the project medical doctor, project midwives and 10 11 interviewers will be done by Survey Solutions data entry software (version 19.12.6) on tablets. This 12 data will be uploaded to a central server on a weekly basis. All questionnaires were programmed and 13 14 have been tested on the Survey Solutions Designer website and include validation codes to promote 15 16 the quality of the data entry in the field. Assignments will be sent once a week to the tablets of the 17 field team and preloaded data collected at an earlier contact moment will be used to lower the amount 18 For peer review only 19 of incorrect data. Paper forms will also be available on the field as a backup. 20 21 22 Further data quality checks will be done in Stata 15.1 (Statacorp, Texas). Missing or inconsistent data 23 outliers will be sent back to the field for revision. 24 25 26 Statistical analysis 27 28 29 We refer to the Statistical Analysis Plan (SAP) of the trial; “Statistical analysis plan: Impact of a prenatal 30 and postnatal balanced energy protein supplement on birth size and postnatal child growth in Burkina 31 32 Faso” published at www.misame3.ugent. 33 34 35 Data monitoring 36 37 http://bmjopen.bmj.com/ 38 Data monitoring and auditing 39 The Data and Safety Monitoring Board (DSMB) is an independent multidisciplinary group whose 40 41 members are not involved in the trial. The board consists of a Belgian endocrinologist, a Belgian 42 43 pediatrician, a Burkinabe pediatrician, a Belgian gynecologist and a Belgian ethicist. 44

on September 29, 2021 by guest. Protected copyright. 45 46 Serious adverse events (SAE) 47 48 FA will be trained to recognize health issues and will refer those participants actively to see the project 49 midwife in the CSPS. All SAEs will be recorded on a case-by-case basis and verbal autopsies will be 50 51 conducted for maternal, neonatal and infant deaths by the field medical doctor. 52 53 54 Patient and Public involvement 55 56 MISAME III has been well accepted by the community because of the previous positive experience they 57 58 had with the MISAME I and II study. Through the formative research, women were involved in the 59 60

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1 2 3 choice of BEP supplement. Workshops will be planned at in the end of the study in order to 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 communicate the results to the community. 6 7 8 Ethics and dissemination 9 10 Ethics approval and consent to participate 11 12 MISAME III has been reviewed and approved by the University Hospital of the Ghent University and 13 14 the Burkinabe ethics committee. Important protocol changes will be noted on ClinicalTrials.gov. When 15 eligible women meet the inclusion criteria, project midwives will explain the background and 16 17 procedure of the complete trial. Written informed consent or assent will be asked from the 18 For peer review only 19 participating women. In case of illiteracy, a thumb print will be asked and witnessed by the recruiting 20 investigator and one extra witness. Participants will be explained that all data collected during the trial 21 22 is confidential and that they are allowed to withdraw at any time. A copy of the informed consent and 23 24 assent can be found on www.misame3.ugent 25 26 Confidentiality 27 28 A data management plan has been put in place to address concerns regarding the General Data 29 Protection Regulation (GDPR) rules. During the trial, the data files containing personal identifying 30 31 information will be stored on the Survey Solutions server. Only the principal investigators and the 32 33 project coordinators will be able to access those files. 34 35 36 Dissemination plan 37 http://bmjopen.bmj.com/ 38 Upon completion of the trial, all anonymized study data will be available upon request. Final results 39 40 will be communicated to the participants, the Burkinabe Ministry of health, the field staff, the BMGF, 41 the Ghent University researchers and students, AfricSanté, healthcare professionals and other relevant 42 43 international public institutions. Papers on the study results will be published in peer-review journals 44 45 and will be available on the project website. All investigators contributing to the realization of the on September 29, 2021 by guest. Protected copyright. 46 project and publication of results will be an author. Other contributors such as the participants, FA and 47 48 field staff members will be mentioned in the acknowledgments. 49 50 51 Ancillary care 52 53 The MISAME III project will pay for ancillary care when participants have health issues and in case the 54 55 costs are not covered by the national health care program. Participants suffering harm from trial 56 participation will be covered. 57 58 59 60

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1 2 3

Discussion BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 5 6 In this paper the protocol of an individually randomized efficacy trial in rural Burkina Faso has been 7 8 described in which pregnant and lactating women in the intervention group will receive a BEP 9 10 supplement together with IFA tablets. The control group will only receive the standard IFA treatment. 11 12 The key features of the present trial are first, the inclusion of a formative research for a better 13 14 understanding of which format of supplement is preferred, what taste is most acceptable, and which 15 16 factors affect adherence in the study population. Second, the supplementation will not only be given 17 during pregnancy but also during the first six months after birth. This will give the opportunity to assess 18 For peer review only 19 the specific value of postnatal supplementation on several outcomes. Third, the observed daily intake 20 21 of intervention and control supplements is a key feature to ensure compliance and to avoid sharing of 22 the supplements with other household members. Fourth, MISAME III has the advantage of being the 23 24 third trial of its kind in the study area. This presents an opportunity to anticipate the issues that arose 25 26 in previous trials. For instance, women in specific villages tended to leave their homes for a longer 27 period to go work on the fields outside the village. This gave problems in the continuation of the 28 29 supplementation in the past and will be taken into consideration. And last, similar studies are being 30 31 conducted in other countries, allowing the comparison of results from different contexts. 32 33 In addition, body composition will allow us to give insight in the mechanism by which prenatal 34 35 supplementation affects birth anthropometry. These analyses will provide additional insights in the 36 37 mechanisms by which the prenatal and postnatal interventions affect birth size and postnatal growth, http://bmjopen.bmj.com/ 38 39 respectively. 40 41 Overall, the MISAME III study is expected to improve the birth outcomes of neonates born from 42 43 malnourished women in rural Burkina Faso. This will strengthen and refine the recommendation of the 44 45 WHO concerning the use of context-specific BEP supplementation during pregnancy and lactation. on September 29, 2021 by guest. Protected copyright. 46 47 Supplementary file 48 49 SPIRIT Checklist 2013. 50 51 52 Abbreviations 53 54 AE; Adverse Events, ANC; Antenatal Care, BEP; Balanced Energy Supplement, BMGF; Bill and Melinda 55 56 Gates Foundation, BMI; Body Mass Index, CSPro; Census and Survey Processing System, CSPS; Centre 57 58 de Santé et de Promotion Social, DHS; Demographic and Health Survey, DSMB; Data and Safety 59 Monitoring Board, EP(D)S; Edinburgh Postnatal (Postpartum) Depression Scale, FA; Femme 60

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1 2 3 Accompagnante, GDPR; General Data Protection Regulation, HDI; Human Development Index, IFA; 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 Iron/Folic Acid, IUGR; Intra Uterine Growth Restriction, LAZ; Length-for-Age Z-score, LBW; Low Birth 6 7 Weight, LMIC; Low and Middle Income Countries, LMP; Last Menstrual Period, LQAS; Lot Quality 8 Assessment Sampling, MISAME; MIcronutriments pour la SAnté de la Mère et de l’Enfant, MMS; 9 10 Multiple-Micronutrient Supplements, RCT; Randomized Controlled Trial, SAE; Serious Adverse Events, 11 12 SGA; Small-for-Gestational Age, UNICEF; United Nations International Children's Emergency Fund, 13 UNIMMAP; UNICEF/WHO/UNU International Multiple Micronutrient, WASH; water, sanitation and 14 15 hygiene, WAZ; Weight-for-Age Z-score, WHO; World Health Organization, WHZ; Weight-for-Height Z- 16 17 score, WLZ; Weight-for-Length Z-score. 18 For peer review only 19 Acknowledgements 20 21 The MISAME Study Group thanks the pregnant women and their families for the time spent in this 22 23 study. We would like to acknowledge the staff of AFRICSanté; including Henri Somé for his support on 24 the CAPI software, the field medical doctor Anderson Compaoré, the data collectors (midwives, 25 26 interviewers and femmes accompagnantes), dr. Alain Hein for the standardization exercises and dr. 27 28 Hermann Lanou for the support during the training. We would like to thank dr. Sheila Isanaka for the 29 collaboration during the formative research. Private sector partner Nutriset (France) is acknowledged 30 31 for the development and production of the BEP supplements. 32 33 34 Competing interests statement 35 KV: none to declare 36 BdK: none to declare 37 LCT: none to declare http://bmjopen.bmj.com/ 38 NDC: none to declare 39 MO: none to declare 40 RG: none to declare 41 42 CL: none to declare 43 LH: none to declare 44 PK: none to declare 45 on September 29, 2021 by guest. Protected copyright. 46 Contributor Statement 47 The authors’ responsibilities were as follows: 48 KV wrote the manuscript; 49 PK, LH, CL, NDC, LCT, KV, BdK designed the study and the protocol; 50 PK, LH, LCT, KV, BdK designed the study material tools; 51 LCT, LH, KV, BdK, MO, RG trained the field data collectors; 52 53 All authors contributed substantially to the manuscript and approved the final version. 54 55 Correspondence 56 [email protected] 57 58 Funding Information 59 This work was supported by the Bill and Melinda Gates Foundation. Grant number OPP1175213. 60

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1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 The trial is registered on clinical trials.gov (identifier: NCT00909974) prior to recruitment. Enrolment 5 started in October 2019. 6 7 8 REFERENCES 9 1. Lee AC, Katz J, Blencowe H, Cousens S, Kozuki N, Vogel JP, et al. National and regional 10 estimates of term and preterm babies born small for gestational age in 138 low-income and middle- 11 12 income countries in 2010. The Lancet Global Health. 2013;1(1):e26-e36. 13 2. Katz J, Lee AC, Kozuki N, Lawn JE, Cousens S, Blencowe H, et al. Mortality risk in preterm and 14 small-for-gestational-age infants in low-income and middle-income countries: a pooled country 15 analysis. The Lancet. 2013;382(9890):417-25. 16 3. van Rossum CTM, Fransen HP, Verkaik-Kloosterman J, Buurma-Rethans EJM, Ocké MC. Dutch 17 National Food Consumption Survey 2007–2010: Diet of children and adults aged 7 to 69 years2011. 18 4. Lee AC, KozukiFor N, Cousens peer S, Stevens review GA, Blencowe H, Silveiraonly MF, et al. Estimates of burden 19 and consequences of infants born small for gestational age in low and middle income countries with 20 21 INTERGROWTH-21st standard: analysis of CHERG datasets. bmj. 2017;358:j3677. 22 5. Stevens B, Buettner P, Watt K, Clough A, Brimblecombe J, Judd J. The effect of balanced 23 protein energy supplementation in undernourished pregnant women and child physical growth in 24 low-and middle-income countries: a systematic review and meta-analysis. Maternal & child nutrition. 25 2015;11(4):415-32. 26 6. World Health Organization. WHO recommendations on antenatal care for a positive 27 pregnancy experience: World Health Organization; 2016. 28 7. Krishna U, Bhalerao S. Placental insufficiency and fetal growth restriction. The Journal of 29 Obstetrics and Gynecology of India. 2011;61(5):505-11. 30 31 8. Mridula D, Mishra C, Chakravorty A. Dietary intake of expectant mother. Indian J Nutr Diet. 32 2003;40(1):24-30. 33 9. Tang AM, Chung M, Dong K, Terrin N, Edmonds A, Assefa N, et al. Determining a global mid- 34 upper arm circumference cutoff to assess malnutrition in pregnant women. Food and Nutrition 35 Technical Assistance. 2016. 36 10. Branca F, Grummer-Strawn L, Borghi E, Blössner Md, Onis Md. Extension of the WHO http://bmjopen.bmj.com/ 37 maternal, infant and young child nutrition targets to 2030. SCN News. 2015(41):55-8. 38 11. Keats EC, Haider BA, Tam E, Bhutta ZA. Multiple-micronutrient supplementation for women 39 40 during pregnancy. Cochrane Database of Systematic Reviews. 2019(3). 41 12. Hambidge KM, Westcott JE, Garcés A, Figueroa L, Goudar SS, Dhaded SM, et al. A 42 multicountry randomized controlled trial of comprehensive maternal nutrition supplementation 43 initiated before conception: the Women First trial. The American journal of clinical nutrition. 44 2019;109(2):457-69. 45 13. Ota E, Hori H, Mori R, Tobe-Gai R, Farrar D. Antenatal dietary education and supplementation on September 29, 2021 by guest. Protected copyright. 46 to increase energy and protein intake. Cochrane Database of Systematic Reviews. 2015(6). 47 14. Imdad A, Bhutta ZA. Maternal nutrition and birth outcomes: Effect of balanced 48 protein-energy supplementation. Paediatric and Perinatal Epidemiology. 2012;26:178-90. 49 50 15. Kramer MS, Kakuma R. Energy and protein intake in pregnancy. Cochrane database of 51 systematic reviews. 2003(4). 52 16. Bill and Melinda Gates Foundation (BMGF). Framework and Specifications for the Nutritional 53 Composition of a Food Supplement for Pregnant and Lactating Women (PLW) in Undernourished and 54 Low-Income Settings. Expert consultation Seatle, US; 2016 2016. 55 17. United Nations Development Programme. Human Development Reports [Internet]. 2019 56 [cited 25 February 2020]. Available from: http://hdr.undp.org/en/countries/profiles/BFA. 57 18. Unicef. The state of the world's children: children with disabilities: UN; 2013. 58 59 60

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1 2 3 19. Roberfroid D, Huybregts L, Lanou H, Henry M-C, Meda N, Menten J, et al. Effects of maternal BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 multiple micronutrient supplementation on fetal growth: a double-blind randomized controlled trial 5 in rural Burkina Faso. The American journal of clinical nutrition. 2008;88(5):1330-40. 6 7 20. INSD MD, Macro I. Enquête démographique et de santé et à indicateurs multiples (EDSBF- 8 MICS IV), Rapport préliminaire, Burkina Faso, 2010. Ouagadougou: INSD. 2011;40. 9 21. Institut National de la Statistique et de la Démographie (INSD) et ICF International. Enquête 10 Démographique et de Santé et à Indicateurs Multiples du Burkina Faso 2010. Calverton, Maryland, 11 USA : INSD et ICF International. 2012. 12 22. MOH. National Iodine Status and Anemia Survey, Burkina Faso. 2014. 13 23. MOH. MdlS. Enquête Nutritionnelle Nationale 2013. 2013. 14 24. Savy M, Martin-Prével Y, Sawadogo P, Kameli Y, Delpeuch F. Use of variety/diversity scores 15 for diet quality measurement: relation with nutritional status of women in a rural area in Burkina 16 17 Faso. European journal of clinical nutrition. 2005;59(5):703. 18 25. Huybregts LF,For Roberfroid peer DA, Kolsteren review PW, Van Camp JH.only Dietary behaviour, food and 19 nutrient intake of pregnant women in a rural community in Burkina Faso. Maternal & child nutrition. 20 2009;5(3):211-22. 21 26. Huybregts L, Roberfroid D, Lanou H, Menten J, Meda N, Van Camp J, et al. Prenatal food 22 supplementation fortified with multiple micronutrients increases birth length: a randomized 23 controlled trial in rural Burkina Faso. Am J Clin Nutr. 2009;90(6):1593-600. 24 27. Cuzzocrea F, Larcan R, Lanzarone C. Gender differences, personality and eating behaviors in 25 26 non-clinical adolescents. Eat Weight Disord-St. 2012;17(4):E282-E9. 27 28. Villar J, Altman D, Purwar M, Noble J, Knight H, Ruyan P, et al. The objectives, design and 28 implementation of the INTERGROWTH-21st Project. BJOG: An International Journal of Obstetrics & 29 Gynaecology. 2013;120:9-26. 30 29. De Onis M, Garza C, Onyango AW, Martorell R. WHO child growth standards: Taylor & Francis 31 Philadelphia, PA; 2006. 32 30. Becquey E, Huybregts L, Zongrone A, Le Port A, Leroy JL, Rawat R, et al. Impact on child acute 33 malnutrition of integrating a preventive nutrition package into facility-based screening for acute 34 35 malnutrition during well-baby consultation: A cluster-randomized controlled trial in Burkina Faso. 36 PLoS medicine. 2019;16(8). 37 31. Salomon L, Alfirevic Z, Da Silva Costa F, Deter R, Figueras F, Ghi T, et al. ISUOG Practice http://bmjopen.bmj.com/ 38 Guidelines: ultrasound assessment of fetal biometry and growth. Ultrasound in Obstetrics & 39 Gynecology. 2019;53(6):715-23. 40 32. Cox JL, Holden JM, Sagovsky R. Detection of postnatal depression: development of the 10- 41 item Edinburgh Postnatal Depression Scale. The British journal of psychiatry. 1987;150(6):782-6. 42 33. Femi A. Perception of performance appraisal and workers’ performance in Wema Bank 43 Headquarters, Lagos. Global Journal of Arts, Humanities and Social Sciences. 2013;1(4):89-101. 44 45 34. Martin-Prével Y, Allemand P, Wiesmann D, Arimond M, Ballard T, Deitchler M, et al. Moving on September 29, 2021 by guest. Protected copyright. 46 forward on choosing a standard operational indicator of women’s dietary diversity. FAO; 2015. 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 Figure 1 Study design of the RCT 5 6 7 8 Assessed for eligibility (census) 9 10 11 Follow-up of eligible women (n=9700) 12 13 14 Pregnancies (n=1776) 15 16 Randomization 17 18 For peer review only 19 20 Intervention (n=888) Control (n=888) 21 22 Live births Live births 23 24 25 26 27 28 29 Intervention (n=800) Control (n=800) 30 31 32 Follow-up for at least 6 months Follow-up for at least 6 months 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 Supplementary file : SPIRIT Checklist 2013. 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 Reporting checklist for protocol of a clinical trial. 7 8 Based on the SPIRIT guidelines. 9 10 Instructions to authors 11 Complete this checklist by entering the page numbers from your manuscript where readers will find 12 each of the items listed below. 13 14 Your article may not currently address all the items on the checklist. Please modify your text to 15 include the missing information. If you are certain that an item does not apply, please write "n/a" 16 and provide a short explanation. 17 18 Upload your completedFor checklist peer as an extra reviewfile when you submit only to a journal. 19 20 In your methods section, say that you used the SPIRITreporting guidelines, and cite them as: 21 22 Chan A-W, Tetzlaff JM, Altman DG, Laupacis A, Gøtzsche PC, Krleža-Jerić K, Hróbjartsson A, Mann H, 23 Dickersin K, Berlin J, Doré C, Parulekar W, Summerskill W, Groves T, Schulz K, Sox H, Rockhold FW, 24 Rennie D, Moher D. SPIRIT 2013 Statement: Defining standard protocol items for clinical trials. Ann 25 26 Intern Med. 2013;158(3):200-207 27 28 Page 29 Reporting Item Number 30 31 Administrative 32 information 33 34 Title #1 Descriptive title identifying the study design, 1 35 population, interventions, and, if applicable, trial 36 acronym 37 http://bmjopen.bmj.com/ 38 Trial registration #2a Trial identifier and registry name. If not yet registered, 2 39 name of intended registry 40 41 Trial registration: #2b All items from the World Health Organization Trial n/a 42 data set Registration Data Set 43 44 Protocol version #3 Date and version identifier 1 45 on September 29, 2021 by guest. Protected copyright. 46 Funding #4 Sources and types of financial, material, and other 20 47 support 48 49 Roles and #5a Names, affiliations, and roles of protocol contributors 1 / 20 50 responsibilities: 51 contributorship 52 53 Roles and #5b Name and contact information for the trial sponsor 20 54 55 responsibilities: 56 sponsor contact 57 information 58 59 60

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1 2 3 Roles and #5c Role of study sponsor and funders, if any, in study 20 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 responsibilities: design; collection, management, analysis, and 5 6 sponsor and funder interpretation of data; writing of the report; and the 7 decision to submit the report for publication, including 8 whether they will have ultimate authority over any of 9 these activities 10 11 Roles and #5d Composition, roles, and responsibilities of the 17 12 responsibilities: coordinating centre, steering committee, endpoint 13 committees adjudication committee, data management team, and 14 15 other individuals or groups overseeing the trial, if 16 applicable (see Item 21a for data monitoring 17 committee) 18 For peer review only 19 Introduction 20 21 Background and #6a Description of research question and justification for 4 22 rationale undertaking the trial, including summary of relevant 23 studies (published and unpublished) examining benefits 24 and harms for each intervention 25 26 Background and #6b Explanation for choice of comparators 4 27 28 rationale: choice of 29 comparators 30 31 Objectives #7 Specific objectives or hypotheses 4 32 33 Trial design #8 Description of trial design including type of trial (eg, 5 34 parallel group, crossover, factorial, single group), 35 allocation ratio, and framework (eg, superiority, 36 equivalence, non-inferiority, exploratory) 37 http://bmjopen.bmj.com/ 38 Methods: 39 Participants, 40 interventions, and 41 42 outcomes 43 44 Study setting #9 Description of study settings (eg, community clinic, 6 45 academic hospital) and list of countries where data will on September 29, 2021 by guest. Protected copyright. 46 be collected. Reference to where list of study sites can 47 be obtained 48 49 Eligibility criteria #10 Inclusion and exclusion criteria for participants. If 6 50 applicable, eligibility criteria for study centres and 51 individuals who will perform the interventions (eg, 52 53 surgeons, psychotherapists) 54 #11a 8 55 Interventions: Interventions for each group with sufficient detail to 56 description allow replication, including how and when they will be 57 administered 58 59 60

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1 2 3 Interventions: #11b Criteria for discontinuing or modifying allocated n/a BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 modifications interventions for a given trial participant (eg, drug dose 5 6 change in response to harms, participant request, or 7 improving / worsening disease) 8 9 Interventions: #11c Strategies to improve adherence to intervention 7 / 10 adherance protocols, and any procedures for monitoring 11 adherence (eg, drug tablet return; laboratory tests) 12 13 Interventions: #11d Relevant concomitant care and interventions that are n/a 14 concomitant care permitted or prohibited during the trial 15 16 Outcomes #12 Primary, secondary, and other outcomes, including the 9 17 specific measurement variable (eg, systolic blood 18 For peerpressure), analysisreview metric (eg, changeonly from baseline, 19 20 final value, time to event), method of aggregation (eg, 21 median, proportion), and time point for each outcome. 22 Explanation of the clinical relevance of chosen efficacy 23 and harm outcomes is strongly recommended 24 25 Participant timeline #13 Time schedule of enrolment, interventions (including 14 26 any run-ins and washouts), assessments, and visits for 27 28 participants. A schematic diagram is highly 29 recommended (see Figure) 30 31 Sample size #14 Estimated number of participants needed to achieve 11 32 study objectives and how it was determined, including 33 clinical and statistical assumptions supporting any 34 sample size calculations 35 36 Recruitment #15 Strategies for achieving adequate participant enrolment 6 http://bmjopen.bmj.com/ 37 to reach target sample size 38 39 Methods: 40 Assignment of 41 42 interventions (for 43 controlled trials) 44 45 Allocation: sequence #16a Method of generating the allocation sequence (eg, 8 on September 29, 2021 by guest. Protected copyright. 46 generation computer-generated random numbers), and list of any 47 factors for stratification. To reduce predictability of a 48 random sequence, details of any planned restriction (eg, 49 50 blocking) should be provided in a separate document 51 that is unavailable to those who enrol participants or 52 assign interventions 53 54 Allocation #16b Mechanism of implementing the allocation sequence 8 55 concealment (eg, central telephone; sequentially numbered, opaque, 56 mechanism sealed envelopes), describing any steps to conceal the 57 sequence until interventions are assigned 58 59 60

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1 2 3 Allocation: #16c Who will generate the allocation sequence, who will 8 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 implementation enrol participants, and who will assign participants to 5 6 interventions 7 Blinding (masking) #17a Who will be blinded after assignment to interventions 9 8 9 (eg, trial participants, care providers, outcome 10 assessors, data analysts), and how 11 12 Blinding (masking): #17b If blinded, circumstances under which unblinding is 9 13 emergency permissible, and procedure for revealing a participant’s 14 unblinding allocated intervention during the trial 15 16 Methods: Data 17 collection, 18 management, and For peer review only 19 20 analysis 21 22 Data collection plan #18a Plans for assessment and collection of outcome, 12 23 baseline, and other trial data, including any related 24 processes to promote data quality (eg, duplicate 25 measurements, training of assessors) and a description 26 of study instruments (eg, questionnaires, laboratory 27 28 tests) along with their reliability and validity, if known. 29 Reference to where data collection forms can be found, 30 if not in the protocol 31 32 Data collection plan: #18b Plans to promote participant retention and complete 13 33 retention follow-up, including list of any outcome data to be 34 collected for participants who discontinue or deviate 35 from intervention protocols 36 http://bmjopen.bmj.com/ 37 Data management #19 Plans for data entry, coding, security, and storage, 17 38 39 including any related processes to promote data quality 40 (eg, double data entry; range checks for data values). 41 Reference to where details of data management 42 procedures can be found, if not in the protocol 43 44 Statistics: outcomes #20a Statistical methods for analysing primary and secondary 17 on September 29, 2021 by guest. Protected copyright. 45 outcomes. Reference to where other details of the 46 statistical analysis plan can be found, if not in the 47 48 protocol 49 50 Statistics: additional #20b Methods for any additional analyses (eg, subgroup and n/a 51 analyses adjusted analyses) 52 53 Statistics: analysis #20c Definition of analysis population relating to protocol 17 54 population and non-adherence (eg, as randomised analysis), and any 55 missing data statistical methods to handle missing data (eg, multiple 56 imputation) 57 58 Methods: 59 Monitoring 60

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1 2 3 Data monitoring: #21a Composition of data monitoring committee (DMC); 17 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 formal committee summary of its role and reporting structure; statement 5 6 of whether it is independent from the sponsor and 7 competing interests; and reference to where further 8 details about its charter can be found, if not in the 9 protocol. Alternatively, an explanation of why a DMC is 10 not needed 11 12 Data monitoring: #21b Description of any interim analyses and stopping n/a 13 interim analysis guidelines, including who will have access to these 14 15 interim results and make the final decision to terminate 16 the trial 17 18 Harms For#22 peerPlans for collecting, review assessing, reporting,only and managing 17 19 solicited and spontaneously reported adverse events 20 and other unintended effects of trial interventions or 21 trial conduct 22 23 Auditing #23 Frequency and procedures for auditing trial conduct, if n/a 24 any, and whether the process will be independent from 25 26 investigators and the sponsor 27 28 Ethics and 29 dissemination 30 31 Research ethics #24 Plans for seeking research ethics committee / 18 32 approval institutional review board (REC / IRB) approval 33 34 Protocol #25 Plans for communicating important protocol 18 35 amendments modifications (eg, changes to eligibility criteria, 36 outcomes, analyses) to relevant parties (eg, http://bmjopen.bmj.com/ 37 investigators, REC / IRBs, trial participants, trial 38 39 registries, journals, regulators) 40 Consent or assent #26a Who will obtain informed consent or assent from 18 41 42 potential trial participants or authorised surrogates, and 43 how (see Item 32) 44 45 Consent or assent: #26b Additional consent provisions for collection and use of n/a on September 29, 2021 by guest. Protected copyright. 46 ancillary studies participant data and biological specimens in ancillary 47 studies, if applicable 48 49 Confidentiality #27 How personal information about potential and enrolled 18 50 participants will be collected, shared, and maintained in 51 order to protect confidentiality before, during, and after 52 53 the trial 54 #28 20 55 Declaration of Financial and other competing interests for principal 56 interests investigators for the overall trial and each study site 57 58 59 60

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1 2 3 Data access #29 Statement of who will have access to the final trial 18 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 dataset, and disclosure of contractual agreements that 5 6 limit such access for investigators 7 Ancillary and post #30 Provisions, if any, for ancillary and post-trial care, and 18 8 9 trial care for compensation to those who suffer harm from trial 10 participation 11 12 Dissemination policy: #31a Plans for investigators and sponsor to communicate trial 18 13 trial results results to participants, healthcare professionals, the 14 public, and other relevant groups (eg, via publication, 15 reporting in results databases, or other data sharing 16 17 arrangements), including any publication restrictions 18 Dissemination policy:For #31b peerAuthorship eligibilityreview guidelines only and any intended use of 18 19 20 authorship professional writers 21 22 Dissemination policy: #31c Plans, if any, for granting public access to the full n/a 23 reproducible protocol, participant-level dataset, and statistical code 24 research 25 26 Appendices 27 28 Informed consent #32 Model consent form and other related documentation 18 29 materials given to participants and authorised surrogates 30 31 Biological specimens #33 Plans for collection, laboratory evaluation, and storage n/a 32 of biological specimens for genetic or molecular analysis 33 in the current trial and for future use in ancillary studies, 34 if applicable 35 36

The SPIRIT checklist is distributed under the terms of the Creative Commons Attribution License CC- http://bmjopen.bmj.com/ 37 BY-ND 3.0. This checklist was completed on 06. March 2020 using https://www.goodreports.org/, a 38 39 tool made by the EQUATOR Network in collaboration with Penelope.ai 40

41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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Manuscript ID bmjopen-2020-038393.R1

Article Type: Protocol

Date Submitted by the 06-Jan-2021 Author:

Ouedraogo, Moctar; AfricSanté http://bmjopen.bmj.com/ Dailey-Chwalibog, Trenton; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering Hanley Cook, Giles; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering Ganaba, Rasmané; AfricSanté Lachat, Carl; Ghent University, Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering Huybregts, Lieven; IFPRI, PHN division

Kolsteren, Patrick; Ghent University, Department of Food Technology, on September 29, 2021 by guest. Protected copyright. Safety and Health, Faculty of Bioscience Engineering

Primary Subject Nutrition and metabolism Heading:

Secondary Subject Heading: Paediatrics

Nutrition < TROPICAL MEDICINE, Community child health < Keywords: PAEDIATRICS, Protocols & guidelines < HEALTH SERVICES ADMINISTRATION & MANAGEMENT

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1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 [Title Page File] 5 6 7 Title: 8 THE EFFECT OF BALANCED ENERGY-PROTEIN SUPPLEMENTATION DURING PREGNANCY AND 9 10 LACTATION ON BIRTH OUTCOMES AND INFANT GROWTH IN RURAL BURKINA FASO : STUDY PROTOCOL 11 12 FOR A RANDOMIZED CONTROLLED TRIAL 13 14 15 16 Authors: 17 Vanslambrouck Katrien a, de Kok Brenda a, Toe Laeticia Celine a c, De Cock Nathalie a, Ouedraogo Moctar d , Dailey- 18 For peer review only 19 Chwalibog Trenton a, Hanley-Cook Giles a, Ganaba Rasmané d, Lachat Carl a, Huybregts Lieven b, 20 a 21 Kolsteren Patrick 22 23 24 25 Authors institutional affiliations: 26 a Department of Food Technology, Safety and Health, Faculty of Bioscience Engineering, Ghent 27 28 University, Ghent, Belgium 29 30 b Poverty, Health and Nutrition Division, International Food Policy Research Institute (IFPRI), 1201 I 31 Street NW, Washington, DC 20006, USA 32 33 c Institut de Recherche en Sciences de la Santé (IRSS), Bobo-Dioulasso, Burkina Faso 34 35 d Africsanté, Bobo-Dioulasso, Burkina Faso 36 37 http://bmjopen.bmj.com/ 38 Word count : 39 40 4.000 words 41 42 Correspondence : 43 44 [email protected] - [email protected] 45 on September 29, 2021 by guest. Protected copyright. 46 Faculty of Bioscience Engineering 47 Department of Food technology, Safety and Health 48 49 Coupure Links 653, geb. A, 9000 Gent 50 51 09/264 61 77 52 53 Other email addresses : 54 55 [email protected] - [email protected] - [email protected] 56 [email protected] – [email protected] – [email protected] - 57 58 [email protected] - [email protected] – [email protected] - 59 60 [email protected]

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 8 [Main Text File] 9 10 11 Abstract 12 13 Introduction: 14 15 Adequate nutrition during pregnancy is crucial to both mother and child. Maternal malnutrition can 16 17 be the cause of stillbirth or lead to poor birth outcomes such as preterm delivery and small-for- 18 gestational-age new-borns.For There peer is a probable review positive effect only of providing pregnant women a 19 20 balanced energy-protein (BEP) food supplement but more evidence is needed. The MISAME III project 21 22 aims to improve birth outcomes and infant growth by testing a BEP supplement during pregnancy and 23 lactation in rural Burkina Faso. This paper describes the study protocol. 24 25 Methods and analysis: 26 27 MISAME III is a 4-arm individually randomized efficacy trial implemented in six rural health center 28 catchments areas in the district of Houndé. Eligible pregnant women, aged between 15 and 40 years 29 30 old and living in the study areas, will be enrolled. Women will be randomly assigned to one of the four 31 32 study groups: (i) prenatal intervention only, (ii) postnatal intervention only, (iii) prenatal and postnatal 33 intervention or (iv) no prenatal or postnatal intervention. The intervention group will receive the BEP 34 35 supplement and iron/folic acid (IFA) tablets while the control group will only receive the IFA tablets 36 37 following the national health protocol. Consumption will be supervised by trained village women on a http://bmjopen.bmj.com/ 38 39 daily basis by means of home visits. The primary outcomes are small-for-gestational age at birth and 40 length-for-age z-score at six months of age. Secondary outcomes will be measured at birth and during 41 42 the first six months of the infants’ life. Women will be enrolled from October 2019 until the total 43 44 sample size is reached. on September 29, 2021 by guest. Protected copyright. 45 Ethics and dissemination: 46 47 MISAME-III has been reviewed and approved by the University Hospital of Ghent and the ethics 48 49 committee of Centre Muraz, Burkina Faso. Informed consent will be obtained. Results will be published 50 in relevant journals and shared with other researchers and public health institutions. 51 52 Trial registration number on Clinical-Trials.gov: NCT03533712. 53 54 Protocol v2 55 56 15/12/2020 57 58 59 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 8 9 10 11 12 Strengths and limitations of this study 13 - This trial will help to fill the evidence gap on the effect of BEP supplements in pregnant and 14 15 lactating women on birth outcomes and infant growth. 16 17 - Formative research to select the most suitable BEP supplement ensured that the selected BEP 18 For peer review only 19 is well-accepted by the study population. 20 - The daily intake of BEP supplements and IFA tablets during pregnancy and lactation will be 21 22 directly observed by study workers. 23 24 - This study will assess the impact of factorial combinations of pre-and postnatal BEP on child 25 growth to elucidate the relative importance of BEP during pregnancy and/or early lactation. 26 27 - Blinding of study participants and staff members will not be possible, as the supplements are 28 29 identifiable. 30 31 32 33 34 Maternal nutrition, Dietary supplements, Pregnancy outcomes, Child growth, Small for 35 Keywords: 36

Gestational Age, Length for Age, Burkina Faso http://bmjopen.bmj.com/ 37 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 Introduction 7 8 9 Pregnancy is a challenging period in the life of many women in low- and middle-income countries 10 11 (LMICs). Maternal mortality remains high and many neonates suffer from premature delivery and/or 12 intrauterine growth retardation, both in length and in weight accumulation (1). An indicator to 13 14 measure neonatal growth is small-for-gestational age (SGA). SGA is defined as a birthweight below the 15 th 16 10 percentile of a standard optimal reference population for a given gestational age and sex (2). SGA 17 is often caused by growth-restriction in the womb and has been associated with neonatal and post- 18 For peer review only 19 neonatal mortality (2). It has also been linked to an increased risk of morbidity later in life, especially 20 21 non-communicable diseases (3). SGA affected 23.3 million term children in LMICs in 2012 (4). Adequate 22 nutrition during pregnancy is crucial for optimal maternal and newborn health (5, 6) and maternal 23 24 malnutrition has been associated with fetal growth restriction (7). An adequate dietary balance is 25 26 necessary to ensure sufficient energy intake for adequate growth of the fetus (8). Unfortunately, 27 maternal undernutrition remains a public health challenge in regions across sub-Saharan Africa and 28 29 Asia (9, 10). 30 31 32 Several types of food supplements have been developed and evaluated over the past years. A positive 33 effect of multiple-micronutrient supplements (MMS) during pregnancy on birth outcomes has been 34 35 found in previous studies (11). Keats et al. concluded in their review that MMS during pregnancy gave 36 37 a probable reduction in SGA and preterm births and can thus be used for future guidance (11). http://bmjopen.bmj.com/ 38 39 According to a multi-country randomized controlled trial (RCT) done in LMICs, a positive effect of lipid- 40 based nutrient supplements on fetal growth-related birth outcomes can be seen when starting 41 42 supplementation before conception or during the first trimester (12). Moreover, the latest evidence 43 44 indicates a possible positive effect of providing pregnant women a balanced energy-protein (BEP) food on September 29, 2021 by guest. Protected copyright. 45 supplement (5) (13) (14) (15). In line with that evidence, the 2016 World Health Organization’s (WHO) 46 47 antenatal care guidelines state that pregnant women in undernourished populations should receive, 48 49 depending on the context, BEP supplements to reduce the risk of stillbirth and SGA (6). Researchers, 50 however, still highlight the limited amount of evidence and a need to evaluate the effect of balanced 51 52 supplements on birth outcomes, such as SGA (5, 13). Experimental trials of high quality and large 53 54 sample sizes, especially in undernourished pregnant women, are thus needed (13). Following this 55 recommendation, compositional guidance for a ready-to-use food supplement for pregnant women 56 57 was developed by the Bill and Melinda Gates Foundation (BMGF) in 2016 (16). 58 59 60

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1 2 3 Two previous projects, MISAME-I & II (MIcronutriments pour la SAnté de la Mère et de l’Enfant), 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 conducted in Burkina Faso, investigated the effect of supplementation during pregnancy and the effect 6 7 on birth outcomes in infants (17) (18). MISAME-I compared the effect of the UNICEF/WHO/UNU 8 international multiple micronutrient preparation (UNIMMAP) with the effect of iron and folic acid (IFA) 9 10 alone on fetal growth in a double-blind RCT and concluded that UNIMMAP modestly, but significantly 11 12 increased fetal growth (17). The second study (MISAME-II) assessed the effect of a lipid-based nutrient 13 supplement fortified with UNIMMAP compared to a UNIMMAP tablet during pregnancy in an open- 14 15 label, individually randomized controlled trial on birth anthropometry. It was found that combining 16 17 energy with micronutrients during the prenatal phase led to larger birth lengths (18). Since MISAME-II 18 used the UNIMMAP Foras a control, peer a complete review assessment of theonly impact of the fortified lipid-based 19 20 UNIMMAP was not possible. Therefore, MISAME-III will study the effect of a BEP supplementation, 21 22 compared to a control group, and extend the supplementation postnatally to investigate the net 23 contribution of pre-and postnatal BEP supplementation on child linear growth up to 6 months of age. 24 25 26 In summary, the MISAME-III study hypothesizes that: (i) providing women with a BEP supplement 27 28 during pregnancy will decrease the incidence of SGA compared to the control group; and, (ii) providing 29 them with a BEP supplement during the postnatal period will increase children’s length by the age of 30 31 six months compared to the control group. 32 33 34 Methods 35 36 37 This protocol has been developed in accordance with the SPIRIT guidelines (Supplementary file 1). http://bmjopen.bmj.com/ 38 39 40 MISAME III study design 41 42 The MISAME-III project is an individually randomized 2×2 factorial efficacy trial aiming to improve birth 43 44 outcomes and infant growth in rural Burkina Faso by testing a BEP supplement during pregnancy and 45 on September 29, 2021 by guest. Protected copyright. 46 lactation (Figure 1). At inclusion, pregnant women will be individually and randomly allocated to a 47 prenatal intervention or control group and a postnatal intervention or control group. The intervention 48 49 group will receive a daily BEP supplement to be consumed under supervision for the duration of 50 51 pregnancy/lactation. Both intervention and control groups will receive the standard IFA tablet through 52 the national antenatal care In addition to the main trial, we propose a number of sub-studies to test 53 54 specific hypotheses in a subsample of pregnant/lactating women and children. MISAME-III began with 55 56 a formative study to identify the preferred product type for the provision of a fortified BEP supplement 57 during the RCT. 58 59 60 Study setting

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1 2 3 The study will be conducted in the district of Houndé in Burkina Faso, a land locked country situated 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 in West-Africa, similar to the previous MISAME studies Burkina Faso has an infant mortality rate of 53 6 7 per 1.000 live births (19), with an estimated LBW prevalence at 14% in 2013 (20). The prevalence of 8 SGA has been estimated to be between 32.2% and 41.6% in the district of Houndé (17). The 9 10 Demographic and Health Survey (DHS) of 2010 reported that 16% of women had a body mass index 11 12 (BMI) below 18.5 kg/m² , which indicates the presence of chronic energy deficiencies in the zone(21). 13 The highest prevalence can be found in the Eastern region, where 31% of women have a BMI lower 14 15 than 18.5 kg/m² i.e., low BMI (22). Moreover, in particular, adolescent Burkinabèe girls between the 16 17 age of 15 and 19 years have a low BMI, with an estimated prevalence of 23% (23). Micronutrient 18 deficiencies also remainFor a major peer problem in bothreview infants and women only of reproductive age in the country 19 20 (23, 24). 21 22 The climate of the country is Sudano-Sahelian, with a dry season from October to March/April and a 23 24 rainy season from May until September/October. The diet is essentially cereal-based (25) with maize 25 26 as the main staple food (26). 27 28 MISAME-III will be conducted in the same health district where the two previous MISAME studies were 29 30 organized. The study villages are concentrated around six health centers, which are within an 31 accessible range from the district hospital. A list of all study sites can be found 32 33 on:www.misame3.ugent. 34 35 36 Study population and recruitment 37 http://bmjopen.bmj.com/ 38 39 Women living in the study villages and aged between 15 and 40 years will be identified through a 40 census. The villages were selected based on their accessibility and distance to the nearest health 41 42 centre, number of facility-based deliveries per year, and their agricultural model as some households 43 44 tend to reside on their fields during the harvest season. Trained village women (femme on September 29, 2021 by guest. Protected copyright. 45 accompagnante: FA), selected in collaboration with the community leaders, will visit the households 46 47 every five weeks to ask about women’s menstruation. In case of amenorrhea, women will be sent to 48 49 the nearest health center for a pregnancy test and a first antenatal consultation by our project midwife 50 when tested positive. An ultrasound examination will be completed soon after inclusion by the project 51 52 medical doctor to assess gestational age. A baseline interview will also be done by the project 53 54 interviewers to assess the household members’ characteristics, household properties, water sanitation 55 and hygiene (WASH) and household food security. 56 57 58 Inclusion criteria: 59  Women between 15-40 years old at study inclusion; 60

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1 2 3 

Confirmed pregnancy by a pregnancy test; BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 5  Women who signed the informed consent form. 6 7 Exclusion criteria: 8 9  Pregnancies > 20 weeks of gestational age; 10 11  Women planning on leaving the area during their pregnancy; 12  Women planning on delivering outside the study area; 13 14  Women who are allergic to peanuts. 15 16 17 FAs will be informed by the project midwife when a participant has been included. FAs will visit 18 pregnant women on aFor daily basis peer to distribute review the BEP supplement only and/or IFA tablet and to supervise 19 20 consumption. During the postnatal period, FAs will distribute the supplements and IFA tablets to the 21 22 intervention group on a daily basis until six weeks after birth. From that moment onwards, they will 23 receive a week’s worth of BEP supplements. The postnatal control group will receive the IFA tablets on 24 25 a daily basis during the first six weeks after birth, and participants will thereafter be visited once a 26 27 week (without any supplementation) to minimize the effect of home visits. The FA will inform women 28 on the supplement’s function, the importance of antenatal visits during pregnancy, maintaining a 29 30 healthy diversified diet, the importance of delivering at a health facility, the importance of exclusive 31 32 breastfeeding, and the introduction of complementary foods at the age of six months. Throughout the 33 study, the FAs will be supervised by project interviewers. Supervision visits will be conducted using Lot 34 35 Quality Assurance Sampling (LQAS) schemes and empty sachets counts to ensure that study participant 36 37 are visited according to the project protocol. http://bmjopen.bmj.com/ 38 39 Manufacturing of twelve fortified BEP supplements and the formative study 40 41 42 Twelve fortified BEP supplements were pre-tested before the start of the RCT during a formative 43 44 research phase. Several food manufacturing companies were invited to produce ready-to-use BEP 45 on September 29, 2021 by guest. Protected copyright. 46 supplements following the compositional guidelines proposed during an expert meeting hosted by the 47 Bill and Melinda Gates Foundation (BMGF) (16) in September 2016 (Table 1). The BEP supplements 48 49 had to be (i) ready to consume, (ii) not need for a cold chain, and (iii) microbiologically stable. 50 51 52 Table 1 The compositional guidelines for macro- and micronutrients 53 Nutrition component Target per serving 54 55 Total energy: 250-500 kcal per serving; 56 57 Fat content: 10-60% of energy intake; 58 Protein content: 16 g (14-18 g) with a digestible indispensable amino 59 acid score (DIAAS) of ≥ 0.9; 60

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1 2 3

Carbohydrates: between 45g and 32g per 100g (added sucrose BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 between 20g and 10g per 100g); 5 Trans fats: <1% of energy intake; 6 7 Fatty Acid (optional): min of 1.3g of n-3 or 300mg 8 docosahexaenoic acid (DHA) + eicosapentaenoic acid 9 (EPA) (of which 200mg DHA) to achieve a healthy n-6: 10 ratio of the supplement of 5:1; 11 Micronutrients: vitamin A, D, E, K, B1 (thiamin), B2 (riboflavin), B3 12 (niacin), B6 (pyridoxine), B9 (folate), B12 and C; 13 minerals: iron, zinc, iodine, calcium, phosphorous, 14 15 copper, and selenium; 16 Optionally: pantothenic acid, manganese, potassium, biotin and 17 choline will be included; 18 For peer reviewThe final composition only of the product will be 19 determined by the selected product as the 20 manufacturing process will influence the 21 macronutrient composition. 22 23 Seven out of 12 supplements were characterized as sweet and five as savoury. Products were produced 24 25 in different forms, including a biscuit, pillow, wafer, bar, paste, instant drink, and soup. 26 27 28 In a first screening step of the formative study, the two most preferred BEP supplements were 29 30 identified by using a combined evaluation approach consisting of a single meal test, sensory evaluation, 31 and focus group discussions, in a convenience sample of 40 pregnant women. In a next step, we 32 33 compared the acceptability of the two pre-selected BEP supplements using a 10-week home-feeding 34 35 study, with 80 pregnant women, to select to most preferred product for the RCT. We refer to both 36

papers for detailed information (27) (28). http://bmjopen.bmj.com/ 37 38 39 Study intervention 40 41 42 At inclusion, pregnant women will be randomly allocated to four different study groups; (i) prenatal 43 intervention only, (ii) postnatal intervention only, (iii) prenatal and postnatal intervention or (iv) no 44 45 prenatal and no postnatal intervention. Prenatal and postnatal supplementation will start right after on September 29, 2021 by guest. Protected copyright. 46 47 inclusion and birth, respectively. The prenatal intervention group will receive the BEP supplement and 48 IFA, the control group IFA tablets alone. The IFA tablets contain 65mg iron and 0.4mg folic acid. The 49 50 postnatal intervention group will receive the BEP supplement for six months in combination with the 51 52 IFA tablets for six weeks following the national protocol of Burkina Faso; the control group will receive 53 IFA alone for six weeks. 54 55 56 Allocation/randomization 57 58 59 We will apply a stratified permuted block randomization schedule to allocate women to the prenatal 60 intervention or control group and in a next step to allocate women to a postnatal intervention and

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1 2 3 control group. Per health center (i.e., stratum), women will be individually randomly in permuted 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 blocks of 8 so that, per block, equal numbers are obtained in the prenatal intervention (n=4) and 6 7 control (n=4) group and equal numbers are also obtained in the postnatal intervention (n=4) and 8 control (n=4) group. The double random sequence will be generated before the start of the study using 9 10 Stata 15.1 (Statacorp, Texas) by an external research analyst. The allocation group will be coded with 11 12 two letters (A or B for the prenatal and Y or Z for the postnatal study group) and placed in a sequentially 13 numbered sealed opaque envelopes by project employees, not in direct contact with participants. At 14 15 study enrollment, the trial midwife will draw the next sealed envelope and allocate the participant to 16 17 the study group defined by the letter code in the envelope. Blinding of participants and community- 18 based project workersFor will notpeer be possible review since the supplements only are identifiable. Field staff 19 20 responsible for measuring primary and secondary study outcomes are not directly involved in the daily 21 22 supplementation of the study participants and can therefore be considered to be partially blinded. 23 24 Outcomes 25 26 27 Primary outcomes of the RCT 28 29 The trial has two primary study outcomes that will be used to assess the impact of the prenatal and 30 31 the postnatal intervention, respectively: 32

33 th st 34  Incidence of SGA, defined as birth weight <10 centile of the Intergrowth 21 35 reference (29); 36 37  Length-for-age z-score (LAZ) calculated using the WHO 2006 growth reference at six http://bmjopen.bmj.com/ 38 months of age (30). 39 40 Secondary outcomes of the RCT 41 42 43 A list of the trial’s secondary outcomes can be found in Table 2. 44 Birthweight measurements will be defined using the Intergrowth 21st reference (29) and child on September 29, 2021 by guest. Protected copyright. 45 anthropometry using the Child Growth Standards developed by the WHO (30). 46 47 Table 2 Secondary outcomes of the RCT on maternal, newborn, and child level 48 49 50 Maternal outcomes Newborn Child 51 52 53 Total and trimester-specific Birth weight (measured within 72 Weight-for-Age Z-score at six 54 prenatal weight gain and hours after birth) months of age (WAZ) (and 9 and 55 gestational weight change 12 months on a subsample) 56 57 58 59 60

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1 2 3 4 Probable and possible maternal Birth length (measured within 72 Weight-for-Length Z-score at six BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 postnatal depression at 2 and 6 hours after birth) months of age (WLZ) (and 9 and 6 months of child age 12 months on a subsample) 7 8 9 Maternal anemia at the third Ponderal or Rohrer’s index at birth Stunting at six months of age 10 antenatal consultation 11 12 Women’s mean and minimum Gestational age at delivery Wasting at six months of age 13 dietary diversity score (measured 14 twice weekly) 15 16 Large-for-gestational age Underweight at six months of age 17 18 For peer review only 19 Chest circumference (measured Duration of exclusive 20 within 72 hours after birth) breastfeeding during the first six 21 months of age 22 23 24 25 26 Head circumference (measured Incidence of child wasting over 27 within 72 hours after birth) first six months of life 28 29 30 Arm circumference (measured Weight gain over first six months 31 within 72 hours after birth) of life 32 33 34 Incidence of preterm birth Child mortality (between birth 35 and six months of age) 36 37 http://bmjopen.bmj.com/ 38 Fetal loss Monthly change in LAZ over first 39 six months of life 40 41 42 Stillbirths Monthly change in WHZ over first 43 six months of life 44 45 on September 29, 2021 by guest. Protected copyright. 46 Monthly change in WAZ over first 47 six months of life 48 49 50 Monthly change in head 51 circumference over first six 52 months of life 53 54 55 Child morbidity symptoms over 56 first six months of life 57 58 59 Anemia at six months of age 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 Hemoglobin concentration at six 7 months of age. 8 9 10 11 Outcomes of the sub-studies 12 13 14 Sub-study 1: Impact of the intervention on neonatal and maternal body composition 2-3 weeks after 15 delivery. 16 17 18 Body compositionFor will peer be determined review in mother-child only dyads by deuterium dilution, and 19 20 analysis of saliva by a Fourier Transform Infrared reader (Agilent FTIR 4500 series). The sub- 21 22 study will also assess if early-gestation maternal body mass index (defined as body weight in 23 kilograms divided by the square of height in meters) modifies the intervention's effect on 24 25 neonatal body composition. 26 27 Sub-study 2: Impact of the intervention on dietary intake. 28 29 30 A dietary assessment study will be conducted, using 24-hour dietary recalls in a subsample of 31 32 women. This sub-study will enable us to assess possible substitution of the prenatal diet by 33 the BEP supplement. 34 35 36 Sub-study 3: Impact of the intervention on breastmilk. 37 http://bmjopen.bmj.com/ 38 Breastmilk samples will be taken in the four study groups to compare the composition and to 39 40 analyze the interaction between the supplementation periods. 41 42 43 Sample size 44 45 on September 29, 2021 by guest. Protected copyright. With an SGA prevalence of 32% and an anticipated decrease of 7%, a sample of 652 subjects per 46 47 prenatal arm is required with α = 0.05 and β = 0.2 (31). To accommodate for possible losses, the 48 49 number of subjects per arm was increased to 888 (total subjects: 1776). Possible losses are based on 50 previous MISAME studies where the prevalence was ~26 %, due to a combination of miscarriage, 51 52 stillbirths, multifetal pregnancies, out-migrants, maternal deaths and incomplete data (18). For the 53 54 analysis of an effect of the postnatal intervention on LAZ at six months of age, the minimally detectable 55 56 effect depends on the presence or absence of an interaction effect between the pre- and postnatal 57 intervention. In the absence of a statistically significant interaction between pre-and postnatal 58 59 intervention, at sample size of 588 children per postnatal study arm would allow us to detect a 60

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1 2 3 difference of 0.18 Z-score (SD=1.1), based on a cross-sectional survey conducted in the Gourcy health 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 district in Burkina Faso (32), between study arms with α=0.05 and β=0.20. This implies that if ~1400 6 7 singleton live births are available, we allow for a maximum loss to follow-up of 16%. In the presence 8 of a statistically significant interaction between pre-and postnatal intervention, a total sample size of 9 10 1176 represents 294 children per factorial combination of the prenatal and postnatal study group (four 11 12 groups in total). A subgroup size of 294 would allow us to detect a difference in LAZ at six months of 13 age of 0.28 assuming a SD of 1.1, α=0.025 (Bonferroni correction for two primary endpoints analyses) 14 15 and β=0.20. 16 17 18 Data collection For peer review only 19 20 Anthropometric and clinical procedures 21 22 23 At enrolment, anthropometric measurements from all women will be taken. Gestational age will be 24 25 determined during an ultrasound consultation by measuring crown-rump length (7-13 weeks), or by 26 calculating the mean of three to four measurements: biparietal diameter, head circumference, 27 28 abdominal circumference, and femur length (12-26 weeks) (33). 29 30 31 During pregnancy, clinical follow up will consist of antenatal visits following the national guidelines. 32 33 34 At birth, anthropometric measurements of all neonates will be assessed in duplicate within the first 72 35 hours of life (in practice, the aim is to measure within the 24 hours of life). After birth, mother and 36 37 child will visit the health care centers monthly for a follow-up on clinical, anthropometric and child http://bmjopen.bmj.com/ 38 39 morbidity measures (signs including fever, vomiting, diarrhea, cough, difficulty breathing and running 40 nose). A subsample will be visited at home by the project interviewers to collect postnatal data at 41 42 months 9 and 12. 43

44 rd 45 Hemoglobin concentrations will be measured in women at enrolment and during the 3 antenatal care on September 29, 2021 by guest. Protected copyright. 46 visit. This will be conducted at six months of age among children. 47 48 49 Baseline questionnaires 50 51 52 Pre- and postnatal maternal depression will be assessed using the standardized Edinburgh Postnatal 53 Depression Scale (EPDS) questionnaire consisting of 10 questions (34). Study midwives will be trained 54 55 for this and the questionnaire will be asked at inclusion and at month 2 and 6 after birth. Socio- 56 57 economic and demographic information from all participants will be collected once included. Trained 58 study interviewers will ask questions on household members' characteristics, household properties, 59 60 WASH environment, and household food security (35). The women’s dietary diversity score will be

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1 2 3 measured in all participating women by the FA during the home visits. This will be enumerated twice 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 a week per participant using the Women’s dietary diversity score with 11 food groups (36). 6 7 8 Table 3 shows the overview of the time schedule and measurements of the trial. 9 10 Quality of all study data will be insured by a thorough training of all field staff. Procedures to handle 11 12 data collection tools (questionnaires, anthropometric and clinical measurement material, and 13 14 laboratory procedures) will be pre-tested in the field during a dry-run of ± three months. 15 Anthropometric measurement standardizations of the field staff will be repeated bimonthly 16 17 throughout the trial. Anthropometric measurements will be taken in duplicate. Newborns will be 18 For peer review only 19 measured within 72 hours after birth (preferably within the 24 hours), and all weighing scales and 20 HemoCue 201+ devices will undergo weekly quality control. A WhatsApp group will be set up where 21 22 problems can be communicated and solved quickly. 23 24 25 All data collection forms of the trial can be found on: www.misame3.ugent. 26 27 Women will be designated as lost to follow up if they move from the study area or withdraw their 28 29 participation. Reasons for discontinuation will be recorded. 30 31 32 Women will be enrolled in the study from October 2019 until the total sample size has been reached. 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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16 Enrolment http://bmjopen.bmj.com/ 17 18 Census X 19 20 Pregnancy X 21 identification 22 23 Pregnancy X

24 confirmatory test on September 29, 2021 by guest. Protected copyright. 25 26 Informed consent and X 27 allocation to study 28 group 29 30 Study groups 31 32 Prenatal BEP + IFA X X 33 34 Prenatal IFA X X 35 36 Postnatal BEP + IFA* X X 37 38 Postnatal IFA X X 39 40 41 42 14 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from

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1 2 3 4 Assessments 5 6 Mothers 7 8 Baseline questionnaire X 9 10 Gestational age X 11 determination 12 For peer review only 13 Skinfold measurements X 14 15 Weight (kg) and arm X X X X X X X X X X X

16 http://bmjopen.bmj.com/ circumference (mm) 17 **/*** 18 19 20 Height (cm) X 21 22 Hemoglobin (g/dL) X X 23 (ANC3)

24 on September 29, 2021 by guest. Protected copyright. 25 Women’s dietary X 26 diversity score (twice 27 weekly) 28 29 Maternal depression X X X 30 31 Infants 32 33 Birth weight (kg) X 34 35 Birth length (cm) X 36 37 Head circumference X X X X X X X 38 (mm) 39 40 41 42 15 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from

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24 on September 29, 2021 by guest. Protected copyright. 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 16 43 For peer review only - http://bmjopen.bmj.com/site/about/guidelines.xhtml 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Page 19 of 39 BMJ Open

1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 Data management and analysis 5 6 7 FAs will use smartphones with computer assisted person interviewing (CAPI) programmed in CSPRO 8 9 (version 7.3.1) to collect data during household visits. The study data collected by the project medical 10 11 doctor, project midwives, and interviewers will be done by Survey Solutions data entry software 12 (version 19.12.6) on tablets. This data will be uploaded to a central server on a weekly basis. All 13 14 questionnaires were programmed and have been tested on the Survey Solutions Designer website and 15 16 include validation codes to promote the quality of the data entry in the field. Assignments will be sent 17 once a week to the tablets of the field team and preloaded data collected at an earlier contact moment 18 For peer review only 19 will be used to lower the amount of incorrect data. Paper forms will also be available on the field as a 20 21 backup. 22 23 Further data quality checks will be conducted in Stata 15.1 (Statacorp, Texas). Missing or inconsistent 24 25 data outliers will be sent back to the field for revision. 26 27 28 Statistical analysis 29 30 We refer to the Statistical Analysis Plan (SAP) of the trial; “Statistical analysis plan: Impact of a prenatal 31 32 and postnatal balanced energy-protein supplement on birth size and postnatal child growth in Burkina 33 34 Faso” published on: www.misame3.ugent. 35 36 37 Data monitoring http://bmjopen.bmj.com/ 38 39 Data monitoring and auditing 40 41 The Data and Safety Monitoring Board (DSMB) is an independent multidisciplinary group whose 42 43 members are not involved in the trial. The board consists of a Belgian endocrinologist, a Belgian 44

pediatrician, a Burkinabè pediatrician, a Belgian gynecologist, and a Belgian ethicist. on September 29, 2021 by guest. Protected copyright. 45 46 47 48 Serious adverse events (SAEs) 49 FAs will be trained to recognize health issues and will actively refer those participants to see the project 50 51 midwife in the CSPS in the event they occur. All SAEs will be recorded on a case-by-case basis and 52 53 verbal autopsies will be conducted for maternal, neonatal, and infant deaths by the field medical 54 doctor. 55 56 57 58 Patient and public involvement 59 60

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1 2 3 MISAME-III has been well accepted by the community, because of the previous positive experiences 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 they had with the MISAME-I and II studies. Through the formative study, women were involved in the 6 7 choice of BEP supplement. Workshops will be planned at in the end of the study in order to 8 communicate the study results to the community. 9 10 11 Ethics and dissemination 12 13 14 Ethics approval and consent to participate 15 MISAME-III has been reviewed and approved by the University Hospital of Ghent University 16 17 (B670201734334) and the Burkinabe ethics (N°2018-22/MS/SG/CM/CEI) committee. Important 18 For peer review only 19 protocol changes will be noted on ClinicalTrials.gov. When eligible women meet the inclusion criteria, 20 project midwives will explain the background and procedure of the complete trial. Written informed 21 22 consent or assent will be asked from the participating women. In case of illiteracy, a thumb print will 23 24 be asked and witnessed by the recruiting investigator and one extra witness. Participants will be told 25 that all data collected during the trial is confidential and that they are allowed to withdraw at any time. 26 27 A copy of the informed consent and assent can be found on: www.misame3.ugent and as 28 29 supplementary file (Supplementary file 2). 30 31 Confidentiality 32 33 A data management plan has been put in place to address concerns regarding the General Data 34 35 Protection Regulation (GDPR) rules. During the trial, the data files containing personal identifying 36 information will be stored on the Survey Solutions server. Only the principal investigators and the 37 http://bmjopen.bmj.com/ 38 project coordinators will be able to access those files. 39 40 41 Dissemination plan 42 43 Upon completion of the trial, all anonymized study data will be available upon request. Final results 44 45 will be communicated to the participants, the Burkinabè Ministry of health, the field staff, the BMGF, on September 29, 2021 by guest. Protected copyright. 46 Ghent University researchers and students, AfricSanté, healthcare professionals and other relevant 47 48 international public institutions. Papers on the study results will be published in peer-reviewed 49 50 journals and will be available on the project website. All investigators contributing to the realization of 51 the project and publication of results will be included as an author. Other contributors such as the 52 53 participants, FA and field staff members will be mentioned in the acknowledgments. 54 55 56 Ancillary care 57 58 59 60

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1 2 3 The MISAME-III project will pay for ancillary care when participants have health issues and in case the 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 costs are not covered by the national health care program. Participants suffering harm due to their 6 7 trial participation will be covered. 8 9 10 11 Discussion 12 13 14 In this paper the protocol of an individually randomized 4-arm efficacy trial in rural Burkina Faso has 15 16 been described in which pregnant and lactating women in the intervention group will receive a BEP 17 supplement together with IFA tablets. The control group will only receive the standard IFA treatment. 18 For peer review only 19 20 The key features of the present trial are first, the inclusion of a formative study for a better 21 22 understanding of which type of supplement is preferred, what taste is most acceptable, and which 23 factors affect adherence in the study population. Second, the supplementation will not only be given 24 25 during pregnancy but also during the first six months after birth. This will give us the opportunity to 26 27 assess the specific value of postnatal supplementation on several outcomes. Third, the observed daily 28 intake of intervention and control supplements is a key feature to ensure compliance and to avoid 29 30 sharing of the supplements with other household members. Fourth, MISAME-III has the advantage of 31 32 being the third trial of its kind in the study area. This presents an opportunity to anticipate the issues 33 that arose in previous trials. For instance, women in specific villages tended to leave their homes for a 34 35 longer period to go work on the fields outside the village. This posed problems in the continuation of 36 37 the supplementation in the past and will be taken into consideration during MISAME-III. Fourth, four http://bmjopen.bmj.com/ 38 39 sub-studies are nested in the main trial that will provide insight into the mechanism by which prenatal 40 BEP supplementation affects birth and infant outcomes. And last, similar studies are being conducted 41 42 in other countries, allowing for comparison between results from different contexts. 43 44 45 The MISAME III study will provide evidence on the impact of BEP supplements on birth and infant size on September 29, 2021 by guest. Protected copyright. 46 using a rigorous study design. The study results will further strengthen and refine WHO’s 47 48 recommendation on the use of context-specific BEP supplementation during pregnancy and lactation. 49 50 51 Abbreviations 52 53 AE; Adverse Events, ANC; Antenatal Care, BEP; Balanced Energy-Protein Supplement, BMGF; Bill and 54 55 Melinda Gates Foundation, BMI; Body Mass Index, CSPro; Census and Survey Processing System, CSPS; 56 57 Centre de Santé et de Promotion Social, DHS; Demographic and Health Survey, DSMB; Data and Safety 58 Monitoring Board, EP(D)S; Edinburgh Postnatal (Postpartum) Depression Scale, FA; Femme 59 60 Accompagnante, GDPR; General Data Protection Regulation, HDI; Human Development Index, IFA;

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1 2 3 Iron/Folic Acid, IUGR; Intra Uterine Growth Restriction, LAZ; Length-for-Age Z-score, LBW; Low Birth 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 Weight, LMIC; Low and Middle Income Countries, LMP; Last Menstrual Period, LQAS; Lot Quality 6 7 Assessment Sampling, MISAME; MIcronutriments pour la SAnté de la Mère et de l’Enfant, MMS; 8 Multiple-Micronutrient Supplements, RCT; Randomized Controlled Trial, SAE; Serious Adverse Events, 9 10 SGA; Small-for-Gestational Age, UNICEF; United Nations International Children's Emergency Fund, 11 12 UNIMMAP; UNICEF/WHO/UNU International Multiple Micronutrient, WASH; water, sanitation and 13 hygiene, WAZ; Weight-for-Age Z-score, WHO; World Health Organization, WHZ; Weight-for-Height Z- 14 15 score, WLZ; Weight-for-Length Z-score. 16 17 18 Acknowledgements For peer review only 19 The MISAME Study Group thanks the pregnant women and their families for the time spent in this 20 21 study. We would like to acknowledge the staff of AFRICSanté; including Henri Somé for his support on 22 23 the CAPI software, the field medical Dr. Anderson Compaoré, the data collectors (midwives, 24 interviewers and femmes accompagnantes), Dr. Alain Hein for the standardization exercises and Dr. 25 26 Hermann Lanou for his support during the training. We would like to thank Dr. Sheila Isanaka for her 27 28 collaboration during the formative research. Our private sector partner Nutriset (France) is 29 acknowledged for donating the BEP supplements. 30 31 32 Competing interests statement 33 KV: none to declare 34 BdK: none to declare 35 LCT: none to declare 36 37 NDC: none to declare http://bmjopen.bmj.com/ 38 MO: none to declare 39 TDC: none to declare 40 GHC: none to declare 41 RG: none to declare 42 CL: none to declare 43 LH: none to declare 44

PK: none to declare on September 29, 2021 by guest. Protected copyright. 45 46 Contributor Statement 47 48 KV wrote the manuscript; 49 PK, LH, CL, NDC, LCT, KV, BdK, TDC, GHC designed the study and the protocol; 50 PK, LH, LCT, KV, BdK designed the study material tools; 51 LCT, LH, KV, BdK, MO, RG trained the field data collectors; 52 TDC, GHC critically reviewed and revised the manuscript; 53 All authors contributed substantially to the manuscript and approved the final version. 54 55 Correspondence 56 [email protected] 57 58 [email protected] 59 60 Funding Information

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1 2 3 This work was funded by the Bill and Melinda Gates Foundation. Grant number OPP1175213. 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 The trial is registered on Clinical Trials.gov (identifier: NCT03533712) prior to recruitment. Enrolment 6 7 started in October 2019. 8 9 Statement 10 Nutriset will not be involved in the design, the implementation, the analysis and the write-up of the 11 results. 12 The BMGF will not be involved in the design, the implementation, the analysis and the write-up of the 13 results. 14 15 REFERENCES 16 17 1. Lee AC, Katz J, Blencowe H, Cousens S, Kozuki N, Vogel JP, et al. National and regional 18 estimates of term andFor preterm peerbabies born smallreview for gestational only age in 138 low-income and middle- 19 income countries in 2010. The Lancet Global Health. 2013;1(1):e26-e36. 20 21 2. Katz J, Lee AC, Kozuki N, Lawn JE, Cousens S, Blencowe H, et al. Mortality risk in preterm and 22 small-for-gestational-age infants in low-income and middle-income countries: a pooled country 23 analysis. The Lancet. 2013;382(9890):417-25. 24 3. van Rossum CTM, Fransen HP, Verkaik-Kloosterman J, Buurma-Rethans EJM, Ocké MC. Dutch 25 National Food Consumption Survey 2007–2010: Diet of children and adults aged 7 to 69 years2011. 26 4. Lee AC, Kozuki N, Cousens S, Stevens GA, Blencowe H, Silveira MF, et al. Estimates of burden 27 and consequences of infants born small for gestational age in low and middle income countries with 28 INTERGROWTH-21st standard: analysis of CHERG datasets. bmj. 2017;358. 29 5. Stevens B, Buettner P, Watt K, Clough A, Brimblecombe J, Judd J. The effect of balanced 30 31 protein energy supplementation in undernourished pregnant women and child physical growth in 32 low-and middle-income countries: a systematic review and meta-analysis. Maternal & child nutrition. 33 2015;11(4):415-32. 34 6. Organization WH. WHO recommendations on antenatal care for a positive pregnancy 35 experience: World Health Organization; 2016. 36 7. Krishna U, Bhalerao S. Placental insufficiency and fetal growth restriction. The Journal of http://bmjopen.bmj.com/ 37 Obstetrics and Gynecology of India. 2011;61(5):505-11. 38 8. Mridula D, Mishra C, Chakravorty A. Dietary intake of expectant mother. Indian J Nutr Diet. 39 40 2003;40(1):24-30. 41 9. Tang AM, Chung M, Dong K, Terrin N, Edmonds A, Assefa N, et al. Determining a global mid- 42 upper arm circumference cutoff to assess malnutrition in pregnant women. Food and Nutrition 43 Technical Assistance. 2016. 44 10. Branca F, Grummer-Strawn L, Borghi E, Blössner Md, Onis Md. Extension of the WHO 45 maternal, infant and young child nutrition targets to 2030. SCN News. 2015(41):55-8. on September 29, 2021 by guest. Protected copyright. 46 11. Keats EC, Haider BA, Tam E, Bhutta ZA. Multiple-micronutrient supplementation for women 47 during pregnancy. Cochrane Database of Systematic Reviews. 2019(3). 48 12. Hambidge KM, Westcott JE, Garcés A, Figueroa L, Goudar SS, Dhaded SM, et al. A 49 50 multicountry randomized controlled trial of comprehensive maternal nutrition supplementation 51 initiated before conception: the Women First trial. The American journal of clinical nutrition. 52 2019;109(2):457-69. 53 13. Ota E, Hori H, Mori R, Tobe-Gai R, Farrar D. Antenatal dietary education and supplementation 54 to increase energy and protein intake. Cochrane Database of Systematic Reviews. 2015(6). 55 14. Imdad A, Bhutta ZA. Maternal nutrition and birth outcomes: effect of balanced protein- 56 energy supplementation. Paediatr Perinat Epidemiol. 2012;26 Suppl 1:178-90. 57 15. Kramer MS, Kakuma R. Energy and protein intake in pregnancy. Cochrane Database of 58 59 systematic reviews. 2003(4). 60

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1 2 3 16. Bill and Melinda Gates Foundation (BMGF). Framework and Specifications for the Nutritional BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 Composition of a Food Supplement for Pregnant and Lactating Women (PLW) in Undernourished and 5 Low-Income Settings. Expert consultation Seatle, US; 2016 2016. 6 7 17. Roberfroid D, Huybregts L, Lanou H, Henry M-C, Meda N, Menten J, et al. Effects of maternal 8 multiple micronutrient supplementation on fetal growth: a double-blind randomized controlled trial 9 in rural Burkina Faso–. The American journal of clinical nutrition. 2008;88(5):1330-40. 10 18. Huybregts L, Roberfroid D, Lanou H, Menten J, Meda N, Van Camp J, et al. Prenatal food 11 supplementation fortified with multiple micronutrients increases birth length: a randomized 12 controlled trial in rural Burkina Faso. Am J Clin Nutr. 2009;90(6):1593-600. 13 19. United Nations Development Programme. Human Development Reports [Internet] 2019. 14 Available from: http://hdr.undp.org/en/countries/profiles/BFA. 15 20. Unicef. The state of the world's children: children with disabilities: UN; 2013. 16 17 21. INSD MD, Macro I. Enquête démographique et de santé et à indicateurs multiples (EDSBF- 18 MICS IV), Rapport préliminaire,For peer Burkina Faso, review 2010. Ouagadougou: only INSD. 2011;40. 19 22. Institut National de la Statistique et de la Démographie (INSD) et ICF International. Enquête 20 Démographique et de Santé et à Indicateurs Multiples du Burkina Faso 2010. Calverton, Maryland, 21 USA : INSD et ICF International. 2012. 22 23. MOH. National Iodine Status and Anemia Survey, Burkina Faso. 2014. 23 24. MOH. MdlS. Enquête Nutritionnelle Nationale 2013. 2013. 24 25. Savy M, Martin-Prével Y, Sawadogo P, Kameli Y, Delpeuch F. Use of variety/diversity scores 25 26 for diet quality measurement: relation with nutritional status of women in a rural area in Burkina 27 Faso. European journal of clinical nutrition. 2005;59(5):703. 28 26. Huybregts LF, Roberfroid DA, Kolsteren PW, Van Camp JH. Dietary behaviour, food and 29 nutrient intake of pregnant women in a rural community in Burkina Faso. Maternal & child nutrition. 30 2009;5(3):211-22. 31 27. Jones L, de Kok B, Moore K, de Pee S, Bedford J, Vanslambrouck K, et al. Acceptability of 12 32 fortified balanced energy protein supplements-Insights from Burkina Faso. Maternal & Child 33 Nutrition. 2020:e13067. 34 35 28. de Kok BM, K., Jones, L., Vanslambrouck, K., Toe, L.C., Ouedraogo, M., Ganaba, R., de Pee, S., 36 Bedford, J., Lachat, C., Kolsteren, P., Isanaka, S. . Acceptability and at-home consumption of two 37 fortified balanced energy-protein supplements among pregnant women in rural Burkina Faso: a http://bmjopen.bmj.com/ 38 mixed methods study. Maternal & Child Nutrition, manuscript in production 2020. 39 29. Villar J, Altman D, Purwar M, Noble J, Knight H, Ruyan P, et al. The objectives, design and 40 implementation of the INTERGROWTH-21st Project. BJOG: An International Journal of Obstetrics & 41 Gynaecology. 2013;120:9-26. 42 30. de Onis M, Branca F. Childhood stunting: a global perspective. Matern Child Nutr. 2016;12 43 Suppl 1:12-26. 44 45 31. Huybregts LF, Roberfroid DA, Kolsteren PW, Van Camp JH. Dietary behaviour, food and on September 29, 2021 by guest. Protected copyright. 46 nutrient intake of pregnant women in a rural community in Burkina Faso. Matern Child Nutr. 47 2009;5(3):211-22. 48 32. Becquey E, Huybregts L, Zongrone A, Le Port A, Leroy JL, Rawat R, et al. Impact on child acute 49 malnutrition of integrating a preventive nutrition package into facility-based screening for acute 50 malnutrition during well-baby consultation: A cluster-randomized controlled trial in Burkina Faso. 51 PLoS medicine. 2019;16(8). 52 33. Salomon L, Alfirevic Z, Da Silva Costa F, Deter R, Figueras F, Ghi T, et al. ISUOG Practice 53 54 Guidelines: ultrasound assessment of fetal biometry and growth. Ultrasound in Obstetrics & 55 Gynecology. 2019;53(6):715-23. 56 34. Cox JL, Holden JM, Sagovsky R. Detection of postnatal depression: development of the 10- 57 item Edinburgh Postnatal Depression Scale. The British journal of psychiatry. 1987;150(6):782-6. 58 35. Femi A. Perception of performance appraisal and workers’ performance in Wema Bank 59 Headquarters, Lagos. Global Journal of Arts, Humanities and Social Sciences. 2013;1(4):89-101. 60

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1 2 3 36. Martin-Prével Y, Allemand P, Wiesmann D, Arimond M, Ballard T, Deitchler M, et al. Moving BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 forward on choosing a standard operational indicator of women’s dietary diversity. FAO; 2015. 5 6 7 8 9 Figure legend 10 Figure 1 : Study design of the RCT 11 12 13 14 15 16 17 18 For peer review only 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 5 6 1. PARTICIPANT INFORMATION SHEET 7 8 9 THE EFFECT OF BALANCED ENERGY-PROTEIN SUPPLEMENTATION DURING PREGNANCY AND 10 LACTATION ON BIRTH OUTCOMES AND INFANT GROWTH IN RURAL BURKINA FASO 11

12 13 Coordinating Investigator : Prof. Dr. Patrick Kolsteren 14 Principal Investigator: Dr. Laeticia Celine TOE 15 Sponsor of the study: Ghent University 16 Participant Number: |__||__||__||__||__| _ |__||__| 17

18 For peer review only 19 20 Dear madam, 21 22 You are invited to participate in a study that wants to study the effect of providing a food supplement 23 during pregnancy and lactation on the growth of your baby. The study is a collaboration between the 24 Institut de Recherche en Sciences de la Santé (IRSS) and AfricSanté in Burkina Faso, Ghent University 25 of Belgium and the International Food Policy Research Institute (IFPRI) in the USA. Before you decide 26 to participate in this study, it is important that you read/ are read and explained to this form, because 27 28 it explains your rights and our responsibilities to you. In this information and consent form, the 29 purpose, examinations, advantages, risks and inconveniences related to this study are explained. The 30 available alternatives and the right to withdraw your consent to participate at any time are also 31 described below. No promises or guarantees can be made about the results of the study. You have the 32 right to ask questions at any time, for example about the possible and/or known risks related to this 33 study. 34 35 36 37 PURPOSE AND DESCRIPTION OF THE STUDY http://bmjopen.bmj.com/ 38 This research study will provide more evidence and insight into how we can improve pregnancy 39 outcomes , birthweight of babies and the growth of newborns. With this study, we aim to assess 40 whether the provision of the food supplement under investigation can improve the growth of fetuses 41 during pregnancy and that of infants during their first 6 months of life. In addition, we will evaluate 42 how the food supplements that are offered to you improve the nutritional quality of the human milk 43 produced for your child, as well as your and your child’s body composition. 44

on September 29, 2021 by guest. Protected copyright. 45 46 We will thus compare the effect of 2 different approaches. One is to give you a tablet during your 47 pregnancy and with the other approach participants will also receive a food supplement next to the 48 tablet. The tablet is what you normally received for any pregnancy according to the guidelines of the 49 ministry of health of Burkina Faso. We will provide the supplement for the entire duration of your 50 pregnancy. After birth some mothers will continue to receive a food supplement for six months. Who 51 receives what is entirely determined by chance. In total, 1776 pregnant women living in the catchment 52 area of the health centers of de Boni, Dohoun, Dougmato II, Karaba, Kari and Koumbia will be recruited 53 to participate in this study. 54

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1 2 3 HOW THE STUDY IS DONE BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 If you are confirmed pregnant and accept to participate in this study, the midwife will do a full medical 5 check-up of you and your baby according to medical standards. She will also inform you about a good 6 7 dietary practices for you during your pregnancy. 8 After the check-up we will provide you either with the tablets, or the food supplements. 9 10 Thereafter, you will receive a daily visit from the community health worker who will ask you to take 11 the tablet or eat the food supplement. A ultrasound examination will be planned for you within 2 12 weeks after your first consultation and will be performed by the project doctor. We will also arrange 13 for you regular check-ups of your pregnancy by the midwife or the medical doctor. 14 15

16 17 EXAMINATIONS IN THE CONTEXT OF THE STUDY 18 If you accept to participateFor inpeer the study andreview if you and your only child meet all the conditions for 19 participating in the study, the following tests and examinations will be performed: 20 21 - We will take your weight t and height and ask you your age at inclusion. 22 - The midwife will invite you for antenatal check-ups according to the guidelines of the ministry 23 of health and the medical doctor will perform an echography of your baby. This is not invasive 24 25 and has no known risks. It is like taking a picture of your baby. As part of the routine check-up 26 we will take fingerpick blood to test whether you are anemic and give you treatment when 27 necessary. 28 - We will also ask you questions about your mental wellbeing, e.g. how you are living your 29 pregnancy and if something bothers you in your everyday life. 30 - After delivery we will again take fingerpick blood to test whether you are anemic and give you 31 treatment when necessary. 32 - When you deliver we will weigh your baby and measure his/her length and head and chest 33 circumferences. 34 35 - After birth we will visit you or ask you to come to the health center every month to have the 36 weight and the length of you baby taken. At that time we will also ask you question on illnesses 37 your child might have had in the previous week and on what he/she has been eating. http://bmjopen.bmj.com/ 38 - We will also perform a finger/heel prick on your baby to see if he/she is anemic and a 39 treatment will be provided if necessary 40 - Halfway through pregnancy in some of the participating women, we will ask you questions 41 about what and how much they have eaten the day before the interview. This should take 42 about half an hour. 43 44 45 When the doctor identifies a medical problem he will see to it that you receive the appropriate on September 29, 2021 by guest. Protected copyright. 46 information and necessary treatment, and will refer you when necessary. 47 48 If your child suffers from a disease or undernutrition, he/she will be treated in the best way possible. 49 In such case, your child will be referred to the local health center or the district hospital, for further 50 physical examination by a medical doctor. 51 52

53 54 VOLUNTARY PARTICIPATION 55 You participate entirely voluntarily in this study. You have the right to refuse to participate in the study. 56 Your decision to participate or not in this study, or to stop your participation in this study will have no 57 influence whatsoever on present and future medical consultations and possible treatments. You also 58 have the right yourself to stop your participation in the study at any time, even after you have signed 59 60

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1 2 3 this informed consent form. The withdrawal of your consent will not cause any disadvantage or loss of BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 advantages / privileges. 5 6 7 RISKS AND INCONVENIENCES 8 Finger-prick blood will be taken from you at the first ante-natal consultation. You will experience a 9 prick in your finger, however the prick is not invasive. Antiseptic measures will be taken to prevent any 10 inflammation/ infection of your finger. 11 12 The food supplement used in this study is safe for you and your baby. The food supplement contains 13 milk, sugar, oil, peanut butter and a mix of vitamins. Previous studies in Houndé that provided similar 14 food supplements during pregnancy have not documented any complications during pregnancy or 15 16 delivery. However, the supplement contains milk, which can cause bloating, flatulence and other 17 digestive discomfort in some people. We encourage you to notify us, should you experience these 18 effects, so we can takeFor measures peer to insure youreview the best comfort only possible. 19 20 All other investigation are routinely done as part of the follow-up of pregnancy. 21 22 Some of you will be asked to donate a small amount of breast milk at two time points during the study 23 (between 1-2 months and 3-4 months age of your child). This will not diminish the amount of milk for 24 25 your child or influence lactating performance. We will ask your permission for this donation again 26 when the time comes. 27 28 In a random sample of all participants, we will assess body composition after delivery in the mother 29 and the infant, using a special water that has been proven safe for such use. If you are chosen for 30 that examination, we will provide detailed explanations of the procedure to you and a specific 31 consent will be ask before we perform the assessment. Your privacy will be respected at all times 32 33 34 35 BENEFITS 36 We expect to show that taking food supplements during pregnancy and lactation will help children 37 grow better and improve the quality of breastmilk. If this would be the case we will have the possibility http://bmjopen.bmj.com/ 38 to change policy to provide supplements to pregnant and lactating women. 39 40

41 42 PROTECTION OF YOUR PRIVATE LIFE 43 Your identity and your participation in this study will be treated strictly confidential. The specific 44 information we obtain from you will not be shared with anybody, except the study investigators and 45 partner institutes. Your identity remains secret since your personal information will only be designated on September 29, 2021 by guest. Protected copyright. 46 by a unique participant number. Your name will not appear in any reports or publication resulting from 47 this study. After the study is completed, you may request information about the study results. As soon 48 as possible (maximum 5 years) after the study is completed, all personal information from participant 49 50 shall be deleted from all databases to ensure complete anonymity. Those anonymized databases shall 51 be shared with other researchers to advance research on mother and child health. This shall be done 52 in strict accordance with international laws and regulations about privacy. 53 54 55 ETHICS COMMITTEE 56 Before starting, this study has been reviewed and approved by an independent Ethics Committee in 57 Belgium, namely the Ethics Committee of the University Hospital in Ghent and it has been reviewed 58 59 and approved by the Ethics Committee Centre Muraz in Burkina Faso. These committees also perform 60

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1 2 3 continuous reviews of the study during its progression to make sure the study is carried out in the BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 safest possible way. 5 6 7 COMPENSATIONS AND INCIDENTS THAT MAY OCCUR IN THIS STUDY 8 9 All costs related to sickness occurring during the study will be reimbursed to you. Investigators shall 10 seek to provide a compensation and/or the best possible treatment in the event of damages/ injuries 11 that may occur as a result of your participation in this study. For any other damages thought to be 12 related to your participation in this study, and occuring after the study has been completed, participant 13 may file a complaint to the relevant jurisdiction, which will treated in accordance with applicable laws 14 15 in Burkina Faso. 16 17 CONTACT PERSONS IN CASE YOU HAVE QUESTIONS ABOUT THIS STUDY 18 For peer review only 19 If you have any questions concerning your participation in this study, or think you have been injured 20 as a result of the study or have a reaction to the study food, inform the village health worker who visits 21 22 you. S/he will bring you immediately in contact with the project coordinator or the project doctor. All 23 village health workers have cell phones to contact these persons directly. You may also contact, now, 24 during or after the study: 25 26 The principle investigator in Burkina Faso: Dr. Laeticia Celine Toe, Tel : +226 71 00 72 72 27 The president of the ethic committee: Dr. Adama Dembélé, Tel: +226 20 97 01 02 28 Or The local project Coordinator : Mr. Moctar Ouédraogo, Tel: +226 70 23 81 98 29 Or project medical doctor: Compaoré S. Anderson Casimir Tel : +226 70 57 51 02 30 31 32 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 Informed consent 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 7 Before you agree to participate in the informal consensus activity, you need to be aware that: 8 9  This study was presented and cleared by the Ethical Review Board of the Ghent University and 10 the ethics committee of Centre Muraz, Burkina Faso. 11 12  This clearance is not to be taken as an obligation to take part in this study. 13 14  Your participation is only voluntary and will be confirmed by signing this form. If you wish, you 15 can withdraw from this study at any point, even after signing this form; you can withdraw by 16 contacting the researcher (below) through email or telephone. You do not have to motivate or 17 explain the decision of withdrawal. In case of withdrawal, your data already collected will be 18 used to assess studyFor outcomes. peer review only 19 20  You can revise your answers to the questions if you wish so. 21  Your input will be stored anonymously; researchers not involved in the data collection will not 22 23 have access to your personal data and name. Anonymized databases shall be shared with other 24 researchers for to advance research on maternal and child health. 25  You can contact the researcher or the coordinator of the project at any time if you wish to obtain 26 more information regarding this study 27 28  There are very limited risks related to your participation in this study. Those are mentioned 29 under the section “ risks and inconveniences” of the information sheet. However, in accordance 30 with the Directive concerning experiments on humans (07/05/2004), a civil liability insurance 31 has been foreseen in the event of any injury or damage occurring and deemed due to your 32 33 participation in this study. 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 Part intended only for the participant 6 7 I, the undersigned, ______8 (name and surname) confirm that I have been informed about the MISAME-III clinical trial and that I 9 10 have received a copy of the Participant Information Sheet and a copy of the consent form. I confirm 11 that I have read and / or understood the four pages of the information sheet for participants. The 12 study responsible gave me enough information about the conditions and duration of the study, and 13 the possible risks and disadvantages. In addition, I had enough time to review the information and to 14 ask questions, and I received satisfactory answers. 15 16  I understand that I can terminate my participation in this study at any time after notifying the 17 study responsible and this decision will not cause any inconvenience to me or my child. 18  For peer review only 19 I am aware of the purposes for which the data is collected, processed and used in this study. 20  I am ready to give information about my medical history and that of my child, or about any 21 medication taken or participation in another study. 22 23  I voluntarily consent to participate in this study and to cooperate with the required examinations 24 and tests. 25 26  I consent to my data being shared anonymously for the benefit of research and maternal and 27 child health. 28 29  I understand that auditors, employee representatives, the Commission for Medical Ethics or 30 competent authorities may inspect my data in order to verify the information collected. By 31 signing this document, I give permission for this control. At all times my privacy will be respected. 32  I understand that the biological samples collected from me and / or my child will be sent to 33 34 Europe or the USA for analysis, and this in strict respect of my confidentiality. 35  I am aware that this study has been approved by an independent commission for medical ethics 36

linked to the University Hospital of Ghent, examined by the Ethics Committee of the Center http://bmjopen.bmj.com/ 37 Muraz in Burkina Faso and that this study will be carried out in accordance with the guidelines 38 39 Good Clinical Practice Guidelines and the Declaration of Helsinki, established for the protection 40 of people participating in clinical trials. 41 42 43 Signature (or fingerprint) of the participant 44 45 ______on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 In case of minority 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 Informed consent of the legal representative 7 8 9 I, the undersigned, ______10 (name and surname), legal representative of 11 ______(name and surname) confirm that I have 12 been informed about the MISAME-III clinical trial and that I have received a copy information sheet 13 for participants and a copy of the consent form. I confirm that I have read and / or understood the 6 14 pages of the information sheet for participants. The study responsible gave me enough information 15 16 about the conditions and duration of the study, the procedures, the advantages and possible 17 disadvantages. In addition, I had enough time to review the information and to ask questions, and 18 received satisfactory Foranswers. peer review only 19 20  I understand that I can withdraw my consent at any time after having informed the study 21 22 managers and this decision will not cause any inconvenience to my child / wife / ward or to 23 myself. 24  I am aware of the purposes for which the data is collected, processed and used in this study. 25 26  I voluntarily consent to my child / wife / ward participating in this study. 27 28  I understand that auditors, employee representatives, the Commission for Medical Ethics or 29 competent authorities may inspect their data in order to verify the information collected. By 30 signing this document, I give permission for this control. At all times his privacy will be respected. 31

32 33 Date: | ___ | ___ | / | ___ | ___ | / | ___ | ___ | ___ | ___ | (dd / mm / yyyy) 34 35 36 Signature (or fingerprint) of the participant's legal representative http://bmjopen.bmj.com/ 37 ______38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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1 2 3 Part intended for the investigator 4 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 5 6 I, the undersigned, ______confirm that I 7 8 have informed, ______9 10 (full name of the participant) and that she has: 11 12 13 Consent to participate in the study 14 Refused to participate in the study 15 16 17 18 Reason for refusalFor (mark peer not filled in review if no reason provided) only 19 ______20 21 ______22 23 24 Date | ___ | ___ | / | ___ | ___ | / | ___ | ___ | ___ | ___ | (dd / mm / yyyy) 25 26 27 Signature: ______28 29 30 31 32 33 34 35 If the participant is unable to read and / or write, an impartial witness should be present during the consent discussion. 36

After having read and explained the information sheet and the informed consent form to the participant. After she has http://bmjopen.bmj.com/ 37 verbally consented to her participation in the study, and has affixed her fingerprint, the witness should complete the name 38 of the participant, add the date, and personally sign and date the consent form. By signing the consent form, the witness 39 certifies that the information in the information sheet and the consent form have been precisely explained and understood 40 by the participant and that the participant has freely given her consent. 41 42 43 Name and first name (s) of witness: 44 45 ______on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 Signature of witness : 51 ______52 53 54

55 56 Date: | ___ | ___ | / | ___ | ___ | / | ___ | ___ | ___ | ___ | (dd / mm / yyyy) 57 58 59 60

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1 2 3 BMJ Open: first published as 10.1136/bmjopen-2020-038393 on 24 March 2021. Downloaded from 4 Contact Principal investigator Contact of project medical doctor 5 Compaoré S. Anderson Casimir 6 TOE Laeticia Celine Tel : +226 70 57 51 02 7 IRSS/DRO site Ecole Jamot Email : [email protected] 8 01 BP 545 Bobo-Dioulasso

9 Tél: +226 71 00 72 72

10 Email: [email protected] 11 12 13 14 15 16 17 18 For peer review only 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 http://bmjopen.bmj.com/ 38 39 40 41 42 43 44 45 on September 29, 2021 by guest. Protected copyright. 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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