Egg Intervention During Pregnancy in Indonesia (PRECODE)

Effect on Pregnancy Outcomes, Infant Growth and Development of an Egg Intervention During Pregnancy in Indonesia

London School of Hygiene and Tropical Medicine, University of Aberdeen, University of Brighton, University College, London, Royal Veterinary College, Birkbeck, University of London, The International Livestock Research Institute (ILRI), Cheikh Anta Diop University, Senegal, National Institute of Nutrition, India, International Centre for Research in Agroforestry, Science Made Simple, Liverpool School of Tropical Medicine, International Initiative for Impact Evaluation, Digital Green Foundation, SOAS, University of London, University of Sheffield

The study consists of two arms: 1) intervention group using eggs as supplementary food given from 2nd trimester of pregnancy to birth, and 2) observational group of pregnant mothers. it aims to assess the effectiveness of improving dietary quality during pregnancy on the epigenetic and stunting related outcomes (growth and development) in infants, who will be followed up until 24 months old

The study aims to assess the impact of improving dietary quality during pregnancy on the epigenetic and stunting related outcomes in infants. The open-label intervention study would be conducted alongside the observational study in the same study setting by recruitment of additional number (n=153) of pregnant women. Thus, a total of 653 pregnant women would be enrolled in the study; 153 women would be randomized to intervention arm and 500 to the control arm who would form an observational cohort of women and newborns as described above. The intervention group women will be provided one egg three times per week from recruitment (2nd trimester) until term. The control group women will receive standard intervention in the form of Ante Natal Care from village midwives or Public Health Centre (IFA tablet, calcium tablet, nutrition counselling).

Weight Gain, Anemia, Iron Deficiency, Birth Weight, Birth Length, Child Development, Zinc Deficiency, Folate Deficiency, B12 Deficiency Vitamin, Amino Acid Deficiency, Mineral Deficiency, Vitamin A Deficiency, Fatty Acid Deficiency, Parasite Infestation, E. Coli Infection, Salmonella Infections, Shigella Infection, Protozoan Infections

A total of 653 pregnant women will be recruited from 40 villages in three sub-districts. Pregnant women will be randomly allocated into the intervention or control groups. Pregnant women in the intervention group will receive boiled eggs three times per week from 2nd trimester (16-20 weeks) until delivery.

In the observational cohort, pregnant mothers in the 2nd trimester (n=500) will be recruited and they will be followed up until their children are 24 months old. The control group women will receive standard intervention in the form of Ante Natal Care from village midwives (Polindes) or Puskesmas (IFA tablet, calcium tablet, nutrition counselling).

The intervention group women (n=153) will be provided one egg three times per week from recruitment (2nd trimester) until term along with the standard Ante Natal Care.

Eggs are boiled until the white and yolk are firm (ca. 8 minutes) to maintain quality and safety and ensure the eggs are safe for consumption.

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess fine and gross motor skills among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess expressive and receptive language among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess behavior among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess executive function among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess empathy among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess problem solving among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess attention among children aged 10 to 14 months

Oxford Neurodevelopment Assessment (OX-NDA) is used to assess social-emotional reactivity among children aged 10 to 14 months

INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess motor development among children aged 20 to 24 months

INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess cognition among children aged 20 to 24 months

INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess language among children aged 20 to 24 months

INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess social-emotional development among children aged 20 to 24 months

MacArthur-Bates Communicative Development Inventories (CDI) is used to assess vocabulary comprehension scale among children aged 10 to 12 months

MacArthur-Bates Communicative Development Inventories (CDI) is used to assess vocabulary production among children aged 10 to 12 months

Genome-wide analysis of epigenetic states using the Illumina Infinium Methylation EPIC 850k Bead Chip (EPIC array) will be performed for selected samples from the core cohort. The outcomes will be the epigenetic state of a large number of genes which are associated with child stunting.

All samples (newborn, children 24 mo, parents) will be analyzed using Next Generation Bisulphite Amplicon Sequencing (BSAS) from Illumina MiSeq platform in targeted regions of the genome. The outcomes will be profiles of specific epigenetic markers of birth anthropometry, adult stature, metabolic state, and cognitive ability.

All measurements will be taken to the nearest 0.1 kg using standard procedures with SECA weighing machine.

All measurements will be taken to the nearest 0.1 kg using standard procedures with SECA weighing machine.

All measurements will be taken to the nearest milimeter using standard procedures with SECA stadiometer/infantometer.

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their hemoglobin.

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum ferritin

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum transferrin receptor

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum zinc

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum retinol

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their RBC folate

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their RBC fatty acids

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers for their serum essential amino acids

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum methylmalonic acid

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum choline

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum betaine

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum RBP

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum hepcidine

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Nutritional status measured by biochemical assessment to the mothers and children. Both the pregnant mothers and children will be measured for their serum homocysteine

Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery

Faecal microbiome would be analyzed using 16S RNA sequencing of the V4 region on the Illumina MiSeq and BSAS.

Inclusion Criteria: Woman is between 16 and 20 weeks of pregnancy based on the date of the first day of her last menstrual period. She is 18-40 years of age. She is planning to remain in the study area over the next 30 months. She is of Sasak ethnicity Exclusion Criteria: She is expecting multiple births. She has a known egg allergy.

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