Egg Intervention During Pregnancy in Indonesia

Study Overview
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Brief Summary
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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

Detailed Description
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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).

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

Conditions
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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

Intervention / Treatment
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* Dietary Supplement: Egg intervention


Participation Criteria
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Eligibility Criteria
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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.

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 40 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Prevention
Allocation: Randomized
Intervention Model: Parallel Assignment
Interventional Model Description: 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.
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| No Intervention: Observational cohort (control group)<br>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). | |
| Experimental: Intervention group<br>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. | Dietary Supplement: Egg intervention<br>* 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.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Prevalence of stunting | Z-score of LAZ <-2 SD based on WHO 2006 | birth until 24 months after delivery | 
| Proportion of children 10-14 months with impaired fine and gross motor skills | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess fine and gross motor skills among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired expressive and receptive language | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess expressive and receptive language among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired behavior | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess behavior among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired executive function | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess executive function among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired empathy | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess empathy among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired problem solving | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess problem solving among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired attention | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess attention among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 10-14 months with impaired social-emotional reactivity | Oxford Neurodevelopment Assessment (OX-NDA) is used to assess social-emotional reactivity among children aged 10 to 14 months | 10-14 months of age | 
| Proportion of children 20-24 months with impaired motor development | INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess motor development among children aged 20 to 24 months | 20-24 months of age | 
| Proportion of children 20-24 months with impaired cognition | INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess cognition among children aged 20 to 24 months | 20-24 months of age | 
| Proportion of children 20-24 months with impaired language | INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess language among children aged 20 to 24 months | 20-24 months of age | 
| Proportion of children 20-24 months with impaired social-emotional development | INTERGROWTH Neurodevelopment Assessment (INTER-NDA) is used to assess social-emotional development among children aged 20 to 24 months | 20-24 months of age | 
| Scores of CDI vocabulary comprehension scale in children 10-12 months | MacArthur-Bates Communicative Development Inventories (CDI) is used to assess vocabulary comprehension scale among children aged 10 to 12 months | 10-12 months of age | 
| Scores of CDI vocabulary production scale in children 10-12 months | MacArthur-Bates Communicative Development Inventories (CDI) is used to assess vocabulary production among children aged 10 to 12 months | 10-12 months of age | 
| Epigenetic state of genes associated with stunting | 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. | parents: 72 h after delivery; baby: 72 h after delivery, 24 month | 
| Epigenetic markers of birth anthropometry, adult stature, metabolic state, and cognitive ability | 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. | parents: 72 h after delivery; baby: 72 h after delivery, 24 month | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Weight gain during pregnancy | All measurements will be taken to the nearest 0.1 kg using standard procedures with SECA weighing machine. | 2nd trimester (16-20 weeks gestation) and 3rd trimester (28-32 weeks gestation) of pregnancy | 
| Birth weight | All measurements will be taken to the nearest 0.1 kg using standard procedures with SECA weighing machine. | 24 hours after birth | 
| Birth length | All measurements will be taken to the nearest milimeter using standard procedures with SECA stadiometer/infantometer. | 24 hours after birth | 
| Hemoglobin concentration | 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 | 
| Serum ferritin concentration | 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 | 
| Serum transferrin receptor concentration | 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 | 
| Serum zinc concentration | 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 | 
| Serum retinol concentration | 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 | 
| RBC folate concentration | 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 | 
| Serum vitamin B12 concentration | Nutritional status measured by biochemical assessment to the mothers for their serum vitamin B12 | Mothers: second and third trimester of pregnancy | 
| RBC fatty acids concentration | 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 | 
| Serum essential amino acids concentration | Nutritional status measured by biochemical assessment to the mothers for their serum essential amino acids | Mothers: second and third trimester of pregnancy | 
| Serum methylmalonic acid concentration | 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 | 
| Serum choline concentration | 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 | 
| Serum betaine concentration | 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 | 
| Serum vitamin B2 concentration | Nutritional status measured by biochemical assessment to the mothers for their serum vitamin B2 | Mothers: second and third trimester of pregnancy | 
| Serum vitamin B6 concentration | Nutritional status measured by biochemical assessment to the mothers for their serum vitamin B6 | Mothers: second and third trimester of pregnancy | 
| Serum vitamin D concentration | Nutritional status measured by biochemical assessment to the mothers for their serum vitamin D | Mothers: second and third trimester of pregnancy | 
| Serum CRP concentration | Subclinical inflammation will be measured by serum CRP in pregnant mothers and children | Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery | 
| Serum AGP concentration | Subclinical inflammation will be measured by serum AGP in pregnant mothers and children | Mothers: second and third trimester of pregnancy; children: 6 months (+/- 2 weeks) after delivery; breastmilk: 3 and 6 months (+/- 2 weeks) after delivery | 
| Serum RBP concentration | 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 | 
| Serum hepcidine concentration | 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 | 
| Serum homocysteine concentration | 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 | 
| Serum HbA1C concentration | Gestational diabetes status will be assesed to the mothers for their serum HbA1C | Mothers: second and third trimester of pregnancy | 
| Fecal myeloperoxidase (MPO) | Gut inflammation from fecal will be measured by faecal myeloperoxidase (MPO) using ELISA | baby: 1, 6, 24 months of age | 
| Fecal α1-antitrypsin (AAT) | Gut inflammation from fecal will be measured by fecal α1-antitrypsin (AAT) using ELISA | baby: 1, 6, 24 months of age | 
| Soil-transmitted helminths infection | Fecal parasites from fecal will be assesed by Kato Katz and confirmed by qPCR | mothers: 3rd trimester (28-32 gestational weeks) of pregnancy; baby: 1, 6, 24 months of age | 
| Bacteria infection | Type of bacteria (Salmonella, Shigella) from fecal will be assesed by culture method | baby: 1, 6, 24 months of age | 
| Gut microbiota | Gut microbiota species (EPEC, ETEC, EHEC, EIEC) from fecal will be assesed using qPCR | baby: 1, 6, 24 months of age | 
| Gut microbiome | Faecal microbiome would be analyzed using 16S RNA sequencing of the V4 region on the Illumina MiSeq and BSAS. | baby: 1, 6, 24 months of age | 
| Intestinal fatty acid binding protein | Intestinal fatty acid binding protein from serum will be measured using ELISA | baby: 6 months of age | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Egg, Pregnancy, Indonesia, Epigenetic, Growth, Child development