Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT02446873 |
| Other study ID # |
201409108 |
| Secondary ID |
|
| Status |
Completed |
| Phase |
N/A
|
| First received |
|
| Last updated |
|
| Start date |
February 2015 |
| Est. completion date |
February 2017 |
Study information
| Verified date |
June 2018 |
| Source |
Washington University School of Medicine |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
The proposed study will fill an important gap in the literature by examining, through a
randomized controlled trial, the effect of egg consumption on biochemical markers of choline,
vitamin B-12, lipids, and amino acids in young children in a poor rural area of Ecuador.
Children from Cotopaxi Province, Ecuador (n=180) will be randomized into one of two groups:
1) intervention, receiving one egg per day for six months; or 2) control. Baseline and
endline data on socio-economic factors, and child diet, morbidities, and anthropometry will
be collected. Blood will also be drawn from the children at these time points for nutrient
biomarker analyses. Through qualitative research the proposed study will provide insight into
the attitudes, beliefs, and use of eggs by mothers and other caregivers during the
complementary feeding period. The University of San Francisco in Quito (USFQ) will be the
lead field coordinator of the research working in partnership with Washington University in
St. Louis, Pan-American Health Organization (PAHO) and University of California, Davis.
Description:
OBJECTIVES
Objective 1: Test the efficacy daily egg consumption for six months on the macro- and
micronutrient status of young children from an indigenous community in Ecuador.
1a. Measure serum concentration of micronutrient biomarkers: choline; betaine; and vitamin
B12.
1. b. Measure serum concentrations of macronutrient biomarkers: lipids (total free and
esterified); and amino acids.
Objective 2: Characterize attitudes, beliefs, and use of eggs during the complementary
feeding period.
2. a. Describe existing attitudes, beliefs, and use of eggs in young child diets in
Ecuador.
2b. Monitor acceptability and tolerance of daily egg consumption among young children.
2c. Assess willingness of caregivers to prepare and give eggs daily to young children.
BACKGROUND AND SIGNIFICANCE
Globally, 165 million children are affected by stunting arising in part from macro- and
micronutrient deficiencies during the complementary feeding period; defined as between 6 and
24 months of age. An estimated 200 million children are not meeting their developmental
potential in part because of poor nutrition and, in particular, nutrient intake inadequacies.
Eggs provide several of these critical nutrients known to be missing in the diets of young
children living in poor countries, yet have been largely underutilized in infant and young
child nutrition.
Eggs provide essential fatty acids, proteins, and many critical micronutrients at levels
above or comparable to other animal source foods, but are relatively more affordable. Eggs
are considered a perfect protein source with an amino acid profile against which other
proteins are compared. Eggs are also highly concentrated in choline, an important precursor
for acetylcholine in neurotransmitter synthesis, phospholipids needed for cell-membrane
signaling, and methyl-group donation in the homocysteine production pathway. The evidence for
the adverse consequences of choline deficiencies come largely from animal studies, though
some studies have shown an association between choline or its metabolites and negative health
and development outcomes in children. Only one study has shown that egg consumption among
young children can be improved through counseling.
Eggs contribute to dietary diversity. They are one of three animal source food groups
together with flesh foods and dairy products, included in the seven food groups used to
assess the indicator of minimum dietary diversity in the WHO Indicators for assessing infant
and young child feeding practices. However, they are underused in the diets of young children
relative to their potential to improve child nutrition, particularly in many African
countries but even in Latin America and the Caribbean. Egg consumption the preceding day
among children 6 to 8.9 months of age was only 7.3%, 15.8% and 22.8% in Africa, Asia and
Latin America, respectively. Among children 12 to 23.9 months of age, consumption increased
to only 13.4% in Africa, 33.0% in Asia and 41.8% in Latin America and the Caribbean.
All evidence to date suggests that eggs are underutilized as complementary foods relative to
their potential to improve young child nutrition and, in particular, levels of key macro and
micronutrients. Research to test the efficacy of dietary egg intakes to improve child
nutrition in developing countries is needed. It is also needed to understand attitudes,
beliefs and use of eggs during the complementary feeding period.
Significance. Despite the potential of eggs to improve young child nutrition, to our
knowledge there have been no randomized controlled trials to test the efficacy of egg
consumption to improve the micronutrient status of young children living in developing
countries and very little evidence from egg-related interventions. There is also very limited
evidence on the attitudes, beliefs, and use of eggs by mothers and other caregivers during
the complementary feeding period. Therefore, the proposed study will fill an important gap in
the literature by examining, through a randomized controlled trial, the effect of egg
consumption on serum concentration of micronutrient biomarkers: B12; choline; and betaine and
macronutrient biomarkers: lipids (total free and esterified); and protein nutrition in young
children in a poor rural area of Ecuador. Through qualitative research the proposed study
will also shed light on the attitudes, beliefs, and use of eggs by mothers and other
caregivers during the complementary feeding period.
EXPERIMENTAL DESIGN & METHODS
University of San Francisco in Quito (USFQ) will coordinate all field activities. A
randomized controlled trial with a parallel design will be applied. A total of 180
mother-infants pairs enrolled in this study: 90 in the control group; and 90 in the egg
intervention group. Inclusion criteria for all infants include: age 6-8 mo; healthy (no
fever, congenital heart condition, or egg allergy); singleton birth; and not severely
malnourished [weight-for-length z (WLZ) > -3). There will be 180 mothers or if mother is not
available, other caregivers of infants (fathers, grandparents, aunts/uncles, siblings, etc.)
enrolled.
Participants (recruitment, screening, and consent process). Mothers/primary caregivers and
children will be recruited in Pastocalle Parish in Cotopaxi Province of Ecuador, where
formative research was conducted to ensure sufficient numbers of infants in this age
category. There will be a rolling admission process over a one-year period to ensure sample
size is reached, and potentially we will extend recruitment to adjacent parishes.
Participants will be recruited through participatory approaches, with the support of local
leaders and the active involvement of community stakeholders. There will be an initial stage
before recruitment in which the field team will build rapport with caregivers using
standardized techniques. Mothers (or other primary caregivers) will be approached and invited
to participate with their children in the study. Study staff will verbally explain the
rationale for the study, all study procedures and their risks and benefits, and will invite
mothers to participate with their children. Eligibility screening will then occur. Screening
questions posed to the mother/primary caregiver will include:
- How old are you?
- What is the child's birthday?
- Is the child healthy, without fever or congenital heart condition, and not severely
malnourished?
- Does the child have any reactions to eating to eggs, a rash, hives, difficulty
breathing?
- Was the birth of this child a singleton birth; he/she does not have a twin or triplet,
etc.
If the primary caregiver reports that the child is malnourished and/or the child has the
appearance of thinness or other symptoms such as edema, the weight and length of the child
will be measured and the weight-for-length Z score determined from the WHO Growth Standards
(2006).
Mothers or primary caregivers of children will be given a written informed consent statement
in Spanish to read and sign, indicating their consent and consent on behalf of their child.
If the mothers/primary caregiver is less than 18 years of age, informed consent process will
be sought from their parents or guardians. Participants with limited literacy will be offered
the option to have the statement read to them. Caregivers will be invited to ask any
questions about the nature of the study during this time. A potential participant will have
approximately one week to decide whether or not to participate and thus, have the opportunity
to discuss with family and friends.
Study field procedures. Children will be randomized through a computer-generated number
sequence program to one of two comparison groups in a 1:1 ratio: 1) control; and 2) egg
supplementation.
The baseline survey will first be administered to all mothers/primary caregivers in community
centers, health care posts or nurseries on assigned dates. Household level socioeconomic,
demographic, water, hygiene, and sanitation information will be collected. Child dietary
intake data will be collected using a food frequency questionnaire of commonly used
complementary foods, supplemented with targeted questions about child and household egg
consumption patterns. Morbidities will be assessed using a two-week recall, with particular
focus on respiratory infections given its high prevalence in this population and any symptoms
that might be associated with egg allergies. Anthropometric measures of child length and
weight will then be taken using international protocols. Phlebotomists from an experienced
and well-equipped laboratory in Ecuador, NETLAB, will collect 3 ml of blood from children. A
numbing cream will be used for the children.
A weekly supply of eggs will be given to the mother in the egg supplementation group with
instructions to give the child one egg per day. Additional messages will be communicated
regarding hygiene practices and egg handling and preparation. Eggs will not require
refrigeration, but will need to be stored in a cool location. Mothers/caregivers from the
control group will be provided vouchers for 6 months of eggs after the completion of the
intervention study. All mothers will receive an attendance card that will track participation
in study events including baseline and endline visits and qualitative assessment as
appropriate. At the completion of the study, all mothers will receive a small personal care
gift (lotion or perfume) in appreciation for their contributions.
Motivational workshops will periodically be held by the study coordinator and team to receive
feedback from the participants and continue to provide rationale for study participation.
These workshops effectively engage caregivers in the study and can minimize
losses-to-follow-up. As well, regular meetings with community leaders and health workers will
be held to communicate and dialogue about study progress and findings.
Eggs will be purchased locally in Pastocalle and distributed to families on a weekly basis by
the study team. (The Food and Agriculture Organization (FAO) will potentially support USFQ to
implement a poultry development project that could linked with this study. With technical
support from Ministry of Agriculture extension workers, families from Pastocalle will receive
and raise chickens for laying eggs.) Eggs for this study will be purchased from these
households and an already established project of egg production developed through the Parish
Council to support individuals with disabilities.
The study coordinator and enumerator team will contact mothers/primary caregivers lost to
follow-up by mobile phones, visits, and through family members and friends, in order to
encourage them to return to the study and to collect information about the reasons they have
left the study.
Nutrient biomarkers: Nutrient biomarkers assessed for this study will include vitamin B12,
choline, betaine, lipids and amino acids. NETLAB in Quito, Ecuador will be responsible for:
1) collecting the blood samples; 2) processing and completing the extraction process for
choline/betaine, lipids and amino acids; 3) shipping these samples to Washington University
Biomedical Mass Spectrometry Resource Facility; and 4) completing the assay for vitamin B12.
Trained phlebotomists will travel to Pastocalle and surrounding parishes periodically to
collect blood samples (3 mL from each child). The ethylenediaminetetraacetic acid (EDTA)
plasma will be obtained by collecting blood into Vacutainer Tubes marked with participant ID
numbers only. The samples will be placed on ice and transported to the NETLAB facility in
Quito.
Samples will be processed and the extraction completed for choline, betaine, lipids, and
amino acids. For the choline and betaine analytes, an aliquot of plasma is mixed with
acetonitrile containing d9-choline and d9-betaine. The acetonitrile extracts are centrifuged,
the supernatant is transferred to a micro-centrifuge tube, and dried to completion under
nitrogen gas. The micro-centrifuge tube is labeled with participant ID# and date of
collection. An extraction process is also completed for processing the lipids and amino
acids. These extracted samples will then be shipped on dry ice to Washington University
Biomedical Mass Spectrometry Resource Facility in St. Louis, MO by Globalex shipping company.
The choline and betaine assays will be tested using LC/MS/MS protocol (Appendix 1). The
remaining sample aliquots will be frozen at -80 degrees C. Vitamin B12 will later be tested
using a competitive chemiluminescent enzyme immunoassay solid phase.
Qualitative research. Similar qualitative research techniques will be applied here as were
used for Focused Ethnographic Studies (FES). The qualitative method is based on grounded
theory. Grounded theory is a unique form of qualitative analysis in that it has an inductive
orientation for constructing theory out of a systematic analysis of collected data. In-depth
interviews and focus groups will be conducted with mothers, grandmothers, and other
caregivers, as well as local health care providers, community leaders, and egg producers to
understand attitudes, perceptions and use of eggs for young child nutrition.
Structured observations will be conducted in the homesteads of consenting caregivers and
child care centers to examine environmental factors, chicken rearing practices, and
complementary food storage and egg preparation and feeding practices. Additionally, we follow
a combination of three methods for the analysis of collected qualitative data: coding in
three stages; abductive reasoning; and schemes generation. Of these three, the core method is
the coding on three stages. First, open coding identifies the basic concepts in individual
responses. Second, axial coding is developed through the generation of categories and
properties that emerge from the pattern of responses and which produces relations between
categories and properties. Third, selective coding integrates and refines the main topics and
the interrelations between them. Coding will be done by USFQ, with support from investigators
from UCD and PAHO to triangulate for quality control.
Data management. USFQ will be responsible for data management of data collected in Pastocalle
and at NETLAB (with UC-Davis), and Washington University will be responsible for data
generated at the Biomedical Mass Spectrometry Resource Facility. Data entry will be initiated
early and continue throughout the study to minimize errors and increase the likelihood of
accurate and comprehensive reporting. When potential errors or missing data are identified,
managers will inquire with enumerators and caregivers/mothers to either provide appropriate
responses or explanatory notes in the data set. Double data entry will be conducted on a
sub-set of surveys to check for entry error.
Sample size and statistical analysis. Sample size was estimated based on hypothesized change
(difference-in-difference with control) in plasma vitamin B12 concentration of 59.4 pmol/L
over the 6 month period based on other previous findings of vitamin B12 supplementation and
the variability in vitamin B12 status among infants. This nutrient was selected for sample
size calculation because sufficient data was available to estimate variability and potential
effects from egg consumption in contrast to the other nutrients that will be studied. We
estimate requiring 90 children per group assuming a 20% attrition rate.
Descriptive statistics will first be applied to examine characteristics of the sample, and
the primary and secondary outcomes. Univariate tests, including chi squared and t-tests, will
then be applied to compare characteristics across trial arms and examine cross-sectional
differences in the outcome variables at baseline and endline. Longitudinal modeling for the
primary biomarker outcomes (continuous serum/plasma nutrient concentration markers) and
secondary growth (LAZ, WAZ, WLZ) and morbidity outcomes (days with diarrhea) will be
conducted using generalized least squares (GLS) with random effects. Logit models will be
applied to examine adjusted odds of threshold outcomes (nutrient deficiencies, stunting,
underweight, wasting, respiratory infection, etc.). All inference analyses for the efficacy
of eggs will use intention-to-treat. Quantitative data analyses were performed with STATA
software (version 11.1; StataCorp, College Station, TX).
For qualitative data analyses, interview transcripts will be uploaded into Nvivo10 software
and coded for themes. Top-line themes will be sub-divided into more detailed categories
according to participants' responses in the interviews. Analyses of these categories and
themes reveal patterns in the interview data that form the basis of the qualitative findings.
A framework matrix analysis of a subset of interviews in the two study arms analyzing key
themes will be applied to investigate any systematic differences in responses across study
arms.
To measure micronutrient intakes against normative levels of adequacy, the full probability
approach will be used. This method allows for a more comprehensive estimate of inadequacy
risk across the entire distribution of intakes when compared to a simple cut-point approach
in which intakes above and below a threshold level are estimated. Child micronutrient intake
will be standardized by subtracting the estimated average requirement (EAR) of the reference
distribution and then dividing by the SD of the reference distribution. The resulting number
will be compared to a standard normal distribution to obtain the probability of inadequacy,
the area to the right under the standard normal curve, and averaged across the sample.
Profile of the Target Population. The target population for this study will be infants ages
6-8 months old from the Pastocalle Parish, which is populated mainly by indigenous
communities, in Cotopaxi Province of Ecuador. Findings from a national health and nutrition
survey, Encuesta Natcional de Salud y Nutriciόn (ENSANUT-ECU) showed 25.3% of children ages
0-5 years nationally, and 42.3% of indigenous children were stunted (HAZ<-2). Other
indicators from this study further demonstrated the nutritional vulnerabilities of
indigenous, young children in Ecuador. Prevalences of anemia (41.6%), zinc deficiency
(32.4%), and vitamin A deficiency (19.3%) all exceeded national averages and were highest in
the 6-11 mo age category. The ENSANUT-ECU found high levels of nutrient intake inadequacies
in the general population for protein, iron, and zinc. Rice comprises nearly one-third
(32.8%) of energy in the diet, with relatively low levels of animal source foods. USFQ
conducted the Chispaz Study examining the efficacy of a micronutrient powder; this study was
carried out in Pastocalle. Thus, there is already an understanding and appreciation for
research in the community.
Risks and Benefits. There are less than minimal risks associated with this study. The risk of
an allergic reaction to eggs is low; expected to be only between 1 to 2% in young children.
The majority of cases are mild consisting of hives soon after consuming eggs. Children with a
known egg allergy will be excluded from the study and any children displaying any symptoms of
an allergic reaction will be referred for treatment and will be excluded from any future
participation in the study. There is a small risk associated with the blood draw for this
study. The study team collecting samples will be well-trained on safe handling of human
biologic specimens and will follow strict hygienic protocols relating to the usage of gloves,
sterile equipment, and alcohol swabs to cleanse the area. A numbing cream is similarly being
explored as an option to minimize pain for the child and psychological distress for the
mother/caregiver. Interviews will be conducted with caregivers in a private setting to
maintain confidentiality.
The children will benefit directly through their participation in this study through the
likely improvements in nutritional status via consumption the consumption of eggs, a high
nutritional quality, nutrient-dense food. The study involves a delayed intervention design,
where eventually all participants will have the opportunity to receive the egg intervention.
Thus, all participants have the opportunity to benefit through the receipt of a high quality
food. There are potential benefits to society in terms of knowledge gained regarding a
food-based, sustainable approach to improving young child nutrition. Globally, nutrition
programming and policies have largely relied on the use of single nutrient or multiple
nutrient supplementation, micronutrient powders, or imported processed complementary foods.
This project aims to test a food-based approach, providing nutrients through eggs in a highly
bioavailable matrix appropriate for young children. Further, the eggs are purchased locally
with likely positive implications for economic development in the region.
Privacy and confidentiality. The consent, interview, anthropometric measurement, and blood
draw processes will occur at community centers, health care posts or children nurseries -as
the field team may determine the most appropriate location in each district of the parish.
The spaces are sufficiently large to ensure confidentiality. Separate, private space in each
of these settings will be designated for interviews and data collection. Only questions
pertaining to child nutrition will be asked to avoid any unnecessary invasion of privacy.
Additional time will be allocated for each consent and interview process to allow
mothers/primary caregivers to ask additional questions as needed. They are also located in
close proximity to households to facilitate convenient access for mothers and caregivers. In
some cases, the field team will visit directly the participants at home for the purpose above
mentioned.
The informed consent form will include mention of the possible use of data and blood samples
for future research, as optional for consent on the part of mothers/caregivers. Two questions
will be included in the baseline survey, the first asking if mothers/caregivers agree that
their data may be used for future research and the second, asking mothers/caregivers if the
blood of the child may be used for future research.
Data will be collected on paper forms and will be stored in locked cabinets in the Instituto
de Investigacion en Salud y Nutricion (ISYN) office. All participants will be assigned a
unique identifier that will be written on all study forms. Access to personal identifying
information will be limited among the research investigators to only the PI, the senior
research team, and the field team the purposes of participant follow-up and tracking. Data
will be entered by the study team at USFQ. Encrypted raw data files will be transferred
periodically to Washington University using password protected storage devices or other
electronic data transfer mechanisms. De-identified datasets will be shared with co-PIs. In
Ecuador, the study will be previously approved by the IRB (Comité de Bioética) of the USFQ
and by the Ministry of Public Health.
