Vitamin A, Stool Microbiota and Vaccine Response in Bangladeshi Infants

Newborn Vitamin A Supplementation, Gut Microbiota and Vaccine Response During the Second Year of Life in Bangladeshi Infants

Vitamin A deficiency (VAD) increases the risk of death from infections in infants and young children. The World Health Organization (WHO) recommends high-dose vitamin A supplementation (VAS) from 6-59 months of age to reduce the risk of death in countries where VAD is common. Such countries include Bangladesh, where this study is being conducted. While providing VAS at 6 months is recommended, providing VAS at birth may also decrease the risk of death since newborn infants are also at risk of VAD. VAS presumably reduces infant mortality by improving the immune response to infection and immunization. Vitamin A particularly affects the development and function of T cells, which develop in the thymus and are a key component of the memory response to infection and immunization. Vitamin A is important for development of an important class of T cells, regulatory T-cells, in the intestine. Regulatory T-cells prevent over-reaction of the immune system to substances the immune system might otherwise treat as harmful such as food or the healthy bacteria in the intestine. VAD could disrupt the normal colonization of the infant's intestinal tract and cause a condition called "dysbiosis" where abnormal bacteria flourish and adversely affect the infant's immune system. Dysbiosis may disrupt the immune response to injectable and oral vaccines. VAS at birth may prevent dysbiosis and thus improve immune function, response to vaccines, and child survival. The investigators recently completed an intervention trial in Bangladeshi infants (NCT01583972) examining the effect of VAS at birth on immune function and response to vaccines administered from birth to 14 wk of age. The present study will recruit infants who completed NCT01583972 when they are from 12 to 24 m of age to determine if VAS at birth affects the responses to these same vaccines when they are measured during the second year of life. The investigators will examine the effect of VAS at birth on gut microbiota measured early in infancy and during the second year of life, and explore the association of the gut microbiota with vaccine response. Mothers of study infants will participate in the study because the breast milk oligosaccharide content strongly affects gut microbiota composition and the "secretor status" of the mother, which can be determined from maternal FUT2 genotype, strongly affects breast milk oligosaccharide content.

Study Overview

• Status: Completed
• Conditions: Vitamin A Deficiency

Detailed Description

Hypotheses and Specific Aims The investigators will test the hypotheses that VAS at birth will (1) improve production of new T cells at 1-2 yr of age; (2) improve T-cell memory responses at 1-2 yr of age to vaccines given early in infancy (birth - 14 wk); and (3) alter intestinal colonization early in infancy (6, 11 and 15 wk) and at 1-2 yr of age to increase Bifidobacterium and other healthy bacteria and decrease Proteobacteria and other harmful bacteria. Because the "secretor status" of the mother affects the carbohydrate content of the breastmilk, which can in turn affect Bifidobacterium growth in the infant gut, the "secretor status" of the mother will be determined. Furthermore these differences in composition of the intestinal bacteria will be associated with greater immunologic responses to oral and systemic vaccines.

Specific Aim 1: Determine if VAS or placebo at birth affect the blood concentration and thymic output of naïve T-cells at 1-2 yr of age. Specific Aim 2: Determine if VAS or placebo at birth affect T-cell mediated responses at 1-2 yr of age to vaccines given early in infancy, including serum and intestinal antibody levels, vaccine-specific proliferative responses by T cells and cytokine production by T cells. Specific Aim 3: Determine if VAS or placebo at birth affect the relative abundance of healthy intestinal bacteria and common harmful bacteria in the feces at 6, 11, and 15 wk of age and at 1-2 yr of age and determine if relative abundance of these bacteria correlates with vaccine responses shortly after vaccination (6, 11, 15 wk) and later in infancy (1-2 yr). As part of Aim 3 the "secretor status" genotype (FUT2 gene) of the mother will be determined from a cheek swab DNA sample.

• Study Type: Observational
• Enrollment (Actual): 258

Contacts and Locations

Study Locations

• Bangladesh
  • Dhaka, Bangladesh, 1212
    • International Centre for Diarrhoeal Disease Research, Bangladesh
• United States
  • California
    • Davis, California, United States, 95616
      • University of California, Davis
    • Davis, California, United States, 95616
      • USDA Western Human Nutrition Research Center

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 1 year to 2 years (Child)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

• Sampling Method: Non-Probability Sample

Study Population

The present study will recruit infants who completed NCT01583972 when they are from 12 to 24 months of age. Mothers of these infants will also be recruited for collection of DNA from a cheek swab.

Description

Inclusion Criteria:

• for infant: completion of NCT01583972
• for mother: mother of study infant

Exclusion Criteria:

• none

Study Plan

How is the study designed?

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Naïve T-cells in peripheral blood	The concentration of naïve T-cells in peripheral blood will be measured once at 52 - 104 wk of age by flow cytometric analysis.	measured once at 52 - 104 weeks of age

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Thymic output of naïve T cells	The thymic output of naïve T cells will be measured once at 52 - 104 wk of age by T-cell receptor excision circle (TREC) levels in DNA purified from peripheral blood mononuclear cells (PBMC).	measured once at 52 - 104 weeks of age
Immune response to vaccines	The immune responses to vaccines, measured at 52 - 104 wk of age, include: (A) Serum immunoglobulin G (IgG) and immunoglobulin A (IgA) responses to oral polio virus vaccine (OPV; given at birth, 6 wk, 10 wk, 14 wk); (B) serum IgG response to tetanus toxoid vaccine (TT; given at 6, 10 and 14 wk) and (C) Hepatitis B virus vaccine (HBV; given at 6, 10 and 14 wk); (D) the IgA response to OPV in stool; the (E) proliferative and (F) cytokine response of peripheral blood cluster of differentiation 4 (CD4) T-cells to OPV, TT, HBV vaccines and to the tuberculosis vaccine (BCG; given at birth). Antibody responses will be measured by ELISA, T-cell proliferation by flow cytometric analysis after 6 days of culture with corresponding vaccine antigens, and cytokines will be measured in supernatant after 3 and 6 d of culture with the same antigens.	Measured once at 52 - 104 weeks of age
Change in the relative abundance of stool bacteria	The relative abundance of stool bacteria will be measured using DNA extracts at four time points (6, 10 and 14 wk, and at one point between 52 and 104 wk) using culture-independent, next generation sequencing (NGS) of bacterial populations, followed by quantitative PCR (QPCR) and Bifidobacterium-specific terminal restriction-fragment-length polymorphism (TRFLP.)	6, 10, 14 and 52-104 weeks of age
Serum Vitamin A status	Vitamin A status will be assessed by serum retinol or retinol binding protein (RBP4) at 52 - 104 wk of age.	measured once at 52-104 weeks of age
Secretor status of the study infant's mother.	The "secretor status" of the mother is determined by the FUT2 genotype. FUT2 is related to FUT1, the gene encoding fucosyltransferase 1, which determines the composition of specific glycans on erythrocytes that determine the ABO (or ABH) and Lewis blood group antigens. The activity of fucosyltransferase 2 (encoded by FUT2) causes the synthesis and secretion of these same glycans into breast milk and other secretions (e.g., saliva and other intestinal secretions).	measured once when infant is 52 - 104 weeks of age

Collaborators and Investigators

• Sponsor: USDA, Western Human Nutrition Research Center
• Collaborators: University of California, Davis; International Centre for Diarrhoeal Disease Research, Bangladesh; Thrasher Research Fund
• Investigators: Principal Investigator: Charles B Stephensen, PhD, USDA, Western Human Nutrition Research Center

Publications and helpful links

General Publications

• Ahmad SM, Huda MN, Raqib R, Qadri F, Alam MJ, Afsar MNA, Peerson JM, Tanumihardjo SA, Stephensen CB. High-Dose Neonatal Vitamin A Supplementation to Bangladeshi Infants Increases the Percentage of CCR9-Positive Treg Cells in Infants with Lower Birthweight in Early Infancy, and Decreases Plasma sCD14 Concentration and the Prevalence of Vitamin A Deficiency at Two Years of Age. J Nutr. 2020 Nov 19;150(11):3005-3012. doi: 10.1093/jn/nxaa260.
• Ahmad SM, Raqib R, Huda MN, Alam MJ, Monirujjaman M, Akhter T, Wagatsuma Y, Qadri F, Zerofsky MS, Stephensen CB. High-Dose Neonatal Vitamin A Supplementation Transiently Decreases Thymic Function in Early Infancy. J Nutr. 2020 Jan 1;150(1):176-183. doi: 10.1093/jn/nxz193.
• Huda MN, Ahmad SM, Alam MJ, Khanam A, Kalanetra KM, Taft DH, Raqib R, Underwood MA, Mills DA, Stephensen CB. Bifidobacterium Abundance in Early Infancy and Vaccine Response at 2 Years of Age. Pediatrics. 2019 Feb;143(2):e20181489. doi: 10.1542/peds.2018-1489.
• Huda MN, Ahmad SM, Kalanetra KM, Taft DH, Alam MJ, Khanam A, Raqib R, Underwood MA, Mills DA, Stephensen CB. Neonatal Vitamin A Supplementation and Vitamin A Status Are Associated with Gut Microbiome Composition in Bangladeshi Infants in Early Infancy and at 2 Years of Age. J Nutr. 2019 Jun 1;149(6):1075-1088. doi: 10.1093/jn/nxz034.

Helpful Links

• International Centre for Diarrhoeal Disease Research, Bangladesh
• United States Department of Agriculture, Western Human Nutrition Research Center

Study record dates

Study Major Dates

• Study Start: October 1, 2014
• Primary Completion (Actual): September 1, 2015
• Study Completion (Actual): September 1, 2015

Study Registration Dates

• First Submitted: December 23, 2013
• First Submitted That Met QC Criteria: January 2, 2014
• First Posted (Estimate): January 6, 2014

Study Record Updates

• Last Update Posted (Estimate): November 18, 2015
• Last Update Submitted That Met QC Criteria: November 17, 2015
• Last Verified: November 1, 2015

More Information

Terms related to this study

• Keywords: vaccine; Infant; immunity; Bangladesh; microbiota; vitamin A
• Additional Relevant MeSH Terms: Eye Diseases; Nutrition Disorders; Avitaminosis; Deficiency Diseases; Malnutrition; Vision Disorders; Night Blindness; Vitamin A Deficiency
• Other Study ID Numbers: PR-13068