The Effect of Oral Iron Supplementation on Gut Microbial Composition: a Secondary Analysis of a Double-Blind, Randomized Controlled Trial among Cambodian Women of Reproductive Age

Study information

Needs review

study design

randomized controlled trial

Citation

PMID PubMed identifier for scientific articles.

37199608

DOI Digital object identifier for electronic documents.

https://doi.org/10.1128/spectrum.05273-22

URI

Authors

Finlayson-Trick E, Nearing J, Fischer JA, Ma Y, Wang S, Krouen H, Goldfarb DM, Karakochuk CD

Journal

Microbiology spectrum

Year

2023

Keywords:

enteropathogen, gut microbiome, iron supplementation

The World Health Organization recommends untargeted iron supplementation for women of reproductive age (WRA) in countries where anemia prevalence is greater than 40%, such as Cambodia. Iron supplements, however, often have poor bioavailability, so the majority remains unabsorbed in the colon. The gut houses many iron-dependent bacterial enteropathogens; thus, providing iron to individuals may be more harmful than helpful. We examined the effects of two oral iron supplements with differing bioavailability on the gut microbiomes in Cambodian WRA. This study is a secondary analysis of a double-blind, randomized controlled trial of oral iron supplementation in Cambodian WRA. For 12 weeks, participants received ferrous sulfate, ferrous bisglycinate, or placebo. Participants provided stool samples at baseline and 12 weeks. A subset of stool samples (n = 172), representing the three groups, were randomly selected for gut microbial analysis by 16S rRNA gene sequencing and targeted real-time PCR (qPCR). At baseline, 1% of women had iron-deficiency anemia. The most abundant gut phyla were Bacteroidota (45.7%) and Firmicutes (42.1%). Iron supplementation did not alter gut microbial diversity. Ferrous bisglycinate increased the relative abundance of Enterobacteriaceae, and there was a trend towards an increase in the relative abundance of Escherichia-Shigella. qPCR detected an increase in the enteropathogenic Escherichia coli (EPEC) virulence gene, bfpA, in the group that received ferrous sulfate. Thus, iron supplementation did not affect overall gut bacterial diversity in predominantly iron-replete Cambodian WRA, however, evidence does suggest an increase in relative abundance within the broad family Enterobacteriaceae associated with ferrous bisglycinate use. IMPORTANCE To the best of our knowledge, this is the first published study to characterize the effects of oral iron supplementation on the gut microbiomes of Cambodian WRA. Our study found that iron supplementation with ferrous bisglycinate increases the relative abundance of Enterobacteriaceae, which is a family of bacteria that includes many Gram-negative enteric pathogens like Salmonella, Shigella, and Escherichia coli. Using qPCR for additional analysis, we were able to detect genes associated with enteropathogenic E. coli, a type of diarrheagenic E. coli known to be present around the world, including water systems in Cambodia. The current WHO guidelines recommend blanket (untargeted) iron supplementation for Cambodian WRA despite a lack of studies in this population examining iron's effect on the gut microbiome. This study can facilitate future research that may inform evidence-based global practice and policy.

Experiment 1

Edit History Delete

Mark reviewed

Your review change was submittedDuplicate this Experiment Add a new Signature

Needs review

Curated date: 2024/03/23

Curator: Manisha28

Revision editor(s): Manisha28

Subjects

Location of subjects: Cambodia

Host species Species from which microbiome was sampled. Contact us to have more species added.: Homo sapiens

Body site Anatomical site where microbial samples were extracted from according to the Uber Anatomy Ontology: Feces Cow dung,Cow pat,Droppings,Dung,Excrement,Excreta,Faeces,Fecal material,Fecal matter,Fewmet,Frass,Guano,Matières fécales@fr,Merde@fr,Ordure,Partie de la merde@fr,Piece of shit,Porción de mierda@es,Portion of dung,Portion of excrement,Portion of faeces,Portion of fecal material,Portion of fecal matter,Portion of feces,Portion of guano,Portion of scat,Portionem cacas,Scat,Spoor,Spraint,Stool,Teil der fäkalien@de,Feces,feces

Condition The experimental condition / phenotype studied according to the Experimental Factor Ontology: Environmental exposure measurement Environmental exposure measurement,environmental exposure measurement

Group 0 name Corresponds to the control (unexposed) group for case-control studies: Placebo group

Group 1 name Corresponds to the case (exposed) group for case-control studies: Ferrous bisglycinate group

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group: Healthy, non pregnant women of reproductive age who received oral supplementation of ferrous bisglycinate capsule containing 18 mg of elemental iron.

Group 0 sample size Number of subjects in the control (unexposed) group: 47

Group 1 sample size Number of subjects in the case (exposed) group: 46

Antibiotics exclusion Number of days without antibiotics usage (if applicable) and other antibiotics-related criteria used to exclude participants (if any): Women who were taking antibiotics in the previous 12 weeks were excluded.

Lab analysis

Sequencing type: 16S

16S variable region One or more hypervariable region(s) of the bacterial 16S gene: V6-V8

Sequencing platform Manufacturer and experimental platform used for quantifying microbial abundance: Illumina

Statistical Analysis

Data transformation Data transformation applied to microbial abundance measurements prior to differential abundance testing (if any).: relative abundances

Statistical test: Beta Binomial Regression

Significance threshold p-value or FDR threshold used for differential abundance testing (if any): 0.05

MHT correction Have statistical tests be corrected for multiple hypothesis testing (MHT)?: Yes

Confounders controlled for Confounding factors that have been accounted for by stratification or model adjustment: Confounders controlled for: "inflammation" is not in the list (abnormal glucose tolerance, acetaldehyde, acute graft vs. host disease, acute lymphoblastic leukemia, acute myeloid leukemia, adenoma, age, AIDS, alcohol consumption measurement, alcohol drinking, ...) of allowed values.inflammation

Alpha Diversity

Shannon Estimator of species richness and species evenness: more weight on species richness: unchanged

Richness Number of species: unchanged

Faith Phylogenetic diversity, takes into account phylogenetic distance of all taxa identified in a sample: unchanged

Signature 1

Edit History Delete

Mark reviewed

Your review change was submitted

Needs review

Curated date: 2024/03/29

Curator: Manisha28

Revision editor(s): Manisha28

Source: Figure 4, Figure 5

Description: Differentially abundant bacterial Taxa in the Ferrous bisglycinate group compared to the Placebo group.

Abundance in Group 1: increased abundance in Ferrous bisglycinate group

NCBI	Quality Control	Links
Enterobacteriaceae
Escherichia/Shigella sp.
Weissella

Revision editor(s): Manisha28

Signature 2

Edit History Delete

Mark reviewed

Your review change was submitted

Needs review

Curated date: 2024/03/29

Curator: Manisha28

Revision editor(s): Manisha28

Source: Figure 4

Description: Differentially abundant bacterial Taxa in the Ferrous bisglycinate group compared to the Placebo group.

Abundance in Group 1: decreased abundance in Ferrous bisglycinate group

NCBI	Quality Control	Links
Enterococcus

Revision editor(s): Manisha28

Experiment 2

Edit History Delete

Mark reviewed

Your review change was submittedDuplicate this Experiment Add a new Signature

Needs review

Curated date: 2024/03/29

Curator: Manisha28

Revision editor(s): Manisha28

Differences from previous experiment shown

Subjects

Group 0 name Corresponds to the control (unexposed) group for case-control studies: Placebo Group

Group 1 name Corresponds to the case (exposed) group for case-control studies: Ferrous sulfate group

Group 1 definition Diagnostic criteria applied to define the specific condition / phenotype represented in the case (exposed) group: Healthy, non pregnant women of reproductive age who received oral supplementation of ferrous sulfate capsule containing 60 mg of elemental iron.