(P22-20-26) Effect of Multiple Micronutrient-Fortified Bouillon Cubes on Fecal Resistomes of Children in Northern Ghana: A Metagenomic Study Nested in a Randomized Controlled Trial

Objectives: Iron-containing fortified foods/supplements are useful public health tools for reducing iron deficiency and anemia. However, excess luminal iron from such interventions may compromise the gut microbiome and influence antimicrobial resistance gene (ARG) carriage, with implications for gastrointestinal morbidity. We assessed the effect of multiple micronutrient (MN)-fortified bouillon cubes on child fecal resistomes (ARGs in the gut).

Methods: In a 38-wk double-blind randomized controlled trial, households of enrolled children aged 2–5 y (N=690) received biweekly bouillon cube rations fortified with either:1) multiple MN––vitamins A: 200 µg retinol/g, B12:1.2 µg/g, B9: 80 µg folic acid/g; zinc: 3 mg/g, iodine: 30 µg/g, and iron: 4 mg/g or 2) iodine only (control). Stool samples collected at baseline and wk 35 from a randomly selected subgroup underwent shotgun metagenomic sequencing, and resistomes were profiled via the Comprehensive Antibiotic Resistance Database. PERMANOVA assessed compositional differences. Difference in differences (DiD) mixed models assessed intervention effects on ARG richness (number of unique ARGs) and normalized ARG abundance (copies per million), including specifications by individual ARGs, antibiotic drug class, and resistance mechanism.

Results: The parent trial reported child bouillon intakes of ~1.5-1.9 g/day and low biweekly reported antibiotic use and diarrhea. In the analytical sample (n =136; mean age = 40 mos), the intervention did not affect resistome composition (PERMANOVA R² = 0.004, p = 0.29) or normalized ARG abundance over time (geometric mean ratio DiD: 0.99; 95% CI: 0.91–1.08; p=0.78). The control group had a 9% reduction in ARG richness relative to the intervention over time (incidence rate ratio DiD: 0.91 95% CI: 0.80–0.92; p < 0.01). Tetracycline resistance genes dominated fecal resistomes, but no intervention effects emerged within ARG drug classes or resistance mechanisms (q > 0.05). Sixteen ARGs showed nominal changes over time (p  < 0.05), but none remained significant after Benjamini-Hochberg corrections for multiple comparisons (q > 0.05).

Conclusions: Multiple MN-fortified bouillon cubes did not alter gut resistomes of children in this context, suggesting minimal risk of ARG-mediated gastrointestinal or infectious morbidity.

Funding Sources: Gates Foundation, via Helen Keller Intl