Coming Into Resistance: Clinically Important Antibiotic Resistance Genes in NICU vs. Healthy Infants

Antibiotic-resistant infections in neonates often arise from the infant's own colonizing bacteria harboring antibiotic resistance genes (ARG). This study evaluates ARG that impact antibiotic selection and drive resistance-related mortality, termed clinically important antibiotic resistance genes (CI-ARG).

To determine differences in gut CI-ARG abundance in children from birth through 3 years old, stratified by neonatal intensive care unit (NICU) admission and antibiotic (abx) exposure in the first two months.

Cohort study (sequencing, analysis completed 2025-2026) using shotgun metagenomic sequencing of stool samples from infants in two studies: "Neonatal Intestinal Microbiome" (+NICU groups, 2015-2022), "The First 1000 Days of Life and Beyond" (-NICU groups, 2018-2021). Infants grouped into four cohorts: +NICU/+abx, +NICU/-abx, -NICU/-abx, and -NICU/+abx. Prespecified CI-ARG abundance compared longitudinally among groups.

NICU, well-baby nursery, Inova Fairfax Hospital, Virginia.

+NICU criteria: admission to the NICU, anticipated length of stay (LOS) of at least five days, maternal age ≥18 years. -NICU: parents with good general health and age ≥18 years enrolled prior to infant birth. Stool samples: adequate stool processing volume, at least one stool sample from before one month and after one year of age, 7-day minimum between serial samples.

NICU versus well-baby nursery; antibiotics versus none in first two months of corrected age.

Predefined primary outcome-longitudinal CI-ARG abundance by group, compared using linear mixed effects models with adjustment for multiple comparisons (Benjamini-Hochberg procedure). Predefined secondary outcomes-differences in overall microbiota and resistome composition.

1000 serial stool samples, 179 infants (birth-39 months). Longitudinal abundance significantly differed among cohorts for 11 CI-ARG (p<0.05); often higher in the +NICU groups in early life, with or without antibiotic exposure. Abundance of ARG conferring resistance to aminoglycosides, macrolides, and fluoroquinolones later decreased below -NICU levels. mecA , which often drives methicillin resistance in Staphylococcus , was detected only in +NICU infants during premature period.

This longitudinal study demonstrates significant longitudinal CI-ARG differences in children grouped by early-life NICU status and/or antibiotic exposure. Future studies should explore the mechanisms driving these CI-ARG changes and evaluate their impact on infection risk and community spread.

Question: How do clinically important antibiotic resistance genes (CI-ARG) differ longitudinally among infants up to 3 years old based on NICU admission and antibiotic exposure in the first two months of life?Findings: In this cohort study of 179 infants through 39 months of age, longitudinal abundance was significantly different for 11 prespecified CI-ARG, often higher in the NICU groups in early life, with or without antibiotic exposure.Meaning: Children with a history of NICU hospitalization, whether due to the NICU environment itself, prematurity, or other factors, may have increased CI-ARG burden early in life, irrespective of direct antibiotic exposure.