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Linehan et al. Microbiome Res Rep. 2025;4:24 https://dx.doi.org/10.20517/mrr.2024.92 Page 15 of 20

particularly in cases where unusual or rare genera are observed, which could suggest a potential link to
[106]
maternal sources . The methodology may also artificially inflate bacterial species abundance due to early
PCR kinetics. Future studies could benefit from targeted qPCR, amplification-free metagenomics, FISH,
SEM, microbial culturing, and control DNA spiking to enhance absolute abundance calculation. The use of
PICRUSt2 to predict metabolic function also has limitations. Its predictions are biased toward established
reference genomes, reducing the identification of rare, environment-specific functions, although the
expansion of genome databases is mitigating this bias. Moreover, the amplicon-based nature of PICRUSt2
limits its ability to discern strain-specific functionality; shotgun metagenomics would overcome this,
allowing for accurate gene abundance measurement and strain-level differentiation. Finally, there is a need
to investigate how multiple perinatal factors interact in shaping the infant microbiome. For example,
[107]
breastfeeding can modify colonization patterns in C-section-born infants . Understanding transmission
routes and the influence of perinatal factors on the mother-infant microbial bond is crucial for developing
therapeutic interventions and next-generation probiotics to support infant health. Finally, it is important to

clarify that while the CS rate in this subset (n = 18) was 44%, the rate across the full recruited cohort (n = 63)
was 36.6%, which aligns closely with the national average in Ireland and in the recruitment hospital during
the study period (~35%) and with more recent figures (~39%) .
[108]
[109]
In conclusion, this study highlights the significant role of maternal sources, particularly the oral and vaginal
microbiomes, in microbial sharing with infants. We observed that, on average, 45 ASVs were shared
between mother-infant oral samples, accounting for 65% of reads in infant oral microbiota. Approximately
15 vaginal bacterial taxa (15% of infant reads) were shared, with significantly higher transmission following
vaginal delivery (Mann–Whitney P = 0.045). Our findings confirm the presence of a distinct meconium
microbiome - comprising taxa such as S. epidermidis, Bifidobacterium, and Streptococcus and support the
absence of a measurable placental microbiome, consistent with negative control profiles.

Perinatal factors, including delivery mode, maternal antibiotic use, and feeding type, influenced microbial
diversity and functional pathways. For instance, maternal antibiotic exposure was associated with increased
abundance of Gemella and Fusobacterium and enrichment of heme biosynthesis pathways in meconium,
while breastfeeding promoted pathways involved in vitamin K2 and B6 biosynthesis. Additionally, feeding
type influenced microbial composition, with Bacteroides enriched in breastfed infants and Blautia and
Dorea in formula-fed infants. These results underscore the complex and dynamic interactions shaping early
microbial colonization, highlighting critical maternal contributions and the modulating role of perinatal
exposures. Future longitudinal studies incorporating strain-resolved metagenomics are necessary to track
microbial development and transmission patterns over time, with the ultimate goal of informing next-
generation probiotic and therapeutic interventions to support infant health.

DECLARATIONS
Authors’ contributions
Performed DNA extractions, prepared 16S rRNA compositional sequencing libraries, and conducted
bioinformatics and statistical analyses: Linehan K
Recruited study participants, assisted with sample collection, and provided clinical metadata: Hurley E,
O’Shea CA
Performed part of the bioinformatics analysis of 16S rRNA sequencing data: Healy K
Designed the study and obtained ethical approval: Dempsey EM, Ryan CA, Ross RP, Stanton C
Drafted the manuscript: Linehan K