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Linehan et al. Microbiome Res Rep. 2025;4:24 https://dx.doi.org/10.20517/mrr.2024.92 Page 13 of 20

detection and minimize contamination, we included negative DNA extraction and sequencing controls and
followed stringent protocols recommended for low biomass samples [43,44] . In our cohort, the core meconium
microbiome consisted of genera such as Staphylococcus, Bifidobacterium, Streptococcus, Enterococcus,
N
c
a
Escherichia-Shigella, Delftia, Afipia, Cutibacterium, n d Rothia. o t a b l y , S . epidermidis, o m m o n i n
colostrum and breast milk, was present in all meconium samples and has been previously identified in
breastfed neonates’ meconium [45-47] . These findings align with studies suggesting that neonatal meconium
reflects microbial communities acquired during and post-birth [3,48-50] . Functional analysis using PICRUSt2
revealed that the pentose phosphate pathway was the most abundant metabolic pathway in meconium. This
pathway is crucial for generating NADPH, which is needed for biosynthetic reactions, and ribose-5-
phosphate for nucleotide synthesis. Other prevalent pathways included L-isoleucine biosynthesis, branched-
chain, aromatic amino acid biosynthesis, and glycolysis, indicating roles in energy production and
biosynthesis of essential molecules. Regarding placental microbiota, we did not detect any non-contaminant
ASVs, aligning with other studies that report the absence of a placental microbiome [51-53] . Exploratory

analysis without decontamination steps showed that ASVs in placental samples matched those in negative
controls, with phyla (Actinobacteriota, Firmicutes, Proteobacteria), families (Bacillaceae,
Corynebacteriaceae, Micrococcaceae, Streptococcaceae, Xanthobacteraceae), and genera (Afipia, Bacillus,
Corynebacterium, Enhydrobacter, Micrococcus, Streptococcus) similar in presence and relative abundance.
These taxa are known contaminants from the laboratory [54,55] . Thus, our findings support the consensus that
microbial colonization typically occurs at birth and that replicating microbes are absent in healthy
[42]
pregnancies without clinical infections .
The second aim of our study was to investigate the contribution of maternal microbial sources (vagina, oral
cavity, and placenta) to their infants’ oral and gut microbiomes. Focusing first on the infant oral
microbiome, its composition was consistent with previous studies, dominated by Streptococcus, Rothia,
Prevotella, Neisseria, Escherichia-Shigella, Gemella, and Haemophilus . Early colonizers like S. salivarius,
[56]
S. oralis, R. mucilaginosa, S. epidermidis, and F. nucleatum were abundant [57-59] . Maternal oral samples,
predominantly containing Prevotella, Streptococcus, Veillonella, Rothia, Neisseria, and Haemophilus, were
similar to findings in other cohorts [60-62] . Species like R. mucilaginosa, H. parainfluenzae, P. melaninogenica,
and F. nucleatum were prevalent [63-65] . The vaginal microbiome demonstrated a unique composition
compared to meconium and oral microbiomes, with significantly lower diversity . Dominant genera
[18]
i
a
i n c l u d e d Peptoniphilus, Lactobacillus, Finegoldia, Corynebacterium, n d Anaerococcus, n d i c a t i n g
community state type 4-A and 4-B with lower lactic acid bacteria and higher anaerobic bacteria [62,66] . Species
such as F. magna and P. faecalis, both associated with bacterial vaginosis, were found in most samples [67,68] .
The metabolic pathways identified were consistent with those reported across vaginal samples . Vertical
[69]
transmission of microbiota is primarily influenced by the maternal gut, but our study focused on the
maternal oral and vaginal contributions to the infant’s oral and gut microbiomes . Overall, infant and
[70]
maternal oral microbiomes were similar, except for Escherichia-Shigella, Cutibacterium, and
Corynebacterium detected in infants. On average, 45 ASVs were shared between mother-infant pairs,
accounting for 65% of reads in infant samples. While perinatal factors did not significantly affect sharing,
Kageyama et al. found greater acquisition of maternal oral bacteria in formula-fed infants . Shared taxa
[71]
included Streptococcus, Veillonella, Neisseria, Haemophilus, and Fusobacterium, consistent with previous
studies . S. oralis was found in all oral samples and is known as a primary colonizer in both infants and
[63]
adults [72,73] . Other highly shared species like H. parainfluenzae and R. mucilaginosa are also commensal
microbiota, while F. nucleatum, associated with periodontal disease, was present in most dyads [74,75] . These
findings align with reports that around 70% of the neonatal oral microbiota is maternally derived [76,77] .
Regarding vaginal-to-oral transmission, approximately 15 bacterial taxa, accounting for 15% of reads in
infant samples, were shared. NB resulted in significantly higher sharing between maternal vaginal and infant
oral microbiomes compared to CS (Mann Whitney, P = 0.045), similar to other studies [19,78] .