Assessment of absolute abundance in mother-infant gut microbiome using marine-sourced bacterial DNA spike-in and comparison with conventional quantification methods

Aim: To evaluate the effectiveness of marine-sourced bacterial DNA spike-in quantification for determining absolute microbial abundance in the gut microbiome of mother-infant pairs and to compare this method with conventional quantification techniques. Methods: We conducted a pilot study involving six mother-infant pairs, applying a DNA spike-in quantification method using bacterial DNA from Pseudoalteromonas sp. APC 3896 and Planococcus sp. APC 3900, isolated from deep-sea fish. We compared our approach with established absolute quantification methods-flow cytometry, total DNA measurement, quantitative PCR (qPCR), and culture-based plate count-to evaluate microbial load and taxonomic composition across mother-infant samples. Results: Our spike-in method accurately estimated microbial loads, producing results consistent with qPCR and total DNA quantification. We observed that mothers exhibited higher total bacteria loads than infants by approximately half a log, while the abundance of Bifidobacterium was comparable in both groups. The spike-in method revealed significant differences in taxonomic composition, highlighting the impact of absolute quantification on microbiome analysis outcomes. Importantly, the method did not alter alpha diversity measures but slightly affected beta diversity analysis, reflecting more precise inter-group differences. Conclusion: Marine-sourced bacterial DNA spike-in offers a reliable, scalable, and accurate approach for absolute microbiome quantification. This method enhances microbiome analysis by addressing biases inherent in relative abundance measures, providing a deeper understanding of microbial dynamics in mother-infant gut microbiomes.