Towards Quantitative Microbiome Community Profiling Using Internal Standards.

Area of Science:

• Microbiology
• Molecular Biology
• Oceanography

Background:

• High-throughput rRNA gene sequencing provides only relative microbial abundances, hindering accurate interpretation of ecological relationships.
• Spurious correlations in microbiome data complicate the assessment of links to biogeochemical rates.
• Existing methods lack quantitative precision for marine microbial community profiling.

Purpose of the Study:

• To develop and validate a quantitative method for estimating microbial abundances in marine environments.
• To address the inherent limitations of relative abundance data in microbiome surveys.
• To improve the accuracy of microbial community profiling for ecological studies.

Main Methods:

• Spiking known amounts of internal DNA standards into seawater samples before high-throughput sequencing.
• Quantitative estimation of microbial abundances per unit volume of filtered seawater.
• Comparison of results with traditional CHEMTAX analysis and flow cytometry counts.

Main Results:

• The internal standard method yielded significantly different community profiles and co-occurrence patterns compared to relative abundance data.
• Quantitative estimates aligned with independent measures like pigment analysis and bacterial cell counts.
• Chloroplast 16S rRNA gene data proved useful for quantifying specific eukaryotic phototrophs.

Conclusions:

• The internal standard method provides a quantitative approach to rRNA gene microbial community profiling.
• This method enhances the ability to link marine microbial communities to quantitative processes.
• Accurate microbial abundance data are crucial for advancing our understanding of marine ecosystems.