High-quality DNA extraction is a crucial step in metagenomic studies.Bias by different isolation kits impairs the comparison across datasets.A trending topic is,however,the analysis of multiple metagenomes from the same patients to draw a holistic picture of microbiota associated with diseases.We thus collected bile,stool,saliva,plaque,sputum,and conjunctival swab samples and performed DNA extraction with three commercial kits.For each combination of the specimen type and DNA extraction kit,20-gigabase(Gb)metagenomic data were generated using short-read sequencing.While profiles of the specimen types showed close proximity to each other,we observed notable differences in the alpha diversity and composition of the microbiota depending on the DNA extraction kits.No kit outperformed all selected kits on every specimen.We reached consistently good results using the Qiagen QiAamp DNA Microbiome Kit.Depending on the specimen,our data indicate that over 10 Gb of sequencing data are required to achieve sufficient resolution,but DNA-based identification is superior to identification by mass spectrometry.Finally,long-read nanopore sequencing confirmed the results(correlation coefficient＞0.98).Our results thus suggest using a strategy with only one kit for studies aiming for a direct comparison of multiple microbiotas from the same patients.

Jacqueline Rehner;Georges Pierre Schmartz;Laura Groeger;Jan Dastbaz;Nicole Ludwig;Matthias Hannig;Stefan Rupf;Berthold Seitz;Elias Flockerzi;Tim Berger;Matthias Christian Reichert;Marcin Krawczyk;Eckart Meese;Christian Herr;Robert Bals;S?ren L.Becker;Andreas Keller;Rolf Müller

Institute of Medical Microbiology and Hygiene,Saarland University,D-66421 Homburg,Germany;Clinical Bioinformatics,Saarland University,D-66123 Saarbriicken,Germany;Department of Human Genetics,Saarland University,D-66421 Homburg,Germany;Helmholtz Institute for Pharmaceutical Research Saarland,D-66123 Saarbriicken,Germany;Clinic of Operative Dentistry,Periodontology and Preventive Dentistry,Saarland University,D-66421 Homburg,Germany;Department of Ophthalmology,Saarland University Medical Center,D-66421 Homburg,Germany;Department of Medicine HI,Saarland University Medical Center,D-66421 Homburg,Germany;Department of Internal Medicine V-Pulmonology,Allergology,Intensive Care Medicine,Saarland University,D-66421 Homburg,Germany

基因组蛋白质组与生物信息学报（英文版）

[1]Jacqueline Rehner;Georges Pierre Schmartz;Laura Groeger;Jan Dastbaz;Nicole Ludwig;Matthias Hannig;Stefan Rupf;Berthold Seitz;Elias Flockerzi;Tim Berger;Matthias Christian Reichert;Marcin Krawczyk;Eckart Meese;Christian Herr;Robert Bals;S?ren L.Becker;Andreas Keller;Rolf Müller-.Systematic Cross-biospecimen Evaluation of DNA Extraction Kits for Long-and Short-read Multi-metagenomic Sequencing Studies)[J].基因组蛋白质组与生物信息学报（英文版）,2022(02):405-417

biospecimen,Kits,metagenomes,gigabase,QiAamp,microbiotas

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