TY  - JOUR
U1  - Zeitschriftenartikel, wissenschaftlich - begutachtet (reviewed)
A1  - Neidhöfer, Claudio
A1  - Bagniceva, Maria
A1  - Wetzig, Nina
A1  - Sieber, Martin A.
A1  - Thiele, Ralf
A1  - Parčina, Marijo
T1  - Pragmatic Considerations When Extracting DNA for Metagenomics Analyses of Clinical Samples
JF  - International Journal of Molecular Sciences
N2  - Microbiome analyses are essential for understanding microorganism composition and diversity, but interpretation is often challenging due to biological and technical variables. DNA extraction is a critical step that can significantly bias results, particularly in samples containing a high abundance of challenging-to-lyse microorganisms. Taking into consideration the distinctive microenvironments observed in different bodily locations, our study sought to assess the extent of bias introduced by suboptimal bead-beating during DNA extraction across diverse clinical sample types. The question was whether complex targeted extraction methods are always necessary for reliable taxonomic abundance estimation through amplicon sequencing or if simpler alternatives are effective for some sample types. Hence, for four different clinical sample types (stool, cervical swab, skin swab, and hospital surface swab samples), we compared the results achieved from extracting targeted manual protocols routinely used in our research lab for each sample type with automated protocols specifically not designed for that purpose. Unsurprisingly, we found that for the stool samples, manual extraction protocols with vigorous bead-beating were necessary in order to avoid erroneous taxa proportions on all investigated taxonomic levels and, in particular, false under- or overrepresentation of important genera such as Blautia, Faecalibacterium, and Parabacteroides. However, interestingly, we found that the skin and cervical swab samples had similar results with all tested protocols. Our results suggest that the level of practical automation largely depends on the expected microenvironment, with skin and cervical swabs being much easier to process than stool samples. Prudent consideration is necessary when extending the conclusions of this study to applications beyond rough estimations of taxonomic abundance.
KW  - human microbiome
KW  - DNA extraction protocols
KW  - automation of sample processing
KW  - microbiome analyses
KW  - microbial community structure
KW  - extraction-linked bias
UN  - https://nbn-resolving.org/urn:nbn:de:hbz:1044-opus-73988
SN  - 1422-0067
SS  - 1422-0067
U6  - https://doi.org/10.3390/ijms241411262
DO  - https://doi.org/10.3390/ijms241411262
PM  - 37511022
VL  - 24
IS  - 14
SP  - 10
S1  - 10
PB  - MDPI
ER  -