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Incomplete cell disruption of resistant microbes.
Robert Starke1,2, Nico Jehmlich3, Trinidad Alfaro4
1Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington, USA. robert.starke@biomed.cas.cz.
Scientific Reports
|April 6, 2019
Summary
Bead-beating and ultra-sonication yield different microbial biomolecule extraction results. Ultra-sonication offers higher energy, but bead-beating may yield more fungal DNA, suggesting potential underrepresentation in ecological studies.
Area of Science:
- Microbiology
- Molecular Biology
- Biotechnology
Background:
- Biomolecule extraction for OMIC analysis of microbial communities is crucial.
- Common methods like bead-beating and ultra-sonication exhibit variable yields and efficiencies.
- Different microbial cell types (bacteria, fungi) require distinct disruption pressures.
Purpose of the Study:
- To compare the disruption efficiency and biomolecule yields of bead-beating and ultra-sonication.
- To evaluate the impact of these methods on different microbial cell types.
- To identify potential biases in microbial community analysis due to extraction techniques.
Main Methods:
- Comparison of bead-beating and ultra-sonication for biomolecule extraction.
- Energy transfer measurement for both methods over 10 minutes.
- Transmission Electron Microscopy (TEM) imaging of microbial cells post-disruption.
- DNA yield and extraction efficiency assessment for bacteria and fungi.
Main Results:
- Ultra-sonication transferred three times more energy than bead-beating in 10 minutes.
- TEM revealed intact gram-positive bacteria and fungi, but destroyed gram-negative bacteria after ultra-sonication.
- Bead-beating yielded higher fungal DNA but showed a three-fold lower extraction efficiency due to larger fungal genomes.
Conclusions:
- Current biomolecule extraction methods may lead to underrepresentation of certain microbes, especially fungi.
- The choice of extraction technique significantly impacts microbial community profiling.
- Further investigation and refinement of extraction protocols are needed for accurate ecological studies.
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