Enzyme treatment of skin samples improves microbiome analysis

Date:
August 10, 2021
Source:
Technical University of Munich (TUM)
Summary:
Healthy skin has a bacterial shield to protect against germs: the
microbiome. This complex assembly of microorganisms was previously
believed to be difficult to decipher. A team of researchers has
now succeeded in using the enzyme benzonase to identify the living
bacteria in skin swabs through sequencing. Their method opens up
new possibilities for diagnosis and treatment in dermatology.



FULL STORY ==========================================================================
In many skin diseases such as atopic dermatitis and acne, the bacterial
layer protecting the skin is damaged. "Our goal is to learn the role
played in such illnesses by the various kinds of skin bacteria," says
Dr. Martin Ko"berle, head of the Dermatoinfectology Laboratory at the
Klinikum Rechts der Isar of the Technical University of Munich (TUM).


==========================================================================
Past efforts by dermatologists to investigate the detailed composition
of the microbiome have hit roadblocks. The reason: In conventional
cultures grown on agar plates, not all bacteria thrive and multiply
equally well. As a result, some slow-growing species can be overlooked entirely. The disadvantage of more recent genetic analytical methods is
that large quantities of DNA sequences from skin cells and fragments
of dead bacteria are captured. This reduces the information value of
the results.

Separating the genetic wheat from the chaff Dr. Ko"berle and the biologist
Dr. Yacine Amar, both part of Prof. Biedermann's team at the Clinic
and Polyclinic for Dermatology and Allergology at TUM, have developed
a method for removing the DNA non-target species in cooperation with an international, interdisciplinary team. They used a special characteristic
of the enzyme benzonase. It destroys the nucleotide chains that carry hereditary information in all living things by breaking them down into
short fragments.

Only live bacteria whose DNA is protected by an outer cell wall escape destruction by the enzyme.

Benzonase has been used for some time, for example to purify proteins:
The enzymes break up all foreign DNA and RNA fragments. These can then
be removed in a centrifuge, leaving the proteins behind. The selection
of skin bacteria functions according to the same principle: Genetic
material from skin cells or dead bacteria is broken up by the enzyme
and can then be separated from the sample. The remaining bacteria can
be destroyed mechanically, permitting the study of their DNA.

"Our experiments showed that, with this method, we can indeed fully
eliminate the non-target DNA and select the skin microbiome," says project leader Yacine Amar. In the lab he initially studied artificial samples containing a mixture of human cells and dead and living bacteria created
using a strict protocol and pre-treated with benzonase. "The process then
used -- known as 16S sequencing - - yielded a highly precise picture
of the composition of the intact bacteria," says the researcher. The
analysis of real skin swabs was just as successful: No residual DNA from
dead bacteria was found in the samples.

Dr. Ko"berle is confident that this approach will also play a key role in future research: "The enzyme-based selection of living skin bacteria can
help us to find microbial biomarkers for certain dermatological illnesses
and also to identify the bacteria that have a positive influence on
the course of the disease. Perhaps they will be used in treatments one
day." The new method for microbiome analysis is already being used in
many cohort studies on skin diseases at the TUM Clinic and Polyclinic
for Dermatology.

========================================================================== Story Source: Materials provided by
Technical_University_of_Munich_(TUM). Note: Content may be edited for
style and length.


========================================================================== Journal Reference:
1. Yacine Amar, Ilias Lagkouvardos, Rafaela L. Silva, Oluwaseun Ayodeji
Ishola, Ba"rbel U. Foesel, Susanne Kublik, Anne Scho"ler, Sebastian
Niedermeier, Rachela Bleuel, Alexander Zink, Klaus Neuhaus,
Michael Schloter, Tilo Biedermann, Martin Ko"berle. Pre-digest of
unprotected DNA by Benzonase improves the representation of living
skin bacteria and efficiently depletes host DNA. Microbiome, 2021;
9 (1) DOI: 10.1186/ s40168-021-01067-0 ==========================================================================

Link to news story: https://www.sciencedaily.com/releases/2021/08/210810130803.htm

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