characterizing-microbiome-dynamics-flow-cytometry-based-workflows

Characterizing Microbiome Dynamics &#8211; Flow Cytometry Based Workflows from Pure Cultures to Natural Communities

The investigation of pure cultures and monitoring of microbial community dynamics is vital to understand and control natural ecosystems and technical ...

Alkaline pretreatment has the potential to enhance the anaerobic digestion of lignocellulosic biomass to biogas. However, the elevated pH of the substrate may require alkalitolerant microbial communities for an effective digestion. Three mixed anaerobic lignocellulolytic cultures were enriched from sediments from two soda lakes with wheat straw as substrate under alkaline (pH 9) mesophilic (37°C) and thermophilic (55°C) conditions. The gas production of the three cultures ceased after 4 to 5 weeks, and the produced gas was composed of carbon dioxide and methane. The main liquid intermediates were acetate and propionate. The physiological behavior of the cultures was stable even after several transfers. The enrichment process was also followed by molecular fingerprinting (terminal restriction fragment length polymorphism) of the bacterial 16S rRNA gene and of the mcrA/mrtA functional gene for methanogens. The main shift in the microbial community composition occurred between the sediment samples and the first enrichment, whereas the structure was stable in the following transfers. The bacterial communities mainly consisted of Sphingobacteriales, Clostridiales and Spirochaeta, but differed at genus level. Methanothermobacter and Methanosarcina genera and the order Methanomicrobiales were predominant methanogenes in the obtained cultures. Additionally, single cellulolytic microorganisms were isolated from enrichment cultures and identified as members of the alkaliphilic or alkalitolerant genera. The results show that anaerobic alkaline habitats harbor diverse microbial communities, which can degrade lignocellulose effectively and are therefore a potential resource for improving anaerobic digestion. 

Characterization of wheat straw-degrading anaerobic alkali-tolerant mixed cultures from soda lake sediments by molecular and cultivation techniques.

Microbial flow cytometry is an established fast and economic technique for complex ecosystem studies and enables visualization of rapidly changing community structures by measuring characteristics of single microbial cells. Cytometric evaluation routines are available such as flowCyBar which are useful for automatic data processing. Here, a cytometric workflow was established which allows to routinely analyze salivary microbiomes on the example of ten oral healthy subjects. First, saliva was collected within a 3-month period, cytometrically analyzed and the evolution of the microbiomes followed as well as the calculation of their intra- and inter-subject similarity. Second, the respective microbiomes were stressed by exposition to high sugar or acid concentrations and immediate changes were recorded. Third, bactericide solutions were tested on their impact on the microbiomes. In all three set ups huge intra-individual variations in cytometric community structures were found to be largely absent, even under stress, while inter-individual diversity was obvious. The bacterial cell counts of saliva samples were found to vary between 3.0×10 and 6.2×10 cells per sample and subject in undisturbed environments. The application of the two bactericides did not cause noteworthy diversity changes but the loss in cell numbers by about 50% was high after treatment. Illumina® sequencing of whole microbiomes or sorted sub-microbiomes revealed typical phyla such as Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes and Fusobacteria. This approach is useful for fast monitoring of individual salivary microbiomes and automatic calculation of intra- and inter-individual dynamic changes and variability and opens insight into ecological principles leading to their sustainment in their individual environment.

Article	Year
Characterization of wheat straw-degrading anaerobic alkali-tolerant mixed cultures from soda lake sediments by molecular and cultivation techniques. Microbial biotechnology\| PubMed ID: 25737100	2015
A cytometric approach to follow variation and dynamics of the salivary microbiota. Methods (San Diego, Calif.)\| PubMed ID: 28842259	2018

Florian Schattenberg

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