Project Details
Projekt
Analysis of microbial relations in vivo using Raman Microscopy (AMIRA)
Subject Area
Analytical Chemistry
Term
from 2018 to 2022
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 397827619
Bacteria rarely occur in their planktonic form in nature, instead they associate into biofilms. Biofilms are dense, highly-structured, highly-diverse meta-organisms, which show incredible abilities not seen in the individual species comprising them. Information about the descend and relative relatedness, i.e. the phylogeny, of these spatially arranged microbial populations is essential for understanding and harnessing these incredibly powerful natural resources. However, to date all phylogenetic methods are inherently invasive, rendering a phylogenetically sensitive observation of ongoing, undisturbed biofilm formation impossible.In this project, we intend to establish cytochrome c-resonant Raman microscopy (CRRM) for the continuous analysis of bacterial relations in native, undisturbed biofilms in vivo and in situ with a resolution at cell level. This will allow for the first time the direct observation of ongoing biofilm formation including phylogenetic information and phenotypic adaptation in complex biofilms in vivo.CRRM was used successfully for structural analysis of biofilms at cell level in vivo. Since cytochromes are known evolutionary markers, it will also allow in vivo phylogeny if calibrated and tested against a well-defined, well-characterized set of bacteria strains in pure planktonic cultures and while associating into biofilm. For this, we have to precisely determine the phylogenetic range of the method under changing conditions and its comparability with the established but invasive standard methods for bacterial phylogeny. We also have to optimize the spectrum analysis procedures and determine the spectral requirements to be met for reliable results under unknown conditions.Ammonium oxidizing bacteria (AOB) will be used as reference. AOB are a subset of bacteria known to be of utmost importance for the Earths nitrogen cycle. In addition, biofilms with a high AOB content are crucial for the efficient treatment of wastewater worldwide, yet unsatisfactory little is known about their formation. Therefore, we will use twenty well-characterized stains of AOB covering the whole order of Nitrosomonadales. AOB will be analyzed in pure planktonic cultures under optimal growth conditions, in pure planktonic cultures suffering environmental stress, and while undergoing biofilm formation under two different flow regimes.Within this project, we intend to implement a fully functional method for the phylogenetically sensitive analysis of complex bacteria biofilms. We will form the first optical phylogenetic distance tree based on non-invasive, contact-free in vivo measurements of individual bacteria cells in situ. Thus, for the first time, studies of undisturbed ongoing formation of single and triple species AOB biofilms in vivo and in situ and the recording of phylogenetic movies of ongoing biofilm formation will be possible.
DFG Programme
Research Grants
