Noninvasive tool for optical online monitoring of individual biomass concentrations in a defined coculture

Abstract

Cocultures bear great potential in the conversion of complex substrates and process intensification, as well as, in the formation of unique components only available due to inter‐species interactions. Dynamic data of coculture composition is necessary for understanding and optimizing coculture systems. However, most standard online determined parameters measure the sum of all species in the reactor system. The kinetic behavior of the individual species remains unknown. Up to now, different offline methods are available to determine the culture composition, as well as the online measurement of fluorescence of genetically modified organisms. To avoid any genetic modification, a noninvasive online monitoring tool based on the scattered light spectrum was developed for microtiter plate cultivations. To demonstrate the potential, a coculture consisting of the bacterium Lactococcus lactis and the yeast Kluyveromyces marxianus was cultivated. Via partial least squares regression of scattered light spectra, the online determination of the individual biomass concentrations without further sampling and analyses is possible. The results were successfully validated by a Coulter counter‐analysis, taking advantage of the different cell sizes of both organisms. The findings prove the applicability of the new method to follow in detail the dynamics of a coculture.

Measurement Techniques to Resolve and Control Population Dynamics of Mixed-Culture Processes

“…There are two main limitations of this technique are mentioned: (i) the overlap of distributions of the bacterial strains; and (ii) changing cell volumes of A. vinelandii depending on culture conditions. The impact of both these limitations on the mixed culture analysis was shown more in detail and confirmed by Geinitz and colleagues, who investigated the mixed culture dynamics of Lactobacillus lactis and Kluyveromyces marxianus [52].…”

“…The applicability of scattered light spectra detection for co-culture performance screening in microtiter plates was published by Geinitz and colleagues ( Figure 3B), who measured the online trends of cell dry weight for the bacterium L. lactis and the yeast K. marxianus under different cultivation conditions [52]. Although both organisms differ in both size and morphology, there are only minor differences in the scattered light spectra (overlay >95%).…”

“…For lower-complexity mixed cultures and especially for defined co-cultures, various strain-specific features have been applied to resolve the population dynamics, which are often case specific. For example, bacteria-yeast co-cultures can be easily resolved using particle counting and size measurements [51,52]. When two co-culture members produce different metabolites, these metabolites can be taken as an indication of population ratios [53,54].…”

“…Optical measurement techniques are currently the most powerful tools that enable the monitoring of mixed cultures and will also have an integral role for active control of these systems ( Table 2). Available technologies are based on fluorescence, absorbance, and scattered light at a single wavelength or with the advanced ability to determine the spectra (Box 2 and Figure 3) [33,52,55,[73][74][75]. All technologies provide insight into the system without any relevant delay [76][77][78].…”

“…For example, Goodacre and colleagues showed in 1996 that Fourier transform-infrared (FTIR) spectroscopy can be used to identify hospital isolates [94]. It was shown that the FTIR spectrum of, for example, Streptococcus pneumoniae differs from that of Streptococcus pyogenes, which should theoretically allow an online signal for each species in combination with a suitable chemometric method, comparable to the scattered light-based method published by Geinitz and colleagues [52]. A first step in this direction was published by Schäwe and colleagues, who investigated the culture dynamic of a Pseudomonas putida and Rhodococcus ruber co-culture [95].…”