Explore the vast range of titles available.

Background Analyses of substrate and metabolites are often bottleneck activities in high-throughput screening of microbial bioprocesses. We have assessed Fourier transform infrared spectroscopy (FTIR), in combination with high throughput micro-bioreactors and multivariate statistical analyses, for analysis of metabolites in high-throughput screening of microbial bioprocesses. In our previous study, we have demonstrated that high-throughput (HTS) FTIR can be used for estimating content and composition of intracellular metabolites, namely triglyceride accumulation in oleaginous filamentous fungi. As a continuation of that research, in the present study HTS FTIR was evaluated as a unified method for simultaneous quantification of intra- and extracellular metabolites and substrate consumption. As a proof of concept, a high-throughput microcultivation of oleaginous filamentous fungi was conducted in order to monitor production of citric acid (extracellular metabolite) and triglyceride lipids (intracellular metabolites), as well as consumption of glucose in the cultivation medium. Results HTS FTIR analyses of supernatant samples was compared with an attenuated total reflection (ATR) FTIR, which is an established method for bioprocess monitoring. Glucose and citric acid content of growth media was quantified by high performance liquid chromatography (HPLC). Partial least square regression (PLSR) between HPLC glucose and citric acid data and the corresponding FTIR spectral data was used to set up calibration models. PLSR results for HTS measurements were very similar to the results obtained with ATR methodology, with high coefficients of determination (0.91–0.98) and low error values (4.9–8.6%) for both glucose and citric acid estimates. Conclusions The study has demonstrated that intra- and extracellular metabolites, as well as nutrients in the cultivation medium, can be monitored by a unified approach by HTS FTIR. The proof-of-concept study has validated that HTS FTIR, in combination with Duetz microtiter plate system and chemometrics, can be used for high throughput screening of microbial bioprocesses. It can be anticipated that the approach, demonstrated here on single-cell oil production by filamentous fungi, can find general application in screening studies of microbial bioprocesses, such as production of single-cell proteins, biopolymers, polysaccharides, carboxylic acids, and other type of metabolites.

Background Oleaginous fungi can accumulate lipids by utilizing a wide range of waste substrates. They are an important source for the industrial production of omega-6 polyunsaturated fatty acids (gamma-linolenic and arachidonic acid) and have been suggested as an alternative route for biodiesel production. Initial research steps for various applications include the screening of fungi in order to find efficient fungal producers with desired fatty acid composition. Traditional cultivation methods (shake flask) and lipid analysis (extraction-gas chromatography) are not applicable for large-scale screening due to their low throughput and time-consuming analysis. Here we present a microcultivation system combined with high-throughput Fourier transform infrared (FTIR) spectroscopy for efficient screening of oleaginous fungi. Results The microcultivation system enables highly reproducible fungal fermentations throughout 12 days of cultivation. Reproducibility was validated by FTIR and HPLC data. Analysis of FTIR spectral ester carbonyl peaks of fungal biomass offered a reliable high-throughput at-line method to monitor lipid accumulation. Partial least square regression between gas chromatography fatty acid data and corresponding FTIR spectral data was used to set up calibration models for the prediction of saturated fatty acids, monounsaturated fatty acids, polyunsaturated fatty acids, unsaturation index, total lipid content and main individual fatty acids. High coefficients of determination (R2 = 0.86–0.96) and satisfactory residual predictive deviation of cross-validation (RPDCV = 2.6–5.1) values demonstrated the goodness of these models. Conclusions We have demonstrated in this study, that the presented microcultivation system combined with rapid, high-throughput FTIR spectroscopy is a suitable screening platform for oleaginous fungi. Sample preparation for FTIR measurements can be automated to further increase throughput of the system.

Background Lactic acid bacteria are commonly used as protective starter cultures in food products. Among their beneficial effects is the production of ribosomally synthesized peptides termed bacteriocins that kill or inhibit food-spoiling bacteria and pathogens, e.g., members of the Listeria species. As new bacteriocins and producer strains are being discovered rapidly, modern automated methods for strain evaluation and bioprocess development are required to accelerate screening and development processes. Results In this study, we developed an automated workflow for screening and bioprocess optimization for bacteriocin producing lactic acid bacteria, consisting of microcultivation, sample processing and automated antimicrobial activity assay. We implemented sample processing workflows to minimize bacteriocin adsorption to producer cells via addition of Tween 80 and divalent cations to the cultivation media as well as acidification of culture broth prior to cell separation. Moreover, we demonstrated the applicability of the automated workflow to analyze influence of media components such as MES buffer or yeast extract for bacteriocin producers Lactococcus lactis B1629 and Latilactobacillus sakei A1608. Conclusions Our automated workflow provides advanced possibilities to accelerate screening and bioprocess optimization for natural bacteriocin producers. Based on its modular concept, adaptations for other strains, bacteriocin products and applications are easily carried out and a unique tool to support bacteriocin research and bioprocess development is provided.

Fungal production of polyunsaturated fatty acids (PUFAs) is a highly potential approach in biotechnology. Currently the main focus is directed towards screening of hundreds strains in order to select of few potential ones. Thus, a reliable method for screening a high number of strains within a short period of time is needed. Here, we present a novel method for screening of PUFA-producing fungi by high-throughput microcultivation and FTIR spectroscopy. In the study selected Mucor fungi were grown in media with different carbon sources and fatty acid profiles were predicted on the basis of the obtained spectral data. FTIR spectra were calibrated against fatty acid analysis by GC-FD. The calibration models were cross-validated and correlation coefficients (R 2 ) from 0.71 to 0.78 with RMSECV (root mean squared error) from 2.86% to 6.96% (percentage of total fat) were obtained. The FTIR results show a strong correlation to the results obtained by GC analysis, where high total contents of unsaturated fatty acids (both PUFA and MUFA) were achieved for Mucor plumbeus VI02019 cultivated in canola, olive and sunflower oil and Mucor hiemalis VI01993 cultivated in canola and olive oil.

Lipases are enzymes that can be secreted by several microorganisms, making interesting the biodiversity exploration for searching new microorganisms able to produce these enzymes. Many agro-industrial residues can be used as potential substrates for production of enzymes. The main objective of this work was the isolation and screening of microorganisms with potential to produce lipases. Among 24 fungi, five were selected as good lipase producers using tributyrin on agar plates and solid state fermentation of soybean bran. Two of them were isolated from soil samples, another two from soybean bran, and one from dairy products. These fungi were identified by microcultivation technique as from Penicillium and Aspergillus genera. Through random amplified polymorphic DNA technique, the most promising strains could be genetically discriminated, selecting two fungi as good lipase producers but genetically different. One isolated from soybean bran could hydrolyze efficiently triglycerides with fatty acids with different chain length. Another isolated from dairy products was only effective to hydrolyze triglycerides with long-chain fatty acids. Two distinct groups could be verified by means of this technique, comprising the most productive strains and the lowest or nonproductive ones in terms of hydrolytic activity.

Infections with antimicrobial-resistant (AMR) bacteria are globally on the rise. In the future, multi-resistant infections will become one of the major problems in global health care. In order to enable reserve antibiotics to retain their effect as long as possible, broad-spectrum antibiotics must be used sparingly. This can be achieved by a rapid microfluidic phenotypic antibiotic susceptibility test, which provides the information needed for a targeted antibiotic therapy in less time than conventional tests. Such microfluidic tests must cope with a low bacteria concentration. On-chip filtering of the samples to accumulate bacteria can shorten the test time. By means of fluorescence microscopy, we examined a novel nanogap filtration principle to hold back Escherichia coli and to perform cultivation experiments with and without antibiotics present. Microfluidic chips based on the nanogap flow principle showed to be useful for the concentration and cultivation of E. coli. With a concentration of 106 cells/mL, a specific growth rate of 0.013 min−1 and a doubling time of 53 min were achieved. In the presence of an antibiotic, no growth was observed. The results prove that this principle can, in future, be used in fast and marker-free antimicrobial susceptibility testing (AST).

Alginate microbeads, produced by emulsion/internal gelation, were studied for the entrapment and microcultivation of microbial cells with biotechnological potential. An anaerobic consortium which was selected for its capacity to degrade complex carbohydrates, and a pure culture of cellulose degrading bacteria were used for entrapment studies. Optimization of conditions for the formation of spherical alginate microbeads in sizes between 20 and 80 μm were examined. The best conditions were achieved by combining rapeseed methyl ester as oil phase and stirring at 100 rpm using a rotation impeller. Calcium alginate microbeads produced under these conditions were shown to present morphological stability, with large pores in the internal matrix that favours microcolony development. Finally, single cells were observed inside the beads after the entrapment procedure and microcolony formation was confirmed after cultivation in cellobiose.