Machine learning applied for metabolic flux-based control of micro-aerated fermentations in bioreactors.

Various bio-based processes depend on controlled micro-aerobic conditions to achieve a satisfactory product yield. However, the limiting oxygen concentration varies according to the micro-organism employed, while for industrial applications, there is no cost-effective way of measuring it at low levels. This study proposes a machine learning procedure within a metabolic flux-based control strategy (SUPERSYS_MCU) to address this issue. The control strategy used simulations of a genome-scale metabolic model to generate a surrogate model in the form of an artificial neural network, to be used in a micro-aerobic fermentation strategy (MF-ANN). The meta-model provided setpoints to the controller, allowing adjustment of the inlet air flow to control the oxygen uptake rate. The strategy was evaluated in micro-aerobic batch cultures employing industrial Saccharomyces cerevisiae yeast, with defined medium and glucose as the carbon source, as a case study. The performance of the proposed control scheme was compared with a conventional fermentation and with three previously reported micro-aeration strategies, including respiratory quotient-based control and constant air flow rate. Due to maintenance of the oxidative balance at the anaerobiosis threshold, the MF-ANN provided volumetric ethanol productivity of 4.16 g·L-1 ·h-1 and a yield of 0.48 gethanol .gsubstrate-1 , which were higher than the values achieved for the other conditions studied (maximum of 3.4 g·L-1 ·h-1 and 0.35-0.40 gethanol ·gsubstrate-1 , respectively). Due to its modular character, the MF-ANN strategy could be adapted to other micro-aerated bioprocesses.

Cost analysis based on bioreactor cultivation conditions: Production of a soluble recombinant protein using Escherichia coli BL21(DE3).

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32 °C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Cost analysis based on bioreactor cultivation conditions: production of a soluble recombinant protein using Escherichia coli BL21(DE3)

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32°C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Cost analysis based on bioreactor cultivation conditions: production of a soluble recombinant protein using Escherichia coli BL21(DE3)

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32°C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Physiological and biochemical responses of a coralline alga and a sea urchin to climate change: Implications for herbivory.

Direct responses to rising temperatures and ocean acidification are increasingly well known for many single species, yet recent reviews have highlighted the need for climate change research to consider a broader range of species, how stressors may interact, and how stressors may affect species interactions. The latter point is important in the context of plant-herbivore interactions, as increasing evidence shows that increasing seawater temperature and/or acidification can alter algal traits that dictate their susceptibility to herbivores, and subsequently, community and ecosystem properties. To better understand how marine rocky shore environments will be affected by a changing ocean, in the present study we investigated the direct effects of short-term, co-occurring increased temperature and ocean acidification on a coralline alga (Jania rubens) and a sea urchin herbivore (Echinometra lucunter) and assessed the indirect effects of these factors on the algal-herbivore interaction. A 21-day mesocosm experiment was conducted with both algae and sea urchins exposed to ambient (24 °C, Low CO2), high-temperature (28 °C, Low CO2), acidified (24 °C, High CO2), or high-temperature plus acidified (28 °C, High CO2) conditions. Algal photosynthesis, respiration, and phenolic content were unaffected by increased temperature and CO2, but calcium carbonate content was reduced under high CO2 treatments in both temperatures, while total sugar content of the algae was reduced under acidified, lower temperature conditions. Metabolic rates of the sea urchin were elevated in the lower temperature, high CO2 treatment, and feeding assays showed that consumption rates also increased in this treatment. Despite some changes to algal chemical composition, it appears that at least under short-term exposure to climate change conditions, direct effects on herbivore metabolism dictated herbivory rates, while indirect effects caused by changes in algal palatability seemed to be of minor importance.

Production and purification of recombinant fragment of pneumococcal surface protein A (PspA) in Escherichia coli

New conjugated vaccines against Streptococcus pneumoniae are being developed using pneumococcal surfaceproteins as carriers. The pneumococcal surface protein A (PspA) was selected as carrier because it is indispensablefor virulence of S. pneumoniae. The PspA can be classified into 3 families according to the homology of proteinsequences, within each family there is immunological cross-reactivity and PspA from family 1 or 2 are present in99% of strains associated with pneumococcal invasive disease. Hence, the purpose of this work was to develop an industrial production and purification process of His-tagged recombinant fragment of PspA in E. coli BL21 (DE3),rfPspA245 from family 1. Fed-batch cultivations in 5-L bioreactors with defined medium were carried out using glycerol as carbon source. Itwas obtained circa 60 g/L of dry cell weight and 3.0 g/L of rfPspA. Cells were disrupted with 96.7% of efficiency by high pressure continuous homogenizer. The clarification step was done by centrifugation. The results ofchromatographic steps were analyzed by densitometry of SDS-PAGE protein bands. Using the chromatographicsequence anion exchange (Q-Sepharose) followed by metal affinity (IMAC-Sepharose), the rfPspA245 was obtained with 67% and 97% of purity respectively for each step and final recovery of 23%. In conclusion, the purification process was developed and rfPspA245 was obtained with high purity, but the recovery should still be improved.