Towards continuous industrial bioprocessing with solventogenic and acetogenic clostridia: challenges, progress and perspectives

Abstract

The sustainable production of solvents from above ground carbon is highly desired. Several clostridia naturally produce solvents and use a variety of renewable and waste-derived substrates such as lignocellulosic biomass and gas mixtures containing H2/CO2 or CO. To enable economically viable production of solvents and biofuels such as ethanol and butanol, the high productivity of continuous bioprocesses is needed. While the first industrial-scale gas fermentation facility operates continuously, the acetone–butanol–ethanol (ABE) fermentation is traditionally operated in batch mode. This review highlights the benefits of continuous bioprocessing for solvent production and underlines the progress made towards its establishment. Based on metabolic capabilities of solvent producing clostridia, we discuss recent advances in systems-level understanding and genome engineering. On the process side, we focus on innovative fermentation methods and integrated product recovery to overcome the limitations of the classical one-stage chemostat and give an overview of the current industrial bioproduction of solvents.

Sporulation in solventogenic and acetogenic clostridia

“…Once sporulation is initiated, cell growth stops, and the cell’s energy is used to generate metabolically inactive cells (Patakova et al 2013 ). These events are undesirable in industrial settings as they negatively impact solvent productivity and cause cell wash-out in a continuous process (Li et al 2020b ; Papoutsakis 2008 ; Vees et al 2020 ). Therefore, various attempts were made, like random mutagenesis, inactivation of early-stage sporulation proteins, and engineering of degenerated strain (Li et al 2020b ) to obtain asporogenous solventogenic strains or to control sporulation.…”

“…Spores can be used for cell immobilization. Low biomass is one of the issues of continuous culture with clostridia, and cell immobilization may prevent wash-out at high dilution rates (Vees et al 2020 ). Spore can be immobilized on porous carriers (Dolejš et al 2014 ; Krouwel et al 1983 ) and microencapsulation (Rathore et al 2015 ) to prevent cell wash-out.…”

“…These issues need to be tackled to enable competitive biofuel prices. According to estimates, the product titer in a bioprocess aiming for the biofuel market needs to reach at least 50 g L -1, and the productivity should amount to 3 g L -1 h -1 to be commercially viable (Vees et al 2020). While in standard batch conditions, solvent productivity has been reported to reach 10 g L -1 h -1 with clostridial fermentation, the clostridial maximum butanol titer in batch fermentation could not exceed 25 g L -1 without the application of in situ solvent removal techniques to prevent toxicity (Annous and Blaschek 1991;Qureshi and Blaschek 2001;Wang et al 2019;Xu et al 2015).…”

Genetic and metabolic engineering challenges of C1-gas fermenting acetogenic chassis organisms

“…In eukaryotic organisms, non-homologous end joining (NHEJ) allows to repair the breaks, creating random mutations when this option is desired. However, in prokaryotes unable to perform NHEJ (Joseph, Kim and Sandoval 2018 ; Vees, Neuendorf and Pflügl 2020 ) such as acetogens, genome editing via CRISPR-Cas9 relies on HR-based replacement of the target sequence with the desired mutant allele and subsequent elimination of the wild-type population through RNA-guided Cas9 cleavage of the parental allele. Thus, conventional allelic exchange mechanisms generate mutants, while CRISPR-Cas9 allows selection of mutants harbouring the desired allele from mixed populations and therefore, immune to cleavage, unlike the wild-type cells.…”

Butanol Tolerance of Lactiplantibacillus plantarum: A Transcriptome Study

“…Butanol emits less unburned hydrocarbon, carbon mono- and dioxide, nitrogen oxides, and other unregulated emissions compared with conventional transport fuels [ 1 ]. It is preferred over ethanol for the spark-ignition engine because of its higher energy density, excellent combustion characteristics, less corrosiveness, lower vapour pressure and volatility, and the opportunity to be blended with gasoline at any concentration [ 2 ]. Butanol is likewise indispensable in the chemical industry for latex paints and plastics [ 3 ] and in pharmacy and cosmetics as a solvent.…”