High Substrate Uptake Rates Empower Vibrio natriegens as Production Host for Industrial Biotechnology

Abstract

The productivity of industrial fermentation processes is essentially limited by the biomass-specific substrate consumption rate (q_S ) of the applied microbial production system. Since q_S depends on the growth rate (μ), we highlight the potential of the fastest-growing nonpathogenic bacterium, Vibrio natriegens, as a novel candidate for future biotechnological processes. V. natriegens grows rapidly in BHIN complex medium with a μ of up to 4.43 h^-1 (doubling time of 9.4 min) as well as in minimal medium supplemented with various industrially relevant substrates. Bioreactor cultivations in minimal medium with glucose showed that V. natriegens possesses an exceptionally high q_S under aerobic (3.90 ± 0.08 g g^-1 h^-1) and anaerobic (7.81 ± 0.71 g g^-1 h^-1) conditions. Fermentations with resting cells of genetically engineered V. natriegens under anaerobic conditions yielded an overall volumetric productivity of 0.56 ± 0.10 g alanine liter^-1 min^-1 (i.e., 34 g liter^-1 h^-1). These inherent properties render V. natriegens a promising new microbial platform for future industrial fermentation processes operating with high productivity.IMPORTANCE Low conversion rates are one major challenge to realizing microbial fermentation processes for the production of commodities operating competitively with existing petrochemical approaches. For this reason, we screened for a novel platform organism possessing characteristics superior to those of traditionally employed microbial systems. We identified the fast-growing V. natriegens, which exhibits a versatile metabolism and shows striking growth and conversion rates, as a solid candidate to reach outstanding productivities. Due to these inherent characteristics, V. natriegens can speed up common laboratory routines, is suitable for already existing production procedures, and forms an excellent foundation for engineering next-generation bioprocesses.

Keywords: Vibrio natriegens; alanine production; glucose consumption rate; growth rate; industrial biotechnology; productivity.

FIG 1

(A and B) Growth of V. natriegens in different standard complex media (A) and in BHIN medium at a low cell density as shaking-flask cultures (B). (C) Growth rates in VN minimal medium with 10 g glucose liter⁻¹ under different NaCl concentrations in shaking flasks. For each condition, at least three independent experiments were performed. Error bars show standard deviations.

FIG 2

Bioreactor cultivations of V. natriegens in BHIN complex medium and VN minimal medium with 10 g glucose liter⁻¹ under aerobic and anaerobic conditions. Biomass formation as g cell dry weight (CDW) liter⁻¹, growth rates (μ), biomass yields (Y_X/S), and biomass-specific glucose consumption rates (q_S) are depicted in panels A to D, respectively. Time courses of biomass formation, glucose consumption, and product formation under aerobic and anaerobic conditions are shown in panels E and F. For each condition, three independent experiments were performed. Error bars show standard deviations.

FIG 3

Course of product formation and volumetric productivity (Q_P) of the respective products and combined Q_P (Σ) of resting high-cell-density cultures (∼20 g CDW liter⁻¹) of V. natriegens in VN medium with 42.5 g glucose liter⁻¹ under anaerobic conditions. Three independent experiments were performed. Error bars show standard deviations.

FIG 4

(A) Alanine yields (Y_P/S) of V. natriegens wild-type (V.n.) and its engineered derivatives, V. natriegens Δdldh (V.n. Δ), V. natriegens Δdldh Δlldh (V.n. ΔΔ), V. natriegens Δdldh Δlldh Δpfl (V.n. ΔΔΔ), and V. natriegens Δdldh Δlldh Δpfl Δmdh (V.n. ΔΔΔΔ) in VN medium with 5 g glucose liter⁻¹ under anaerobic conditions. (B) Alanine yield (Y_P/S) and volumetric productivity (Q_P) of zero-growth production at a high cell density with V. natriegens Δdldh Δlldh Δpfl Δmdh. At least three independent experiments were performed. Error bars show standard deviations.