Abstract
Since filamentous fungi rapidly adjust their metabolic properties to environmental changes, a rigorous standardization and characterization of cultivation conditions is necessary to obtain meaningful and reproducible results. In batch cultures, which are commonly characterized according to the classical growth curve in textbooks (i.e., lag, exponential, stationary, and declining phase), this is of special difficulty. Although various studies in literature report atypically shaped growth curves of filamentous fungi in batch culture, systematic investigations on this topic are scarce and deviations are barely mentioned in textbooks. Summarizing approximately a decade of observations of growth characteristics from bioreactor batch grown filamentous fungi – in particular two strains (CBS123.823 and CBS123.824) of Penicillium ochrochloron – we demonstrate with a series of highly standardized bioreactor batch culture experiments that the classical growth curve failed to describe growth dynamics of the studied fungi in this work. The nature of the first exhausted nutrient was of remarkable importance for the resulting shape of the growth curve. In all experiments, online respirometry proved to be a powerful tool to distinguish growth phases and revealed more physiological states than expected from the mere biomass curve. In this respect we discuss why “atypical” shaped growth curves often remain unrecognized and that they might be the rule rather than the exception. Acknowledging the importance of the correct presentation of this complex topic in textbooks, we also propose a modified growth curve scheme to sensitize students for potential alternative shaped growth curves.
Highlights
Books must follow sciences, and not sciences books (Francis Bacon)This article is a plea for paying more attention to growth conditions and growth curves when growing filamentous fungi in submerged culture
For most bioreactor batch cultivations, where ammonium was used as nitrogen source and glucose as carbon-source, a filamentous growing preculture was produced using a HEPES-glucose medium (Gallmetzer et al, 1998) with the following composition: glucose × 1 H2O (400), (NH4)2SO4 (6.25), NH4Cl (12.5), KH2PO4 (5.8), MgSO4 × 7 H2O (1.6), HEPES (1000), 10 mL L−1 trace element solution; pH 7.3 adjusted with 10 M NaOH
For bioreactor batch experiments with nitrate as nitrogen source the preculture medium was as follows: glucose × 1 H2O (400), NaNO3 (25), NaCl (12.5), KH2PO4 (5.8), MgSO4 × 7 H2O (1.6), NaSO4 (6.25), HEPES (1000), 10 mL L−1 of trace element solution; pH 7.0 adjusted with 10 M NaOH
Summary
Not sciences books (Francis Bacon)This article is a plea for paying more attention to growth conditions and growth curves when growing filamentous fungi in submerged culture. The focus of this article is the delimitation and the characterization of the growth phases – and if applicable the physiological states within a growth phase – of our model filamentous fungus Penicillium ochrochloron in batch cultures with different nutrient limitations. It is worth considering the origin of the classical growth curve for batch cultures. As stated in the outstanding review of Wanner and Egli (1990), Buchanan (1918) summarized the knowledge of his time about the dynamics of bacterial growth in batch culture He proposed a growth curve consisting of seven growth phases, which nowadays is often simplified to the well-known four growth phases: lag, exponential, stationary and declining phase (e.g., Figure 1A). It was assumed that growth in batch culture stops immediately after depletion of the first essential nutrient – irrespective of the nature of this essential nutrient
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