Changes in morphology of Rhizopus chinensis in submerged fermentation and their effect on production of mycelium-bound lipase

Teng, Yun; Xu, Yan; Wang, Dong

doi:10.1007/s00449-008-0259-8

Cited by 35 publications

(16 citation statements)

References 42 publications

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“…For example, dispersed hyphae enhance production of certain acids (fumaric acid), proteins (amylases, neo-fructosyltransferase, and phytases) and secondary metabolites (penicillin) [15, 16]. However, the rheological consequences of dispersed growth elevate medium viscosity, which, in turn, can cause extensive temperature and nutrient concentration gradients within bioreactors as a result of heat and mass transfer limitations [17, 18].…”

Section: Introductionmentioning

confidence: 99%

A quantitative image analysis pipeline for the characterization of filamentous fungal morphologies as a tool to uncover targets for morphology engineering: a case study using aplD in Aspergillus niger

Cairns

Feurstein

Zheng

et al. 2019

Biotechnol Biofuels

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Background Fungal fermentation is used to produce a diverse repertoire of enzymes, chemicals, and drugs for various industries. During submerged cultivation, filamentous fungi form a range of macromorphologies, including dispersed mycelia, clumped aggregates, or pellets, which have critical implications for rheological aspects during fermentation, gas/nutrient transfer, and, thus, product titres. An important component of strain engineering efforts is the ability to quantitatively assess fungal growth phenotypes, which will drive novel leads for morphologically optimized production strains. Results In this study, we developed an automated image analysis pipeline to quantify the morphology of pelleted and dispersed growth (MPD) which rapidly and reproducibly measures dispersed and pelleted macromorphologies from any submerged fungal culture. It (i) enables capture and analysis of several hundred images per user/day, (ii) is designed to quantitatively assess heterogeneous cultures consisting of dispersed and pelleted forms, (iii) gives a quantitative measurement of culture heterogeneity, (iv) automatically generates key Euclidian parameters for individual fungal structures including particle diameter, aspect ratio, area, and solidity, which are also assembled into a previously described dimensionless morphology number MN, (v) has an in-built quality control check which enables end-users to easily confirm the accuracy of the automated calls, and (vi) is easily adaptable to user-specified magnifications and macromorphological definitions. To concomitantly provide proof of principle for the utility of this image analysis pipeline, and provide new leads for morphologically optimized fungal strains, we generated a morphological mutant in the cell factory Aspergillus niger based on CRISPR-Cas technology. First, we interrogated a previously published co-expression networks for A. niger to identify a putative gamma-adaptin encoding gene ( aplD ) that was predicted to play a role in endosome cargo trafficking. Gene editing was used to generate a conditional aplD expression mutant under control of the titratable Tet-on system. Reduced aplD expression caused a hyperbranched growth phenotype and diverse defects in pellet formation with a putative increase in protein secretion. This possible protein hypersecretion phenotype could be correlated with increased dispersed mycelia, and both decreased pellet diameter and MN. Conclusion The MPD image analysis pipeline is a simple, rapid, and flexible approach to quantify diverse fungal morphologies. As an exemplar, we have demonstrated that the putative endosomal transport gene aplD plays a crucial role in A. niger filamentous growth and pellet formation during submerged culture. This suggests that endocytic components are ...

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Section: Introductionmentioning

confidence: 99%

A quantitative image analysis pipeline for the characterization of filamentous fungal morphologies as a tool to uncover targets for morphology engineering: a case study using aplD in Aspergillus niger

Cairns

Feurstein

Zheng

et al. 2019

Biotechnol Biofuels

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“…In other cases freely dispersed mycelium seems favorable and regarded as prerequisite to ensure high productivity (Papagianni and Mattey, 2006). This holds for the production of enzymes such as amylase, neo-fructosyltransferase, or phytase (Teng et al, 2009) or the production of penicillin (Vecht-Lifshitz et al, 1990). Here, the mycelial form allows an increased oxygen supply of the cells stimulating growth and production (Wittler et al, 1986).…”

Section: Introductionmentioning

confidence: 99%

Morphology engineering of Aspergillus niger for improved enzyme production

Driouch

Sommer

Wittmann

2010

Biotech & Bioengineering

160

129

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Supplementation with silicate microparticles was used as novel approach to control the morphological development of Aspergillus niger, important as the major world source of citric acid and higher-value enzymes, in submerged culture. With careful variation of size and concentration of the micromaterial added, a number of distinct morphological forms including pellets of different size, free dispersed mycelium, and short hyphae fragments could be reproducibly created. Aluminum oxide particles similarly affected morphology, showing that this effect is largely independent of the chemical particle composition. Image analysis of morphological development of A. niger during the cultivation process showed that the microparticles influence the morphology by collision-induced disruption of conidia aggregates and probably also the hindrance of new spore-spore interactions in the very early stage of the process. Exemplified for different recombinant A. niger strains enzyme production could be strongly enhanced by the addition of microparticles. Linked to the formation of freely dispersed mycelium, titers for glucoamylase (GA) expressed as intracellular enzyme (88 U/mL) and fructofuranosidase secreted into the supernatant (77 U/mL), were up to fourfold higher in shake flasks. Moreover, accumulation of the undesired by-product oxalate was suppressed by up to 90%. The microparticle strategy could be successfully transferred to fructofuranosidase production in bioreactor, where a final titer of 160 U/mL could be reached. Using co-expression of GA with green fluorescent protein, enzyme production was localized in the cellular aggregates of A. niger. For pelleted growth, protein production was maximal only within a thin layer at the pellet surface and markedly decreased in the pellet interior, whereas the interaction with the microparticles created a highly active biocatalyst with the dominant fraction of cells contributing to production.

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“…In concordance with our observation, a freely dispersed mycelium from A. niger SKAn1015 induced by the supplementation of silicate microparticles was linked to an increase of a fructofuranosidase activity displaying the microparticles itself an important physiological role (Driouch et al 2010). On the contrary, the performance of a synthetic mycelium-bound lipase activity from Rhizopus chinensis was favoured by fully entangled mycelial filaments (Teng et al 2009). However, as it will be discussed later, the lipase production by filamentous fungi should be analyzed from a holistic point of view.…”

Section: Biomass Morphology and Lipase Productionmentioning

confidence: 99%

Orchestration an extracellular lipase production from Aspergillus niger MYA 135: biomass morphology and fungal physiology

et al. 2021

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The impact of biomass morphology and culture conditions on fungal fermentation was widely reviewed in the literature. In this work, we presented three independent experiments in order to evaluate the influence of some of those input factors on a lipase production separately by using the Aspergillus niger MYA 135 and the two-stage fermentation technique. Regarding the culture modality, the biomass was pre-grown in a first reactor. Then, the washed mycelium was transferred to a second reactor to continue the study. Firstly, linear effects of fungal morphology and several physiological parameters on a lipase production were explored using the Plackett–Burman design. The dispersed fungal morphology was confirmed as a proper quality characteristic for producing an extracellular lipase activity. Concerning the impact of the carbon source on the biomass pre-growth, the sucrose (E = 9.923, p < 0.001) and the l-arabinose (E = 4.198, p = 0.009) presented positive and significant effects on the enzyme production. On the contrary, the supplementation of 0.05 g/L CaCl2 displayed a highly negative and significant effect on this process (E = − 7.390, p < 0.001). Secondly, the relationship between the enzyme production and the input variables N:C ratio, FeCl3 and olive oil was explored applying the central composite design. Among the model terms, the N:C ratio of the production medium had the most negative and significant influence on the enzyme synthesis. Thus, it was concluded that a low N:C ratio was preferable to increase its production. In addition, the bifunctional role of FeCl3 on this fungus was presented. Thirdly, a prove of concept assay was also discussed.

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Changes in morphology of Rhizopus chinensis in submerged fermentation and their effect on production of mycelium-bound lipase

Cited by 35 publications

References 42 publications

A quantitative image analysis pipeline for the characterization of filamentous fungal morphologies as a tool to uncover targets for morphology engineering: a case study using aplD in Aspergillus niger

A quantitative image analysis pipeline for the characterization of filamentous fungal morphologies as a tool to uncover targets for morphology engineering: a case study using aplD in Aspergillus niger

Morphology engineering of Aspergillus niger for improved enzyme production

Orchestration an extracellular lipase production from Aspergillus niger MYA 135: biomass morphology and fungal physiology

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