Morphology engineering - Osmolality and its effect on Aspergillus niger morphology and productivity
Tóm tắt
The filamentous fungus Aspergillus niger is a widely used strain in a broad range of industrial processes from food to pharmaceutical industry. One of the most intriguing and often uncontrollable characteristics of this filamentous organism is its complex morphology, ranging from dense spherical pellets to viscous mycelia depending on culture conditions. Optimal productivity correlates strongly with a specific morphological form, thus making high demands on process control. In about 50 2L stirred tank cultivations the influence of osmolality on A. niger morphology and productivity was investigated. The specific productivity of fructofuranosidase producing strain A. niger SKAn 1015 could be increased notably from 0.5 to 9 U mg-1 h-1 around eighteen fold, by increasing the culture broth osmolality by addition of sodium chloride. The specific productivity of glucoamylase producing strain A. niger AB1.13, could be elevated using the same procedure. An optimal producing osmolality was shown to exist well over the standard osmolality at about 3.2 osmol kg-1 depending on the strain. Fungal morphology of all cultivations was examined by microscope and characterized by digital image analysis. Particle shape parameters were combined to a dimensionless Morphology number, which enabled a comprehensive characterization of fungal morphology correlating closely with productivity. A novel method for determination of germination time in submerged cultivations by laser diffraction, introduced in this study, revealed a decelerated germination process with increasing osmolality. Through the introduction of the versatile Morphology number, this study provides the means for a desirable characterization of fungal morphology and demonstrates its relation to productivity. Furthermore, osmolality as a fairly new parameter in process engineering is introduced and found to affect fungal morphology and productivity. Osmolality might provide an auspicious and reliable approach to increase the productivity in industrial processes. Because of the predictable behavior fungal morphology showed in dependence of osmolality, a customization of morphology for process needs seems feasible.
Tài liệu tham khảo
Grimm LH, Kelly S, Krull R, Hempel DC: Morphology and Productivity of Filamentous Fungi. Appl Microbiol Biotechnol. 2005, 69 (4): 375-384. 10.1007/s00253-005-0213-5.
Lubertozzi D, Keasling JD: Developing Aspergillus as a host for heterologous expression. Biotechnol Adv. 2009, 27 (1): 53-75. 10.1016/j.biotechadv.2008.09.001.
Papagianni M: Fungal morphology and metabolite production in submerged mycelial processes. Biotechnol Adv. 2004, 22 (3): 189-259. 10.1016/j.biotechadv.2003.09.005.
Papagianni M, Mattey M, Kristiansen B: Citric acid production and morphology of Aspergillus niger as functions of the mixing intensity in a stirred tank and a tubular loop bioreactor. Biochem Eng J. 1998, 2: 197-205. 10.1016/S1369-703X(98)00032-1.
Papagianni M, Mattey M, Kristiansen B: The influence of glucose concentration on citric acid production and morphology of Aspergillus niger in batch and glucostat culture. Enzyme Microb Technol. 1999, 25: 710-717. 10.1016/S0141-0229(99)00102-7.
Paul GC, Priede MA, Thomas CR: Relationship between morphology and citric acid production in submerged Aspergillus niger fermentations. Biochem Eng J. 1999, 3: 121-129. 10.1016/S1369-703X(99)00012-1.
Punt PJ, van Biezen N, Conesa A, Albers A, Mangnus J, van den Hondel C: Filamentous fungi as cell factories for heterologous protein production. Trends Biotechnol. 2002, 20 (5): 200-206. 10.1016/S0167-7799(02)01933-9.
Casas López JL, Sánchez Pérez JA, Fernández Sevilla JM, Rodríguez Porcel EM, Chisti Y: Pellet morphology, culture rheology and lovastatin production in cultures of Aspergillus terreus. J Biotechnol. 2005, 116 (1): 61-77. 10.1016/j.jbiotec.2004.10.005.
Gupta K, Mishra P, Srivastava P: A correlative evaluation of morphology and rheology of Aspergillus terreus during lovastatin fermentation. Biotechnol Bioprocess Eng. 2007, 12 (2): 140-146. 10.1007/BF03028640.
Kim JH, Lebeault JM, Reuss M: Comparative study on rheological properties of mycelial broth in filamentous and pelleted forms. Appl Biochem Biotechnol. 1983, 18 (1): 11-16.
Pazouki M, Panda T: Understanding the morphology of fungi. Bioproc Biosys Eng. 2000, 22 (2): 127-143.
Gibbs PA, Seviour RJ, Schmid F: Growth of Filamentous Fungi in Submerged Culture: Problems and Possible Solutions. Crit Rev Biotechnol. 2000, 20 (1): 17-48. 10.1080/07388550091144177.
Kaup B-A, Ehrich K, Pescheck M, Schrader J: Microparticle-enhanced cultivation of filamentous microorganisms: Increased chloroperoxidase formation by Caldariomyces fumago as an example. Biotechnol and Bioeng. 2008, 99 (3): 491-498. 10.1002/bit.21713.
Krull R, Cordes C, Horn H, Kampen I, Kwade A, Neu TR, Nörtemann B: Morphology of filamentous fungi - Linking cellular biology to process engineering using Aspergillus niger. Adv Biochem Eng/Biotechnol. 2010, 121 (116): 1-21.
Zhang Z, Jin B, Kelly J: Effects of cultivation parameters on the morphology of Rhizopus arrhizus and the lactic acid production in a bubble column reactor. Eng Life Sci. 2007, 7: 490-496. 10.1002/elsc.200700002.
McIntyre M, Muller C, Dynesen J, Nielsen J: Metabolic engineering of the morphology of Aspergillus. Adv Biochem Eng/Biotechnol. 2001, 73: 103-128. 10.1007/3-540-45300-8_6.
Driouch H, Sommer B, Wittmann C: Morphology engineering of Aspergillus niger for improved enzyme production. Biotechnol Bioeng. 2009, 105 (6): 1058-1068.
Kelly S, Grimm LH, Bendig C, Hempel DC, Krull R: Effects of fluid dynamic induced shear stress on fungal growth and morphology. Proc Biochem. 2006, 41: 2113-2117. 10.1016/j.procbio.2006.06.007.
Lin P-J, Scholz A, Krull R: Effect of volumetric power input by aeration and agitation on pellet morphology and product formation of Aspergillus niger. Biochem Eng J. 2010, 49 (2): 213-220. 10.1016/j.bej.2009.12.016.
Rocha-Valadez JA, Galindo E, Serrano-Carreón L: The influence of circulation frequency on fungal morphology: A case study considering Kolmogorov microscale in constant specific energy dissipation rate cultures of Trichoderma harzianum. J Biotechnol. 2007, 130 (4): 394-401. 10.1016/j.jbiotec.2007.05.001.
Znidarsic P, Pavko A: The Morphology of Filamentous Fungi in Submerged Cultivations as a Bioprocess Parameter. Food Technol Biotechnol. 2001, 39: 237-252.
Wucherpfennig T, Kiep KA, Driouch H, Wittmann C, Krull R: Morphology and Rheology in Filamentous Cultivations. Adv Appl Microbiol. Edited by: Allen I, Laskin SS, Geoffrey MG. 2010, Academic Press, 72: 89-136.
Allaway AEaJ DH: The influence of cations on glucose transport and metabolism by, and the loss of sugars alcohols from, the fungus Dendryphiella salina. New Phytol. 1970, 69: 581-593.
Allaway AEaJ DH: The effect of cations on glucose utilization by, and on the growth of, the fungus Dendryphiella salina. New Phytol. 1971, 70: 511-518. 10.1111/j.1469-8137.1971.tb02552.x.
Bobowicz-Lassociska T, Grajek W: Changes in protein secretion of Aspergillus niger caused by the reduction of the water activity by potassium chloride. Acta Biotechnol. 1995, 15 (3): 277-287. 10.1002/abio.370150305.
Fiedurek J: Effect of osmotic stress on glucose oxidase production and secretion by Aspergillus niger. J Basic Microbiol. 1998, 38 (2): 107-112. 10.1002/(SICI)1521-4028(199805)38:2<107::AID-JOBM107>3.0.CO;2-1.
Lee GM, Park SY: Enhanced specific antibody productivity of hybridomas resulting from hyperosmotic stress is cell line-specific. Biotechnol Lett. 1995, 17 (2): 145-150. 10.1007/BF00127978.
Lee M, Lee G: Effect of hypoosmotic pressure on cell growth and antibody production in recombinant Chinese hamster ovary cell culture. Cytotechnology. 2001, 36 (1): 61-69. 10.1023/A:1014032701800.
Oh SKW, Vig P, Chua F, Teo WK, Yap MGS: Substantial overproduction of antibodies by applying osmotic pressure and sodium butyrate. Biotechnol and Bioeng. 1993, 42 (5): 601-610. 10.1002/bit.260420508.
Ozturk SS, Palsson BO: Effect of medium osmolarity on hybridoma growth, metabolism, and antibody production. Biotechnol and Bioeng. 1991, 37 (10): 989-993. 10.1002/bit.260371015.
Shen D, Kiehl TR, Khattak SF, Li ZJ, He A, Kayne PS, Patel V, Neuhaus IM, Sharfstein ST: Transcriptomic responses to sodium chloride-induced osmotic stress: A study of industrial fed-batch CHO cell cultures. Biotechnol Progr. 2010, 26 (4): 1104-1115.
Fernandez RC, Ottoni CA, da Silva ES, Matsubara RM, Carter JM, Magossi LR, Wada MA, de Andrade Rodrigues MF, Maresma BG, AE M: Screening of -fructofuranosidase-producing microorganisms and effect of pH and temperature on enzymatic rate. Appl Microbiol Biotechnol. 2007, 75: 87-93. 10.1007/s00253-006-0803-x.
Maiorano A, Piccoli R, da Silva E, de Andrade Rodrigues M: Microbial production of fructosyltransferases for synthesis of pre-biotics. Biotechnol Lett. 2008, 30 (11): 1867-1877. 10.1007/s10529-008-9793-3.
Zuccaro A, Götze S, Kneip S, Dersch P, Seibel J: Tailor-Made Fructooligosaccharides by a Combination of Substrate and Genetic Engineering. ChemBioChem. 2008, 9 (1): 143-149. 10.1002/cbic.200700486.
Emmler M, Jungebloud A, Göcke Y, Cordes C, Horn H, Hempel DC: Apparent Delay of Product Secretion by Product Adsorption in Aspergillus niger. Eng Life Sci. 2006, 6 (5): 488-491. 10.1002/elsc.200620152.
Hooley P, Fincham DA, Whitehead MP, Clipson NJW: Fungal Osmotolerance. Adv Appl Microbiol. Edited by: Allen I, Laskin JWB, Geoffrey MG. 2003, Academic Press, 53: 177-211.
Griffin DH: Fungal Physiology. 1994, New York.: Wiley-Liss, 2
Beever RE, Laracy EP: Osmotic adjustment in the filamentous fungus Aspergillus nidulans. J Bacteriol. 1986, 168 (3): 1358-1365.
Tresner HD, Hayes JA: Sodium chloride tolerance of terrestrial fungi. Appl Microbiol. 1971, 22 (2): 210-213.
Money NP: Wishful Thinking of Turgor Revisited: The Mechanics of Fungal Growth. Fungal Genetics and Biology. 1997, 21 (2): 173-187. 10.1006/fgbi.1997.0976.
Clipson NJW, Jennings DH: Dendryphiella salina and Debaryomyces hansenii: models for ecophysiological adaptation to salinity by fungi which grow in the sea. Can J Bot. 1992, 2097-2105. 70
Paul G, Thomas C: Characterisation of mycelial morphology using image analysis. Adv Biochem Eng/Biotechnol. 1998, 1-59.
Park J-C, Matsuoka H, Takatori K, Kurata H: Adaptation of Aspergillus niger to acidic conditions and its relationship to salt stress and miconazole. Mycological Research. 1996, 100 (7): 869-874. 10.1016/S0953-7562(96)80037-2.
Park J-C, Nemoto Y, Homma T, Jing W, Chen Y, Matsuoka H, Ohno H, Takatori K, Kurata H: Adaptation of Aspergillus niger to short-term salt strees. Appl Microbiol Biotechnol. 1993, 40 (2): 394-398.
Kim Y, Nandakumar MP, Marten MR: Proteome map of Aspergillus nidulans during osmoadaptation. Fungal Genetics and Biology. 2007, 44 (9): 886-895. 10.1016/j.fgb.2006.12.001.
Kralj Kuncic M, Kogej T, Drobne D, Gunde-Cimerman N: Morphological Response of the Halophilic Fungal Genus Wallemia to High Salinity. Appl Environ Microbiol. 2010, 76 (1): 329-337. 10.1128/AEM.02318-09.
Galindo E, Larralde-Corona CP, Brito T, Córdova-Aguilar MS, Taboada B, Vega-Alvarado L, Corkidi G: Development of advanced image analysis techniques for the in situ characterization of multiphase dispersions occurring in bioreactors. J Biotechnol. 2005, 116 (3): 261-270. 10.1016/j.jbiotec.2004.10.018.
Villena GK, Fujikawa T, Tsuyumu S, Gutiérrez-Correa M: Structural analysis of biofilms and pellets of Aspergillus niger by confocal laser scanning microscopy and cryo scanning electron microscopy. Bioresour Technol. 2010, 101 (6): 1920-1926. 10.1016/j.biortech.2009.10.036.
Papagianni M: Quantification of the fractal nature of mycelial aggregation in Aspergillus niger submerged cultures. Microbial Cell Factories. 2006, 5 (1): 5-10.1186/1475-2859-5-5.
Tucker K, Thomas C: Effect of biomass concentration and morphology on the rheological parameters of Penicillum chrysogenum fermentation broths. Trans Inst Chem Eng. 1993, 71: 111-117.
Hille A, Neu TR, Hempel DC, Horn H: Effective diffusivities and mass fluxes in fungal biopellets. Biotechnol Bioeng. 2009, 103 (6): 1202-1213. 10.1002/bit.22351.
McIntyre M, Müller C, Dynesen J, Nielsen J: Metabolic Engineering of the Morphology of Aspergillus. Adv Biochem Eng/Biotechnol. 2001, 73: 103-128. 10.1007/3-540-45300-8_6.
Wösten HAB, Moukha SM, Sietsma JH, Wessels JGH: Localization of growth and secretion of proteins in Aspergillus niger. J Gen Microbiol. 1991, 137 (1): 2017-2023.
Wongwicharn A, McNeil B, Harvey L: Effect of Oxygen Enrichment on Morphology, Growth, and Heterologous Protein Production in Chemostat Cultures of Aspergillus niger B1-D. Biotech Bioeng. 1999, 65 (4): 416-424. 10.1002/(SICI)1097-0290(19991120)65:4<416::AID-BIT6>3.0.CO;2-Z.
Amanullah A, Blair R, Nienow AW, Thomas CR: Effects of Agitation Intensity on Mycelial Morphology and Protein Production in Chemostat Cultures of Recombinant Aspergillus oryzae. Biotechnol Bioeng. 1999, 62 (4): 434-446. 10.1002/(SICI)1097-0290(19990220)62:4<434::AID-BIT6>3.0.CO;2-D.
Grimm LH, Kelly S, Völkerding II, Krull R, Hempel DC: Influence of Mechanical Stress and Surface Interaction on the Aggregation of Aspergillus niger Conidia. Biotechnol Bioeng. 2005, 92 (7): 879-888. 10.1002/bit.20666.
O'Mahony RJ, Burns ATH, Millam S, Hooley P, Fincham DA: Isotropic growth of spores and salt tolerance in Aspergillus nidulans. Mycological Research. 2002, 106 (12): 1480-1486. 10.1017/S0953756202006949.
Osherov N, May GS: The molecular mechanisms of conidial germination. FEMS Microbiol Lett. 2001, 199 (2): 153-160. 10.1111/j.1574-6968.2001.tb10667.x.
Grimm LH, Kelly S, Hengstler J, Göbel A, Krull R, Hempel DC: Kinetic studies on the aggregation of Aspergillus niger conidia. Biotechnol Bioeng. 2004, 87 (2): 213-218. 10.1002/bit.20130.
Trinci AP: A kinetic study of the growth of Aspergillus nidulans and other fungi. J Gen Microbiol. 1969, 57 (1): 11-24.
Dantigny P, Bensoussan M, Vasseur V, Lebrihi A, Buchet C, Ismaili-Alaoui M, Devlieghere F, Roussos S: Standardisation of methods for assessing mould germination: A workshop report. Int J Food Microbiol. 2006, 108 (2): 286-291. 10.1016/j.ijfoodmicro.2005.12.005.
Judet D, Bensoussan M, Perrier-Cornet J-M, Dantigny P: Distributions of the growth rate of the germ tubes and germination time of Penicillium chrysogenum conidia depend on water activity. Food Microbiol. 2008, 25 (7): 902-907. 10.1016/j.fm.2008.05.007.
Araujo R, Rodrigues AG: Variability of Germinative Potential among Pathogenic Species of Aspergillus. J Clin Microbiol. 2004, 42 (9): 4335-4337. 10.1128/JCM.42.9.4335-4337.2004.
Papagianni M, Mattey M: Morphological development of Aspergillus niger in submerged citric acid fermentation as a function of the spore inoculum level. Application of neural network and cluster analysis for characterization of mycelial morphology. Microbial Cell Factories. 2006, 5 (1): 3-10.1186/1475-2859-5-3.
Carlsen M, Spohr AB, Nielsen J, Villadsen J: Morphology and physiology of an alpha-amylase producing strain of Aspergillus oryzae during batch cultivations. Biotechnol Bioeng. 1996, 49 (3): 266-276.
Mattern I, van Noort J, van den Berg c, Archer D, Roberts I, van den Hondel C: Isolation and characterization of mutants of Aspergillus niger deficient in extracellular proteases. Mol Gen Genomics. 1992, 234: 332-336.
van Hartingsveldt W, Mattern I, van Zeijl C, Pouwels P, van den Hondel C: Development of a homologous transformation system for Aspergillus niger based on the pyrG gene. Mol Gen Genet. 1987, 206: 71-75. 10.1007/BF00326538.
Withers J, Swift R, Wiebe M, Robson G, Punt P, van den Hondel C, Trinci A: Optimization and stability of glucoamylase production by recombinant strains of Aspergillus niger in chemostat culture. Biotech Bioeng. 1998, 4 (59): 407-418.
Huggett A, Nixon D: Use of glucose oxidase, peroxidase, and O-dianisidine in determination of blood and urinary glucose. Lancet. 1957, 273 (6991): 368-370.
Becker J, Klopprogge C, Schroder H, Wittmann C: Metabolic Engineering of the Tricarboxylic Acid Cycle for Improved Lysine Production by Corynebacterium glutamicum. Appl Environ Microbiol. 2009, 75 (24): 7866-7869. 10.1128/AEM.01942-09.
ImageJ. [http://rsbweb.nih.gov/ij/]
Collins TJ: ImageJ for microscopy. BioTechniques. 2007, 43 (1 Suppl): 25-30.