Comparative Analysis of Growth and Morphology among Citric Acid Producing and Non-Producing Strains of Aspergillus niger
EMSL Project ID
16995
Abstract
The primary source of commercial citric acid production is the fermentation of sugars by the filamentous fungus Aspergillus niger. Empirical determinations of process conditions and strain selection have resulted in industrial strains that can convert 90% of a supplied sugar to citric acid. These conditions include high glucose concentrations, low pH and very low levels of Mn2+ in the culture media. Under these conditions, the normally filiform hyphae of A. niger are not present and the organism grows as a clump of cells or ‘pellet’ lacking any long hyphal extensions. Despite the fact that pelleted growth of filamentous fungi is the optimal morphology for the production of many different compounds by several fungal species, very little cell biological data exists that defines how the fungus grows from spores to form a pellet. A study performed by the Fungal Biotechnology Team at PNNL identified genes that are differentially expressed during pelleted growth or during filamentous growth in the presence of Mn2+ by subtractive hybridization of cDNA (Dai et al. Appl. Environ. Microbiol. Vol. 70, No. 4, 2004). More than half of the identified genes are only present in other fungi and most code for proteins with little or no peptide sequence similarity to proteins with known function. The objective of this work will be to define, from spore germination to pellet formation, the growth parameters of a strain that is optimized for citric acid production (An11414) and compare these data to other A. niger strains. These will include the 11414 parental strain, An1015, as well as a non production strain An9029 and a strain used for protein production AnNRRL3122. Each strain will be examined in a normal minimal medium as well as citric acid production media with and without additional Mn2+. Growth parameters will include hyphal length and diameter measurements as well as numbers of nuclei and septa per germling at earliest stages. The results of this study will serve two main purposes. First, a better understanding of the growth associated with pellet development will aid us in adapting this morphology to other systems. For example, these data will help us further define the effects of Mn2+ addition by providing details about the nuclear number and cell size under these conditions. Secondly, this study will aid in defining functions for the genes identified by Dai et al. 2004. Our group is currently in the process of creating null mutations of each of these genes. Detailed knowledge of the phenotype associated with pellet formation will be necessary to accurately explain the effects these mutations may have on pellet formation and citric acid production. The bulk of the counting of cells and collection of data will be performed in the Applied Biotechnology Laboratory in PSL 522. The EMSL resources needed for this study include access to a fluorescence microscope with a CCD camera for image acquisition and analysis as well as a confocal microscope for collecting Z-series images from fungal pellets.
Project Details
Project type
Exploratory Research
Start Date
2005-11-30
End Date
2006-11-07
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members