Quantitative trait loci (QTL) underlying phenotypic variation in bioethanol-related processes in Neurospora crassa.

Bioethanol production from lignocellulosic biomass has received increasing attention over the past decade. Many attempts have been made to reduce the cost of bioethanol production by combining the separate steps of the process into a single-step process known as consolidated bioprocessing. This requires identification of organisms that can efficiently decompose lignocellulose to simple sugars and ferment the pentose and hexose sugars liberated to ethanol. There have been many attempts in engineering laboratory strains by adding new genes or modifying genes to expand the capacity of an industrial microorganism. There has been less attention in improving bioethanol-related processes utilizing natural variation existing in the natural ecotypes. In this study, we sought to identify genomic loci contributing to variation in saccharification of cellulose and fermentation of glucose in the fermenting cellulolytic fungus Neurospora crassa through quantitative trait loci (QTL) analysis. We identified one major QTL contributing to fermentation of glucose and multiple putative QTL's underlying saccharification. Understanding the natural variation of the major QTL gene would provide new insights in developing industrial microbes for bioethanol production.

Habitat-Specific Clock Variation and Its Consequence on Reproductive Fitness.

The circadian clock controls daily activities at the cellular and organismic level, allowing an organism to anticipate incoming stresses and to use resources accordingly. The circadian clock has therefore been considered a fitness trait in multiple organisms. However, the mechanism of how circadian clock variation influences organismal reproductive fitness is still not well understood. Here we describe habitat-specific clock variation (HSCV) of asexual reproduction in Neurospora discreta, a species that is adapted to 2 different habitats, under or above tree bark. African (AF) N. discreta strains, whose habitat is above the tree bark in light-dark (LD) conditions, display a higher rhythmicity index compared with North American (NA) strains, whose habitat is under the tree bark in constant dark (DD). Although AF-type strains demonstrated an overall fitness advantage under LD and DD conditions, NA-type strains exhibit a habitat-specific fitness advantage in DD over the LD condition. In addition, we show that allelic variation of the clock-controlled gene, Ubiquinol cytochrome c oxidoreductase (NEUDI_158280), plays a role in HSCV by modulating cellular reactive oxygen species levels. Our results demonstrate a mechanism by which local adaptation involving circadian clock regulation influences reproductive fitness.

Developing elite Neurospora crassa strains for cellulosic ethanol production using fungal breeding.

The demand for renewable and sustainable energy has generated considerable interest in the conversion of cellulosic biomass into liquid fuels such as ethanol using a filamentous fungus. While attempts have been made to study cellulose metabolism through the use of knock-out mutants, there have been no systematic effort to characterize natural variation for cellulose metabolism in ecotypes adapted to different habitats. Here, we characterized natural variation in saccharification of cellulose and fermentation in 73 ecotypes and 89 laboratory strains of the model fungus Neurospora crassa. We observed significant variation in both traits among natural and laboratory generated populations, with some elite strains performing better than the reference strain. In the F1 population N345, 15% of the population outperformed both parents with the top performing strain having 10% improvement in ethanol production. These results suggest that natural alleles can be exploited through fungal breeding for developing elite industrial strains for bioethanol production.

Macromolecular Interactions Control Structural and Thermal Properties of Regenerated Tri-Component Blended Films.

With a growing need for sustainable resources research has become highly interested in investigating the structure and physical properties of biomaterials composed of natural macromolecules. In this study, we assessed the structural, morphological, and thermal properties of blended, regenerated films comprised of cellulose, lignin, and hemicellulose (xylan) using the ionic liquid 1-allyl-3-methylimidazolium chloride (AMIMCl). Attenuated total reflectance Fourier transform infrared (ATR-FTIR) analysis, scanning electron microscopy (SEM), atomic force microscopy (AFM), X-ray scattering, and thermogravimetric analysis (TGA) were used to qualitatively and quantitatively measure bonding interactions, morphology, and thermal stability of the regenerated films. The results demonstrated that the regenerated films' structural, morphological, and thermal character changed as a function of lignin-xylan concentration. The decomposition temperature rose according to an increase in lignin content and the surface topography of the regenerated films changed from fibrous to spherical patterns. This suggests that lignin-xylan concentration alters the self-assembly of lignin and the cellulose microfibril development. X-ray scattering confirms the extent of the morphological and molecular changes. Our data reveals that the inter- and intra-molecular interactions with the cellulose crystalline domains, along with the amount of disorder in the system, control the microfibril dimensional characteristics, lignin self-assembly, and possibly the overall material's structural and thermal properties.