Acetic acid inhibition of yeast fermentation has a negative impact in several industrial processes. As an initial step in the construction of a Saccharomyces cerevisiae strain with increased tolerance for acetic acid, mutations conferring resistance were identified by screening a library of deletion mutants in a multiply auxotrophic genetic background. Of the 23 identified mutations, 11 were then introduced into a prototrophic laboratory strain for further evaluation. Because none of the 11 mutations was found to increase resistance in the prototrophic strain, potential interference by the auxotrophic mutations themselves was investigated. Mutants carrying single auxotrophic mutations were constructed and found to be more sensitive to growth inhibition by acetic acid than an otherwise isogenic prototrophic strain. At a concentration of 80 mM acetic acid at pH 4.8, the initial uptake of uracil, leucine, lysine, histidine, tryptophan, phosphate, and glucose was lower in the prototrophic strain than in a non-acetic acid-treated control. These findings are consistent with two mechanisms by which nutrient uptake may be inhibited. Intracellular adenosine triphosphate (ATP) levels were severely decreased upon acetic acid treatment, which likely slowed ATP-dependent proton symport, the major form of transport in yeast for nutrients other than glucose. In addition, the expression of genes encoding some nutrient transporters was repressed by acetic acid, including HXT1 and HXT3 that encode glucose transporters that operate by facilitated diffusion. These results illustrate how commonly used genetic markers in yeast deletion libraries complicate the effort to isolate strains with increased acetic acid resistance.

Studies reporting on potentially toxic interactions between aqueous fullerene nanoparticles (nC 60 ) and microorganisms have been contradictory. When known confounding factors were avoided, growth yields of Saccharomyces cerevisiae and Escherichia coli cultured in the presence and absence of independently prepared lots of underivatized nC 60 were found not to be significantly different.The increasing use of nanomaterials in industrial processes and commercial products is expected to lead to accumulation of these materials in the environment. Because the consequences of increased environmental exposure are unclear, it is important that studies be undertaken to determine potential risks (4). Both deleterious effects (5,21,26,27,30) and a lack of toxicity (7, 14, 18) have been reported for aqueous nanoparticles of underivatized aqueous fullerene nanoparticles (nC 60 ). Only some of these conflicting observations have been rationalized (9). Accurately assessing doses of nanoparticles in cell culture systems can be problematic (24). The variety of methods used to prepare nC 60 also complicates interpretations of otherwise similar toxicological evaluations, because different preparation methods produce nC 60 particles with different physicochemical properties (2, 16).With specific reference to microorganisms, conflicting data have also been previously reported (19). At least four factors confound assessments of toxicity. First, it is now recognized that tetrahydrofuran (THF) used in nC 60 preparation generates toxic derivatives (13,22,29). Unless these derivatives and trace THF are removed, observed toxicity cannot be ascribed to nC 60 alone. Reports from studies that found antibacterial activity by the use of THF-solubilized nC 60 prior to this discovery are thus difficult to interpret (6,(15)(16)(17). Second, in aqueous media, hydrophobic nC 60 particles tend to agglomerate as a function of the solution condition. For some microbiological media, this leads to precipitation of nC 60 (17) and hence a reduction in the actual exposure dose. Binding of organic components in complex media to nC 60 particles can reduce nC 60 bioavailability or lead to agglomeration (15). Both effects would result in false-negative assessments of potential growth inhibition (3). Third, negative results reported from studies where C 60 powder was used directly without prior solubilization may reflect a lack of bioavailability (8,20,23,25). Fourth, potential inhibitory effects toward one or few species in mixed cultures could be masked by other species when growth is assessed at the community level (8,20,25).In light of these complications and a lack of studies done with fungi, which comprise a significant component of the soil microbial community, the toxicity of nC 60 towards the yeast Saccharomyces cerevisiae and Escherichia coli was assessed based on a simple growth endpoint under conditions where the aforementioned confounding factors were avoided. Pure microbial cultures were grown in minimal media to which carefully washed and characte...