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Novel Mutations within the embB Gene in Ethambutol-Susceptible Clinical Isolates of Mycobacterium tuberculosis. Ann S. G. Lee, 2004.Genetic analysis of the embB gene revealed mutations in 17 (68%) of 25 ethambutol (EMB) resistant isolates (M306I, M306V, M306L, Q497R) but also in 4 (20%) of 20 EMB-susceptible isolates of Mycobacterium tuberculosis, namely, an ATG  ATM substitution resulting in M306I, G406N, and the novel alterations M423I and A659T .
Oxygen- and Glucose-Dependent Regulation of Central Carbon Metabolism in Pichia anomala. Elisabeth Fredlund, 2004.We investigated the regulation of the central aerobic and hypoxic metabolism of the biocontrol and non-Saccharomyces wine yeast Pichia anomala . In aerobic batch culture, P . anomala grows in the respiratory mode with a high biomass yield (0.59 g [dry weight] of cells g of glucose–1) and marginal ethanol, glycerol, acetate, and ethyl acetate production . Oxygen limitation, but not glucose pulse, induced fermentation with substantial ethanol production and 10-fold-increased ethyl acetate production . Despite low or absent ethanol formation, the activities of pyruvate decarboxylase and alcohol dehydrogenase were high during aerobic growth on glucose or succinate . No activation of these enzyme activities was observed after a glucose pulse . However, after the shift to oxygen limitation, both enzymes were activated threefold . Metabolic flux analysis revealed that the tricarboxylic acid pathway operates as a cycle during aerobic batch culture and as a two-branched pathway under oxygen limitation . Glucose catabolism through the pentose phosphate pathway was lower during oxygen limitation than under aerobic growth . Overall, our results demonstrate that P . anomala exhibits a Pasteur effect and not a Crabtree effect, i.e., oxygen availability, but not glucose concentration, is the main stimulus for the regulation of the central carbon metabolism . Characterization of the Stringent Response and relBbu Expression in Borrelia burgdorferi. Julia Bugrysheva, 2003.The stringent response is a global bacterial response to nutritional stress mediated by (p)ppGpp . We previously found that both noninfectious Borrelia burgdorferi strain B31 and infectious B . burgdorferi strain N40 produced large amounts of (p)ppGpp during growth in BSK-H medium and suggested that the stringent response was triggered in B . burgdorferi under these conditions . Here we report that (p)ppGpp levels in B . burgdorferi growing in BSK-II or BSK-H medium are not further increased by nutrient limitation or by serine hydroxamate-induced inhibition of protein synthesis and that the presence of (p)ppGpp during growth of N40 in BSK-H medium is not associated with decreased 16S rRNA synthesis . Decreased 16S rRNA synthesis was associated with the decreased growth rate of N40 seen during coculture with tick cells, which are growth conditions that were previously shown to decrease (p)ppGpp levels . One-half as much of the mRNA of the gene encoding the Rel protein of B . burgdorferi (relBbu) was produced by B31 as by N40 during in vitro growth (2 ± 0.5 and 4 ± 0.8 fg of relBbu mRNA/ng of total Borrelia RNA, respectively) . Although the amounts of N40 relBbu mRNA were identical during growth in vitro and in rat peritoneal chambers, they were markedly decreased during growth in nymphal ticks . In contrast to the lack of change in relBbu mRNA levels, larger amounts of a 78-kDa protein that was cross-reactive with antibodies to Bacillus subtilis RelBsu were detected in immunoblots of N40 lysates after growth in rat peritoneal chambers than after growth in vitro . Differences in the level of production of (p)ppGpp between B31 and N40 could not be explained by differences in relBbu promoters since identical transcriptional start sites 309 nucleotides upstream from the B31 and N40 relBbu ATG start codon and identical  70-like promoters were identified by primer extension and sequencing analysis . relBbu complemented an Escherichia coli CF1693 relA spoT double mutant for growth on M9 minimal medium, and the transformed cells produced relBbu mRNA . These results indicate that relBbu is functional and that its transcription and translation and production of (p)ppGpp are affected by environmental conditions in strains N40 and B31 . They also suggest that in B . burgdorferi, an organism with few rRNA operons that grows slowly, the role of (p)ppGpp may differ from the classic role played by this molecule in E . coli and that (p)ppGpp may not be responsible for growth rate control .
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