Caldicellulosiruptor bescii DSM6725 stress responses to pH and osmolarity variation

Caldicellulosiruptor bescii is a hyper thermophile (70-80^oC) that can utilize a wide range of substrates. However, inhibitors present in biomass, generated through pretreatment or as a product of metabolism can limit growth and bioconversion to products. Such inhibitory chemicals like sugar or lignin degradation products and weak acids can form potential targets for tolerance studies. Complete inhibition of C. bescii growth at 200mM concentrations of organic acids like acetic acid (Baronofsky et.al., 1984); 50mM NaCl (Farkas et.al. 2013) and fermentation ends around pH 5.0 have been previously reported. In this study, we examined responses to acid and osmotic stresses using various compounds to study C. bescii growth and physiology. C. bescii DSM6725 was grown in fermenters at 75 ⁰C, pH was lowered to 6.0 at mid-log phase for treated group using sulfuric acid while controls were maintained at pH 7.2. Surprisingly increased cell growth was observed in treated fermenters as compared to controls. RNA-seq data analysis using DESeq2, from samples collected pre and post acid stress, detected ABC transporters, especially the xylose sugar ABC transporters as the most significantly differentially expressed genes. To test dependence of increased growth in fermenters on sulfate from sulfuric acid, bottled experiments with and without sodium sulfate were conducted and negligible growth differences were observed. pH optima experiments are ongoing and will be reported.  Preliminary NaCl osmolarity in the range 150-350 mOsm indicated longer lag phases above 200 mOsm with similar end-point growth densities across range and form the basis for more detailed ongoing studies.