Activity of ethanol-stressed Oenococcus oeni cells: a flow cytometric approach

Aims:  To study the effect of ethanol on Oenococcus oeni activity at the single cell level.
Methods and Results:  The active extrusion of the fluorescent probe carboxy fluorescein (cF) was used to assess the metabolic activity of ethanol-stressed O. oeni cells. Subsequent flow cytometric analysis revealed that O. oeni cells extrude the accumulated cF upon energizing with l -malic acid. However, O. oeni cells exposed to 12% (v/v) ethanol for 1 h showed a decreased capacity for active extrusion of cF. Moreover, two subpopulations could be distinguished, one of which being able to extrude cF and the other one remaining cF fluorescent. Growing cells in the presence of 8% (v/v) ethanol resulted in robust cells that maintained the capacity to actively extrude cF after being exposed to 12% (v/v) ethanol, which in turn correlated with the high levels of ATP observed in these ethanol stressed, malolactic fermentation (MLF) performing cells.
Conclusion:  From our results, it becomes evident that active extrusion of cF can be used to assess malolactic activity in O. oeni .
Significance and Impact of the Study:  The present study provides information for the development of a rapid method to assess the malolactic activity of individual O. oeni cells performing MLF during wine production.     

Characterization of germination receptors of Bacillus cereus ATCC 14579

Specific amino acids, purine ribonucleosides, or a combination of the two is required for efficient germination of endospores of Bacillus cereus ATCC 14579. A survey including 20 different amino acids showed that l-alanine, l-cysteine, l-threonine, and l-glutamine are capable of initiating the germination of endospores of B. cereus ATCC 14579. In addition, the purine ribonucleosides inosine and adenosine can trigger germination of the spores. Advanced annotation of the B. cereus ATCC 14579 genome revealed the presence of seven putative germination (ger) operons, termed gerG, gerI, gerK, gerL, gerQ, gerR, and gerS. To determine the role of the encoded putative receptors in nutrient-induced germination, disruption mutants were constructed by the insertion of pMUTIN4 into each of the seven operons. Four of the seven mutants were affected in the germination response to amino acids or purine ribonucleosides, whereas no phenotype could be attributed to the mutants with disrupted gerK, gerL, and gerS loci. The strain with a disrupted gerR operon was severely hampered in the ability to germinate: germination occurred in response to l-glutamine but not in the presence of any of the other amino acids tested. The gerG mutant showed significantly reduced l-glutamine-induced germination, which points to a role of this receptor in the l-glutamine germination signaling pathway. gerR, gerI, and gerQ mutants showed reduced germination rates in the presence of inosine, suggesting a role for these operons in ribonucleoside signaling. Efficient germination by the combination of l-glutamine and inosine was shown to involve the gerG and gerI operons, since the germination of mutants lacking either one of these receptors was significantly reduced. Germination triggered by the combination of l-phenylalanine and inosine was lost in the gerI mutant, indicating that both molecules are effective at the GerI receptor.     

Impact of sorbic acid on germinant receptor-dependent and -independent germination pathways in Bacillus cereus

Amino acid- and inosine-induced germination of Bacillus cereus ATCC 14579 spores was reversibly inhibited in the presence of 3 mM undissociated sorbic acid. Exposure to high hydrostatic pressure, Ca-dipicolinic acid (DPA), and bryostatin, an activator of PrkC kinase, negated this inhibition, pointing to specific blockage of signal transduction in germinant receptor-mediated germination.     

Seasonal changes in serum dehydroepiandrosterone,androstenedione, and testosterone levels in the squirrel monkey (Saimiri boliviensis boliviensis)

The squirrel monkey (Saimiri boliviensis boliviensis) has a well-defined breeding season during which adult males undergo androgen-dependent morphological changes, with acquisition of active spermatogenesis. To assess the hormonal events of this annual cycle, blood samples were obtained weekly from ten adult males, and serum was assayed for testosterone (T), androstenedione (ΔA), and dehydroepiandrosterone (DHEA). A significant seasonal variation was noted in mean serum T (P < 0.02), ΔA (P < 0.02), and DHEA (P < 0.001) concentrations. Mean ΔA concentrations increased from a nonbreeding season nadir of 91.4 ± 12.9 ng/ml (mean ± standard error) to a prebreeding concentration of 139 ± 10.5 ng/ml and breeding season peak of 167.5 ± 15.4 ng/ml (P < 0.05). Mean DHEA concentrations increased from a nonbreeding season nadir of 8.3 ± 0.8 to a breeding season peak of 14.3 ± 1.2 (P < 0.001). Mean T levels in the nonbreeding (52.2 ± 11.6 ng/ ml) and prebreeding season (48.6 ± 7.4) were similar. However, T significantly increased during the breeding season to 103.5 ± 12.8 ng/ml (P < 0.05). Progressive changes in body weight and morphology paralleled the rise in serum ΔA levels. The pattern of peripheral serum androgen concentrations throughout the year would suggest annual activation of the hypothalamic-pituitary-adrenal and/or hypothalamic-pituitary-gonadal axes.     

Restasis® for the treatment of 'dry eye' in Aotus nancymaae

Background  This case report describes the treatment of three male owl monkeys ( Aotus nancymaae ) diagnosed with chronic dry eye with a topical cyclosporine product, Restasis^®, approved for use in humans. These owl monkeys had ocular disease resulting from procedures performed at a biotechnology company. They were moved to the Center for Neotropical Primate Research and Resources at University of South Alabama to be incorporated into the breeding colony.
Materials and methods  Schirmer tear testing was performed initially and during the course of treatment to monitor efficacy of twice daily administered Restasis^®. The goals of treatment were to reduce pain and/or distress and if possible to quantitatively increase tear production.
Results and discussion  All animals had improvements in conjunctival inflammation and had an increase in tear production.     

Deletion of the sigB Gene in Bacillus cereus ATCC 14579 Leads to Hydrogen Peroxide Hyperresistance

The sigB gene of Bacillus cereus ATCC 14579 encodes the alternative sigma factor σ^B. Deletion of sigB in B. cereus leads to hyperresistance to hydrogen peroxide. The expression of katA, which encodes one of the catalases of B. cereus, is upregulated in the sigB deletion mutant, and this may contribute to the hydrogen peroxide-resistant phenotype.