Staphylococcal growth and enterotoxins (A—D) and thermonuclease synthesis in the presence of dehydrated garlic

E. GONZÁLEZ-FANDOS, M.L. GARCÍA-LÓPEZ, M.L. SIERRA AND A. OTERO. 1994. The inhibition of Staphylococcus aureus growth and enterotoxin and thermonuclease production by various concentrations of garlic ( Allium sativum ) was studied in BHI broth. The growth of Staph. aureus was inhibited by dehydrated garlic at levels of 1.5% (w/v) and over. Enterotoxins A, B and C₁ were only detectable in broth containing < 1% of garlic while enterotoxin D was produced at a level of 2%. Garlic also inhibited thermonuclease (TNAse) production, complete inhibition being observed at levels ≥ 1.5%. TNAse was not always detected when enterotoxin was present.     

Probiotic properties of vaginal lactic acid bacteria to prevent metritis in cattle

AIMS: The isolation of bovine vaginal lactic acid bacteria (LAB) and the screening of their beneficial properties to select those that could be used as probiotics in the prevention of bovine metritis were performed. METHODS AND RESULTS: Out of 76 Lactobacillus sp. and seven Streptococcus sp. strains, a small number showed high- and medium hydrophobicity when the microbial adhesion to hydrocarbons method (MATH) was applied. In the agar plate diffusion test, a large number of strains inhibited vaginal bovine Escherichia coli 99/14 and human E. coli. This inhibition was due to acid. Only a few strains inhibited Actinomyces pyogenes 96/393, a pathogen isolated from bovine metritis. This inhibition remained after neutralization. The taxonomic identification of the selected strains was carried out by an amplified ribosomal DNA restriction analysis (ARDRA). Most of the strains were identified as Lactobacillus fermentum, a few as Lactobacillus gasseri and one as Lactobacillus rhamnosus. CONCLUSIONS: Bovine vaginal lactobacilli strains have differential surface properties. The strains selected are capable of inhibiting specific metritis pathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results can be applied for future studies to design a probiotic product to prevent metritis in dairy postpartum cows.     

Interactions of 17β-Hydroxysteroid Dehydrogenase Type 10 and Cyclophilin D in Alzheimer's Disease

The nucleus-encoded 17β-hydroxysteroid dehydrogenase type 10 (17β-HSD10) regulates cyclophilin D (cypD) in the mitochondrial matrix. CypD regulates opening of mitochondrial permeability transition pores. Both mechanisms may be affected by amyloid β peptides accumulated in mitochondria in Alzheimer's disease (AD). In order to clarify changes occurring in brain mitochondria, we evaluated interactions of both mitochondrial proteins in vitro (by surface plasmon resonance biosensor) and detected levels of various complexes of 17β-HSD10 formed in vivo (by sandwich ELISA) in brain mitochondria isolated from the transgenic animal model of AD (homozygous McGill-R-Thy1-APP rats) and in cerebrospinal fluid samples of AD patients. By surface plasmon resonance biosensor, we observed the interaction of 17β-HSD10 and cypD in a direct real-time manner and determined, for the first time, the kinetic parameters of the interaction (k_a 2.0?×?10⁵ M¹s^?1, k_d 5.8?×?10⁴ s^?1, and K_D 3.5?×?10^–10 M). In McGill-R-Thy1-APP rats compared to controls, levels of 17β-HSD10–cypD complexes were decreased and those of total amyloid β increased. Moreover, the levels of 17β-HSD10–cypD complexes were decreased in cerebrospinal fluid of individuals with AD (in mild cognitive impairment as well as dementia stages) or with Frontotemporal lobar degeneration (FTLD) compared to cognitively normal controls (the sensitivity of the complexes to AD dementia was 92.9%, that to FTLD 73.8%, the specificity to AD dementia equaled 91.7% in a comparison with the controls but only 26.2% with FTLD). Our results demonstrate the weakened ability of 17β-HSD10 to regulate cypD in the mitochondrial matrix probably via direct effects of amyloid β. Levels of 17β-HSD10–cypD complexes in cerebrospinal fluid seem to be the very sensitive indicator of mitochondrial dysfunction observed in neurodegeneration but unfortunately not specific to AD pathology. We do not recommend it as the new biomarker of AD.

     

The T→C mutation at position +96 of the untranslated region 3′ to the terminating codon of the β-globin gene is a rare polymorphism that does not cause a β-thalassemia as previously ascribed

We have observed a TC mutation at position +96 of the untranslated region 3 to the terminating codon of the -globin gene in members of two Czech families and one black family. Data from initial studies suggested that this change was the cause of a -thalassemia, but continued analyses have provided convincing evidence that this mutation is a simple polymorphism.     

Anti-Cancer Potential of MAPK Pathway Inhibition in Paragangliomas–Effect of Different Statins on Mouse Pheochromocytoma Cells

To date, malignant pheochromocytomas and paragangliomas (PHEOs/PGLs) cannot be effectively cured and thus novel treatment strategies are urgently needed. Lovastatin has been shown to effectively induce apoptosis in mouse PHEO cells (MPC) and the more aggressive mouse tumor tissue-derived cells (MTT), which was accompanied by decreased phosphorylation of mitogen-activated kinase (MAPK) pathway players. The MAPK pathway plays a role in numerous aggressive tumors and has been associated with a subgroup of PHEOs/PGLs, including K-RAS-, RET-, and NF1-mutated tumors. Our aim was to establish whether MAPK signaling may also play a role in aggressive, succinate dehydrogenase (SDH) B mutation-derived PHEOs/PGLs. Expression profiling and western blot analysis indicated that specific aspects of MAPK-signaling are active in SDHB PHEOs/PGLs, suggesting that inhibition by statin treatment could be beneficial. Moreover, we aimed to assess whether the anti-proliferative effect of lovastatin on MPC and MTT differed from that exerted by fluvastatin, simvastatin, atorvastatin, pravastatin, or rosuvastatin. Simvastatin and fluvastatin decreased cell proliferation most effectively and the more aggressive MTT cells appeared more sensitive in this respect. Inhibition of MAPK1 and 3 phosphorylation following treatment with fluvastatin, simvastatin, and lovastatin was confirmed by western blot. Increased levels of CASP-3 and PARP cleavage confirmed induction of apoptosis following the treatment. At a concentration low enough not to affect cell proliferation, spontaneous migration of MPC and MTT was significantly inhibited within 24 hours of treatment. In conclusion, lipophilic statins may present a promising therapeutic option for treatment of aggressive human paragangliomas by inducing apoptosis and inhibiting tumor spread.     

Characterization and subcellular targeting of GCaMP-type genetically-encoded calcium indicators

Genetically-encoded calcium indicators (GECIs) hold the promise of monitoring [Ca(2+)] in selected populations of neurons and in specific cellular compartments. Relating GECI fluorescence to neuronal activity requires quantitative characterization. We have characterized a promising new genetically-encoded calcium indicator-GCaMP2-in mammalian pyramidal neurons. Fluorescence changes in response to single action potentials (17+/-10% DeltaF/F [mean+/-SD]) could be detected in some, but not all, neurons. Trains of high-frequency action potentials yielded robust responses (302+/-50% for trains of 40 action potentials at 83 Hz). Responses were similar in acute brain slices from in utero electroporated mice, indicating that long-term expression did not interfere with GCaMP2 function. Membrane-targeted versions of GCaMP2 did not yield larger signals than their non-targeted counterparts. We further targeted GCaMP2 to dendritic spines to monitor Ca(2+) accumulations evoked by activation of synaptic NMDA receptors. We observed robust DeltaF/F responses (range: 37%-264%) to single spine uncaging stimuli that were correlated with NMDA receptor currents measured through a somatic patch pipette. One major drawback of GCaMP2 was its low baseline fluorescence. Our results show that GCaMP2 is improved from the previous versions of GCaMP and may be suited to detect bursts of high-frequency action potentials and synaptic currents in vivo.     

EVOLUTION OF MICROALGAE IN HIGHLY STRESSING ENVIRONMENTS: AN EXPERIMENTAL MODEL ANALYZING THE RAPID ADAPTATION OF DICTYOSPHAERIUM CHLORELLOIDES (CHLOROPHYCEAE) FROM SENSITIVITY TO RESISTANCE AGAINST 2,4,6‐TRINITROTOLUENE BY RARE PRESELECTIVE MUTATIONS1

The increasing rates of global extinction due to human activities necessitate studies of the ability of organisms to adapt to the new environmental conditions resulting from human disturbances. We investigated the evolutionary adaptation of a microalga to sudden environmental change resulting from exposure to novel toxic chemical residues. A laboratory strain of Dictyosphaerium chlorelloides (Naum.) Kom. and Perm. (Chlorophyceae) was exposed to increasing concentrations of the modern contaminant 2,4,6‐trinitrotoluene (TNT). When algal cultures were exposed to 30 mg·L ^{? 1} 1 Received 9 July 2001. Accepted 23 July 2002.
TNT, massive lysis of microalgal cells was observed. The key to understanding the evolution of microalgae in such a contaminated environment is to characterize the TNT‐resistant variants that appear after the massive lysis of the TNT‐sensitive cells. A fluctuation analysis demonstrated unequivocally that TNT did not facilitate the appearance of TNT‐resistant cells; rather it was found that TNT‐resistant cells appeared spontaneously by rare mutations under nonselective conditions, before exposure to TNT. The estimated mutation rate was 1.4 × 10 ^{? 5} mutants per cell division. Isolated resistant mutants exhibited a diminished fitness in the absence of TNT. Moreover, the gross photosynthetic rate of TNT‐resistant mutants was significantly lower than that of wild‐type cells. Competition experiments between resistant mutants and wild‐type cells showed that in small populations, the resistant mutants were driven to extinction. The balance between mutation rate and the rate of selective elimination determines the occurrence of about 36 TNT‐resistant mutants per million cells in each generation. These scarce resistant mutants are the guarantee of potential for adaptation.