Mutation to auxotrophy and prototrophy as related to symbiotic effectiveness in Rhizobium leguminosarum and R. trifolii

1969 ◽  
Vol 15 (6) ◽  
pp. 611-622 ◽  
Author(s):  
E. A. Schwinghamer
Keyword(s):  
Rhizobium Leguminosarum ◽  
Auxotrophic Mutant ◽  
Metabolic Inhibitors ◽  
Nutritional Requirement ◽  
Metabolic Defects ◽  
Resistant Mutants ◽  
The Relationship ◽  
Derivatives Of ◽  
Very High

Auxotrophs were isolated from four effective strains of Rhizobium leguminosarum and R. trifolii for a study of the relationship between metabolic defects and loss of ability for symbiosis. Most auxotrophs were isolated indirectly by prior isolation of mutants for resistance to metabolic inhibitors, especially D-alanine and D-histidine. The likelihood of some nutritional requirement was greatly increased among such resistant mutants, and was very high for derivatives of the R. trifolii strains. The complexity of growth factor requirement varied greatly between auxotrophs, and few had simple requirements. The most common requirement was for vitamins, notably thiamine. Partial or full restoration of effectiveness in symbiosis often accompanied reversion to prototrophy in some ineffective auxotrophs. The frequency of some degree of restoration among prototrophs varied from 0% to 100%, depending on the auxotrophic mutant involved. In one ineffective auxotroph of R. trifolii strain T1 the level of reversion (partial to complete) in vitro varied considerably and generally paralleled the degree of restoration of effectiveness. Biochemical deficiency appeared to be meaningfully related to impaired symbiosis in some auxotrophs but the relationship was probably incidental in most others. These auxotroph–prototroph studies are considered from the standpoint of relationship to several areas of research on Rhizobium.

The relationship between the digestibility of a sward and the herbage consumption of grazing calves

1968 ◽  
Vol 70 (1) ◽  
pp. 47-51 ◽  
Author(s):  
J. Hodgson
Keyword(s):  
Food Intake ◽  
Linear Relationship ◽  
Full Range ◽  
Chromic Oxide ◽  
Regression Equations ◽  
Faecal Output ◽  
Herbage Intake ◽  
The Relationship ◽  
Very High

SUMMARYNitro-chalk was applied to S.23 ryegrass swards, at approximately monthly intervals, at two contrasting levels in 1961 and three levels in 1962. Steer calves, 3–6 months old, grazed in rotation round a series of four paddocks on each treatment. Paddocks were trimmed and fertilizer applied after each grazing. The herbage intakes of the calves were determined on three occasions in 1961 and four occasions in 1962. Faecal output was estimated by chromic oxide dilution. In vitro digestibility determinations were carried out on samples of herbage clipped from the swards.There was a close linear relationship between the digestibility of the herbage and the amount eaten, over the full range of digestibility encountered (68–82%). Regression equations calculated within seasons and fertilizer levels, and within years, did not differ significantly. The regressions of faecal output on herbage digestibility were not significantly different from zero.The observed relationship between herbage digestibility and herbage intake may reflect (a) the greater sensitivity of young ruminants than adult stock to changes in the digestibility of the diet, and (b) the reduced importance of the chemostatic control of food intake, except at very high levels of digestibility, in young rapidly growing animals.

scholarly journals The binding and processing of monoclonal human IgG1 by cells of a human macrophage-like cell line (U937)

Blood ◽  
1983 ◽  
Vol 62 (3) ◽  
pp. 652-662 ◽  
Author(s):  
RJ Kurlander ◽  
JE Gartrell
Keyword(s):  
Cell Line ◽  
Metabolic Inhibitors ◽  
Human Macrophage ◽  
Reversible Binding ◽  
In Vitro Binding ◽  
Irreversible Reduction ◽  
Vitro Binding ◽  
Human Igg1 ◽  
The Relationship

Abstract The goal of these experiments was to assess the relationship between the binding and processing of IgG by Fc-receptor-bearing cells. Cells of the U937 human macrophage-like cell line were incubated with 125I- labeled monomers, dimers, oligomers (composed of 2–4 IgG1 subunits), and HP (heavy polymers composed of 5 or more subunits per polymer) of monoclonal human IgG1 in vitro. Binding was assessed by spinning cells through a layer of phthalate oils. Internalization of IgG1 was assessed by quantitating residual binding to cells after surface-bound IgG was removed by a brief treatment with a solution containing 0.25 M acetic acid and 0.5 M sodium chloride. Catabolism was assessed by measuring the release of radioactive fragments of IgG1, which were not precipitated by 10% trichloroacetic acid. Unstimulated U937 bound about 10,000 molecules per cell of IgG1 monomer, with an equilibrium binding constant (Ka) of 5 X 10(8) M-1. After stimulation with a conditioned medium in vitro, binding per cell was increased 3–7--fold, and the Ka was decreased 2–4--fold. Both unstimulated and stimulated cells internalized and catabolized labeled IgG1 HP, but stimulated cells internalized and digested much more IgG1 HP per cell than unstimulated cells. Both monomers and dimers of IgG1 were internalized and degraded very slowly by stimulated cells, even though both preparations readily bound to cells. In contrast, oligomers and (to an even greater extent) IgG1 HP were internalized and degraded much more rapidly. Internalization of IgG1 HP was markedly inhibited by incubation at 4 degrees C, but not by incubation with a variety of metabolic inhibitors. Catabolism was inhibited by chloroquine and monensin (inhibitors of lysosomal acidification) and by cytochalasin (an inhibitor of microfilament polymerization). Binding to the surface of cells was not markedly inhibited by any agent tested. The capacity of cells to bind labeled IgG1 was markedly reduced by prior incubation in the presence of unlabeled IgG1. This reduction was in part due to the steric blockade of receptors caused by the avid, but reversible, binding of IgG1. In addition, IgG1 oligomers or HP (but not IgG1 monomers or dimers) also caused an irreversible reduction in the number of Fc receptors by a process analogous to receptor down-regulation, as observed in other receptor--ligand systems.

scholarly journals Pharmacodynamic Assessment Based on Mutant Prevention Concentrations of Fluoroquinolones To Prevent the Emergence of Resistant Mutants of Streptococcus pneumoniae

2007 ◽  
Vol 51 (11) ◽  
pp. 3810-3815 ◽  
Author(s):  
Tomoyuki Homma ◽  
Toshihiko Hori ◽  
Giichi Sugimori ◽  
Yoshinori Yamano
Keyword(s):  
Streptococcus Pneumoniae ◽  
Population Analysis ◽  
Pharmacodynamic Model ◽  
Time Curve ◽  
Concentration Time ◽  
Mutant Prevention Concentration ◽  
Resistant Mutants ◽  
The Relationship ◽  
Concentration Curves

ABSTRACT The objective of this study was to investigate the relationship between pharmacokinetic and pharmacodynamic parameters, on the basis of the mutant prevention concentration (MPC) concept, and the emergence of resistant mutants of Streptococcus pneumoniae to fluoroquinolone antibacterials. Some clinical isolates with various MIC and MPC values of moxifloxacin and levofloxacin were exposed under conditions simulating the time-concentration curves observed when moxifloxacin (400 or 80 mg, once a day) or levofloxacin (200 mg, twice a day) was orally administered by using an in vitro pharmacodynamic model. The decrease in susceptibility was evaluated by altering the population analysis profiles after moxifloxacin or levofloxacin treatment for 72 h. When the area under the concentration-time curve from 0 to 24 h (AUC0-24)/MPC and peak concentration (C max)/MPC were above 13.41 and 1.20, respectively, complete eradication occurred and no decrease in susceptibility was observed. On the other hand, when AUC0-24/MPC and C max/MPC were below 0.84 and 0.08, respectively, the susceptibility decreased. However, the time inside the mutant selective window and the time above the MPC did not show any correlation with the decrease in susceptibility. These results suggest that AUC0-24/MPC and C max/MPC are important parameters for predicting the emergence of resistant mutants and that higher values indicate greater effectiveness.

scholarly journals The binding and processing of monoclonal human IgG1 by cells of a human macrophage-like cell line (U937)

Blood ◽  
1983 ◽  
Vol 62 (3) ◽  
pp. 652-662
Author(s):  
RJ Kurlander ◽  
JE Gartrell
Keyword(s):  
Cell Line ◽  
Metabolic Inhibitors ◽  
Human Macrophage ◽  
Reversible Binding ◽  
In Vitro Binding ◽  
Irreversible Reduction ◽  
Vitro Binding ◽  
Human Igg1 ◽  
The Relationship

The goal of these experiments was to assess the relationship between the binding and processing of IgG by Fc-receptor-bearing cells. Cells of the U937 human macrophage-like cell line were incubated with 125I- labeled monomers, dimers, oligomers (composed of 2–4 IgG1 subunits), and HP (heavy polymers composed of 5 or more subunits per polymer) of monoclonal human IgG1 in vitro. Binding was assessed by spinning cells through a layer of phthalate oils. Internalization of IgG1 was assessed by quantitating residual binding to cells after surface-bound IgG was removed by a brief treatment with a solution containing 0.25 M acetic acid and 0.5 M sodium chloride. Catabolism was assessed by measuring the release of radioactive fragments of IgG1, which were not precipitated by 10% trichloroacetic acid. Unstimulated U937 bound about 10,000 molecules per cell of IgG1 monomer, with an equilibrium binding constant (Ka) of 5 X 10(8) M-1. After stimulation with a conditioned medium in vitro, binding per cell was increased 3–7--fold, and the Ka was decreased 2–4--fold. Both unstimulated and stimulated cells internalized and catabolized labeled IgG1 HP, but stimulated cells internalized and digested much more IgG1 HP per cell than unstimulated cells. Both monomers and dimers of IgG1 were internalized and degraded very slowly by stimulated cells, even though both preparations readily bound to cells. In contrast, oligomers and (to an even greater extent) IgG1 HP were internalized and degraded much more rapidly. Internalization of IgG1 HP was markedly inhibited by incubation at 4 degrees C, but not by incubation with a variety of metabolic inhibitors. Catabolism was inhibited by chloroquine and monensin (inhibitors of lysosomal acidification) and by cytochalasin (an inhibitor of microfilament polymerization). Binding to the surface of cells was not markedly inhibited by any agent tested. The capacity of cells to bind labeled IgG1 was markedly reduced by prior incubation in the presence of unlabeled IgG1. This reduction was in part due to the steric blockade of receptors caused by the avid, but reversible, binding of IgG1. In addition, IgG1 oligomers or HP (but not IgG1 monomers or dimers) also caused an irreversible reduction in the number of Fc receptors by a process analogous to receptor down-regulation, as observed in other receptor--ligand systems.

scholarly journals HGG-33. EXPLOITING METABOLIC DEFECTS WITH NAMPT INHIBITORS IN DIPG

2021 ◽  
Vol 23 (Supplement_1) ◽  
pp. i24-i24
Author(s):  
Ranjithmenon Muraleedharan ◽  
Collin Heer ◽  
Ranjini Sundam ◽  
Charles Brenner
Keyword(s):  
Cell Lines ◽  
Treatment Options ◽  
Pediatric Brain Tumors ◽  
Diffuse Intrinsic Pontine Glioma ◽  
Metabolic Inhibitors ◽  
Synthetic Lethal ◽  
Metabolic Defects ◽  
Metabolic Defect ◽  
H3k27m Mutation

Abstract Diffuse intrinsic pontine glioma (DIPG) are universally lethal pediatric brain tumors with limited treatment options. We recently performed synthetic lethal drug screen with a panel of DNA repair and metabolic inhibitors in vitro, in patient-derived DIPG cells and isogenic cell lines engineered to contain key DIPG-associated mutations. Nearly 80% of DIPGs harbor a recurrent H3K27M mutation in H3.3 (H3F3A) or H3.1 (HIST1H3B) histones. This has prompted us to consider H3K27M mutation-induced exploitable defects for a therapeutic gain. This screen identified synthetic lethal interactions between H3K27M mutations and the nicotinamide phosphoribosyl transferase (NAMPT) inhibitor, FK866. The association between H3K27M mutations and NAMPTi sensitivity was validated in follow-up studies using isogenic WT and H3K27M-mutant expressing pairs of human immortalized astrocytes and neural progenitor cells (NPCs). In addition, we tested the effects of FK866 in a panel of unique DIPG patient-derived tumor neurospheres in short-term viability assays. We found that FK866 induced depletion of NAD and exhibited toxicity towards H3K27M mutant DIPG cell lines with an IC50 of ~2.5 nM. These findings were consistent across three structurally unique NAMPT inhibitors. Finally, we found that inducible expression of mutant H3K27M results in a gradual, 50% decrease in cellular NAD levels over the course of five days, suggesting that H3K27M mutations may induce an inherent NAD metabolic defect that is exploited by NAMPTi’s. We now seek to understand the mechanistic basis for the observed metabolic defect and NAMPTi sensitivity associated with mutant H3K27M. In addition, we aim to identify specific NAMPTi and DNA damaging agent combinations which maximally exploit this H3K27M-associated NAD metabolic defects.

scholarly journals Synthesis and in vitro cytotoxicity of acetylated 3-fluoro, 4-fluoro and 3,4-difluoro analogs of D-glucosamine and D-galactosamine

2016 ◽  
Vol 12 ◽  
pp. 750-759 ◽  
Author(s):  
Štěpán Horník ◽  
Lucie Červenková Šťastná ◽  
Petra Cuřínová ◽  
Jan Sýkora ◽  
Kateřina Káňová ◽  
...  
Keyword(s):  
Prostate Cancer Cell ◽  
Cancer Cell Line ◽  
Prostate Cancer Cell Line ◽  
In Vitro Cytotoxicity ◽  
Metabolic Inhibitors ◽  
Human Prostate Cancer Cell ◽  
Complete Series ◽  
Derivatives Of ◽  
Complex Glycan

Background: Derivatives of D-glucosamine and D-galactosamine represent an important family of the cell surface glycan components and their fluorinated analogs found use as metabolic inhibitors of complex glycan biosynthesis, or as probes for the study of protein–carbohydrate interactions. This work is focused on the synthesis of acetylated 3-deoxy-3-fluoro, 4-deoxy-4-fluoro and 3,4-dideoxy-3,4-difluoro analogs of D-glucosamine and D-galactosamine via 1,6-anhydrohexopyranose chemistry. Moreover, the cytotoxicity of the target compounds towards selected cancer cells is determined. Results: Introduction of fluorine at C-3 was achieved by the reaction of 1,6-anhydro-2-azido-2-deoxy-4-O-benzyl-β-D-glucopyranose or its 4-fluoro analog with DAST. The retention of configuration in this reaction is discussed. Fluorine at C-4 was installed by the reaction of 1,6:2,3-dianhydro-β-D-talopyranose with DAST, or by fluoridolysis of 1,6:3,4-dianhydro-2-azido-β-D-galactopyranose with KHF2. The amino group was introduced and masked as an azide in the synthesis. The 1-O-deacetylated 3-fluoro and 4-fluoro analogs of acetylated D-galactosamine inhibited proliferation of the human prostate cancer cell line PC-3 more than cisplatin and 5-fluorouracil (IC50 28 ± 3 μM and 54 ± 5 μM, respectively). Conclusion: A complete series of acetylated 3-fluoro, 4-fluoro and 3,4-difluoro analogs of D-glucosamine and D-galactosamine is now accessible by 1,6-anhydrohexopyranose chemistry. Intermediate fluorinated 1,6-anhydro-2-azido-hexopyranoses have potential as synthons in oligosaccharide assembly.

Large chromosomal inversion correlated with spectinomycin resistance in Lactococcus lactis subsp. lactis bv. diacetylactis S50

2008 ◽  
Vol 54 (2) ◽  
pp. 143-149 ◽  
Author(s):  
Milan Kojic ◽  
Branko Jovcic ◽  
Jelena Begovic ◽  
Djordje Fira ◽  
Ljubisa Topisirovic
Keyword(s):  
Lactococcus Lactis ◽  
Chromosomal Inversion ◽  
High Concentration ◽  
Lactococcus Lactis Subsp ◽  
Ribosomal Operons ◽  
One Step ◽  
Resistant Mutants ◽  
High Concentrations ◽  
Derivatives Of

A large chromosomal inversion that confers resistance to high concentrations of the antibiotic spectinomycin in Lactococcus lactis subsp. lactis bv. diacetylactis S50 was identified by pulsed field gel electrophoresis. The same type of inversion was identified in 4 independent experiments and in 4 different derivatives of strain S50, indicating the same position and the same mechanism of recombination as a response to antibiotic selective pressure in all derivatives. An analysis of ribosomal operons in strain S50 and mutants revealed that ribosomal operons are not endpoints of the recombination. Spectinomycin-resistant mutants appeared in a population of S50 derivatives at a high frequency of 2 × 10−7. These spectinomycin-resistant mutants were not able to compete successfully with the wild-type strain during 25 generations (48 h) of co-culture in vitro, indicating that inversion had a significant fitness cost. Results demonstrate that as a mechanism of genome plasticity, inversion can be directly involved in one-step development of the adaptation to a high concentration of spectinomycin.

scholarly journals Furazolidone resistance in Salmonella gallinarum: the relationship between in vitro and in vivo determinations of resistance

1981 ◽  
Vol 87 (1) ◽  
pp. 71-79 ◽  
Author(s):  
H. Williams Smith ◽  
J. F. Tucker ◽  
M. Lovell
Keyword(s):  
Minimum Inhibitory Concentration ◽  
Inhibitory Concentration ◽  
Salmonella Gallinarum ◽  
Experimental Infections ◽  
Resistant Strains ◽  
Resistant Mutants ◽  
The 1950S ◽  
The Relationship

SummaryOf 22 strains of Salmonella gallinarum isolated from recent outbreaks of infection in poultry in Greece (15), Amman (3), Kenya (2), Lebanon (1) and Yemen (1), 20 were more resistant to furazolidone in vitro than 6 strains that had been isolated in the U.K. in the 1950s; the minimum inhibitory concentration of furazolidone was approximately 0·3 μg/ml for the sensitive strains and 1·3 or 2·5 μg/ml for the more resistant strains.Furazolidone given continuously in the food did not control experimental infections in chickens caused by most of the strains that had been classed as more resistant by the in vitro tests. Chloramphenicol, trimethoprim and sulphadiazine or mixtures of the latter two were the best antibiotics for treating these infections, but they were less satisfactory than furazolidone for treating infections caused by the furazolidone-sensitive strains.As a group, the furazolidone-resistant strains and furazolidone-resistant mutants of one of the sensitive strains were less virulent for chickens than the sensitive strains.

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