Study of Phylogenetic Relationships Among Fusarium oxysporum f. sp. dianthi Isolates: Confirmation of Intrarace Diversity and Development of a Practical Tool for Simple Population Analyses.

Fusarium wilt, caused by Fusarium oxysporum f. sp. dianthi, is the most important disease of carnation worldwide. Knowing the diversity of the F. oxysporum f. sp. dianthi population present in a carnation growing area is a key component of preventing dramatic losses in production. Sequence analyses of partial ß-tubulin, translation elongation factor 1α genes, and the full-length ribosomal DNA intergenic spacer (IGS) were conducted to resolve phylogenetic relationships in a wide collection of Spanish F. oxysporum f. sp. dianthi isolates, along with some representatives from Italy. We found that, among the three different gene regions, the IGS sequence was the best choice to resolve phylogenetic relationships among F. oxysporum f. sp. dianthi isolates. The phylogenetic tree generated with the complete IGS region was the only one showing a clear clustering of isolates according to the molecular group (virulence grouping) and the vegetative compatibility group. In order to develop a more practical tool based on a shorter DNA sequence to quickly analyze diversity in F. oxysporum f. sp. dianthi populations, we examined IGS nucleotide alignments and identified a region of approximately 300 bp that accumulates enough "informative" changes to resolve intraspecific relationships and determine pathogenic variants in F. oxysporum f. sp. dianthi. Moreover, the "condensed" alignment of this short IGS region showing only the informative positions revealed the existence of virulence group-discriminating positions. In addition to clarifying the phylogenetic relationships among F. oxysporum f. sp. dianthi isolates of the recently described race groups by using multigene genealogies, we have developed simple tools for the phylogenetic analyses of F. oxysporum f. sp. dianthi populations and the determination of the molecular group of uncharacterized F. oxysporum f. sp. dianthi isolates.

Comparative kinetic behaviour and regulation by fructose-1,6-bisphosphate and ATP of pyruvate kinase from erythrocytes, reticulocytes and bone marrow cells.

1. Kinetic and regulatory properties of pyruvate kinase have been studied in haemolysates of erythrocytic populations from blood and bone marrow of rats. 2. Pyruvate kinase from normal rat erythrocytes showed sigmoidal kinetics vs phosphoenolpyruvate. In contrast, the enzyme from reticulocytes and erythroid-rich bone marrow cells behaved as hyperbolic. 3. The enzyme activities were always inhibited by ATP. Activation by fructose-1,6-bisphosphate was only observed in erythrocytes. 4. These kinetic differences suggest changes in pyruvate kinase isozymes in cells of the erythrocytic line of rats.

A comparative kinetics and regulatory study of pyruvate kinase from rat erythrocytes, reticulocytes and bone marrow cells.

The kinetics of pyruvate kinase (PK) at various phosphoenol pyruvate (PEP) concentrations, has been studied in cells of the erythrocytic line. The enzyme from erythrocytes shows positive cooperativity, it behaves as michaelian in reticulocytes and shows negative cooperativity in bone marrow cells. ATP exerts an inhibitory effect in all cases. The activator effect of fructose 1,6-bisphosphate (FBP) was found only in erythrocytes.

[Glycolytic regulatory enzymes in erythrocytes, reticulocytes and immature bone marrow cells of rats]. / Estudio de las enzimas reguladoras glucolíticas en eritrocitos, reticulocitos y células inmaduras de méduloa ósea de rata.

Kinetic characteristics of glycolytic kinases in cells of the erythrocytic line have been studied in rats. Erythrocyte Hexokinase (HK) shows low Km, while in reticulocytes a high Km enzyme is also found. The low Km HK is inhibited by higher than physiological concentrations of 2,3-bisphosphoglycerate. The ATP inhibition of 6-phosphofructokinase (PFK) from erythrocytes and reticulocytes, and the deinhibitory effect shown by AMP and cyclic-AMP, are lower in situ than in vitro. The sigmoidal fructose-6-phosphate saturation curve for PFK is observed in vitro but not in situ. Under this approach the enzyme shows hyperbolic kinetics, lower response to positive allosteric effectors and a decrease in activity. An antagonistic effect is shown by cyclic-AMP and cyclic-GMP on the in vitro sigmoidal behaviour of PFK. Phosphoglycerate kinase (PGK) is an hyperbolic enzyme, with similar Km values, in erythrocytes, reticulocytes and bone marrow cells. Pyruvate kinase shows sigmoidal kinetics and activation by fructose-1,6-bisphosphate (FBP) in erythrocytes; a non rectangular hyperbolic kinetics with negative cooperativity, lack of response to FBP and inhibition by ATP and L-alanine, was observed in the bone marrow enzyme.