Cryptic speciation in Fusarium subglutinans

Fusarium isolates that form part of the Gibberella fujikuroi species complex have been classified using either a morphological, biological, or phylogenetic species concept. Problems with the taxonomy of Fusarium species in this complex are mostly experienced when the morphological and biological species concepts are applied. The most consistent identifications are obtained with the phylogenetic species concept. Results from recent studies have presented an example of discordance between the biological and phylogenetic species concepts, where a group of F. subglutinans sensu stricto isolates, i.e., isolates belonging to mating population E of the G. fujikuroi complex, could be sub-divided into more than one phylogenetic lineage. The aim of this study was to determine whether this sub-division represented species divergence or intraspecific diversity in F. subglutinans. For this purpose, we included 29 F. subglutinans isolates belonging to the E-mating population that were collected from either maize or teosinte, from a wide geographic range. DNA sequence data for six nuclear regions in each of these isolates were obtained and used in phylogenetic concordance analyses. These analyses revealed the presence of two major groups representing cryptic species in F. subglutinans. These cryptic species were further sub-divided into a number of smaller groups that appear to be reproductively isolated in nature. This suggests not only that the existing F. subglutinans populations are in the process of divergence, but also that each of the resulting lineages are undergoing separation into distinct taxa. These divergences did not appear to be linked to geographic origin, host, or phenotypic characters such as morphology.     

Effect of abscisic acid on gibberellin biosynthesis in Fusarium moniliforme

The effect of ABA on gibberellin biosynthesis by Fusarium moniliformecultured on inorganic medium was studied. ABA had no apparenteffect on gibberellin levels. (Received October 24, 1969; )     

Description of Gibberella sacchari and neotypification of its anamorph Fusarium sacchari

We described the teleomorph of Fusarium sacchari as Gibberella sacchari, sp. nov. This species can be separated from other species of Gibberella on the basis of the longer, narrower ascospores found in G. sacchari and by sexual cross fertility. Female-fertile mating type tester strains were developed that can be used for making sexual crosses with this heterothallic fungus under laboratory conditions. The anamorph, Fusarium sacchari, was neotypified.     

Occurrence ofFusarium sacchari var.subglutinans and its mycotoxin production ability in broiler feed

Spores ofFusarium sacchari var.subglutinans isolated from broiler feed BR1 were obtained at an average concentration of 1.5/mg in 25% of tested samples. The spore concentration was increased from 1 to 100/mg of solid substrate (BR2; biscuit) or to 1/nL of Sabouraud broth after 3 weeks of cultivation. Mycotoxin analyses of these three substrates showed negative reactions for T-2 toxin and zearalenone but a positive reaction for deoxynivalenol (DON) which was found in concentrations of 5 ppm in Sabouraud broth, 50 ppm in BR2 and 220 ppm in biscuit. Therefore, ourF. sacchari isolate appeared to be a DON producer.     

Transmission ratio distortion in an interspecific cross between Fusarium circinatum and Fusarium subglutinans

Previously, an interspecific cross between Fusarium circinatum and Fusarium subglutinans was used to generate a genetic linkage map. A ca. 55 % of genotyped markers displayed transmission ratio distortion (TRD) that demonstrated a genome-wide distribution. The working hypothesis for this study was that TRD would be non-randomly distributed throughout the genetic linkage map. This would indicate the presence of distorting loci. Using a genome-wide threshold of α = 0.01, 79 markers displaying TRD were distributed on all 12 linkage groups (LGs). Eleven putative transmission ratio distortion loci (TRDLs), spanning eight LGs, were identified in regions containing three or more adjacent markers displaying distortion. No epistatic interactions were observed between these TRDLs. Thus, it is uncertain whether the genome-wide TRD was due to linkage between markers and genomic regions causing distortion. The parental origins of markers followed a non-random distribution throughout the linkage map, with LGs containing stretches of markers originating from only one parent. Thus, due to the nature of the interspecific cross, the current hypothesis to explain these observations is that the observed genome-wide segregation was caused by the high level of genomic divergence between the parental isolates. Therefore, homologous chromosomes do not align properly during meiosis, resulting in aberrant transmission of markers. This also explains previous observations of the preferential transmission of F. subglutinans alleles to the F₁ progeny.     

Characterization of gibberellin producing strains of Fusarium moniliforme based on DNA polymorphism

The gibberellins are one of the major groups of growth promoting hormones and are secondary metabolites of the fungus Fusarium moniliforme (Perfect stage: Gibberella fujikuroi). Sixteen strains of Fusarium from different geographical regions and different hosts were analysed for their ability to produce gibberellins (GA) and for genetic relatedness by random amplified polymorphic DNA (RAPD). Range of gibberellin production varied between 28.9 to 600.0 mg g^-1 dry weight of mycelium in different strains of Fusarium. RAPD analysis showed completely different pattern between high, moderate and low producing strains. High producers formed nearly identical RAPD patterns, whereas the low and moderate producers gave heterologous amplification patterns. Since Fusarium pallidoroseum was in another group, it was possible to distinguish between different species of the genus Fusarium by RAPD. These investigations may find an application in the diagnosis of unknown Fusarium species and in distinguishing isolates of Gibberella fujikuroi within the section of Liseola. This revised version was published online in June 2006 with corrections to the Cover Date.     

Gibberellin biosynthesis and its regulation

The GAs (gibberellins) comprise a large group of diterpenoid carboxylic acids that are ubiquitous in higher plants, in which certain members function as endogenous growth regulators, promoting organ expansion and developmental changes. These compounds are also produced by some species of lower plants, fungi and bacteria, although, in contrast to higher plants, the function of GAs in these organisms has only recently been investigated and is still unclear. In higher plants, GAs are synthesized by the action of terpene cyclases, cytochrome P450 mono-oxygenases and 2-oxoglutarate-dependent dioxygenases localized, respectively, in plastids, the endomembrane system and the cytosol. The concentration of biologically active GAs at their sites of action is tightly regulated and is moderated by numerous developmental and environmental cues. Recent research has focused on regulatory mechanisms, acting primarily on expression of the genes that encode the dioxygenases involved in biosynthesis and deactivation. The present review discusses the current state of knowledge on GA metabolism with particular emphasis on regulation, including the complex mechanisms for the maintenance of GA homoeostasis.     

Gibberellin homeostasis in tobacco is regulated by gibberellin metabolism genes with different gibberellin sensitivity

Gibberellins are phytohormones that regulate growth and development of plants. Gibberellin homeostasis is maintained by feedback regulation of gibberellin metabolism genes. To understand this regulation, we manipulated the gibberellin pathway in tobacco and studied its effects on the morphological phenotype, gibberellin levels and the expression of endogenous gibberellin metabolism genes. The overexpression of a gibberellin 3-oxidase (biosynthesis gene) in tobacco (3ox-OE) induced slight variations in phenotype and active GA(1) levels, but we also found an increase in GA(8) levels (GA(1) inactivation product) and a conspicuous induction of gibberellin 2-oxidases (catabolism genes; NtGA2ox3 and -5), suggesting an important role for these particular genes in the control of gibberellin homeostasis. The effect of simultaneous overexpression of two biosynthesis genes, a gibberellin 3-oxidase and a gibberellin 20-oxidase (20ox/3ox-OE), on phenotype and gibberellin content suggests that gibberellin 3-oxidases are non-limiting enzymes in tobacco, even in a 20ox-OE background. Moreover, the expression analysis of gibberellin metabolism genes in transgenic plants (3ox-OE, 20ox-OE and hybrid 3ox/20ox-OE), and in response to application of different GA(1) concentrations, showed genes with different gibberellin sensitivity. Gibberellin biosynthesis genes (NtGA20ox1 and NtGA3ox1) are negatively feedback regulated mainly by high gibberellin levels. In contrast, gibberellin catabolism genes which are subject to positive feedback regulation are sensitive to high (NtGA2ox1) or to low (NtGA2ox3 and -5) gibberellin concentrations. These two last GA2ox genes seem to play a predominant role in gibberellin homeostasis under mild gibberellin variations, but not under large gibberellin changes, where the biosynthesis genes GA20ox and GA3ox may be more important.     

Complete genetic linkage maps from an interspecific cross between Fusarium circinatum and Fusarium subglutinans

The Gibberella fujikuroi complex includes many plant pathogens of agricultural crops and trees, all of which have anamorphs assigned to the genus Fusarium. In this study, an interspecific hybrid cross between Gibberella circinata and Gibberella subglutinans was used to compile a genetic linkage map. A framework map was constructed using a total of 578 AFLP markers together with the mating type (MAT-1 and MAT-2) genes and the histone (H3) gene. Twelve major linkage groups were identified (n=12). Fifty percent of the markers showed significant deviation from the expected 1:1 transmission ratio in a haploid F(1) cross (P <0.05). The transmission of the markers on the linkage map was biased towards alleles of the G. subglutinans parent, with an estimated 60% of the genome of F(1) individuals contributed by this parent. This map will serve as a powerful tool to study the genetic architecture of interspecific differentiation and pathogenicity in the two parental genomes.     

Fusarium temperatum sp. nov. from maize, an emergent species closely related to Fusarium subglutinans

A large number of Fusarium isolates closely related to F. subglutinans were collected from maize in Belgium. We used a robust polyphasic approach to describe a new biological species, Fusarium temperatum, within the Gibberella fujikuroi species complex. F. temperatum can be distinguished from F. subglutinans and from other Fusarium species within the Gibberella fujikuroi species complex with AFLP fingerprint profile, differences in the translation elongation factor 1-α and β-tubulin DNA sequence and interspecies mating compatibility analyses. Intraspecies mating compatibility suggests that sexual reproduction might be common for field isolates of F. temperatum, and reliable female fertile mating population tester strains were proposed for this heterothallic species.     

Influence of an inhibitor of gibberellin biosynthesis, paclobutrazol, on apple seed gibberellin content

Tissue-culture-propagated own-rooted cv. Spartan apple trees (Malus domestica Borkh.) planted in 1979 were treated in 1983 and 1985 via a soil-line trunk drench with the plant growth retardant paclobutrazol [(2RS, 3RS)-1-(4-chlorophenyl)-4.4-dimethyl-2-(1,2, 4-triazol-1-yl) pentan-3-ol]. Seeds of immature fruits from untreated and treated trees were sampled in 1989 ca 75 days after full bloom. After seeds were freeze-dried, gibberellins (GAs) were extracted, purified and fractionated via C₁₈ reversed-phase high-performance liquid chromatography (HPLC). Gibberellins A₁, A₃, A₄, A₇, A₈, A₉, A₁₅, A₁₇, A₁₉, A₂₀, A₂₄, A₃₄, A₃₅, A₄₄, A₅₁, A₅₃, A₅₄, A₆₁, A₆₂, A₆₃ and A₆₈ were identified by using C₁₈ HPLC, gas chromatography-selected ion monitoring and Kovats retention indices. Eight of the GAs identified were also quantified by using deuterated internal standards. The paclobutrazol applications caused a 55% reduction of vegetative shoot elongation in 1989, but both treated and untreated trees had developed a biennial bearing pattern by that time (heavy bloom or “on year’in 1989). Levels of early 13-hydroxylation pathway GAs, viz. GA₅₃, GA₁₉, GA₂₀, GA₁ and also GA₃, were not altered by treatment. However, GA₄, GA₇ and GA₉ were increased 13.4, 6.5 and 3.8 times, respectively, in seeds of fruit from treated compared to untreated trees.     

Cytochromes P450 in gibberellin biosynthesis

The gibberellins (GAs) are an important class of plant growth regulators that are active in many aspects of plant growth and development. GAs are synthesized by a complex pathway involving three enzyme classes spanning different subcellular compartments. One of these enzyme classes is the cytochrome P450s which catalyze a number of oxidation steps in the middle part of the pathway. Mutants in these cytochrome P450-mediated steps in a number of species have been crucial in isolating the genes encoding these enzymes and have also played an important role in understanding GA physiology. GAs are also synthesized by fungi, in a biosynthesis pathway largely catalyzed by cytochrome P450s. The fungal pathway appears to have evolved independently to that of higher plants.

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Isolation of beauvericin as an insect toxin from Fusarium semitectum and Fusarium moniliforme var. subglutinans

Nonpolar methylene chloride-soluble extracts from the mycelia of Fusarium semitectum and Fusarium moniliforme var. subglutinans were toxic to Colorado potato beetles. The major toxic metabolite was isolated and found to be the cyclodepsipeptide, beauvericin. This is the first report of the isolation of beauvericin from the genus Fusarium.     

Microbial transformation of ginsenosides Rb1, Rb3 and Rc by Fusarium sacchari

Aims: This study examined the transformation pathways of ginsenosides G‐Rb₁, G‐Rb₃, and G‐Rc by the fungus Fusarium sacchari. Methods and Results: Ginsenosides G‐Rb₁, G‐Rb₃ and G‐Rc were isolated from leaves of Radix notoginseng, and their structural identification was confirmed using NMR. Transformation of G‐Rb₁, G‐Rb₃ and G‐Rc by Fusarium sacchari was respectively experimented. Kinetic evolutions of G‐Rb₁, G‐Rb₃ and G‐Rc and their metabolites during the cell incubation were monitored by HPLC analysis. High‐performance liquid chromatography (HPLC) was used for monitoring the transformation kinetics of bioactive compounds during F. sacchari metabolism. Conclusions: Ginsenoside C‐K was transformed by F. sacchari from G‐Rb₁ via G‐Rd or via G‐F₂, or from G‐Rb₁ via firstly Rd and then G‐F₂, and C‐Mx was transformed by F. sacchari or directly from Rb₃, or from Rb₃ via Gy‐IX, while G‐Mc was transformed by F. sacchari directly from G‐Rc. Furthermore, C‐K could be also formed from G‐Rc via notoginsenoside Fe (N‐Fe). Significance and Impact of the Study: The results showed an important practical application in the preparation of ginsenoside C‐K. As our precious research indicated C‐K possessed much more antitumor activities than C‐Mx and G‐Mc, so according to the transformation pathways proposed by this work, the production of antitumor compound C‐K may be performed by biotransformation of G‐Rb₁ previously isolated from PNLS.     

Gibberellin biosynthesis mutations and root development in pea

Dwarf mutants of pea (Pisum sativum), with impaired gibberellin (GA) biosynthesis in the shoot, were studied to determine whether the roots of these genotypes had altered elongation and GA levels. Mutations na, lh-2, and ls-1 reduced GA levels in root tips and taproot elongation, although in lh-2 and ls-1 roots the reduction in elongation was small (less than 15%). The na mutation reduced taproot length by about 50%. The roots of na plants elongated in response to applied GA(1) and recombining na with mutation sln (which blocks GA catabolism) increased GA(1) levels in root tips and completely restored normal root development. In shoots, Mendel's le-1 mutation impairs the 3beta-hydroxylation of GA(20) to the bioactive GA(1), resulting in dwarfism. However, GA(1) and GA(20) levels were normal in le-1 roots, as was root development. The null mutation le-2 also did not reduce root GA levels or elongation. The results support the theory that GAs are important for normal root elongation in pea, and indicate that a 3beta-hydroxylase gene other than LE operates in pea roots.     

Sites of gibberellin biosynthesis in pea seedlings

Potential sites of gibberellin biosynthesis in 10-day-old `Alaska' pea (Pisum sativum L.) seedlings were investigated using a cell-free ezyme system capable of incorporating [¹⁴C]-mevalonic acid into ent-kaurene. In peas, ent-kaurene is assumed to be a committed intermediate in the gibberellin biosynthetic pathway. Comparative results from enzyme assays using extracts from shoot tips, leaf blades, internodes, and root tips indicate that the highest capacity for ent-kaurene (and presumably gibberellin) synthesis is in those tissues with the greatest potential for growth. The highest rates were obtained with extracts prepared from the fifth (youngest) internode, the fourth (youngest) expanded leaf, and the shoot tip itself. This report represents the first direct evidence that the enzymes responsible for early stages in gibberellin biosynthesis occur in internode tissues with potential for rapid elongation.     

The genes for gibberellin biosynthesis in wheat

The gibberellin biosynthesis pathway is well defined in Arabidopsis and features seven key enzymes including ent-copalyl diphosphate synthase (CPS), ent-kaurene synthase (KS), ent-kaurene oxidase (KO), ent-kaurenoic acid oxidase (KAO), GA 20-oxidase, GA 3-oxidase, and GA 2-oxidase. The Arabidopsis genes were used to identify their counterparts in wheat and the TaCPS, TaKS, TaKO, and TaKAO genes were cloned from Chinese Spring wheat. In order to determine their chromosome locations, expression patterns and feedback regulations, three TaCPS genes, three TaKS genes, three TaKO genes, and three TaKAO genes were cloned from Chinese Spring wheat. They are mainly located on chromosomes 7A, 7B, 7D and 2A, 2B and 2D. The expression patterns of TaCPS, TaKS, TaKO, and TaKAO genes in wheat leaves, young spikes, peduncles, the third and forth internodes were investigated using quantitative PCR. The results showed that all the genes were constitutively expressed in wheat, but their relative expression levels varied in different tissues. They were mainly transcribed in stems, secondly in leaves and spikes, and the least in peduncles. Feedback regulation of the TaCPS, TaKS, TaKO, and TaKAO genes was not evident. These results indicate that all the genes and their homologs may play important roles in the developmental processes of wheat, but each of the homologs may function differently in different tissues or during different developmental stages.