Biology and epidemiology of Fusarium circinatum

Fusarium circinatum is the causal agent of the disease known as pitch canker of pine. The fungus causes resinous cankers on stems and branches of mature trees, dieback of female flowers and cones, as well as root rot and pre- and post emergence damping off of seedlings. Little is known regarding the epidemiology and biology of F. circinatum in South African pine seedling nurseries, where the fungus has been causing major economic losses since its introduction into the country in the early 1990s. The objectives of this study were, therefore, to study the infection biology and epidemiology of F. circinatum on pine seedlings, the organism’s saprophytic biology. I also considered approaches to rapidly diagnose plants infected by F. circinatum and its relatedness to other species. Much research has been done on the pitch canker disease and the causal agent F. circinatum. Chapter 1 of this thesis aimed to summarize the available knowledge on the pitch canker fungus and its biology, ecology and epidemiology. Trials to screen for resistance of Pinus spp. to the pitch canker fungus have been conducted by many research groups and also by industries that rely on Pinus spp. for pulp and wood production. In Chapter 2, parameters for such trials, including optimal wounding methods, spore concentrations, plant physiological considerations and time elapsed between wounding and inoculation, were investigated. Temperature and ambient humidity are considered important factors in plant disease epidemiology. The effect of these factors on pitch canker epidemics has not yet been studied. In Chapter 3, a survey of disease incidence in pine nurseries from different geographic areas in South Africa with different climates is presented. This was done by correlating disease incidence data from the nurseries with temperature and humidity measurements. The saprophytic biology of a plant pathogen is of great importance in its epidemiology. The extent of any plant pathogen’s saprophytic survival determines the initial inoculum levels at the onset of an epidemic. In Chapter 4, I investigated the saprophytic growth and survival of F. circinatum in various substrates, temperatures and moisture regimes. Fusarium circinatum is morphologically similar to fungi referred to as Fusarium subglutinans sensu lato. Distinguishing F. circinatum from other species in this group has in the past required pathogenicity tests and sexual crosses, which are labour intensive and time consuming. In Chapter 5, a molecular diagnostic technique, based on real-time PCR, with which identification of Fusarium spp. commonly occurring in South African nurseries is possible, was developed. Fusarium circinatum and other fungi referred to as F. subglutinans sensu lato are members of the Gibberella fujikuroi species complex. Molecular taxonomic studies have shown that F. subflutinans sensu lato is a polyphyletic taxon. The objective of the study presented in Chapter 6 was to resolve the taxon phylogenetically with the use of multiple loci. The studies in the individual chapters of this thesis present individual aspects of the biology, ecology, epidemiology and molecular ecology of Fusarium circinatum. Each chapter represents an independent entity and consequently repetition between chapters has been unavoidable.Dissertation (MSc)--University of Pretoria, 2007.Microbiology and Plant PathologyUnrestricte

Biosynthesis of polyphenols in Norway spruce as a defense strategy against attack by the bark beetle associated fungus Ceratocystis polonica

The family Pinaceae (order Coniferales) includes many environmentally and economically important softwood tree genera. The most important Pinaceae species endemic to Europe is the Norway spruce tree (Picea abies). This tree species is frequently subject to severe attacks by the scolytid bark beetle Ips typographus and its microbial associate, the blue stain fungus, Ceratocystis polonica. Spruce trees have structural and chemical defense strategies against invasion by the beetle-fungus complex, including cells with autofluorescent inclusion bodies containing aromatic compounds such as stilbene glucosides and flavan-3-ols. The goal of our research was the elucidation of the pathways leading to the formation of stilbene glucosides and flavan-3ols in spruce and to determine their role in the defense of this tree against infection by the bark beetle associate C. polonica. Research in this thesis demonstrated that the biosynthesis of the major tetrahydroxystilbenes in spruce, astringin and isorhapontin, proceeds via resveratrol and is enhanced by fungal infection. Furthermore, it was also shown that flavan-3-ol biosynthesis, which is driven by three newly characterized leuconanthocyanidin reductase enzymes, is up-regulated in response to infection by C. polonica. However, although C. polonica is slightly inhibited in its growth by stilbenes and flavan-3-ols, the fungus has adaptive mechanisms to overcome the toxic effects of phenolic spruce defenses. In this context we showed that C. polonica can detoxify stilbene glycosides by biotransformation into stilbene lactones, aglycones as well as stilbene dimers. By comparing in vitro biotransformation rates of C. polonica stains which differed in their virulence against Norway spruce, we could show that the polyphenol detoxification strategy and velocity has a significant effect on the ecological fitness of this fungus during spruce infection and colonization

Roles of plant volatiles in defence against microbial pathogens and microbial exploitation of volatiles

Plants emit a large variety of volatile organic compounds during infection by pathogenic microbes, including terpenes, aromatics, nitrogen‐containing compounds, and fatty acid derivatives, as well as the volatile plant hormones, methyl jasmonate, and methyl salicylate. Given the general antimicrobial activity of plant volatiles and the timing of emission following infection, these compounds have often been assumed to function in defence against pathogens without much solid evidence. In this review, we critically evaluate current knowledge on the toxicity of volatiles to fungi, bacteria, and viruses and their role in plant resistance as well as how they act to induce systemic resistance in uninfected parts of the plant and in neighbouring plants. We also discuss how microbes can detoxify plant volatiles and exploit them as nutrients, attractants for insect vectors, and inducers of volatile emissions, which stimulate immune responses that make plants more susceptible to infection. Although much more is known about plant volatile–herbivore interactions, knowledge of volatile–microbe interactions is growing and it may eventually be possible to harness plant volatiles to reduce disease in agriculture and forestry. Future research in this field can be facilitated by making use of the analytical and molecular tools generated by the prolific research on plant–herbivore interactions.A. H. and T. A. are funded by South African National Research Council Incentive Funds (2019) and the University of Pretoria, and J. G. is funded by the Max Planck Society.https://wileyonlinelibrary.com/journal/pce2020-10-01hj2020BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologyZoology and Entomolog

Fungal associates of the tree-killing bark beetle, Ips typographus, vary in virulence, ability to degrade conifer phenolics and influence bark beetle tunneling behavior

The bark beetle Ips typographus carries numerous fungi that could be assisting the beetle in colonizing live Norway spruce (Picea abies) trees. Phenolic defenses in spruce phloem are degraded by the beetle's major tree-killing fungus Endoconidiophora polonica, but it is unknown if other beetle associates can also catabolize these compounds. We compared the ability of five fungi commonly associated with I. typographus to degrade phenolic compounds in Norway spruce phloem. Grosmannia penicillata and Grosmannia europhioides were able to degrade stilbenes and flavonoids faster than E. polonica and grow on minimal growth medium with spruce bark constituents as the only nutrients. Furthermore, beetles avoided medium amended with phenolics but marginally preferred medium colonized by fungi. Taken together our results show that different bark beetle-associated fungi have complementary roles in degrading host metabolites and thus might improve this insect's persistence in well defended host tissues.acceptedVersio

Rust Infection of Black Poplar Trees Reduces Photosynthesis but Does Not Affect Isoprene Biosynthesis or Emission

Poplar (Populus spp.) trees are widely distributed and play an important role in ecological communities and in forestry. Moreover, by releasing high amounts of isoprene, these trees impact global atmospheric chemistry. One of the most devastating diseases for poplar is leaf rust, caused by fungi of the genus Melampsora. Despite the wide distribution of these biotrophic pathogens, very little is known about their effects on isoprene biosynthesis and emission. We therefore infected black poplar (P. nigra) trees with the rust fungus M. larici-populina and monitored isoprene emission and other physiological parameters over the course of infection to determine the underlying mechanisms. We found an immediate and persistent decrease in photosynthesis during infection, presumably caused by decreased stomatal conductance mediated by increased ABA levels. At the same time, isoprene emission remained stable during the time course of infection, consistent with the stability of its biosynthesis. There was no detectable change in the levels of intermediates or gene transcripts of the methylerythritol 4-phosphate (MEP) pathway in infected compared to control leaves. Rust infection thus does not affect isoprene emission, but may still influence the atmosphere via decreased fixation of CO2

Continuous replanting could degrade soil health in short-rotation plantation forestry

PURPOSE OF REVIEW : Continuous replanting of land with the same or similar plant species can result in the accumulation of harmful soil microbes, which can lead to crop failure. In this review, we explore the influence of constant replanting on the health of short-rotation forestry soil, focusing on the accumulation of deleterious microbes and the decline of beneficial microbes. We also suggest possible practical solutions to address this problem and consider future research that could be conducted to better understand and reduce the build-up of deleterious soil microbes in short-rotation forestry soil. RECENT FINDINGS : Compelling evidence that continuous replanting of the same tree species in short-rotation plantation forestry might contribute to the build-up of deleterious soil microbes is still lacking. However, our assessment of existing soil microbiome data from global short-rotation plantation environments suggests a high risk of an accumulation of harmful microbes and a loss of beneficial microbes in plots that were continually replanted with the same tree species. Based on this evidence, and that from agriculture, we propose further research to acquire a better understanding of the build-up of harmful soil microbes in short-rotation plantation forestry, and suggest crop rotation and intercropping strategies to avoid this malady in the future. SUMMARY : The accumulation of microbes detrimental to plantation trees and the decline of microbes beneficial to these trees are realistic risks when plantations are continually replanted with the same tree species. Extensive research is necessary to evaluate the impact of short continuous planting rotations on the biodiversity of soil microbes in plantations and to develop strategies that would alleviate the build-up of detrimental microbes.Open access funding provided by University of Pretoria. This work was financially supported by the University of Pretoria and the Tree Protection Cooperative Programme (TPCP), Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria, South Africa.https://link.springer.com/journal/40725hj2023BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant PathologyPlant Production and Soil ScienceZoology and Entomolog

Phenolic compound degradation by Pseudomonas syringae phylogroup 2 strains

It has recently been shown that Pseudomonas syringae strains pathogenic to woody hosts belonging to phylogroup (PG) 2 lack phenolic compound degradation pathways such as the beta-ketoadipate and protocatechuate pathways. The aim of this study was to analyse a selection of P. syringae PG 2 genomes, including those used previously to determine if they had other phenolic compound degradation pathways and to determine whether or not they were functional. Twenty-one publicly available genomes of PG 2 strains were analyzed. These strains had previously been isolated from both woody and herbaceous hosts. Phenolic degradation enzymes were present in 5 (23%) of the strains analysed, originating from both woody and herbaceous hosts. Hypothetical pathways were proposed to determine if catechol, anthranilate and benzoic acid were degraded by these strains. Both spectrophotometric and HPLC were used to determine phenolic compound degradation. The five strains with phenolic degradation enzymes were able to metabolize catechol, and HRI-W 7924 and MAFF 301072 could also metabolize anthranilate and benzoate, respectively. The study showed that even though some PG 2 strains lack the beta-ketoadipate and protocatechuate pathways, they still have phenolic compound degrading enzymes that may play a role in virulence.The Horticultural Knowledge Group (HORTGRO) and National Research Foundation (NRF).https://link.springer.com/journal/421612019-07-01am2019BiochemistryForestry and Agricultural Biotechnology Institute (FABI)Microbiology and Plant PathologyZoology and Entomolog

Flavanone-3-hydroxylase plays an important role in the biosynthesis of spruce phenolic defenses against bark beetles and their fungal associates

Conifer forests worldwide are becoming increasingly vulnerable to attacks by bark beetles and their fungal associates due to the effects of global warming. Attack by the bark beetle Ips typographus and the blue-stain fungus it vectors (Endoconidiophora polonica) on Norway spruce (Picea abies) is well known to induce increased production of terpene oleoresin and polyphenolic compounds. However, it is not clear whether specific compounds are important in resisting attack. In this study, we observed a significant increase in dihydroflavonol and flavan-3-ol content after inoculating Norway spruce with the bark beetle vectored fungus. A bioassay revealed that the dihydroflavonol taxifolin and the flavan-3-ol catechin negatively affected both I. typographus and E. polonica. The biosynthesis of flavan-3-ols is well studied in Norway spruce, but little is known about dihydroflavonol formation in this species. A flavanone-3-hydroxylase (F3H) was identified that catalyzed the conversion of eriodictyol to taxifolin and was highly expressed after E. polonica infection. Down-regulating F3H gene expression by RNA interference in transgenic Norway spruce resulted in significantly lower levels of both dihydroflavonols and flavan-3-ols. Therefore F3H plays a key role in the biosynthesis of defense compounds in Norway spruce that act against the bark beetle-fungus complex. This enzyme forms a defensive product, taxifolin, which is also a metabolic precursor of another defensive product, catechin, which in turn synergizes the toxicity of taxifolin to the bark beetle associated fungus.The Max Planck Institute for Chemical Ecology and the University of Pretoria RDP program.http://www.frontiersin.org/Plant_Scienceam2019Forestry and Agricultural Biotechnology Institute (FABI)Zoology and Entomolog

Flavanone-3-Hydroxylase Plays an Important Role in the Biosynthesis of Spruce Phenolic Defenses Against Bark Beetles and Their Fungal Associates

Conifer forests worldwide are becoming increasingly vulnerable to attacks by bark beetles and their fungal associates due to the effects of global warming. Attack by the bark beetle Ips typographus and the blue-stain fungus it vectors (Endoconidiophora polonica) on Norway spruce (Picea abies) is well known to induce increased production of terpene oleoresin and polyphenolic compounds. However, it is not clear whether specific compounds are important in resisting attack. In this study, we observed a significant increase in dihydroflavonol and flavan-3-ol content after inoculating Norway spruce with the bark beetle vectored fungus. A bioassay revealed that the dihydroflavonol taxifolin and the flavan-3-ol catechin negatively affected both I. typographus and E. polonica. The biosynthesis of flavan-3-ols is well studied in Norway spruce, but little is known about dihydroflavonol formation in this species. A flavanone-3-hydroxylase (F3H) was identified that catalyzed the conversion of eriodictyol to taxifolin and was highly expressed after E. polonica infection. Down-regulating F3H gene expression by RNA interference in transgenic Norway spruce resulted in significantly lower levels of both dihydroflavonols and flavan-3-ols. Therefore F3H plays a key role in the biosynthesis of defense compounds in Norway spruce that act against the bark beetle-fungus complex. This enzyme forms a defensive product, taxifolin, which is also a metabolic precursor of another defensive product, catechin, which in turn synergizes the toxicity of taxifolin to the bark beetle associated fungus

Bark beetle attack history does not influence the induction of terpene and phenolic defenses in mature Norway spruce (Picea abies) trees by the bark beetle-associated fungus Endoconidiophora polonica

Terpenes and phenolics are important constitutive and inducible conifer defenses against bark beetles and their associated fungi. In this study, the inducible defenses of mature Norway spruce (Picea abies) trees with different histories of attack by the spruce bark beetle, Ips typographus were tested by inoculation with the I. typographusassociated fungus Endoconidiophora polonica. We compared trees that had been under previous attack with those under current attack and those that had no record of attack. After fungal inoculation, the concentrations of mono-, sesqui-, and diterpenes in bark increased 3- to 9-fold. For the phenolics, the flavan-3-ols, catechin, and gallocatechin, increased significantly by 2- and 5-fold, respectively, while other flavonoids and stilbenes did not. The magnitudes of these inductions were not influenced by prior bark beetle attack history for all the major compounds and compound classes measured. Before fungal inoculation, the total amounts of monoterpenes, diterpenes, and phenolics (constitutive defenses) were greater in trees that had been previously attacked compared to those under current attack, possibly a result of previous induction. The transcript levels of many genes involved in terpene formation (isoprenyl diphosphate synthases and terpene synthases) and phenolic formation (chalcone synthases) were significantly enhanced by fungal inoculation suggesting de novo biosynthesis. Similar inductions were found for the enzymatic activity of isoprenyl diphosphate synthases and the concentration of their prenyl diphosphate products after fungal inoculation. Quantification of defense hormones revealed a significant induction of the jasmonate pathway, but not the salicylic acid pathway after fungal inoculation. Our data highlight the coordinated induction of terpenes and phenolics in spruce upon infection by E. polonica, a fungal associate of the bark beetle I. typographus, but provide no evidence for the priming of these defense responses by prior beetle attack.Zwillenberg-Tietz Foundation and the Max Planck Society.https://www.frontiersin.org/journals/plant-sciencedm2022Forestry and Agricultural Biotechnology Institute (FABI)Zoology and Entomolog