Comparing lodgepole pine defence responses against mountain pine beetle and Grosmannia clavigera

• Author / Creator

  Pu, Duowen

• The mountain pine beetle (Dendroctonus ponderosae Hopkins; MPB) is a bark beetle that poses a significant threat to pine species in western North America. This threat is evident in the ongoing MPB epidemic, which has resulted in significant losses of lodgepole pine (Pinus contorta var. latifolia) forests. One reason for the effectiveness of MPB attacks on lodgepole pine is their symbiotic relationship with ophiostomatoid fungal associates like Grosmannia clavigera (Robinson-Jeffrey and Davidson) Zipfel, de Beer and Wingfield. Recent research used plant defense hormone profiling to cast doubt on the contribution that G. clavigera, makes to MPB's capacity to overcome lodgepole pine defenses during mass attack. These analyses showed that G. clavigera-inoculated lodgepole pine synthesize significantly increased levels of jasmonate (JA) and the active conjugate jasmonate-isoleucine (JA-Ile) as well as the ethylene (ET) precursor 1-aminocyclopropane-1-carboxylate (ACC). JA and ET are the hormone signature of plant response to necrotrophic fungal pathogens. In contrast, lodgepole pine trees subjected to MPB mass attack exhibited significantly increased levels of JA and JA-Ile but not ACC during the mass attack phase, as expected for plant responses to herbivore insects. The lack of ACC synthesis in lodgepole pines during the mass attack phase suggests that ophiostomatoid fungal symbionts have not begun to colonize host tissues during mass attack, and as such do not contribute to MPB’s capacity to overcome lodgepole pine defenses during this critical phase of insect-host interactions. Building upon this recent study, my study investigated expression patterns of jasmonic acid (JA) and ethylene (ET) biosynthesis and signaling genes in MPB-attacked versus G. clavigera-inoculated lodgepole pines. I hypothesized that if G. clavigera does not significantly contribute to MPB’s capacity to overcome tree defenses, the tree’s response to MPB during the mass attack phase would involve JA biosynthesis andiiisignaling genes but not ET biosynthesis and signaling genes. Since increased expression of genes coding for biosynthetic enzymes often precedes increased levels of their metabolite products, analyzing transcript abundance profiles for JA and ET biosynthesis genes enabled me to explore potential roles for JA and ET in lodgepole pine’s response to MPB during the transition from mass attack to colonization. Based on previous transcriptome profiling experiments, I identified, cloned and carried out in silico analyses of six lodgepole pine cDNAs putatively involved in JA- and ET-mediated responses : lipoxygenase (PcLOX) and allene oxide synthase (PcAOS) for JA biosynthesis, jasmonate ZIM-domain (PcJAZ) for JA signaling, and 1-aminocyclopropane-1-carboxylate oxidase (PcACO1 and PcACO2) and 1-aminocyclopropane-1-carboxylic acid synthase (PcACS) for ET biosynthesis. I then carried out transcript abundance profiling by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) in secondary xylem and secondary phloem harvested from mature lodgepole pine trees that were either mass attacked by MPB or inoculated with G. clavigera, comparing these treatments with both unwounded controls and mock-attacked or mock-inoculated controls. JA-related genes PcLOX and PcJAZ showed significant upregulation in MPB-attacked, G. clavigera-inoculated and mock-treated trees, consistent with the model that chewing herbivorous insects, necrotrophic fungal pathogens and mechanical wounding all trigger the JA pathway. I observed significant upregulation of ET biosynthesis genes PcACO1 and PcACO2 following G. clavigera inoculation, and in in response to wounding and MPB attack. These significant increases in PcACO1 and PcACO2gene expression mirrored measured ACC levels in G. clavigera-inoculated trees, but not in mock- or MPB-attacked samples. The increases in PcACO1 and PcACO2gene transcript abundance in MPB-attacked trees, which were only measured at the later time point and not earlier time point, could be interpreted to mean that G. clavigera colonization wasivsufficient to trigger plant perception of the fungal pathogen during the transition from mass attack to colonization, but that increased ET biosynthesis gene expression had not yet translated into increased ACC levels. Alternatively – or additionally –increased expression of ET biosynthesis genes may have been associated with production of traumatic resin ducts, a classic response of conifer species to wounding, herbivore and pathogen attack. ET has also been recently implicated in plant repair responses. These transcript profiling experiments reveal an additional layer of complexity in the roles for ET in host responses to MPB attack that require additional experiments to resolve.

• Subjects / Keywords

  • mountain pine beetle
  • Grosmannia clavigera
  • Lodgepole Pine

• Graduation date

  Spring 2024

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/r3-vzy7-vz25

• License

  This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.