Tesis: "Phytophthora cinnamomi"

1

Finlay, Annabelle Ruth. "Microbial suppression of Phytophthora cinnamomi." Thesis, Queen's University Belfast, 1987. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317116.

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2

King, Michaela. "The phosphite responsive transcriptome of phytophthora cinnamomi." Thesis, King, Michaela (2007) The phosphite responsive transcriptome of phytophthora cinnamomi. PhD thesis, Murdoch University, 2007. https://researchrepository.murdoch.edu.au/id/eprint/132/.

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Phosphite has been used to effectively control the soil borne plant pathogen Phytophthora cinnamomi in many horticultural crops, forest trees and natural ecosystems. However, the molecular mechanisms behind phosphite action on this pathogen are poorly understood. Several studies have shown that phosphite inhibits growth and zoospore production of P. cinnamomi and in addition induces significant physiological and metabolic changes in the mycelium. As an approach to understanding the mechanisms and relevance of these changes in the pathogen, the effect of phosphite on gene expression was investigated using microarray analysis. To construct the microarray, RNA was extracted from phosphite-treated (40 ug/ml) mycelium of P. cinnamomi isolate MP 80. The chosen phosphite concentration inhibited the mycelial growth by 70% but provided sufficient mycelium for RNA extractions after 4 days growth at 25C. The mRNA was reverse transcribed into cDNA and cloned into lambda to construct a library consisting of 2 million pfu of which 80 % were recombinant phage. The inserts were sequenced for a random selection of clones from the library. The nucleotide sequences generated revealed a range of different P. cinnamomi genes being expressed and demonstrated that the cDNA library provided a good representation of the transcripts expressed in P. cinnamomi. The types of genes found to be expressed in the mycelium of P. cinnamomi included genes encoding GTP binding proteins involved in vesicle transport, structural proteins involved in maintaining cell membrane integrity,elicitors, phosphatases and ribosomal proteins. Over nine thousand cDNA transcripts were randomly selected from the cDNA library and prepared by PCR amplification and purification for microarray construction. Custom made cDNA arrays containing 9216 cDNA transcripts were constructed and probed with RNA from untreated mycelium and mycelium grown in medium with 40 ug/ml phosphite. Two genes, EF-1 alpha and cinnamomin gene, identified by qRT-PCR as being constitutively expressed were also positioned on the arrays as positive controls. In the process of identifying constitutively expressed genes, qRT PCR revealed that phosphite down-regulated a gene encoding ubiquitin-conjugating enzyme, a component of the ubiquitin/proteasome pathway involved in the removal of abnormal and short lived-regulatory proteins and rate limiting enzymes. From the arrays a further seventy-two transcripts with altered patterns in gene expression (fold change > 2) were identified. The majority of the cDNA transcripts spotted on the array were down-regulated with changes in gene expression ranging from 2- to 3.5-fold. Thirty-two cDNA transcripts were up-regulated with changes in gene expression ranging from 2- to 16-fold. Characterisation by sequencing revealed that the most highly induced transcripts coded for ADP-ribosylation factors, an ABC cassette transporter and a glycosyl transferase. A transcript encoding a vitamin B6 biosynthesis protein was also identified as up-regulated by 2.9-fold. In contrast, the down-regulated transcripts coded for cellulose synthase I, annexin, glutamine synthetase, metallothionein and an alternative oxidase. The results are discussed in terms of possible roles and mechanism(s) of phosphite action within the mycelium of P.cinnamomi. This work is the first comprehensive screen for phosphite regulated-gene expression in P. cinnamomi and represents a significant step towards an understanding of the mode of action of phosphite on this organism. This thesis provides valuable information on the molecular interaction between phosphite and P. cinnamomi, which in future studies may stimulate the discovery of novel methods and cellular targets for the control of plant pathogenic Oomycetes.

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3

King, Michaela. "The phosphite responsive transcriptome of Phytophthora cinnamomi /." King, Michaela (2007) The phosphite responsive transcriptome of phytophthora cinnamomi. PhD thesis, Murdoch University, 2007. http://researchrepository.murdoch.edu.au/132/.

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Phosphite has been used to effectively control the soil borne plant pathogen Phytophthora cinnamomi in many horticultural crops, forest trees and natural ecosystems. However, the molecular mechanisms behind phosphite action on this pathogen are poorly understood. Several studies have shown that phosphite inhibits growth and zoospore production of P. cinnamomi and in addition induces significant physiological and metabolic changes in the mycelium. As an approach to understanding the mechanisms and relevance of these changes in the pathogen, the effect of phosphite on gene expression was investigated using microarray analysis. To construct the microarray, RNA was extracted from phosphite-treated (40 ug/ml) mycelium of P. cinnamomi isolate MP 80. The chosen phosphite concentration inhibited the mycelial growth by 70% but provided sufficient mycelium for RNA extractions after 4 days growth at 25C. The mRNA was reverse transcribed into cDNA and cloned into lambda to construct a library consisting of 2 million pfu of which 80 % were recombinant phage. The inserts were sequenced for a random selection of clones from the library. The nucleotide sequences generated revealed a range of different P. cinnamomi genes being expressed and demonstrated that the cDNA library provided a good representation of the transcripts expressed in P. cinnamomi. The types of genes found to be expressed in the mycelium of P. cinnamomi included genes encoding GTP binding proteins involved in vesicle transport, structural proteins involved in maintaining cell membrane integrity,elicitors, phosphatases and ribosomal proteins. Over nine thousand cDNA transcripts were randomly selected from the cDNA library and prepared by PCR amplification and purification for microarray construction. Custom made cDNA arrays containing 9216 cDNA transcripts were constructed and probed with RNA from untreated mycelium and mycelium grown in medium with 40 ug/ml phosphite. Two genes, EF-1 alpha and cinnamomin gene, identified by qRT-PCR as being constitutively expressed were also positioned on the arrays as positive controls. In the process of identifying constitutively expressed genes, qRT PCR revealed that phosphite down-regulated a gene encoding ubiquitin-conjugating enzyme, a component of the ubiquitin/proteasome pathway involved in the removal of abnormal and short lived-regulatory proteins and rate limiting enzymes. From the arrays a further seventy-two transcripts with altered patterns in gene expression (fold change > 2) were identified. The majority of the cDNA transcripts spotted on the array were down-regulated with changes in gene expression ranging from 2- to 3.5-fold. Thirty-two cDNA transcripts were up-regulated with changes in gene expression ranging from 2- to 16-fold. Characterisation by sequencing revealed that the most highly induced transcripts coded for ADP-ribosylation factors, an ABC cassette transporter and a glycosyl transferase. A transcript encoding a vitamin B6 biosynthesis protein was also identified as up-regulated by 2.9-fold. In contrast, the down-regulated transcripts coded for cellulose synthase I, annexin, glutamine synthetase, metallothionein and an alternative oxidase. The results are discussed in terms of possible roles and mechanism(s) of phosphite action within the mycelium of P.cinnamomi. This work is the first comprehensive screen for phosphite regulated-gene expression in P. cinnamomi and represents a significant step towards an understanding of the mode of action of phosphite on this organism. This thesis provides valuable information on the molecular interaction between phosphite and P. cinnamomi, which in future studies may stimulate the discovery of novel methods and cellular targets for the control of plant pathogenic Oomycetes.

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4

au, M. King@murdoch edu, and Michaela King. "The phosphite responsive transcriptome of phytophthora cinnamomi." Murdoch University, 2007. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20080526.104656.

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Phosphite has been used to effectively control the soil borne plant pathogen Phytophthora cinnamomi in many horticultural crops, forest trees and natural ecosystems. However, the molecular mechanisms behind phosphite action on this pathogen are poorly understood. Several studies have shown that phosphite inhibits growth and zoospore production of P. cinnamomi and in addition induces significant physiological and metabolic changes in the mycelium. As an approach to understanding the mechanisms and relevance of these changes in the pathogen, the effect of phosphite on gene expression was investigated using microarray analysis. To construct the microarray, RNA was extracted from phosphite-treated (40 ug/ml) mycelium of P. cinnamomi isolate MP 80. The chosen phosphite concentration inhibited the mycelial growth by 70% but provided sufficient mycelium for RNA extractions after 4 days growth at 25C. The mRNA was reverse transcribed into cDNA and cloned into lambda to construct a library consisting of 2 million pfu of which 80 % were recombinant phage. The inserts were sequenced for a random selection of clones from the library. The nucleotide sequences generated revealed a range of different P. cinnamomi genes being expressed and demonstrated that the cDNA library provided a good representation of the transcripts expressed in P. cinnamomi. The types of genes found to be expressed in the mycelium of P. cinnamomi included genes encoding GTP binding proteins involved in vesicle transport, structural proteins involved in maintaining cell membrane integrity,elicitors, phosphatases and ribosomal proteins. Over nine thousand cDNA transcripts were randomly selected from the cDNA library and prepared by PCR amplification and purification for microarray construction. Custom made cDNA arrays containing 9216 cDNA transcripts were constructed and probed with RNA from untreated mycelium and mycelium grown in medium with 40 ug/ml phosphite. Two genes, EF-1 alpha and cinnamomin gene, identified by qRT-PCR as being constitutively expressed were also positioned on the arrays as positive controls. In the process of identifying constitutively expressed genes, qRT PCR revealed that phosphite down-regulated a gene encoding ubiquitin-conjugating enzyme, a component of the ubiquitin/proteasome pathway involved in the removal of abnormal and short lived-regulatory proteins and rate limiting enzymes. From the arrays a further seventy-two transcripts with altered patterns in gene expression (fold change > 2) were identified. The majority of the cDNA transcripts spotted on the array were down-regulated with changes in gene expression ranging from 2- to 3.5-fold. Thirty-two cDNA transcripts were up-regulated with changes in gene expression ranging from 2- to 16-fold. Characterisation by sequencing revealed that the most highly induced transcripts coded for ADP-ribosylation factors, an ABC cassette transporter and a glycosyl transferase. A transcript encoding a vitamin B6 biosynthesis protein was also identified as up-regulated by 2.9-fold. In contrast, the down-regulated transcripts coded for cellulose synthase I, annexin, glutamine synthetase, metallothionein and an alternative oxidase. The results are discussed in terms of possible roles and mechanism(s) of phosphite action within the mycelium of P.cinnamomi. This work is the first comprehensive screen for phosphite regulated-gene expression in P. cinnamomi and represents a significant step towards an understanding of the mode of action of phosphite on this organism. This thesis provides valuable information on the molecular interaction between phosphite and P. cinnamomi, which in future studies may stimulate the discovery of novel methods and cellular targets for the control of plant pathogenic Oomycetes.

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5

Reitmann, Anandi. "Identification of pathogenicity genes in Phytophthora cinnamomi." Diss., University of Pretoria, 2014. http://hdl.handle.net/2263/79179.

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6

Gilovitz, Joshua. "Screening Lambertia for resistance to Phytophthora cinnamomi." Thesis, Gilovitz, Joshua (2007) Screening Lambertia for resistance to Phytophthora cinnamomi. Honours thesis, Murdoch University, 2007. https://researchrepository.murdoch.edu.au/id/eprint/32597/.

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7

Wheeler, Margaret Anne. "Reproductive and molecular biology of Eucalyptus marginata Donn ex Smith /." Access via Murdoch University Digital Theses Project, 2003. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20040723.140250.

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8

McCarren, Kathryn. "Saprophytic ability and the contribution of chlamydospores and oospores to the survival of Phytophthora cinnamomi." Thesis, McCarren, Kathryn (2006) Saprophytic ability and the contribution of chlamydospores and oospores to the survival of Phytophthora cinnamomi. PhD thesis, Murdoch University, 2006. https://researchrepository.murdoch.edu.au/id/eprint/190/.

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Phytophthora cinnamomi has been recognised as a key threatening process to Australia's biodiversity by the Commonwealth's Environment Protection and Biodiversity Conservation Act 1999. Despite over 80 years of extensive research, its exact mode of survival is still poorly understood. It is widely accepted that thin- and thick-walled chlamydospores are the main survival propagules while oospores are assumed to play no role in the survival of the pathogen in the Australian environment, yet evidence is limited. The saprophytic ability of the pathogen is still unresolved despite the important role this could play in the ability of the pathogen to survive in the absence of susceptible hosts. This thesis aimed to investigate chlamydospores, oospores and the saprophytic ability of P. cinnamomi to determine their contribution to survival. Phytophthora cinnamomi did not show saprophytic ability in non-sterile soils. The production of thick-walled chlamydospores and selfed oospores of P. cinnamomi in vitro was documented. Thick-walled chlamydospores were sporadically formed under sterile and non-sterile conditions in vitro but exact conditions for stimulating their formation could not be determined. The formation of thick-walled chlamydospores emerging from mycelium of similar wall thickness was observed, challenging the current knowledge of chlamydospore formation. Selfed oospores were abundant in vitro on modified Ribeiro's minimal medium in one isolate. Three other isolates tested also produced oospores but not in large numbers. Although the selfed oospores did not germinate on a range of media, at least 16 % were found to be viable using Thiozolyl Blue Tetrazolium Bromide staining and staining of the nuclei with 4', 6-diamidino-2-phenylindole.2HCl (DAPI). This indicated the potential of selfed oospores as survival structures and their ability to exist dormantly. The ability of phosphite to kill chlamydospores and selfed oospores was studied in vitro. Results challenged the efficacy of this chemical and revealed the necessity for further study of its effect on survival propagules of P. cinnamomi in the natural environment. Phosphite was shown to induce dormancy in thin-walled chlamydospores if present during their formation in vitro. Interestingly, dormancy was only induced by phosphite in isolates previously reported as sensitive to phosphite and not those reported as tolerant. Chlamydospores were produced uniformly across the radius of the colony on control modified Ribeiro's minimal medium but on medium containing phosphite (40 or 100 mcg ml-1), chlamydospore production was initially inhibited before being stimulated during the log phase of growth. This corresponded to a point in the colony morphology where mycelial density changed from tightly packed mycelium to sparse on medium containing phosphite. This change in morphology did not occur when the pathogen was grown on liquid media refreshed every four days, and chlamydospores were evenly distributed across the radius of these colonies. This trend was not observed in selfed oospores produced in the presence of phosphite. Selfed oospore production was found to be inhibited by phosphite at the same concentrations that stimulated chlamydospore production. Isolates of P. cinnamomi were transformed using a protoplast/ polyethylene glycol method to contain the Green Fluorescent Protein and geneticin resistance genes to aid in future studies on survival properties of the organism. Although time constraints meant the stability of the transgene could not be determined, it was effective in differentiating propagules of the transformed P. cinnamomi from spores of other microrganisms in a non-sterile environment. Two different sized chlamydospores (approximately 30 mcg diameter and < 20 mcg diameter) were observed in preliminary trials of transformed P. cinnamomi inoculated lupin roots floated in non-sterile soil extracts and these were easily distinguished from microbial propagules of other species. The growth and pathogenicity was reduced in two putative transformants and their ability to fluoresce declined over ten subcultures but they still remained resistant to geneticin. This study has improved our knowledge on the survival abilities of P. cinnamomi in vitro and has provided a useful tool for studying these abilities under more natural glasshouse conditions. Important implications of phosphite as a control have been raised.

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9

McCarren, Kathryn. "Saprophytic ability and the contribution of chlamydospores and oospores to the survival of Phytophthora cinnamomi." McCarren, Kathryn (2006) Saprophytic ability and the contribution of chlamydospores and oospores to the survival of Phytophthora cinnamomi. PhD thesis, Murdoch University, 2006. http://researchrepository.murdoch.edu.au/190/.

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Phytophthora cinnamomi has been recognised as a key threatening process to Australia's biodiversity by the Commonwealth's Environment Protection and Biodiversity Conservation Act 1999. Despite over 80 years of extensive research, its exact mode of survival is still poorly understood. It is widely accepted that thin- and thick-walled chlamydospores are the main survival propagules while oospores are assumed to play no role in the survival of the pathogen in the Australian environment, yet evidence is limited. The saprophytic ability of the pathogen is still unresolved despite the important role this could play in the ability of the pathogen to survive in the absence of susceptible hosts. This thesis aimed to investigate chlamydospores, oospores and the saprophytic ability of P. cinnamomi to determine their contribution to survival. Phytophthora cinnamomi did not show saprophytic ability in non-sterile soils. The production of thick-walled chlamydospores and selfed oospores of P. cinnamomi in vitro was documented. Thick-walled chlamydospores were sporadically formed under sterile and non-sterile conditions in vitro but exact conditions for stimulating their formation could not be determined. The formation of thick-walled chlamydospores emerging from mycelium of similar wall thickness was observed, challenging the current knowledge of chlamydospore formation. Selfed oospores were abundant in vitro on modified Ribeiro's minimal medium in one isolate. Three other isolates tested also produced oospores but not in large numbers. Although the selfed oospores did not germinate on a range of media, at least 16 % were found to be viable using Thiozolyl Blue Tetrazolium Bromide staining and staining of the nuclei with 4', 6-diamidino-2-phenylindole.2HCl (DAPI). This indicated the potential of selfed oospores as survival structures and their ability to exist dormantly. The ability of phosphite to kill chlamydospores and selfed oospores was studied in vitro. Results challenged the efficacy of this chemical and revealed the necessity for further study of its effect on survival propagules of P. cinnamomi in the natural environment. Phosphite was shown to induce dormancy in thin-walled chlamydospores if present during their formation in vitro. Interestingly, dormancy was only induced by phosphite in isolates previously reported as sensitive to phosphite and not those reported as tolerant. Chlamydospores were produced uniformly across the radius of the colony on control modified Ribeiro's minimal medium but on medium containing phosphite (40 or 100 mcg ml-1), chlamydospore production was initially inhibited before being stimulated during the log phase of growth. This corresponded to a point in the colony morphology where mycelial density changed from tightly packed mycelium to sparse on medium containing phosphite. This change in morphology did not occur when the pathogen was grown on liquid media refreshed every four days, and chlamydospores were evenly distributed across the radius of these colonies. This trend was not observed in selfed oospores produced in the presence of phosphite. Selfed oospore production was found to be inhibited by phosphite at the same concentrations that stimulated chlamydospore production. Isolates of P. cinnamomi were transformed using a protoplast/ polyethylene glycol method to contain the Green Fluorescent Protein and geneticin resistance genes to aid in future studies on survival properties of the organism. Although time constraints meant the stability of the transgene could not be determined, it was effective in differentiating propagules of the transformed P. cinnamomi from spores of other microrganisms in a non-sterile environment. Two different sized chlamydospores (approximately 30 mcg diameter and < 20 mcg diameter) were observed in preliminary trials of transformed P. cinnamomi inoculated lupin roots floated in non-sterile soil extracts and these were easily distinguished from microbial propagules of other species. The growth and pathogenicity was reduced in two putative transformants and their ability to fluoresce declined over ten subcultures but they still remained resistant to geneticin. This study has improved our knowledge on the survival abilities of P. cinnamomi in vitro and has provided a useful tool for studying these abilities under more natural glasshouse conditions. Important implications of phosphite as a control have been raised.

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10

Pilbeam, Ros. "Effects of phosphite on disease development and histological responses in Eucalyptus marginata infected with Phytophthora cinnamomi." Thesis, Pilbeam, Ros (2003) Effects of phosphite on disease development and histological responses in Eucalyptus marginata infected with Phytophthora cinnamomi. PhD thesis, Murdoch University, 2003. https://researchrepository.murdoch.edu.au/id/eprint/260/.

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Phosphite is currently used for the management of Phytophthora cinnamomi in native plant communities. A greater understanding of how phosphite affects the host-pathogen interaction is required in order to determine the most effective treatment. This thesis aimed to investigate the effects of applied phosphite concentration on phytotoxicity, in planta concentration of phosphite, disease development and anatomical responses of Eucalyptus marginata. Spraying the foliage to run-off with 7.5 and 10 g phosphite/L led to the development of severe leaf necrosis within 7 days, with greater than 60% of the leaf area damaged. Moderate phytotoxicity was observed after treatment with 5 g phosphite/L. In planta concentration of phosphite in stems, lignotubers and roots did not differ significantly between applied concentrations of phosphite. Stem tissue contained the largest concentration of phosphite at one week after spraying, with approximately 210 and 420 g phosphite/g dry weight detected after treatment with 5 and 10 g phosphite/L, respectively. In a subsequent field trial, the applied concentration of phosphite was found to affect the duration of effectiveness of phosphite in protecting E. marginata seedlings from stem colonisation by P. cinnamomi. Plants were wound-inoculated with P. cinnamomi at 6-monthly intervals after spraying with phosphite. The 2.5 and 5 g phosphite/L treatments were effective against colonisation by P. cinnamomi when inoculated 0 and 6 months after spraying, but only the 5 g phosphite/L treatment inhibited P. cinnamomi within 12 months of spraying. Phosphite had no effect on colonisation by P. cinnamomi when plants were inoculated at 17 months after spraying. The in planta concentration of phosphite detected in the leaves, stems and roots of plants treated with 5 g phosphite/L did not differ significantly between the time of harvest or tissue type at 0.2 and 6 months after spraying. P. cinnamomi remained viable in plants treated with phosphite.Treatment with 2.5 and 5 g phosphite/L when P. cinnamomi was well established in the stems was ineffective at preventing the death of E. marginata. Between 45 and 89% of plants were girdled on the day of spraying. Spraying plants with 2.5 and 5 g phosphite/L when conditions were less favourable for the pathogen reduced the mortality of E. marginata for up to 10 months. E. marginata seedlings responded to damage by P. cinnamomi with the production of kino veins and woundwood. Bark lesions were in the process of being sloughed off by 7 months after inoculation in plants that remained alive. In plants of a resistant (RR) clonal line and susceptible (SS) clonal line, phosphite treatment inhibited lesion extension in stems, but lesions did not indicate the amount of stem colonised by P. cinnamomi. The pathogen was isolated from up to 17 cm beyond the lesion front in the RR clonal line. Treatments that reduced the mortality of E. marginata were 5 g phosphite/L in the RR clonal line (RR/5) and 10 g phosphite/L in the SS clonal line (SS/10). Uninoculated plants were wounded with liquid nitrogen to determine the microscopic responses to injury in the absence of the pathogen. Wound closure was achieved within 21 days of wounding, with callus formation and vascular cambium regeneration. A wound periderm separated wounded tissue from healthy tissue, adjacent to a lignified boundary zone. Two types of phellem were observed - thin-walled phellem (TnP) and thick-walled phellem (TkP). The first-formed TnP layers contained variable-shaped cells, while subsequent layers were more cubical in shape. Multiple TnP layers developed up to 42 days after wounding, with TkP cells sandwiched between the TnP layers. Genotype and phosphite treatment did not affect the wound responses. Inoculated plants with a restricted lesion extension also formed a wound periderm to separate damaged tissue from healthy tissue. Phosphite treatment stimulated the responses to P. cinnamomi in both clonal lines. Early development of the wound periderm was visible by 6 days after phosphite treatment. It waspreceded by the formation of a ligno-suberised boundary zone in the cambial zone and in phloem parenchyma cells existing prior to injury. Suberin was not detected in the SS/0 treatment. TnP layers completely surrounded lesioned tissue in plants still alive by 24 days after phosphite treatment. Extensive callus production was evident in the SS/10, RR/5 and RR/10 treatments. Temperature affected the post-inoculation efficacy of phosphite and anatomical responses of E. marginata. At 20 degrees C lesion extension was restricted in both clonal lines of E. marginata, irrespective of phosphite treatment. Greater than 70% of inoculated plants in all treatments produced a ligno-suberised boundary zone at 20 degrees C and between 30 and 70% formed a wound periderm. At 28 degrees C, lesion extension was reduced in phosphite-treated plants at 7 days after treatment. However, lesions continued to extend up to 5 mm per day in the SS clonal line and very few SS plants formed a wound periderm at the lesion front. This contrasted with the strong responses to abiotic wounding observed in uninoculated SS plants at 28 degrees C. The most extensive responses to P. cinnamomi were detected in the RR/5 treatment at 28 degrees C, with a ligno-suberised boundary zone and differentiated TnP of a wound periderm observed in greater than 70% of plants. This treatment resulted in significantly less girdled plants than all other treatments at 28 degrees C, including the RR/0 treatment. At 23 and 24 degrees C, there was no significant difference in acropetal lesion extension or circumferential lesion spread between clonal lines. The inoculation technique and environmental conditions may have resulted in too high a disease pressure for a full expression of resistance in the RR clonal line. This thesis demonstrates that phosphite has the potential to enhance the resistance of young E. marginata and enable them to survive infection by P. cinnamomi. However, its effectiveness is dependent upon a number of factors, including host resistance, environmental conditions, the applied phosphite concentration and the timing of application.

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11

Pilbeam, Ros. "Effects of phosphite on disease development and histological responses in Eucalyptus marginata infected with Phytophthora cinnamomi." Murdoch University, 2003. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20040820.140206.

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Phosphite is currently used for the management of Phytophthora cinnamomi in native plant communities. A greater understanding of how phosphite affects the host-pathogen interaction is required in order to determine the most effective treatment. This thesis aimed to investigate the effects of applied phosphite concentration on phytotoxicity, in planta concentration of phosphite, disease development and anatomical responses of Eucalyptus marginata. Spraying the foliage to run-off with 7.5 and 10 g phosphite/L led to the development of severe leaf necrosis within 7 days, with greater than 60% of the leaf area damaged. Moderate phytotoxicity was observed after treatment with 5 g phosphite/L. In planta concentration of phosphite in stems, lignotubers and roots did not differ significantly between applied concentrations of phosphite. Stem tissue contained the largest concentration of phosphite at one week after spraying, with approximately 210 and 420 µg phosphite/g dry weight detected after treatment with 5 and 10 g phosphite/L, respectively. In a subsequent field trial, the applied concentration of phosphite was found to affect the duration of effectiveness of phosphite in protecting E. marginata seedlings from stem colonisation by P. cinnamomi. Plants were wound-inoculated with P. cinnamomi at 6-monthly intervals after spraying with phosphite. The 2.5 and 5 g phosphite/L treatments were effective against colonisation by P. cinnamomi when inoculated 0 and 6 months after spraying, but only the 5 g phosphite/L treatment inhibited P. cinnamomi within 12 months of spraying. Phosphite had no effect on colonisation by P. cinnamomi when plants were inoculated at 17 months after spraying. The in planta concentration of phosphite detected in the leaves, stems and roots of plants treated with 5 g phosphite/L did not differ significantly between the time of harvest or tissue type at 0.2 and 6 months after spraying. P. cinnamomi remained viable in plants treated with phosphite.Treatment with 2.5 and 5 g phosphite/L when P. cinnamomi was well established in the stems was ineffective at preventing the death of E. marginata. Between 45 and 89% of plants were girdled on the day of spraying. Spraying plants with 2.5 and 5 g phosphite/L when conditions were less favourable for the pathogen reduced the mortality of E. marginata for up to 10 months. E. marginata seedlings responded to damage by P. cinnamomi with the production of kino veins and woundwood. Bark lesions were in the process of being sloughed off by 7 months after inoculation in plants that remained alive. In plants of a resistant (RR) clonal line and susceptible (SS) clonal line, phosphite treatment inhibited lesion extension in stems, but lesions did not indicate the amount of stem colonised by P. cinnamomi. The pathogen was isolated from up to 17 cm beyond the lesion front in the RR clonal line. Treatments that reduced the mortality of E. marginata were 5 g phosphite/L in the RR clonal line (RR/5) and 10 g phosphite/L in the SS clonal line (SS/10). Uninoculated plants were wounded with liquid nitrogen to determine the microscopic responses to injury in the absence of the pathogen. Wound closure was achieved within 21 days of wounding, with callus formation and vascular cambium regeneration. A wound periderm separated wounded tissue from healthy tissue, adjacent to a lignified boundary zone. Two types of phellem were observed thin-walled phellem (TnP) and thick-walled phellem (TkP). The first-formed TnP layers contained variable-shaped cells, while subsequent layers were more cubical in shape. Multiple TnP layers developed up to 42 days after wounding, with TkP cells sandwiched between the TnP layers. Genotype and phosphite treatment did not affect the wound responses. Inoculated plants with a restricted lesion extension also formed a wound periderm to separate damaged tissue from healthy tissue. Phosphite treatment stimulated the responses to P. cinnamomi in both clonal lines. Early development of the wound periderm was visible by 6 days after phosphite treatment. It waspreceded by the formation of a ligno-suberised boundary zone in the cambial zone and in phloem parenchyma cells existing prior to injury. Suberin was not detected in the SS/0 treatment. TnP layers completely surrounded lesioned tissue in plants still alive by 24 days after phosphite treatment. Extensive callus production was evident in the SS/10, RR/5 and RR/10 treatments. Temperature affected the post-inoculation efficacy of phosphite and anatomical responses of E. marginata. At 20°C, lesion extension was restricted in both clonal lines of E. marginata, irrespective of phosphite treatment. Greater than 70% of inoculated plants in all treatments produced a ligno-suberised boundary zone at 20°C and between 30 and 70% formed a wound periderm. At 28°C, lesion extension was reduced in phosphite-treated plants at 7 days after treatment. However, lesions continued to extend up to 5 mm per day in the SS clonal line and very few SS plants formed a wound periderm at the lesion front. This contrasted with the strong responses to abiotic wounding observed in uninoculated SS plants at 28°C. The most extensive responses to P. cinnamomi were detected in the RR/5 treatment at 28°C, with a ligno-suberised boundary zone and differentiated TnP of a wound periderm observed in greater than 70% of plants. This treatment resulted in significantly less girdled plants than all other treatments at 28°C, including the RR/0 treatment. At 23 and 24°C, there was no significant difference in acropetal lesion extension or circumferential lesion spread between clonal lines. The inoculation technique and environmental conditions may have resulted in too high a disease pressure for a full expression of resistance in the RR clonal line. This thesis demonstrates that phosphite has the potential to enhance the resistance of young E. marginata and enable them to survive infection by P. cinnamomi. However, its effectiveness is dependent upon a number of factors, including host resistance, environmental conditions, the applied phosphite concentration and the timing of application.

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12

Harland, Chad S. "F-actin and integrin like proteins in Phytophthora cinnamomi." Thesis, University of Canterbury. Biological Sciences, 2007. http://hdl.handle.net/10092/1895.

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Tip growth is the primary form of growth in hyphal organisms and some plant cells. Tip growth in hyphae is highly dependent on F-actin, which acts to regulate and support growth. One of the models suggested for tip growth, the amebae model of tip growth, suggests that F-actin may also be the primary source of protrusive force for tip growth in some conditions, and that proteins with a similar function to animal integrins would be present an involved in tip growth (Heath and Steinberg 1999). In this thesis we examine the role of F-actin in the growth of the oomycete Phytophthora cinnamomi and the effects on growth of the F-actin disrupting compound Latrunculin B. We demonstrate that F-actin plays a critical role in the tip growth of Phytophthora cinnamomi with it's disruption causing rapid cessation in directional growth, followed by significant subapical swelling. Further more we examine Phytophthora cinnamomi for the presence of an B4 integrin like protein that has been previously reported in the oomycete Achlya bisexualis (Chitcholtan & Garrill 2005) and show that the B4 integrin like protein is not present in Phytophthora cinnamomi. These experiments help further our understanding of tip growth in Phytophthora cinnamomi an economically important plant pathogen.

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13

Armistead, Rodney. "The impact of Phytophthora cinnamomi on the yellow-footed antechinus (mardo) (Antechinus flavipes leucogaster) (Marsupialia: Dasyuridae) /." Murdoch University Digital Theses Program, 2008. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20100330.90319.

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14

Dunne, Christopher Philip. "Control of Sudden Death in Cultivated Proteas from the Southwest of Western Australia." Murdoch University, 2004. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20041207.140807.

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Phytophthora cinnamomi Rands is a common and devastating pathogen of cultivated proteas worldwide. Webb (1997) described a Sudden Death plant disease of proteas in Western Australia (WA) protea plantations. Proteas that suffer the syndrome display symptoms such as stunted growth, wilting, chlorosis and often death. In the current study, a number of protea plantations in the southwest of WA were visited to quantify the extent that P. cinnamomi was attributing to deaths of cultivated proteas. The survey indicated that P. cinnamomi is the major cause of Sudden Death in proteas. A range of other fungi (Fusarium, Botryosphaeria, Pestalotiopsis, Alternaria) and pests (nematodes, mealy bug, scale insects) were also identified to be contributing to protea death and decline in WA plantations. In many cases the factors contributing to protea disease appeared complex, with a range of physical factors or nutritional imbalances commonly associated with these pathogens and pests. As P. cinnamomi was the major cause of death of cultivated proteas the remainder of the experiments described in this dissertation investigated its control in horticultural plantings. Biofumigation has the potential to become an important technique in an overall integrated management approach to P. cinnamomi. In this thesis, biofumigation refers to the suppression of pathogens and pests by the incorporation of Brassica plants into the soil. Two biofumigants (Brassica juncea (L.) Czern., B. napus L.) were screened for their effect on the in vitro growth of five common Phytophthora species (P. cinnamomi, P. cactorum (Lebert & Colin) Schroeter., P. citricola Sawada, P. cryptogea Pethyb. & Laff. and P. megasperma Drechsler). Growth was determined by the measuring dry weight and radial growth of vegetative hyphae. B. juncea was found to be superior in its suppressive effect compared to B. napus. There was also significant variation in the sensitivity of the Phytophthora species to the suppressive effects of the biofumigants. P. cinnamomi was the most sensitive of the five species investigated. Where the rates of the biofumigant were sufficient to suppress growth of Phytophthora, the suppressive effect was mostly fungicidal. To determine how B. juncea and B. napus affect the infective ability and survival of P. cinnamomi, their effects on sporangia and chlamydospores production in soil was investigated in vitro. P. cinnamomi colonised Miracloth discs were added to soil amended with the two Brassica species, before being removed every two days over an eight day period for the determination of sporangia production, chlamydospore production and infective ability. Only the soils amended with B. juncea significantly reduced sporangia production in P. cinnamomi. Both Brassica species increased the percentage of aborted or immature sporangia and reduced the infective ability of the pathogen. Neither Brassica species had any effect on zoospore release or chlamydospore production in P. cinnamomi. Soil cores and soil leachate were collected from biofumigant-amended field soils to determine the inoculum potential and infective ability of the pathogen under glasshouse conditions. Amending the soil with both Brassica species had an immediate suppressive effect on the inoculum potential and infective ability of the P. cinnamomi. However, after this initial suppression there was a gradual increase in the recovery of the pathogen over the monitoring period of four weeks. To determine if the suppression would result in decreased disease incidence in a susceptible host, Lupinus angustifolius L. seeds were planted in the biofumigant amended soil. B. juncea amended soils reduced the disease incidence of P. cinnamomi by 25%. B. napus had no effect on disease incidence in L. angustifolius. Although the current study had demonstrated that biofumigants could suppress the growth, sporulation and infection of P. cinnamomi, it was unclear if this would equate to a reduction in disease incidence when applied in the field. A field trial was conducted on a protea plantation in the southwest of Western Australia that compared biofumigation with B. juncea to chemical fumigation (metham sodium) and soil solarisation. The three soil treatments were used in an integrated management approach to control P. cinnamomi that included the use of a hardwood compost, mulch and water sterilisation. All treatments were monitored during their application to ensure the treatments were conducted successfully. The three soil treatments significantly reduced the recovery of the pathogen and the infective ability of the pathogen to a soil depth of 20 cm. Metham sodium was the most suppressive soil treatment and soil solarisation was the least suppressive treatment. Only the metham sodium treatment resulted in a significant reduction in the incidence of root rot in Leucadendron salignum P.J. Bergius x laureolum (Lam.) Fourc (c.v. Safari Sunset) over the monitoring period of three years. Another field trial was conducted on the same protea plantation to compare the effectiveness of B. juncea and B. napus, without the use of other control strategies, to reduce the incidence of P. cinnamomi infection of Leucadendron Safari Sunset. The concentration of isothiocyanates was monitored for seven days after the incorporation of the biofumigants. Although both Brassica species reduced the recovery and infective ability of the pathogen, neither biofumigant reduced the incidence of root rot in Leucadendron Safari Sunset. In conclusion, P. cinnamomi is the most common and devastating pathogen in WA protea plantations. The current study demonstrated that P. cinnamomi is sensitive to the suppressive nature of biofumigants. Biofumigants can suppress the in vitro growth, sporulation, infective ability of P. cinnamomi and reduce the incidence of the disease caused by the pathogen in the glasshouse. Of the two Brassica species investigated, B. juncea was superior in its ability to control P. cinnamomi compared to B. napus. When applied in the field, biofumigation using B. juncea was found to be more suppressive that soil solarisation, but not as effective as metham sodium.

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15

au, D. Huberli@murdoch edu, and Daniel Huberli. "Phenotypic variation of two localised populations of Phytophthora cinnamomi from Western Australia and how they impact on Eucalyptus marginata resistance." Murdoch University, 2001. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20070827.91902.

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Phytophthora cinnamomi is an introduced soilborne phytopathogen to Western Australia (WA) and impacts on 2000 of the approximately 9000 plant species indigenous in the southwest of WA. Amongst these is Eucalyptus marginata (jarrah), the dominant and economically important hardwood timber species of the jarrah forest. This thesis aimed to investigate the morphological, pathogenic and genotypic variation in two local WA populations of P. cinnamomi isolates. The populations were selected from areas where jarrah clonal lines selected for resistance to P. cinnamomi may be used in the rehabilitation of infested jarrah forest and rehabilitated bauxite minesites in the southwest of WA. Resistance against a range of isolates using different inoculation methods. Seventy-three isolates of P. cinnamomi were collected from diseased jarrah and Corymbia calophylla (marri) trees from two populations located 70 km apart and these were examined for phenotypic and genotypic variation. Microsatellite DNA analysis showed that all isolates were of the same clonal lineage. In P. cinnamomi for the first time I show that there is a broad and continuous variation in the morphology and pathology between two populations of one clonal lineage, and that all phenotypes varied independently from one another. No relationship was found between morphological and pathogenic characters. The ability of isolates in both populations to cause deaths ranged from killing all plants within 59 days to plants being symptomless 182 days after inoculation. Single and multiple paragynous antheridia formed along with amphigynous ones in mating studies with all WA isolates and a sample of worldwide isolates. Developmental studies and cytological examination showed fertilisation tubes developed asynchronously or synchronously from both antheridial types and indicated that either antheridial type contributed a nucleus for fertilisation of the oosphere. This is the first report of paragynous antheridial associations in P. cinnamomi. Antheridial variation is a characteristic that needs to be adjusted in the taxonomic Phytophthora identification keys. In underbark and zoospore stem inoculations of three 1.5-year-old jarrah clonal lines (two ranked as resistant (RR) and one as susceptible (SS) to P. cinnamomi in the original selection trials) at 15, 20, 25 and 30°C, it was found that the method of inoculation did not produce comparable results, particularly at 25 and 30°C. At these temperatures, all three clonal lines had 100% mortality when inoculated underbark, but when inoculated with zoospores, one RR line had 60% survival and the SS and remaining RR line had 100% mortality. Generally, the level of resistance of all clonal lines declined with increasing temperature. Lesion development was measured at 20, 25 and 30°C for 4 days in detached branches of an RR and SS clonal line inoculated underbark with four different P. cinnamomi isolates. Detached branches were found to be a potential screen for jarrah resistance to P. cinnamomi and to allow the identification of susceptible and resistant clonal lines at 30°C. Lesion and colonisation development of P. cinnamomi isolates were assessed in situ (late autumn) of seed-grown and clonal lines of 3.5 to 4.5 year-old jarrah trees growing in a rehabilitated minesite jarrah forest in underbark inoculation of lateral branches (1995) or simultaneously in lateral branches and lateral roots (1996). Trees were underbark inoculated in lateral branches and lateral roots. Colonisation was more consistent as a measure of resistance than lesion length over the two trials because it accounted for the recovery of P. cinnamomi from macroscopically symptomless tissue beyond lesions, which on some occasions, was up to 6 cm. In the two trials, one RR clonal line consistently had small lesion and colonisation lengths in branches and roots. In contrast, the remaining two RR clonal lines had similar lesion and colonisation lengths to the SS clonal line and may, therefore, not be suitable for use in the rehabilitation of P. cinnamomi infested areas. The relative rankings of the jarrah clonal lines by colonisation lengths were similar between branch and root inoculations. Branch inoculations are a valid option for testing resistance and susceptibility of young jarrah trees to P. cinnamomi. The pathogen was recovered on Phytophthora selective agar 36 months after inoculation from 50% of samples with lesions and 30% of symptomless samples in a series of growth cabinet, glasshouse and field experiments. However, up to 11% of samples with and without lesions and from which P. cinnamomi was not initially isolated contained viable pathogen after leaching the plant material in water over 9 days. This indicates that the pathogen could be present as dormant structures, such as chlamydospores, where dormancy needs to be broken for germination to occur, or fungistatic compounds in the tissue need to be removed to allow the pathogen to grow, or both. These results have important implications for disease diagnosis and management, disease-free certification and quarantine clearance. No clonal line of jarrah was found to be 100% resistant using different inoculation methods, environmental conditions and when challenged by individuals from a large range of P. cinnamomi isolates. Even the most promising RR line had individual replicates that were unable to contain lesions or died with time. This suggests that further screening work may be required using more isolates varying in their capacity to cause disease and a broader range of environmental conditions. Jarrah clonal lines that survive such rigorous screening could then be expected to survive planting out in a range of environments in the jarrah forest and rehabilitated bauxite minesites.

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16

Dunne, Christopher P. "Control of sudden death in cultivated proteas from the Southwest of Western Australia /." Access via Murdoch University Digital Theses Project, 2004. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20041207.140807.

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17

Jayasekera, Arunodini Uthpalawanna. "Interactions between Phytophthora cinnamomi and Acacia pulchella : consequences on ecology and epidemiology of the pathogen /." Murdoch University Digital Theses Program, 2006. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20061129.134500.

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18

au, N. Williams@murdoch edu, and Nari Michelle Anderson. "DNA methods for the detection of Phytophthora cinnamomi from soil." Murdoch University, 2006. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20070820.130155.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip ¥ microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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19

Sampaio, Ana Rita Brito Chedas. "Selecção de plantas com efeito alelopático para controlar Phytophthora cinnamomi." Master's thesis, ISA/UL, 2017. http://hdl.handle.net/10400.5/13857.

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Mestrado em Engenharia Agronómica - Protecção das Plantas - Instituto Superior de Agronomia - UL O declínio dos Montados de sobro e azinho é uma doença que tem sido descrita desde a década de 80 do século passado e é influenciada pela interacção de factores bióticos e abióticos. Inúmeros estudos mostram uma associação entre espécies do género Phytophthora e o declínio dos Montados, sendo que Phytophthora cinnamomi é a espécie isolada com mais frequência nos solos desses ecossistemas. P. cinnamomi é um patogénio do solo, da classe Oomycota, que causa podridão radicular e, consequentemente, a morte da planta. A sua erradicação dos solos é muito difícil. Os produtos fitofarmacêuticos utilizados até ao momento não apresentam eficácia no seu controlo, pois o patogénio encontra-se disseminado nos solos e apresenta uma elevada gama de hospedeiros. Tais condicões favorecem a sua sobrevivência. Dado que a luta química para o controlo de P. cinnamomi tem-se mostrado ineficaz, é necessário procurar alternativas mais sustentáveis. Este trabalho teve como objectivo apresentar um primeiro contributo para a selecção de plantas com um efeito alelopático sobre P. cinnamomi. Para tal, seleccionaram-se doze espécies das seguintes famílias: Fabaceae, Poaceae, Lamiaceae e Brassicaceae. Avaliou-se a susceptibilidade das espécies selecionadas a P. cinnamomi em ensaios em estufa, tendo-se observado que três espécies foram infectadas. Realizaram-se ensaios in vitro de modo a testar os extractos aquosos radiculares das plantas e determinar os seus efeitos na actividade do patogénio através do seu crescimento micelial, produção de esporângios e clamidósporos bem como da viabilidade e germinação de zoósporos. Como resultado, seleccionou-se as espécies Eruca sativa e Raphanus raphanistrum por inibirem quase totalmente o crescimento e desenvolvimento do patogénio, O efeito inibitório total fna actividade do patogénio foi observado no extracto combinado das duas espécies. Por outro lado, os extractos de gramíneas, em particular de Lolium rigidum, tiveram um efeito promotorr do patogénio N/A

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20

Anderson, Nari Michelle. "DNA methods for the detection of Phytophthora cinnamomi from soil." Thesis, Anderson, Nari Michelle (2006) DNA methods for the detection of Phytophthora cinnamomi from soil. PhD thesis, Murdoch University, 2006. https://researchrepository.murdoch.edu.au/id/eprint/42/.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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21

Anderson, Nari Michelle. "DNA methods for the detection of Phytophthora cinnamomi from soil." Anderson, Nari Michelle (2006) DNA methods for the detection of Phytophthora cinnamomi from soil. PhD thesis, Murdoch University, 2006. http://researchrepository.murdoch.edu.au/42/.

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This project assesses two aspects of DNA detection of Phytophthora species from soil samples. Firstly, a nested PCR protocol was established with both primary and nested PCR specific for P. cinnamomi detection. PCR amplification of P. cinnamomi DNA isolated from soil was optimised with the addition of bovine serum albumin and formamide. This was found to improve both the specificity and sensitivity of PCR amplification of DNA in the presence of inhibitors co-extracted along with the target DNA from soil samples. The application of diagnostic nested PCR with the addition of BSA and formamide was verified by comparison with routine culture based detection methods. In all cases, nested PCR detection incorporating BSA and formamide was found to be considerably more sensitive than the culture based detection methods. The second component of this thesis investigates the simultaneous detection of multiple species of Phytophthora using microarray analysis. Microarray based detection has been previously limited by variable and inconsistent hybridisation intensities across the diversity of probes used in each array. In this study a novel concept for the differentiation of detection targets using duplex melting kinetics is introduced. A microarray assay was developed on a PamChip microarray enabling the differentiation of target Phytophthora species using the melting kinetics of probe-target duplexes. In the majority of cases the hybridization kinetics of target and non-target duplexes differed significantly. Analysis of the melting kinetics of duplexes formed by probes with target and non-target DNA was found to be an effective method for determining specific hybridization and was independent of fluctuations in hybridization signal intensity. This form of analysis was more robust than the traditional approach based on hybridisation intensity, and allowed the detection of individual Phytophthora species and mixtures there of.

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22

Palmer, Bryony. "The dispersal of Phytophthora cinnamomi by the woylie (Bettongia penicillata)." Thesis, Palmer, Bryony (2009) The dispersal of Phytophthora cinnamomi by the woylie (Bettongia penicillata). Honours thesis, Murdoch University, 2009. https://researchrepository.murdoch.edu.au/id/eprint/32592/.

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23

Phillips, Tom. "Detection of Phytophthora cinnamomi from bulk water and soil samples." Thesis, Phillips, Tom (2008) Detection of Phytophthora cinnamomi from bulk water and soil samples. Honours thesis, Murdoch University, 2008. https://researchrepository.murdoch.edu.au/id/eprint/32595/.

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24

Christie, John Barry. "Determining the phenotypic resistance mechanisms in avocado against Phytophthora cinnamomi." Diss., University of Pretoria, 2012. http://hdl.handle.net/2263/31497.

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The avocado (Persea americana Mill.) is an economically important crop worldwide. The most important disease of avocado is Phytophthora root rot, which is caused by Phytophthora cinnamomi Rands. Currently, phosphonate trunk injections provide satisfactory disease control; however, the possibility of reduced sensitivity and eventually resistance to this fungicide is lurking on the horizon. Furthermore, consumer demands for “organic” fruit has been increasing over the past decade, emphasising the need to use root rot-resistant rootstocks. Due to a lack of understanding of the interaction between these two organisms, screening for specific resistant mechanisms is not possible and consequently only partially resistant rootstocks are currently commercially available. The aim of this thesis was therefore to address this need by investigating phenotypic traits in avocado rootstocks that could play a role in resistance against P. cinnamomi. Dissertation (MSc)--University of Pretoria, 2012. Microbiology and Plant Pathology MSc Unrestricted

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25

Khaliq, Ihsanul. "Range expansion of Phytophthora, particularly Phytophthora cinnamomi into colder environments: adaptation, a changing environment or both?" Thesis, Khaliq, Ihsanul (2019) Range expansion of Phytophthora, particularly Phytophthora cinnamomi into colder environments: adaptation, a changing environment or both? PhD thesis, Murdoch University, 2019. https://researchrepository.murdoch.edu.au/id/eprint/43119/.

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Alpine and sub-alpine regions were long considered free of Phytophthora species, especially Phytophthora cinnamomi due to restrictions on their growth from low temperatures. However, P. cinnamomi was isolated from a sub-alpine area ‘Barrington Tops National Park’ in the 1990s. Subsequent Australia wide surveys detected 68 Phytophthora species in Australia. Of these, 33 Phytophthora species, including P. cinnamomi, were detected in the alpine and sub-alpine areas on Kosciuszko National Park (KNP) alone. This suggested that Phytophthora species had adapted to cold environments. This project investigated the ability of Phytophthora species to produce infective propagules (zoospores) and cause disease at increasingly lower temperatures. Phytophthora cinnamomi was selected as a ‘test’ species due to its national and international significance. Initially, preliminary surveys were conducted in the sub-alpine and alpine areas of KNP and Tasmania to obtain living Phytophthora isolates. The lower temperature limit for growth and sporulation of Mediterranean (one isolate was from a sub-alpine area) P. cinnamomi isolates was determined and phenotypic plasticity experiments were established in an attempt to ‘train’ them to produce infective propagules and cause disease at increasingly lower temperatures. Finally, the distribution patterns of Phytophthora and vascular plants species in relation to disturbance and elevation were determined across elevation gradients in KNP. Preliminary surveys resulted in the isolation of eight Phytophthora species, including two new species that were formally described. Phytophthora cinnamomi was shown to produce infective propagules at temperatures lower (7.5 °C) than originally established (10 °C), and in a shorter time compared to original isolates when ‘trained’ under cold conditions. This suggests that P. cinnamomi responds rapidly to selection pressure and adapts to new environments. Although P. cinnamomi produced infective propagules at 7.5 °C, the pathogen could not be isolated from plants grown at 7.5 °C after three months. Therefore, more work is required to establish disease development at 7.5 °C and below. Results of surveys along elevation gradients showed Phytophthora and vascular plant species exhibited a linearly monotonic decline with increasing elevation on roads, but not in native vegetation. However, the elevation range of Phytophthora species was higher than vascular plants on both roads and in native vegetation. Phytophthora species did not show any habitat preference and exhibited similar composition and frequency on roads and in native vegetation; vascular plants showed the opposite trend with greater frequency in native vegetation. This suggests that Phytophthora richness at the plot level mimics that of vascular plants. A changing climate may permit invasion by other Phytophthora species not yet present.

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26

Schoenbaum, Elizabeth. "Genotypic Characterization of Phytophthora cinnamomi from Ornamental Crops in North Carolina." NCSU, 2008. http://www.lib.ncsu.edu/theses/available/etd-11042008-100454/.

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Forty-two Phytophthora cinnamomi isolates from Camellia spp., Ilex spp., Juniperus spp., and Rhododendron spp. were characterized for mating type, mefenoxam fungicide sensitivity, and aggressiveness on Rhododendron âHino de Giriâ. Isolates collected from Camellia spp. were of the A1 mating type, while isolates from the other host plants were A2. All isolates were sensitive to mefenoxam at 100 ppm and all but one was sensitive at 1 ppm. Isolates from Rhododendron spp. scored higher average foliar disease and root rot ratings, while A1 isolates from Camellia spp. had the lowest average foliar disease and root rot ratings. The population sample of 42 isolates was also examined for DNA sequence polymorphisms in two nuclear loci, beta-tubulin (Btu) and a portion of the intergenic spacer (IGS) region of the nuclear rDNA repeat, and one mitochondrial DNA locus, cytochrome c oxidase subunit 1 (COX 1). Six base substitutions were found among the 42 isolates with a multi-locus data set. Isolates grouped into four haplotypes. Haplotype grouping corresponded to isolate mating type, plant host, and heterozygosity in the Btu locus. Our inferred multilocus rooted gene genealogy revealed a putative ancestral lineage representing the most frequently sampled haplotype in the population. This haplotype contained A2 isolates collected from Ilex spp., Juniperus spp., and Rhododendron spp.. Isolates of the A1 mating type diverged more recently in the genealogy. There is an increase in heterozygosity at the Btu locus that coincides with the appearance of the A1 mating type. These findings increase our understanding of the population structure of P. cinnamomi.

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27

Monteiro, Brígida Trigo de Miranda Strecht. "Interacção in vitro entre Quercus suber L. e Phytophthora cinnamomi Rands." Doctoral thesis, Universidade de Aveiro, 2007. http://hdl.handle.net/10773/979.

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Doutoramento em Biologia Este trabalho circunscreve-se na área da fitopatologia e tem por objectivo principal a contribuição para o levantamento de alguns mecanismos relacionados com a resistência e a susceptibilidade de Quercus suber ao agente etiológico Phytophthora cinnamomi. Para a iniciação de culturas in vitro de Q. suber propõe-se uma abordagem de desinfecção superficial por aplicação de Peróxido de Hidrogénio. Tanto do ponto de vista quantitativo, como qualitativo, este método produziu resultados melhores em termos de indução e manutenção de porções aéreas e de tecido caloso de Q. suber in vitro (p < 0,001) e taxas de descontaminação sempre superiores a 88,8%. Os tecidos vegetais cresceram melhor em meio nutritivo de Gresshoff e Doy modificado. Por aplicação de uma combinação de dois métodos para produção de zoósporos e duas estirpes patogénicas, obtiveram-se suspensões de zoósporos de P. cinnamomi. O método se CHAMBERS et al. (1995) e a estirpe H1000 contribuíram com os melhores resultados (104 zoósporos.mL-1). A infecção das culturas (plântulas micropropagadas, porções aéreas e tecido caloso) forneceu quadros sintomatológicos de infecção em tudo semelhantes ao que sucede na interacção in vivo. Foram eleitos entre os clones de tecido caloso disponíveis um resistente (proveniente de Montemor-o-Novo) e um susceptível (proveniente de Ponte-de-Sôr). Foram analisados alguns parâmetros químicos e bioquímicos (Cl- , SO4 2-, NO3 - , NO2 - , HPO4 3-, F- , ião oxalato, Na+ , K+ , NH4 + , Mg2+, Ca2+ por electroforese capilar e perfis peptídicos em electroforese de geles de poli-acrilamida em condições desnaturantes) após a interacção do tecido caloso (a crescer em meios com diferentes composições hormonais) com os zoósporos de P. cinnamomi. Na presença do agente patogénico as quantidades dos iões NH4 + , NO2 - , NO3 - e F- eram vestigiais, dos iões K+ , Ca2+ e Na+ diminuíam, do ião Mg2+ mantinhamse, mais ou menos estáveis, dos iões Cl- e SO4 2- diminuíam no tecido resistente e mantinham-se constantes no tecido susceptível, e do anião HPO4 2- mantinham-se constantes no tecido resistente e diminuíam no tecido susceptível. Para o tecido susceptível os ganhos em número de bandas são maiores entre os 205-100 kDa e para o tecido resistente entre os 13-5 kDa. Nos pesos moleculares entre 100-60 kDa, 60-40 kDa e 40-13 kDa, o número de bandas é sempre superior no tecido resistente e este é o que apresenta maiores perdas ao longo da interacção. Correlacionando o número de bandas dos perfis peptídicos com as concentrações em iões foram obtidas três correlações positivas (Mg2+/40-13 kDa; Cl- /100-60 kDa e Cl- /60-40 kDa) e duas negativas (K+ /13-5 kDa e Oxalato/205-100 kDa). Neste modelo de interacção foi encontrada maior relevância nas variações do número de bandas nos perfis peptídicos (60-40 kDa>40-13 kDa>100-60 kDa>205-100 kDa>13-5 kDa), seguida da relevância dos catiões (K+ >Na+ >Mg2+>Ca2+) e dos aniões (SO4 2->Cl- >HPO4 2 ). Igualmente relevante o número de bandas entre 13 a 5 kDa e a concentração em ião oxalato (com contributos com 52,08% e 60,99%, respectivamente). This is a work in the area of the phytopathology and the main goal is to contribute to the finding of some mechanisms related to the resistance and the susceptibility of Quercus suber to the pathogen Phytophthora cinnamomi. To initiate Q. suber in vitro cultures the application of hydrogen peroxide as a superficial disinfection agent is proposed. This method was better from the qualitative and quantitative point of view to the induction and maintenance of Q. suber in vitro cultures (p < 0,001) and the decontamination level was over 88%. The tissues grew better in modified Gresshoff and Doy medium. P. cinnamomi zoospores suspensions were obtained by combining two zoospores production methods and two pathogenic isolates. The bests results were achieved with CHAMBERS et al. (1995) method and H1000 isolate (104 zoospores.mL-1). The infection of the cultures (micropropagated plantlets, shoots and calli) showed several symptomatologic degrees of infection comparable to the in vivo interaction. One calli clone resistant (from Montemor-o-Novo) and other susceptible (from Ponte-de-Sôr) were elected from the available clones. Some chemical and biochemical parameters (Cl - , SO 4 2-, NO 3 - , NO 2 - , HPO 4 3-, F - , oxalate ion, Na + , K + , NH 4 + , Mg2+, Ca2+ by capillary electrophoresis and the polypeptide profile determination by denaturant polyacrilamide gels electrophoresis) were analysed after calli cultures (growing in different hormonal compositions) and P. cinnamomi zoospores interaction. In the presence of the pathogen the quantities of NH 4 + , NO 2 - , NO 3 - and F- ions were almost nulls, K + , Ca2+ and Na+ ions diminished, Mg2+ ion maintained more or less stable, Cl - and SO 4 2- ions diminished in the resistant tissue and maintained constant in the susceptible tissue, and HPO 4 2- ion maintained constant in the resistant tissue and diminished in the susceptible tissue. Gains in the number of bands in the challenged susceptible tissue were grater between 205-100 kDa and in the susceptible tissue between 13-5 kDa. The number of bands in the molecular weights between 100-60 kDa, 60-40 kDa and 40-13 kDa was always superior in the resistant tissue and this always showed grater losses along the interaction. By correlating the number of band obtained in SDS PAGE with the ion concentrations we obtained tree positive correlations (Mg2+/40-13 kDa; Cl - /100-60 kDa and Cl - /60-40 kDa) and two negative correlations (K + /13-5 kDa and Oxalate/205-100 kDa). In this interaction model grater relevancy was found in the polypeptide profile bands numbers variations (60-40 kDa > 40-13 kDa > 100-60 kDa > 205-100 kDa > 13-5 kDa), followed by the relevancy of the cations (K + > Na + > Mg2+ > Ca2+) and of the anions (SO 4 2- > Cl - > HPO 4 2 ). The band number between 13-5 kDa and the oxalate concentration were equally important (with 52,08% and 60,99% contributions, respectively).

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Stasikowski, Patricia. "Biochemical effects of phosphite on the phytopathogenicity of Phytophthora cinnamomi Rands." Thesis, Stasikowski, Patricia (2012) Biochemical effects of phosphite on the phytopathogenicity of Phytophthora cinnamomi Rands. PhD thesis, Murdoch University, 2012. https://researchrepository.murdoch.edu.au/id/eprint/14887/.

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Phosphite, a chemical analogue of orthophosphate, controls disease symptoms and spread of Oomycete plant pathogens, particularly those caused by Phythophthora spp. Phosphite can be applied to horticultural and native plant species as a foliar spray or trunk injection and results in in planta phosphite concentrations of between 25 - 425 μg g-1 dry weight (equivalent to 0.3 – 6.0 mM). However, despite its extensive use it is not known why phosphite is biostatic towards oomycetes, although several mechanisms have been proposed. This thesis aims to devise and test a biochemical model of phosphite action that could account for the observed effects of phosphite on the interaction between Phytophthora cinnamomi and a susceptible plant host. However, prior to this it was necessary to devise a test to assess the concentration of phosphite in planta and to establish that phosphite needs to be present at the plant /pathogen interface in order to have an effect. A silver nitrate staining method was developed and its ability to detect phosphite was assessed in a variety of native Australian and horticultural plants. The method demonstrated that phosphite concentrations of between 1 and 3 mM were present in the tips of the roots of lupins that had been foliar sprayed with 0.5 % phosphite (equivalent to 62 mM) and that in most instances, these concentrations were sufficient to completely control the development of disease symptoms. As phosphite is chemically similar to orthophosphate its presence in a cell is likely to interfere with many aspects phosphate metabolism in both plant and pathogen. In order to discern the mechanism of action of phosphite it was important to separate the antipathogenic effects from the general/ pleotropic effects. A bioassay was devised whereby the roots of lupin seedlings were inoculated with filter paper discs that had been colonised with P. cinnamomi isolate MP94-48 and then treated with phosphite or other chemicals that would be expected to reduce its pathogenicity. The extent of lesion development and the root growth below the point of inoculation were the two parameters by which the effect of the chemicals on pathogenicity was assessed. Increasing either the concentration of orthophosphate (0 – 100 mM) or phosphite (0 – 10 mM) in the growth medium of P. cinnamomi colonised discs reduced lesion development on inoculated lupin seedling roots. Orthophosphate concentrations of between 3 – 10 mM, in combination with 1 mM phosphite did not reduce the extent of lesion development. In contrast, plants inoculated with discs treated with concentrations of orthophosphate above 10 mM together with 3 and 10 mM phosphite, lesions were reduced when compared to plants inoculated with discs treated with phosphite alone. The inhibition of phosphatase activity in P. cinnamomi is often proposed to be a primary effect of phosphite. Treatment of P. cinnamomi colonized discs with the phosphatase inhibitors okadaic acid, sodium fluoride, and a mixture of inhibitors containing sodium vanadate, sodium molybdate, sodium tartrate and imidazole, neither decreased nor increased the development of lesions, and no change in the degree of phosphorylation of cytosolic proteins could be detected by Pro-Q Diamond phosphoproteins staining. The addition of the kinase inhibitor staurosporine (0.1 - 1 mM) reduced lesion development on lupins and this effect was augmented slightly, but significantly, by the addition of phosphite (3 mM). It was not possible to draw any conclusions from the results of experiments testing the effect of addition of exogenous cAMP or the phosphatase inhibitor phenyl arsine oxide to colonised discs on the ability of P. cinnamomi to produce lesions in lupins. These results suggest that phosphorylation reactions and cascades may not be the primary control mechanism in either initiation or inhibition of phytopathogenesis. However, the addition of glucose (30 mM) increased pathogenicity and the development of lesions. As evidence exists that abscisic acid (ABA) increases the susceptibility of plants to infection by Phytophthora spp. and that ABA signaling involves phospholipase D (PLD) the effect of inhibitors on this signaling pathway were tested on the ability of P. cinnamomi to produce lesions. Primary, 2o and 3o butyl alcohol, as well as the guanine nucleotide exchange factor (GEF) inhibitor brefeldin A, and ABA itself were added to cultures of P. cinnamomi. The application of either 1o, 2o or 3o butyl alcohol to P. cinnamomi colonised discs had no effect on lesion development, which would be expected were the generation of phosphatidic acid per se was vital to pathogenicity. However, Brefeldrin A (10 - 250 μM) had a highly significant and concentration dependent effect on the development of lesion on lupins. These results suggested that a member of the Ras-superfamily (such as an ADP ribosylation factor) is likely to be involved in the development of lesions and that the exchange of GDP for GTP on this protein is required for pathogenesis. The results from the bioassays of addition of exogenous ABA to P. cinnamomi colonised discs were ambiguous and additional experimentation is needed to elucidate the role of ABA in the phytopathogenesis of P. cinnamomi. Calcium ion signatures and cytosolic gradients are known to be important components of many signal transduction pathways and increased soil calcium can limit the development of Phytophthora disease. The effect of external calcium ion concentration and the calcium channel blockers ruthenium red (RR), lanthanum chloride (La3+) and the calcium ion chelator EGTA on the development of lesions was investigated. The results of the bioassays indicated that external calcium ion concentration, RR and La3+ (100 μM) reduced lesion development significantly as did EGTA (1 mM) and that this reduction was further enhanced in the presence of phosphite. The combined role of phosphite and external calcium ion concentration was further investigated in a glasshouse pathogenicity trial using P. cinnamomi and the Australian plant Banksia leptophilia in a factorial nested pot design with foliar phosphite and soil calcium sulphate concentration as independent variables. The results one year post-inoculation confirmed that when foliar phosphite (0.1% - 0.3%) was used in conjunction with soil supplementation with calcium sulphate (3 – 30 mM) disease symptoms and lesion development were significantly reduced and general plant health was improved. The combined results of these experiments suggested not only a role for calcium ion concentration and signaling in pathogenicity but, together with the 35-fold increase in PPi concentration, imply that inhibition of the calcium dependent ATPase responsible for regulating cytosolic Ca 2+ concentration may be the cause of the antipathogenic effect of phosphite in P. cinnamomi. Calcium dependent ATPases are known to be involved in the gravitropic response of roots as well as the polar growth of pollen tubes (i.e. presence of the Ca2+ channel blocker La3+ results in inhibition of the gravitropic and polar response). Preliminary results of the effect of phosphite on the gravitropism of lupin seedling roots indicate that phosphite does inhibit the gravitropic response, suggesting that there is a causal link between the mechanism of action of phosphite and calcium-dependent ATPases.

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29

Kunadiya, Manisha B. "New tools for the detection of Phytophthora cinnamomi in environmental samples." Thesis, Kunadiya, Manisha B (2018) New tools for the detection of Phytophthora cinnamomi in environmental samples. PhD thesis, Murdoch University, 2018. https://researchrepository.murdoch.edu.au/id/eprint/45563/.

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Phytophthora cinnamomi (Rands) is one of the world’s most invasive plant pathogens and accurate diagnosis of its presence in plants and soil using molecular tools is very important. Few of the existing primers were found to discriminate between P. cinnamomi and a number of newly described species, and for those that could do so, the sensitivity was inadequate. Further, for my research, primers to detect both DNA and RNA were required, and the existing primers based on non-protein coding gene regions were inappropriate. New primers were developed based on the cytochrome oxidase subunit 2 (cox2) gene, a mitochondrial gene without introns and suitable for the RT-qPCR assay and applicable to both DNA and RNA. Procedures were modified to minimize loss of nucleic acids during extraction. These primers were specific for P. cinnamomi and able to detect as little as 150ag DNA. An exception was the closely related P. parvispora, which showed late amplification at high DNA concentrations. Primers were successfully used to detect infection in plant materials and in a range of soil types. The rate of decay of P. cinnamomi DNA and RNA in different soil types, under wet or dry conditions were also studied. P. cinnamomi DNA can survive in soil with no living host plant roots, for 378 days or more if the soil is dry, but only up to 90 days if it is wet. P. cinnamomi RNA can persist in soil for only 3 days or less in both dry and wet soil; in wet silty loam it could not be recovered after ~30 minutes. The clay content of the soil also affected the survival time of the DNA. Although RNA analysis is very accurate for the detection of living P. cinnamomi, the high cost of the analysis makes it impractical for widespread use at present. The new primers have already been adopted by the Centre for Phytophthora Science and Management as part of a best-practice protocol used to determine is P. cinnamomi is still present following eradication activities on Alcoa mine sites.

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30

Gouveia, Maria Eugénia. "Doença da tinta do castanheiro. Avaliação da resistência a Phytophthora cinnamomi Rands." Master's thesis, Universidade Técnica de Lisboa, Instituto Superior de Agronomia, 1993. http://hdl.handle.net/10198/4321.

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É apresentada uma nova metodologia de avaliação da resistência do castanheiro 11 Phytophthora cinnamomi, fungo mais frequentemente associado 11 doença da tinta. Envolve a inoculação de micelio de P. cinnamomi em ramos destacados de castanheiro, quando os lançamentos apresentam 30-40cm de crescimento e a incubação em condições adequadas de temperatura e humidade a que se segue a avaliação da dimensão da lesão. JNICT

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31

Gutierrez, Rodriguez Edwin Antonio. "Tolerância a Phytophthora cinnamomi de portaenxertos de abacateiro e propagação in vitro /." Jaboticabal, 2016. http://hdl.handle.net/11449/144407.

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Orientador: Renata Aparecida de Andrade Coorientador: Rita de Cássia Panizzi Banca: Priscila Lupino Gratão Banca: Eduardo Custódio Gasparino Banca: Tatiana Eugenia Cantuarias Avilés Banca: Simone Rodrigues da Silva Abstract: The tests in this study aimed to approach the answer to the question : The progeny of avocado tolerant rootstocks keeps the tolerance to to Phytophthora cinnamomi. Moreover aimed at addressing issues related to the in vitro establishment of explants of Duke 7 and Toro canyon cultivars. Resumo: Os testes relacionados neste trabalho objetivaram se aproximar da resposta da pergunta: A progênie de matrizes de abacateiro tolerantes a Phytophthora cinnamomi mantém a tolerância dos parentais. Alem disso visaram abordar aspectos relacionados à introdução in vitro de explantes dos materiais Duke 7 e Toro canyon. Doutor

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32

Dobrowolski, Mark Paul. "Population and sexual genetics of Phytophthora cinnamomi in Australia using microsatellite markers." Thesis, Dobrowolski, Mark Paul (1999) Population and sexual genetics of Phytophthora cinnamomi in Australia using microsatellite markers. PhD thesis, Murdoch University, 1999. https://researchrepository.murdoch.edu.au/id/eprint/3327/.

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Phytophthora cinnamomi is a plant pathogen that causes dieback disease in southern Australia. It threatens the biodiversity of many natural ecosystems due to the susceptibility of the native vegetation. If methods of control are to be successful then we must appreciate the genetic variation in the pathogen and the ways in which this variation is generated. Previously, the only genetic markers available to study P. cinnamomi were isozymes, which showed that isolates in Australia were one of three isozyme types. In this thesis I describe the development of microsatellite DNA markers for P. cinnamomi. Five microsatellites were successfully developed into markers for the nuclear genome and protocols for their use were established. Research into microsatel1ites for the mitochondrial genome is also presented though this was unsuccessful in providing markers useful for population genetic studies. The developed micro satellite markers were used to study inheritance in sexual progeny of four P. cinnamomi crosses. All but one of 201 progeny germinated were outcrosses. A large amount of non-Mendelian inheritance of the microsatellite alleles was observed. This could be explained by a high frequency of imperfect meiosis (e.g., nondisjunction, unequal crossing over) leading to additions and deletions in the chromosome complement of the sexually derived progeny. A population genetic study of three intensively sampled P. cinnamomi disease fronts located in southwest Australia is also presented. A total of 647 isolates were analysed from these hierarchically sampled sites with the micro satellite markers along with 133 culture collection isolates from across Australia. This analysis revealed that P. cinnamomi in Australia consists of three clonal lineages, with no sexual reproduction evident, even though both mating types co-occur. However, within these clonal lineages I found evidence for frequent mitotic recombination (mitotic crossing over). This mechanism for producing genetic variation may explain phenotypic variation known to occur within the identified clonal lineages.

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33

Maurel, Marion. "Impact écophysiologique sur jeunes chênes et châtaigniers de l'infection racinaire par Phytophthora cinnamomi." Paris 11, 2001. http://www.theses.fr/2001PA112157.

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L'impact des infections racinaires par p. Cinnamomi sur l'écophysiologie du châtaignier, du chêne pédoncule, du chêne rouge et du chêne vert a été étudié en conditions semi-controlées. L’intensité des effets observés s'explique essentiellement par le niveau de dégâts racinaires. Parmi les chênes, moins sensibles a p. Cinnamomi que le châtaignier, les plus fortes réponses physiologiques ont été observées chez le chêne vert et les moins importantes chez le chêne pédoncule. Les effets provoqués sont une baisse de la conductance stomatique et de la transpiration avec, à plus long terme, un ajustement de la biomasse aérienne. Chez le châtaignier, la conductance stomatique et la transpiration baissent linéairement avec la sévérite des dégâts racinaires. La surface foliaire et le rapport entre biomasse de racines saines et biomasse aérienne baissent a un seuil d'environ 50% de racines nécrosées. Par contre, le potentiel hydrique foliaire, la conductance hydraulique spécifique sol-feuille et la photosynthèse ne baissent que lorsque le système racinaire est presque totalement détruit. Chez le chêne vert, la sévérite des dégâts racinaires a été plus difficile à estimer et est moins clairement corrélée à l'intensité des effets observés. Le découplage entre la baisse de conductance stomatique et celle du potentiel hydrique de base et de la conductance hydraulique suggère l'existence d'un signal racinaire transmis jusqu'aux feuilles. L’hypothèse de l'intervention d'elicitines ou de l'acide abscissique a été testée par des tests biologiques sur feuilles de châtaignier excisées et des dosages d'acide abscissique dans la sève. Les elicitines ne semblent pas jouer le rôle de molécule signal mais l'intervention de l'acide abscissique n'est pas à exclure. Par ses effets négatifs sur le fonctionnement hydrique de l'arbre et son effet d'affaiblissement à long terme, p. Cinnamomi pourrait être, dans les processus de dépérissement, un facteur de prédisposition, notamment à la sécheresse

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Lucas, Anne. "Water stress and disease development in Eucalyptus marginata (jarrah) infected with Phytophthora cinnamomi." Thesis, Lucas, Anne (2003) Water stress and disease development in Eucalyptus marginata (jarrah) infected with Phytophthora cinnamomi. PhD thesis, Murdoch University, 2003. https://researchrepository.murdoch.edu.au/id/eprint/167/.

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The south-west of Western Australia has a Mediterranean climate and flora endemic to this area, including the keystone species, jarrah (Eucalyptus marginata), have adapted to the droughted summer conditions. The introduction of an exotic soil borne pathogen, Phytophthora cinnamomi, has challenged the survival of this and many other species. The expectation might be that plants stressed by drought are more susceptible to disease and this study examined the development of disease caused by P. cinnamomi in E. marginata and the significance of water status to that development. Seedlings of E. marginata, clonal plants resistant to P. cinnamomi and clonal plants susceptible to P. cinnamomi, were subjected to different watering regimes in a number of field and glasshouse experiments. To determine the level of drought stress that could be imposed on container-grown E. marginata seedlings without killing them, a preliminary experiment progressively lowered the moisture levels of the substrate in their containers, until the plants reached wilting point, at which time moisture was restored to a predetermined droughted level and the process repeated. With each subsequent droughting the wilting point was lower until it was found that the seedlings could survive when only 5% of the moisture lost from container capacity to wilting point was restored. No deaths had occurred after seedlings had been maintained at this low level for 14 days (Chapter 2). Based on these findings, the level of droughting maintained in all experiments conducted under controlled glasshouse conditions was 10% restoration. After testing the appropriateness of underbark inoculation, and a zoospore inoculation method for which no wounding was necessary, a new, non-invasive stem inoculation technique was developed. Stems were moistened in a pre-treatment, then agar plugs colonized with P. cinnamomi mycelium were held against the stem with wads of wet cotton wool and bound in place with tape. This technique resulted in a high proportion of infection in E. marginata (Chapter 4) without the need for underbark inoculation or the use of zoospores (Chapter 3). It was successfully used in a large field trial in a rehabilitated bauxite mine site with 2-year-old E. marginata clonal plants, resistant to P. cinnamomi (Chapter 5). Inoculation was in late spring after the winter and spring rainfall. This timing was to allow comparison of disease development in stressed plants under normal droughted summer conditions compared with itsdevelopment in non-stressed, irrigated plants. However, two months after inoculation, the area was deluged with unseasonal and abnormally heavy summer rainfall, negating any difference in the treatments and causing an outbreak of P. cinnamomi in the soil from an adjacent infested site. This resulted in the infection and death of some noninoculated control clones. Monitoring of the site continued for twelve months and the advance of P. cinnamomi at the site was mapped. To test the effect of drought on the expression of P. cinnamomi under more controlled conditions, a series of glasshouse experiments was set up that simulated two possible summer conditions; drought or drought followed by abnormally high summer rainfall. These experiments utilised E. marginata seedlings and clonal plants, some resistant and some susceptible to P. cinnamomi. Plants were inoculated with P. cinnamomi prior to or after droughting. Results were compared to those of control plants that had not experienced water deficit. In both seedlings and clonal plants, the greatest extent of colonization was found in plants which had experienced no water deficit. These results indicated that drought stress played a role in inhibiting the in planta development of P. cinnamomi in all genotypes (Chapter 8). This finding was consistent for both clones, susceptible and resistant to P. cinnamomi. Most recoveries were made from non-stressed clonal plants, resistant to P. cinnamomi (Chapter 6) and more colonization was found in non-stressed clonal plants, susceptible to P. cinnamomi (Chapter 7), than was recorded for droughted plants. The results of the field trial showed that P. cinnamomi was not recovered from some inoculated stems, which had obvious lesions, when segments were plated onto selective agar. This led to an intensive in vitro investigation into improved methods of recovery. Dark brown exudates from some segments of inoculated stems stained the surrounding agar onto which they were plated, suggesting the presence of phenolic compounds. Recovery of the pathogen from stems increased by about 10% when segments were first soaked in distilled water to leach out the phenolic compounds, then replated onto agar. Other recovery methods were also tested, including (1) baiting with Pimelea ferruginea leaves floated on the surface of water or soil filtrate, in which the infected stem segments were immersed and (2) the application of different light and temperature regimes. It was clearly shown that exudates from infected stems of field grown E. marginata inhibited the outgrowth of P. cinnamomi onto the agar. To counter the possible toxic effect that oxidized phenolics had on the growth of the P. cinnamomi, an antioxidant was added to the agar. P. cinnamomi was grown on media whichincorporated exudates from infected stems and different concentrations of ascorbic acid, with and without adjusted pH levels. There was a pronounced pH effect, with less growth on media with lower pH and no significant increase in growth of the mycelium with increased ascorbic acid concentration on pH adjusted agar (Chapter 9). The inhibitory effect of the exudates from the stem segments led to an investigation of the possibility that, if seedlings to be planted in the rehabilitation process could be pre-treated with phenolic compounds to render them more resistant, they may have an advantage when establishing in areas where there was a potential threat of P. cinnamomi. E. marginata seeds were germinated and the seedlings grown hydroponically in a constant temperature growth room. Different concentrations of synthetic catechol, a phenolic compound naturally occurring in E. marginata, were added to the nutrient solution. Roots remained immersed in the catechol solutions for three days, before being inoculated at the root tip with zoospores of P. cinnamomi. Roots in higher concentrations of catechol were less colonized than those in lower concentrations, indicating an increased resistance to the pathogen (Chapter 10). Further work is required to determine if seedlings treated before being planted in areas threatened by an outbreak of P. cinnamomi have a greater capacity for survival, and for how long the protection persists. The improved recovery of P. cinnamomi from infected plants is important for accurate assessment of the spread of the disease in an area and for the subsequent implementation of management strategies of containment and control. An outbreak of P. cinnamomi can impact on the revegetation of rehabilitated mine sites and the aetiology of the pathogen in mine sites needs to be more fully understood. The interaction of plant defences with the invasive pathogen has been examined in a range of environments in the field, the glasshouse, in a hydroponics system and in vitro. The results indicate that summer droughting increases the resistance of E. marginata to P. cinnamomi. However, more work is required to understand the mechanisms involved. The study also indicates that clones of E. marginata, selected as resistant to P. cinnamomi, are not resistant under all conditions and that environmental interactions should be further investigated. Lastly, for effective management strategies to be implemented it is critical that the pathogen can be confidently isolated from plants. It was shown that exudates from infected hosts inhibit the recovery of P. cinnamomi. Recovery methods that can overcome these inhibitory compounds are required. The findings invite further research into the complexity of host-pathogen relationships.

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35

Lucas, Anne. "Water stress and disease development in Eucalyptus marginata (jarrah) infected with Phytophthora cinnamomi." Murdoch University, 2003. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20040820.13290.

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Ortega, Ramírez Eddy. "Detección de Phytophthora cinnamomi en raíces de Persea americana "palto" por Nested-PCR." Bachelor's thesis, Universidad Nacional Mayor de San Marcos, 2015. https://hdl.handle.net/20.500.12672/8818.

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Publicación a texto completo no autorizada por el autor Establece una metodología de detección de ADN de Phytophthora cinnamomi en muestras de raíces de Persea americana “palto” procedente del Sector III del Proyecto Especial CHAVIMOCHIC, La Libertad-Perú. Se establece un protocolo de amplificación de regiones del espaciador transcrito interno (ITS) del ADNr por Nested-PCR, que permite el diagnóstico rápido y confiable de P. cinnamomi. Los iniciadores PPF/PPR específicos del orden Pythiales se emplean en la primera reacción de PCR (Simple-PCR), mientras que los iniciadores PcinnF/PcinnR específicos de P. cinnamomi se usan en la segunda reacción de PCR (Nested-PCR). La estandarización de las pruebas de amplificación se realiza a partir de ADN extraído de micelio en cultivo puro de la cepa FM2C1R1 aislada previamente de la zona de estudio. Se obtienen amplificaciones del ADNr de P. cinnamomi por Simple-PCR desde 100 ρg de ADN extraído de raíces; mientras que en Nested-PCR es posible desde los 10 ρg del ADN previamente amplificado. El método de extracción y amplificación a partir de ADN de raíces es suficientemente sensible para detectar al patógeno sin necesidad de mayores purificaciones del ADN extraído. La dilución del ADN extraído o el incremento de la concentración de MgCl2 en la reacción no mejoran considerablemente la visualización de amplicones. De un total de 30 muestras de raíces de palto analizadas en el área de estudio, 22 resultan positivas para el patógeno en Nested-PCR. La confiabilidad de la Nested-PCR es verificada mediante secuenciación de los productos de amplificación y análisis filogenético. Esta investigación es la primera en establecer una metodología de detección de ADN de P. cinnamomi en raíces de palto y proporciona una herramienta molecular aplicable para la prevención y monitoreo del patógeno en el cultivo de palto. Tesis

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Collins, Sarah. "Long-term survival of Phytophthora cinnamomi in rehabilitated bauxite mines and adjacent jarrah." Thesis, Collins, Sarah (2006) Long-term survival of Phytophthora cinnamomi in rehabilitated bauxite mines and adjacent jarrah. PhD thesis, Murdoch University, 2006. https://researchrepository.murdoch.edu.au/id/eprint/32426/.

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Gresser, Mark P. "Effects of thinning and burning rehabilitation on Phytophthora cinnamomi and small mammal populations." Thesis, Gresser, Mark P. (2009) Effects of thinning and burning rehabilitation on Phytophthora cinnamomi and small mammal populations. Honours thesis, Murdoch University, 2009. https://researchrepository.murdoch.edu.au/id/eprint/32758/.

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Perhaps the most important function that can be fulfilled by ecological studies is the identification of factors that threaten the survival of flora and fauna and subsequently influence their distribution and abundance. In the jarrah forest of southwestern Western Australia, one particular human activity that threatens the survival of wildlife is bauxite mining, which involves the removal of all vegetation from an area, disturbance of the soil profile and subsequent revegetation. In particular, the thinning-and-burning of revegetated minesites, which is conducted to increase water runoff in catchments and accelerate the development of natural jarrah forest structure within rehabilitation, may be having an important influence on the distribution and abundance of small mammals inhabiting the jarrah forest and the way they use their habitat. One particular small mammal that might be affected in this way is the nectivorous Western Pygmy-possum Cercartetus concinnus, of the family Burramyidae. In this study, Western Pygmy-possums were captured in pit-traps and subjected to diet analysis (by pollen swabbing and scat analysis), radio-tracking (to identify patterns of nest selection and movement) and fluorescent powder tracking (to identify patterns of microhabitat selection). The incidence and severity of Phytophthora Die back within the mining landscape and its flow-on impacts on the Western Pygmy-possum were also assessed. The aim was to determine the impacts of mining activity, particularly the thinning and burning of rehabilitation, on the Western Pygmy-possum. As the first detailed study on the Western Pygmy-possum in the jarrah forest, this study has greatly increased our current knowledge about the species, especially in relation to diet and nest selection. It appeared that Western Pygmy-possums rely on unmined forest for suitable nest sites in the form of large trees, but used rehabilitation for their nocturnal activities, which likely mostly consists of feeding. Although it was clear mining has a negative impact on the Western Pygmy-possum by reducing the abundance of suitable nest sites, there was no consistent evidence that the thinning-and-burning of restoration has an important impact. However, it was revealed that thinning-and-burning has a significant effect on vegetation structure, especially by reducing ground leaf matter and increasing understorey density. Notably, restoration that had not been thinned-and-burned was more similar to unmined forest than was thinned-and-burned restoration, and thus it was revealed that thinning-and-burning was not effective in accelerating the development of natural jarrah forest structure within restored minesites. Importantly, no significant results were obtained about the importance of structural vegetation features for the way Western Pygmy-possums use their habitat, which supports the idea that the availability of plant species that are an important source of food in the form of nectar and pollen is likely the greatest limiting factor for the distribution of the Western Pygmy-possum. There was no evidence that Phytophthora Dieback affects the Western Pygmy-possum, but this was most likely because there was a lack of susceptible species in rehabilitation. While the results of this study have important implications for the future management of minesites in the jarrah forest, in particular by highlighting the importance of unmined forest as a source of nest sites and revealing that thinning-and-burning is not achieving its aims, this study has wider significance. In particular, the information gained from this study will be of increasing importance in Western Australia as water becomes a scarcer resource. Climate change predictions are that rainfall and water run-off will continue to decline in south-west Western Australia over the next 100 years and consequently thinning is increasingly being considered as a way of boosting the State's water supply.

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Sykes, Melissa. "Do zoospores of Phytophthora cinnamomi produce enzymes such as cutinases, cellulases and pectinases?" Thesis, Sykes, Melissa (1995) Do zoospores of Phytophthora cinnamomi produce enzymes such as cutinases, cellulases and pectinases? Honours thesis, Murdoch University, 1995. https://researchrepository.murdoch.edu.au/id/eprint/32817/.

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Bekker, Theo Frederik. "Efficacy of water soluble silicon for control of phytophthora cinnamomi root rot of avocado." Diss., Pretoria : [s.n.], 2007. http://upetd.up.ac.za/thesis/available/etd-09172007-084901.

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41

Gouveia, Maria Eugénia. "Métodos moleculares na identificação caracterização e detecção de Phytophthora cambivora (Petri) Buisman e Phytophthora cinnamomi Rands associadas com a Doença da Tinta do Castanheiro." Doctoral thesis, UTAD, 2004. http://hdl.handle.net/10198/4292.

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Um total de 49 isolados de Phytophthora spp. associados com a doença da tinta do castanheiro foram obtidos, pelos métodos clássicos de isolamento, em diferentes situações de crescimento do castanheiro na região Norte de Portugal: viveiros, plantações jovens, soutos adultos e do solo. Os isolados de Phytophthora foram identificados por PCR-RFLP da região ITS-rDNA amplificada pelos “primers” universais ITS6 (Cooke & Duncan, 1997) e ITS4 (White et al., 1990). A restrição enzimática do fragmento amplificado foi realizada com as enzimas Alu I, Msp I e Taq I cujo perfil de restrição permite a identificação ao nível da espécie no género Phytophthora. A população de Phytophthora associada com a doença da tinta foi caracterizada quanto ao tipo de compatibilidade sexual, polimorfismo RAPD e sensibilidade ao metalaxil. P. cinnamomi é a espécie predominante em viveiro, plantações jovens, soutos adultos e no solo junto das plantas mortas. Apenas quatro isolados foram identificados como P. cambivora e dois deles foram obtidos em plantas de castanheiro com sintomas característicos da doença no colo da planta. Todos os isolados (P. cambivora e P. cinnamomi) são A2 e 9 % dos isolados de P. cinnamomi evidenciaram sensibilidade reduzida ao metalaxil. A caracterização RAPD permitiu a separação das espécies de Phytophthora associadas à doença da tinta embora os primers “10-mer” utilizados não tivessem possibilitado a caracterização dos isolados de P. cinnamomi. O estudo RAPD permitiu, no entanto, identificar um fragmento de amplificação característico de P. cinnamomi. O fragmento foi purificado clonado e sequenciado (SCAR) e desenharam-se primers específicos (Cin1, Cin2 e Cin3) para a detecção e identificação de P. cinnamomi por PCR. Os primers utilizados na combinação Cin2/Cin1 e Cin3/Cin1 proporcionam um produto de amplificação PCR com aproximadamente 650 pb. Com esta metodologia de diagnóstico não se detectaram reacções cruzadas com as espécies de P. cactorum, P. capsici, P. cinnamomi var. parvispora, P. citricola, P. cryptogea, P. nicotianae, P. quercina, P. syringae e ainda com Cryphonectria parasitica, Pythium sp. e Castanea sativa. O método PCR-Diagnóstico foi aplicado com sucesso, em raízes de castanheiro, que cresceram em substrato inoculado com P. cinnamomi, em tecidos vegetais (folhas de castanheiro) e na água da técnica armadilha de detecção de Phytophthora no solo. A total of 49 isolates of Phytophthora spp. associated with ink disease of chestnut were obtained by classical methods from diverse chestnut growing regions of the north of Portugal: nurseries, young and old chestnut groves, and also in the soil near dead plants of sweet chestnuts. Phytophthora isolates were identified by PCR-RFLP of ITS-rDNA amplified by the universal primer pair ITS6 (Cooke & Duncan, 1997) and ITS4 (White et al., 1990). Restriction digestions of the amplified DNA product were performed with restriction enzymes Alu I, Msp I, and Taq I, which allowed individual species identification. The population of Phytophthora was characterised in terms of mating type, metalaxil resistance and RAPD polymorphisms. P. cinnamomi is the most prevalent species in nurseries, orchards and in soil of chestnut groves. Only four isolates were identified as P. cambivora and two of them were obtained from typical rot crown of chestnut. All the isolates (P. cambivora and P. cinnamomi) were mating type A2 and 9 % of P. cinnamomi were metalaxil tolerant. Although RAPD polymorphisms allowed species differentiation the “10-mer” primers used were not suitable for P. cinnamomi characterization. The RAPD analyses allowed, however, the identification of a fragment characteristic of P. cinnamomi. The fragment was purified, cloned and sequenced (SCAR) and then species-specific primers were designed (Cin1, Cin2, and Cin3) for detection and identification of P. cinnamomi by PCR. Each primer pairs Cin2/Cin1 and Cin3/Cin1 produced an amplicon of approximately 650 bp from all tested P. cinnamomi isolates. Both primer pairs revealed no undesirable cross-reaction with a diverse range of Phytophthora species (P. cactorum, P. capsici, P. cinnamomi var parvispora, P. citricola, P. cryptogea, P. nicotianae, P. quercina, P. syringae), Cryphonectria parasitica, Pythium sp., and Castanea sativa. The PCR-Diagnostic assay based on the species-specific primers was successfully used to detect P. cinnamomi in roots of chestnut plants, which were grown in artificially infected substrate with this pathogen, in bait tissue (chestnut leaves) and in the water of the same bait test. Parcialmente FCT SFRH/BD/1474 e Programa PRODEP Acção 5.3

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Aberton, Michael J., and lswan@deakin edu au. "The use of phosphite as a control for Phytophthora cinnamomi in southeastern Victorian vegetation communities." Deakin University. School of Biological and Chemical Sciences, 2005. http://tux.lib.deakin.edu.au./adt-VDU/public/adt-VDU20060921.150649.

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One of the major aims of the research presented in this thesis was to assist managers of native vegetation communities in southeastern Australia in understanding the dynamics of P. cinnamomi with an important ecological species, Xanthorrhoea australis. It trialed the use of phosphite in large-scale field applications to establish the usefulness of this management option for the first time on Victorian flora. This thesis describes the process of disease development within mature X. Australia plants. For the first time it was shown that within X. australis plants, secondary disease symptoms are related to the percentage of stem that has been infested by the disease. It was evident that after initial invasion the pathogen moves via root xylem and throughout the plant within vascular to the stem, especially within the desmium. The research shows that the pathogen could not be isolated consistently even though it was considered to be responsible for disease symptoms. Trials of a control fungicide (Foli-R-fos 200) shows that protection occurs in many susceptible plants when 2 and 6g a.i./L phosphite is applied. Phytotoxicity occurred in native plants at Anglesea and within controlled environment trials when using ≥ 6g a.i./L. It will be shown that 2g a.i./L phosphite controls disease in sprayed plots within heathlands at Anglesea and a recently burnt coastal woodland community at Wilsons Promontory. The proportion of healthy X. australis plants treated with phosphite was significantly higher than the proportion in control plots without phosphite. The research shows that phosphite was recovered from leaves of three species treated with Foli-R-fos 200 in the field. For the first time it has been shown that seed germination was reduced in two species when high concentrations of phosphite were applied. The first documentation of the effect that phosphite has on soil properties showed that nitrogen and oxidised organic carbon were the only parameters to alter significantly. This thesis provides answers to some important questions, answers that can now be used by managers in formulating better policies and actions at an operational level. There has been a dire need in Victoria to address many issues regarding P. cinnamomi and this thesis provides relevant and informative approaches to disease control, and a better understanding of the disease progress.

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Daniel, Rosalie, and mikewood@deakin edu au. "Aspects of the interaction between Xanthorrhoea australis and Phytophthora cinnamomi in south-western Victoria, Australia." Deakin University. School of Biological and Chemical Sciences, 2002. http://tux.lib.deakin.edu.au./adt-VDU/public/adt-VDU20051201.144848.

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Diseases in natural ecosystems are often assumed to be less severe than those observed in domestic cropping systems due to the extensive biodiversity exhibited in wild vegetation communities. In Australia, it is this natural biodiversity that is now under threat from Phytophthora cinnamomi. The soilborne Oomycete causes severe decline of native vegetation communities in south-western Victoria, Australia, disrupting the ecological balance of native forest and heathland communities. While the effect of disease caused by P. cinnamomi on native vegetation communities in Victoria has been extensively investigated, little work has focused on the Anglesea healthlands in south-western Victoria. Nothing is known about the population structure of P. cinnamomi at Anglesea. This project was divided into two main components to investigate fundamental issues affecting the management of P. cinnamomi in the Anglesea heathlands. The first component examined the phenotypic characteristics of P. cinnamomi isolates sampled from the population at Anglesea, and compared these with isolates from other regions in Victoria, and also from Western Australia. The second component of the project investigated the effect of the fungicide phosphonate on the host response following infection by P. cinnamomi. Following soil sampling in the Anglesea heathlands, a collection of P, cinnamomi isolates was established. Morphological and physiological traits of each isolate were examined. All isolates were found to be of the A2 mating type. Variation was demonstrated among isolates in the following characteristics: radial growth rate on various nutrient media, sporangial production, and sporangial dimensions. Oogonial dimensions did not differ significantly between isolates. Morphological and physiological variation was rarely dependant on isolate origin. To examine the genetic diversity among isolates and to determine whether phenotypic variation observed was genetically based, Random Amplified Polymorphic DNA (RAPD) analyses were conducted. No significant variation was observed among isolates based on an analysis of molecular variance (AMQVA). The results are discussed in relation to population biology, and the effect of genetic variation on population structure and population dynamics. X australis, an arborescent monocotyledon indigenous to Australia, is highly susceptible to infection by P. cinnamomi. It forms an important component of the heathland vegetation community, providing habitat for native flora and fauna, A cell suspension culture system was developed to investigate the effect of the fungicide phosphonate on the host-pathogen interaction between X. australis and P. cinnamomi. This allowed the interaction between the host and the pathogen to be examined at a cellular level. Subsequently, histological studies using X. australis seedlings were undertaken to support the cellular study. Observations in the cell culture system correlated well with those in the plant. The anatomical structure of X australis roots was examined to assist in the interpretation of results of histopathological studies. The infection of single cells and roots of X. australis, and the effect of phosphonate on the interaction are described. Phosphonate application prior to inoculation with P. cinnamomi reduced the infection of cells in culture and of cells in planta. In particular, phosphonate was found to stimulate the production of phenolic material in roots of X australis seedlings and in cells in suspension cultures. In phosphonate-treated roots of X australis seedlings, the deposition of electron dense material, possibly lignin or cellulose, was observed following infection with P. cinnamomi. It is proposed that this is a significant consequence of the stimulation of plant defence pathways by the fungicide. Results of the study are discussed in terms of the implications of the findings on management of the Anglesea heathlands in Victoria, taking into account variation in pathogen morphology, pathogenicity and genotype. The mode of action of phosphonate in the plant is discussed in relation to plant physiology and biochemistry.

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Mahomed, Waheed. "Sequencing ESTs of the avocado transcriptome to study the tolerant response to Phytophthora cinnamomi." Diss., University of Pretoria, 2012. http://hdl.handle.net/2263/31150.

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Avocado (Persea americana Mill.) is an important crop whose cultivation is severely threatened by Phytophthora cinnamomi Rands. The South African avocado industry makes an important contribution to the world’s avocado supply, and is one of the world’s largest exporters. If the current Phytophthora root rot problem is not addressed soon, the losses encountered by the avocado industry may become so extensive that it results in job losses. The scant information that is available for P. cinnamomi interaction studies indicate that there is no gene-for-gene interaction yet described between the pathogen and host. Avocado genomics are not well understood either and there is not much sequence data available for this basal angiosperm. The data available comprises of sequence that was generated in marker studies on fruit and flowering organs. It is now possible to generate large amounts of sequence data using highthroughput sequencing platforms and identify defence-related genes. The identification of defence-related genes in a tolerant rootstock will allow us to characterize the avocado-P. cinnamomi interaction on a molecular level. The aim of this MSc was to identify defence-related genes in a tolerant rootstock and characterize their expression in order to understand the avocado-P. cinnamomi interaction. Chapter 1 provides a comprehensive overview of the advances in molecular work conducted on avocado thus far. A background of avocado rootstock development is provided with details of molecular markers developed for use in avocado. Additionally, an introduction is also given to high-throughput sequencing and its application to non-model crops such as avocado. Chapter 2 describes the mRNA isolation and EST pyrosequencing of avocado roots. Gene annotation of metabolic, cell wall associated and stress response genes are provided along with the characterisation of defence-related genes. Chapter 3 reports of the expression profiling of defence-related genes obtained from avocado root ESTs. The expression of nine defence-related genes are studied over six time points in P. cinnamomi infected R0.09 tolerant avocado roots. Chapter 4 provides a general discussion of the result obtained in this study along with future applications of the sequencing data produced. Dissertation (MSc)--University of Pretoria, 2012. Genetics MSc Unrestricted

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Eshraghi, Leila. "Genetic analysis of host and phosphite mediated resistance to Phytophthora cinnamomi in Arabidopsis thaliana." Thesis, Eshraghi, Leila (2012) Genetic analysis of host and phosphite mediated resistance to Phytophthora cinnamomi in Arabidopsis thaliana. PhD thesis, Murdoch University, 2012. https://researchrepository.murdoch.edu.au/id/eprint/12737/.

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Phosphite (Phi), an analogue of phosphate (Pi) is highly effective for the control of Phytophthora cinnamomi, a devastating necrotrophic pathogen worldwide. This study describes the effect of phosphite (Phi) on the induction of defence responses in Phytophthora cinnamomi-infected Arabidopsis thaliana accessions Ler and Col-0, and mutants defective in salicylic acid (SA), jasmonic acid (JA), ethylene (ET), abscisic acid (ABA), phosphate starvation response (PSR) and auxin response signalling pathways. The inoculation of the resistant Col-0 with P. cinnamomi induced a rapid increase in callose deposition (by 6 h after inoculation) and hydrogen peroxide (H2O2) production (by 24 h after inoculation) whereas inoculation of susceptible Ler showed a delayed and reduced response. Treatment of Ler with Phi produced a response to P. cinnamomi inoculation similar to that observed in Col-0 in terms of timing and magnitude suggesting Phi primes the plant for a rapid and intense response to infection involving heightened activation of a range of defence responses. A reliable method for measuring disease progression is important when evaluating susceptibility in host–pathogen interactions. A sensitive quantitative polymerase chain reaction (QPCR) assay was developed for the quantitative measurement of P. cinnamomi DNA (biomass) in planta that avoids problems caused by variation in DNA extraction efficiency and degradation of host DNA during host tissue necrosis. Purified plasmid DNA, containing the pScFvB1 mouse gene, was added during DNA extraction and the pathogen’s biomass was normalized based on plasmid DNA rather than host DNA or sample fresh weight. It was demonstrated that normalization of pathogen DNA to sample fresh weight or host DNA in samples with varying degrees of necrosis led to an overestimation of the pathogen’s biomass. Inoculation of mutants in the SA, JA, and ET defence signalling pathways did not affect the resistance of Col-0 suggesting alternative pathways are involved. A high level susceptibility was observed in the aba2-4 mutant suggesting a role for ABA signalling in the induction of resistance to P. cinnamomi. Phi treatment of aba2-4 increased resistance but not to the wild type levels indicating a possible role for ABA-dependent and ABA independent signalling in Phi mediated resistance. Application of Phi to non-inoculated A. thaliana seedlings elevated transcription of defence genes in the SA (PR1 and PR5) and JA/ET (THI2.1 and PDF1.2) pathways. Furthermore, analysis of gene expression in Col-0 revealed that either Phi or P. cinnamomi caused the down-regulation of the transcriptional level of AtMYC2 (a positive regulator of ABA signalling which also negatively regulates JA-related genes) and increased the transcriptional abundance of PDF1.2. Together these results suggest that the resistance response of Col-0 and Phi treatment both act partially through an ABA dependent mechanism which is independent of the antagonism between ABA and elements of the JA/ET pathway such as PDF1.2. Phosphite has been suggested to interfere with various plant processes including Pi homeostasis therefore the potential involvement of the Pi and auxin signalling pathways in resistance to P. cinnamomi was investigated using several PSR and auxin response pathway mutants. The mutants tir1-1, an auxin response mutant deficient in the auxin-stimulated SCF (Skp1−Cullin−F-Box) ubiquitination pathway and phr1-1, a mutant defective in response to Pi starvation were highly susceptible to P. cinnamomi compared to their parental background Col-0. Complementation restored resistance to the level observed in Col-0. Moreover, inhibition of auxin transporters by TIBA (2,3,5-triiodobenzoic acid) led to a significant increase in susceptibility of Lupinus angustifolius seedlings to P. cinnamomi supporting the importance of the auxin signalling pathway in P. cinnamomi resistance. The 26S proteasome subunits mutants; rpn10-1 (Defective in ubiquitin/26S proteasome-mediated proteolysis) and pbe1-1 (proteasome subunit beta type-5-A) were also susceptible to P. cinnamomi. The rpn10-1 mutant has also been associated with the auxin signalling pathway and the susceptibility of rpn10-1 and pbe1-1 indicates that the 26S proteasome and auxin signalling could play a role in resistance to P. cinnamomi. Given the apparent involvement of auxin and PSR signalling in the resistance to P. cinnamomi, the possible involvement of these pathways in Phi mediated resistance was also investigated. Application of Phi at both low and high concentrations attenuated some of the Pi starvation inducible genes such as At4, AtACP5 and AtPT2. However, in phosphate sufficient plants, Phi treatment mimicked Pi starvation responses in terms of enhanced expression of PHR1, AUX1, AXR1, AXR2 and SGT1B; suppression of primary root elongation, and increased root hair formation. Together, these results suggest that the auxin response pathway, particularly auxin sensitivity and transport, plays a role in the plant’s resistance to P. cinnamomi and suggest that phosphite-mediated resistance may in some part be through its effect on stimulation of the auxin response pathway.

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Simmons, Donna. "The impact of Phytophthora cinnamomi on reptile communities in banksia woodlands of Western Australia." Thesis, Simmons, Donna (2011) The impact of Phytophthora cinnamomi on reptile communities in banksia woodlands of Western Australia. Honours thesis, Murdoch University, 2011. https://researchrepository.murdoch.edu.au/id/eprint/32587/.

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47

Favas, Melissa. "The role of active compounds produced by actinomycetes in the control of Phytophthora cinnamomi." Thesis, Favas, Melissa (1994) The role of active compounds produced by actinomycetes in the control of Phytophthora cinnamomi. Honours thesis, Murdoch University, 1994. https://researchrepository.murdoch.edu.au/id/eprint/32819/.

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The overall aim of this project was to screen actinomycetes for the ability to produce antibiotics inhibitory to P. cinnamomi in vitro. Some of the more promising isolates were then screened in vivo in a glasshouse pot-trial. The actinomycete isolates used had been previously isolated from jarrah forest soils suppressive to P. cinnamomi, ie. areas where P. cinnamomi was present in the soil but there was little evidence of disease. They had been isolated using dry heat pretreatments ( 120°C for one hour) designed to select for less frequently isolated actinomycetes especially the genus Microbispora. Isolates were screened against P. cinnamomi in vitro for the production of secondary metabolites inhibitory to this pathogen. Of the isolates screened 40% caused strong inhibition of P. cinnamomi , 13% caused medium inhibition and 46% caused no inhibition of P. cinnamomi. Of the 40% of isolates that caused inhibition in vitro only ±4.0% of those tested for inhibition of P. cinnamomi in the pot-trial showed a potential for inhibiting P. cinnamomi in soil. The pot-trial was performed using actinomycete isolates that had shown different levels of inhibition in vitro. Eight actinomycetes that had produced an antibiotic inhibitory to P. cinnamomi in vitro were tested in the pot-trial, 6 showing strong inhibition and 2 medium inhibition, but only 2 of these isolates a Streptomyces, isolate 97, and a maduromycete, isolate 55A, suppressed P. cinnamomi in vivo. In addition 2 of the 4 isolates with no in vitro inhibition of P. cinnamomi showed some activity against P. cinnamomi in the pot-trial. The results emphasised the fact that the production of an antibiotic in vitro does not automatically mean that the isolate will have the ability to control the pathogen in vivo. The actinomycete isolates 97 and 55A which had shown strong inhibition in the in vitro screening were also screened against a range of fungal plant pathogens from different taxonomic groups. The fungal pathogens included representatives from the ascomycetes, hyphomycetes, basidiomyeetes and oomycetes. Eight other species of Phytophthora were also included. Isolate 97 showed the ability to strongly inhibit all of the fungal pathogens except Pythium ultimum which was only partially inhibited. Isolate 55A did not have the ability to inhibit Pythium ultimum but did inhibit all other pathogens to different degrees. Both isolates though showed that the antibiotics they produced were active over a range of different fungal taxonomy groups. The metabolites from Streptomyces isolate 97 which showed strong activity against a range of fungal pathogens were extracted and purified from a broth culture filtrate. The antibiotic compound did not lose its ability to inhibit P cinnamomi through the extraction or purification process. Two antibiotic compounds were extracted, YB1 and YB2, both of which were inhibitory to P cinnamomi although YB2 was the stronger of the two. Antibiotics YBI and YB2 were also inhibitory to a few bacterial plant pathogens increasing the range of pathogens that could be inhibited by isolate 97. Due to the wide range of pathogens inhibited by Streptomyces isolate 97 and its metabolites an attempt was made to further identify the isolate to species. There was not sufficient time to determine all the data required to identify the isolate by numerical classification, which requires data from 139 unit characters but a simple outdated key was used in the attempt. The use of the older key did not result in the identification of the isolate but the isolate was characterised morphologically and with respect to carbon source utilisation.

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com, kathrynmccarren@hotmail, and Kathryn McCarren. "Saprophytic ability and the contribution of chlamydospores and oospores to the survival of Phytophthora cinnamomi." Murdoch University, 2006. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20060807.92625.

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Phytophthora cinnamomi has been recognised as a key threatening process to Australias biodiversity by the Commonwealths Environment Protection and Biodiversity Conservation Act 1999. Despite over 80 years of extensive research, its exact mode of survival is still poorly understood. It is widely accepted that thin- and thick-walled chlamydospores are the main survival propagules while oospores are assumed to play no role in the survival of the pathogen in the Australian environment, yet evidence is limited. The saprophytic ability of the pathogen is still unresolved despite the important role this could play in the ability of the pathogen to survive in the absence of susceptible hosts. This thesis aimed to investigate chlamydospores, oospores and the saprophytic ability of P. cinnamomi to determine their contribution to survival. Phytophthora cinnamomi did not show saprophytic ability in non-sterile soils. The production of thick-walled chlamydospores and selfed oospores of P. cinnamomi in vitro was documented. Thick-walled chlamydospores were sporadically formed under sterile and non-sterile conditions in vitro but exact conditions for stimulating their formation could not be determined. The formation of thick-walled chlamydospores emerging from mycelium of similar wall thickness was observed, challenging the current knowledge of chlamydospore formation. Selfed oospores were abundant in vitro on modified Ribeiros minimal medium in one isolate. Three other isolates tested also produced oospores but not in large numbers. Although the selfed oospores did not germinate on a range of media, at least 16 % were found to be viable using Thiozolyl Blue Tetrazolium Bromide staining and staining of the nuclei with 4´, 6-diamidino-2-phenylindole.2HCl (DAPI). This indicated the potential of selfed oospores as survival structures and their ability to exist dormantly. The ability of phosphite to kill chlamydospores and selfed oospores was studied in vitro. Results challenged the efficacy of this chemical and revealed the necessity for further study of its effect on survival propagules of P. cinnamomi in the natural environment. Phosphite was shown to induce dormancy in thin-walled chlamydospores if present during their formation in vitro. Interestingly, dormancy was only induced by phosphite in isolates previously reported as sensitive to phosphite and not those reported as tolerant. Chlamydospores were produced uniformly across the radius of the colony on control modified Ribeiros minimal medium but on medium containing phosphite (40 or 100 µg ml-1), chlamydospore production was initially inhibited before being stimulated during the log phase of growth. This corresponded to a point in the colony morphology where mycelial density changed from tightly packed mycelium to sparse on medium containing phosphite. This change in morphology did not occur when the pathogen was grown on liquid media refreshed every four days, and chlamydospores were evenly distributed across the radius of these colonies. This trend was not observed in selfed oospores produced in the presence of phosphite. Selfed oospore production was found to be inhibited by phosphite at the same concentrations that stimulated chlamydospore production. Isolates of P. cinnamomi were transformed using a protoplast/ polyethylene glycol method to contain the Green Fluorescent Protein and geneticin resistance genes to aid in future studies on survival properties of the organism. Although time constraints meant the stability of the transgene could not be determined, it was effective in differentiating propagules of the transformed P. cinnamomi from spores of other microrganisms in a non-sterile environment. Two different sized chlamydospores (approximately 30 µm diameter and < 20 µm diameter) were observed in preliminary trials of transformed P. cinnamomi inoculated lupin roots floated in non-sterile soil extracts and these were easily distinguished from microbial propagules of other species. The growth and pathogenicity was reduced in two putative transformants and their ability to fluoresce declined over ten subcultures but they still remained resistant to geneticin. This study has improved our knowledge on the survival abilities of P. cinnamomi in vitro and has provided a useful tool for studying these abilities under more natural glasshouse conditions. Important implications of phosphite as a control have been raised.

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Coelho, Valentim. "Efeito do fosfonato de potássio na protecção das raízes do castanheiro (Castanea sativa Mill.) contra Phytophthora cinnamomi." Master's thesis, Instituto Politécnico de Bragança, Escola Superior Agrária, 2009. http://hdl.handle.net/10198/1200.

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A Doença da Tinta do Castanheiro, é considerada como uma das principais causas do desaparecimento dos soutos. Phytophthora cinnamomi e P. cambivora são as duas espécies associadas à Doença da Tinta do Castanheiro, sendo P. cinnamomi a espécie preponderante na doença da tinta em Portugal. Os agentes patogénicos que causam a Doença da Tinta no Castanheiro provocam uma situação de difícil solução, pois estes possuem formas de sobrevivência e de disseminação que lhes permite manterem-se no solo quase indefinidamente. Os meios de luta disponíveis para combater as doenças provocadas por Phytophthora, não têm, até hoje, resolvido de forma eficiente e duradoura os problemas sanitários das culturas e das florestas atacadas por estes parasitas. Actualmente os fosfitos, sais ou os ésteres do ácido fosforoso, por estarem associados com os mecanismos biológicos de resistência e induzirem na planta mecanismos de defesa são uma forma alternativa no combate contra P. cinnamomi. Com este trabalho estudou-se o efeito da aplicação foliar de fosfonato potássico em plantas jovens de castanheiro que cresceram em substrato inoculado com P. cinnamomi. Estudou-se ainda um método indirecto, por inoculação de P. cinnamomi na parte aérea da planta, para determinar o efeito protector nas raízes e o efeito in vitro do fosfonato potássico e fosetil-Al em diferentes isolados de Phytophthora e outros fungos associados com o castanheiro. Os resultados obtidos mostram que a aplicação foliar do fosfonato potássico, protegeu as raízes dos castanheiros, não evidenciando as planta tratadas com fosfonato potássico sintomas da Doença da Tinta. Todas as plantas que cresceram em substrato inoculado com P. cinnamomi e sem aplicação foliar de fosfonato potássico evidenciaram sintomas característicos da doença com epinastia e necrose das folhas. A análise estatística evidenciou diferenças significativas entre tratamentos em todos os parâmetros relacionados com as raízes. O peso seco das raízes secundárias foi o parâmetro fisiológico mais afectado, tendo as plantas sem tratamento com fosfonato potássico menor comprimento radicular e menor numero de raízes assim como grande extensão de podridão radicular. A protecção conferida pelo fosfonato de potássio avaliada por inoculação de P. cinamomi na parte aérea da planta revelou que o comprimento da lesão é superior nas plantas não tratadas com fosfonato potássico ao contrário do verificado em plantas não tratadas tendo sido considerado uma metodologia adequada para avaliar o efeito protector da substância utilizada. A análise da toxicidade in vitro revelou que os valores de EC50 variam entre 0,64 μg/ml e 31,56 μg/ml para P. cinnamomi e 9,92 μg/ml e 22,44 μg/ml para P. cambivora. O fosetil-Al apresentou baixa toxicidade in vitro nas diferentes espécies de Phytophthora. Chestnut Ink Disease is considered one of the most important causes of the disappearance of the chestnut orchards. The two associated species to the chestnut ink disease are Phytophthora cinnamomi and P. cambivora, being the first one the foremost important cause of this disease in Portugal. The pathogenic agents related to the ink disease in chestnut bring out a situation of complex resolution, due to their survival and spreading ways, that allow them to remain in the soil almost indefinitely. The available control means against diseases caused by Phytophthora, haven’t been able, so far, to resolve, in a long-lasting and efficient way the health problems of crops and forests infected by these parasites. Currently, phosphites, salts or esters of phosphorous acid, due to their relation to the biological resistance mechanisms, as well as their ability to induce defense mechanisms in plants, are an alternative way to control P. cinnamomi. The aims of this work are to evaluate the effect of foliar application of potassium phosphonate in young plants of chestnut in the radicular protections against Phytophthora. An indirect method was also tested, by P. cinnamomi inoculation in the aerial part of the plant, to determine the protective effect on roots and in vitro effect of potassium phosphonate and fosetil-Al in different Phytophthora isolates and other fungi associated with the chestnut. The achieved results showed that the plants treated with foliar application of potassium phosphonate didn´t show the symptoms of ink disease, leading to the conclusion that this product did protect the chestnut roots against this disease. All the plants grown in substrate inoculated with P. cinnamomi and without foliar application of potassium phosphonate showed symptoms of the disease with epinasty and necrosis of leaves. A statistic analysis provides significant differences between treatments in all the root related parameters. Dry root weight was the most affected physiological parameter, and the plants without treatment with potassium phosphonate have lower root length and lower number of roots. The potassium phosphonate protection action, evaluated with the inoculation of P. cinamomi in branches of the plant revealed that the length of the lesion is higher in plants not treated with potassium phosphonate due to the lack of the protection granted by this compound, thus proving to be an adequate methodology to evaluate the protective effect of the used substance. The in vitro toxicity analyses revealed EC50 ranging from 0,64 mgL-1 to 31,56 mgL-1 for P. cinnamomi and 9,92 mgL-1 to 22,44 mgL-1 for P. cambivora. Fosetyl-Al showed low toxicity in vitro in different species of Phytophthora.

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Jackson, Tania. "Role of host defences in controlling the growth of Phytophthora cinnamomi in phosphite treated clonal Eucalyptus marginata plants resistant and susceptible to P. cinnamomi." Thesis, Jackson, Tania (1997) Role of host defences in controlling the growth of Phytophthora cinnamomi in phosphite treated clonal Eucalyptus marginata plants resistant and susceptible to P. cinnamomi. Honours thesis, Murdoch University, 1997. https://researchrepository.murdoch.edu.au/id/eprint/32814/.

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