The Nature of the Resistance of Oats to the Take-all Fungus: II. INHIBITION OF GROWTH AND RESPIRATION OF OPHIOBOLUS GRAMINIS SACC. AND OTHER FUNGI BY A CONSTITUENT OF OAT SAP

Growth of Ophiobohu gramimt and of O. gramims var. avenae isinhibited by concentrations of 3·3-4·0µg./ml.,and respiration by concentrations of 55µg./nil. of a partiallypurified substance from oat-leaf sap. The two varieties appearto be equally sensitive. The filtrate of boiled sap is inhibitorybut here dilution of the sap permits better growth of isolatesof var. avenae. Sap from oat roots is inhibitory to O.graminisonly, and fractionation of the sap shows that the inhibitorcan be masked by a growth stimulant. Inhibition of growth andrespiration can be reduced by glutathione and ascorbic acid,particularly if the inhibitor and reducing agent are previouslyincubated together for a few hours, suggesting that the inhibitoris inactivated on reduction. The capacity of var. avenae toovercome inhibition in the favourable medium provided by thecrude sap more readily than can the type variety is suggestedas the cause of the slight differential activity of the filtrateof leaf sap and the full differential activity of the root sap.Susceptibility of oats to var. avenae would thus be due to conditionsenabling the fungus to overcome toxicity rather than to an absenceof toxicity. Activity of the inhibitor against growth and respiration ofa number of fungi and a few other organisms has been tested.Bacteria and oat and barley roots are not affected but abouthalf of the fungi tested are inhibited although none is as sensitiveas O. gramims. No members of the fungi imperfecti tested aresensitive.     

An Enzymic Basis for Pathogenic Specificity in Ophiobolus graminis

Avenacin, the glucosidic inhibitor present in oat roots, isacted on by a specific glucosidase produced by Ophiobolus-graminisvar. avenae, which destroys biological activity, as does hydrolysiswith 0.1 N. HCl. Neither O. graminis itself nor any other fungustested produces this enzyme. It is suggested that the resistanceof oats to O. graminis and its susceptibility to var. avenaedepend on an inhibitor-inactivating enzyme complex, and thisis compared with the antibiotic-inactivating enzyme complexfound in penicillin-producing moulds and resistant penicillinase-producingbacterial strains; such a complex may be concerned in othercases of pathogenic specificity. The way in which a varietyof O. graminis pathogenic to oats may have arisen is discussed.     

Time-course of Biosynthesis of Phenolics and Lignin in Roots of Wheat Genotypes Differing in Manganese Efficiency and Resistance to Take-all Fungus

Inhibition of lignin biosynthesis in Triticum aestivum L. rootsby Mn deficiency has been suggested as the mechanism of reducedresistance of Mn-deficient wheat roots to infection by the take-allfungus (Gaeumannomyces graminis var. tritici). This study evaluatedphenolics and lignin accumulation in roots of wheat genotypesdiffering in Mn efficiency (measured as growth and yield inMn-deficient soils) and take-all resistance. Seedlings of theMn-inefficient, take-all sensitive genotype Bayonet and theMn-efficient, more take-all resistant genotype C8MM were grownin nutrient solution without added Mn for 18 d and then transferredto a Mn-deficient sandy soil fertilized with Mn at 0 or 30 mgkg^-1. Both genotypes had Mn-deficient roots and shoots at thetime of transfer to the soil. Roots of both genotypes were inoculatedwith the take-all fungus 0, 1, 3 and 7 d after transfer. Twenty-fourhours after inoculation, take-all fungus penetrated the rootstele of take-all sensitive Bayonet but not of more resistantC8MM wheat. Rates of phenolics and lignin accumulation in rootsdeclined steadily during growth in soil for up to 8 d, werehigher in mature, fully differentiated parts of the root systemcompared to distal, younger root tissue, and were higher inBayonet than in C8MM. Manganese fertilization did not significantlyinfluence rates of phenolics and lignin accumulation but reduceddepth of radial penetration by hyphae in both genotypes. Therate of phenolics accumulation was positively (r = 0·91to 0·96) correlated with the rate of lignin accumulation.Mn-efficient C8MM had a higher rate of lignin accumulation perunit of phenolics than Mn-inefficient Bayonet over a wide rangeof phenolics synthesis rates. From this we suggest that C8MMhas a more efficient mechanism for conversion of phenolics tolignin, the trait which appears related to higher take-all resistanceof this genotype.Copyright 1994, 1999 Academic Press Gaeumannomyces graminis var. tritici, lignin, manganese, phenolics, resistance, roots, Triticum aestivum     

Failure of in vitro growth inhibition by cysteine to differentiate between Australian Gaeumannomyces graminis isolates pathogenic and non-pathogenic to oats

Radial growth of oat and non oat-attacking Australian isolates of Gaeumannomyces graminis was greatly inhibited by increasing concentration of DL-cysteine in basal medium agar, and growth was completely inhibited by cysteine concentrations of 3 μM. As a group, isolates of G. graminis var. tritici (both oat and non oat-attacking forms) were more inhibited than isolates of G.graminis var.avenae at 1 μM cysteine, but differences did not occur at other concentrations. Isolates of a lobed-hyphodiate fungus similar to G. graminis var. graminis were more tolerant of cysteine than other isolates. The findings indicate that in vitro inhibition of Australian G. graminis isolates by cysteine is not useful for differentiation between oat and non oat-attacking types, and is unlikely to be fundamentally related to the ability of isolates to attack oats.     

Infection process of Gaeumannomyces graminis var. graminis on the roots and culms of rice

Crown sheath rot, caused by the ascomycete Gaeumannomyces graminis var. graminis that infects the root and the base of the culm of rice, causes early grains maturation, tiller death and reduced yield. As a paucity of information exists in the literature on the rice‐G. graminis var. graminis interaction at the microscopic level, this study aimed to gain novel insights into the infection process of this pathogen in the root and culm of rice using both light and scanning electron microscopy. In the roots, the fungus initially colonized the epidermal, exodermal and sclerenchyma cells. At 15 days after inoculation (dai), fungal hyphae colonized the cortex and clusters of perithecia were observed in the roots. At 20 dai, the fungus reached the central cylinder, and an intense fungal colonization at the base of the culm was observed that resulted in the formation of a mycelial mat on both adaxial and abaxial surfaces of the leaf sheaths. At 25 dai, fungal growth was noticed in the parenchyma cells, vascular bundles and airspaces. Perithecia emerged through the base of prophyllum and from the first leaf sheath at 30 dai. The results of this study provide new insights into the infection process of G. graminis var. graminis in rice and may help to find better control measures in reducing crown sheath rot development.     

Transformation of Gaeumannomyces graminis with β‐Glucuronidase Reporter and Hygromycin Resistance Genes

Take‐all disease is caused by Gaeumannomyces graminis, (Sacc.) Arx & D. Olivier, a soil‐borne fungus, which colonizes the root and crown tissue of many members of the Poaceae plant family. This fungus is able to grow along the surface of roots as darkly pigmented runner hyphae, which has the ability to penetrate the root. Here, we describe a genetic transformation of G. graminis var. graminis by using polyethylene glycol (PEG)‐based protoplast transformation. Fungus cells were transformed with a plasmid, pHPG, containing the gusA reporter gene that codes for β‐glucuronidase (GUS) and the hph gene for hygromycin resistance as the selectable marker. A de novo transformant selection assay was developed to identify the putative transformants that were expressing the hph gene. In addition, the transformed cells maintained the ability to infect the plant tissues. The GUS‐expressing fungus can be used to study fungal infection processes including fungal penetration, colonization and the role(s) of melanin during pathogenesis. Thus, this study is the first report of G. graminis var. graminis transformed with a visibly detectable reporter gene that provides a useful tool to a better understanding of host–Gaeumannomyces interactions.     

Pathogenicity of ‘take-all’ fungus to oats: Its relationship to the concentration and detoxification of the four avenacins

Data for inhibition of the growth of Gaeumannomyces graminis var. tritici (Ggt) and var. avenae (Gga), Phialophora radicicola and Fusarium avenaceum, caused by avenacins, are presented. The avenacins found in all oat species examined are sufficient in quantity to totally suppress growth of wheat ‘take-all’ (Ggt), even old roots containing 25 μg/g (fr. wt). Fungal variants that can also attack oats [var. avenae (Gga)] show considerable variations in their tolerance to avenacin A-1, ec₅₀ values being 5–80 μg/ml. Nevertheless, all Gga isolates maintained some growth at avenacin A-1 concentrations as high as 200 μg/ml and it is this ability to grow, albeit slowly, at high concentrations that is the critical difference between Gga and Ggt strains. The pathogenicity towards oats of a range of isolates of Gga is related to the fungicidal activity of avenacins. Gga pathogenicity is shown to increase with poor nutrition of the oat hosts (poor illumination, lack of minerals). Fungal detoxification of avenacins produces mono-deglucosylavenacin A-1, bis-deglucosylavenacin A-1 and, in one case, tris-deglycosylavenacin A-1. Ggt strains left avenacin A-1 almost unaffected giving only traces of mono-deglucosyl product. Gga strains bring about mono- and bis-deglucosylation whilst Fusarium avenaceum causes mainly bis-deglucosylation. Mono-deglucosylavenacin is shown to be less inhibitory to Gga than is avenacin A-1, whilst the bis-deglucosyl compound is still less inhibitory.     

Reduction of Take-all Inoculum by Rotation with Lupins, Oats or Field Peas

The feasibility of use of lupins, oats and field peas as alternative rotation crops to reduce inoculum of the take-all fungus (Gaeumannomyces graminis var. tritici) (under Western Australian field conditions) and disease in following wheat was investigated with a one year field trial, the soil from which was used in two succeeding pot experiments. The possible mechanisms of reduction of inoculum and disease by these crops were examined testing the soil for pathogen and disease suppression. Rotation with lupins or oats for two seasons reduced (P <0.05) inoculum of the take-all fungus and lupins, oats or field peas reduced (P <0.05) disease in following wheat. Lupins alone reduced inoculum and disease, (P <0.1) after one season. No apparent suppression of the pathogen in the absence of host plants was recorded after one season of rotation, but after two seasons, lupins, oats or field peas all suppressed (P <0.02) growth of the pathogen within soil. However only field pea soil suppressed take-all in comparison with the wheat control. Although after two seasons all rotation crops were effective in reducing inoculum and disease the mechanisms of reduction appear to differ between the rotation crops used in this study.     

Methoxyhydroquinone, a Growth Inhibitor of Ophiobolus graminis, in Leaves of Oat Seedlings

A methanol extract of leaves of oat seedlings grown in sand cultures in the dark contained a compound which inhibited the growth of Ophiobolus graminis. The inhibitory factor was isolated and proved to be present in the plant as methoxyhydroquinone glucoside. The glucoside was readily hydrolysed to the corresponding aglucone. The methoxyhydroquinone, or possibly its oxydation product, methoxy-P-benzoquinone, was inhibitory to both Ophiobolus graminis var. graminis and Ophiobolus graminis var. avenae, whereas Fusarmm oxysporum var. lycopcrsici was not affected. Synthetic methoxyhydroquinone at 80 mg/l gave a 100% inhibition of Ophiobolus graminis var. graminis. After being exposed to 80 mg/l of the inhibitor for 24 h the mycelium was unable to initiate growth when transferred to a fresh nutrient solution. Only extracts from young leaves showed inhibitory activity, extracts from mature leaves giving no inhibition. The hydroquinone, or its glucoside, was not detected in roots of young seedlings, where avenacin was the only antifungal compound present.     

A comparative study of Phialophora radicicola, an avirulent fungal root parasite of grasses and cereals

The biology and infection-behaviour of a typical isolate of Phialophora radicicola Cain have been compared with those of a representative isolate of Ophiobolus graminis (Sacc.) Sacc. Both species can utilize a nitrate source of nitrogen and both require thiamine and biotin for growth on inorganic nitro-gen; P. radicicola, but not O. graminis, was able to synthesize biotin when grown on asparagine as a nitrogen source. The pH range for good growth of P. radicicola in nutrient solution was narrower than that for O. graminis, and its growth rate on agar was only one-third. P. radicicola was the more active decomposer of cellulose, and its cellulolysis adequacy index was I.66 as com-pared with a value of 0.33 for 0. graminis. In agreement with prediction from Garrett's (I966) hypothesis on the cellulolysis adequacy index, saprophytic survival of P. radicicola in wheat straw was shortened by additional soil nitrogen, which prolongs survival of O. graminis.P. radicicola was found to spread ectotrophically over the roots of wheat, oats and barley by runner hyphae indistinguishable from those of O. graminis, but cortical infection caused no necrosis and no discernible check to growth of the infected cereals, nor any significant decrease in grain yield of inoculated wheat grown to maturity. Pre-existing infection of wheat roots by P. radicicola retarded spread of infection by O. graminis; inoculation of several grass species with P. radicicola reduced the extent of infection by O. graminis of wheat following the grasses.     

Use of a root assessment tray for the detection of take-all on lateral roots of wheat seedlings

A root assessment tray was designed for the meticulous assessment of take-all on wheat seedling roots from soil bioassays. Subsequently, the detection of lateral root infections (in addition to the more obvious infections on main axes of seminal roots) resulted in increased estimates of propagule numbers of the take-all fungus (Gaeumannomyces graminis var.tritici) for 196 of the 368 soil samples bioassayed in a field study conducted in Western Australia between 1984 and 1986.     

Water use and phosphorus and potassium status of wheat seedlings colonized byGaeumannomyces graminis orPhialophora radicicola

Summary The water consumption and levels of phosphorus, potassium, and total minerals were measured for wheat seedlings colonized byGaeumannomyces graminis var.tritici, Phialophora radicicola var.radicicola, orPhialophora radicicola var.graminicola. Infection byG. graminis resulted in a considerable reduction in water consumption, and reduced level of phosphorus when the supply of phosphorus to the seedlings was plentiful. Colonization byP. radicicola var.radicicola increased levels of phosphorus and potassium, but these increases varied according to the isolate of the fungus and the supply of phosphorus and potassium available to the seedlings. Colonization byP. radicicola var.graminicola resulted in reduced water consumption by the seedlings.The results are discussed in relation to stelar cell wall thickening in wheat roots colonized byP. radicicola, and the effects on nutrient uptake of mycorrhizal root systems.     

Effects of Salinity on Growth, Water Relations and Ion Accumulation of the Subtropical Perennial Halophyte, Atriplex griffithii var. stocksii

The effects of salinity on growth, water relations, glycinebetainecontent, and ion accumulation in the perennial halophyte Atriplexgriffithii var. stocksii were determined. The following questionswere addressed: (1) What effect does salinity have on growthresponses at different ages? (2) Is A. griffithii an ion accumulator?(3) Does A. griffithii accumulate glycinebetaine in responseto salinity? Atriplex griffithii plants were grown in pots at0, 90, 180 and 360  m M NaCl in sand culture in a plantgrowth chamber and plants were harvested after 30, 60 and 90d. Plant total dry weight was significantly inhibited at 360m M NaCl. Root growth showed a substantial promotion at 90 mM NaCl. The water potential and osmotic potential of shootsbecame more negative with increasing salinity and time of growth.The Na⁺and Cl^-content in both shoots and roots increased withincreases in salinity. Increased treatment levels of NaCl induceddecreases in Ca⁺, K⁺and Mg²⁺in plants. Atriplex griffithii accumulateda large quantity of ions, with the ash content reaching 39%of the dry weight in leaves. Inorganic ion accumulation is significantin osmotic adjustment and facilitates water uptake along a soil-plantgradient. Glycinebetaine concentration was low in roots, andin stems it increased with increases in salinity. Total amountsof glycinebetaine in leaves increased with increases in salinity,and its concentration increased substantially at 360 m M NaCl.Copyright 2000 Annals of Botany Company Atriplex griffithii, glycinebetaine, growth, ions, water relations.     

Use of Polymerase Chain Reaction To Detect the Take-All Fungus, Gaeumannomyces graminis, in Infected Wheat Plants

Gaeumannomyces graminis, the causative agent of take-all disease of wheat, barley, and oats, was detected in infected wheat seedlings by using the polymerase chain reaction to amplify Gaeumannomyces-specific DNA fragments. Nested primers and two rounds of amplification were used to amplify two fragments, approximately 287 and 188 bp in size, from G. graminis-infected wheat seedlings. The use of nested primers greatly decreased the number of nonspecific amplification products. Polymerase chain reaction products were not obtained with DNA from seedlings infected with several other phytopathogenic fungi or with DNA from uninfected seedlings. Amplified products were visualized on agarose gels, and their identities were confirmed by DNA hybridization. This method did not require culturing the fungus and has potential for detecting G. graminis in infested wheat, barley, or oat fields.     

Control of Ophiobolus patch in Agrostis turf using avirulent fungi and take-all suppressive soils in pot experiments

The incorporation of avirulent fungi such as Gaeumannomyces graminis var. graminis, an avirulent isolate of G. graminis var. tritici, a Phialophora sp. with lobed hyphopodia synonymous with Phialophora radiciola var. radicicola sensu Deacon and P. radicicola var. graminicola at the time of seeding Agrostis turf in pots of sterilised soil completely controlled Ophiobolus patch disease. The addition of a 5 mm layer of take-all suppressive (TAS) soils, artifically developed by the repeated addition of live mycelium of the varieties avenae, tritici and graminis of G. graminis to soil, controlled the disease to a lesser extent. However, a 20 mm layer of a TAS soil developed from live mycelium of G. g. avenae almost completely suppressed the disease. A survey of 66 golf and bowling greens in four states of Australia showed that P. r. graminicola was the most prevalent avirulent fungus.     

The Influence of Leaf Water Status and ABA on Leaf Growth and Stomata of Phaseolus Seedlings with Hypoxic Roots

The objective of this study was to assess the relative rolesof leaf water status and root-sourced signals in mediating beanleaf responses to root hypoxia. To do so, the roots of beanplants under varied VPD (0.95 kPa to 0.25 KPa) were made hypoxic.Under all conditions, leaf growth rates and stomatal conductanceswere reduced. There was a transitory decline in leaf water potentialat high VPD which accounted for the initial reduction in leafgrowth rates and stomatal conductance. At low VPD, no waterdeficits were detected. Leaf growth inhibition and reduced stomatalconductance under low VPD treatments were unrelated to leafwater status and must be induced by some other factor. In vitrogrowth of leaf discs was reduced by xylem sap collected fromhypoxic roots. Exogenously applied ABA, at high concentrationsin KCl and sucrose, or at low concentrations diluted in xylemsap from aerated plants, inhibited in vitro growth of leaf discs.Applications of ABA in the transpiration stream reduced stomatalconductance.     

Xylem Sap from Actinidia chinensis: seasonal Changes in Composition

Seasonal variation was followed in the content of total a-aminoacids, arginine, calcium, total carbohydrate, ß-galactosidaseactivity, magnesium, nitrate, phosphatase activity, phosphateand sulphate in vacuum-extracted xylem sap from Actinidia chinensisvar. hispida, the Chinese gooseberry or kiwifruit. There wasa marked increase in the concentration of most sap componentsjust prior to leaf emergence followed by a rapid decrease afterthe leaves had expanded. Experiments with excised extensionshoots showed that much of this spring-time increase in concentrationsof sap components was due to mobilization of nutrients withinthe shoot itself. Sap from the trunk and the older brancheschanged less in composition than did sap from the younger partsof the plant. The amplitude and direction of trends in concentrationof sap from the different parts of the plant varied with nutrientand with time. Analysis of vacuum-extracted xylem sap collectedduring periods of rapid transpiration from early summer onwardsgives a reliable indication of the composition of the transpirationstream. Actinidia chinensis, Chinese gooseberry, kiwifruit, storage reserves, xylem sap, amino acids, arginine, calcium, carbohydrate, ßgalactosidase, magnesium, nitrate, phosphatase, phosphate, potassium, sulphate     

Patterns of Root Colonization in Epacridaceous Plants Collected from Different Sites

Root colonization was studied in ten species of the Epacridaceaeat three sites in Victoria by morphological and cross-inoculationexperiments. The sites and genera chosen were Cranbourne [Epacrisimpressa Labill. andLeucopogon ericoides(Smith) R. Br.] andRye [L. parviflorus(Andrews) Lindley] on the Mornington Peninsula,and the Grampians[Astroloma conostephioides(Sond.) Benth.,A.humifusum(Cav.) R. Br.,A pinifolium(R. Br.) Benth,Brachylomadaphnoides(Smith) Benth.,E. impressa, E. impressavar.grandifloraBenth.andStyphelia adscendensR. Br.] in western Victoria. For morphologicalstudies, samples of roots from each species at each site werecleared and stained and examined microscopically. For cross-inoculationstudies, cuttings from each site were struck in potting mediuminoculated with soil from the same and other sites. The ericoidmycorrhizae in the roots of plants found at or grown in Cranbourneand Rye soils were similar. Both were significantly differentfrom the internal hyphae found in the roots of plants foundat or grown in Grampians soils, which were three times largerin diameter and formed dense coils which filled the host celland invaded adjacent epidermal cells. This suggests that morethan one fungus is involved in the relationships, that the MorningtonPeninsula sites had a different fungus from the Grampians siteand that host specificity is low. Vesicular structures werealso found commonly on plants at the Grampians site, in contrastwith other sites. Epacridaceae; root; fungus; mycorrhiza; morphology; inoculation     

Detoxification of Benzoxazolinone Allelochemicals from Wheat by Gaeumannomyces graminis var. tritici, G. graminis var. graminis, G. graminis var. avenae, and Fusarium culmorum

The ability of phytopathogenic fungi to overcome the chemical defense barriers of their host plants is of great importance for fungal pathogenicity. We studied the role of cyclic hydroxamic acids and their related benzoxazolinones in plant interactions with pathogenic fungi. We identified species-dependent differences in the abilities of Gaeumannomyces graminis var. tritici, Gaeumannomyces graminis var. graminis, Gaeumannomyces graminis var. avenae, and Fusarium culmorum to detoxify these allelochemicals of gramineous plants. The G. graminis var. graminis isolate degraded benzoxazolin-2(3H)-one (BOA) and 6-methoxy-benzoxazolin-2(3H)-one (MBOA) more efficiently than did G. graminis var. tritici and G. graminis var. avenae. F. culmorum degraded BOA but not MBOA. N-(2-Hydroxyphenyl)-malonamic acid and N-(2-hydroxy-4-methoxyphenyl)-malonamic acid were the primary G. graminis var. graminis and G. graminis var. tritici metabolites of BOA and MBOA, respectively, as well as of the related cyclic hydroxamic acids. 2-Amino-3H-phenoxazin-3-one was identified as an additional G. graminis var. tritici metabolite of BOA. No metabolite accumulation was detected for G. graminis var. avenae and F. culmorum by high-pressure liquid chromatography. The mycelial growth of the pathogenic fungi was inhibited more by BOA and MBOA than by their related fungal metabolites. The tolerance of Gaeumannomyces spp. for benzoxazolinone compounds is correlated with their detoxification ability. The ability of Gaeumannomyces isolates to cause root rot symptoms in wheat (cultivars Rektor and Astron) parallels their potential to degrade wheat allelochemicals to nontoxic compounds.     

黄绿绿僵菌对两种稻田蜘蛛捕杀褐飞虱作用的影响

在室内研究了喷施黄绿绿僵菌Metarhizium anisopliae var. acridum Driver et Milner孢子悬液对稻田蜘蛛捕杀褐飞虱作用的影响。结果表明,黄绿绿僵菌不感染拟水狼蛛Pirata subpiraticus Bsenberg et Strand和食虫沟瘤蛛Ummeliata inse cticeps Bsenberg et Strand,只对蜘蛛喷施菌液不影响它们的捕杀褐飞虱的能力。而对褐飞虱喷施黄绿绿僵菌液后,褐飞虱活力明显下降,导致蜘蛛对其捕杀效果显著提高。在喷施黄绿绿僵菌106 、107、108/mL孢子浓度后,拟水狼蛛的平均捕杀量分别为10.5头/d、11.1头/d和11.4头/d,食虫沟瘤蛛的平均捕杀量分别为3.8头/d、4.3头/d和4.7头/d,均显著大于对照组。对蜘蛛和褐飞虱同时喷施黄绿绿僵菌不影响前者的捕杀力。这些结果提示,在稻田施用黄绿绿僵菌防治稻飞虱对蜘蛛天敌没有不良影响。