Thickening and browning of cortical cell walls in seminal roots of wheat seedlings infected with Gaeumannomyces graminis var. tritici

This paper examines a little studied reaction of wheat roots to invasion by Gaeumannomyces graminis var. tritici, namely the thickening and browning of cortical cell walls. Examination was confined to distal segments of young seminal roots grown in sand. Browning and thickening of walls of cells were associated with the lignification of tissue and appeared to be a response of living or recently live cells to invasion by weakly virulent isolates. Wheat genotypes differed in their ability to thicken and lignify cell walls, and the difference between two wheat cultivars appeared to be under simple genetic control. There was some evidence that cortical browning temporarily retarded radial invasion by hyphae of G. graminis into young seminal roots of wheats.     

Evidence for resistance in wheat cultivars grown in sand culture to the take-all pathogen, Gaeumannomyces graminis var. tritici

A laboratory method to inoculate seedlings uniformly with Gaeumannomyces graminis var. tritici is described. Resistance is defined via the rate of hyphal entry into the vascular tissue of host seedlings, and is measured by direct observation and by early stelar lesion development in seminal roots. The two scores for resistance are compared and evaluated, for infection with an isolate of low virulence. Evidence was obtained for resistance in the roots of wheat seedlings to G. graminis.     

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 shading and powdery mildew infection on senescence of the root cortex and coleoptile of wheat and barley seedlings,and implications for root- and foot-rot fungi

Summary Nuclear and cytoplasmic staining methods were used to study natural senescence of the root cortex and coleoptile of wheat and barley seedlings grown in glasshouse conditions. Coleoptiles of barley senesced more slowly than those of wheat, paralleling the known difference in rates of root cortex senescence in these cereals. The coleoptiles and root cortices of both cereals senesced more slowly in shaded than in unshaded conditions, but infection of the shoots of barley byErysiphe graminis had little effect on root cortex senescence. The results are discussed in relation to infection by root- and foot-rot fungi. Previous reports on the effects of illumination on take-all infection (Gaeumannomyces graminis) are explained. It is suggested that natural senescence of the coleoptile might affect establishment of infection by the eyespot fungus,Pseudocercosporella herpotrichoides, either directly or through the activities of competing microorganisms.     

Comparison of rates of natural senescence of the root cortex of wheat (with and without mildew infection), barley,oats and rye

Summary In comparative tests in a glasshouse, the cortex of oat and rye roots senesced more slowly than the cortex of wheat and barley roots. Of the cereals tested, wheat showed the most rapid rate of root cortical senescence, and the rate was unaffected by inoculation of leaves withErysiphe graminis. The results are discussed in relation to infection by root pathogens.     

Effect of zinc supply on growth of three species of Eucalyptus seedlings and wheat

Summary Effects of zinc supply on shoot and root dry weight, root length, zinc concentrations and carbonic anhydrase activity were measured in 52 day old seedlings ofEucalyptus maculata, E. marginata, E. patens and wheat grown in a zinc deficient soil in the glasshouse.Symptoms of zinc deficiency in the eucalyptus and wheat appeared within 20 to 35 days. Eucalypt seedlings had short internodes and small necrotic leaves, reduced dry weight of shoots and roots, root length and zinc concentrations in young leaves; the measurable level of leaf carbonic anhydrase activity decreased to zero. Similar responses also occurred in wheat.The level of zinc fertilizer required for normal growth of Eucalyptus seedlings is therefore likely to be similar to that used for wheat and other agricultural crops.     

Disease in wheat genotypes naturally infected with Gaeumannomyces graminis var. tritici

Wheat genotypes consisting of seven homozygous lines and 40 segregate families were studied at two sites naturally infested with the take-all pathogen, Gaeumannomyces graminis var. tritici. The numbers of seminal and coronal roots infected with G. graminis and other root pathogens were recorded. The genotypes differed in infection with G. graminis, with little evidence of genotype × environment interactions. The incidence of G. graminis and Rhizoctonia solani was negatively associated, but the association did not greatly influence differences between wheats in infection with G. graminis. The distribution of R. solani was negatively associated with the severity of take-all at only one site. Of symptoms of infection with G. graminis, wheat genotypes differed most in incidence of deadheads, but differences were not consistent over environments, and were associated with earliness of maturity. Wheats differed more in expression of disease than in infection with G. graminis. The course of disease was deduced from associations between the incidence of pathogens and components of plant growth and yield. G. graminis was the dominant pathogen at both sites, and caused a yield loss of 0–15% at one site, and an average 62% loss at the other. More components of yield were affected where disease was most severe.     

Programmed cell death in the root cortex of soybean root necrosis mutants

The soybean root necrosis (rn) mutation causes a progressive browning of the root soon after germination that is associated with accumulation of phytoalexins and pathogenesis-related proteins and an increased tolerance to root-borne infection by the fungal pathogen, Phytophthora sojae. Grafting and decapitation experiments indicate that the rn phenotype is root-autonomous at the macroscopic level. However, the onset and severity of browning was modulated in intact plants by exposure to light, as was the extent of lateral root formation, suggesting that both lateral roots and the rn phenotype could be directly or indirectly controlled by similar shoot-derived factors. Browning first occurs in differentiated inner cortical cells adjacent to the stele and is preceded by a wave of autofluorescence that emanates from cortical cells opposite the xylem poles and spreads across the cortex. Before any visible changes in autofluorescence or browning, fragmented DNA was detected by TUNEL (T erminal deoxynucleotidyl transferase-mediated dU TP-digoxigenin n ick e nd l abeling) in small clusters of inner cortical cells that subsequently could be distinguished cytologically from neighboring cells throughout rn root development. Inner cortical cells overlying lateral root primordia in either Rn or rn plants also were stained by TUNEL. Features commonly observed in animal cell apoptosis were confirmed by electron microscopy but, surprisingly, cells with a necrotic morphology were detected alongside apoptotic cells in the cortex of rn roots when TUNEL-positive cells were first observed. The two morphologies may represent different stages of a common pathway for programmed cell death (pcd) in plant roots, or two separate pathways of pcd could be involved. The phenotype of rn plants suggests that the Rn gene could either negatively regulate cortical cell death or be required for cortical cell survival. The possibility of a mechanistic link between cortical cell death in rn plants and during lateral root emergence is discussed.     

The effect of CCCP on ion fluxes in the stele and cortex of maize roots

Summary The accumulation of ⁸⁶Rb labelled potassium by isolated stelar and cortical tissues from 7-day-old roots of Zea mays has been compared with the levels accumulated by these tissues in the intact root. Cortical tissues have similar uptake eapacities in these two conditions whereas stelar tissues only exhibit an uptake capacity in the intact root system. The uncoupler carbonylcyanide m-chlorophenylhydrazone caused a considerable decrease in the uptake of potassium by these tissues. In the intact root system it prevented ions from the bathing medium reaching the stelar tissues. The efflux pattern from preloaded isolated stelar and cortical tissues was considerably altered by the inhibitor, a promotion of the efflux occurring in both of these tissues.It is concluded that stelar tissues only accumulated ions when these are supplied through the root symplasm and that the stelar plasmalemma has only a limited uptake capacity per se. Stelar uptake is thus a reflection of vacuolar accumulation across the tonoplast. There is no evidence in the present study of a carrier-mediated active secretion of ions across the stelar plasmalemma. The fact that the efflux was promoted rather than depressed by the uncoupler supports the postulate that a passive leakage is the final stage in the transport of ions across the plant root.     

Fungal communities and health status of roots of winter wheat cultivated after oats and oats mixed with other crops

Health status of winter wheat roots and thecomposition of wheat root fungi were studiedover 1996-1999 following the cultivation ofoats in a pure stand and mixed with otherplants as forecrops. The infection of wheatroots by >Gaeumannomyces graminis wasobserved to be largely dependent on the kind offorecrop; the best being oats in a pure stand,and then oats with pea or lupin mixtures. Inthe emergence and shooting phases, saprophyticfungi were dominant, while in the stage of harddough stage mainly pathogenic fungi, especially>G. graminis were common. The pathogenicfungi were mostly represented by >G.graminis and >Fusarium spp., while >Rhizoctonia spp. were much less frequent.The composition of the fungal communitydepended considerably on the forecrop anddevelopment phase of the plant. The kind offorecrop significantly affected the frequencyof infection by >G. graminis. The highestnumber of isolates was obtained from wheat rootsof crops grown after a mixture of oats andbarley.     

Effect of different soil types,pH and phosphorus levels on the wheat seedling and the incidence ofPythium graminicolum

Summary In the present study, the effect of different types of soil, pH and phosphorus levels on root, shoot development and browning extent on shoot of wheat seedlings growth in soil inoculated withPythium graminicolum; was studied. Out of the four soil types the development of root and shoot was better in Domatta soil where as the disease development was more in black clay soil. The pH values below 5.5 and above 8.0 are favourable for the root and shoot development whereas these values are unfavourable for the development of the Pythium and consequently for infection by it. The percentage of browning of the collar region was more in soils near neutral (pH 6.5–7.1) and it was less in acidic (pH 5.0) or alkaline (pH 8.6) soils. The development of wheat roots and shoot in soil with 30 lb/acre phosphorus was good as compared to that in soil with P₀, P_15,45 and P₆₀ lb/acre in inoculated soil. The percentage of extent of browning on collar region was less at 15 lb and 30 lb/acre P, due to less development of the disease.     

Ectomycorrhizal fungi increase the vitality of Norway spruce seedlings under the pressure of Heterobasidion root rot in vitro but may increase susceptibility to foliar necrotrophs

We tested if root colonisation by ectomycorrhizal fungi (EMF) could alter the susceptibility of Norway spruce (Picea abies) seedlings to root rot infection or necrotic foliar pathogens. Firstly, spruce seedlings were inoculated by various EMF and challenged with Heterobasidion isolates in triaxenix tubes. The ascomycete EMF Meliniomyces bicolor, that had showed strong antagonistic properties towards root rot causing Heterobasidion in vitro, protected spruce seedlings effectively against root rot. Secondly, spruce seedlings, inoculated with M. bicolor or the forest humus, were subjected to necrotrophic foliar pathogens in conventional forest nursery conditions on peat substrates. Botrytis cinerea infection after winter was mild and the level of needle damage was independent of substrate and EMF colonisation. Needle damage severity caused by Gremminiella abietina was high in seedlings grown in substrates with high nutrient availability as well as in seedlings with well-established EMF communities. These results show that albeit M. bicolor is able to protect spruce seedlings against Heterobasidion root rot in axenic cultures it fails to induce systemic protection against foliar pathogens. We also point out that unsterile inoculum sources, such as the forest humus, should not be considered for use in greenhouse conditions as they might predispose seedlings to unintended needle damages.     

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.     

Effect of Ophiobolus graminis infection on the assimilation and distribution of 14C in wheat

The uptake of ¹⁴C and movement of ¹⁴C-labelled assimilates in wheat plants inoculated with Ophiobolus graminis was examined following exposure of the second youngest leaf to ¹⁴CO₂. Autoradiographs of plants with infected seminal roots showed that assimilates were not translocated past the sites of root infection but accumulated in the undamaged portions of infected root systems, in particular the developing crown roots. There was no evidence that assimilates accumulated in the vicinity of O. graminis lesions. The net assimilation of ¹⁴CO₂ by wheat plants over a 5 h feeding period was not significantly affected by O. graminis infection. However, infection reduced the amount of ¹⁴C lost through respiration. Infection delayed the transfer of labelled assimilates from the fed leaf to the remainder of the plant but increased the proportion translocated to the roots. The latter effect was not apparent when infected plants were continuously irrigated during, and for 20 h following, the feeding period.     

Evidence for proteins specific for vascular elements in intact and cultured tissues and cells of maize

A group of antigenically distinct proteins characteristic for the tissue complex of the vascular cylinders was found in maize (Zea mays L.) seedlings using an immunofiltration technique. Specific stelar antigens present in the fully developed stele (vascular cylinder) of the primary root were also found in steles extracted from adventitious roots and from the mesocotyl but were absent, within the limits of sensitivity of the immunodiffusion tests employed, in root cortex and epidermis. Some of the stelar antigens were also evident in the meristem of the primary root and were present in traces in the scutellum, the mesocotyl node, and the primary leaves plus coleoptile. The specific stelar antigens could be traced in 13- and 15-day-old developing embryos and were definitely expressed by the 21 st day after pollination. Several stelar-specific antigens were found in embryo-derived callus tissues and in stem-derived cells maintained in serial suspension culture. Higher resolution of the stelar antigens by a modified technique of crossed immunoelectrophoresis was used to demonstrate several minor stelar antigens that were presumably characteristic exclusively of the completely differentiated stele. This technique along with sequential immunoprecipitation of labelled proteins provided a semiquantitative estimate of the specific stelar antigens in the meristem and the stele of the primary root, and in suspension-cultured cells which were devoid of noticeable signs of vascular differentiation.     

Detection of Latent Infection of Wheat Leaves Caused by Blumeria graminis f.sp. tritici Using Nested PCR

Wheat powdery mildew caused by Blumeria graminis f.sp. tritici (Bgt) is an important and destructive disease worldwide. Detection of latent infection of wheat seedlings is critical to estimate initial inoculum potential of epidemics in the fields. To improve the conventional method, a nested PCR approach had been established in this study to detect latent infections of wheat leaves caused by Bgt. The DNA primer sets including external and internal primer pairs for the nested PCR were designed followed by testing their specificities to Bgt by using Bgt and other fungal species of wheat. Sensitivity test demonstrated that the nested PCR could detect as low as 0.1 fg template DNA and about 10,000 times more sensitive than the standard PCR. Results of artificial inoculation experiments showed that the nested PCR assay can detect a low level of latent infection of wheat seedlings 2 days earlier than did standard PCR. The incidences of latent infection of wheat seedlings determined by the nested PCR linearly correlated with those by the conventional incubation method (r² = 0.66, P = 0.0023). The incidences of latent infection detected with nested PCR were higher than that with the conventional method. This study provides an accurate method to efficiently estimate the initial inoculum potential of wheat powdery mildew epidemics in the fields.     

Structure of Syncytia Induced by Heterodera avenae Woll. in Roots of Susceptible and Resistant Wheat (Triticum aestivum L.)

The structure of syncytia induced by Heterodera avenae in roots of susceptible Triticum aestivum cv. ‘Capa’ and resistant wheat AUS 10894 was investigated. In susceptible plants the first syncytia were observed 4 days after placing the seedlings into a substrate containing nematode cysts. They were located on different root levels. In resistant plants syncytia were first found after 14 days, exclusively in the root maturity zone. In both wheats syncytia consisted mostly of stelar parenchymatic cells. The syncytia induced in roots of Triticum aestivum AUS 10894 were characterized by less immediate contact with the host tracheary elements than the syncytia induced in roots of susceptible Triticum aestivum cv. ‘Capa’. In both wheats a stimulation of pericycle cell divisions, giving rise to lateral roots, was found in parts where the syncytia developed.     

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.     

Effect of Ophiobolus graminis infection on the growth of wheat and barley

Glasshouse experiments are reported, in which the development of young wheat and barley plants was examined following inoculation with Ophiobolus graminis (Sacc.) Sacc. The dry weight, leaf area, tiller number and water content of the shoot were reduced by infection. Reductions were equally severe in wheat and barley. The seminal root system of both was severely attacked and its growth retarded. Inoculated plants, however, translocated a greater proportion of their total assimilates to the root system and produced more adventitious roots than healthy plants. As a result there was an increase in both the number and proportion of healthy roots on these plants following the initial infection of their root systems. This effect was more pronounced in barley than in wheat. It is suggested that this may in part account for the reported relative tolerance of barley to take-all attack under field conditions.     

Antifungal Activity of Aqueous Extracts of the Leaves of Cowparsnip and Comfrey

We found that extracts from the leaves of medicinal comfrey and cowparsnip strongly inhibit the germination of Erysiphe graminisconidia and uredospores of Puccinia graminis. Spraying wheat seedlings with these extracts, in contrast to the irrigation of soil, markedly diminished infection in plants with powdery mildew. Antifungal activity in vitroand protective activity (when plants were sprayed) correlated with the level of phenolic compounds in these extracts. Experiments with healthy plants have demonstrated that the photosynthetic apparatus of wheat plants is stimulated by extracts. Spraying seedlings with the extracts resulted in an increased rate of O₂evolution calculated per unit of chlorophyll, an increase in the ratio (F_M– F_T)/F_Tin the experiments that recorded slow fluorescence induction, an increase in the relative light intensity of band A, and a decrease of relative intensity of band Cin experiments with thermoluminescence of wheat leaves. These results provide evidence that the protective activity of comfrey and cowparsnip extracts is associated with their action on the pathogenic fungus and with the activation of natural defense reactions of the host plant.