Uredospore Germination and Development of Some Cereal Rusts from South-central Chile at Constant Temperatures

The minimum and maximum temperatures for germination of uredospores of Puccinia striiformis, P. recondite, P. coronata and P.gramnis isolates from south-central Chile were 0°C and 26°C, 0°C and 32°C, 8°C and 30°C, and 4°C and 34°C, respectively, whereas the shortest latent period was 8.5 days for P. struformis at 20°C, 5 days for P. recondite at 26°C, 5.5 days for P. coronata at temperatures, reaching the minimum and maximum threshold temperatures at 0.006°C and 23°C for P. strüformis, 0.31°C for P. recondita, 2.87°C and 30°C for P. coronata, and 1.72°C and 32°C for P. graminis. rspectively. The cardinal temperatures give no explanation as to the observed sequential appearance of these rusts during the growing season. Other phenomena like the systemic mycelial growth of P.striiformis might be involved here. At temperatures between 19°C and 22°C, the average daily increase of the area of sporulation of P.striiformis in wheat leaves varied between 9.05 and 22.48 mm²/infection site. This variation was due to substrate (plant) history and environmental factors.     

Effect of rust infection on the protein components of wheat

The three wheat varieties KSML-3, HD-2009 and WG-377 were grown in rust-free plots and in plots infected with the rusts Puccinia striiformis and P. recondita. Total protein extracted from the grain samples was analysed quantitatively by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) to determine effects of rust infection on the protein components. In varieties HD-2009 and WG-377 rust infection increases the proportion of albumin-globulin and gliadin proteins while decreasing the high M_r glutenins. The proteins of variety KSML-3 were less affected by rust infection than were those of the other two varieties. The consequent possible effects on the functional quality of the grain could be a weakening of dough mixing resistance and an increase in dough extensibility. The difference in varietal response to rust infection could be important in selection of seed for cultivation in rust prone locations.     

Relatedness Among Agronomically Important Rusts Based on Mitochondrial Cytochrome b Gene and Ribosomal ITS Sequences

A fragment of the mitochondrial cytochrome b (cyt b) gene of 13 agronomically important plant pathogenic Basidiomycetes was sequenced, including several Puccinia spp., Uromyces appendiculatus, Phakopsora pachyrhizi, Hemileia vastatrix and Rhizoctonia solani. The deduced amino acid sequences (residues 142–266) were used to study the relatedness of these pathogens as compared to other species of the Basidiomycetes, Ascomycetes and Oomycetes. The relatedness was also studied at nuclear level using the Internal Transcribed Spacers (ITS) in the ribosomal DNA. Phylogenic trees were constructed with the maximum parsimony (MP) and the neighbour‐joining (NJ) methods. On the basis of both cytochrome b and ITS sequences, the Puccinia species pathogenic to graminaceous crop plants, such as Puccinia recondita f. sp. tritici, Puccinia graminis f. sp. tritici, Puccinia striiformis f. sp. tritici, Puccinia coronata f. sp. avenae, Puccinia hordei, P. recondita f. sp. secalis and Puccinia sorghi, together with Puccinia horiana from Chrysanthemum, were very closely related to each other, whereas Puccinia arachidis (from peanut) was closely related to U. appendiculatus (from beans) but more distant from the other Puccinia species. Both rusts on soybean (P. pachyrhizi) and coffee (H. vastatrix) were outside the Puccinia cluster. All rusts were separated from other Basidiomycetes such as R. solani and the strobilurin producing species Strobilurus tenacellus and Mycena galopoda. Our results demonstrate that the amino acid sequence of the mitochondrial cytochrome b is a valid tool to study phylogenic relatedness among plant pathogenic Basidiomycetes and supports taxonomic grouping based on morphological structures and host specificity. Because of their high variability, ITS sequences were able to discriminate Puccinia species, which were identical on the basis of the cytochrome b amino acid sequence. Thus, ITS sequences could better show differences among species or within a species, whereas cytochrome b is more suitable than ITS for phyologenic inference at family or genus level. In addition, the sequence data obtained during this study represent essential information for easy isolation of the cyt b gene and detection of point mutations conferring resistance to Qo Inhibitor fungicides that eventually may evolve.     

Aphanocladiutn album, a fungus inducing teliospore production in rusts

The hyphomycetous fungus Aphanocladiutn album can grow over and around uredia of the rusts Puccinia coronata, P. hordei, P. graminis f.sp. avenae and P. recondita f.sp. triticina when host plants are kept under very humid conditions, but not on such plants not infected with rusts; uredia are adversely affected and telia develop in their vicinity. Plants inoculated with these rusts and with five isolates of A. album (one from a dead insect) showed: (1) much earlier development of telia on detached and non-detached rusted leaves inoculated with A. album than on corresponding leaves not thus inoculated; (2) telial induction by A. album in some isolates of rust species which hitherto had rarely or never produced telia; (3) precocious telial formation, in comparison with controls, when A. album spores were sprayed on leaves as much as 3 days before and 9 days after rust inoculation, and occasionally after uredia had already matured. As affected leaves remained green until the whole leaf became moribund, senescence is apparently not the factor inducing telia formation. The normal-appearing teliospores of some isolates were induced to germinate, whereas others did not. Rhamnus palaestina inoculated with basidiospores of one isolate of A. album-treated P. coronata f.sp. avenae produced pycnia and fertile aecia. The importance of A. album as a working tool in rust research and as a possible means for biological control of rust epiphytotics is discussed.     

Tissue culture of the alternate hosts of wheat,rye and oat rusts and their response to in vitro inoculation with the rust pathogens

Alternate host plants of cereal rust fungi are necessary for studying the rust sexual cycle and pathogenicity. These plants are usually difficult to propagate through cloning, while seed-propagated plants may have variable responses to the pathogen. To overcome these obstacles, tissue culture, under controlled and aseptic conditions, was utilized for clonal propagation and in vitro inoculation of the following species: Rhamnus palaestinus Boiss., the alternate host of oat (Avena spp.) crown rust (Puccinia coronata Corda); Thalictrum speciosissimum L., the alternate host of brown leaf rust of wheat (Puccinia recondita f. sp. tritici Eriks. & Henn.); and Lycopsis arvensis L., the alternate host of rye (Secala spp.) leaf rust (Puccinia recondita f. sp. recondita Rob. & Desm.). Shoot culture procedures for initial establishment and proliferation were developed for all three alternate host species. Shoot cultures were multiplied at rates ranging from 0.3 to 1.7 shoots/week. Successful infection following inoculation with teliospores of the corresponding rust fungi was obtained for R. palaestinus and T. speciosissimum but not for L. arvensis. The hardening and acclimatization efficiency of rooted T. speciosissimum and L. arvensis was of 80–90%. The propagation efficiency for R. palaestinus was not successful because of the low rate and poor quality of its rooting. It is concluded that the in vitro system might be used as an alternative method for inoculation and multiplication of alternate hosts of cereal rusts, although more experimentation is needed to define accurately the appropriate conditions for the proper infection response. This revised version was published online in June 2006 with corrections to the Cover Date.     

Vertical Concentrations of Fungal Spores above Wheat Fields

Kramer-Collins volumetric spore samplers were used to measure concentrations of Puccinia graminis, P. recondita, Erysiphe graminis, Cladosporium, and Alternaria spores and fungal hyphal fragments within the canopy and at 1, 3, 6 and 14 m above ground level over wheat fields near Manhattan, Kansas, USA. The largest numbers of spores of each of the named fungi and hyphal fragments were trapped during hours when free moisture was not present on host leaf surfaces. As wind velocity increased, the number of spores and hyphal fragments trapped at all heights increased. Airspora trapped at the various sampling heights under all combinations of biometeorological conditions were calculated as ratios. Location and severity of infection or frequency of occurrence of the parent fungi greatly affected the percentage of propagules released in the canopy and escaped into the atmosphere. Less than 40% of P. recondita urediniospores released from tillering plants in the fall and trapped at 20–25 cm were trapped at 1 m, while only 6% of those trapped at 10 cm were trapped at 1 m. Ratios for the average number of spores trapped at 3 m: 1 m during the entire sampling period were similar for P. recondita (0.54), Alternaria (0.57), and E. graminis (0.48). However, the higher ratios that occurred with spores of P. graminis (0.72) and Cladosporium (0.76), and hyphal fragments (0.77) indicate considerable mixing of locally released airspora with airspora from exogenous sources.     

Early molecular diagnosis and detection of Puccinia striiformis f. sp. tritici in China

Aims: Wheat stripe (yellow) rust, caused by Puccinia striiformis f. sp. tritici (Pst), is the most important foliar disease on wheat in China. Early molecular diagnosis and detection of stripe rust will provide a useful aid to the accurate forecast and seasonal control of this destructive disease. Our objective was to develop PCR assays for the rapid identification and detection of P. striiformis. Methods and Results: The genomic DNA of P. striiformis and P. triticina were amplified by a pair of primers derived from conserved β‐tubulin gene sequence. A 235‐bp specific DNA fragment of P. striiformis was isolated and purified. Based on its sequence, another two primer sets were designed successfully to obtain new sequence‐characterized amplified region (SCAR) markers of P. striiformis, which could be amplified in all test isolates of P. striiformis, whereas no DNA fragment was obtained in other nontarget wheat pathogens. The detection limit of the primer set YR (f)/YR (r1) was 2·20 pg μl^?1. The new SCAR markers of P. striiformis can also be detected in Pst‐infected wheat leaves postinoculated for 2 days. Conclusions: Our assays are significantly faster than the conventional methods used in the identification of P. striiformis. Significance and Impact of the Study: Development of a simple, high‐throughput assay kit for the rapid diagnosis and detection of wheat stripe rust would be anticipated in a further study.     

Host and environmental effects on the penetration of oats by Puccinia graminis avenae and Puccinia coronata avenae

The frequency of penetration from appressoria of Puccinia graminis avenae and P. coronata avenae varied among Avena species and between oat cultivars, although both rusts produced susceptible infection type pustules on the cultivars tested. Penetration on cv. Garry was significantly less than that on the Avena species (A. barbata, A.fatua and A. sterilis) studied and penetration of these Avena species was significantly less than on the cvs Algerian and Fulmark. When the rusts were allowed to develop into pustules on seedlings which had been inoculated with fixed amounts of inoculum, there was a direct relationship between number of pustules produced and penetration frequency. The effects of temperature, light and dew period on penetration from appressoria of ‘single race’ and ‘mixed race’ inocula was also studied on these cultivars and species. Penetration by P. graminis avenae was greatest at 30–35 °C and at light intensities of 5625 lux and above, whereas that by P. coronata avenae was greatest at 20 °C and was unaffected by artificial light intensities up to n 250 lux. Maximal penetration by P. graminis avenae and P. coronata avenae was observed after inoculated plants had been exposed to dew periods of 16 and 12 h respectively. Some penetration was observed after a dew period of 8 h. The time taken for each rust to attain maximum penetration varied from 36 to 52 h after inoculation, depending on the environment, and was usually less for P. coronata avenae than for P. graminis avenae.     

Host and environmental effects on post-penetration development of Puccinia graminis avenae and P. coronata avenae

After inoculation of Avena sterilis and the oat cultivars Algerian and Garry with Puccinia graminis avenae the time required for the eruption of pustules of the rust was markedly less at 30–35 than at 20–25 ^oC. In addition, the area of pustules and number of uredospores produced were significantly greater at 30–35 than at 20 ^oC. Peaks of uredospore production occurred between 12 and 22 days after inoculation. In comparable experiments, the time required for pustules of P. coronata avenae to erupt, and the size of pustules, were relatively insensitive to change of temperature, although weight of uredospores produced was greater at 20 than at 30 ^oC. Peaks of uredospore production occurred between 14 and 18 days after inoculation. Both rusts showed straight-line relationships between pustule area and number of uredospores produced. The percentage of infection foci that developed into pustules was similar with both rusts and on all the oat cultivars examined. Both rusts produced susceptible reaction types on all the hosts tested. Pustules of P. graminis avenae were smaller and fewer and generation time longer on cv. Garry than on cv. Algerian or Avena sterilis and the numbers of pustules per unit of inoculum of both rusts were greatest on Algerian, least on Garry. It is suggested that these quantitative differences in phases of the infection process contribute towards the ‘slow-rusting’ reaction of cv. Garry.     

Effect of temperature, light and host on prepenetration development of Puccinia graminis avenae and Puccinia coronata avenae

The influence of temperature and light on prepenetration development of single and mixed isolates of Puccinia graminis avenae and Puccinia coronata avenae was studied on 0–2% water agar and on leaves of three oat cultivars and on three non-cultivated species of Avena. Germination of uredospores of P. graminis avenae and P. coronata avenae occurred best at 10–30^oC and at 20^oC respectively. The optimum temperature for germ-tube growth and for appressorial formation was 20^oC for both rusts. An inverse relationship was observed between light intensity and prepenetration development with maximal germination of uredospores, germ-tube growth and appressorial formation occurring in darkness. Under optimum conditions maximum percentage germination and appressorium formation of both rusts was attained within 4 and 12 h after inoculation respectively. The proportion of germinated uredospores of crown rust which gave rise to appressoria was about twice that observed for stem rust. No significant differences were observed in prepenetration development between the single and mixed race inocula of the two rusts. Although germination of uredospores was significantly greater on water agar than on oat leaves, there were no significant differences in prepenetration development of the rusts on the various oat cultivars and species examined. Consequently, the data failed to indicate the presence of resistance mechanisms operating during the prepenetration phase of the infection process on the cultivars and species examined.     

Role of topography sensing for infection-structure differentiation in cereal rust fungi

Over 90% of the germ tubes of Puccinia graminis tritici (wheat stem rust) and Puccinia hordei (barley brown rust) differentiate appressoria on encountering stomata.There has been controversy as to the role of host topographical signals in the highly precise and efficient induction of these infection structures over stomata by cereal rusts. In the present study, polystyrene replicas of microfabricated silicon wafers, bearing precise microtopographies of defined dimensions, were used to investigate the influence of ridge spacing and height on infection-structure induction by P. graminis tritici and P. hordei. It was found that artificial topographical signals alone can induce a reproducibly high percentage (83–86%) of germ tubes to differentiate infection structures. Multiple, closely spaced (1.5 μm) ridges which were 2.0 μm high provided the most inductive topography. Differentiation on flat surfaces and over single ridges was < 4%. Appressorium induction commonly initiated a cascade of differentiation events involving the formation of infection pegs, vesicles, infection hyphae, and occasionally haustorial mother cells. It is suggested that the close spacing of cell junctions associated with the dumbbell-shaped guard cells of cereal stomatal complexes provide inductive signals for infection-structure formation by cereal rusts in vivo. Received: 17 October 1996 / Accepted: 5 December 1996     

Versuche zum Herbizideinsatz in gesäten Kohlarten

In den Jahren 1987 bis 1989 wurden die Auswirkungen und Effekte eines kombinierten Befalls des Winterweizens mit Sitobion avenae (Fabr.) und Puccinia recondita Rob.ex Desm. f.sp.tritici Erikss. untersucht. Dabei wurde das Verhalten der Schaderregerpopulationen und die Einflußnahme auf das Ertragsgeschehen bei separatem und simultanem Befall erfaßt. Während bei starkem Befall der oberen Blattetagen mit P.recondita eine Förderung der Aphidenpopulation an den Ähren der befallenen Pflanzen registriert werden konnte, war bei schwachem Braunrostbefall keine Einflußnahme auf die Blattläuse nachweisbar. Durch das kombinierte Auftreten beider Schaderreger kann es zu Reduktionen bei der Kornmasse/Ähre und bei der Tausendkornmasse kommen, die größer sind, als die Summe der Verluste bei Einzelbefall. Ähren stark braunrostbefallener Winterweizenpflanzen wurden zeitiger von S.avenae angeflogen und schneller besiedelt als diese von gesunden Kontrollpflanzen. Als Ursache für die Förderung der Populationsentwicklung von S.avenae werden durch P.recondita induzierte Veränderungen im Angebot an freien Aminosäuren in der Wirtspflanze diskutiert.     

The detection and molecular mapping of a major gene for non-specific adult-plant disease resistance against stripe rust (Puccinia striiformis) in wheat

A major gene determining non-specific adult-plant disease resistance against stripe rust (Puccinia striiformis) designated Yrns-B1 was mapped by using a cross between ’Lgst.79–74’ (resistant) and ’Winzi’ (susceptible). Analyzing F₃ lines of two consecutive experimental years contrary modes of inheritance were observed due to the intermediate character of the gene and the difference in the disease pressure during the seasons. Using the disease scoring data of both experimental years independently two maps were constructed detecting Yrns-B1 20.5 and 21.7 cM, respectively, proximal to the wheat microsatellite (WMS) marker Xgwm493 on the short arm of chromosome 3BS. The genetic relationships to other major genes or to quantitative trait loci controlling adult plant disease resistance against rusts in wheat are discussed. Received: 27 May 1999 / Accepted: 28 September 1999     

The genetics of non-host disease resistance in wheat to barley yellow rust

Non-host resistance is investigated as a potential source of durable resistance. However, the genetics of non-host resistance between closely related plant species and their corresponding pathogens would indicate that in these interactions, non-host resistance primarily involves major genes that operate on a gene-for-gene principal similar to that seen in host resistance. Wheat is a non-host of the barley-attacking form of the fungus responsible for yellow rust, i.e. Puccinia striiformis f. sp. hordei. While P. striiformis f. sp. hordei is generally unable to infect wheat, a partial susceptibility was exhibited by the wheat variety Chinese 166. Consequently, in the cross Lemhi × Chinese 166 two major QTLs for resistance to P. striiformis f. sp. hordei were identified: one on chromosome 1D and a second on 2B. These two QTLs accounted for 43.5% and 33.2% of the phenotypic variance for resistance to barley yellow rust, respectively. In addition, two QTLs of smaller effect were also identified: one on chromosome 5A, contributing 5.1% of the variance and a second on chromosome 6A, contributing 10.9% to the phenotype. The QTL on 6A was derived from the susceptible variety, Chinese 166. In all cases the resistance towards P. striiformis f. sp. hordei was associated with a visual chlorosis/necrosis response typical of race-specific host resistance.     

Stimulation of antioxidative enzymes and polyamines during stripe rust disease of wheat

Content of polyamines and activities of antioxidative enzymes in response to stripe rust disease caused by Puccinia striiformis have been studied in two wheat (Triticum aestivum L.) cultivars PBW 343 (resistant) and HD 2329 (susceptible). Various infection stages ranging from traces to 100 % were collected. Infection leads to stimulation of peroxidase (POD), superoxide dismutase (SOD), catalase, diamine oxidase and polyamine oxidase activities along with increase in putrescine, spermidine and spermine content while ascorbate, tocopherol and chlorophyll content decreased in HD 2329 and no visible symptoms appeared in PBW 343. Histochemical localization pattern of POD and SOD activities revealed correlation with lignin deposition in cell walls.     

Molecular Polymorphism of the Wheat Leaf Rust Fungus in Morocco Using Amplified Fragment Length Polymorphism

The genetic variability and collection structure of the wheat leaf rust fungus Puccinia recondita collected from four agro‐ecological areas of Morocco, Abda‐doukala, Chaouia‐Tadla, Gharb and Tangérois were investigated by amplified fragment length polymorphism (AFLP) markers. A set of five AFLP primers combinations which generated 253 polymorphic loci were used. Hierarchical partitioning revealed that bread wheat collections of Puccinia recondita form a single collection. No significant variation was observed between durum wheat collections of Puccinia recondita; they maintained most of the genetic variability within rather among collections. The distribution pattern of genetic variation of Puccinia recondita collections seems to be the result of high gene flow and the mixed reproduction system.     

SEPTAL PORES IN THE HETEROBASIDIOMYCETIDAE,PUCCINIA GRAMINIS AND P. RECONDITA

The septal pores in uredial mycelium of Puccinia graminis and P. recondita lack the septal swelling and septal pore cap (dolipore-parenthosome configuration) typically associated with the pores of previously investigated Homobasidiomycetidae and the Tremellales among the Heterobasidiomycetidae. The pores in young hyphae of these two species of Puccinia are characterized by the presence of a cytoplasmic matrix which apparently occludes the pore and acts as a plug, thus preventing the migration of organelles from cell to cell. Large vesicles are typically present at the periphery of the pore matrix and the matrix may be very incompletely bounded by a membrane. Nuclei and other cytoplasmic structures migrate from cell to cell through an opening in the septum lateral to the pore. The available evidence indicates that this peripheral gap in the septum results from a breakdown of a portion of an initially complete septum rather than from incomplete septum formation. In addition to the centripetally formed septa, the hyphae of P. graminis and P. recondita are further compartmentalized by shallow infoldings of the lateral wall and limited unilateral septum formation. There is apparent free passage of cellular material between adjacent compartments.     

The Expression of Race-Specific Resistance of Wheat Seedlings to Puccinia recondita

A range of wheat cultivars with resistance factors effective against Puccinia recondita f. sp. tritici was studied to investigate the mode of action and expression of resistance at first and third seedling leaf stages. In most cultivars, resistance to isolate 74/2 resulted in extremely low levels of infection, apparently linked with a predominantly hypersensitive response by the host. In seedlings of cultivars Mans Fundin and Sterna, however, race-specific resistance was expressed as increased pathogen latent period and intermediate infection levels.     

Sugar and Amino Acid Content of Poa pratensis Infected with Ustilago striiformis and Urocystis agropyri

Leaf and root tissues of Poa pratensis L. showing distinct morphological changes associated with infection by Ustilago striiformis (West.) Niessl var. poae (stripe smut) or Urocystis agropyri (Pruess) Schroet. (flag smut) were analyzed for total soluble sugars and free amino acids. Decreases in the quantity of soluble sugars in stripe-smutted plants were not significant, indicating that the presence of U. striiformis sori in leaf tissue does not interfere with photosynthesis of the host. Severe decreases in the quantity of soluble sugars in flag-smutted plants suggest that U. agropyri either directly impairs photosynthesis or effectively metabolizes the photosynthate. Both pathogens caused significant decreases in total free amino acids in leaf and root tissues. The negligible decrease in soluble sugars in stripe-smutted plants was associated with a disproportionate decrease in free amino acids, suggesting that the pathogen either metabolizes amino acids or inhibits their synthesis. The severe decrease in amino acids in leaves and roots of U. agropyri-infected plants is believed due to carbohydrate starvation. It is probable that morphological changes in U. agropyri-infected plants, including reduced branching, dwarfed leaves and root systems, and the inability to produce inflorescences, are probably the direct result of severely reduced levels of sugars and amino acids. The reduced branching, somewhat smaller root systems, and inhibition of inflorescence production on plants infected by U. striiformis probably are associated to some extent with decreases in amino acids. The upright elongated leaves of U. striiformis-infected plants, however, cannot be attributed to the decrease in amino acid content and are suggestive of hyperauxiny.     

Urate Oxidase from the Rust <Emphasis Type="Italic">Puccinia recondita</Emphasis> Is a Heterotetramer with Two Different-Sized Monomers

Uricase (urate: oxygen oxidoreductase; EC 1.7.3.3) from the rust Puccinia recondita was purified to electrophoretic homogeneity. Preparations with a specific activity of 8.4 U/mg were used for characterization of the enzyme, which showed a strong similarity to other plant and fungal urate oxidases. The enzyme had a pH optimum of 9.0, a K _m of 35 μM for urate, and it was inhibited only by oxonate and xanthine. A molecular mass of 152 kDa was estimated for the native protein. SDS-PAGE analysis revealed a striking difference to most urate oxidases, since two different-sized subunits were detected. These results suggest that P. recondita uricase is a tetramer with two types of subunits. Received: 21 February 2001 / Accepted: 30 July 2001