Specificity and host specialization of Puccinia chondrillina

Various strains of Puccinia chondrillina, the Chondrilla rust, were collected during surveys in the Mediterranean, undertaken to discover biological control agents suitable for use in Australia against skeleton weed, Chondrilla juncea. Only the strain collected at Vieste (S.E. Italy) was highly virulent against the common Australian form of C. juncea. The specificity of this strain of the rust was tested by inoculation of many unrelated cultivated plants, related cultivated plants and closely related wild plants; all were immune to the rust. These tests satisfied the Australian plant quarantine authorities and this strain of the rust has now been introduced into Australia as a biological control agent for C. juncea.     

Host-specificity of Uromyces heliotropii, a fungal agent for the biological control of common heliotrope (Heliotropium europaeum) in Australia

Common heliotrope, Heliotropium europaeum (Boraginaceae), a summer annual, is a serious weed of pastures in Australia causing poisoning in sheep and other grazing animals. The weed is native to Mediterranean and Middle Eastern regions of Eurasia where it is attacked by several arthropods and pathogens including the rust fungus Uromyces heliotropii. The host-specificity of a virulent and effective strain of the rust from Turkey has been tested by inoculating 96 plants of importance to the Australasian region using both microscopic and macroscopic observations of the reaction of host and non-host plants. The test plants included several members of the Boraginaceae and related families as well as species of Heliotropium native to Australia. The infection was mostly limited to European species of Heliotropium. U. heliotropii was thus found to represent a safe introduction and has recently been introduced to Australia for the biological control of common heliotrope.     

Genetic evidence for the identity of Caulerpa racemosa (Forsskål) J. Agardh (Caulerpales,Chlorophyta) in the Adriatic Sea

Three taxa of Caulerpa racemosa that differ morphologically and genetically have been reported in the Mediterranean Sea. The ‘invasive variety’ was recorded for the first time in the early 1990s in Libya. In less than 10 years, it was found in almost all parts of the Mediterranean. The first record of C. racemosa in the Croatian part of the Adriatic Sea was in 2000 and as many as 35 different localities were established by the end of 2004. In terms of morphology, the specimens from the Adriatic Sea resemble the ‘invasive variety’. To confirm this, we analysed populations from two different localities (the island of Mljet and the peninsula of Pelje?ac) using the ITS1-5.8S-ITS2 region of 18S-5.8S-26S rDNA unit as a molecular marker. The ITS1-5.8S-ITS2 region from nine individual plants was amplified by PCR, cloned in a pCR 2.1 vector and sequenced. Phylogenetic comparison of sequences from specimens found in the Adriatic Sea with specimens of the same and similar taxa, found both inside and outside the Mediterranean, provided genetic evidence that C. racemosa populating the Adriatic Sea corresponds to the Mediterranean C. racemosa var. cylindracea (Sonder) Verlaque, Huisman?&?Boudouresque, i.e. to the ‘invasive variety’.     

Interactions Between Rhizoctonia Root Rot and the Foliar Common Bean Diseases Anthracnose and Rust

The effects of co‐inoculation of Rhizoctonia solani and Colletotrichum lindemuthianum or Uromyces appendiculatus at different inoculum levels were studied on the disease dynamics and on the growth of bean plants under greenhouse conditions. Bean seeds were sown in R. solani‐infested soil. Additional experiments in which seedlings were transplanted to infested soil were also carried out. Conidial suspensions of C. lindemuthianum or uredospores of U. appendiculatus were inoculated onto leaves at plant developmental stages V2 and V3, respectively. Interactions between root rot and the aerial diseases were observed depending on the inoculum levels and on the timing of R. solani inoculation. Anthracnose severity tended to be higher on R. solani‐infected plants. Conversely, R. solani infection significantly reduced diameter of pustules and rust severity. When seedlings were transplanted to soil infested with low levels of R. solani, root rot severity and density of R. solani in the soil were magnified at high levels of C. lindemuthianum or U. appendiculatus. In these experiments, a synergistic interaction between root rot and anthracnose was observed to affect the plant dry weight. Antagonistic effects on the plant dry weight were found for the combination root rot/rust only when seeds were sown in infested soil.     

The Lr34 adult plant rust resistance gene provides seedling resistance in durum wheat without senescence

The hexaploid wheat (Triticum aestivum) adult plant resistance gene, Lr34/Yr18/Sr57/Pm38/Ltn1, provides broad‐spectrum resistance to wheat leaf rust (Lr34), stripe rust (Yr18), stem rust (Sr57) and powdery mildew (Pm38) pathogens, and has remained effective in wheat crops for many decades. The partial resistance provided by this gene is only apparent in adult plants and not effective in field‐grown seedlings. Lr34 also causes leaf tip necrosis (Ltn1) in mature adult plant leaves when grown under field conditions. This D genome‐encoded bread wheat gene was transferred to tetraploid durum wheat (T. turgidum) cultivar Stewart by transformation. Transgenic durum lines were produced with elevated gene expression levels when compared with the endogenous hexaploid gene. Unlike nontransgenic hexaploid and durum control lines, these transgenic plants showed robust seedling resistance to pathogens causing wheat leaf rust, stripe rust and powdery mildew disease. The effectiveness of seedling resistance against each pathogen correlated with the level of transgene expression. No evidence of accelerated leaf necrosis or up‐regulation of senescence gene markers was apparent in these seedlings, suggesting senescence is not required for Lr34 resistance, although leaf tip necrosis occurred in mature plant flag leaves. Several abiotic stress‐response genes were up‐regulated in these seedlings in the absence of rust infection as previously observed in adult plant flag leaves of hexaploid wheat. Increasing day length significantly increased Lr34 seedling resistance. These data demonstrate that expression of a highly durable, broad‐spectrum adult plant resistance gene can be modified to provide seedling resistance in durum wheat.     

Widespread resistance of Mediterranean island ecosystems to the establishment of three alien species

Although some invasive plants are cosmopolitan, not all ecosystems are invaded to the same degree. Yet there is little experimental work on how ecosystem resistance to invasion at the establishment phase differs among ecosystems. We conducted two field sowing experiments in two consecutive years to examine establishment of the deciduous tree Ailanthus altissima, the succulent subshrub Carpobrotus spp. and the annual geophyte Oxalis pes‐caprae in coastal dunes, shrublands and oldfields in more than 200 sites across six Mediterranean Basin islands differing in climatic conditions and local species richness. Establishment success (i.e. percentage of plots with at least one seedling) and rates (i.e. seedling to sown seed ratio) were low, especially for Ailanthus even when accounting for differences in seed viability. Oxalis was capable of producing a new cohort of seedlings the year following planting. By contrast, all Ailanthus seedlings and half the Carpobrotus seedlings died following the first summer. Differences in establishment success and rates among ecosystems were species‐, island‐ and year‐dependent. Differences in precipitation and mean temperature were associated with differences in establishment rates across sites. Establishment rates tended to be positively correlated with cumulative precipitation and negatively with mean T^a. Unexpectedly, native species richness was not a good predictor of seedling establishment, except for higher Oxalis establishment success in species rich habitats. By conducting field sowing tests at multiple sites across a region we found that except for Oxalis, Mediterranean island ecosystems are quite resistant to invader establishment. These results suggest that differences in the degree of invasion between ecosystems and islands might be more dependent upon the influence of invasion event factors (e.g. propagule pressure) or factors acting at a later life‐history stages rather than differences in the resistance imposed by ecosystems to invader recruitment. Moreover, our results support the notion that in Mediterranean ecosystems invasions are highly idiosyncratic events and strongly dependent on water availability conditions.     

A new strain of the rust fungus Puccinia chondrillina for biological control of skeleton weed in Australia

A strain of the rust fungus Puccinia chondrillina introduced into Australia in 1971 for the biological control of skeleton weed, Chondrilla juncea, attacked only the narrow-leaf form of this weed. Studies were, therefore, undertaken in the Mediterranean region to discover strains of the rust which will attack the other two Australian forms of skeleton weed, intermediate- and broad-leaf. Several strains of the rust were found to be highly virulent against the intermediate form. One of these strains from Manisa, western Turkey, was both highly specific to its host and safe for use as a biological control agent. This strain has recently been introduced into Australia for the control of the intermediate form of Chondrilla.     

The Effects of Temperature on the Respiratory Responses of Groundsel (Senecio vulgaris) to Infection by Rust (Puccinia lagenophorae)

Groundsel (Senecio vulgaris) was grown in either a warm (20°C) or a cool (8°C) controlled environment and infected with Puccinia lagenophorae. Dark respiration, measured over the range 6 to 18°C, was higher in leaves of healthy plants grown under low temperatures than in those of plants grown under high temperatures. Infection increased the rates of dark respiration in the region of sporulating lesions in both sets of plants, but the greater increase in plants grown under warm conditions resulted in both sets having similar respiration rates across the range 6 to 18°C. The conclusion that the magnitude of the respiratory increase following rust infection depends upon the conditions under which plants were grown is supported by literature on other rust diseases and has implications for the utilization of carbohydrate reserves and the survival of both rust and host populations over winter.     

Host-pathogen interactions in natural populations of Linum marginale and Melampsora lini

Summary The epidemiology of rust caused by the fungus Melampsora lini and the effects of infection by this pathogen on its host, the herbaceous perennial Linum marginale, were determined in the field and in garden experiments. There was considerable natural variability in disease levels over the four years (1986–1989) of the study. In two years (1986, 1989) major rust epidemics occurred. In the field, the main effect of disease was to reduce survivorship during the winter following infection. Plants which were heavily infected during the 1986 or 1989 growing seasons had reduced survivorship relative to more lightly infected plants. Melampsora lini infections did not appear to affect survivorship in either 1987 or 1988. Flowering was correlated with environmental factors and the number of stems a plant possessed. A severe drought in 1987 completely inhibited flowering. In the other three years the number of flowers produced by a plant was strongly positively correlated with the number of stems it possessed. Disease levels had no consistent effect on flowering. Controlled garden experiments were also used to examine the response of seedlings and adult plants to infection. These showed that both the timing and severity of disease appears to determine the effect of M. lini infections on L. marginale. Early, severe infection reduced growing season and overwintering survivorship as well as capsule production. However, plants in the field were most often infected only after flowering had begun, and the predominant effect of infection was a reduction in overwintering survivorship. The high variability in disease levels from year to year and the deferred nature of the effect of the rust on its host have significant implications for the design of experiments aimed at assessing the role of diseases in plant communities.     

Drought resistance and recovery of photosystem II activity in a Mediterranean semi-deciduous shrub at the seedling stage

Phlomis fruticosa is one of the main species of post-fire Mediterranean communities, occupying areas with post-fire grazing or altered precipitation pattern and consequently water stress imposed on seedlings of co-occurring species. Seedlings of woody perennials often differ from their mature individuals regarding their resistance of photosynthetic performance to environmental stresses. Such differences have been reported for tree species but there is a lack of information regarding shrub species. In the present study, we tried to detect changes in (PSII) activity imposed by water stress in P. fruticosa seedlings as well as its capability for recovery after rehydration. Maximum PSII photochemical efficiency decreased only under severe water stress exactly as in mature plants in the field. However, leaf chlorophyll (Chl) content was almost stable regardless of leaf relative water content (RWC). We assume that the photoprotective chlorophyll loss process, reported for many mature Mediterranean species (including P. fruticosa), has not yet been developed at the seedling stage. On the other hand, photoprotection through an increase of the relative amount of non-Q_B-reducing centers was found during dehydration. Non-photochemical quenching (NPQ) contributed to protection from photodamage until moderate water stress but was significantly suppressed under severe water stress. Both processes were reversed after rehydration. The above characteristics enabled seedlings not only to survive during aggravating drought but also to maintain a considerable part of their effective quantum yield and perform significant electron transport even at extremely low relative water content (RWC). This was confirmed with measurements in a semi-natural environment (pots) and under real steady state conditions regarding adaptation of the photosynthetic machinery to prevailing light intensities.     

The Biogeography and Phylogeny of Unicellular Cyanobacterial Symbionts in Sponges from Australia and the Mediterranean

The distribution, host associations, and phylogenetic relationships of the unicellular cyanobacterial symbionts of selected marine sponges were investigated with direct 16s rDNA sequencing. The results indicate that the symbionts of the marine sponges Aplysina aerophoba, Ircinia variabilis, and Petrosia ficiformis from the Mediterranean, four Chondrilla species from Australia and the Mediterranean, and Haliclona sp. from Australia support a diversity of symbionts comprising at least four closely related species of Synechococcus. These include the symbionts presently described as Aphanocapsa feldmannii from P. ficiformis and Chondrilla nucula. A fifth symbiont from Cymbastela marshae in Australia is an undescribed symbiont of sponges, related to Oscillatoria rosea. One symbiont, Candidatus Synechococcus spongiarum, was found in diverse sponge genera in the Mediterranean Sea and the Indian, Pacific, and Southern oceans, whereas others were apparently more restricted in host association and distribution. These results are discussed in terms of the biodiversity and biogeographic distributions of cyanobacterial symbionts.This revised version was published online in November 2004 with corrections to Volume 48.     

Catabolic activity of two phosphoric diester hydrolases in wheat leaves inoculated with brown rust,Puccinia recondita

Changes in total phospholipids and activities of two phosphoric diester hydrolases,i.e. phospholipase C and phospholipase D have been studied in inoculated wheat leaves of 25-day old plants, interacting differentially with three races of brown rust,Puccinia recondita, during early stages of pathogenesis. With progressive rust iafection the lipids and phospholipids were found to decrease in all the three interactions except at later stages (48 h and 72 h) in race 104-interaction. However, total phospholipids at various stages were found to be higher in resistant interaction (race 63) as compared to the other two interactions. An initial increase in activities of phospholipase C and D in all the three interactions was followed by a decline at the intermediate stages which continued in case of phospholipase C activity while a reverse trend was observed in phospholipase D activity towards later stages of infection. Further the enzymatic activities at various stages were found to be higher in the susceptible and intermediate interactions (with races 77 and 104 respectively) as compared to the resistant interaction.     

A rust-inducible gene from flax (fis1) is involved in proline catabolism

A gene fis1 from flax (Linum usitatissimum), which is induced in mesophyll cells at the site of rust (Melampsora lini) infection, is also expressed in vascular tissue, particularly in floral structures of healthy plants. This paper reports that the promoter controlling this expression is contained within 282 bp 5′ to the coding region and that fis1 gene induction is specifically by the rust pathogen and not by other fungal pathogens or by wounding. The fis1 gene has 73% homology with an Arabidopsis gene which encodes delta-1-pyrroline-5-carboxylate dehydrogenase (P5CDH) which is a part of the proline degradation pathway. Transgenic flax plants that either over-express fis1 or show reduced fis1 expression due to RNA-mediated gene silencing have an unaltered morphology. However, plants with reduced fis1 expression have markedly increased sensitivity to exogenous proline and show alteration in epidermal cell morphology, callose deposition and the production of hydrogen peroxide during proline-induced death. These lines, which show a biologically significant level of fis1 suppression, have an unaltered reaction to either virulent or avirulent rust infections, as do fis1 over-expression lines. These data indicate that the fis1 gene plays a role in proline metabolism and most likely encodes for a P5CDH enzyme. However, the precise role of fis1 and P5C catabolism in the development of rust disease remains unclear.     

Biosystematics,genetics and upper temperature tolerance ofGigartina teedii (Rhodophyta) from the Atlantic and Mediterranean

Plants ofGigartina teedii from the mediterranean isolated into laboratory culture showedPolysiphonia-type life histories with consistent formation of dioecious gametangial plants, as previously reported for Atlantic isolates. Male and female plants from the Atlantic and Mediterranean were almost completely compatible in terms of cystocarp formation on female plants, and carpospores from positive crosses always formed plants that released viable tetraspores. Sex-linked inheritance of branching pattern was found in all strains, but showed varying degrees of expression. Female plants were more branched than male plants and it is suggested that this may be an adaptation for spermatial capture.G. teedii plants showed differences in morphology in culture that are considered to be genetically-based. Preliminary studies of tip elongation showed that Mediterranean strains may have up to three times the elongation rates of Atlantic strains at 15°C, . Such genetic variation in fully-interbreeding strains suggests that populations of this species in the Atlantic and Mediterranean are genecodemic. All strains showed an upper temperature tolerance of 31°C when tested at 1°C intervals from 29—34°C. An upper temperature tolerance of 31–32°C was found for the related speciesG. intermedia from Korea and Japan, butG. johnstonii from the Gulf of California showed an upper tolerance of 32–33°C.     

Genetic polymorphism in Chondrilla (Asteraceae) in southern European Russia and the nature of Chondrilla juncea L

We studied genetic diversity in 54 populations of nine sexual and apomictic species of the genus Chondrilla (C. acantholepis, C. ambigua, C. brevirostris, C. canescens, C. graminea, C. juncea, C. laticoronata, C. latifolia and C. pauciflora) in SE European Russia and neighboring territories of NW Kazakhstan. We analysed the trnT–trnF region of plastid DNA and the internal transcribed spacer of ribosomal DNA (ITS1–5.8S–ITS2) using statistical parsimony, maximum likelihood and neighbor net methods. Two major evolutionary lineages, roughly corresponding to the two subgenera traditionally recognized in the region, were revealed. Within the first evolutionary lineage (subgenus Brachyrhynchus), the sexual diploid C. ambigua and its putatively hybrid apomictic derivatives C. brevirostris, C. laticoronata and C. pauciflora could be recognized. Their identity was also confirmed by analyses of ISSR markers. The second evolutionary lineage (subgenus Chondrilla) comprises C. juncea, C. acantholepis, C. canescens, C. graminea and C. latifolia in European Russia, but analyses of morphological variability and the genealogy of plastid and nuclear markers favor their treatment as the single facultatively apomictic species C. juncea. The results demonstrate that an apomictic mode of reproduction does not necessarily result in the formation of genetically separated microspecies.     

Identification and validation of molecular markers linked to the leaf rust resistance gene Lr19 in wheat

A leaf rust resistance gene Lr19 on the chromosome 7DL of wheat derived from Agropyron elongatum was tagged with random amplified polymorphic DNA (RAPD) and microsatellite markers. The F₂ population of 340 plants derived from a cross between the leaf rust resistant near-isogenic line (NIL) of Thatcher (Tc + Lr19) and leaf rust susceptible line Agra Local that segregated for dominant monogenic leaf rust resistance was utilized for generating the mapping population. The molecular markers were mapped in the F₂ derived F₃ homozygous population of 140 seedlings. Sixteen RAPD markers were identified as linked to the alien gene Lr19 among which eight were in a coupling phase linkage. Twelve RAPD markers co-segregated with Lr19 locus. Nine microsatellite markers located on the long arm of chromosome 7D were also mapped as linked to the gene Lr19, including 7 markers which co-segregated with Lr19 locus, thus generating a saturated region carrying 25 molecular markers linked to the gene Lr19 within 10.2 ± 0.062 cM on either side of the locus. Two RAPD markers S265₅₁₂ and S253₇₃₇ which flanked the locus Lr19 were converted to sequence characterized amplified region markers SCS265₅₁₂ and SCS253₇₃₆, respectively. The marker SCS265₅₁₂ was linked with Lr19 in a coupling phase and the marker SCS253₇₃₆ was linked in a repulsion phase, which when used together mimicked one co-dominant marker capable of distinguishing the heterozygous resistant seedlings from the homozygous resistant. The molecular markers were validated on NILs mostly in Thatcher background isogenic for 44 different Lr genes belonging to both native and alien origin. The validation for polymorphism in common leaf rust susceptible cultivars also confirmed the utility of these tightly linked markers to the gene Lr19 in marker-assisted selection.     

TECHNICAL ADVANCE: Transformation of the flax rust fungus,Melampsora lini: selection via silencing of an avirulence gene

Rust fungi cause devastating diseases on many important food crops, with a damaging stem rust epidemic currently affecting wheat production in Africa and the Middle East. These parasitic fungi propagate exclusively on plants, precluding the use of many biotechnological tools available for other culturable fungi. In particular the lack of a stable transformation system has been an impediment to the genetic manipulation required for molecular analysis of rust pathogenicity. We have developed an Agrobacterium‐mediated genetic transformation procedure for the model flax rust fungus Melampsora lini, which infects flax (Linum usitatissimum). Selection of transgenic rust lines is based on silencing of AvrL567, which encodes a rust effector protein that is recognised by the flax L6 immune receptor. The non‐transgenic rust line is unable to infect flax plants expressing L6, while silenced transgenic lines are virulent on these plants, providing an effective selection system. This directly confirms that the cloned AvrL567 gene is responsible for flax rust virulence phenotypes, and demonstrates the utility of this system to probe rust gene function.