Showing papers in "Molecular Plant Pathology in 2003"

Alternaria spp.: from general saprophyte to specific parasite.

Abstract: SUMMARY Alternaria species are mainly saprophytic fungi However, some species have acquired pathogenic capacities collectively causing disease over a broad host range This review summarizes the knowledge on pathogenic strategies employed by the fungus to plunder the host Furthermore, strategies employed by potential host plants in order to ward off an attack are discussed Taxonomy:Alternaria spp kingdom Fungi, subkingdom Eumycotera, phylum Fungi Imperfecti (a non-phylogenetic or artificial phylum of fungi without known sexual stages whose members may or may not be related; taxonomy does not reflect relationships), form class Hypomycetes, Form order Moniliales, form family Dematiaceae, genus Alternaria Some species of Alternaria are the asexual anamorph of the ascomycete Pleospora while others are speculated to be anamorphs of Leptosphaeria Host Range: Most Alternaria species are common saprophytes that derive energy as a result of cellulytic activity and are found in a variety of habitats as ubiquitous agents of decay Some species are plant pathogens that cause a range of economically important diseases like stem cancer, leaf blight or leaf spot on a large variety of crops Latent infections can occur and result in post-harvest diseases or damping-off in case of infected seed Useful Website:

Fusarium oxysporum: exploring the molecular arsenal of a vascular wilt fungus

Abstract: SUMMARY Taxonomy: Vascular wilt fungus; Ascomycete although sexual stage is yet to be found. The most closely related teleomorphic group, Gibberella, is classified within the Pyrenomycetes. Host range: Very broad at the species level. More than 120 different formae speciales have been identified based on specificity to host species belonging to a wide range of plant families. Disease symptoms: Initial symptoms of vascular wilt include vein clearing and leaf epinasty, followed by stunting, yellowing of the lower leafs, progressive wilting of leaves and stem, defoliation and finally death of the plant. In cross-sections of the stem, a brown ring is evident in the area of the vascular bundles. Some formae speciales are not primarily vascular pathogens but cause foot- and rootrot or bulbrot. Economic importance: Causes severe losses on most vegetables and flowers, several field crops such as cotton and tobacco, plantation crops such as banana, plantain, coffee and sugarcane, and a few shade trees. Control: Use of resistant varieties is the only practical measure for controlling the disease in the field. Under greenhouse conditions, soil sterilization can be performed. Alternative control methods with potential for the future include soil solarization and biological control with antagonistic bacteria or fungi. Useful websites: http://www.fgsc.net/fus.htm, http://www-genome.wi.mit.edu/annotation/fungi/fusarium/, http://www.cbs.knaw.nl/fusarium/database.html

Soft rot erwiniae: from genes to genomes

Abstract: SUMMARY The soft rot erwiniae, Erwinia carotovora ssp. atroseptica (Eca), E. carotovora ssp. carotovora (Ecc) and E. chrysanthemi (Ech) are major bacterial pathogens of potato and other crops world-wide. We currently understand much about how these bacteria attack plants and protect themselves against plant defences. However, the processes underlying the establishment of infection, differences in host range and their ability to survive when not causing disease, largely remain a mystery. This review will focus on our current knowledge of pathogenesis in these organisms and discuss how modern genomic approaches, including complete genome sequencing of Eca and Ech, may open the door to a new understanding of the potential subtlety and complexity of soft rot erwiniae and their interactions with plants. Taxonomy: The soft rot erwiniae are members of the Enterobacteriaceae, along with other plant pathogens such as Erwinia amylovora and human pathogens such as Escherichia coli, Salmonella spp. and Yersinia spp. Although the genus name Erwinia is most often used to describe the group, an alternative genus name Pectobacterium was recently proposed for the soft rot species. Host range: Ech mainly affects crops and other plants in tropical and subtropical regions and has a wide host range that includes potato and the important model host African violet ( Saintpaulia ionantha ). Ecc affects crops and other plants in subtropical and temperate regions and has probably the widest host range, which also includes potato. Eca , on the other hand, has a host range limited almost exclusively to potato in temperate regions only. Disease symptoms: Soft rot erwiniae cause general tissue maceration, termed soft rot disease, through the production of plant cell wall degrading enzymes. Environmental factors such as temperature, low oxygen concentration and free water play an essential role in disease development. On potato, and possibly other plants, disease symptoms may differ, e.g. blackleg disease is associated more with Eca and Ech than with Ecc. Useful websites: http://www.scri.sari.ac.uk/TiPP/Erwinia.htm, http://www.ahabs.wisc.edu:16080/pernalab/erwinia/index.htm, http://www.tigr.org/tdb/mdb/mdbinprogress.html, http://www.sanger.ac.uk/Projects/E_carotovora/.

Xanthomonas citri: breaking the surface

Abstract: SUMMARY Taxonomy: Bacteria; Proteobacteria, Gammaproteobacteria; Xanthomonadales; Xanthomonadaceae, Xanthomonas. Microbiological properties: Gram-negative, obligately aerobic, straight rods, motile by a single polar flagellum, yellow pigment. Related species: X. campestris , X. axonopodis , X. oryzae , X. albilineans . Host range: Affects Rutaceous plants, primarily Citrus spp., Fortunella spp., and Poncirus spp., world-wide. Quarantined pathogen in many countries. Economically important hosts are cultivated orange, grapefruit, lime, lemon, pomelo and citrus rootstock. Disease symptoms: On leaves, first appearance is as oily looking, 2‐10 mm, similarly sized, circular spots, usually on the abaxial surface. On leaves, stems, thorns and fruit, circular lesions become raised and blister-like, growing into white or yellow spongy pustules. These pustules then darken and thicken into a light tan to brown corky canker, which is rough to the touch. On stems, pustules may coalesce to split the epidermis along the stem length, and occasionally girdling of young stems may occur. Older lesions on leaves and fruit tend to have more elevated margins and are at times surrounded by a yellow chlorotic halo (that may disappear) and a sunken centre. Sunken craters are especially noticeable on fruit, but the lesions do not penetrate far into the rind. Defoliation and premature abscission of affected fruit occurs on heavily infected trees. Useful websites: ;

Root-knot nematode parasitism and host response: molecular basis of a sophisticated interaction

Abstract: SUMMARY Taxonomy: Eukaryota; Metazoa; Nematoda; Chromadorea; order Tylenchida; Tylenchoidea; Heteroderidae; genus Meloidogyne. Physical properties: Microscopic-non-segmented worms. Meloidogyne species can reproduce by apomixis, facultative meiotic parthenogenesis or obligate mitotic parthenogenesis. Obligate biotrophic parasites inducing the re-differentiation of plant cells into specialized feeding cells. Hosts: Meloidogyne spp. can infest more than 3000 plant species including vegetables, fruit trees, cereals and ornamental flowers. Symptoms: Root swellings called galls. Alteration of the root vascular system. Disease control: Cultural control, chemical control, resistant cultivars. Agronomic importance: Major threat to agriculture in temperate and tropical regions.

Pathogen derived elicitors: searching for receptors in plants.

Abstract: SUMMARY Recognition of potential pathogens is central to plants' ability to defend themselves against harmful microbes. Plants are able to recognize pathogen-derived molecules; elicitors that trigger a number of induced defences in plants. Microbial elicitors constitute a bewildering array of compounds including different oligosaccharides, lipids, peptides and proteins. Identifying the receptors for this vast array of elicitors is a major research challenge. Only in a very few cases has the cognate receptor for a particular elicitor been identified. Biochemical studies have resulted in the characterization of some elicitor binding proteins that may be part of the recognition complex. Transmembrane receptor-like protein kinases (RLKs) constitute one of the most likely categories of receptors involved in pathogen perception. Some of these serine/threonine kinases have been identified as resistance or R genes, others as induced by pathogens or elicitors. One of the RLKs belonging to a leucine rich repeat (LRR) class of putative receptor kinases was recently identified as a receptor for bacterial flagellin, and the underlying signal pathway leading to activation of defence genes was elucidated. These and other recent studies have revealed intriguing similarities in elicitor recognition and defence signalling processes in plant and animal hosts suggesting a common evolutionary origin of eukaryotic defence mechanisms.

Cytosolic HSP90 and HSP70 are essential components of INF1-mediated hypersensitive response and non-host resistance to Pseudomonas cichorii in Nicotiana benthamiana.

Abstract: SUMMARY Mitogen-activated protein kinases (MAPKs) play pivotal roles in the signal transduction pathway of plant defence responses against pathogens. A search for MAPK-interacting proteins revealed an interaction between a Nicotiana benthamiana MAPK, SIPK (NbSIPK) and cytosolic Hsp90 (NbHsp90c-1) in yeast twohybrid assay. To study the function of Hsp90 in disease resistance, we silenced NbHsp90c-1 in N. benthamiana by virus-induced gene silencing (VIGS) with Potato virus X (PVX). NbHsp90c-1 silenced plants exhibited: (1) a stunted phenotype, (2) no hypersensitive response (HR) development after infiltration with the Phytophthora infestans protein INF1 and a non-host pathogen Pseudomonas cichorii that normally triggers HR in N. benthamiana , (3) compromised non-host resistance to P. cichorii , and (4) consistently reduced transcription levels of PR (pathogenesis related) protein genes. Similar phenotypes were observed also for plants in which a cytosolic Hsp70 ( NbHsp70c-1 ), a gene for another class of molecular chaperon, was silenced. Hsp90 was isolated as a MAPK-interacting protein in yeast two-hybrid assay, therefore we tested the effect of NbHsp90c-1 silencing as well as NbHsp70c-1 silencing on the HR development caused by infiltration of a hyperactive potato MAPKK (StMEK1 DD ). No difference in the timing or extent of HR was found among NbHsp90c-1 silenced, NbHsp70c-1 silenced and control plants. This result indicates that observed impairment of INF1- and P. cichoriimediated HR development in NbHsp90c-1 silenced and NbHsp70c-1 silenced plants was not caused by the abrogation in MAPK function downstream of active MAPKK that leads to HR. These findings suggest essential roles of Hsp90 and Hsp70 in plant defence signal transduction pathway upstream or independent of the MAPK cascade.

The nutrient supply of pathogenic fungi: a fertile field for study

Abstract: SUMMARY Phytopathogenic fungi must feed on their hosts to propagate and cause disease. Their ability to access the rich nutrient supply offered by living plants is one of the most obvious properties that distinguish pathogens from saprophytes. Successful invasion by pathogens depends as much on their ability to utilize the available nutrient sources offered by plants as on their ability to penetrate plants and evade defensive mechanisms. Here, we review current knowledge on the nature of the nutrient supplies utilized by pathogens during infection. The available evidence is rudimentary in most cases. There is much evidence to suggest that fungal metabolism can be divided into at least two phases. The first is based on lipolysis and occurs during germination and penetration of the host. The second phase uses glycolysis and predominates during the invasion of host tissue. We also propose, mainly on theoretical grounds, that a third phase of nutrition occurs late in infection during which new spores are produced. Contrary to early assumptions, the nitrogen sources available to some pathogens appear to be abundant. The idea that nitrogen starvation is a cue that controls fungal gene expression during infection may need to be reassessed. Very little is known about the micronutrient (Fe, S, P) or vitamin supply. The knowledge gained from this research may enable the design of new antifungal strategies targeting potential weaknesses in fungal metabolism and will also impact on agronomic practices.

Pathogenic fungi: leading or led by ambient pH?

Abstract: SUMMARY Pathogenic fungi have successfully attacked a wide range of hosts, which has forced them into ambient-adaptation. pH is one of the major ambient traits affecting the activity of pathogenicity factors secreted by the pathogen, hence, a pH sensing-response system was developed to enable the pathogen to tailor its arsenal to best fit its host. The pacC palA, B, C, F, H and I apparatus was first identified in Aspergillus nidulans and later found in other fungi. Secreted pathogenicity factors, such as cell wall degrading enzymes, were recognized to be controlled by environmental pH and later shown to be regulated by the pH regulatory system, either directly or by harbouring the pacC consensus sequence. The ability of the pathogen to actively increase or decrease its surrounding pH allows it to select the specific virulence factor, out of its vast arsenal, to best fit a particular host.

Downy mildew of Arabidopsis thaliana caused by Hyaloperonospora parasitica (formerly Peronospora parasitica)

Abstract: SUMMARY Downy mildew of Arabidopsis is not a hugely destructive disease of an important crop plant, neither is it of any economic importance. The most obvious symptom, the aerial conidiophores, might, at a glance to the casual observer, be mistaken for the trichomes normally present on the leaves. However, a huge research effort is being devoted to this humble pathosystem which became established as a laboratory model in the 1990s. Since then, enormous progress has been made in cloning and characterizing major genes for resistance (RPP genes) and in defining many of their downstream signalling components, some of them RPP-gene specific. Resistance is generally associated with an oxidative burst and a salicylic acid dependent hypersensitive reaction type of programmed cell death. Biological and chemical induction of systemic acquired resistance (SAR) in Arabidopsis protecting against downy mildew were demonstrated early on, and investigations of mutants have contributed fundamentally to our understanding of host–pathogen interactions and the mechanisms of plant defence. This review will attempt to collate the wealth of information which has accrued with this pathosystem in the last decade and will attempt to predict future research directions by drawing attention to some still unanswered questions. Taxonomy:Hyaloperonospora Constant. parasitica (Pers.:Fr) Fr. (formerly Peronospora parasitica ), Kingdom Chromista, Phylum Oomycota, Order Peronosporales, Family Peronosporaceae, Genus Hyaloperonospora , of which it is the type species. The taxonomy of the group of organisms causing downy mildew of brassicas has undergone a number of revisions since Corda (1837) originally coined the genus Peronospora. All isolates pathogenic on brassicas were described initially as P. parasitica but Gaumann (1918) classified isolates from different brassicaceous hosts distinctly and thus defined 52 new species based on conidial dimensions and host range. After much debate it was decided to revert to the aggregate species of P. parasitica for all brassica-infecting downy mildews, whilst recognizing that these show some isolate-specific differences (Yerkes and Shaw, 1959). The latest re-examination of P. parasitica by Constantinescu and Fatehi (2002) has placed isolates of P. parasitica and five other downy mildew species in a clear new subgroup on the basis of their hyaline conidiospores, recurved conidiophore branch tips and ITS1, ITS2 and 5.8S rDNA sequence comparisons; meriting the coining of the new genus ‘Hyaloperonospora Constant’. The class Oomycetes in the Kingdom Chromista (Straminipila) comprises fungus-like organisms with heterokont zoospores (i.e. possessing two types of flagellae, whiplash and tinsel). The Oomycetes have non-septate hyphae with cellulose-based walls containing very little or no chitin. The latter is regarded as a major distinction separating the Oomycetes from the true fungi, and reports of the presence of chitin had generally been regarded as due to small amounts of contamination (Gams et al., 1998). However, in view of recent studies by Werner et al. (2002) showing a chitin synthase gene in an Oomycete and demonstrating the presence of the polymer itself by an interaction with wheat germ agglutinin (WGA), it is perhaps safe to say that we have not seen the last taxonomic revision which will affect this group! The families within the Oomycetes show a clear evolutionary trend to a lesser absolute dependence on an aqueous environment and some members of the Peronosporales, e.g. H. parasitica, have no zoosporic stage in the life cycle. Host range: Isolates infecting Arabidopsis thaliana have so far proven to be non-pathogenic on other crucifers tested but exist in a clear gene-for-gene relationship with different host ecotypes. Disease symptoms: Infections are first apparent to the naked eye as a carpet or ‘down’ of conidiophores covering the upper and lower surfaces of leaves and petioles. This symptom is characteristic of this group of diseases and lends it its name. Useful websites: (links to references on Oomycetes), (TAIR, The Arabidopsis Information Resource).

Polymyxa graminis and the cereal viruses it transmits: a research challenge.

Abstract: UNLABELLED SUMMARY Polymyxa graminis is a eukaryotic obligate biotrophic parasite of plant roots that belongs to a poorly studied discrete taxonomic unit informally called the 'plasmodiophorids'. P. graminis is non-pathogenic, but has the ability to acquire and transmit a range of plant viruses which cause serious diseases in cereal crop species and result in significant yield reductions. The viruses are protected from the environment within P. graminis resting spores ('cysts') that may remain dormant but viable for decades (probably until a suitable host plant is encountered). The persistent, soil-borne nature of these diseases makes the use of virus-resistant crop varieties currently the only practical and environmentally friendly means of control. USEFUL WEBSITES http://www.rothamsted.bbsrc.ac.uk/ppi/links/pplinks/plasmod/index.html, http://www.dpvweb.net/, http://www.rothamsted.bbsrc.ac.uk/ppi/Iwgpvfv/index.html, http://www.rothamsted.bbsrc.ac.uk/ppi/links/pplinks/bymoviruses/index.html, http://oak.cats.ohiou.edu/~braselto/plasmos/

The Fusarium graminearum MAP1 gene is essential for pathogenicity and development of perithecia

Abstract: SUMMARY Fusarium graminearum is the causal agent of ear blight disease of cereals. Infection occurs at anthesis when ascospores and/or conidia directly penetrate exposed anther and ovary tissue. The hemibiotrophic hyphae colonize floral tissues and developing grains to cause premature ear senescence. During infection, Fusarium hyphae can also produce hazardous trichothecene mycotoxins, thereby posing a threat to human and animal health and safety. The Fusarium MAP1 gene was identified using a PCR approach by its homology to a known pathogenicity gene of Magnaporthe grisea, the mitogen-activated protein kinase gene PMK1. Gene replacement F. graminearum map1 mutants were non-pathogenic on wheat flowers and roots, and also could not infect wounded wheat floral tissue or tomato fruits. Unlike the wild-type strain, map1 mutant inoculations did not compromise grain yield. Map1 mutants lost their ability to form perithecia in vitro, but their rate of asexual conidiation was unaffected. DON mycotoxin production in planta was still detected. Collectively, the observed phenotypes suggest that the Map1 signalling protein controls multiple events in disease establishment and propagation. Novel approaches to control Fusarium ear blight disease by blocking perithecial development are discussed.

Cotton leaf curl disease, a multicomponent begomovirus complex

Abstract: SUMMARY Cotton leaf curl is a serious disease of cotton and several other malvaceous plant species that is transmitted by the whitefly Bemisia tabaci. The disease is, at this time, endemic throughout Pakistan and epidemic in Western India. Affected cotton plants exhibit a range of symptoms such as leaf curling, stunted growth and a poor yield of cotton fibre. In addition, affected plants may develop leaf-like outgrowths from the veins on the undersides of leaves. A number of distinct begomoviruses have been shown to be associated with infected plants, all of which require a satellite component (DNA β) to induce symptoms in cotton. DNA β components are a group of recently identified, symptom modulating, single-stranded satellite molecules. An additional, satellite-like component, DNA 1, is invariably found in diseased plants, although it is not required for disease development. Taxonomy: The viruses associated with the CLCuD complex on the Indian subcontinent, five of which have been identified thus far (Table 1), are all single component begomoviruses (genus Begomovirus family Geminiviridae). The satellite (DNA β) and satellite-like (DNA 1) components have yet to be classified, although the DNA 1 components are closely related to, and thought to have originated from, components of a second group of single-stranded DNA viruses, the nanoviruses (family Nanoviridae). Table 1. Begomoviruses associated with cotton leaf curl disease on the Indian subcontinent. Species Acronym Reference Cotton leaf curl Alabad virus CLCuAV Zhou et al. (1998) Cotton leaf curl Kokhran virus CLCuKV Zhou et al. (1998); Mansoor et al. (2003b) Cotton Leaf curl Multan virus CLCuMV Zhou et al. (1998); Briddon et al. (2000); Mansoor et al. (2003b) Cotton leaf curl Rajasthan virus CLCuRV Radhakrishnan et al. unpublished. Papaya leaf curl virus PaLCuV Mansoor et al. (2003b) Physical properties: The begomoviruses associated with CLCuD, like all geminiviruses, have geminate (twinned) particles, approximately 18–20 nm in diameter and 30 nm long, consisting of two incomplete T = 1 icosahedra joined together in a structure with 22 pentameric capsomers and 110 identical protein subunits. It is probable, although not conclusively proven, that the DNA 1 and DNA β components, being half the size of the viral component, are encapsidated in monomeric, rather than geminate particles. Disease symptoms: Symptoms in cotton usually appear within 2–3 weeks of inoculation by B. tabaci (determined experimentally (Singh et al., 1997)) and are initially characterized by a deep downward cupping of the youngest leaves. This is followed by either upward or downward curling of the leaf margins, swelling and darkening of the veins as well as the formation of enations on the veins, which frequently (dependant on variety) develop into cup-shaped, leaf-like structures (Fig. 1). Figure 1. Symptoms induced in cotton (Gossypium hirsutum) induced by CLCuD. The upper two panels are leaves of plants experimentally infected with CLCuMV and CLCuD DNA β showing vein swelling, vein darkening, the formation of enations and the leaf-like outgrowths on the veins on the undersides of leaves. The bottom photograph was taken in a field in Haryana state, India, in 2001 showing the typical foliar symptoms of CLCuD. Download figure to PowerPoint Disease control: Control of CLCuD is mainly based on insecticide treatments against the insect vector (Bemisia tabaci). Roguing, the removal of affected plants, particularly of ratoon cotton from the previous seasons crop, is recommended but appears to have little affect in reducing the incidence of the disease. More recently, resistant cotton cultivars have been introduced that were developed by conventional breeding/selection. After initially showing promise in the control of CLCuD, recent reports have suggested that the virus complex has overcome the resistance. Useful websites: , .

Plant pathogenic Verticillium species: how many of them are there?

Abstract: SUMMARY Two of the currently widely accepted species in the section Nigrescentia of the genus Verticillium are major plant pathogens inducing wilt diseases in a wide range of mainly dicotyledonous hosts. Three species closely related to these two are less important wilt pathogens and soil saprophytes. A sixth species, V. theobromae, causes the cigar end of banana. Molecular and genetic studies have shown that these species represent a complex pool of discrete lineages of varying degrees of relatedness with unknown levels of gene flow between them. Most isolates are haploid, but some are thought to be amphihaploid interspecific hybrids. Until our understanding of this complex is much improved, it seems most appropriate to add only one new species, for wilt isolates primarily associated with potato and producing dark-resting-mycelium in bundles (currently known as V. albo-atrum Grp2). It is suggested that the following be retained: (i) V. dahliae to include all isolates which produce only microsclerotia, (ii) V. albo-atrum to cover the majority of isolates producing only dark-resting-mycelium (and not in bundles), and (iii) V. nigrescens, V. nubilum, V. tricorpus and V. theobromae for the minor wilt pathogens/saprophytes and the non-wilt pathogen.

Molecular, ecological and evolutionary approaches to understanding Alternaria diseases of citrus

Abstract: SUMMARY Alternaria fungi cause four different diseases of citrus: Alternaria brown spot of tangerines, Alternaria leaf spot of rough lemon, Alternaria black rot of several citrus fruits and Mancha foliar of Mexican lime. The first three diseases are caused by the small-spored species, Alternaria alternata and the causal agents can only be differentiated using pathogenicity tests, toxin assays or genetic markers. Mancha foliar is caused by the morphologically distinct, large-spored species A. limicola. Substantial progress has been made in understanding the biology, ecology, population biology, systematics, molecular biology and biochemistry of the interactions between these pathogens and citrus. Epidemiological studies have focused on brown spot of tangerines and their hybrids and have contributed to the development of a model of disease development which has improved control and reduced fungicide use. Studies of the population genetics, host specificity and ecology of A. alternata from different ecological niches on citrus have revealed host specific forms of the pathogen which cause disease on different citrus species, the existence of three phylogenetic lineages of the fungus which cause brown spot world-wide, and closely related non-pathogenic isolates which colonize healthy citrus tissue. The role of host-specific toxins in Alternaria diseases of citrus has been extensively studied for over 20 years, and these pathosystems have become model systems for host-pathogen interactions. Recent molecular research has started to unravel the genetic basis of toxin production and the host susceptibility to toxin, and the role of extracellular, degradative enzymes in disease.

Characterization of a chorismate mutase from the potato cyst nematode Globodera pallida.

Abstract: SUMMARY Some plant endoparasitic nematodes are biotrophic and induce remarkable changes in their hosts in order to ensure a continuous supply of food. Proteins secreted from oesophageal gland cells have been implicated in this pathogenic process. A potentially secreted chorismate mutase has been isolated from the potato cyst nematode Globodera pallida. The gene encoding this protein is expressed in the subventral oesophageal gland cells of the nematode, and the mRNA derived from this gene is only present in the early parasitic stages. Sequence analysis of this gene shows that, like other genes involved in the host-parasite interaction of plant parasitic nematodes, it is likely to have been acquired by horizontal gene transfer from bacteria. The presence of a signal peptide in the deduced amino acid sequence of the G. pallida chorismate mutase and its expression in the subventral oesophageal gland cells suggest that it is secreted from the nematode, pointing to a role for the protein in the host-parasite interaction. The shikimate pathway, of which chorismate mutase is normally a part, is not found in animals but is present in plants and bacteria. In plants it gives rise to a variety of compounds which are important in amino acid synthesis and defence signalling pathways, as well as auxins, which have been implicated in the early development of nematode feeding sites. The potential roles of a nematode chorismate mutase are discussed.

Quantification of induced resistance against Phytophthora species expressing GFP as a vital marker: β-aminobutyric acid but not BTH protects potato and Arabidopsis from infection

Abstract: SUMMARY Induced resistance was studied in the model pathosystem Arabidopsis-Phytophthora brassicae (formerly P. porri) in comparison with the agronomically important late blight disease of potato caused by Phytophthora infestans. For the quantification of disease progress, both Phytophthora species were transformed with the vector p34GFN carrying the selectable marker gene neomycine phosphotransferase (nptII) and the reporter gene green fluorescent protein (gfp). Eighty five per cent of the transformants of P. brassicae and P. infestans constitutively expressed GFP at high levels at all developmental stages both in vitro and in planta. Transformants with high GFP expression and normal in vitro growth and virulence were selected to quantify pathogen growth by measuring the in planta emitted GFP fluorescence. This non-destructive monitoring of the infection process was applied to analyse the efficacy of two chemical inducers of disease resistance, a functional SA-analogue, benzothiadiazole (BTH), and beta-aminobutyric acid (BABA) which is involved in priming mechanisms of unknown nature. BABA pre-treatment (300 microm) via soil drench applied 24 h before inoculation completely protected the susceptible Arabidopsis accession Landsberg erecta (Ler) from infection with P. brassicae. A similar treatment with BTH (330 microm) did not induce resistance. Spraying the susceptible potato cultivar Bintje with BABA (1 mm) 2 days before inoculation resulted in a phenocopy of the incompatible interaction shown by the resistant potato cultivar Matilda while BTH (1.5 mm) did not protect Bintje from severe infection. Thus, in both pathosystems, the mechanisms of induced resistance appeared to be similar, suggesting that the Arabidopsis-P. brassicae pathosystem is a promising model for the molecular analysis of induced resistance mechanisms of potato against the late blight disease.

Active defence responses associated with non-host resistance of Arabidopsis thaliana to the oomycete pathogen Phytophthora infestans.

Abstract: SUMMARY The molecular basis of non-host resistance, or species-specific resistance, remains one of the major unknowns in the study of plant-microbe interactions. In this paper, we describe the characterization of a non-host pathosystem involving the model plant Arabidopsis thaliana and the economically important and destructive oomycete pathogen Phytophthora infestans. Cytological investigations into the early stages of this interaction revealed the germination of P. infestans cysts on Arabidopsis leaves, direct penetration of epidermal cells, formation of infection vesicles and occasionally secondary hyphae, followed by a typical hypersensitive response. P. infestans biomass dynamics during infection of Arabidopsis was monitored using kinetic PCR, revealing an increase in biomass during the first 24 h after inoculation, followed by a decrease in the later stages. Transgenic reporter lines and RNA blot analyses were used to characterize the defence responses induced following P. infestans infection. Significant induction of PDF1.2 was observed at 48 h after inoculation, whereas elevated levels of PR gene expression were detected three days after inoculation. To further characterize this defence response, DNA microarray analyses were carried out to determine the expression profiles for c. 11 000 Arabidopsis cDNAs 16 h after infection. These analyses revealed a significant overlap between Arabidopsis non-host response and other defence-related treatments described in the literature. In particular, non-host response to P. infestans was clearly associated with activation of the jasmonate pathway. The described Arabidopsis-P. infestans pathosystem offers excellent prospects for improving our understanding of non-host resistance.

Genetic and forma specialis diversity in Blumeria graminis of cereals and its implications for host-pathogen co-evolution.

Abstract: SUMMARY The grass powdery mildew fungus, Blumeria graminis is classified into eight formae speciales (ff.spp.) based on strict host specialization. However, evidence suggests that host ranges extend to more than one genus and are particularly diverse among samples from the Middle East, the proposed centre of origin and diversification of crop plants. This study investigated whether geographical origin, host species or both determine the genetic variation in B. graminis that is found in cereals, sampled from Europe, Asia and North America, and whether there is any evidence for co-evolution between pathogen and host. Phylogenetic analysis of nucleotide sequence variation within the ribosomal DNA Internal Transcribed Spacer (ITS) regions and the beta-tubulin (tub2) gene gives rise to two dendrograms with different topologies. In both trees, isolates of B. graminis from cultivated cereals are grouped according to their principal host genus. This grouping was supported by amplified fragment length polymorphism (AFLP) analysis and cross-infectivity tests. However, there was no evidence of co-evolution. There was far greater divergence between ff.spp. in tub2 sequences than ITS regions and a faster rate of mutation of tub2, especially in the third base position of exons. It is proposed that variation in the rDNA-ITS regions is constrained either by their functional role in the processing of rDNA precursor molecules or by concerted evolution, hence limiting their use in phylogenetic studies. AFLP data suggests an overall lack of correlation between geographical and genetic distances. This may be related to the long distance dispersal exhibited by B. graminis.

Agrobacterium tumefaciens mediated transformation of the oomycete plant pathogen Phytophthora infestans

Abstract: Agrobacterium tumefaciens is widely used for plant DNA transformation and, more recently, has also been used to transform yeast and filamentous fungi. Here we present a protocol for Agrobacterium-mediated DNA transformation of the oomycete Phytophthora infestans, the causal agent of potato late blight. Binary T-DNA vectors containing neomycin phosphotransferase (npt) and beta-glucuronidase (gus) fused to oomycete transcriptional regulatory sequences were constructed. Seven days of co-cultivation followed by transfer to a selective medium containing cefotaxim to kill Agrobacterium and geneticin to select for transformants, resulted in geneticin resistant colonies. Under optimal conditions with Agrobacterium supplemented with a ternary plasmid carrying a constitutive virG gene and in the presence of acetosyringone as inducer, up to 30 transformants per 10(7) zoospores could be obtained. The majority of these transformants contained a single T-DNA copy randomly integrated at a chromosomal locus. Using a similar protocol, geneticin resistant transformants of two other oomycetes species were obtained, Phytophthora palmivora and Pythium ultimum.

Potato leafroll virus: a classic pathogen shows some new tricks.

Abstract: SUMMARY Taxonomy: PLRV is the type species of the genus Polerovirus, in the family Luteoviridae. Isolates are known from most continents, presumably all spread in potato material derived from the Andean region of South America. Physical properties: PLRV particles are isometric and c. 25 nm in diameter. They contain one major (c. 23 kDa) and one minor (c. 80 kDa) protein. The genome is a single 5.8 kb positive sense RNA that has neither a 5′-cap nor 3′ poly(A) but carries a VPg. Host range: PLRV has a limited host range; about 20 largely solanaceous species have been infected experimentally. PLRV is a common pathogen of potato, and closely related isolates are occasionally found in tomato, but no other crops are affected. Symptoms: Infection, especially from infected seed potato stocks, causes leafrolling and stunting, the extent depending on the potato cultivar. Biological properties: The biology of PLRV is that of a classic luteovirus. Its isometric particles are persistently transmitted by aphids in a non-propagative manner, it multiplies largely in phloem tissue and disease symptoms reflect this localization. A decade or so of molecular study has revealed the many features of PLRV that are characteristic of its family. Key attractions: In recent years some interesting features of PLRV have emerged that are the focus of further investigation. These are, its phloem confinement, its movement in infected plants, its ability to suppress gene silencing and new ideas about the structure of its particles. This review describes the background to PLRV and points towards these new developments.

The role of lipopolysaccharides in induction of plant defence responses.

Abstract: SUMMARY Lipopolysaccharides (LPS) are ubiquitous, indispensable components of the cell surface of Gram-negative bacteria that apparently have diverse roles in bacterial pathogenesis of plants. As an outer membrane component, LPS may contribute to the exclusion of plant-derived antimicrobial compounds promoting the ability of a bacterial plant pathogen to infect plants. In contrast, LPS can be recognized by plants to directly trigger some plant defence-related responses. LPS also sensitize plant tissue to respond more rapidly or to a greater extent to subsequently inoculated phytopathogenic bacteria. Sensitization is manifested by an accelerated synthesis of antimicrobial hydroxycinnamoyl-tyramine conjugates, in the expression patterns of genes coding for some pathogenesis-related (PR) proteins, and prevention of the hypersensitive reaction caused by avirulent bacteria. The description at the molecular level of the various effects of LPS on plants is a necessary step towards an understanding of the signal transduction mechanisms through which LPS triggers these responses. A definition of these signal transduction pathways should allow an assessment of the contribution that LPS signalling makes to plant disease resistance in both natural infections and biocontrol.

Pathogenicity and other genomic islands in plant pathogenic bacteria

Abstract: SUMMARY Pathogenicity islands (PAIs) were first described in uropathogenic E. coli. They are now defined as regions of DNA that contain virulence genes and are present in the genome of pathogenic strains, but absent from or only rarely present in non-pathogenic variants of the same or related strains. Other features include a variable G+C content, distinct boundaries from the rest of the genome and the presence of genes related to mobile elements such as insertion sequences, integrases and transposases. Although PAIs have now been described in a wide range of both plant and animal pathogens it has become evident that the general features of PAIs are displayed by a number of regions of DNA with functions other than pathogenicity, such as symbiosis and antibiotic resistance, and the general term genomic islands has been adopted. This review will describe a range of genomic islands in plant pathogenic bacteria including those that carry effector genes, phytotoxins and the type III protein secretion cluster. The review will also consider some medically important bacteria in order to discuss the range, acquisition and stabilization of genomic islands.

In situ PCR detection of phytoplasma DNA in embryos from coconut palms with lethal yellowing disease.

Abstract: SUMMARY DNA of the lethal yellowing (LY) phytoplasma was detected in 13 of 72 embryos from fruits of four diseased Atlantic tall coconut palms by polymerase chain reaction (PCR) assays employing phytoplasma universal rRNA primer pair P1/P7, nested LY group-specific rRNA primer pair 503f/LY16Sr or LY phytoplasma-specific nonribosomal primer pair LYF1/R1. Phytoplasma distribution in sectioned tissues from six PCR positive embryos was determined by in situ PCR and digoxigenin-11-deoxy-UTP (Dig) labelling of amplification products. Dig-labeled DNA products detected by colourimetric assay were clearly evident on sections from the same three embryos investigated in detail by in situ PCRs employing primer pairs P1/P7 or LYF1/R1. Deposition of blue-green stain on sections as a result of each assay was restricted to areas of the embryos corresponding to the plumule and cells ensheathing it. By comparison, similarly treated embryo sections derived from fruits of a symptomless Atlantic tall coconut palm were consistently devoid of any stain. Presence of phytoplasma DNA in embryo tissues suggests the possible potential for seed transmission which remains to be demonstrated.

Deciphering host resistance and pathogen virulence: the Arabidopsis/Pseudomonas interaction as a model

Abstract: SUMMARY The last decade has witnessed steady progress in deciphering the molecular basis of plant disease resistance and pathogen virulence. Although contributions have been made using many different plant and pathogen species, studies of the interactions between Arabidopsis thaliana and Pseudomonas syringae have yielded a particularly significant body of information. The present review focuses on recent findings regarding R gene products and the guard hypothesis, RAR1/SGT1 and other examples where protein processing activity is implicated in disease resistance or susceptibility, the use of microarray expression profiling to generate information and experimental leads, and important molecular- and genome-level discoveries regarding P. syringae effectors that mediate bacterial virulence. The development of the Arabidopsis-Pseudomonas model system is also reviewed briefly, and we close with a discussion of characteristics to consider when selecting other pathosystems as experimentally tractable models for future research.

Beet yellows virus: the importance of being different.

Abstract: SUMMARY Taxonomic relationship: Type member of the genus Closterovirus, family Closteroviridae. A member of the alphavirus-like supergroup of positive-strand RNA viruses. Physical properties: Virions are flexuous filaments of approximately 1300 nm in length and approximately 12 nm in diameter that are made up of a approximately 15.5 kb RNA and five proteins. The major capsid protein forms virion body of helical symmetry that constitutes approximately 95% of the virion length. The short virion tail is assembled by the minor capsid protein, Hsp70-homologue, approximately 64-kDa protein, and approximately 20-kDa protein. Viral proteins: The 5'-most ORFs 1a and 1b encode leader proteinase and RNA replicase. The remaining ORFs 2-8 are expressed by subgenomic mRNAs that encode 6-kDa membrane protein, Hsp70 homologue, approximately 64-kDa protein, minor and major capsid proteins, approximately 20-kDa protein, and approximately 21-kDa protein, respectively. Hosts: The principal crop plants affected by Beet yellows virus (BYV) are sugar beet (Beta vulgaris) and spinach (Spinacea oleracea). In addition, BYV was reported to infect approximately 120 species in 15 families. Most suitable propagation species are Nicotiana benthamiana, Tetragonia expansa, and Claytonia perfoliata.

In planta induced genes of Puccinia triticina.

Abstract: SUMMARY Wheat leaf rust disease, caused by the biotrophic fungus Puccinia triticina, is a result of complex interactions requiring the coordinated activities of the two organisms involved. In an effort to understand the molecular basis of wheat-rust interactions, we isolated and characterized cDNA corresponding to in planta induced fungal genes (PIGs) from susceptible wheat leaves infected with P. triticina by using suppression subtractive hybridization to construct a cDNA library. 350 clones were sequenced, of which 104 were unique. Forty-four cDNA clones encode ribosomal proteins, comprising the single largest category of clones isolated. Twenty-five of these ribosomal protein genes are likely to be of fungal origin, as was suggested by sequence homology. Hybridization of 56 selected non-ribosomal protein clones to rust germling cDNA or genomic DNA probes showed that at least 44 were of fungal origin, demonstrating that the library was highly enriched for fungal cDNA. Differential expression analysis identified 26 non-ribosomal protein genes that were induced in rust-infected leaves. At least 21 of the induced genes were from the rust fungus, indicating that the majority of the induced genes were rust PIGs that are likely to play a role in parasitism. Some of the induced genes share homology to known PIGs or virulence genes in other fungi, suggesting similarities in parasitism among different fungi. Eight clones correspond to novel PIGs that have not been reported in any organism. This paper reports, for the first time, the isolation of P. triticina PIGs and discusses the use of total rust genomic DNA to identify the source of genes.

Identification of cytokinins produced by the plant parasitic nematodes Heterodera schachtii and Meloidogyne incognita

Abstract: SUMMARY The presence of different types of cytokinins was analysed in exudates and lysates of stage-2 juveniles of Heterodera schachtii and Meloidogyne incognita and in mixed stages of Caenorhabditis elegans. For all species, cytokinins were detected in lysates and exudates in which benzyladenine and zeatin-type cytokinins were the most prominent forms. The production of cytokinins by Meloidogyne was much higher than by Heterodera, and the detected levels were in a range which interfered with the physiological activities of the host plant. The presence of 5-methoxy-N,N-dimethyltryptamine hydrogen oxalate did not affect hormone production by H. schachtii, whereas resorcinol slightly stimulated hormone production by M. incognita. The exuded cytokinins may play a role in feeding site induction, more particularly in cell cycle activation and in establishing the feeding site as a nutrient sink.

Potato virus X: a model system for virus replication, movement and gene expression.

Abstract: SUMMARY Considerable research has focused on the cis- and trans-acting components required for various aspects of the potato virus X (PVX) infection process. In addition, the development of PVX-based vectors has facilitated analyses of the PVX infection process and provided diverse technological applications. As a result, the PVX system will continue to serve as a model for analyses of processes such as virus movement, RNA replication, and gene silencing, and as a tool for protein expression.

Isolation of genes expressed during compatible interactions between leaf rust (Puccinia triticina) and wheat using cDNA-AFLP.

Abstract: SUMMARY The rust fungi are obligate biotrophic pathogens that depend on living host tissue for their growth. In compatible interactions they go through a number of developmental stages to form intercellular hyphae and haustoria within host cells, through which they obtain their nutrients. Here we have exploited the cDNA-AFLP technique to isolate fragments of wheat and rust genes that are expressed at specific defined time-points during the infection process. A number of these sequences were used as probes in Northern hybridizations and in RT-PCR to confirm their expression patterns, and were also characterized by PCR analysis and Southern hybridizations to determine whether they are of fungal or wheat origin. A cDNA library was constructed from pooled RNAs extracted from days 5 and 7 after inoculation, and this library was screened to isolate full-length cDNAs of selected sequences. Sequence analysis of these cDNA fragments and clones revealed similarities amongst the fungal genes to a chitinase, a sorbitol utilization protein, an arabinitol dehydrogenase and a proteasome regulatory unit, whilst in wheat, we identified sequences with homology to a katanin and a cell enlargement protein.