Host/parasite interaction in blackspot disease of roses caused by Diplocarpon rosae Wolf

The utilization of amino acids by Diplocarpon rosae Wolf was investigated and compared with amino acid compositions of leaves of resistant and of tolerant roses. No amino acid appeared to determine resistance of roses to blackspot on a nutritional basis, but phenylalanine marginally increased resistance when taken up by wounded leaves of resistant or tolerant varieties. There is an accumulation of phenolic compounds in response to infection by D. rosae. Most phenolic compounds are toxic to D. rosae in vitro, particularly ellagic acid and p-coumaric acid, but their presence and quantity in healthy rose leaves could not be related to disease resistance. Inhibition of germination on rose leaves of conidia of the fungus was determined in a manner that suggested production of a phytoalexin. It is suggested that resistance of roses to D. rosae may be related to accumulation of polyphenols and possibly to the production of a phytoalexin.     

The toxicity of sulphur dioxide to Diplocarpon rosae Wolf causing blackspot of roses

Germination of conidia of Diplocarpon rosae Wolf is inhibited in a solution made by dissolving 35 p.p.m. sulphur dioxide in water. Inhibition is permanent after exposure for 3 hr. Gaseous SO₂ in air under controlled conditions at 100μg./m³ markedly reduces infection of rose leaves after exposure for 2 days. Blackspot of roses is checked or eliminated in areas where SO₂ pollution of air exceeds 100/μ./m.³.     

Genetic analysis of resistance to blackspot (Diplocarpon rosae) in tetraploid roses

  Diplocarpon rosae is the causal agent of rose blackspot, one of the most severe diseases of field-grown roses. The genetics of resistance to this pathogen was investigated in crosses between tetraploid rose genotypes. The hybrid breeding line 91/100-5, which exhibits a broad resistance to all isolates tested so far, was selfed to produce an F₂ population, backcrossed to the susceptible tetraploid variety ‘Caramba’ and crossed to the susceptible varieties ‘Heckenzauber’, ‘Pariser Charme’ and ‘Elina’. Infection experiments resulted in segregation ratios consistent with the presence of a single dominant resistance locus in the duplex configuration in the hybrid 91/100-5. This suggests, together with previous data on the race structure of the fungus, a “gene-for-gene” type of interaction in the pathosystem Diplocarpon/Rosa. We propose to designate this gene Rdr1, which is the first resistance gene described in the genus Rosa. The advantages and limitations of such an interaction type for future rose breeding programmes and for marker-assisted selection strategies are discussed. Received: 29 August 1997 / Accepted: 19 September 1997     

Efficacy of triazoles and strobilurins in controlling black spot disease of roses caused by Diplocarpon rosae

Black spot disease is an important disease of roses with worldwide occurrence. It is caused by Diplocarpon rosae , an ascomycetous fungus and its control relies on fungicides. The effects of strobilurins and triazoles on D. rosae development within the host and on disease symptoms have not been well studied. Strobilurins completely inhibited germination of conidia when applied protectively 1 week before inoculation or on the same day with the inoculum. They were, however, not effective in eradicating the disease when applied after the fungus was established in the host. Triazoles reduced the germination rate of the conidia when applied protectively and they inhibited disease symptom development when applied after the fungus was established in the host but before symptom expression. Application of triazoles after symptom development suppressed further development of the disease, but in the case of treatment with myclobutanil yellowing and defoliation still occurred. Post-infection application of triazoles led to the apparent breakdown of subcuticular mycelia, intercellular mycelia, and hyphae in the epidermal cells, while the effects of strobilurins were limited to the subcuticular mycelia. Triazoles were more effective than strobilurins because they are more systemic.     

Resistance of roses to pathotypes of Diplocarpon rosae

Colonies of Diplocarpon rosae derived from single conidia were isolated on malt extract agar, multiplied (at 23°C) and stored (at ?20°C) on surface‐sterilised leaf discs of a universally susceptible rose, ‘Frensham’. The resistance of 16 species and cultivars of Rosa to different isolates of D. rosae was assessed using surface‐sterilised leaf discs. Four pathotypes of D. rosae were distinguished on the basis of host range. One species and one hybrid were resistant to all pathotypes. Two species and two cultivars were susceptible to all pathotypes. Four species and six cultivars were interpreted as having vertical resistance because they were strongly resistant to some but not all pathotypes. Only species and hybrids of the section Cinnamomeae were resistant to the pathotype identified as CW1 whereas only roses of other origins were resistant to the pathotype DA2.     

Expression of a chitinase transgene in rose (Rosa hybrida L.) reduces development of blackspot disease (Diplocarpon rosae Wolf)

Blackspot, caused by the Ascomycete fungus Diplocarpon rosae, is the most widespread and pernicious disease of cultivated roses. While some species of rose possess resistance to D. rosae, none of the modern-day rose cultivars are fully resistant to the pathogen. In the current study, Biolistic gene delivery was used to introduce a rice gene, encoding a basic (Class I), chitinase into embryogenic callus of the blackspot-susceptible rose (Rosa hybrida L.) cv. Glad Tidings. The plasmid used for transformation carried the neomycin phosphotransferase (nptII) gene facilitating the selection and regeneration of transgenic plants on medium containing 250 mg/l kanamycin. Southern analysis confirmed integration of 2–6 copies of the chitinase gene into the rose genome; gene expression was confirmed by enzyme assay. Bioassays demonstrated that expression of the chitinase transgene reduced the severity of blackspot development by 13–43%. This degree of resistance to the pathogen correlated with the level of chitinase expression in the transgenic rose plants. The introduction of disease defence genes into rose provides a method of producing blackspot-resistant rose cultivars sought by breeders and growers.     

Microscopic and biochemical evidence of differentially virulent field isolates of Diplocarpon rosae causing black spot disease of roses

Black spot disease caused by Diplocarpon rosae is one of the most widespread diseases of roses that are very difficult to control due to the generative reproduction and complex genetic constitution of roses and up to now the control of black spot still requires intensive use of systemic fungicides. Here we report for the first time evidence of differentially virulent field isolates of D. rosae. Using a combination of fungal structures, disease symptoms and host cells protein expression pattern analysis we here provide direct biochemical evidence that tropical field isolates of D. rosae are more virulent and caused disease symptoms earlier than their temperate counterparts. The tropical fungal field isolates strongly induced an excessive accumulation of ROS and repressed activity of pathogenesis-related proteins such as peroxidases, chitinase and phenylalanine ammonia lyase compared to their temperate counterparts. These findings bring insights into a hidden pathogenic characteristic of tropical D. rosae field isolates compared to their temperate counterparts and open a novel dimension of parameters to be considered when controlling black spot disease of roses by fungicides in tropical versus temperate regions. Interestingly, we found that treatment of rose leaves with ROS (H₂O₂) prior to fungal inoculation promoted plant defense response regardless of the isolate virulence based on protein expression pattern analysis, suggesting that ROS (H₂O₂) can be efficiently incorporated into black spot disease management.     

Influence of phylloplane colonizing biocontrol agents on the black spot of rose caused by Diplocarpon rosae

Abstract

An attempt was made to study the biocontrol efficacy of antagonistic microorganisms in phylloplane of rose cv. Edward to manage the black spot (Diplocarpon rosae) disease. Eight antagonistic microorganisms were tested in vivo against the black spot pathogen. Among these, Trichoderma viride and Pseudomonas fluorescens pf1 reduces the mycelial growth significantly. These two biocontrol agents were evaluated for their ability to induce defense-related enzymes and chemicals in plants. Increased activity of phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenoloxidase (PPO) and total phenolics were recorded in all the biocontrol agents treated leaves. P. fluorescens Pf1 recorded early and increased synthesis of the entire defense-related enzymes and total phenol within 6 days. The application of biocontrol agents induced the defense-related enzymes involved in phenyl propanoid pathway in addition to direct antagonism, which collectively contribute for enhanced resistance against invasion of Diplocarpon rosae in rose.     

Prediction of the Diplocarpon rosae secretome reveals candidate genes for effectors and virulence factors

Rose black spot is one of the most severe diseases of field-grown roses. Though R-genes have been characterised, little information is known about the molecular details of the interaction between pathogen and host. Based on the recently published genome sequence of the black spot fungus, we analysed gene models with various bioinformatic tools utilising the expression data of infected host tissues, which led to the prediction of 827 secreted proteins. A significant proportion of the predicted secretome comprises enzymes for the degradation of cell wall components, several of which were highly expressed during the first infection stages. As the secretome comprises major factors determining the ability of the fungus to colonise its host, we focused our further analyses on predicted effector candidates. In total, 52 sequences of 251 effector candidates matched several bioinformatic criteria of effectors, contained a Y/F/WxC motif, and did not match annotated proteins from other fungi. Additional sequences were identified based on their high expression levels during the penetration/haustorium formation phase and/or by matching known effectors from other fungi. Several host genotypes that are resistant to the sequenced isolate but differ in the R-genes responsible for this resistance are available. The combination of these genotypes with functional studies of the identified candidate effectors will allow the mechanisms of the rose black spot interaction to be dissected.     

Detailed description of developmental growth stages of Diplocarpon rosae in Rosa: a core building block for efficient disease management

Black spot disease of roses caused by the ascomycetous fungus Diplocarpon rosae teleomorph (Marssonina rosae anamorph) is a widespread and devastating disease. Despite considerable progress in the management of black spot disease in the recent years, it is still unclear by which mechanisms this fungus colonises and invades the host system, and without a good knowledge of such infection machineries, it not possible to fully overcome the challenges of D. rosae infection. By exploring research contributions up to date, we highlight in this review the ultrastructure of D. rosae infection cycle in the host cell by emphasising on several aspects related to its in vitro and in vivo germinations, the infection mechanisms of the fungus, the different fungal structures formed in the host cells and the optimum storage conditions for D. rosae to retain its viability and pathogenicity over time. Here, attention is particularly focused on the asexual life cycle of D. rosae, with the sexual cycle being briefly discussed. In addition, a new dimension of research approaches to effectively control black spot disease of roses, that is, how to accurately use the advanced biotechnology tools to speed up the current state of the disease management, is proposed here.     

Identification of molecular markers linked to Rdr1, a gene conferring resistance to blackspot in roses

Blackspot resistance in the tetraploid rose genotype 91/100–5 had been characterised previously as a single dominant gene in duplex configuration. In the present study a tetraploid progeny (95/3) segregating for the presence of the blackspot resistance gene Rdr1 were screened with 868 RAPD and 114 AFLP primers/primer combinations. Seven AFLP markers were found to be linked to Rdr1 at distances between 1.1 and 7.6 cM. The most closely linked AFLP marker was cloned and converted into a SCAR marker that could be screened in a larger population than the original AFLP and was linked at a distance of 0.76 cM. The cloned fragment was used as an RFLP probe to locate the marker on a chromosome map of diploid roses. This is the first report of markers linked to a resistance gene in roses, and the possibilities of using them for a marker-assisted selection for blackspot resistance as well as for map-based cloning approaches are discussed. Received: 23 December 1999 / Accepted: 25 March 2000     

Epidemiological aspects of ageing.

A major societal challenge is to improve quality of life and prevent or reduce disability and dependency in an ageing population. Increasing age is associated with increasing risk of disability and loss of independence, due to functional impairments such as loss of mobility, hearing and vision; a major issue must be how far disability can be prevented. Ageing is associated with loss of bone tissue, reduction in muscle mass, reduced respiratory function, decline in cognitive function, rise in blood pressure and macular degeneration which predispose to disabling conditions such as osteoporosis, heart disease, dementia and blindness. However, there are considerable variations in different communities in terms of the rate of age-related decline. Large geographic and secular variations in the age-adjusted incidence of major chronic diseases such as stroke, hip fracture, coronary heart disease, cancer, visual loss from cataract, glaucoma and macular degeneration suggest strong environmental determinants in diet, physical activity and smoking habit. The evidence suggests that a substantial proportion of chronic disabling conditions associated with ageing are preventable, or at least postponable and not an inevitable accompaniment of growing old. Postponement or prevention of these conditions may not only increase longevity, but, more importantly, reduce the period of illnesses such that the majority of older persons may live high-quality lives, free of disability, until very shortly before death. We need to understand better the factors influencing the onset of age-related disability in the population, so that we have appropriate strategies to maintain optimal health in an ageing population.     

Epidemiological, clinical, and pathomorphological characteristics of epidemic poliomyelitis-like disease caused by enterovirus 71.

In May-September, 1975, an outbreak of epidemic disease clinically and pathomorphologically simulating nearly all known forms of poliomyelitis occurred predominantly among young infants in Bulgaria. Most cases presented benign aseptic meningitis, sometimes with a short period of general cerebra- symptoms. Paralytic forms, such as bulbar polioencephalitis, anterior poliomyelitis, isolated pareses of the facial nerve, occasional cases of encephalomyocarditis, etc., were observed in about 21% of all cases. Over one-fourth of the paralytic cases with bulbar symptoms ended fatally. In March, 1976 another sporadic fatal case of this disease was examined. No new cases occurred in 1977. Histopathological examinations in all fatal cases regularly revealed lesions in the grey matter of the medulla and spinal cord typical of acute anterior poliomyelitis and bulbar polioencephalitis, with some peculiar features of localization and depth of the involvement of the brain stem. The similarity to poliomyelitis and precariously rapid increase in the incidence led to the decision to urgently vaccinate the entire human population with Sabin's live poliovirus vaccine simultaneously in the whole country in order to produce interference with the circulating agent. This aim appeared to have been achieved partially because soon the number of new cases of the disease began to decrease; however, no sharp and complete break in the curve of the incidence occurred. By the time of mass vaccination, the results of virological examinations started before were not yet available. Later, comprehensive complete evidence was obtained that over 25 fatal and many other typical cases of the disease were associated with an enterovirus proved to be antigenically related to enterovirus 71. The diseases in Bulgaria, 1975, differ considerably in the frequency of paralytic forms and in severity from the epidemics caused by enterovirus 71 in Sweden, 1973, Australia, 1972--1973, USA, 1969--1972, and Japan, 1972--1973.     

Epidemiological aspects of strongyloidiasis in Brazil

The objective of this review was to outline an epidemiological profile of Strongyloides stercoralis by parasitological and serological diagnosis in inhabitants, and to associate this profile with different immunosupression situations, in Brazil, over 20 years (1990-2009). The occurrence of S. stercoralis using parasitological methods was 5·5%, being 4·8% in rural and 5·0% in urban areas, characterizing the country as hyperendemic. There was a diversity of techniques used as a diagnostic tool and only 39·1% of the studies presented results based on at least 1 specific method. The occurrence increased with age, being 12·1%, for those over 60 that suggests an epidemiological condition of concern for the elderly population. Of the seroepidemiological studies in the general population the mean positivity in serum samples was 21·7% and 29·2%, using an immunofluorescence antibody test and enzyme-linked immunosorbent assay (ELISA), respectively. The occurrence of strongyloidiasis in immunosuppressed individuals was 11·8% by parasitological methods and 19·5% using immunological methods. Considering that Brazil is a tropical country and that the character of chronicity and autoinfection of the parasite that can result in severe forms of hyperinfection or dissemination makes strongyloidiasis an important medically and socially neglected problem.