BSPP Presidential Meeting 1999

Biotic Interactions in Plant-pathogen Associations

P. H. Gregory Prize Offered Paper Competition

A technique for screening soil samples for biocontrol agents of late blight of potato.
Elinor Thompson

5th Floor, Darwin Building, University College London, Gower Street, London WC1E 6BT, UK.

A method was developed for the evaluation of biocontrol of potato late blight. Communities of soil micro-organisms were tested for inhibition of Phytophthora infestans disease on whole tubers using a technique that eliminated the need for prior, time-consuming culture of single potential antagonists. Soil samples from three different areas were incubated for 7 days in a medium of sterilised loam enriched with sterile powdered potato periderm, according to the method developed by Schisler et al. For isolating antagonists of Fusarium sambucinum [1]. The soil mixtures were applied to potato tubers that had been allowed to develop P. infestans infection for 2 days or, alternatively, the mixtures were inoculated with the pathogen and re-incubated for 2 days before application of the mix to tubers. Disease severity was assessed by measuring necrosis after 2 weeks. Consortia of isolated micro-organisms from soils that inhibited late blight were then tested in aqueous suspension for disease control on infected tubers.

When inoculated with the pathogen before application to tubers, one soil prevented all disease in two-thirds of the inoculation sites and considerably reduced disease in the remainder (compared with controls, reduction in mean depth and width of lesions, 73% and 82%, respectively; P=0.001). In tubers allowed to develop P. infestans infection for 2 days, there was a small reduction in necrosis after incubation with the same soil, and some mixtures of ten and five isolates randomly selected from this sample significantly reduced blight. All three soils yielded potent antagonists of P. infestans in agar-plate duel culture, the most active of which were identified as Pseudomonas fluorescens, Pseudomonas aeruginosa, Chryseomonas luteola and Flavimonas oryzihabitans. The technique was therefore successful in identifying soils and micro-organisms that were inhibitory to P. infestans disease. The method allows the evaluation of the activity of large populations of bacteria or fungi without previous culture or identification because a range of test soils, and the microflora within them, are cultured and applied to tubers in an identical soil medium.

[1] Schisler, D. A., Burkhead, K. D., Slininger, P. J. & Bothast, R. J. (1998). Selection, characterization, and use of microbial antagonists for the control of Fusarium dry rot of potatoes. In: Plant-Microbe Interactions and Biological Control. Edited by G. J. Boland & L. D. Kuykendall. New York: Marcel Dekker; 199-222.

The effects of environmental factors on light leaf spot epidemics on winter oilseed rape in the UK.
Tijs Gilles

IACR-Rothamsted, Harpenden, Hertfordshire, AL5 2JQ, UK.

Pyrenopeziza brassicae can cause severe epidemics of light leaf spot on winter oilseed rape in the UK, but the severity of epidemics varies between seasons and regions. P. brassicae produces wind-dispersed ascospores and splash-dispersed conidia. Mature apothecia releasing ascospores have been observed on debris in the autumn. The ascospores could, therefore, have a role in causing primary infections, but are not known to be an infective inoculum. From infected plants within crops, light leaf spot is likely to spread by splash-dispersal of the conidia over short distances within crops. The effects of environmental factors on the maturation of apothecia and conidial infection of oilseed rape could explain the differences in severity of epidemics between seasons and regions, but have not been studied in great detail. In controlled environments, lesions were observed when less than 300 ascospores were inoculated on a leaf, but not when less than 700 conidia were inoculated on a leaf. On average, 50 lesions were observed on a leaf when c. 2 103 ascospores or c. 0.6106 conidia were inoculated on a leaf. Apothecia matured on petiole debris at temperatures of 6 to 17C, but not at 22C. An interruption in wetness by a dry period of 4 days delayed maturation by c. 4 days. Light leaf spot symptoms developed on inoculated plants at temperatures from 4 to 20C, but not at 24C. Spore pustules developed only when leaf wetness duration was above a certain minimum and this was temperature dependant.

Modelling cross protection between plant virus strains.
Xu-Sheng Zhang and John Holt

Natural Resources Institute, University of Greenwich, Central Avenue, Chatham Maritime, Kent, ME4 4TB, UK. E-Mail:

A mathematical framework was developed for plant virus disease epidemics that involve cross protection between virus strains. Examples of pathosystems with continuous and discontinuous host availability were considered: citrus tristeza virus disease and barley yellow dwarf virus disease (BYDV), respectively. Analysis showed that competing strains could not coexist in the long term. The virus with the higher basic reproductive number, R0 = (transmission rate)/(loss rate), always excluded the other eventually. A change in either the transmission or the loss rates could shift the balance of dominance between competing virus strains. A mild strain could become dominant if phytosanitation led to preferential removal of hosts infected with the severe strain, or if the transmission efficiency of the mild strain was increased, e.g. by a change in vector genotype. As one strain replaced another over time, the strongest competition occurred when the incidence of both strains was similar: at this time disease incidence was lower than when either strain was present alone. Seasonal resetting of virus incidence to a low level to represent an annual, temperate cropping system made no important qualitative difference to dynamic outcomes. However, discontinuous host availability provided a greater opportunity for strains to replace each other than in more stable perennial cropping systems. The process by which one strain of BYDV replaced another in New York State was discussed.

The genetic control of resistance to turnip mosaic virus in Brassica.
Rachel L. Rusholme1,2, A.G. Sharpe1, C.E. Jenner3, S.L. Hughes3, I.A. Parkin1, J.A. Walsh3 and D.J. Lydiate1

1 AAFC Saskatoon Research Centre, 107 Science Place, Saskatoon, SK. S7N OX2. Canada. 

2 John Innes Centre, Norwich Research Park, Norwich. NR4 7UH. UK.

3 Horticulture Research International, Wellesbourne, CV35 9EF. UK.

The Brassica A genome carries a number of genes that confer resistance to different spectra of the twelve distinct pathotypes of turnip mosaic virus (TuMV). Genetic mapping is identifying the genomic positions of individual resistance genes. This information will allow gene pyramiding and the marker-assisted selection of durable resistance to TuMV in Brassica. The genetic dissection of resistance determinants in near-isogenic lines carrying defined resistance genes will facilitate the biological evaluation of different resistance mechanisms. In combination with the ability to construct hybrid virus genomes carrying defined segments of two or more TuMV isolates, this represents an excellent system for resolving the molecular biology of pathogenesis and host resistance for an economically significant crop pathogen. High resolution mapping of Brassica genes for resistance to TuMV is also bringing the cloning of these genes within reach.

Pasteuria penetrans a pathogen of root-knot nematode Meloidogyne spp. and its use as biological control agent.
Lund Badaruddin

Department of AES, University of Newcastle upon Tyne NE1 7RU, UK.

Abstract withdrawn

Substantial reductions of wheat diseases by straw.
Barry S. Rogers-Gray

The University of Reading, School of Plant Sciences, 2 Earley Gate, Whiteknights, Reading RG6 2AU, UK. 

Over a period of three years, winter wheat (cultivar Mercia) was grown in plots with or without straw added (1 kg/m2). Plants from straw treated plots had consistently reduced diseases: Septoria tritici blotch (Mycosphaerella graminicola), mildew (Erysiphe graminis), brown rust (Puccinia recondita) and foot rot (Fusarium). Early on M. graminicola was worse in straw treated plots. Plant establishment and height, but not leaf area per tiller, were lower in straw treated plots. Fertiliser regimes differed between years and soil and leaf nitrogen were recorded. There was no obvious link between nitrogen and any disease. The photosynthetic viability and numbers of leaves scored for disease were similar between treatments. A fungicide, chlorothalonil, was applied in one crop; its effects did not interact with straw. Late in the season straw treated plants had significantly higher leaf silica (P<0.01). In the glasshouse, plants supplied with silicon had lower E. graminis (P<0.001) and higher leaf silica. In pot experiments, silicon had inconsistent effects on M. graminicola. M. graminicola was not suppressed by straw in pot experiments. A prior inoculation of M. graminicola only primed plant defences against a subsequent attack in the presence of adequate silicon.

Comparison of morphological, cultural and biological characteristics between Japanese and European strains of Monilinia fructigena (Aderh. & Ruhl.) Honey.
Gerard C.M. van Leeuwen1,2, R.P. Baayen2, I. Holb3 and M.J. Jeger4

1Laboratory of Phytopathology, Wageningen University, P.O. Box 8025, 6700 EE Wageningen, the Netherlands;

2 Plant Protection Service, P.O. Box 9102, 6700 HC Wageningen, the Netherlands;

3Department of Plant Protection, Debrecen Agricultural University, P.O. Box 36, H-4015 Debrecen, Hungary;

4Department of Agriculture and Horticulture, Wye College, University of London, Wye, Ashford, Kent TN25 5AH, United Kingdom.

Monilinia fructigena (Aderh. & Ruhl.) Honey is one of the main fruit rot pathogens in pome fruit culture. M. fructigena is endemic in Europe, Central Asia and the Far East including Japan. Previous research revealed that the rDNA ITS1-2 base pair sequence differed between European and Japanese M. fructigena strains by five transitions (1). It was our aim to investigate to what extent this genetic difference is expressed in phenotypic and biological differences between both geographical groups. A set of 5-6 representative isolates of each group was used in all experiments. Colony growth rate was compared on potato dextrose agar (PDA) at 22 oC under 12 h light/ 12 h dark regime (nuv). The size of conidia cultured on cherry agar (CHA) was determined, as well as that of conidia obtained from infected pear fruits (cv. Conference). The formation of stromatal (sclerotial) plates on CHA was quantified by an Image Analyser. The lesion growth rate and sporulation intensity of the pathogen on apple and pear were compared. The mean growth rate of Japanese strains on PDA was significantly higher than that of European strains (t-test, P < 0.05). Average length and width of conidia was significantly smaller in Japanese strains (t-test, P < 0.05). Intensive stroma formation occurred in all Japanese strains, whilst two of the European strains did not show any after 21 days incubation in darkness. Clear differences in lesion growth rate or sporulation intensity on fruits were not found.

(1) Fulton, C.E., Van Leeuwen, G.C.M., and Brown, A.E., 1999. Genetic variation among and within Monilinia species causing brown rot of stone and pome fruits. Eur. J. Pl. Pathol. 105: 495-500

Modelling the growth of soil-borne fungi in response to carbon and nitrogen
Angelique Lamour, F. Van Den Bosch, A.J. Termorshuizen & M.J. Jeger

Wageningen Agricultural University, Laboratory of Phytopathology, P.O. Box 8025,6700 EE Wageningen, The Netherlands. 

Growth of soil-borne fungi is poorly described and understood, largely because non-destructive observations on hyphae in soil are difficult to make. Mathematical modelling can help in the understanding of fungal growth. Except for a model by Paustian & Schnrer, fungal growth models do not consider carbon and nitrogen contents of the supplied substrate, although these nutrients have considerable effects on hyphal extension in soil. We introduce a fungal growth model in relation to soil organic matter decomposition dealing with the detailed dynamics of carbon and nitrogen. Substrate with a certain carbon:nitrogen ratio is supplied at a constant rate, broken down and then taken up by fungal mycelium. The nutrients are first stored internally in metabolic pools and then incorporated into structural fungal biomass. Standard mathematical procedures were used to obtain overall-steady-states of the variables (implicitly from a cubic equation) and the conditions for existence. Numerical computations for a wide range of parameter combinations show that at most one solution for the steady-state is biologically meaningful, specified by conditions for existence. These conditions specify a constraint, namely that the 'energy' (in terms of carbon) invested in breakdown of substrate should be less than the 'energy' resulting from breakdown of substrate, leading to a positive carbon balance. The biological interpretation of the conditions for existence is that for growth the 'energy' necessary for production of structural fungal biomass and for maintenance should be less than the mentioned positive carbon balance in the situation where all substrate is colonised. In summary, the analysis of this complicated fungal growth model gave results with a clear biological interpretation.

Resistance to turnip mosaic virus in Brassica.
Sara L. Hughes1,2, R.L. Rusholme3,4, D.J. Lydiate4, M.J. Kearsey2 and J.A. Walsh1

Horticulture Research International, Wellesbourne, Warwick, CV35 9EF, UK.

2 University of Birmingham, Edgbaston, Birmingham, B15 2TT, UK.

3 John Innes Centre, Norwich Research Park, Colney Lane, Norwich, NR4 7UK, UK.

4 AAFC Saskatoon Research Centre, 107 Science Place, Saskatoon, SK S7N OX2, Canada. 

Turnip mosaic virus (TuMV), a member of the Potyvirus genus, is one of the most important viruses that infects field grown vegetable crops. Its wide host range includes economically important Brassica crops e.g. cabbage, broccoli, swede, spring and winter oilseed rape. The symptoms induced in TuMV-infected plants range from chlorotic spots to severe stunting and necrotic lesions, which significantly reduce the marketability of such plants. It also reduces crop yield and can cause plant death, sometimes destroying crops completely. The non-persistent stylet-borne manner in which TuMV is transmitted by aphids causes insecticides to be ineffective, as brief probes are enough to transmit the virus. This, along with the wide host range of TuMV, makes it extremely difficult to control. Natural plant resistance offers the most effective and environmentally friendly way of controlling this virus. To use plant resistance efficiently in combating TuMV, it is important to characterise sources of resistance in terms of genetic inheritance and interaction with different TuMV isolates. Markers linked to resistance genes and map positions of these genes must be identified to enable the production of crops, using marker-assisted selection, with combinations of genes that confer resistance to a broad spectrum of TuMV isolates.

Is the cassava mosaic epidemic in Uganda the result of a new Bemisia tabaci (Genn.) biotype ?
Maruthi, M. N1., Seal, S1. & Colvin, J1.

1Pest Management Department, Natural Resources Institute, the University of Greenwich, Chatham, UK.

The cassava whitefly, Bemisia tabaci, is the vector of cassava mosaic viruses (family Geminiviridae) which cause cassava mosaic disease (CMD). In the past decade, an epidemic of severe CMD has been spreading steadily southwards across Uganda and now moved into neighbouring countries. Harrison et al. (1997, Ann. Appl. Biol., 131: 437-448) reported that a new recombinant geminivirus, called the Uganda variant (UgV) which was the driving force behind the epidemic. However, questions remain related to the association of unusually high numbers of B. tabaci with the epidemic zone (Legg & Ogwal, 1998, J. Appl. Ent., 122: 169-178). A possible explanation is that the B. tabaci population associated with the epidemic is a better adapted, more fecund, biotype causing an increased transmission rate and thereby the rapid spread of the severe CMD. 

To investigate this, reproductive compatibility, fecundity, developmental period and random amplified polymorphic DNA (RAPD) variability were examined in cultures of B. tabaci collected from the epidemic and non-epidemic zone sites. In a series of reciprocal crosses set up between combinations of ten B. tabaci cultures from the two zones, results indicated that there was no mating barrier. The offspring of these crosses were self-crossed to confirm that the hybrids were fertile. Genetic fingerprints of the whitefly populations generated by RAPD-PCR amplification with six 10-mer primers were collated by cluster analysis and the epidemic and non-epidemic zone cultures clustered into a single large group. The fecundity and developmental period of the epidemic and non-epidemic zone B. tabaci were compared on four cassava varieties and no statistically significant differences were obtained either in their fecundity, nymphal development times or numbers surviving to adult eclosion.