BSPP Presidential Meeting 1999

Biotic Interactions in Plant-pathogen Associations

Offered Posters

Percolation, heterogeneity and the saprotrophic invasion of soil by the fungal pathogen Rhizoctonia solani.
Doug Bailey, Wilfred Otten, and Chris Gilligan

Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK.

The ability of certain soil-borne fungal plant pathogens to spread saprotrophically between discrete sites of organic matter on or within the soils is well known and can have a profound effect on the outcome of an epidemic. The extent of saprotrophic spread depends on the chance or probability of the fungus growing from a colonised site to a neighbouring uncolonised site. Based on this probability, percolation theory predicts criteria for either invasive or non-invasive spread of the pathogen amongst a population of sites. We use percolation theory to demonstrate the existence of a threshold probability of transmission for the fungus, Rhizoctonia solani between neighbouring sites in two simple experimental systems involving either agar or dead poppy seeds. We conclude that percolation theory can be used to link the growth of individual mycelial colonies to the formation of patches that result from the colonisation of particulate organic matter.


Effect of host biomass and growth stage on the development of Septoria tritici blotch in wheat.
C.M. Bennett, F.J . Bannon and B.M. Cooke

Department of Environmental Resource Management, Faculty of Agriculture, University College Dublin, Belfield, Dublin 4, Ireland. 
E-Mail
: finian.bannon@ucd.ie

Many factors are known to influence the development of Septoria tritici, including plant growth stage at time of infection, which is of significant importance and has a considerable effect on subsequent disease development and grain yield of wheat. The present investigation examined the effect of sowing date and time of inoculation on spore load build-up and development of Septoria tritici blotch (STB) throughout a growth season (1998/99) on the winter wheat cv. Sponsor. Inoculations at different times throughout the season were carried out to manipulate spore load in the crop due to varying substrate availability. Dry matter was estimated at time of inoculation and at various times throughout the season; estimates of spore load were carried out concurrently. Disease development was monitored by assessing disease severity on the flag, second and third leaves. Preliminary results show that sowing date significantly affected spore load density. Transformed data (log+1) for the number of spores per g biomass showed a highly significant interaction between sowing date and time of inoculation. It was concluded, however, that disease levels of STB were too low to cause significant yield effects. Analysis of data revealed that the relationship between the amount of biomass present at inoculation and the development of spore load in the crop is a complex scenario composed of many possible confounding factors.


Effects of temperature and wetness duration on infection of oilseed rape by ascospores of A-group or B-group Leptosphaeria maculans (Stem canker).
J.E. Biddulph1, B. D. L. Fitt1, M. Jedryczka2, J. S. West1 and S.J. Welham1

1 IACR-Rothamsted, Harpenden, Herts. AL5 2JQ, UK
2
Institute of Plant Genetics, 60-749 Poznan, Poland

Phoma lesions produced on oilseed rape leaves by B-group ascospores were smaller and less distinctive than lesions produced by A-group ascospores of Leptosphaeria maculans, when leaves were inoculated with ascospore suspensions obtained from infected debris from Poland or the UK. Both A-group ascospores and B-group ascospores of L. maculans were able to infect leaves of oilseed rape and produce lesions at temperatures from 4 to 20oC and leaf wetness durations greater than 4 h. However, the greatest number of lesions were produced with a leaf wetness duration of 48h at temperatures of 20oC for A-group and >12oC for B-group ascospores. As leaf wetness durations and temperatures decreased below the optimal values, the numbers of lesions produced decreased. For example, very few lesions were produced at 4oC (and only with a leaf wetness duration >48 h) or with a leaf wetness duration of 4h (and only at temperatures >12oC). There was no evidence that the maximum number of lesions produced in relation to number of ascospores inoculated differed between A-group and B-group L. maculans. The incubation period (from inoculation to the appearance of the first lesion) of B-group L. maculans was shorter than that of A-group L. maculans; at 20oC, lesions appeared within 2 days rather than 5 days from inoculation. As temperature decreased below 20oC, the length of the incubation period increased.


Selection of biological control agents for control of Allium white rot
Simon P Budge, Tina J Payne, John P Clarkson & John M Whipps

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

White rot caused by S. cepivorum is one of the most important diseases of Alliums world-wide. However, control is difficult as there are few effective fungicides and fumigation methods are costly, unreliable and becoming restricted. This lack of control measures has resulted in a large reduction in disease-free land suitable for growing alliums. No single control measure is likely to be successful in controlling white rot and a fully integrated system of disease management is more likely to achieve a long-term solution. Part of such a system could include biological control.

A step-wise screening system to select fungi with potential to control white rot was developed. Fungi isolated from diseased onions were initially assessed on agar drops for their ability to prevent mycelial growth and development of sclerotia or reduce survival of mature sclerotia. Of the109 isolates tested, virtually all prevented mycelial growth and 45 caused softness in over 90% sclerotia when inoculated at the time of sclerotial initiation. These isolates were chosen for assessment in two further tests designed to evaluate their ability to degrade mature sclerotia in soil or prevent disease development on onion seedlings. In both tests, isolates were applied either as spore suspensions or wheat bran germling inoculum. In the first test, the isolates were applied to soil containing bags of sclerotia and the number of healthy sclerotia assessed after 8 weeks. In the second test, the isolates were applied to soil containing sclerotia in which salad onions were sown and disease assessed after 8 weeks.

In the sclerotia infection tests, 8 isolates reduced the viability of mature sclerotia by over 50% and in the seedling bioassay, over 50% disease control was achieved by 7 isolates. In both assays, greater activity was obtained with the wheat bran germling inoculum than with spore suspensions. The 15 best performing isolates were retested in both assays. Four isolates consistently ranked in the top five for activity and gave similar levels of control, with viability of sclerotia being reduced by over 60% and disease controlled on onion seedlings by over 50%. These four antagonists are currently being assessed in different soil types and against different white rot isolates before testing in small scale field trials.


Raspberry bushy dwarf virus in Scotland
Chard, J1, Irvine, S1, McGavin, W2, Nevison, I3, Roberts, A3, Langrell, S1 & Jones, A T2

1Scottish Agricultural Science Agency, East Craigs, Edinburgh, EH12 8NJ
2
Scottish Crop Research Institute, Invergowrie, Dundee, DD2 5DA
3Biomathematics and Statistics Scotland, James Clerk Maxwell Building, Kings Buildings, Edinburgh, EH9 3JZ

A survey of raspberry fruiting plantations was done in 1998 to determine whether raspberry bushy dwarf virus (RBDV) was established in plantations in Scotland. Raspberry-producing holdings were selected according to geographical area and size. Samples (202), each comprising 60 stems per stock, were obtained from 77 holdings and tested by ELISA. ELISA-positive stems from each infected stock were grafted onto an indicator cultivar, Glen Clova, to establish whether the virus was a resistance-breaking (RB) isolate.

RBDV was detected in 22 % of stocks, with levels of infection ranging from 2 80%. No RBDV was found in any of the stocks of cv. Glen Clova tested (33) or in cv. Glen Clova plants successfully grafted with samples from 14 infected stocks. This shows that no RB isolates were detected. The percentage of infected stocks increased with time from planting date. Overall, there was a higher incidence of RBDV in stocks from Perthshire than from Angus.

In order to investigate possible sources of RBDV infection, samples of all raspberry stocks entered for the lowest certified grade (Standard Grade) in Scotland were tested in 1999. No RBDV was detected in any of the samples. RBDV was found rarely in samples from wild Rubus.


Forecasting onion downy mildew
John P Clarkson, Roy Kennedy & Kath Phelps

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

Peronospora destructor causes downy mildew which is a major disease of onions world-wide. Control relies on up to seven routine applications of protectant and eradicant fungicides but maintaining control throughout the life of the crop and timing applications effectively is difficult. In addition, reducing fungicide applications on onions is extremely desirable for the environment and consumer. A forecasting model for onion downy mildew called Downcast has been developed based on the prediction of sporulation and infection periods according to weather criteria. Although this model has been refined, many such periods may be predicted within one season which can result in little reduction in fungicide use. The latent period of P. destructor has been identified as another major limiting factor in downy mildew epidemics. If the length of latent period could be determined after an infection event, this would allow predictions of when P. destructor had developed sufficiently in plants to sporulate. In combination with Downcast, this may help disease prediction and further reduce fungicide applications. This work set out to develop a simple latent period model for P. destructor based on temperature.

Onion plants infected with P. destructor were placed in controlled environment cabinets at different temperatures between 6 and 25C at 80% relative humidity and 12hr day/night. At regular intervals, plants were removed and misted overnight to promote any downy mildew sporulation. A model was then derived from the time taken for 50% of the plants removed to show sporulation of P. destructor at each temperature. To test the model, plants infected with P. destructor were placed outside immediately after inoculation every week and observed for sporulation of downy mildew daily. Observation of sporulation not only indicated completion of the latent period, but also indicated suitable overnight conditions for spore production. Temperature and other weather parameters were measured with a logger. The sporulation observed on exposed plants was then compared with predictions by the latent period model adjusted to the nearest night suitable for sporulation. This was determined by observation of onion plants infected for 2 weeks in the glasshouse (latent period completed) which were exposed outside daily.

The optimum temperature for downy mildew development was approximately 21C where sporulation was observed on 50% of plants after 8 days. Sporulation on 50% of plants was seen after 9 days at 23C but no sporulation ever occurred on plants incubated at 25C. The relationship between length of latent period and temperature (T) was determined as a rate (1/time to 50% of plants with sporulating downy mildew) and was quadratic. From 21 weekly exposures of infected onion plants, the latent period model predicted 12 within 1 day, 7 within 3 days and 2 within 4 or 5 days of observed sporulation. There was therefore good agreement between the model predictions (using the nearest night suitable for sporulation) and observations of sporulation. Further work will now test the efficacy of combining the latent period model with Downcast to reduce fungicide sprays.


Synthesis of infectious transcripts from a full-length cDNA clone of grapevine virus A and molecular characterization of the viral proteins and RNAs.
N. Galiakparov,1 O. Aziz,1 E. Tanne,1 I. Sela2 and R. Gafny1

1Agriculture Research Organization, Bet Dagan Israel and 2 Faculty of Agriculture, The Hebrew University of Jerusalem, Rehovot, Israel

Vitivirus is a newly established genus of plant viruses. Grapevine virus A (GVA) is the type species of the genus. GVA is associated with Kober stem grooving of the grapevine rugose wood disease complex. GVA has a single-stranded RNA genome of 7349 nucleotides and a polyA tail at the 3 terminus. The GVA genome includes 5 open reading frames.

To investigate the role of the various GVA encoded genes we have constructed a full-length cDNA clone of GVA, from which infectious RNA transcripts can be produced. Initial characterization of the clone indicated that a coat protein deficient mutant replicated in Nicotiana benthamiana protoplasts but failed to infect intact plants; suggesting that the coat protein is required for cell to cell movement of the virus. This is the first report of infectious transcripts for a member of the Vitivirus genus.

RNA probes derived from different regions of the GVA genome were used to detect GVA associated RNAs. Four double-stranded RNAs (dsRNA) with 3 termini common with the genomic RNA were detected. These are suggested to be the dsRNAs molecules corresponding to the ORFs 2-5 subgenomic mRNAs. A group of 3 dsRNAs that have a common 5 termini with the GVA genomic RNA were also detected. The role of these unusual, 5 coterminal RNAs, in the virus life cycle is unknown.


Identification of three viruses on Pisum sativum (pea) in South Africa
A.E.C. Jooste, G. Pietersen and G.G.F. Kasdorf

ARC-Plant Protection Research Institute, Private Bag X134, Pretoria, 0001, South Africa

Three virus isolates obtained from pea plants (Pisum sativum) from different regions in South Africa during the 1991-1995 growing seasons were identified by a number of techniques including serology, electron microscopy, cytopathological studies, host range studies, PCR, cloning, sequencing and sequence comparisons. The first pea sample (91/0394) was collected in the George district, Western Cape, and spherical particles were detected in the plant with electron microscopy. The infected plant showed distinctive symptoms and was identified as Pea enation mosaic virus (PEMV) based on serological results with antiserum to the PEMV 3LM strain. A specific antiserum was prepared to the virus. A second pea plant (94/1969) from a pea producer in the Brits district of the North West Province, showed severe mosaic symptoms. Electron microscopy revealed particles typical of Broad bean wilt virus (BBWV). Antisera to known isolates of BBWV were used to positively identify this virus as BBWV serotype II. As in the case of PEMV an antiserum was made to this isolate. The third virus (95/0931) was collected from field trails at the Vegetable and Ornamental Institute at Roodeplaat, Pretoria. The plant, with yellow mosaic and vein clearing, showed flexuous particles under the electron microscope and was tested in ELISA to five Potyviruses. The test was positive for Bean yellow mosaic virus (BYMV). Serological cross reactivity between different Potyviruses prompted the characterisation of this virus on a molecular level. The 3' untranslated region (UTR) and part of the coat protein were amplified by IC-RT-PCR: a PCR product of 714 bp, the expected size for BYMV was obtained. This was cloned and sequenced. The nucleic acid sequence results were aligned with cognate sequences of several Potyviruses. The nucleotide and derived amino acid sequences of isolate 95/0931 showed a 99% and 100% respective similarity to comparable regions of the Pea mosaic virus-I (PMV-I) strain previously sequenced. The PMV group shares 90% and 96% similarity with the BYMV subgroup at the nucleotide and amino acid level, respectively, and is accepted as a strain of BYMV. Isolate 95/0931 is therefore identified as a strain of BYMV most closely related to PMV.


Forecasting light leaf spot of winter oilseed rape in the UK
N. Evans1, B.D.L. Fitt1, P. Gladders2, S.J. Welham1, J.A. Turner3 and K. Sutherland4

1IACR Rothamsted, Harpenden, AL5 2JQ, UK.
2
ADAS Boxworth, Cambridge, CB3 8NN, UK
3
Central Science Laboratory, MAFF, Sand Hutton, York, YO41 1LZ, UK
4
SAC, Craibstone Estate, Bucksburn, Aberdeen, AB21 9YA, UK

Light leaf spot (Pyrenopeziza brassicae) is a serious disease of winter oilseed rape crops in the UK. Wind blown ascospores from crop debris of the previous season are thought to initiate infection in newly sown crops. The pathogen enters a hemibiotrophic phase during which infections are not readily visible. Localised spread in the spring occurs through splash-dispersed conidia. Assessment data from regions of the UK were used to produce a model to predict the risk of a crop in a specific region of the UK developing light leaf spot. The forecast is based on crop and weather factors. At the start of the season, a prediction is made for each region using the average weather conditions expected for that region. This forecast is then updated periodically to take account of deviations in actual weather away from the expected values. Three factors form the basis of the model: amount of pod disease the previous summer, autumn temperatures and the number of winter rain days above the regional average. The model is currently in a third year of evaluation. Recently, an interactive model has made available to growers over the internet (http://www.iacr.bbsrc.ac.uk/res/depts/plantpath/Lls/index.html). This allows growers to make the forecast more crop specific through the input of information on cultivar, sowing date and autumn fungicide applications.


The cause and incidence of sweet potato virus disease in Uganda.
Richard W. Gibson, R. O. M. Mwanga, V. Aritua & E. E. Carey.

Natural Resources Institute, Medway University Campus, Central Ave., Chatham Maritime, Kent, ME4 4TB, UK. 
E-Mail:
r.w.gibson@gre.ac.uk

Sweet potato virus disease (SPVD) is the main disease of sweet potato in Uganda and is caused by the synergistic interaction of two viruses, Sweet potato feathery mottle potyvirus (SPFMV) (Potyviridae), and Sweet potato chlorotic stunt crinivirus (SPCSV) (Closteroviridae). In common Ugandan sweet potato cultivars, SPFMV by itself causes no obvious symptoms, SPCSV may cause a yellowing or reddening of middle leaves and slight stunting but combined infection with the two viruses induces the very severe symptoms of SPVD. Symptoms vary with plant genotype but typically include stunted plants with small distorted leaves, the latter often also being distorted, narrow (strap-like) and crinkled with a chlorotic mosaic and/or vein-clearing giving affected plants an overall pale appearance.


Spread of fungi through soil: can soil physical conditions affect epidemics?
Wilfred Otten1
, Chris Gilligan1, Darroch Hall1, Kirsty Harris2, Iain Young2 and Karl Ritz2

1Department of plant sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK.
E-Mail:
wo200@cus.cam.ac.uk
2
Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, Scotland, UK.

The invasion of soil and spread of diseases caused by soil borne fungi such as Rhizoctonia solani depend on the ability of fungal hyphae to colonise available hosts such as roots, hypocotyls or fragments of organic matter. Until now, not much work has been undertaken that quantifies the spread of fungi through soil with explicit characterisation of the soil structure within which fungal hyphae spread and biological interactions occur. We quantified the spread through soil of the fungal plant pathogen Rhizoctonia solani and conclude that physical conditions, which are influenced by soil type, tillage and water content can have profound effects on the dynamics of epidemics through their influence on fungal growth, and may account for why epidemics occur in some soils and not in others. The effects are mediated through the connectivity and tortuosity of the air-filled pore space which can be controlled by aggregate size distribution, bulk-density and water content.


Optimising the use of fungicides to control stem canker of oilseed rape
J.S. West1*, P.K. Leech1, B.D.L. Fitt1, S.J. Welham1, A. Penaud2, and A. Prs2, B. Poisson2

1IACR - Rothamsted, Harpenden, AL5 2JQ, UK
2
CETIOM, -BP4, 78850 Thiverval-Grignon, France

*
E-Mail: jon.west@bbsrc.ac.uk

Ascospores of Leptosphaeria maculans infect leaves of oilseed rape to cause leaf spots, from which the fungus can grow to infect the stem. Yield losses due to early senescence and lodging result if the stem infections reach a threshold severity prior to harvest. Host resistance alone has not been sufficient to control the disease in the UK where two forms of the fungus [A group (Tox+) and B group (Tox0)] occur, each with different pathogenicity groups. Currently, approved fungicides do not eradicate mycelium once it is inside the stem, so disease management relies on protection of the leaves. Fungicides on the leaf surface degrade and are diluted by leaf expansion, while new leaves are unprotected. The mild, wet climate of the UK and parts of France promotes the release of a succession of spores from different parts of stem debris. Therefore, one or more well-timed fungicide applications are needed. Weather data from different seasons were compared with the corresponding airborne spore numbers, leaf lesion incidence and canker incidence and severity. At early crop growth stages it is important to apply fungicides very promptly, if infections occur during this period. However, when an epidemic is late and the crop is more mature when leaf spotting first appears, a longer period of time exists in which a fungicide application can achieve economic control. Accurate forecasting of fungicide application can therefore not only improve disease control but also decrease fungicide use when the risk of crop damage is small.


Plant pathology in the new HGCA R&D strategy for cereals
Roger H. Williams & Graham J. Jellis

HGCA, Caledonia House, 223 Pentonville Road, London, N1 9HY, UK

The objective of the HGCA programme of levy-funded R&D is to improve the competitiveness and profitability of the UK cereals industry whilst limiting its environmental impact. Within the framework of the previous cereals R&D strategy, published in 1995, research projects dealing with disease control have contributed significantly to achieving this objective. Since 1995, the UK cereals industry has experienced a number of important changes including:

  • Common Agricultural Policy reform;
  • Increased consumer concerns for the environment and for food safety;
  • The introduction of increasingly sophisticated technology;
  • The introduction of hybrid wheat and, potentially, genetically modified cereals.
  • Consequently, it has been necessary to review the cereals R&D strategy to ensure the continued effectiveness of levy-funded research. The new cereals R&D strategy contains many significant changes in direction and priority from the previous strategy. Nevertheless, plant pathology remains an important component with relevance to each of the six key areas of research as illustrated below:

  1. Reducing production costs. Maximising efficiency of fungicide applications to optimise use of labour and machinery.
  2. Meeting user requirements. Ensuring that crop management in the field minimises the risk of mycotoxin formation.
  3. Farming systems and the environment. Determining the implications of changes in crop rotations for disease management. Optimising disease control in organic cereal production.
  4. Variety evaluation and crop improvement. Assessing disease resistance in new varieties.
  5. Crop management. Anticipating and responding to changes in disease threats, and assessing the benefits of new fungicides.
  6. Advanced technologies. Developing tools for pathogen detection and their use in forecasting disease threats. Implementing decision support systems.

Scientists wanting to contribute to the HGCA R&D programme should contact R&D staff early in the preparation of applications to discuss project ideas. Submission of concept notes (1 or 2 pages) is strongly encouraged. Application forms and further information can be found at http://www.hgca.com.


The study on the pathogen of tobacco black death disease

Guangmin Zhang & Zhifa Wang
Shandong Agricultural University, Taian, Shandong 271018, P. R. China

Tobacco black death is one of the major tobacco diseases in the Weifang region of the Shandong province of China. The range of growth temperature of the pathogen was 3-38 C, and suitable was 25-30 C, and the optimum temperature for growth was 28 C. The pathogen could infect 19 species in 9 families of 33 species in families of plants by inoculation in greenhouse. The disease symptoms on Nicotianae tobaccum and N. glutinosa was the same as that in the field. But on other hosts were local necrosis or wilt. On the PDA medium, the colonies had the characteristics of circular shape, smooth edge, and were 81 mm in diameter cultured for 10 days under 25 C. Aerial mycelia grew tightly from white to greyish-white. A few isolates were in dark grey on top of colonies but the colour on the bottom were the same as others. Branchy mycelia grew rapidly and chlamydospores were formed in aged mycelia. Acervuli grew intensively under epidermal layers of hosts, and were 37.8 146.1 μm in diameter. Scattered setae were dark grey and erect and their tops were sharp. Each seta, 66.5-181.4 x 3.2-6.1 μm, had 35 septa. Cylindrical conidiophores grew intensively on cultured mycelia or acervulus, and conidiogenous cell was phialide. Half moon-shaped conidia had the characteristics of single cell, no colour, two sharp ends, 16.2-24.3 x 3.5-6.2 μm. Then conidia germinated and produced germ tubes from its one end or two ends, a septum was formed in the middle. Light or dark brown, ovaide appressoria with entire or slightly irregular margin, 7.6-10.1 x 5.1-56.5 μm, were produced on the top of germ tubes or their branches. 

Based on the characters described above the fungus was named as Colletotrichum capsici (Syd) Butle & Bisby f. nicotianae G.M. Zhang & G.Z. Jiang f.nov., which parasitised the stem and leaf of Nicotiana tobaccum L.


Yield loss of winter oilseed rape in relation to severity of stem canker (Leptosphaeria maculans) in the UK.
Y. Zhou1, B. D. L. Fitt1, S. J. Welham1, P. Gladders2, C. E. Sansford3 & J. S. West1

1IACR-Rothamsted, Harpenden, Hertfordshire AL5 2JQ, UK
2
ADAS Boxworth, Cambridge CB3 8NN, UK
3
Central Science Laboratory, MAFF, Sand Hutton, York YO41 1LZ, UK
 

The relationships between yield loss and incidence or severity of stem canker in winter oilseed rape were analysed using data from experiments at Rothamsted in 1991/92, Withington in 1992/93, Boxworth in 1993/94 and Rothamsted in 1997/98. Critical point models and area under disease progress curve (AUDPC) models were better than multiple point models for describing relationships between yield (t ha-1) and incidence or severity of stem canker. Since yield in t ha-1 is influenced by many factors other than disease, % yield loss was calculated. Critical point models and AUDPC models relating % yield loss to stem canker were constructed and models for different experiments were compared. A critical point model relating % yield loss to stem canker incidence (GS 6.3-6.4), and AUDPC models for incidence or severity from growth stages 4.8 to 6.4 were derived from the combined data sets for the three experiments. The relationships between % yield loss and % plants with different stem canker severity scores at different growth stages, and between % yield loss and incidence or severity of both basal stem cankers and upper stem lesions were also analysed.