^*email: mailto:jon.west@bbsrc.ac.uk

ABSTRACT

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.

INTRODUCTION

The fungus Leptosphaeria maculans (Desm.) Ces & De Not. causes stem canker (blackleg) of winter oilseed rape. Pseudothecia are produced on stubble and release ascospores from the autumn onwards in the UK and France. The spores infect leaves to cause leaf-spots, from which the fungus can grow to infect the stem (Hammond et al.,1985). Severe stem infections, in which over 50% of the stem is girdled, can cause a substantial loss in yield due to early senescence and lodging. Two forms of the fungus occur in the UK; A (Tox⁺) which causes damaging cankers on the stem and B (Tox⁰) which infects the pith (Johnson and Lewis, 1994). Hammond and Lewis (1986) found that the most damaging cankers are formed from leaf infections occurring up to the onset of rapid stem extension. Additionally, the younger leaves (i.e. the first six) appear to be more susceptible to leaf infections than those produced later (McGee and Petrie, 1979). Therefore, it is important to apply fungicides, if infections occur during this period. It has been thought that fungicides do not affect L. maculans once it is inside the stem so it is important to protect the leaves (Gladders, 1988). However, it is not known up to which position in the leaf/petiole, that the fungus is susceptible to a fungicide application. Some reports indicate that fungicides applied to control leaf infections have not always prevented the development of cankers (Rawlinson and Muthyalu, 1979) and Hammond (1985) speculated that the fungus within the leaf may escape fungicides. If this is so, as some fungicides promote leaf retention rather than abscission, late fungicide applications may even be counterproductive by allowing the fungus to reach the stem. Fungicides protecting the leaves degrade and are diluted by leaf expansion, while new leaves are unprotected from later spore releases. Thus, several low dose applications would be needed to give full protection of all leaves, but for practical and economic reasons, only one or two fungicide applications are normally applied (and these may also control other diseases such as light leaf spot). Due to the small size of young leaves and the relatively warm temperatures of early autumn, it is important to apply fungicides very promptly if infections occur. Biddulph et al. (1998) showed that lesions start to appear after only 3 days at 20ºC, 5 days at 16ºC, and 13 days at 8ºC. Hammond (1985) found that at ≈≈12.5ºC lesions start to appear after 8 days, and in infections which were not confined by the host response, entered minor veins by 14 days. At 4ºC, Hammond (1985) found that the systemic phase of infection started after 30 days, with lesion appearance after 40 days. Thus, infections can be advancing towards the stem before any leaf lesions are obvious, while in other infections, the lesion is confined by the plant, although it may sporulate. Therefore, fungicide applications, in response to the first observation of leaf spots (commonly once 10% incidence is reached) may either be too late to prevent stem infection or not needed. Often spray applications, based on leaf spot presence, are delayed by bad weather and so are unlikely to control systemic infections.

The choice of timing of spray applications must depend upon the number of spores in the air (pathogen), favourable infection conditions (environment) and the resistance and age of the crop (host). This paper gives new perspectives of how the number of spores in the air can be predicted and monitored, as part of a disease forecasting scheme.

METHODS

Maturation of pseudothecia was assessed on stubble collected immediately after harvest and incubated in trays outdoors. Periodically, five pseudothecia from each of five stubble sections were excised, place on slides and examined microscopically. Pressure was applied to the coverslip to squash the pseudothecia and reveal their contents which were graded according to the scale of maturation used by CETIOM; where class A = undifferentiated, B = immature asci present but spores are undifferentiated, C = <8 spores per ascus and < 4 cells per spore, D = 8 spores per ascus or > 4 cells per spore. (Additionally, E = empty/discharged fruiting body).

Furthermore, the numbers and pattern of ascospore discharge, detected by Burkard spore samplers located next to sources of inoculum, were compared with the leaf spot incidence observed over two seasons and two sites.

RESULTS

Maturation of pseudothecia - (Table 1) The first pseudothecia were observed at the sites of stem cankers; later infections (stem lesions) tended to produce pseudothecia later, which matured later (data not shown). This resulted in variable data on pseudothecia maturation; e.g. at St Pathus, class D pseudothecia were observed before class A, which means that regular assessments of relatively large numbers of pseudothecia would be required for a reliable forecasting scheme.

Table 1: Relationship between pseudothecial maturation, spore release and disease incidence

( * on plants with severe cankers only)

Ascospore discharge - Comparisons of records of airborne ascospores trapped and leaf spot incidence observed over several seasons (e.g. Fig. 1) revealed that leaf spotting was noticed before any major releases of spores were detected. It appears that the first spores released, although relatively few in number, lead to a relatively high incidence of leaf spotting, because they are deposited when the plant is at its most vulnerable stage.

Fig 1: The pattern of ascospore numbers in the air and leaf lesion incidence in a winter oilseed rape crop at Rothamsted, 1997/98

In the example (Fig. 1) leaf spotting was first noticed on 23/10/97 and was most prevalent that autumn. This led to the appearance of stem cankers from mid-April (data not shown). As in other seasons/sites, a decrease in the incidence of leaf spotting (in untreated plots) was noticed in the winter despite the presence of spores in the air at this time. Although at low temperatures it takes over a month for leaf lesions to appear on newly infected leaves, and older (diseased) leaves are shed at this time, in response to shading and frost, the reduction in leaf infection is also an indication that host resistance may increase with age.

DISCUSSION

An early sowing of the crop can be used to evade infection at the most sensitive period for the crop (LePage and Penaud, 1995) but there can be problems of early flowering and the formation of too dense a crop canopy. The crops in this study were sown in late August, as standard practice. Improving host resistance remains an important aim in the control of stem canker. As some leaf lesions are confined while others may lead to stem infections, knowledge of the aggressivity/compatibility of local populations of the pathogen on the predominant cultivar grown in a region would aid spray decisions. Work on this is in progress as part of the IMASCORE project (Balesdent and Rouxel, 1998; Balesdent et al., 1998)

Previously, timing of spore release was thought to be due to climatic factors immediately prior to the spore release. Additionally, it appears that the severity of infection in the previous crop influences pseudothecial maturation, since the first pseudothecia were observed at the sites of severe stem cankers, while sites of upper stem lesions produced pseudothecia later. There is potential for the population structure of the pathogen to change during the season as pseudothecia from different parts of the inoculum residue mature. This has also been suggested by Thürwächter (personal communication) and Hammond (1985). Pseudothecial matration depends upon temperature, wetness and humidity (Poisson, 1997) and the maturation time may also be influenced by host resistance (Pérès and Poisson, 1997) and any applied chemicals. An investigation of pseudothecial maturation (Pérès and Poisson, 1997) indicated that the first emissions of ascospores occurred 16-19 rain-days after harvest and when the average temperature had dropped to 14ºC. A combination of the date of first spore capture and weather conditions was used to produce a preliminary model to advise on spray timing at different sites. The monitoring of pseudothecial maturation, or modelling the severity of disease the previous season combined with the weather in late summer/early autumn, are possible methods for predicting the risk of spore release and the first occurrence of leaf-spotting. Additionally, the use of a Burkard spore sampler could augment a forecasting service, giving a warning of spore release in indicator areas.

The incidence of leaf infection has been correlated to the subsequent incidence of stem canker in some studies but not in others. Possible factors which increase the incidence of stem canker over that suggested by the leaf spot data are; direct infection of the stem by conidia splashed from stubble on the soil surface (little is known or has been measured), direct infection of the stem by ascospores, and undetected symptomless (biotrophic) leaf infections. Factors which may reduce the later incidence of stem canker over that suggested by leaf spot incidence are; leaf drop prior to stem colonisation, and total confinement of leaf lesions preventing systemic spread. Large differences appear to occur in symptom expression between cultivars and between crop growth stages and this has reduced the general level of understanding in the past. Additionally, further work on the importance of secondary infections by conidia should be established.

Accurate forecasting of disease severity can therefore not only enhance disease control by improved fungicide application but also reduce fungicide use when the risk of crop damage is low. Severity of disease the previous season combined with the weather in late summer/early autumn could be used to predict the first occurrence of leaf infections.

CONCLUSION

Knowledge of disease levels in the previous season, cultural practices, and cultivar resistance is required in the development of general crop risk indices. Monitoring the pseudothecial maturation/timing of spore release and favourable infection conditions can be used to pinpoint spray application timing.

ACKNOWLEDGEMENT

This study is part of the IMASCORE project, funded by the European Union (Fair contract CT96-1669; co-ordinator M-H Balesdent).

REFERENCES

1. Balesdent M-H, Rouxel T. 1998 IMASCORE: un project de recherche européen sur le “phoma du colza”. Les Rencontres Annuelles du CETIOM - Colza, 3 December 1998, 30-31

2. Balesdent M-H, Jedryczka M, Jain L. Mendes-Pereira E, Bertrandy J, Rouxel T. 1998 Conidia as a substrate for internal transcribed spacer-based PCR identification of members of the Leptosphaeria maculans species complex. Phytopathology 88: 1210-1217

3. Biddulph JE, Fitt BDL, Welham SJ. 1998 Effects of temperature and wetness duration on infection of oilseed rape leaves by ascospores of Leptosphaeria maculans (stem canker). 7^th International Congress of Plant Pathology, Edinburgh, Scotland, 9-16 August 1998, (2.5.13)

4. Gladders P. 1988 The contribution and value of pesticides to disease control in combinable break crops. pp 29-50 in: Control of Plant Diseases:cost and benefits, Clifford, BC, Lester, E, ed. Oxford, Blackwell Scientific Publications.

5. Hammond KE. 1985 Systemic infection of Brassica napus L spp. oleifera (Metzger) Sink. by Leptosphaeria maculans (Desm.) Ces. et de Not. PhD. Thesis, University of East Anglia, Norwich

6. Hammond KE, Lewis BG. 1986 The timing and sequence of events leading to stem canker disease in populations of Brassica napus var. oleifera in the field. Plant Pathology 35: 551 - 564.

7. Johnson RD, Lewis BG. 1994 Variation in host range, systemic infection and epidemiology of Leptosphaeria maculans. Plant Pathology 43: 269 - 277.

8. LePage R, Penaud A. 1995 Tout se joue avec le premier pic d’ascospores. CETIOM - Oléoscope N^o 28, 23-27

9. McGee DC, Petrie GA. 1979 Seasonal patterns of ascospore discharge by L. maculans in relation to blackleg of oilseed rape. Phytopathology 69: 586 - 589.

10. Pérès A, Poisson B. 1997 Phoma du colza: avancées en épidémiologie. CETIOM - Oléoscope N^o 40, 37-40

11. Poisson B. 1997 Etudes relatives “la maturation des perétheces de Leptosphaeria maculans sur les pailles de colza d’hiver nécrosèes au collet". ANPP - 5ème conférence sur les maladies des plantes, Tours, 3-4-5 Décember 1997, Tome 1 p345-352

12. Rawlinson CJ, Muthyalu G. 1979 Diseases of winter oilseed rape - occurrence, effect and controls. Journal of Agricultural Science, Cambridge 93: 593-606