DEFRA Project HH3227TFV

Clubroot control using novel and sustainable methods

Project Manager: Rob Harling, SAC

  1. Summary of Progress

SUMMARY

In year 3 (05-06) of this project, two glasshouse and three field experiments were conducted to test treatments for efficacy against clubroot, using calabrese modules (rooted seedlings of 4-6 weeks old) throughout. We continued to refine treatments for controlling clubroot – the strategy has been to test as many treatments as possible in the glasshouse, eliminate those that were ineffective, and take those that were effective forward into the field. Further refinements have been made to the treatments this year, both in the glasshouse and in the field, by manipulating dose rates of products and combining treatments. We aim to end up with the best treatments currently available, and to make recommendations to growers accordingly: this is a practical project designed for immediate uptake by growers.

The most effective treatments were those containing calcium (lime), i.e. calcium carbonate, both as quarried product and as sugarbeet waste lime (marketed as LimeX by British Sugar), calcium oxide and calcium cyanamide (marketed as Perlka). These products were more effective than the fungicide Shirlan (fluazinam) in field trials. In the controlled conditions of the glasshouse, these calcium products almost completely eliminated disease; Shirlan was also effective in the glasshouse. Under moderate disease conditions in the Lincolnshire field trial, LimeX and calcium cyanamide gave 97 and 95% control respectively. Under severe disease pressure in Fife, these treatments gave 18 and 33% control respectively. Fineness of the lime products is essential to their success, as this ensures quick release of the calcium ions and a rapid rise in pH which needs to be above 7.5. In glasshouse tests, we were able to manipulate Ca2+ levels independently of pH and it is clear that control requires both high Ca2+ and pH levels together to achieve inhibition of the early stage of root infection.

We have obtained real-time PCR primers under an agreement with CSL York and have successfully used these to detect P. brassicae in natural and artificially infested soils.

EXPERIMENTAL DETAILS AND RESULTS

Experiments 1 and 2 below were carried out by ADAS, 3-6 were carried out by SAC.

  1. Fungicide drenches applied to calabrese modules in trays pre-planting (glasshouse experiment)

This is an efficient, cost-effective and environmentally-friendly way of applying treatments as it uses very small amounts of product. However, it renders new roots unprotected once they grow out into infested soil upon transplanting. Although we tried many novel products in the previous two years of the project, only conventional fungicides showed any potential when applied in this way.

The fungicides Shirlan (fluazinam), Ranman (cyazofamid) and Amistar (azoxystrobin) were tested at four different rates each (3,6,9,12l/ha). In addition, a drench of calcium hydroxide (CaOH2) at four rates (0.5,1.0, 1.5 and 2.0g/l) was tested; CaOH2 provides Ca2+ ions and increases pH, both of which reduce infection.

Results: Only Ranman gave consistently significant reductions in disease (60 – 80% reduction, depending on dose) when the treated modules were subsequently planted into infested soil. One of the CaOH2 treatments also gave a significant reduction but this was considered anomalous as there was no dose response observed.

Ranman has consistently been the best fungicide in our glasshouse and field trials, but unfortunately the manufacturer has no plans to support an application to the Pesticides Safety Directorate for approval for its use against clubroot; it is currently only approved for blight in potato. Although the other two products were ineffective, higher rates of application might be considered in future work as no phytotoxicity was observed even at 12 l/ha.

  1. Treatments for clubroot control in calabrese: field experiment Lincolnshire

The site chosen was a grower’s farm at Kirton, Lincs, naturally infested with clubroot. Last year’s most effective treatments were those containing calcium – these products provided Ca2+ ions and also raised soil pH levels. This year we tested four Ca–containing products, calcium oxide, calcium carbonate, sugarbeet waste lime (LimeX) and calcium cyanamide, all at different doses. Other treatments tested were rye companion planting alongside the calabrese (as a decoy for pathogen spores), chitin (crushed seashells, combined with LimeX) and the fungicide Shirlan, which, if effective, would be the easiest fungicide product to gain approval for, as the manufacturer has indicated interest in doing so, and it already has approval for soil application for other crops.

Results: Natural clubroot levels at this site were much less than at the Fife site (see below), untreated plots showed 30% disease compared with 93% at Fife. Starting pH at Lincs was 8.0, compared with 6.5 at Fife, the higher pH at Lincs being beneficial to control.

All calcium products gave good control of clubroot with the best treatments, LimeX at 4t/ha and calcium cyanamide at 1t/ha (split doses of 0.5 and 0.5t/ha applied two weeks apart) giving 97 and 95% control respectively. Addition of the fungicide Shirlan to these calcium products had no additional control effect – in fact a high dose of Shirlan actually worsened disease above the controls; an effect we saw last year in Fife due to phytotoxicity.

Companion planting with rye gave some small but non significant control, and also interfered with calabrese establishment because of competition.

Chitin did not have any effect this year, unlike last year.

  1. Treatments for clubroot control in calabrese: field experiment Fife, Scotland

The field site at Crail was chosen in consultation with East of Scotland Growers (ESG), Cupar, Fife, who provided site preparation, module transplanting and application of treatments. As at the Lincs site, treatments concentrated on the use of calcium-based products and some treatment combinations including fungicides. Other treatments were spent mushroom compost at high incorporation rate (90t/treated ha), Shirlan and Amistar, chitin and rye companion planting. Unlike at Lincs they were applied using special tractor-mounted equipment which incorporated the treatments into the soil in a band 20cm wide by 10 cm deep. The calabrese modules were transplanted into the middle of this band, and this had the effect of concentrating the treatments around the roots of the plants as the roots grew out of the modules. This technique has been used successfully for clubroot control in Australia, and ESG have been experimenting with it for their own crops in Scotland.

In addition, this site was chosen for an experiment to determine the speed of calcium release and pH rise in the soil following treatment application.

This was a very late crop, planted 21 July and harvested 18 October, hence the delay in preparing this report.

Results: The site proved to be severely infested with clubroot, and too much of a challenge for most of the control treatments. Untreated disease levels were 93%. High temperatures and drought for three weeks at the start of the trial undoubtedly stressed the crop and made disease worse. Nonetheless, three treatments gave significant but small reductions in disease: calcium cyanamide, LimeX and Shirlan plus chitin in combination, with 33, 18 and 18% reduction respectively.

Baseline (starting) pH and calcium levels at this site were 6.5 and 1720mg/l respectively. The most effective treatments at raising pH and available calcium levels were calcium oxide and LimeX, giving a rapid rise in pH at 3 days to 8.7 and 7.8 respectively, and 7200 and 6500 mg/l Ca2+ respectively.

  1. Treatments applied to the soil (glasshouse experiment)

This experiment evaluated treatments applied to infested soil in pots, as opposed to treatments applied to the modules themselves (i.e. 1. above) – soil treatments are likely to be more effective because roots grow out into infested soil and thus make contact with both pathogen and treatment. In module treatments, the active ingredient is contained in the module only, thus leaving the growing roots unprotected. Treatments were applied to the soil at transplanting, or 2 weeks prior to transplanting (for those requiring time to become active).

A total of 48 different treatments were tested, with replication, making this a large and time consuming experiment. Promising treatments identified in the first two years of the project were taken forward in year 3 and tested in combination as well as individually, to determine if 2- and 3-way combinations were more effective than individual treatments.

Results: Once again, the most effective treatments all contained calcium, i.e. CaO, CaCO3, sugarbeet waste lime (LimeX), calcium cyanamide and chitin: these treatments completely, or almost completely, eliminated disease in these pot trials – results were more dramatically effective in the controlled conditions of the glasshouse, compared to the field. Besides calcium-based products, three other treatments significantly reduced disease: the fungicide Shirlan, the biological control agent Pseudomonas chlororaphis, and spent mushroom compost. Regarding treatment combinations, the only combinations which were effective were those containing the above calcium products and there was no improvement in the control effect achieved in combination – it’s clearly not possible to improve upon the (near) 100% control obtained with the calcium products used alone.

  1. Treatments for clubroot control in calabrese: field experiment using clubroot-infested microplot beds at SAC Aberdeen

At SAC Aberdeen, beds are maintained with clubroot infestation by growing Brassicas every year; these beds are predominately used for trialling cultivars for clubroot resistance. Plot sizes are smaller than with conventional field experiments, 1.2 x 1.8 m vs 1.8 x 5 or 10m and employ seedlings rather than modules because of the smaller plot size. We established a trial this year with the intention of applying the same treatments to the same plots again next year to look at their effectiveness in reducing clubroot inoculum over a 2-year period. We assessed disease this year using the traditional Chinese cabbage bait test, before and after treatments were applied, and will do so again next year.

Results: Four treatments reduced clubroot levels in the beds as assessed by the bait test: calcium carbonate (53% reduction compared to control), Shirlan (63% reduction), calcium cyanamide (53%) and the combination chitin, calcium carbonate and Shirlan (65%). Chitin when used alone had no effect, therefore in the combination treatment, the reduction was due to the calcium carbonate and Shirlan. Calcium cyanamide killed the seedlings which then had to be replanted, therefore the result with this treatment has to be considered in view of the fact that the plants had less time to become infected than with other treatments.

  1. PCR diagnostic test for pathogen detection

Last year we obtained for the first time in the project, real-time PCR primers for P. brassicae under a commercial agreement from CSL York. At CSL these primers are used to detect and quantify the pathogen in plants, but our aim is to use them for detection and quantification in soil. In this 3rd year of the project, we used these primers to successfully detect the pathogen in three naturally infested field soils as well as artificially infested compost. DNA for these tests was extracted from 10g soil samples using a commercially available kit, PowerSoil Max, from MoBio. Pathogen levels were not quantified as this requires calibration of pathogen levels – this is in progress.

We obtained a protocol for field sampling and DNA extraction from 60g soil samples from Alison Lees, SCRI Dundee; this protocol was designed with BioSS to be statistically valid for sampling soil-borne pathogens for PCR testing. Using this protocol, which employs a Retsch mill for soil grinding (an expensive tool, which SAC Aberdeen have invested in), DNA was successfully extracted from 60g soil samples from naturally infested fields, and from soil which had been artificially inoculated with 103 to 107Pb spores per gram of soil. During year 4, the real time primers will be applied to these samples to create a calibration curve for quantifying the pathogen in soils of unknown infestation levels.

CONCLUSIONS

Treatments applied to modules were not able to protect roots from becoming infected once they had grown away from the treated root ball and out into infested soil – which is not surprising. The one exception to this was the fungicide Ranman, but unfortunately, the UK industry is unlikely to see this product approved for Brassicas. Shirlan has some effectiveness as a soil, but not a module treatment, and if rates in modules were to be increased, then it may have some effect, although there are known phytoxicity issues with this product (not, however, seen at 12l/ha this year, which is a high rate, although we did see phytotoxicity at 9l/ha in the field in 2005).

Field and glasshouse experiments once again confirmed the effectiveness of calcium-based products for clubroot control. These products are long established and work through a combination of raising Ca2+ levels and pH in the soil – but, as growers will testify, they are not always effective. As part of this project, we have conducted a literature review of the use of calcium products and, together with the results obtained in this project, will soon be in a position to provide new recommendations to growers for the use of these products to increase their effectiveness.

One of the new findings has been the importance of a calcium and pH burst in the soil, shortly after planting, to inhibit root infection; at present, growers apply lime at least a month in advance with the belief that a time delay is necessary for the lime to act (release calcium and raise pH), but as our soil analyses show, lime acts very quickly, within 3 days of application if in fine powder form (this is vital), and by tying-in other scientific reports, we have found that application of calcium and raising the pH within the first 7 days is crucial if infection is to be avoided. These recommendations will provide new information on what products work best, when to apply them, and how much to apply. As part of our forthcoming team meeting we will discuss these recommendations, but are confident of being able to conclude them by the end of the project. The efficient use of calcium products is consistent with Defra’s aims for promoting alternatives to pesticides.

The PCR test will provide a useful adjunct to the control measures by assisting growers with identifying affected fields and, importantly, for quality control in the production of modules on nurseries.