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Causes of leaf spotting in Chinese

cabbage

Dennis Phillips Department of Agriculture

Western Australia

Project Number: VG02053

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VG02053 This report is published by Horticulture Australia Ltd to pass on information concerning horticultural research and development undertaken for the vegetable industry. The research contained in this report was funded by Horticulture Australia Ltd with the financial support of the Vegetable Industry. All expressions of opinion are not to be regarded as expressing the opinion of Horticulture Australia Ltd or any authority of the Australian Government. The Company and the Australian Government accept no responsibility for any of the opinions or the accuracy of the information contained in this report and readers should rely upon their own enquiries in making decisions concerning their own interests. ISBN 0 7341 0899 0 Published and distributed by: Horticultural Australia Ltd Level 1 50 Carrington Street Sydney NSW 2000 Telephone: (02) 8295 2300 Fax: (02) 8295 2399 E-Mail: [email protected] © Copyright 2004

FINAL REPORT FOR HORTICULTURE AUSTRALIA LIMITED

PROJECT NUMBER VGO2053

CAUSES OF LEAF SPOTTING IN CHINESE CABBAGE

JOHN BURT AND DENNIS PHILLIPS DEPARTMENT OF AGRICULTURE

WESTERN AUSTRALIA MAY 2004

May 2004 Causes of leaf spotting in Chinese cabbage - VGO2053

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CAUSES OF LEAF SPOTTING IN CHINESE CABBAGE The purpose of this report is to communicate the findings of Project VGO2053, which investigated the cause and control of leaf spotting, or ‘black dot’, which has caused problems with the marketing of Chinese cabbage grown in Western Australia for the past 20 years.

This report was funded by Horticulture Australia Limited and the Department of Agriculture Western Australia.

Project Leader: Dennis Phillips (Research and Development Officer, Department of Agriculture, Locked Bag No 4, Bentley Delivery Centre, Perth, Western Australia 6983.

Telephone: 08 9368 3319. E-mail: [email protected]

Principal Researcher: John Burt (Research and Development Officer, Department of Agriculture, Locked Bag No 4, Bentley Delivery Centre, Perth, Western Australia 6983).


Supporting Researcher: Dr Guijun Yan, School of Plant Biology, Faculty of Natural and Agricultural Sciences, The University of Western Australia, Crawley, Perth, Western Australia 6009. Telephone: 08 9380 1240. E-mail [email protected]

Chemists: Dr David Allen and Katrina Walton (Land Resources, Natural Resources, Chemistry Centre of WA, 125 Hay St East Perth, Western Australia 6004).


Technician: David Gatter (Department of Agriculture, Locked Bag No 4, Bentley Delivery Centre, Perth, Western Australia 6983).


ACKNOWLEDGMENTS The financial assistance of Horticulture Australia is gratefully acknowledged, without which this project would not have been possible.

The authors acknowledge the cooperation and assistance of Andrew and Mick Tedesco, Wanneroo, Galati and Sons, Mandogalup, Tom Mitchell, Tony Cosentino, West Gingin and Fernando Pessotto, Manjimup who assisted with this study. David Howey of Sumitomo Ltd gave useful information on the fungicide procymidone while Aileen Reid and Jo Blunn are thanked for their assistance with data preparation and processing.

DISCLAIMER Any recommendations contained in this publication do not necessarily represent current Horticulture Australia policy. No person should act on the basis of the contents of this publication, whether as to matters of fact or opinion or other content, without first obtaining specific independent professional advice in respect of the matters set out in this publication. The Chief Executive Officer of the Department of Agriculture and the State of Western Australia accept no liability whatsoever by reason of negligence or otherwise arising from the use or release of this information or any part of it.

© State of Western Australia, 2004.

HORTICULTURE AUSTRALIAPROJECT NUMBER:

VGO2053


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TABLE OF CONTENTS

Page

1. MEDIA SUMMARY .......................................................................................... 1

2. TECHNICAL SUMMARY .............................................................................. 1

3. INTRODUCTION ............................................................................................... 2

4. LITERATURE REVIEW ................................................................................. 3

5. RESEARCH WORK .......................................................................................... 4

5.1 Trial 1 - Field plants .................................................................................. 5

5.2 Observation 1A - Plants in pots .............................................................. 12

5.3 Observation 1B - Field plants .................................................................. 16

5.4 Observation 2 - Anatomical observations ............................................. 19

5.5 Observation 3 - Growers’ survey ............................................................ 21

5.6 Observation 4 - Plants in pots .................................................................. 22

5.7 Trial 2 - Field plants .................................................................................. 24

6. RESULTS AND DISCUSSION ...................................................................... 32

7. TECHNOLOGY TRANSFER ........................................................................ 32

8. RECOMMENDATIONS .................................................................................. 33

9. LITERATURE CITED ...................................................................................... 33

10. APPENDICES ....................................................................................................... 34

10.1 Research plans and soil amendments ...................................................... 34

10.2 Crop management and weather ................................................................ 40

10.3 Nutrient analysis ........................................................................................... 46


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1. MEDIA SUMMARY In 2003, the Department of Agriculture Western Australia investigated the cause and control of ‘black dot’ in Chinese cabbage (Brassica rapa L. subsp. pekinensis) with funding assistance from Horticulture Australia Limited.

‘Black dot’ had intermittently caused problems in Western Australian grown Chinese cabbage for the past twenty years. ‘Black dot’ is a condition in which thousands of small black dots (up to 1 mm diameter) are present on the leaves of Chinese cabbage heads (see Figure 2 - cover page). The green outside wrapper leaves on heads are affected, as well as up to the ten leaves further into the head. The condition is unsightly and is usually followed by leaf yellowing and marginal necrosis of older leaves. This prevents or reduces the marketability of Chinese cabbage on domestic and export markets. Tests conducted in previous years had not shown any pathogens to be associated with this condition.

The research project was successful in identifying the cause of the condition and was able to eliminate symptoms from growers’ crops by a simple change in disease management practices. Through a combination of planned research and careful observation, the research team proved that ‘black dot’ symptoms were caused by spraying fungicides containing the active ingredient ‘procymidone’. This is sold under the common trade names of Sumisclex® and Fortress®. Products containing procymidone are not registered for disease control in Chinese cabbage and growers are advised that crop damage will occur if they are used on this crop.

The project also investigated the response of heading Chinese cabbage to a range of soil pH treatments and trace element applications. The conclusion reached was that these factors did not contribute to the expression nor severity of ‘black dot’ in this crop.

The research proved the value of a systematic approach to problem solving backed by industry research funds, in determining the cause of a mystery that had plagued the industry for more than twenty years.

The findings from the work should eliminate crop losses from ‘black dot’ and ensure that Chinese cabbage can be produced more profitably for local and export markets in future.

2. TECHNICAL SUMMARY In 2003, the Department of Agriculture Western Australia investigated the cause and control of ‘black dot’ in Chinese cabbage (Brassica campestris pekinensis) with funding assistance from Horticulture Australia Limited.

‘Black dot’ had intermittently caused problems in Western Australian grown Chinese cabbage for the past twenty years. ‘Black dot’ is a condition in which thousands of small black dots (up to 1 mm diameter) are present on the outside leaves of Chinese cabbage heads. The green outside wrapper leaves on heads are affected as well as up to the ten leaves further into the head. The condition is unsightly and is usually followed by leaf yellowing and marginal necrosis of old leaves. This prevents or reduces the marketability of Chinese cabbage on domestic and export markets. Tests in previous years had not shown any pathogens to be associated with this condition.

Anatomical observations at cellular level conducted by the research team concluded that ‘black dot’ is probably a physiological disorder. The work demonstrated that black dots were small groups of dead cells associated with hairs on the leaf surface known as trichomes. These are widely recognised as excretory organs for toxins in the plant. The symptoms suggested that the dead cells may be the result of the plant's attempts to excrete toxic constituents. This was corroborated by results from later observations and trials.


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When the project started, field observations led the research team to believe that ‘black dot’ was caused by nutritional factors and was often related to high soil pH and recent liming. To test this hypothesis, a field trial was planted from seed on 17 January 2003 on a grower’s property in Wanneroo. It investigated the incidence of ‘black dot’ in plots to which lime or sulphur had been added to adjust the pH from 4.5 to 8.5 (measured in water). Various soil applied trace elements were also compared in the pH 7.5 treatment. ‘Black dot’ only occurred in the past week of the trial and was present on every plant. It was concluded that factors other than pH and plant nutrients were causing 'black dot'.

Observations on other plantings of Chinese cabbage at the trial site and information on spray history supplied by the grower led us to test two fungicides and one insecticide, to determine whether ‘black dot’ was due to a phytotoxicity from one or more of these pesticides. Initially this was done in unreplicated pot and field observations at Wanneroo and South Perth. The pot work used a typical nursery potting mix and the field plants were grown in soil on the trial property. It was found on both sites, only six days after the initial spraying, that the fungicide with the active ingredient procymidone (Sumisclex®) resulted in typical ‘black dot’ symptoms. The fungicide carbendazim (Howzat®), or the insecticide fipronil (Regent®) resulted in no ‘black dot’ symptoms.

The pot studies showed that ‘black dot’ symptoms also occurred similarly with two registered products that contained 50% w/v procymidone (Sumisclex® or Fortress®). The fungicides benomyl (Marvel®) and ipriodione (Rovral®) did not result in ‘black dot’ symptoms. It was also found that procymidone would result in symptoms of ‘black dot’, when used with the wetting agents Spraygard® or Agral®, or with no wetting agent.

A field trial to confirm these observations was transplanted on 5 September 2003 on a vegetable farm in Wanneroo. This showed that procymidone, when applied early, late or regularly in the crop, resulted in ‘black dot’. Treated heads of Chinese cabbage were unmarketable, whereas the untreated plants had no ‘black dot’ symptoms and all heads were marketable.

Initial observations on other brassicas, such as cabbage, broccoli and cauliflower showed that they were also affected by ‘black dot’ when sprayed with procymidone, but the effects on yields and quality were not determined.

The findings from this trial were communicated to a select group of growers in May 2003, who were collaborating with the project researchers. They immediately stopped using procymidone. Thereafter, ‘black dot’ has not been seen on their commercial plantings of Chinese cabbage for the past 12 months.

The findings from this work should enable Chinese cabbage to be produced more profitably for local and export markets.

3. INTRODUCTION ‘Black dot’ is a condition (Figure 2 - see cover photo) in which thousands of small black spots are present on the lamina (blade) of Chinese cabbage leaves. It often occurs on up to the ten oldest leaves on mature heads. Black dot symptoms are associated with yellowing of frame and head wrapper leaves of crops close to harvest. This is often obvious in a growing crop from a considerable distance. The symptoms prevent or reduce the marketability of Chinese cabbage on domestic and export markets.

The symptoms can be present throughout the year, but are often more damaging in summer. Chinese cabbage is now grown increasingly for domestic markets throughout the year. Anecdotal evidence at


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the start of the study was that it had become worse in the previous three years, but it has caused a problem for the past twenty years. It usually became obvious as early as three to four weeks after planting.

McKay et al. (1990) observed ‘black dot’ in winter harvested Chinese cabbage in the Perth area in the mid 1980s, but he conducted no work on this problem at that time. Dennis Phillips conducted a leaf analysis survey with a number of growers in the Perth region in 1989 to determine the cause of a petiole spotting problem known as ‘Gomasho’ and also noted symptoms consistent with ‘black dot’ in this survey. He found no relationship between nutrient levels and ‘black dot’, although there was a weak positive relationship between ‘black dot’ and high leaf levels of manganese. The sample size was small in this study and no definite conclusions could be drawn. In 2001 and 2002, Dennis Phillips conducted preliminary leaf analyses on plants affected by ‘black dot’. These showed a relationship between ‘black dot’ and low levels of iron and copper.

Based on observations on growers’ properties, ‘black dot’ appeared to be more common where soils had recently been limed. Liming is more common now than in the 1980s. It is widely used as a treatment to prevent infection caused by the soil-borne disease, clubroot.

A grower in Wanneroo who had experienced ‘black dot’ over a 20 year period had tried various treatments in small observations, but these had no effect. These included fenamiphos (Nemacur®), which controls nematodes and metham sodium (various brands), which is a general soil fumigant. He also trialled over 20 varieties in proprietary seed company trials and they all showed susceptibility to ‘black dot’. Plots were also compared with and without poultry manure. Superphosphate was applied to some beds and compared with a nil application. This grower volunteered his property for the conduct of field trials reported in this study.

In late 2002, ‘black dot’ was considered by members of the project team to be caused by nutritional factors, related to high soil pH and recent liming. The effect on ‘black dot’ of varying pH levels in main plots and added trace elements in sub-plots was therefore examined in this study as well as the effects of a proprietary trace element product. This work was conducted in the 2002/2003 summer.

Tests were also conducted to determine whether primary pathogens could be isolated. Numerous laboratory tests have failed to show any pathogen associated with this condition in the past.

The aim of the project was to determine the cause of ‘black dot’ and to find practical control measures. If successful, this would enable Chinese cabbage to be produced more economically for local and export markets.

4. LITERATURE REVIEW Information on Chinese cabbage is difficult to obtain from the non-English speaking countries that grow this crop, or it is not well reported in scientific journals. It would appear that, world-wide, research work on Chinese cabbage is limited. A comparison with work in other countries is confusing, because the same leaf spotting problem may have different names. There is reference to ‘black speck’ on head cabbage (a related Brassica species) in trials at Florida University (Strandberg, 1969) and Pennsylvania University (1981), and in Holland in 1997. This was reported to be a leaf spot condition of unknown cause.

A number of research papers were published by Japanese researchers in the 1970s and 1980s referring to a petiole spotting condition known in Japan as ‘Gomasho’ (Takahashi, 1981; Yoshida et al., 1984; Kato et al., 1977 and Matsumata, 1981). It was difficult to determine from translated abstracts if ‘Gomasho’ was similar to or the same as the condition we investigated in this study. However, these symptoms were reported as occurring on petioles (mid-ribs) of outer leaves and were described as


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being the size of a sesame seed (the literal translation of ‘Gomasho’). Work done by Phillips and Gersbach (1989) determined from cultivar responses and symptoms that ‘Gomasho’ occurred in Chinese cabbage grown in Western Australia. Dennis Phillips, one of the authors of the present study, concluded that ‘Gomasho’ has different symptoms to those under investigation in this study.

‘Black dot’ is the name unofficially given by the project team conducting this study for ease of communication and reporting. This problem is probably non-pathogenic and therefore does not have an official Latin name for comparative identification. It is therefore difficult to find information on this problem in other areas, including Australia.

There are no reports in the literature from the Eastern States of Australia of any problems that are similar to ‘black dot’. Bruce Tomkins of the Institute of Horticultural Development, Knoxfield, Victoria and Leigh James of Windsor, NSW have both worked on Chinese cabbage in the past and claim to have not seen ‘black dot’.

5. RESEARCH WORK The initial plan was to conduct a field trial to test the hypothesis that ‘black dot’ was associated with high pH soil, through its effect on minor element availability or uptake. The results of this work were to determine the direction that the project took for its duration. This was done and the results of the first trial and field observations led to the project taking different directions to those originally envisaged. The project was concluded after two trials and five observations had been completed as follows:

Table 1 Research activities conducted over the course of the project term

Research design Time Researchers Area Comments Main aims

Replicated trial 1 27 November 2002 to 25 March 2003

Burt et al. Vegetable property, Wanneroo

Field trial To determine if ‘black dot’ is related to nutritional factors and soil pH

Observation 1A 31 March 2003 to 16 June 2003

Burt et al. Dept of Agriculture, South Perth

Pot plants, in open

To determine if ‘black dot’ is caused by a fungicide phytotoxicity

Observation 1B 31 March 2003 to 10 June 2003


Field, similar to Observation 1A

To determine if ‘black dot’ is caused by a fungicide phytotoxicity

Observation 2 March to April 2003

Yan et al. UWA, Crawley, Perth

Laboratory observations of anatomy

To characterise the symptoms of ‘black dot’ at cellular level

Observation 3 February to April 2003

Burt et al. Growers in Baldivis, Gingin and Manjimup

Field-commercial production observations

To determine if ‘black dot’ is related to nutritional factors

Observation 4 29 May to 26 August 2003

Burt et al. Dept of Agriculture, South Perth

Pot plants, in open

To determine if ‘black dot’ is caused by a fungicide phytotoxicity other than procymidone

Replicated trial 2 5 September to 5 November 2003


Field trial To confirm that ‘black dot’ is caused by two common formulations of the fungicide procymidone.


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5.1 TRIAL 1-FIELD PLANTS

Aims (1) To determine, in a replicated field trial, if the incidence and severity of ‘black dot’ in Chinese

cabbage is influenced by soil pH.

(2) To assess, in an unreplicated observation, whether the application of iron, copper or manganese, before planting would reduce the incidence or severity of ‘black dot’, especially on an alkaline soil.

(3) To determine, in an unreplicated side observation, whether the regular applications of a proprietary trace element mixture (Librel BMX) after planting, would reduce the incidence and severity of ‘black dot’.

Site A commercial vegetable farm 22.5 km directly north from Perth CBD (Latitude 32°S 116°E) and 12 km east of the Indian Ocean coastline.

The site had been cropped with vegetables for the preceding 40 years. It had been cropped with lettuce and Chinese cabbage on a two year rotation, with only one crop per year, for at least the past 10 years. The area forms part of the Swan Coastal Plain and has a Karrakatta sand soil type.

The Karrakatta soil type is characterised by aeolian, siliceous deep sands, with a typical analysis of 81% coarse sand, 18% fine sand, 2% clay and a bulk density of 1.5.

In March 2002, the soil pH was 5.0 (measured in 1:5 soil:water) and the site was therefore treated by the grower with crushed limestone at 5 t/ha. By early December 2002, the soil pH had an alkaline reaction and ranged from 7.1 to 8.0 (measured in water) and 6.4 to 7.3 (measured in calcium chloride). The analyses also showed high levels of calcium (1,200 to 1,600 mg/kg), good levels of phosphorus (43 to 98 mg/kg), magnesium (51 to 75 mg/kg), manganese (13 to 17 mg/kg) and zinc (11 to 13 mg/kg), a medium level of iron (48 to 56 mg/kg) and copper (3.6 to 4.6 mg/kg), a low level of potassium (8 to 13 mg/kg), sulphur (5 to 6 mg/kg) and boron (0.1 mg/kg), and a desirable very low level of sodium (1 to 5 mg/kg), cobalt (0.06 mg/kg) and cadmium (0.04 mg/kg) . The organic carbon level ranged from 0.86 to 1.55% in four composite samples over the trial area.

Experimental design The trial was set out in a randomised block design with five soil pH treatments and four replications (see Table 2). The number of plots for statistical analysis was 20. The plot size of one treatment was double the size of the others to accommodate four observational sub-plots (see Table 2) to test the effect on ‘black dot’ of three soil applied trace elements against the untreated control. The untreated control was the only sub-plot to be included in the final statistical analysis.

An observation, outside of the area occupied by the replicated trial (see Appendix 11.1, Table A6) examined the effect on ‘black dot’ of a proprietary trace element spray (Librel BMX), at two rates, when applied three times, post-planting.


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Treatments On 14 November, 2002, two sites on the property were soil surveyed and analysed for pH by the Chemistry Centre of Western Australia. Twenty-four samples per plot were taken from the 0 to 25 cm depth to produce a composite sample. The soil was analysed for pH (water and CaCl2), electrical conductivity (EC), organic carbon, Total N, NH4-N and NO3–N, P and K. One of the two sites (designated as bay 14) was selected as the trial site because its pH was more uniform than the other.

The pH varied from plot to plot on the trial site and was adjusted by adding varying amounts of hydrated lime to raise the pH, or sulphur to lower the pH. The target (pH) treatments are shown in Table 2, while the trial layout, actual levels within the plots, rates of soil amendment application and calculations to determine rates are shown in Tables A1, A2 and A3 in Appendix A.

Table 2. Target pH treatments and sub-plots

Treatments pH treatment (measured in water)

Chemicals added to produce the desired pH Sub-plots

1 8.5 Various rates of hydrated lime. Nil

2* 7.5 Various rates of hydrated lime or dusting sulphur

A B C D

3 6.5 Various rates of dusting sulphur Nil



* Treatment 2 had double length plots (see Table 2). These were divided into four sub-plots, with the following randomised treatments applied before planting:

• A: No additional trace elements. • B: Copper sulphate at 50 kg/ha (43.4 g per sub-plot). • C: Manganese sulphate at 100 kg/ha (86.8 g per sub-plot). • D: Ferrous sulphate at 500 kg/ha (434.0 g per sub-plot).

On 17 December, 2002, we applied hydrated lime or sulphur to produce the five pH treatments, and applied trace elements to four sub-plots in Treatment 2. The rates were based on the pH of each plot on 14 November and a formula supplied by Dr D. Allen of the Chemistry Centre of WA (see Appendix Table A.2). The formula used the pH, buffering capacity (1.8 me%), bulk density of soil (1.5 g/mL), depth of soil (25 cm), per cent purity (% calcium carbonate by weight) and equivalent weigh (50 g CaCO3/mol of H+) to estimate the rates. We also considered the previous experience by Mr A. McKay (Department of Agriculture, Western Australia) on soils of the Swan Coastal Plain.

The lime or sulphur applications were immediately incorporated with a rotary hoe. For the 30 days before planting, the soil was watered three times per day to activate the chemical treatments to produce the desired pH levels. In the same period, weeds were controlled with a rotary hoe, in the reverse direction to the previous cultivation.

Composite samples of soil were taken on 3 January 2003 at the 0 to 20 cm depth and sampled for pH in Treatments 1 and 5 to determine whether there had been the desired change in soil pH 17 days after application of lime and sulphur. The analysis showed that the desired pH changes were occurring (Appendix, Table A.4). We therefore felt confident that we could obtain the desired pH treatments and that the trial could be continued.


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Further pH samplings were done 1, 26, 51 days after planting, and at the end of the trial (Appendix Table A.5). The differences in pH were consistent between treatments and a good range of pH was obtained to test the effects of pH and associated nutritional effects on ‘black dot’. As expected, it was not possible to obtain the desired high alkaline treatment of pH 8.5, but we obtained levels that were only slightly less than this. The desired pH of 7.5 was obtained with Treatment 2. We supplied too much sulphur with all the acidic treatments and this was most evident in Treatment 3 where the desired pH was 6.5. The pH remained fairly stable in the various treatments from planting to harvesting.

Trial layout The total length of the trial was 40.5 m in one bay, 6.73 m wide. The lime and sulphur soil amendments were applied to plots 2.25 m wide by 6.73 m long. The central 1.6 m of each of these plots was planted with four rows of Chinese cabbage seed in rows 40 cm apart. For Treatments 1 and 3 to 5, this gave 68 plants/plot with 24 for harvest and 20 for mid life plant sampling buffered by 2 plants/row between plots. Treatment 2 had a plot size of 2.25 m wide by 13.46 m long, comprising four sub plots each 3.36 m long, allowing 24 plants for final harvest and assessment in each sub plot with two plants/row buffers around each sub plot. Within the rows, plants were 36 cm apart.

Side observation Librel BMX was applied 26, 42 and 54 days after planting to an area of 10.0 by 6.75 m at the southern side of the trial (see Appendix Table A.6), with a buffer of 5.0 m between the trial and the observation. Librel BMX contains the six main trace elements-boron (0.875%), copper (1.7%), iron (3.35%), manganese (1.7%), molybdenum (0.02%) and zinc (0.6%). Librel BMX at 0.5 kg per hectare was applied to an area of 50 m2 at two, four and six weeks after planting, using a knapsack sprayer, taking care not to spray during a southerly wind. Based on grower’s experience, this was a low rate to prevent burning of the leaves. The rate on the label of 1.0 kg/ha was also applied to an area of 10 m2 to see if burning would occur with this rate.

Planting and establishment The trial area was rotary hoed before planting. Chinese cabbage seeds were vacuum planted with three seeds per site on 16 January 2003. The trial bay was situated within a commercial crop of Chinese cabbage with older and younger plantings in bays on either side.

Crop management For details of crop management, see Appendix 10.2.


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Figure 3. Trial plants growing well six weeks after sowing.

Harvesting and data collection Extensive plant tissue analysis of leaves and petioles from all treatments was conducted at the mid growth stage (27 February) and at harvest, (25 March) to assist in identifying trends and nutrient relationships which may have influenced 'black dot' incidence and severity. At each date, the elements N, P, K, Na, Ca, Mg, S, B, Cu, Fe, Mn and Zn were measured in head wrapper blades and petioles as well as blades and petioles of 10 leaves into the head where the head wrapper was nominated as leaf 1. A composite sample of 20 leaves or petioles was used for most of these tests.

Plants were harvested on 25 March, 2003, 68 days after planting. A total of 24 plants in the middle of each plot were sampled. The heads were individually weighed and graded for quality and marketable or reject weights. Each plant was rated for the incidence and severity of ‘black dot’ on a scale of 1-5, where 1=absent; 5=severe. The plant parts chosen for 'black dot' rating by this method were outer head wrapper leaves, and leaves 5 and 10 into the head, where the outer head wrapper was designated as leaf 1.

Results Water from the two bores used to irrigate the trial was collected on 17 February and analysed by the Chemistry Centre of Western Australia. Results of this test are presented in Appendix B, Table B.1. Trace elements were not at a toxic level and those that were essential elements would have been supplied in useful amounts for the nutrient needs of the plants.

Some growth measurements and observations of the treatments were made while they were growing and these are presented in Appendix Tables A7 to A9.


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The symptoms observed in the trial plots included: (1) Small ‘black dots’ on the leaves (see Figure 4) with up to 2000 spots on each leaf. (2) Necrotic marginal leaf burning at the lowest pH. (3) Yellowing of the outer leaves in the three weeks before harvest.

Figure 4. Leaf affected by ‘black dot’.

All plots were harvested on 25 March and 10 heads from each plot were scored for presence and severity of blade and petiole spotting separately on head wrapper and internal leaves. ‘Black dot’ was widespread and severe on head wrapper blades of all treatments, while petiole spots were also common. The two types of spots did not always occur together in the same treatment or plant. Most, if not all petiole spotting was considered to be the condition widely referred to in the literature as ‘Gomasho’ (see Figure 5). ‘Black dot’ did not appear in the trial until a few days before harvest, much later than we expected.

Figure 5. Typical leaf affected by petiole spotting or ‘Gomasho’.


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At harvest, there were no ‘black dot’ symptoms from 12 leaves in towards the centre of the head. The occurrence of 'black dot' and ‘Gomasho’ symptoms at the level of the tenth leaf into the head were too inconsistent to justify statistical analysis, but analyses for the outer head wrapper and the fifth leaf were significant.

The severity of ‘black dot’ became less (lower score) with the progression from outer wrapper leaves towards five leaves into the head, as shown by the decreasing scores in Table 3. This shows that 'black dot' was significantly (p < 0.001) worse (higher score) in the pH 7.5 treatment than at all other pH levels for the outer head wrapper. The fifth leaf into the head showed that the pH 7.5 treatment had significantly (p < 0.05) higher 'black dot' symptoms than the pH treatments, 4.5, 5.5 and 6.5.

Table 3. 'Black dot' severity score for head wrapper and fifth leaf into the head as influenced by soil pH

Plant part Soil pH

4.5 5.5 6.5 7.5 8.5 LSD

Head wrapper 2.75 3.05 2.9 3.68 3.1 0.3739 (p < 0.001)

5th leaf 1.475 1.65 1.625 1.95 1.8 0.2840 (p < 0.05)

It was noticed that the Librel BMX treatment of 1.0 kg/ha had slight burning compared with the treatment at 0.5 kg/ha.

The sub plots within the pH 7.5 treatment produced some significant differences between soil treatments with minor elements and their effect on ‘black dot' and ‘Gomasho’ severity. Application of manganese to the soil prior to planting at 100 kg/hectare gave relatively lower ‘black dot’ severity in head wrapper leaves at harvest compared to the other treatments. However, the differences were small and all heads had levels of ‘black dot' which rendered them unmarketable. This difference could not be detected in the fifth leaf into the head (Table 4).

Similarly, small, but significant differences in ‘Gomasho’ levels were detected in the fifth leaf, with greater severity where iron and copper had been applied. However, this difference was not reproduced in head wrapper leaves (Table 5).

Table 4. Effect of soil applied minor elements on ‘black dot' severity scores (1-5 where 1=absent; 5=severe) on head wrapper and fifth leaves into the head

Plant part Copper Iron Manganese Untreated LSD (p < 0.05)

Head wrapper 3.5 3.725 3.1 3.675 0.4133

5th leaf 1.825 1.875 1.675 1.95 Not significant

Table 5. Effect of soil applied minor elements on 'Gomasho' severity scores (1-5 where 1=absent; 5=severe) on head wrapper and fifth leaves into the head

Plant part Copper Iron Manganese Untreated LSD (p < 0.05)

Head wrapper 3.1 3.1 2.575 2.85 Not significant

5th leaf 1.95 2.2 1.75 1.725 0.3451


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Nutrient analysis of leaf blades and petioles The results of nutrient tests on all plant samples from all treatments are presented in Appendix C Tables C1-C12. The data show many interesting relationships between soil pH and nutrient uptake in Chinese cabbage and for this reason is worthy of reporting. REML analysis was used for the data from the February sampling and the standard ANOVA technique for the April data, because variances were not found to be uniformly distributed for the February data. This aspect of the work may be published in full separately to this report because the results did not directly contribute to elucidating the causes of ‘black dot’.

Diseases Typical ‘black dot’ symptoms were checked by plant pathologist Peter Wood at the Department of Agriculture, Western Australia. He reported that “The bacteriological tests on the lesions of the above sample have now been finalised”. Results indicate that the isolates are neither Xanthomonas sp. (Black Rot) nor Pseudomonas sp. (Bacterial Leaf Spot). As there was also no microscopic evidence of fungal mycelium associated with the lesions I am fairly certain that the symptoms are not the result of infection by a plant pathogenic organism’.

At harvest, plants with typical ‘black dot’ symptoms were submitted for plant pathology analyses by the Department of Agriculture. There were no fungal pathogens isolated from the leaf spots. Alternaria alternata was present as a secondary organism.

Other plantings The grower whose property hosted the pH trial used Sumisclex® once only (on 14 April) on his other plantings. By the end of April, we informed him of our findings with the first spraying in observations 1A and 1B. He did not use Sumisclex® after this time. Observations of all the plantings made before mid April were affected by ‘black dot’. Plantings made from mid April 2003 to May 2004 were not sprayed with Sumisclex® and were not affected by ‘black dot’.

Conclusions ‘Black dot’ was not seen on any of the commercial weekly plantings of the grower from December 2002 to mid March 2003, or on the trial planting to just before harvesting at the end of March 2003. After 20 March, it then appeared severely on all plantings and in the trial. It was not seen on one bay where a proprietary chemical company was conducting a spraying trial.

This bay was not sprayed with the pesticides used by the grower on his commercial plantings, or on the Department of Agriculture trial. The grower stated that for the first time in five months, he had just started to use fungicides to control general diseases. Fungicides were not used on the bay leased by the proprietary firm.

In Figure 6 the area to the right of the picture was the spray trial operated by a proprietary firm and was not sprayed by the grower. The area on the left was sprayed by the grower with a fungicide and is heavily damaged by ‘black dot’.


12

Figure 6. The ‘black dot’ plants on the left of the picture were sprayed with procymidone. The green

plants in the proprietary spraying trial in the right bay were not sprayed with procymidone and have no ‘black dots’.

The evidence pointed to damage from pesticides, especially the fungicides used by the grower in recent weeks. These included Sumisclex®(procymidone) and Howzat® (carbendazim). It was decided to immediately conduct observations in pots (Observation 1A) and in the field (Observation 1B) and at South Perth to determine whether the spraying of these pesticides would induce ‘black dot’.

5.2 OBSERVATION 1A-PLANTS GROWN IN POTS On 8 April 2003, two weeks after the results were obtained from the first trial, a pot observation was started with the variety Kasumi at the Department of Agriculture, South Perth. Surplus seedlings at the two-leaf stage were transplanted from a commercial planting, sown on 30 March, from the grower in Wanneroo.

Aims • To determine whether the fungicides procymidone (Sumisclex®) or carbendazim (Howzat®), or

the insecticide fipronil (Regent®) would result in ‘black dot’ symptoms when sprayed on Chinese cabbage.

• To determine whether the application of a fast working soil conditioner (Tipa Gold®) to pots would result in symptoms of ‘black dot’ on Chinese cabbage.


13

In mid March, and before we had obtained the results from the first trial, we were still thinking that there was a link between ‘black dot’ and pH. We decided to conduct a small observation after the first trial, with Tipa Gold®. This is believed to contain potassium carbonate and is applied by some growers through their trickle irrigation systems to quickly raise the pH in acidic soils.

Treatments There were six plants in each treatment (see Figure 7). Plants were initially placed into 10 cm pots. These established successfully and were potted up after three weeks into 20 cm pots. Sand from the Wanneroo farm was used as the potting mixture and there were no drainage problems in the pots with this media. The pH was 6.6 (measured in water).

Figure 7. Pots two weeks after planting and before spraying.

The plants were placed on slats in the open, in an area that was wind-protected, but not shaded from buildings.

Sumisclex® was applied both at the standard off-label permit rate for cabbage and at double the off-label permit rate. Howzat® was applied at the rate used by the grower, and at double the rate used by the grower. Regent® was used at the standard registered rate for brassicas and at double the registered rate. All pesticides were applied with Spraygard® (334 g/L di-1-p-menthene) which was the wetting agent used by the grower in the first trial. The pesticide treatments (see Table 6) were first applied with a hand-sprayer as foliar sprays 16 days after planting on 24 April, and repeated on 28 April, 2 May, 9 May and 20 May.

The Tipa Gold® treatment was applied to the sand in the pots soon after potting-up into 20 cm pots. This is normally applied in the field at the rate of 3.5 mL per metre of row. This was considered to be equivalent to 1 mL per pot. This was compared with 2 mL per pot and a very high rate of 5 mL per pot. These were watered in with 200 mL of water per pot. There were two plants with each rate of Tipa Gold®.


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Table 6. Treatments in pot plant observation 1A at South Perth, planted on 8/04/03

Treatment Wetting agent Insecticide Fungicides Soil conditioner

1 Nil Nil Nil Nil

2 Spraygard (334 g/L di-1-p-menthene) 1 mL/2L

Regent (200 g/L fiprinol) 0.5 mL/2L

Nil Nil

3 Spraygard 1 mL/2L


Howzat (500 g/L carbendazim) 2 mL/2L

Nil

4 Spraygard 1 mL/2L


Sumisclex (500 g/L procymidone) 2 mL/2L

Nil

5 Spraygard 2 mL/2L

Regent (200 g/L fiprinol) 1 mL/2L

Nil Nil

6 Spraygard 2 mL/2L


Howzat (500 g/L carbendazim) 4 mL/2L

Nil

7 Spraygard 2 mL/2L


Sumisclex (500 g/L procymidone) 4 mL/2L

Nil

8 Nil Nil Nil Tipa Gold 1, 2 or 5 mL per pot

The plants were watered daily and fertilised weekly with Phostrogen Improved®. This contains the following nutrients: 14.0% nitrogen, 4.4% phosphorus, 22.5% potassium, 1.3% magnesium, 0.75% calcium, plus sulphur, boron, copper, iron, manganese, molybdenum and zinc. It was applied initially at half-strength.

Results The plants were first noted with ‘black dot’ symptoms on 30 April 2003, six days after spraying. The only plants to be affected were all the plants in the two Sumisclex® treatments. With continued spraying, these plants were noticeably stunted and yellow. Close examination showed thousands of small ‘black dots’ on each leaf. The high rate of Sumisclex® damaged the plants more than the off-label permit rate (on cabbage) for Sumisclex®. The other pesticide treatments were not affected by ‘black dot’.

Plants were harvested on 16 June, 69 days after planting (see Table 7). The 20 cm pot was probably not an optimum size, and the average head weights were below average for field grown Chinese cabbage.


15

Table 7. Harvest head weights (kg) on 16 June 2003

Treatments showing head weights (kg) for individual plants

Plants Control

Spraygard +

Regent

(low rate)

Spraygard+

Regent +

Howzat (low rate)

Spraygard+

Regent +

Sumisclex (low rate)

Spraygard+

Regent +

(high rate)

Spraygard +

Regent +

Howzat (high rate)

Spraygard +

Regent +

Sumisclex (high rate)

Tipa Gold

1 0.98 0.99 0.96 1.11 0.90 0.72 0.39 0.71 (low rate)

2 0.63 0.87 0.75 0.53 0.68 0.92 0.40 0.52 (low rate)

3 0.59 0.87 0.99 0.38 0.93 1.02 0.22 0.37 (medium rate)

4 0.70 0.77 0.79 0.61 0.93 1.06 0.36 0.75 (medium rate)

5 0.58 0.90 0.81 0.63 0.93 1.09 0.20 0.49 (high rate)

6 0.69 0.90 1.05 0.78 1.09 0.82 0.44 0.37 (high rate)

‘Black dot’ Nil Severe Nil Nil Nil Nil Very severe Nil

Marketability Unmarket-

able - too small

Marketable Marketable Unmarket-

able - ‘black dot’

Marketable Marketable Unmarket-

able - ‘black dot’

Unmarket-able -

too small

Total kg 4.17 5.30 5.35 4.04 5.46 5.63 2.01 3.21

Average head weight (kg) 0.70 0.88 0.89 0.67 0.91 0.94 0.34 0.54

The only plants that were affected by ‘black dot’ were the Sumisclex® treatments and all plants were unmarketable. Plants sprayed with the high rate of Sumisclex® were severely stunted and the reject average head weight was half that of the lower rate. All the plants in the Regent® and Howzat®) treatments were marketable. The nil spraying treatment and the Tipa Gold treatment were unmarketable, due to small head size.

As there was no ‘black dot’ in the Tipa Gold treatment, and due to the small number of pots with the three sub-treatments, it was not considered necessary to measure the pH of the soil after harvesting.

Two plants, out of the total of six plants, in each Sumisclex® treatment were only sprayed for the first three applications. It was decided to see if these plants would recover if they were not sprayed after 2 May. They were therefore not sprayed on the 9 and 20 May. However, they were still unmarketable due to ‘black dot’ when harvested.

Conclusions Severe ‘black dot’ symptoms were induced a few days after spraying with procymidone (Sumisclex®) and this caused yellowing and stunting, especially with the high rate of Sumisclex®. There were no ‘black dot’ symptoms after spraying with carbendazim (Howzat®) or fipronil (Regent®).

The use of a fast acting soil conditioner (Tipa Gold®) at ‘normal’, high and very high rates did not induce ‘black dot’ symptoms.


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5.3 OBSERVATION 1B - FIELD PLANTS On 9 April 2003, two weeks after the results were obtained from the first trial, a field observation was commenced with the variety Kasumi at the vegetable farm in Wanneroo, where Trial 1 had been conducted. The aims of the observation were similar to Observation 1A at South Perth and were as follows, with the main aim of investigating the effect of two fungicides on ‘black dot’.

Aims 1. To determine whether the fungicides procymidone (Sumisclex®) or carbendazim (Howzat®), or

the insecticide fipronil (Regent®) would result in ‘black dot’ symptoms when sprayed on Chinese cabbage.

2. To determine whether the application of a fast working soil conditioner (Tipa Gold) to soil would result in symptoms of ‘black dot’.

3 To determine whether the wetting agent Spraygard®) can induce symptoms of ‘black dot’, when applied alone at normal, high and very high rates.

4 To determine whether high rates of manganese sulphate will induce symptoms of ‘black dot’.

The reason for testing Tipa Gold was detailed in the report for Observation 1A. Spraygard® was used by the grower as the standard wetting agent at low rates for all spray applications, including Trial 1. At high rates, Spraygard® can act as an anti-transpirant, so we decided to determine whether it has a phytotoxic effect on Chinese cabbage at high rates. We also included high rates of manganese as a soil drench, as toxic levels are reported to produce spotting on the leaves.

There were 24 plants in each treatment of the pesticide plots, eight plants per treatment with the Tipa Gold plots, four plants per treatment with the manganese plots and six plants per treatment with the Spraygard plots. The pH was 6.5 (measured in water).

The pesticide treatments were the same as with the pot plant observation, except that a nil spraying treatment was not used, except in the side buffers. Sumisclex® was applied both at the standard off-label permit rate (for cabbage) and at double the off-label permit rate. Howzat® was applied at the rate used by the grower and at double the rate used by the grower. Regent® was used at the standard registered rate for brassicas and at double the registered rate. All pesticides were applied with a wetting agent (Spraygard®). The pesticide treatments (see Table 9) were first applied as foliar sprays, with a small hand-sprayer. The first pesticide spray was applied on 23 April, 24 days after sowing. Further sprays were made on 30 April, 5 May, 21 May and 28 May.

The Spraygard® treatments were applied at the same time as the pesticide treatments. The Tipa Gold® treatment and manganese sulphate treatments were applied around the plants as a soil drench, 18 days after sowing.

The plants were watered daily and fertilised weekly with the growers fertilising program, which was similar to Trial 1. The grower did not spray any pesticides onto the observation. There were no obvious pests and diseases.

Results The plants were first noted with ‘black dot’ symptoms on 30 April 2003, seven days after spraying. The only plants affected were all the plants in the two Sumisclex® treatments. With continued spraying, these plants were noticeably stunted and yellow. Close examination showed thousands of small ‘black dots’ on each leaf. The high rate of Sumisclex® damaged the plants more than the off-label permit rate for Sumisclex® for use on cabbage. The other pesticide treatments were not affected by ‘black dot’.


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Plants were harvested on 11 June 2003, 73 days after planting. There was no ‘black dot’ or ‘Gomasho’ with the treatments Tipa Gold®, manganese sulphate or Spraygard®. As there was no ‘black dot’ in these treatments, and due to the small number of plants with the three sub-treatments, these plants were not weighed at harvesting, but comments on the plants at harvesting are shown in Table 8.

Table 8. Condition of plants at harvesting of the non-pesticide treatments

Plot Treatment Rate Appearance at harvest

A Tipa Gold 8 mL/2L Good yields and quality

B Tipa Gold 40 mL/2L Good yields and quality

C Tipa Gold 80 mL/2L Some black flecks on internal petioles (not ‘black dot’).

D Manganese sulphate 2.5 g/m2 Good yields and quality

E Manganese sulphate 10 g/m2 Good yields and quality

F Manganese sulphate 50 g/m2 Stunted, pale, some leaf spotting (not ‘black dot’)

G Spraygard 1 mLl/2L Good yields and quality

H Spraygard 10 mL/2L Good yields and quality

I Spraygard 50 mL/2L Good yields and quality

With the pesticide treatments, the central 10 plants were harvested per plot and assessed for ‘black dot’ and other problems. The only plants that were affected by ‘black dot’ were the Sumisclex® treatments and all plants were unmarketable (see Table 9). These treatments also had lower total head weight than the other pesticide treatments. The plants in the Regent® and Howzat® treatments were marketable and had good head weights. Plants were much bigger than those harvested from the pot plant observation. Stunting of the plants sprayed with the high rate of Sumisclex®, compared with the lower rate of Sumisclex®, was also less pronounced in the field observation. There was no ‘Gomasho’.

Table 9. Total and marketable weights (kg) for the pesticide treatments (ten heads per plot)

Treatments showing individual head weights (kg)

Plant

Spraygard 1 mL/2L +

Regent 0.5 mL/2L

Spraygard 1 mL/2L

+ Regent

0.5 mL/2L +

Howzat 2 mL/2 L

Spraygard 1 mL/2L

+ Regent

0.5 mL/2L +

Howzat 2 mL/2 L

+ Sumisclex 2 mL/2 L)

Spraygard 1 mL/2L +

Regent 1 mL/2L)

Spraygard 2 mL/2L

+ Regent

1 mL/2L +

Howzat 4 mL/2 L

Spraygard 1 mL/2L

+ Regent

1 mL/2L +

Sumisclex 4 mL/2 L)

1 2.00 1.76 1.47 2.01 1.64 1.54

2 1.37 1.81 1.82 1.42 1.93 1.43

3 2.17 1.83 1.52 2.11 1.86 1.41

4 1.32 1.77 1.56 1.72 1.92 1.68

5 2.03 1.70 1.67 2.20 2.10 1.50

6 1.74 1.85 1.81 1.82 2.00 1.95


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Table 9. Continued

Treatments showing individual head weights (kg)

Plant

Spraygard 1 mL/2L +

Regent 0.5 mL/2L

Spraygard 1 mL/2L

+ Regent

0.5 mL/2L +

Howzat 2 mL/2 L

Spraygard 1 mL/2L

+ Regent

0.5 mL/2L +

Howzat 2 mL/2 L

+ Sumisclex 2 mL/2 L)

Spraygard 1 mL/2L +

Regent 1 mL/2L)

Spraygard 2 mL/2L

+ Regent

1 mL/2L +

Howzat 4 mL/2 L

Spraygard 1 mL/2L

+ Regent

1 mL/2L +

Sumisclex 4 mL/2 L)

7 2.22 2.19 1.60 1.67 1.71 1.49

8 3.00 2.02 1.55 1.34 2.29 1.71

9 2.14 2.30 0.90 1.78 2.36 1.47

10 3.00 2.01 2.13 2.26 1.97 1.32

Total weight per plot (kg) 19.1 17.5 14.6 16.7 18.0 14.1

Average weight per plot (kg) 1.91 1.75 1.46 1.67 1.8 1.41

Total marketable weight (kg)

19.1 17.5 0 16.7 18.0 0

Conclusions Severe ‘black dot’ symptoms were induced a few days after spraying with procymidone (Sumisclex®) and this caused yellowing and stunting. There were no ‘black dot’ symptoms after spraying with carbendazim (Howzat®) or fipronil(Regent®). At harvesting and after five separate sprays, plants in the Howzat®) and (Regent®) plots produced good marketable yields, whereas the plants in the Sumisclex® plots were not marketable, due to severe damage from ‘black dot’.

The use of a fast acting soil conditioner (Tipa Gold®), manganese sulphate or Spraygard® did not induce ‘black dot’ symptoms, especially when these were applied at high rates. However, there were other phytotoxic effects with very high rates of Tipa Gold® and manganese sulphate.

The grower whose property hosted the pH trial used Sumisclex® once only (on 14 April) on his other plantings. In late April, we informed him of our findings with the first spraying in observations 1A and 1B. He did not use Sumisclex® after this time. Observations of all the plantings made before mid April were affected by ‘black dot’. Plantings made from late April 2003 to May 2004 were not sprayed with Sumisclex® and were not affected by ‘black dot’.

Recommendation

It was decided that the next step in this investigation was to test other brands of fungicide containing the active ingredient procymidone in observation plots (see 5.6, Observation 4) .


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5.4 OBSERVATION 2 - ANATOMICAL OBSERVATIONS Principal research worker: Dr Guijun Yan, University of Western Australia

Introduction In parallel with this work, a small contract was let to collaborators at the University of WA to conduct microscopy studies on the symptoms and prepare a report including photographs of the symptoms at cellular level. This research was aimed at characterising the symptoms of ‘black dot’ under a microscope and to develop methodologies for its control.

General description The symptoms of ‘black dot’ appear as small black circular spots mostly occurring on the undersides of exposed older leaves. As the leaves grow older, the lesions penetrate both surfaces. A few lesions are also visible on the upper leaf surface. The peppery spots are normally less than 1 mm in diameter. They are distributed in large areas of the outer layers of leaves, while none of them can be seen on the young inner leaves. When affected leaves were cut into pieces and cultured in Petri dishes in a constant temperature room for more than one week, the lesions showed no further development and no fungal spores were produced. Therefore, ‘black dot’ is unlikely to be infectious, as no fungus or bacterium symptoms were observed. There were no signs of specific patterns of insect travel or insect larvae or eggs. ‘Black dot’ is different from ‘Gomasho’ as it mainly happens on leaf blades whereas ‘Gomasho’ occurs on the leaf petioles.

Detailed description

Older leaves (completely exposed) The spots mainly occurred on leaf blades and some of the dots have yellowish halos in the centre. Using a dissection microscope, the edges of the lesions were clear and the centres were sunken and dry. It could be seen clearly that most of the dots reflected light under a light source and therefore it is concluded that only the cells inside the epidermis were damaged and most of the epidermis cells are left intact. Tearing off the epidermis with a lesion also confirmed that the epidermis was not damaged.

Hand sectioning and observation under a light microscope indicated that the colour of the affected palisade and sponge mesophyll cells were dark brown with only the cell walls left and no contents.

On veins, the lesions were rare, they appeared to be strongly related to surface hairs (trichomes). A total of 84% of lesions were located at the base of the hair on the veins. Affected hairs became hollow (without cell content) and eventually detached from the blade.

Younger leaves (half exposed) Most the lesions occurred on the veins and 92% of them were associated with hairs. They were right at the base of the hairs. Affected hairs lost their cell content and eventually detached from the blade.

Conclusions According to anatomical observations, ‘black dot’ on Chinese cabbage is more likely to be a physiological disorder (i.e. an enzymatic browning type reaction was induced from the plant). The exposed outer leaves were often seriously affected by the spotting, whereas the inner leaves were less affected. The lesions were not associated with the position of the stomata.


20

This work demonstrated that ‘black dots’ were small groups of dead cells on the lamina, or were associated with hairs on the leaf veins known as trichomes. Trichomes are widely recognised as excretory organs for toxins in the plant. The symptoms suggested that the dead cells may be the result of the plants attempts to excrete toxic constituents or breakdown products of the fungicide

Table 10. The occurrence of ‘black dot’ on the leaves of Chinese cabbage and its association with leaf surface hairs on the veins

Total dots

(a)

Dots onvein

(b)

Related tohairs on the veins

(c)

Proportion of dots

on vein:

b/a (%)

Proportion of dots

associated with hairs on

veins: c/b (%)

Old leaves Leaf 1 512 53 49 10.35 92.45

Leaf 2 887 44 33 4.96 75.00

Average 7.65 83.73

Younger leaves Leaf 1 78 78 68 100.00 87.18

Leaf 2 45 45 41 100.00 98.11

Average 53 53 52 100.00 92.13

Dots were recorded from about 20 mm along the main vein under a dissecting microscope.

Figure 8. Distribution of ‘black dot’ lesions

on the under-surface of older leaves. Note they are mainly on the leaf blade (x 6.5).

Figure 9. Distribution of ‘black dot’ lesions on the under-surface of younger leaves. Note they are mainly on the veins (x 6.5).

Figure 10. A lesion on an of older leaf with a

yellowish dry halo (x 50). Figure 11. A lesion on a younger leaf associated

with a hair (x 50).


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Figure 12. The lesion under epidermis cells.

Note the damaged cells have no content and only walls left (x 200).

Figure 13. The damaged cells curved in and were easily broken (x 100).

Figure 14. The lesion on torn-off epidermis.

Note the cell wall for the cells in the centre of the lesion broke down (x 1560).

Figure 15. The lesion under stomata (arrowed) (x 600).

5.5 OBSERVATION 3 - GROWERS’ SURVEY

Aims • To survey crops of three Chinese cabbage farms, to monitor ‘black dot’ in relation to their

management.

• To survey the leaves of a number of growers Chinese cabbage crops to determine whether ‘black dot’ is affected by one or more nutrient deficiencies or toxicities.

The properties were located at:

• Mandogalup, 35 km south of Perth.

• Gingin, 100 km north of Perth.

• Manjimup, 300 km south of Perth.

These growers planted from February and March 2003 onwards for export. Chinese cabbage was also observed on other growers’ properties.


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Results The survey on growers’ farms from February to April 2003 showed that ‘black dot’ was found in all cases where growers had recently used procymidone. ‘Black dot’ was not observed in Manjimup, where procymidone was not used. In early May, we were confident that ‘black dot’ was due to a phytotoxicity caused by procymidone. It was suggested that the two growers in our survey program should cease using this fungicide. This resulted in no evidence of ‘black dot’ symptoms on their new plantings from May 2003 to May 2004.

The original intention of doing intensive leaf analyses on a few growers properties was discontinued when it appeared that ‘black dot’ was due to a phytotoxicity. Our efforts were instead directed to obtaining conclusive proof that fungicides containing procymidone caused the symptoms by conducting confirmatory experiments with pot plants and in the field.

Damage from ‘black dot’, following an early application of procymidone, was also noticed in a commercial planting of cabbages at Mandogalup.

5.6 OBSERVATION 4 - PLANTS IN POTS The second pot observation was planted with the variety Yuki at South Perth on 29 May 2003.

Aims 1. To determine whether another proprietary product containing procymidone (Fortress®) would

result in ‘black dot’ symptoms similar to Sumisclex® (Observations 1A and 1B) when sprayed on Chinese cabbage (see G in Table 11). There are other formulations of procymidone available in Western Australia, but Fortress® and Sumisclex® are the only pesticides with an off-label permit for cabbages.

2. To determine whether ‘black dot’ symptoms would be obtained with procymidone (Sumisclex®) when applied with either of two wetting agents (Agral® or Spraygard® ), or with no wetting agent (see J, K, L in Table 11). Agral® has been one of the main wetting agents used by growers in Western Australia on vegetables for the past 20 years. Spraygard® was the wetting agent used by the grower when spraying pesticides in Trial 1 at Wanneroo

3. To determine the effects of spraying Sumisclex® on ‘black dot’ symptoms, as a single spray, applied early, mid-season or late, compared with five sprays throughout the growing period (see L, M, N, O, P in Table 11) and a nil treatment (see I in Table 11).

4 To determine whether other fungicides that could be sprayed on Chinese cabbage such as benomyl (Marvel®) and ipriodione(Rovral®) would result in ‘black dot’ symptoms (see F and H in Table 11).

5. To determine whether procymidone (Sumisclex®) will result in ‘black dot’ symptoms with some other brassicas, such as swedes, canola and cabbage (see A, B, C, D, E in Table 11).

Yuki is a well known export variety of Chinese cabbage and is especially popular in Victoria and grown by some growers in Western Australia. It is suited for production in cool conditions. A grower in Mandogalup who has often produced this variety in the past, stated that it is very susceptible to ‘black dot’. The variety of swedes was Champion Purple Top. The cabbage variety was Green Gold.

Seed was sown into 20 cm pots filled with a proprietary potting mixture at the Department of Agriculture, South Perth. The spraying times and chemical rates are shown in Table 11. The plants were watered daily and fertilised regularly with Thrive®. It contains the following nutrients: nitrogen 30.0%, phosphorus 4.5%, potassium 8.0%, plus boron, copper, iron, manganese, molybdenum and zinc.


23

The plants were assessed for ‘black dot’ symptoms on 26 August 2003 (see Table 11). Due to the small number of plants, it was decided that yields would not be recorded.

Table 11. ‘Black dot’ assessment on 26/08/03 after spraying various plants with various pesticides

Item Plants Number of plants Chemical Active

Ingredient Sprays Spray dates

Assessment of ‘black dot’ - including spots, marginal necrosis

and yellowing

A Cabbage 5 Sumisclex 1.0 mL/L +

Agral at 3 drops per L

Procymidone 3 9/07/03 23/07/03 6/08/03

Medium

B Cabbage 5 Nil Nil Nil Nil

C Canola 3 Sumisclex 1.0 mL/L+


Procymidone 1 mid 9/07/03 Nil

D Swedes 4 Nil Nil Nil Nil

E Swedes 4 Sumisclex 1.0 mL/L +


Procymidone 1 14/08/03 Nil

F Chinese cabbage

6 Rovral 1.0 mL/L

Ipriodione 2 9/07/03 23/07/03

Nil

G Chinese cabbage

6 Fortress 1.0 mL/L

Procymidone 2 9/07/03 23/07/03

High

H Chinese cabbage

6 Marvel 0.5 g/L

Benomyl 2 9/07/03 23/07/03

Nil

I Chinese cabbage

6 Nil Nil Nil Nil

J Chinese cabbage

6 Sumisclex 1.0 mL/L

Procymidone 1 early 29/05/03 Severe on oldest, senescent leaves only. Existing leaves were not affected and plants showed fairly good vigour.

K Chinese cabbage

6 Sumisclex 1.0 mL/L + SprayGard


L Chinese cabbage

6 Sumisclex 1.0 mL/L +



M Chinese cabbage



Procymidone 5 regularly

29/05/03 9/07/0323/07/03 6/08/0314/08/03

Severe on all leaves. Low biomass.

N Chinese cabbage



Procymidone 1 mid 9/07/03 Severe on older leaves.

O Chinese cabbage



Procymidone 1 mid 23/07/03 Mild on old and medium leaves.

P Chinese cabbage



Procymidone 1 late 14/08/03 Mild on all leaves. Plants similar in size to plants that were unsprayed.


24

Results 1. Fortress® (procymidone) caused typical ‘black dot’ symptoms. This contains the same strength

of procymidone (50%) as Sumisclex®. This suggests that it is the active ingredient of procymidone which causes the ‘black dot’ symptoms. The chemical companies that manufacture Sumisclex® (Sumitomo Ltd) and Fortress® (Crop Care Ltd) were unable to provide information on the ingredients of the formulation other than procymidone. It is possible that Sumisclex® and Fortress® are identical or have a similar formulation. Symptoms from ‘black dot’ have also been seen on a commercial property at Mandogalup after spraying with Fortress®.

2. Symptoms from ‘black dot’ were similar with Sumisclex® when used with Agral® or Spraygard®, or no wetting agent. This again suggests that the primary cause for ‘black dot’ is from the active ingredient procymidone.

3. Damage from ‘black dot’ was seen when Sumisclex® (procymidone) was applied early, mid-season or late in the crop. The most severe damage was when the crop was sprayed regularly. When applied once and early in the life of the crop, it appeared that the outer leaves were damaged, but that the plants recovered and could have been marketed successfully. Plants that were sprayed mid or late were affected with ‘black dot’ at a mild level. However, temperatures were low (less than 18°C) in July and August when the plants were sprayed. The plants were therefore less damaged than occurred with a late application in Trial 1, when temperatures were over 35°C.

4. Fungicides with the active ingredients benomyl (Marvel®) and ipriodione (Rovral®) did not cause ‘black dot’ symptoms when sprayed on Chinese cabbage. It was also noticed with Marvel®) and (Rovral®) on commercial properties in West Gingin and Mandogalup respectively that these pesticides were not associated with ‘black dot’ symptoms.

5. Sumisclex® caused ‘black dot’ symptoms on cabbages, but not on swedes and canola. This was only a minor observation, but it was noted in a large planting of cabbages at Mandogalup that all plants similar symptoms of ‘black dot’ from an early spraying with Sumisclex®. The planting was observed five weeks later and the damage was confined to the oldest leaves.

Conclusion

Fungicides containing procymidone as the active ingredient are most likely to be responsible for ‘black dot’ symptoms in Chinese cabbage.

5.7 TRIAL 2 - FIELD PLANTS

Background Conclusions from the pot and field observations led us to proceed with a confirmatory randomised, replicated, trial to test the following hypothesis: fungicides containing the active ingredient procymidone (Fortress® and Sumisclex®) will result in ‘black dot’ symptoms on the leaf blades of Chinese cabbage when sprayed on the crop.

Site The site was at an established market garden at Wanneroo on aeolian coastal sands of the Swan Coastal Plain, 24.5 km directly north from Perth CBD and 12 km east of Indian Ocean coastline. This property is owned by the same growers where Trial 1 was conducted and is close to this property.

The grey/yellow soil was of Karrakatta soil type. This consists of deep silicaceous sands. A typical analysis has 81% coarse sand, 18% fine sand 1% clay and a bulk density of 1.5g/mL.


25

The soil is treated every three years with crushed limestone at 5 t/ha. The Chinese cabbage is planted in spring and follows a lettuce crop that is harvested in winter. This has been the pattern for more than 10 years. The trial was situated within a commercial crop of Chinese cabbage.

Aims The main aims were:

(1) To confirm in a randomised, replicated trial that ‘black dot’ in Chinese cabbage is caused by a phytotoxicity from using fungicides that contain procymidone.

(2) To determine in a randomised, replicated trial the effect on yields and quality of Chinese cabbage when procymidone (Sumisclex® or Fortress®) is applied regularly, compared with early or late in the crop, or a nil application.

A subsidiary aim was: to determine in a small observation whether procymidone will cause ‘black dot’ in broccoli and cauliflower.

Experimental design The trial was a randomised block design with seven treatments (as shown below) and four replications.

Treatments 1. Control-procymidone was not applied.

2. Sumisclex®, early applications at 1mL/L on 18 September and 25 September.

3. Sumisclex®, regular applications, at 1mL/L on 18 September, 2 October, 16 October and 30 October.

4. Sumisclex®, late applications at 1mL/L on 23 and 30 October.

5. Fortress®, early applications at 1mL/L on 18 and 25 September.

6. Fortress®, regular applications, at 1mL/L on 18 September, 2 October, 16 October and 30 October.

7. Fortress® late applications at 1mL/L on 23 October and 30 October.

All treatments were applied without wetting agents.

The number of plots for statistical analysis was 28.

Total length of trial = 52.2 m in 1 bay, 6.75 m wide.

Plot size: Treatments were applied to plots 1.68 m wide by 3.73 m long, with four rows of Chinese cabbage per plot. Each treatment had two buffer rows on one side and four buffer rows on the other side (see Table 2). Along the rows, there were four contiguous lines of 10 plants (3.73 m). The outer two plants on each end were buffers and were not sprayed. The number of sprayed plants were 24 per treatment and 20 were used for harvesting for yields and quality. There was a surplus of four sprayed plants per plot to allow for losses from any diseases such as Sclerotinia, pests and off-types.

Planting occurred on 5 September 2003.

Total number of plants was 1,120.

With the subsidiary observation, eight broccoli and eight cauliflower plants were planted on 22nd

September in extended buffer areas beyond plot 21 and between plots 7 and 15 respectively.


26

Figure 16. Trial 12 days after transplanting.

Crop management

The plants established and grew well (see Figure 16, two weeks after transplanting and Figure 1, cover photo, of plants four weeks after transplanting). For detailed information, see Appendix 10.2.

Results All plots were rated for ‘black dot’ (see Figure 17) severity at just over the half way stage to maturity (35 days), by which time, those treatments which had already been sprayed with procymidone showed high levels of ‘black dot’ severity (Table 12). Plots where procymidone had been sprayed also showed reduced growth and vigour.


27

Table 12. ‘Black dot’ and vigour ratings on 10 October (35 days after planting)

Treatments AMean score for ‘black dot’ BMean vigour score

Control 0 10

Fortress® early 6.75 8

Fortress® regularly 7 8

Fortress® late 0 10

Sumisclex® early 7 8

Sumisclex® regularly 7 8

Sumisclex® late 0 10

AScores: B 0-nil 1 very poor vigour 5-moderate 5 moderate vigour 10-very severe 10 excellent vigour

Figure 17. Plant damaged by ‘black dot’, showing dots and associated scorching.

An assessment of the plants immediately before harvest showed that the plants in the unsprayed plots produced good yields, quality and marketability, whereas the plants in the procymidone plots were all unmarketable. The marketability rating for the unsprayed control was significantly better than all procymidone sprayed treatments (Table 13).


28

Table 13. Assessment of marketability per plot (scores 1-5 where 1 = unmarketable, 5 = excellent) , immediately before harvest

Treatment Mean score*

Control 4.75A

Sumisclex® early 1.5

Fortress® early 1.5

Sumisclex® regularly 1.25

Fortress® regularly 1.25

Sumisclex® late 1

Fortress® late 1

LSD p < 0.001 0.8696

*Scores 1 very bad and unmarketable

2 bad and unmarketable 3 fair 4 good and marketable 5 very marketable

An assessment of the plants immediately before harvest showed that the plants in the unsprayed plots showed no external bolting, whereas the plants in the procymidone plots showed more bolting, especially with the early sprayed plots (see Table 14). These were sprayed 13 and 20 days after planting. The regularly sprayed plots also received procymidone 13 days after planting, but there was then a break of two weeks before the next spraying. The two sprays within three weeks of planting probably resulted in a greater shock to the plants, which resulted in more bolted plants with the early spraying with procymidone compared with the plots regularly sprayed with procymidone.

Table 14. Number of external bolted plants immediately before harvest

Treatment Rep1 Rep 2 Rep 3 Rep 4

Number of external

bolted plantsper plot of 24 plants average

Control 0 0 0 0 0

Sumisclex® early 4 6 4 6 5.0

Fortress® early 9 9 9 0 6.75

Sumisclex® regularly 2 1 0 0 0.75

Fortress® regularly 2 3 0 0 1.25

Sumisclex® late 1 1 4 0 1.5

Fortress® late 2 0 4 6 3.0

As well as being affected by more bolted plants, the early sprayed treatment also showed more plants which were affected by soft rot in the three oldest leaves.


29

The leaves affected by ‘black dot’ extended to up to the 12 oldest leaves with the plants that were regularly sprayed and up to the 5 oldest leaves with the early and late sprayed treatments.

There was no ‘Gomasho’ in the plants.

It was not possible to harvest the plants at the optimum time and the plants were therefore harvested five days later on 5 November, 61 days after planting.

Statistical analysis of total head weight data at harvest showed that average head weights per plot were significantly (p < 0.05) reduced by all sprayed treatments, compared with the unsprayed control (see Table 15). The early sprayed treatments of Sumisclex® and Fortress® (see Figure 19) produced significantly (p < 0.001) lower head weights and yield than all other treatments.

The unsprayed control treatment (see Figure 18) had a high total yield (see Table 16). It was the only treatment that produced marketable heads. Figures 19, 20 and 21 show the poor condition of the early and late applications of procymidone at harvest.

Table 15. Average total plant weights harvested from 20 plants per plot (kg)

Treatment Mean

Control (unsprayed) 65.07

Sumisclex® early 44.90

Fortress® early 44.55

Sumisclex® regularly 52.33

Fortress® regularly 53.73

Sumisclex® late 59.95

Fortress® late 57.35

LSD p < 0.001 5.53

LSD p < 0.05 4.04

Table 16. Average marketable total head weights (kg) harvested from 20 plants per plot

Treatment Rep1 Rep 2 Rep 3 Rep 4 Average

Control 62.7 67.2 58.2 72.2 65.08

Sumisclex® early 0 0 0 0 0

Fortress® early 0 0 0 0 0

Sumisclex® regularly 0 0 0 0 0

Fortress® regularly 0 0 0 0 0

Sumisclex® late 0 0 0 0 0

Fortress® late 0 0 0 0 0


30

Figure 18. Control treatment just before harvesting.

Figure 19. Fortress® - early application plot with unsprayed buffers on each side.


31

Figure 20. Fortress® - late application plot.

Figure 21. Sumisclex® - late application plot.


32

The broccoli and cauliflower plants in an observation plot adjacent to the main trial which were sprayed with Sumisclex® were severely damaged by ‘black dot’, whereas the unsprayed plants were unaffected.

6. RESULTS AND DISCUSSION Registration records held by the West Australian Department of Agriculture show that Sumisclex® (procymidone) was released in Western Australia in June 1981 and that it was never registered for brassicas. Fortress® and other brands of fungicide containing the active ingredient procymidone have been registered in more recent times. Procymidone is currently registered with an off-label permit for other brassicas such as broccoli, cabbage and cauliflowers for the control of Sclerotinia. It is a wettable powder in suspension.

Procymidone is not registered for use on Chinese cabbage. There is an off-label permit for use on cabbage. Growers may interpret this to mean that they can use procymidone on Chinese cabbage.

The final trial demonstrated that ‘black dot’ is caused by pesticides that contain procymidone i.e. Fortress® and Sumisclex®. As ‘black dot’ was found with both Sumisclex® and Fortress®, this indicates that the phytotoxicity is most likely caused by the active ingredient and not by the formulation. Procymidone resulted in ‘black dot’, with or without the use of a wetting agent. Damage is worse when the fungicides are applied in the first few weeks after planting. However, all plants that were sprayed with procymidone, at any time, were unmarketable. There was evidence from an observation that procymidone causes less phytotoxicity if sprayed in winter.

Other fungicides, such as benomyl, carbendazim and ipriodione, and the insecticide fiprinil did not result in ‘black dot’.

A few plants of broccoli, cabbage and cauliflower were planted in observations. These were also affected by ‘black dot’. This was also seen to damage cabbages in all plants of a grower’s planting. It would appear that procymidone should not be sprayed on any plants of the Brassiceae family.

7. TECHNOLOGY TRANSFER

INDUSTRY NEWSLETTERS Two articles were published in the West Australian ‘Better Brassicas Newsletter during 2003. An article on the conclusions from the project will be placed in the winter 2004 edition of the ‘WA Vegetable Growers’ magazine and the winter 2004 edition of the West Australian ‘Better Brassicas’ Newsletter.

PUBLICATIONS A Bulletin is being produced from the information collected in this project and presented in this report. The Bulletin is in the form of a best practice guide for the growing, post harvest handling and marketing of Chinese cabbage.


33

8. RECOMMENDATIONS From the results of the trials, the following recommendations were developed:

• Fungicides containing the active ingredient procymidone should not be used for disease control in Chinese cabbage, because it causes ‘black dot’ symptoms and stunts plants.

• Initial observations also suggest that procymidone may adversely affect other brassicas, when sprayed for disease control. The trial work conducted with the Chinese cabbage would need to be repeated with these crops to prove this.

• The off-label permit for use of procymidone on some brassicas should be reviewed.

9. LITERATURE CITED Galati, Angi and McKay, Allan. A Horticultural Research and Development Corporation Final

Report. Carrot Yield Decline, 1996, Experiment 92PE68. Lime reduces cavity spot disease of carrots in sandy soils.

Daly, P. and Tomkins, B. 1995. Production and Post-Harvest Handling of Chinese cabbage, RIRDC Publication 97/1.

Jimenez, M. et al. Chinese cabbage cultivars vary in susceptibility to post-harvest development of black speck, Acta Horticulturae, 1998, No.467.

Kato, T., Muneyasu, T. and Nakayama, N., 1977. Prevention of ‘Gomasho’ in Chinese cabbage. Japanese Society for Horticultural Science. Proceedings, Spring meeting:220-221.

Lelystad, Black in White Cabbage- a Colour Problem, PAV Bulletin, Vollegrondsgroenteteelt, November, 1997.

Matsumata, T. 1981. The Present Status of Chinese Cabbage Growing in Japan. Proceedings of the First International Symposium, Asian Vegetable Research and Development Centre, Taiwan.

National Academy of Sciences & National Academy of Engineering, Water Quality Criteria, United States Environmental Protection Agency, Washington DC, Report No EPA-R-373-033.

Phillips, Dennis, Gersbach, Noel. Factors Influencing petiole spotting (‘Gomasho’) in Chinese Cabbage, Acta Horticulturae, 1989, No.247.

Strandberg, J.O, Darby, J.F., Walker, J.C. and Williams, P.H., 1969. Black Speck, a Non Parasitic Disease of Cabbage. Phytopathology, Volume 59.

Takehashi, K, 1981. Physiological Disorders in Chinese Cabbage. Proceedings of the First International Symposium, Asian Vegetable Research and Development Centre, Taiwan.

Yoshida, T., Ootomo, J. and Okimori, A., 1984. Bulletin 48 of the Hiroshima Prefectural Agricultural Experiment Station.


34

10. APPENDIX

10.1 RESEARCH PLANS AND SOIL AMENDMENTS

Table A1. Randomised plots and sub-plots in main trial, plus Librel BMX observation for Replicated Field Trial 1

Trial

Replications Treatments

T2 B T5

T2 A T4

T2 C 1

T3 T2 D

T1

T2 A T5 T4

T2 C T2 B

2 T1 T3

T2 D T2 C T2 D

T4 T5

T2 A 2

T2 B T1 T3

T2 A T3

T2 B T1

T2 D 4

T5 T2 C

T4

Observation Librel BMX at 1.00 kg/ha Librel BMX at 0.50 kg/ha

Table A2. Soil pH (measured in water) on 14 November 2002, before treatments were applied, and calculations for rates of hydrated lime and sulphur to produce desired pH rates for Replicated Field Trial 1

Treatments Soil pH ∆pH Soil pH ∆pH Soil

pH ∆pH Soil pH ∆pH

1 8.5 7.4 - 1.1 7.7 - 0.8 8.0 - 0.5 8.0 - 0.5 2A +Trace elements

7.5 7.1 - 0.4 7.5 0 7.6 + 0.1 7.8 + 0.3

2B+Trace elements

7.5 7.2 - 0.3 7.8 + 0.3 7.6 + 0.1 7.7 + 0.2

3 6.5 7.3 + 0.8 7.6 + 1.1 7.7 + 1.2 7.9 + 1.4 4 5.5 7.2 + 1.7 7.4 + 1.9 7.7 + 2.2 7.7 + 2.2 5 4.5 7.2 + 2.7 7.2 + 2.7 7.6 + 3.1 7.6 + 3.1 Buffering capacity

1.8 1.9 1.7 1.7

Multiplication factor - Sulphur 1296 1368 1224 1224

Multiplication factor - lime 5400 5700 5100 5100


35

Table A3. Amount of hydrated lime or sulphur to obtain the desired pH treatments for Field Trial 1

Treatments Desired pH Rep 1 Rep 2 Rep 3 Rep 4

T/ha Kg/plot T/ha Kg/plot T/ha Kg/plot T/ha Kg/plot

1 8.5 5.94 HL

8.99 HL 4.56 HL

6.90HHL

2.55 HL

3.86 HL 2.55 HL

3.86HL

2A +Trace elements 7.5 2.16

HL 3.27 HL 0 0 0.12 S 0.18 S 0.37 S 0.56 S

2B+Trace elements 7.5 1.62

HL 2.45 HL 0.41 S 0.62 S 0.12 S 0.18 S 0.25 S 0.38 S

3 6.5 1.04 S 1.58 S 1.50 S 2.27 S 1.47 S 2.22 S 1.71 S 2.59 S

4 5.5 2.20 S 3.33 S 2.60 S 3.94 S 2.69 S 4.07 S 2.69 S 4.07 S

5 4.5 3.50 S 5.30 S 3.69 S 5.59 S 3.79 S 5.74 S 3.79 S 5.74 S

HL = Cockburn hydrated lime S = Dusting sulphur

The rates were calculated from formulas supplied by Dr David Allen of the Chemistry Centre of WA. Based on previous experience on the Swan Coastal Plain, the sulphur rates were increased by 20% and the hydrated lime rates were increased by 100%. The latter was due to normal high buffering at an alkaline pH.

Table A4. pH sampling 17 days after Treatments 1 and 5, to test pH changes in Field Trial 1

Treatment

Desired pH

Rep 1 Rep 2 Rep 3 Rep 4

1 8.5 8.8 8.5 9.1 8.5

5 4.5 5.6 5.6 5.4 5.7

Table A5. Soil pH test results for Field Trial 1

Desired pH (measured in water)

14 Nov 17 Jan 11 Feb 13 Mar 1 Apr Ave

Days before planting 63 Days before soil amendment 33

Days after planting 1 23 53 Days after harvesting 4 Days after soil amendment 28 50 70 89

Treatment 1 8.5 Rep 1 7.4 8.5 8.0 8.3 8.5 Rep 2 7.7 8.4 7.6 8.3 8.5 Rep 3 8.0 8.2 7.7 8.1 8.2 Rep 4 8.0 8.3 7.7 8.3 8.5

Ave 7.785 8.35 7.75 8.25 8.425 8.19


36

Table A5. Continued

Desired pH (measured in water)

14 Nov 17 Jan 11 Feb 13 Mar 1 Apr Ave

Treatment 2A 7.5 Rep 1 7.1 7.8 7.6 7.6 7.9 Rep 2 7.5 7.7 7.2 7.7 7.6 Rep 3 7.6 7.6 7.1 7.8 7.6 Rep 4 7.8 7.4 7.3 7.5 7.8

Ave 7.5 7.625 7.3 7.65 7.725 7.55

Treatment 2B 7.5 Rep 1 7.2 7.7 7.5 7.6 7.7 Rep 2 7.8 6.9 7.0 7.6 7.4 Rep 3 7.6 7.3 7.2 7.6 7.6 Rep 4 7.7 7.6 7.3 7.5 7.9

Ave 7.575 7.375 7.25 7.575 7.65 7.46


Ave 7.625 5.35 6.075 5.5 6.025 5.74


Ave 7.5 4.975 5.625 4.775 4.95 5.08


Ave 7.4 4.275 4.6 3.95 3.95 4.19

Observation (untreated) 7.7 7.9 7.3 7.6 7.7 7.625


37

Table A6. Buffer and sample rows across the bay in Field Trial 1

Rows 1&2 3&4 5 6 Rows 7 to 10 11 12 Rows

13&14 Rows 15&16

Western plots Western

plots Central plots Eastern

plots Eastern plots

B S T S B EB EB T B S S B T EB EB B S T S B

5 m buffer

Observation with Librel BMX at 1.0 kg/ha

Observation with Librel BMX at 0.5 kg/ha

Buffer row EB - Extra buffer row

T - Tractor row S - Sample row

Table A7. Stage of plant growth on 11 February 2003 in Field Trial 1

Treatments - main plots and

sub-plots (Treatment 2)

Desired pH (measured in

water) Rep 1 Rep 2 Rep 3 Rep 4

Average formain plots

and sub-plots

1 8.5 1.5 1.5 2.0 2.0 1.75 2 (average) 7.5 3.0 1.5 1.5 2.0 2.0

3 6.5 1.5 2.5 2.0 1.5 1.875 4 5.5 2.0 2.5 2.0 2.0 2.125 5 4.5 1.5 1.5 1.5 1.5 1.5

Average for main plots 1.9 1.9 1.8 1.8

Legend 1.5 plants at 2 leaf stage 3.0 plants at 4 leaf stage 4.5 plants at 6 leaf stage

The number of stunted plants were assessed on 13 March, 56 days after planting (see Table A8). A total of 68 plants were assessed for each plot.

Table A8. Number of stunted plants on 13 March 2003 in Field Trial 1

Treatments- main plots and





and sub-plots

1 8.5 1 2 0 0 0.75 2A (nil) 7.5 3 0 1 1 1.25 2B (Cu) 7.5 1 0 9 .0 2.5 2C (Mn) 7.5 0 0 1 0 0.25 2D (Fe) 7.5 0 1 3 0 1.0

2 (average) 1 0.25 3.5 0.25 1.25 3 6.5 9 4 2 3 4.5 4 5.5 0 5 2 2 2.25 5 4.5 9 12 13 33 16.75



38

Table A9. Vigour ratings on 13 March in Field Trial 1

Treatments - main plots and





and sub-plots A

1 8.5 7.0 7.0 8.0 6.5 7.125 2A (nil) 7.5 7.0 7.5 6.0 8.0 7.125 2B (Cu) 7.5 7.0 7.5 6.0 7.0 6.875 2C (Mn) 7.5 7.0 7.5 6.5 8.0 7.25 2D (Fe) 7.5 7.0 7.0 5.0 8.0 6.75

2 (average) 7.0 7.375 5.875 7.75 7.0 3 6.5 8.0 8.0 7.0 6.0 7.25 4 5.5 8.0 8.0 7.0 6.5 7.375 5 4.5 5.5 7.0 5.5 5.5 5.875

Librel BMX 1 kg/ha

7.0 some leaf

scorch Librel BMX

0.5 kg/ha 7.0


A 5.0 moderate vigour 7.5 good vigour 10.0 excellent vigour

Table A10. Plan of treatments for Observation 1B

Buffer A B C Buffer Tipa Gold soil drench

8 mL/2L 8 plants

Tipa Gold soil drench40 mL/2L 8 plants

Tipa Gold soil drench 80 mL/2L 8 plants

Buffer 1 2 3 Buffer Spraygard 1 mL/2 L

+ Regent 0.5 mL/2 L 24 plants

Spraygard 1 mL/2 L+ Regent 0.5 mL/2 L+ Howzat 2 mL/2 L

24 plants

Spraygard 1 mL/2 L + Regent 0.5 mL/2L + Howzat 2 mL/2 L

24 plants

Buffer D E F Buffer MnSO4 soil drench

2.5 g/m2 4 plants

MnSO4 soil drench 10 g/m2 4 plants

MnSO4 soil drench 50 g/m2 4 plants

Buffer 4 5 6 Buffer Spraygard 2 mL/2 L

+ Regent 1 mL/2 L 24 plants

Spraygard 2 mL/2 L+ Regent 1 mL/2 L + Howzat 4 mL/2 L

24 plants

Spraygard 2 mL/2 L + Regent 1 mL/2L

+ Sumisclex 4 mL/2 L 24 plants

Buffer G H I Buffer Spraygard

1 mL/2 L 6 plants

Spraygard 10 mL/2 L

6 plants

Spraygard 50 mL/2 L

6 plants


39

Table A11. Trial Plan for Field Trial 2 (buffers are not shown) Broccoli observation

Rep 3 Rep 4

Plot 21 Sumisclex® regularly Plot 28 Fortress® late

Plot 20 Control Plot 27 Sumisclex® late

Plot 19 Fortress® late Plot 26 Fortress® early

Plot 18 Sumisclex® early Plot 25 Sumisclex® regularly

Plot 17 Fortress® regularly Plot 24 Control

Plot 16 Sumisclex® late Plot 23 Fortress® regularly

Plot 15 Fortress® early Plot 22 Sumisclex® early

Cauliflower observation

Rep 1 Rep 2

Plot 7 Fortress® early Plot 14 Fortress® early

Plot 6 Sumisclex® early Plot 13 Control

Plot 5 Sumisclex® late Plot 12 Sumisclex® late

Plot 4 Fortress® regularly Plot 11 Fortress® regularly

Plot 3 Control Plot 10 Sumisclex® regularly

Plot 2 Fortress® late Plot 9 Sumisclex® early

Plot 1 Sumisclex® regularly Plot 8 Fortress® late

Table A12. Buffer and treatment rows across the bay for Field Trial 2

Rows 1&2 Rows 3 to 6 Rows 7 to 10 Rows 11 to 14 Rows 15&16

Buffer rows

Treatment rows (‘X’)

Buffer rows

Treatment rows (‘Y’)

Buffer rows

* *

* *

2 plants buffer unsprayed

* *

* *

* *

* *


* *

* *

*

*

*

*

*

*

*

*

*

*

*

*

Tractor row

6 p l a n t s s p r a y e d

*

*

*

*

*

*

*

*

*

*

*

*

* * * * * *

Tractor row

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

Tractor row

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

* * * * * *

Tractor row

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*

*


*

*

*

*

*

*

*

*


*

*

*

*


40

APPENDIX 10.2 CROP MANAGEMENT AND WEATHER FOR FIELD TRIALS

Field Trial 1

Weed Control At planting, the residual herbicide chlorthal (Dacthal®) was applied at 10 kg/ha, but further hand-weeding was needed three times in the next six weeks. Portulaca weed (Portulacaria oleracea) was the most common weed, followed by Amaranthus species and Galinsoga parviflora (potato weed).

Re-planting with 29 transplants (surplus plants) was required two to three weeks after planting to replace plants that had not survived.

Two weeks after planting, the grower thinned by hand to one plant per site.

Fertilising and irrigation On 8 December 2003, nearly six weeks before planting, the grower applied magnesium and a ‘trace element mixture’ by fertigation to the whole trial site area. This consisted of magnesium sulphate at 50 kg/ha, manganese sulphate at 25 kg/ha; borax, copper sulphate, ferrous sulphate and zinc sulphate at 18 kg/ha; molybdenum sulphate at 2 kg/ha.

Based on the soil analysis, the soil had a good level of phosphorus following a lettuce crop and this nutrient was therefore not applied during cropping with Chinese cabbage.

Prilled potassium nitrate was banded at 160 kg/ha on 20 and 30 January, and on 3 February.

Ammonium nitrate was banded at 200 kg/ha on 3, 6, 10 and 13 February.

From planting to harvesting, the area was fertigated weekly through the sprinklers with boron at 1 kg/ha and magnesium sulphate at 10 kg/ha.

Every Tuesday, from 18 February, potassium nitrate at 97.5 kg/ha, ammonium nitrate at 28 kg/ha and calcium nitrate at 20 kg/ha were applied by fertigation. Every Friday from 21 February, potassium sulphate at 90 kg/ha and ammonium nitrate at 65 kg/ha were applied by fertigation.

The trial was irrigated with ‘butterfly’ sprinklers’(6 m by 6 m spacing), two to three times per day at 120% replacement of evaporation, in accordance with Department of Agriculture recommendations.

Five weeks after planting, a full nutrient analysis was taken from the main bores used in the trial. Results of this analysis are presented in Table B1. The salinity in both bores was low. The water was acidic and also contained a high level of sulphate. This may have contributed to the need by the grower to lime every three years and thereby increase the pH to reduce clubroot. The potassium level in the top bore was high. Both bores contained high levels of magnesium. As with the potassium, these were not toxic and would have contributed to the nutrient needs of the plants. The calcium level was also high, especially in the top bore.

The sulphur level in the water was high and this was odorous after irrigation. Ross Brennan (pers.comm.) reported that in the West Australian ‘Wheatbelt’ sulphur toxicity had not been seen in canola (oil-seed plant in Brassiceae) even where high amounts of gypsum had been applied to the soil.


41

Table B1. Analysis of the two main bores on 17 February 2003

Analyte Unit Top bore Bottom bore

Recommended maximum

concentration (mg/L)A

pH measured in water 5.8 6.0

Electrical conductivity

mS/m 60 38

Aluminium Mg/L 0.18 0.21 5.0

Barium mg/L 0.07 0.03

Boron mg/L 0.14 0.02 2.0

Cadmium mg/L < 0.005 < 0.005 0.01

Calcium mg/L 25 6.4

Chromium mg/L < 0.002 < 0.002 0.10

Cobalt mg/L < 0.005 < 0.005 0.05

Copper mg/L < 0.002 < 0.002 0.20

Iron mg/L 0.27 0.32 5.0

Lead mg/L < 0.02 < 0.02 5.0

Magnesium mg/L 12.0 9.4

Manganese mg/L 0.045 0.006 0.20

Molybdenum mg/L < 0.01 < 0.01 0.01

Nickel mg/L < 0.01 < 0.01 0.20

Phosphorus mg/L 0.4 < 0.1

Potassium mg/L 13.0 2.5

Sulphur mg/L 40 14

Sulphate mg/L 119 41

Vanadium mg/L < 0.005 < 0.005 0.10

Zinc mg/L 0.01 0.03 2.0 A National Academy of Sciences, Pratt (1972)

Pesticides Pesticides were applied by the grower late in the afternoon, mainly to control diamond back moth, aphids and Sclerotinia disease as follows:


42

Table B2. Pesticides applied - Field Trial 1 Insecticides Fungicides

Date Chemical name Active ingredient Chemical name Active ingredient

27 January Orthene acephate Nil Nil 3 February Nitofol methamidophos Nil Nil 10 February Regent fiprinol Nil Nil 17 February Orthene acephate Nil Nil 24 February Success spinosad Nil Nil 24 February Xenthari Bacillus

thuriengiensis Nil Nil

3 March Orthene acephate Nil Nil 6 March Regent fiprinol Nil Nil 10 March Regent fiprinol Howzat carbendazim 17 March Orthene acephate Sumisclex procymidone

Spraygard® (166 g/L D-1-P-menthene) at 300 to 500 mL/1000L/ha was used as a wetting agent (non-ionic sticker and spreader) with all pesticides. At these low rates, Spraygard® was not used as an extender and did not have its own with-holding period.

Climate The weather records were obtained from an official Bureau of Meteorology site, 15 km north of Perth and about 8 km from the trial area (see Table B3.).

Table B3. Weather data 8 km from Field Trial 1.

Data 16 to 31 January 1 to 28 February 1 to 28 March Comments

Rainfall (mm) 1.4 14.2 16.4 Typically low rainfall for season.

Average rainfall (mm) 9.2 13.3 19.1 Average maximum temperatures (°C) 2003

33.0 33.5 32.3 Temperatures were higher than average by 2.2 to 2.9°C.

Long-term (130 years) average maximum temperatures (°C)

30.2 31.3 29.4

Highest maximum temperature (°C) 2003

38.5 41.7 43.2

Average minimum temperatures (°C) 2003

19.0 20.5 20.1 Temperatures were higher than average by 1.9 to 3.9°C.

Long-term (130 years) average minimum temperatures (°C)

17.1 17.4 16.0

Lowest minimum temperature (°C) 2003

15.3 17.5 15.1


43

Plot sampling Samples for nutrient leaf analysis were taken at five weeks after planting and at harvest. A leaf analysis was conducted on a composite 20 plants per plot or on 10 leaves per sub-plot in Treatment 2, 40 days after planting and at harvest.

1 and 2. Sample separately the head wrapper and about 10 leaves in. 3 and 4. With both 1 and 2, analyse the petiole and blade separately.

Samples were sent immediately to Chemistry Laboratory in paper bags and dried at up to 60°C for 24 hours. The leaves were analysed for: Total N, NO3-N, NH4-N, P, K, Ca, Mg, S, Na, B, Cu, Fe, Mn, Mo and Zn.

Field Trial 2

Crop management Beds were prepared by the grower one week before planting by rotary hoeing. Plants of the Ming Emperor variety and approximately three weeks old were transplanted on 5 September from nursery grown plants. The residual herbicide chlorthal (Dacthal®) was applied to the plants after planting to control weeds. This gave good weed control and no hand-weeding was required for the duration of the trial.

The trial area was selected on 17 September, where plants were growing well and uniformly. The soil was tested from 20 sites and was found to have an optimum pH of 6.0 to 6.5 (1:5 water).

Plants grew well and no plant replacements were necessary.

Spraying treatments were applied from 18 September to 30 October 2003. A wetting agent was not used with the pesticides. Spraying occurred after 3 p.m., after the last irrigation and when the foliage was dry.

Fertilising The grower applied a ‘trace element mix’-manganese sulphate at 25 kg/ha, borax, copper sulphate, ferrous sulphate at 18 kg/ha, zinc sulphate at 18 kg/ha, molybdenum sulphate at 2 kg/ha, plus magnesium sulphate at 50 kg/ha to the whole trial site area before planting.

The soil had a high level of phosphorus following a lettuce crop and this nutrient was therefore not applied during cropping with Chinese cabbage.

The transplants were dipped in potassium nitrate at 40 g/L plus MAP at 5 g/L immediately before transplanting.

For the first week, for two applications, prilled potassium nitrate at 36 kg/ha plus urea at 24 kg/ha were applied by banding. For the second week, two applications of prilled potassium nitrate at 200 kg/ha were applied. For the next 10 days, ammonium nitrate, each at 200 kg/ha, were applied by banding for three applications.

For the final three weeks, potassium sulphate at 90 kg/ha plus ammonium nitrate at 65 kg/ha were applied on Wednesdays, and potassium nitrate at 97.5 kg/ha, plus ammonium nitrate at 28 kg/ha, were applied on Saturdays by fertigation.

For the final three weeks, magnesium sulphate at 15 kg/ha plus borax at 1 kg/ha were applied weekly by fertigation.


44

Librel BMX was applied with weekly pesticide sprays (see Table B4.) for seven applications.

The pesticides spray program is shown in Table B4. A proprietary trace elements mixture (Librel BMX) was also included with the pesticides.

Table B4. Spray applications to all plots in Field Trial 2

Trace elements Insecticides Fungicides Date Chemical

name Active

ingredients Chemical

name Active

ingredient Chemical

name Active

ingredient

9 Sept Librel BMX boron copper,iron,

manganese, molybdenum

and zinc

Nudrin methomyl Rovral ipriodione



and zinc

Rogor dimethoate Marvel benomyl



and zinc

Nudrin methomyl Polyram metiram

3 Oct Librel BMX boron copper,iron,


and zinc

Nudrin methomyl Dithane mancozeb

10 Oct Librel BMX boron copper,iron,


and zinc

Fastac cypermethrin Marvel benomyl

Pirimor pirimicarb Polyram metiram 16 Oct Librel BMX boron copper,

iron, manganese,

molybdenumand zinc

Rogor dimethoate Dithane mancozeb

Success spinosad 22 Oct Librel BMX boron copper,

iron, manganese,

molybdenumand zinc

Chess Polyram metiram

Regent Rovral ipriodione

Climate The weather records were obtained from an official Bureau of Meteorology site, 15 km north of Perth and about 13 km from the trial area (see Table B6).


45

Table B5. Weather data 13 km from the trial site for Field Trial 2

Data 5 to 30 September 1 to 31 October 1 to 5 November

Rainfall (mm) 77.6 41.1 21.8 Average rainfall (mm) 80.1 55.4 Average maximum temperatures (°C) 2003

21.2 24.6 29.4

Long-term (130 years) average maximum temperatures (°C)

20.1 22.3 24.9

Highest maximum temperature (°C) 2003

28.6 32.4 32.1

Average minimum temperatures (°C) 2003

11.9 13.6 15.1

Long-term (130 years) average minimum temperatures (°C)

8.9 11.1 13.2

Lowest minimum temperature (°C) 2003

9.2 8.4 12.6

Rainfall was slightly above average. Temperatures were above average.


46

APPENDIX 10.3. NUTRIENT ANALYSIS OF LEAF BLADES AND PETIOLES

Tables C1 to C12 show leaf analyses of different nutrients which were analysed for different plant parts, such as the blades or petioles for the tenth leaf or the head wrapper, at mid-growth and at harvest. They also show the analyses for nutrients at various levels of pH at the mid growth stage and/or harvest. The tables only show areas where significant differences were obtained. Please see also Table 3 in the main text for information on levels of significance for the relationship between pH and ‘black dot’.

The analyses show good levels of phosphorus and trace elements, and adequate to moderate levels of nitrogen, potassium, calcium and magnesium.

The results of these analyses are outside the scope of this report and a separate scientific paper will be needed to cover the relationship between pH, nutrient uptake and yields of Chinese cabbage.

Table C1. Nitrogen

Plant part Growth stage

Blade 10th Petiole 10th Blade head wrapper

Petiole head wrapper LSD p < 0.05

Mid growth 3.789 1.855 3.475 1.484 0.3424

Harvest 3.113 1.587 3.182 1.24 0.3913

Table C2. Phosphorus Stage: Mid growth

Plant part Desired Soil

pH Actual pH Blade 10th Petiole 10th Blade head

wrapper Petiole head

wrapper LSD p <

0.05

4.5 4.2 0.7350 0.645 0.6925 0.6325

5.5 5.1 0.7875 0.665 0.8225 0.7575

6.5 5.7 0.8250 0.6425 0.8750 0.7700

7.5 7.5 0.7078 0.5478 0.6675 0.5125

8.5 8.2 0.6200 0.525 0.5400 0.4575 0.10656

Stage: Harvest

Plant Part Blade 10th Petiole 10th Blade head wrapper


0.6440 0.5245 0.6861 0.5155 0.05274

Desired Soil pH

4.5 5.5 6.5 7.5 8.5 LSD p < 0.05

0.6281 0.6494 0.6658 0.5131 0.5061 0.09807


47

Table C3. Potassium




Mid growth 3.415 3.643 3.062 4.758 0.396

Harvest 3.642 3.222 3.172 3.672 0.2301

Table C4. Calcium




Mid growth 0.4867 0.5272 1.2505 1.0195 0.1588

Harvest 0.310 0.643 0.505 1.040 0.0773

Growth stage: Mid growth

Desired Soil pH

4.5 5.5 6.5 7.5 8.5 LSD p< 0.05

0.7125 0.8544 0.8862 0.8017 0.8500 0.1773

Table C5. Magnesium



0.1662 0.1202 0.2145 0.1505 0.01537

Desired Soil pH

4.5 5.5 6.5 7.5 8.5 LSD p< 0.05

0.1344 0.1512 0.1731 0.1712 0.1844 0.0243

Table C5. Manganese

Stage: Mid growth

Plant part Desired Soil pH -

pH - actual Blade 10th Petiole 10th Blade head

wrapper Petiole head

wrapper LSD p <

0.05

4.5 4.2 73.5 42.5 125.75 53.0

5.5 5.1 49.5 27.0 75.25 35.25

6.5 5.7 33.0 18.25 52.75 24.25

7.5 7.5 19.7 7.7 24.0 10.17

8.5 8.2 17.75 13.5 21.0 9.42 24.92


48

Stage: Harvest

Plant part Desired Soil

pH Blade 10th Petiole 10th Blade head wrapper


4.5 44.25 34.5 62.25 60.5

5.5 30.5 21.25 52.75 32.25

6.5 26.75 18.25 35.83 23.5

7.5 21.0 12.25 23.25 9.43

8.5 19.25 14.5 24.68 11.15 12.648

Table C7. Iron




Mid growth 66.05 37.2 88.55 34.7 16.052

Harvest 69.2 38.9 115.1 51.0 19.24

Table C8. Copper

Stage: Mid growth



7.614 4.129 6.355 3.46 0.7

Desired Soil pH

4.5 5.5 6.5 7.5 8.5 LSD p< 0.05

6.094 6.194 6.269 4.523 3.869 0.7818

Stage: Harvest



7.91 3.19 8.72 2.37 1.292

Desired Soil pH

4.5 5.5 6.5 7.5 8.5 LSD p< 0.05

6.64 6.53 6.13 4.01 4.41 1.275


49

Table C9. Boron

Stage: Mid growth



32.1 29.3 43.85 30.65 3.232

Desired soil pH

4.5 5.5 6.5 7.5 8.5 LSD p< 0.05

33.81 35.13 36.88 32.82 31.25 3.598

Stage: Harvest



29.7 29.15 44.46 32.4 3.302

Table C5. Sodium

Stage: Mid growth

Plant part Desired soil

pH Actual pH -

Blade 10th Petiole 10th Blade headwrapper

Petiole head wrapper

LSD p < 0.05

4.5 4.2 0.4875 0.8925 0.6900 1.4100

5.5 5.1 0.5175 0.9975 0.7900 1.7075

6.5 5.7 0.4525 0.8150 0.7750 1.5950

7.5 7.5 0.1363 0.4363 0.4750 0.8225

8.5 8.2 0.3450 0.3625 0.4525 0.8200 0.3444

Stage: Harvest

Plant part Desired soil

pH Actual pH

Blade 10th Petiole 10th Blade headwrapper

Petiole head wrapper

LSD p < 0.05

4.5 4.2 0.3330 0.825 0.570 0.913

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