TECHNICAL FIELD

The invention relates to the use of nickel, in the form of nickel ions, for the improvement of plant growth and health by conferring resistance to plant pathogens and pests. In particular, the invention relates to the use of nickel for preventing or controlling an infestation of a plant pathogen or pest. The plant pathogen may be a microorganism such as a fungus, bacteria or virus. The plant pest may be nematodes, insects, arachnids or molluscs. The invention also relates to the use of nickel to increase fruit yield and to increase secondary metabolites in plants.

BACKGROUND OF THE INVENTION

Nickel is known to be an essential element for all plant species because nickel is a constituent of urease which is an enzyme engaged in urea hydrolysis. When urea is hydrolysed by urease ammonia is liberated which participates in various anabolic processes, particularly glutamine synthesis. ¹ Nickel deficiency has a severe impact on plants which suggests that there may be other essential roles for nickel. ²

It is generally considered that there is sufficient nickel in most soils, but some researchers have noted that there is evidence of insufficient levels of nickel in plants. Acute nickel deficiencies are known to occur in plantings of pecans, ³ and it has been shown that nickel, zinc and copper are absorbed by the same carrier channel in roots and therefore competitively inhibit each other's uptake from the soil. ⁴ It was concluded that severe nickel deficiency is induced by high amounts of zinc in soil, which is a side effect of more than 50 years of zinc application to pecan orchards to correct zinc deficiencies.

Foliar application of nickel to plants is known for treating some specific plant disorders and diseases. Nickel salts have been used to treat a number of fungal diseases including cereal rusts, pecan scab, rice blast and sheath spot ⁵ , and nickel has been shown to increase the resistance of plants to phytophthora, a type of oomycetes. ⁸ Nickel has also been shown to be effective in treating mouse-ear which is a physiological disorder in pecans. ³

There has been one report ⁶ of nickel toxicity in Xanthomonas oryzae pv.oryzae and, as a result, a decrease in bacterial blight in rice seedlings following spraying with nickel. The authors report that nickel is toxic to the bacteria and therefore blight in the plants is reduced.

In contrast to the above reports of nickel use, the applicant has now found evidence that nickel can be used for improving plant resistance to many plant diseases and pathogens and also enhances fruit yield and growth and the production of secondary metabolites.

It is therefore an object of the invention to provide a method for improving plant resistance to plant pathogens and pests, or to enhance fruit yield and growth and the production of secondary metabolites, using a nickel salt, or to at least provide a useful alternative to existing methods.

SUMMARY OF THE INVENTION

In a first aspect of the invention there is provided a method of improving plant resistance to a plant pathogen or pest comprising applying to a plant a composition containing a nickel salt. The pathogen is a microorganism which may be a bacterium, a virus or certain types of fungi. The pest may be any insect, arachnid or mollusc.

The nickel composition is preferably in the form of an aqueous solution formulated for application as a foliar spray, or as a nutrient growth medium for hydroponically grown plants.

In preferred embodiments, the nickel salt is nickel sulfate.

The concentration of nickel in the composition may vary depending on rate of application. For instance, it may be preferable to apply a concentrated composition to a leaf where the leaf is not fully covered in the composition. If the leave is fully covered by the composition then the preferred range would be 10 to 500 ppm.

The bacteria to which resistance in the plant may be improved include Xanthomonas spp., Acidovorax spp., Erwinia spp., Burkholderia spp., Pectobacterium spp., Candidatus Phytoplasma spp., Clavibacter spp., Spiroplasma spp., Dickeya spp., Candidatus Liberibacter spp,., Ralstonia spp., Rhodococcus spp,., Pantoea spp., Agrobactehum spp., Xy!e!!a spp., and Pseudomonas spp.. The viruses to which resistance in the plant may be improved include positive-strand viruses including Bromoviridae, Closteroviridae, Luteoviridae, and Potyviridae; negative-sense RNA including Bunyaviridae and Rhabdoviridae; double-stranded RNA viruses including Reovirus and Caulimovirus; and single-stranded DNA viruses including Geminivirus.

The fungi to which resistance in the plant may be improved include Ascomycetes including Fusarium spp., Thielaviopsis spp., Veriticullium spp., Magnaporthe grisea and Sclerotinia sclerotiorum; and Basidiomycetes including Ustilago spp., Rhizoctonia spp., Puccini a spp., and Armillaria spp..

The nematodes to which resistance in the plant may be improved include Globodera spp., Belonolaimus spp., Xiphenema spp., Trichodorus spp., Rotylenchulus spp., Tylenchulus spp., Pratylenchus spp., Radopholus spp., Meloidogyne spp., Heterodera spp., Naccobus spp., Bursaphelenchus spp., Ditylenchus spp., Anguina spp., and Aphelenchoides spp..

The insects to which resistance in the plant may be improved include aphids (superfamily Aphidoidea), scale insects (order Hem iptera), psyllid (family Psyllidae), whitefly (family Alyrodidae), mealybugs (family Pseudococcidae) , thrips (order Thysanoptera), grasshoppers (order Orthoptera), weevils (superfamily Curculionoidea), and beetles (order Coleoptera). The arachnids to which resistance in the plant may be improved include mites (subclass

Acari).

The molluscs to which resistance in the plant may be improved include snails and slugs (class Gastropoda) and woodlice (order Isopoda).

The plant may be any plant in the division Pinophyta (conifers) and Magnoliophyta

(flowering plants) .

In a second aspect there is provided a method of increasing fruit set and the quantity of flowers produced by plants comprising applying to a plant a composition containing a nickel salt. The nickel composition is preferably applied before leaf fall in autumn for deciduous plants that flower in early spring or before flower development for evergreen plants and deciduous plants that flower in late spring or summer. The nickel composition is preferably in the form of an aqueous solution formulated for application as a foliar spray, or as a nutrient growth medium for hydroponically grown plants.

In a third aspect there is provided a method of increasing the production of secondary metabolites of plants by applying to a plant a composition containing a nickel salt.

In a further aspect of the invention there is provided a method of correcting a nickel deficiency in a plant comprising determining the nickel content of one or more leaves of the plant, determining the amount of nickel sufficient to correct the nickel deficiency by application of a composition containing a nickel salt, and applying the composition to the plant

DETAILED DESCRIPTION

The invention relates to the use of a nickel salt for improving plant resistance to an infestation of a plant pathogen or pest. The applicant has found that the application of nickel prevents or reduces (in some cases dramatically reduces) the impact of certain plant pathogens or pests. The applicant has also found that the application of nickel enhances flowering and fruit yield and the production of secondary metabolites. These effects appear to be observable, and therefore relevant, for numerous plant species including, without limitation to, conifers and flowering plants.

While it has been observed that nickel is beneficial against some fungal diseases, the mechanism of action is poorly understood and it was thought that the foliar application of nickel was effective due to the toxicity of nickel. However, the applicant considers that the nickel is effective because it is important for the SARS (systemic acquired resistance) and ISR (induced systemic resistance) pathways in plants. The applicant has found that nickel application is not effective at controlling some fungi. Thus, it is uncertain whether nickel would be effective against all fungi. However, it is likely that nickel application would be effective against more types of fungi than the rusts and other fungi which to date are the only types of fungi that have been successfully treated by nickel application. The invention is therefore applicable to the treatment or control of those fungi that are not already known to be susceptible to nickel treatment or would not be expected to be susceptible to nickel treatment. Such fungi include, but are not limited to, Fusarium spp., Thielaviopsis spp., Veriticullium spp., Magnaporthe grisea, Sclerotinia sclerotiorum; Ustilago spp., Rhizoctonia spp., Puccinia spp., and Armillaria spp..

The plant pathogen to which this invention relates may alternatively be a bacterium or a virus. The bacteria include, but are not limited to, Xanthomonas spp., Acidovorax spp., Erwinia spp., Burkholderia spp., Pectobacterium spp., Candidatus Phytoplasma spp., Clavibacter spp., Spiroplasma spp., Dickeya spp., Candidatus Libehbacter spp,,. Ralstonia spp,., Rhodococcus spp,,. Pantoea spp., Agrobactehum spp., Xylella spp,,. and Pseudomonas spp, . The viruses include, but are not limited to, positive-strand viruses including Bromoviridae, Closteroviridae, Luteoviridae, and Potyviridae, negative-sense RNA viruses including Bunyaviridae and Rhabdoviridae, double-stranded RNA viruses including Reovirus and Caulimovirus, and single-stranded DNA viruses including Geminivirus.

The plant pest may be any type of nematode, insect, arachnid or mollusc. For the purpose of this invention, nematodes include, but are not limited to Globodera spp., Belonolaimus spp., Xiphenema spp., Trichodorus spp., Rotylenchulus spp., Tylenchulus spp., Pratylenchus spp., Radopholus spp., Meloidogyne spp., Heterodera spp., Naccobus spp., Bursaphelenchus spp., Ditylenchus spp., Anguina spp., and Aphelenchoides spp. Insects include, but are not limited to, aphids (superfamily Aphidoidea), scale insects (order Hemiptera), psyllid (family Psyllidae), whitefly (family Alyrodidae), mealybugs (family Pseudococcidae) , thrips (order Thysanoptera), grasshoppers (order Orthoptera), weevils (superfamily Curculionoidea), and beetles (order Coleoptera). Arachnids include, but are not limited to, spider mites (subclass Acari). Molluscs include, but are not limited to, snails and slugs (class Gastropoda) and woodlice (order Isopoda).

The nickel containing composition may be applied to the plant in any suitable manner.

In certain embodiments of the invention, the nickel is applied to the plant in the form of an aqueous solution of one or more nickel salts. The nickel salt is typically nickel sulfate, but may also be nickel nitrate, nickel chloride or nickel complexes such as nickel citrate, or mixtures thereof. It will be appreciated that various factors may influence the choice of nickel salt including solubility in water and ease of absorption by the plant.

The concentration of the nickel in the composition will be within any suitable range to enable the plant to absorb or otherwise access sufficient nickel to be effective against the pathogen or pest, but not too much such that the plant's health is compromised. In certain embodiments of the invention, the concentration of nickel in the composition is in the range 10 to 500 ppm, but may be in any other suitable range, such as range 50 to 250 ppm, depending on the application method, the type of plant, and the pathogen or pest to be controlled. The composition may additionally comprise one or more other plant nutrients. Such nutrients may be selected from the group comprising salts or complexes of calcium, potassium, magnesium, iron, copper, manganese, zinc and nickel, anions of molybdenum, boron, chlorine, sulphur and phosphorus, and nitrogen sources such as ammonium nitrate and urea. Any other desirable chemical may be included in the composition. In some preferred embodiments, the nickel containing composition may additionally include a potassium, nitrogen or zinc source. The addition of potassium and nitrogen to a nickel containing foliar spray is useful for flowering and fruit set. The addition of zinc is useful in foliar sprays since it avoids the problem of over application of zinc to soils. The foliar application of nickel within a month before or following fertilisation with a nitrogen source may be beneficial for controlling pest species as some pest infestations are associated with the application of nitrogen fertilisers.

The application of nickel to the plant in the form of an aqueous composition is preferably by way of a foliar spray, i.e. the foliage of the plant is sprayed with the composition. The nickel may alternatively be applied by incorporation into the nutrient feed solution of hydroponically grown plants. In some embodiments, a combination of application methods may be used, e.g. a nickel containing hydroponic feed solution and a foliar spray.

For best results, nickel may be applied at different times of the year. For example, plants may preferably be sprayed during summer to encourage plant growth. They may also be sprayed before blossom formation to encourage flowering. For certain plant species and certain pathogens or pests, best results may be achieved by applying nickel multiple times before, during or after the growing season.

The applicant investigated the application of nickel to a variety of plant species including figs, lemons, limes, grapefruit, pears, plums, raspberries, and blackberries. The garden soil was fertilised with citrus fertiliser, aglime, compost from food waste, weeds and other plant clippings. Mineral deficiencies were noted for a number of plants. Plum trees severely attacked by leaf curl aphids had leaves with reddish margins with brownish green centres. Citrus trees and some other trees often had pale yellowish leaves especially in spring which was difficult to correct by the application of fertiliser.

It had previously been considered that the careful application of fertilisers and pesticides where appropriate was sufficient for the enhancement of plant health and for optimising fruit quality and yield for most gardens. Trace elements required by plants, and not provided by standard fertilisers, have been expected to be naturally occurring in most garden soils. However, the applicant has found that the application, particularly the foliar application, of nickel salts dramatically improved plant growth and fruit yield.

Following the application of a nickel foliar spray to thornless blackberry canes, which had been growing to 1 to 1.5 metres in length for over a decade, instead grew to 9 metres. Raspberry canes that previously produced a single shoot started to grow laterals and the autumn fruit yield was substantially higher. Previously it had been observed that the grapefruit had only one growth of new shoots during spring but upon foliar application of nickel several growth cycles were observed. Flowering was also more prolific in a number of the fruit trees.

In the second season following application of nickel, the fig and pear trees produced approximately 100% more fruit and the plum tress about 600% more fruit than in previous seasons. The raspberry canes grew about 150% to 300% taller with a very good supply of berries produced throughout summer. Multiple new blackberry canes appeared whereas in previous seasons there had been only one or two new canes.

In contrast, in the second season, none of the grapefruit trees had flowers and it is surmised that this may be due to lack of foliar application of nickel and possibly potassium prior to the expected spring flowering. However, in the third season with the application of other nutrients to the foliar spray there was a very large increase in blossom yield.

The effect of foliar application on pear slugs was also examined. It was noticed that pear slugs were absent on pear trees which had three biweekly applications of summer oil to treat codling moth once fruit formation commenced. Plum trees which had not been treated were attacked by pear slugs, but only two pear slugs were observed on the pear trees. The plum trees were then treated with nickel foliar spray, whereas the pear trees remained untreated. Usually, the pear slug will have two cycles of infestation with the second infestation typically being the worst. With the second infestation no pear slugs were found on the plum trees, but after the application of summer oil was discontinued the pear trees were severely infested with some leaves showing 80% damage.

A kowhai plant, a deciduous shrub, was subject to severe infestation of the kowhai moth caterpillar each spring on the appearance of the new growth, so much so that most of the leaves were stripped from the plant. However, for two years following the application of nickel, kowhai moth infestations were drastically decreased with very little foliar stripping occurring.

Experiments were undertaken to investigate the effect of adding nickel salts to hydroponic solutions for controlling greenhouse pests, in particular spider mites and aphids. The plants were grown using the nutrient film technique where a shallow stream containing the nutrients flows past the roots of the plants contained in a PVC gully. A 70 L tank of water was dosed with the nutrient solution so that the conductivity of the solution was about 1.5 mS cm ^"1 . The conductivity of the tank was checked once a week and more nutrient solution was added to maintain a conductivity of 1.5 mS cm ^"1 . About once a month the solution was completely replaced with clean water and nutrient.

Strawberries, tomatoes, peppers, dill, basil, rock melon, honeydew, water melon, lettuce and cucumber were grown in three gullies. As with the foliar application of nickel to plants as described above, the effect of adding nickel to the growth media was dramatic. Spider mite infestation was controlled to a high degree, as was aphid infestation, except for three cases where foliar application of additional nickel proved more effective.

Many modern food crops have problems with pathogens and pests, although all have essentially the same metabolism as their wild progeny. For instance, citrus plants are subject to the following pests and diseases in the USA: rust mites, spider mites, broad mites, scale insects, whiteflies, aphids, mealybugs, citrus root weevils, orangedog, grasshoppers, katydids, termites, Asian cockroach, Caribbean fruit fly, flower thrips, citrus root weevil, nematodes, citrus greening, citrus canker, phytophthora, brown rot of fruit, greasy spot, melaose, citrus black spot, citrus scab, alternaria brown spot, postbloom fruit drop, various viroids, blight, and tristeza. ⁷ Organic farmers have noted that applying nitrogen containing fertilisers increases the incidence of disease and pests species.

Plant pests and diseases are also becoming increasingly problematic to treat. PSA (Pseudomonas syringae pv actinidiae) is a disease of kiwifruit vines that cannot be eradicated and is difficult to control. Often vines have to be pulled up after 10 years of growth. Fire blight (Erwinia amylovora) is a disease of Rosaceae, but effects mainly pears and apples. It is becoming difficult to control requiring the application or even the injection of antibiotics. Citrus greening disease (Candidatus Liberibacter spp.) threatens the entire citrus industry in the United States and there is no known treatment for the disease.

Without being bound by theory, it is considered that since nickel is an essential part of urease the application of high nitrogen fertilisers without the concomitant application of nickel is disadvantageous. Some modern agriculture practices may be harmful as well. For example, the application of zinc in particular appears to inhibit the uptake of nickel. ⁸ It may be that plants can readily absorb nitrogen compounds through the roots but that the uptake of nickel is much slower, leading to nickel deficiency in modern high yielding crops. The mechanism of how this affects pests and pathogens remains speculative, but since it is known that nickel is critical for the function of urease in plants, excess nitrogen (possibly as urea) could outgrow the supply of nickel important for controlling pests and diseases in plants. Lack of nickel could also disrupt plant defence mechanisms. Plants have two main signalling molecules that are activated when plants are attacked by pests or diseases. Salicylic acid signals the attack of biotropic pathogens and jasmonate signals the attack of necrotrophs including wounding and herbivory from plant pests, unless ethylene is also present in which case it can also activate defences against necrotrophs. ⁹ It is recognised that activation of one of the pathways can be antagonistic against activation of the other pathway and some pathogens can use this to their advantage. Activation of these pathways gives enhanced resistance against pests and diseases. For salicylic acid, the resistance pathways is known as Systemic Acquired Resistance (SAR) and for jasmonate, Induced Systemic Resistance (ISR). Since nickel seems to have a broad-spectrum effect, it is likely that nickel plays an important role in SAR and ISR pathways. In tobacco plants, SAR activation results in significant reduction of disease symptoms caused by the fungi Phytophthora parasifica, Cercospora nicotianae, and Peronospora tabacina, the viruses tobacco mosaic virus (TMV) and tobacco necrosis virus (TNV), and the bacteria Pseudomonas syringae pv tabaci and Erwinia carotovora. Interestingly, the activation is not effective against the fungal diseases Botrytis cinerea or Alternaria alternate. The applicant also found that nickel application alone appears to be ineffective against some fungal pathogens, namely powdery mildew (probably Podosphaera xanthii) and peach leaf curl (Taphrina defomans).

It has recently been realised that ureases have important toxic properties independent of their enzyme capabilities. The ureases "moonlight" as potent toxins for pathogens and pests. ¹⁰ The applicant speculates that lack of nickel in plants does not allow the plant to produce enough ureases to adequately fight diseases and pests. But this may not be the only mode for which nickel is helpful in controlling plant pests and diseases.

Since activation of the SAR and ISR pathways is an effective response against many pests or diseases, ^11,12 it is expected that nickel application according to the invention will be effective in reducing or eliminating many diseases and pathogens in different kingdoms in the domain eukarya which includes plantae, animalia, fungi and algae as well those in the domain bacteria. As activation of the SAR pathway has been shown to be effective in reducing viral infections in plants, it is expected that nickel application will be effective in reducing or eliminating viral infections in plants.

Support for the invention is found in the experiments described in the Examples below. Example 1 describes an experiment designed to observe the infestation of green peach aphids on hydroponically grown dill. Example 2 describes an experiment relating to the treatment of aphids on hydroponically grown jalapeno peppers. Example 3 is an experiment showing the control of spider mite infestation on hydroponically grown cucumber, melon and strawberry plants. The addition of nickel to the hydroponic nutrient media proved effective in controlling both aphids and spider mites. However, the water melon and cucumber became infected with spider mites and the jalapeno pepper became infected with green peach aphids five months after planting. As a gully hydroponic system was used these plants were further downstream from other plants and may have been deprived of adequate nickel. In contrast, the rock melon and honeydew melon were both upstream of any other plant and no sign of spider mite infestation of the rock melon was observed and only a small outbreak of spider mite infestation of the honeydew melon was observed and the infestation did not spread to the rest of the plant.

The green peach aphid infestation of the jalapeno pepper plant was controlled with three foliar sprays of 200 ppm aqueous nickel sulfate (NiSC .ehhO). The effect was dramatic. The jalapeno pepper went from a severe infestation with aphids being plentiful on both the underside of the leaves and on new shoots to almost complete elimination of aphids, with only one or two aphids left on the whole plant, approximately two weeks after the last foliar application of nickel.

The effect on spider mites was also dramatic with no infestations observed on the rock melon and strawberries and only a small outbreak, which never spread, observed on the honeydew melon. However, spider mites did become established on the cucumber and water melon but the spread of the aphids was not dramatic and some foliar application of nickel salts kept the infestation under control until the end of the season. It is not apparent why these two species should be more susceptible to infestation, but it was noted that in the case of the rock melon and honeydew melon the plants were on the upstream end of the gully and therefore were not competing for micro-nutrients, whereas for the water melon and cucumber they were on the downstream end of the gully and may have been competing for micro- nutrients. Another possible explanation is that nickel absorption through the roots by cucumber, rock melon and jalapeno peppers was being blocked by other nutrients in the hydroponic solution or that absorption of nickel is less facile in these plant species.

The nickel concentrations used in the hydroponic solution was formulated based on the nickel content of leaves for a number of species in northern Europe. ¹³ The content used was at the low end. Higher nickel concentrations in the hydroponic solutions may be more suitable for certain plant species.

In Example 4 it was found that strawberry plants that are very susceptible to infestation by spider mites and to a lesser extent by aphids showed no sign of either pest. The roots of tomato plants showed no signs of aphid infestation even though in previous years the roots had been infested by aphids.

Pear slugs appeared to be controlled by the application of a nickel foliar spray as shown in Example 5. Pear slug injury to plants occurs in two peaks with the second peak much more destructive than the first. Treatment of the plum tree during the first peak with a foliar spray containing 200 ppm aqueous nickel sulfate was effective in completely eliminating pear slugs on the plum tree during the second more destructive peak. In contrast, the untreated pear tree was severely infested with pear slugs during the second peak, especially on the new season growth, with some leaves showing up to 80% damage.

The pear tree was also infected with fire blight in the summer of 2015. Canker blight, an infection phase of fire blight, is always present when fire blight was a problem in the previous season. ¹⁴ However, as described in Example 6, after spraying the pear tree with 200 ppm nickel sulfate in the late summer of 2015, the pear tree showed no symptoms of fire blight, including canker blight, throughout the summer of 2016. Similar results were obtain in 2017 (see Example 13) although only on one of the grafts which was not treated with nickel. Example 7 describes the treatment of a kowhai plant with 50 ppm aqueous nickel sulfate which resulted in a dramatic reduction in damage from kowhai moth caterpillars. Throughout the previous decade, the plant had been subjected to foliage stripping each spring from kowhai moth caterpillars, which ensured that very little growth occurred during spring, summer and autumn. However, after foliar application of nickel no damage from kowhai moth caterpillars was observed and the plant tripled in size in the spring and summer following nickel application. At the beginning of autumn leave were stripped from some of the most recent growth perhaps indicating depletion of nickel after a good growing season. Kowhai plants are legumes and nickel is not only essential for urease activity, but nickel is required for bacteria involved in nitrogen fixation in root nodules on the legumes.

A number of fruit trees were sprayed with aqueous nickel sulfate in the autumn of 2015 as described in Example 8. A dramatic increase in fruit yield was observed for the plum, pear and fig trees. A very dramatic increase in the length of blackberry and raspberry canes was observed and, with the raspberries especially, a sustained cropping of berries occurred throughout the summer, especially the late summer.

The citrus fruit plants, which included two varieties of grapefruit and one variety of lime, did not flower throughout the whole of the summer of 2016. This was unexpected because in the two previous seasons flowering had occurred with better yields after foliar application of nickel in the spring. In 2016 there was no foliar application of nickel before flowering and this may have resulted in plant resources going towards growth rather than flowering and fruiting. From these results it appears likely that fruiting and/or growth can be controlled by the application of foliar sprays at certain times of the year especially for citrus plants. In plants deficient in nickel dramatic increases in yields of fruit occurred with the application of nickel containing foliar sprays. In Example 15 the grapefruit were treated with a composition containing various plant nutrients as well as nickel. This resulted in many bud clusters on the trees in the spring of 2016.

In Example 9 jalapeno peppers were tasted before and after treatment with nickel. The pepper fruit which had no pungency before treatment became very pungent. Capsaicin, the cause of pungency in chili peppers, is a secondary metabolite that is an organic compound not directly involved in growth, development or reproduction of the plant. Many secondary metabolites are present in plants and many act as a deterrent to pathogens and pests. In the case of chili peppers capsaicin acts as a deterrent to predation by small mammals. ¹⁵ It is only after the aphid infestation and subsequent spraying with nickel that the capsaicin content of the peppers increased . Many secondary metabolites of plants are important natural products. Nickel application and possibly stressing the plant should increase the concentration of some of these metabolites. The taste of many plant foods is dependent on secondary metabolites, for instance tannins in grapes or indeed capsaicin in peppers. If viticulture is used as an example, the taste and flavour of wines could be controlled through the application of nickel and the stressing of the plant by for instance withholding water.

In Example 10 leaf analysis of kiwifruit vines showed that plants deficient in nickel showed signs of PSA infection whereas plants that were not deficient did not show signs of PSA infestation. This is consistent with the prior evidence of the importance of nickel content in plant defense mechanism

Examples 11 and 12 show that infestations with spider mite on cucumbers were controlled by addition of nickel to hydroponic solutions used to grow the cucumbers. This again is a significant result as spider mites are important horticultural pests in hothouses.

Leaf curl plum aphids were controlled in spring by the application of nickel to plum trees in autumn. For a decade prior to this leaf curl plum aphids had been increasingly problematic to control eventually causing the death of two grafts on the plum. This shows not only the effectiveness of nickel to control these aphids but also the translocation of nickel from the leaves in autumn to be used in the new leaf growth in spring.

Example 15 shows that treatment with nickel and other plant nutrients can cause a surprising extensive autumn flowering in a young orange plant where typically orange trees flower in spring.

Spraying of a white sapote (Example 17) with nickel and other plant nutrients formed very many flower clusters along the woody branches of the tree.

Any reference to prior art documents in this specification is not to be considered an admission that such prior art is widely known or forms part of the common general knowledge in the field.

As used in this specification, the words "comprises", "comprising", and similar words, are not to be interpreted in an exclusive or exhaustive sense. In other words, they are intended to mean "including, but not limited to".

The invention is further described with reference to the following examples. It will be appreciated that the invention as claimed is not intended to be limited in any way by these examples.

EXAMPLES

Example 1: Treatment of green peach aphids (Myzus persicae) on hydroponically grown dill

In mid-summer green peach aphids had been observed to be feeding on hydroponically-grown dill both on the stems and the leaves.

The hydroponic nutrient solution was based on that of Hoaglund, ¹⁶ but with added nickel according to the following table: Salt Amount g/L

Calcium nitrate 360

Potassium nitrate 200

Potassium dihydrogen phosphate 50

Magnesium sulphate 160

Iron EDTA 6

Sodium borate 1.3

Copper sulphate 0.04

Manganese sulphate 0.9

Zinc sulphate 0.2

Nickel sulphate 0.02

Ammonium molybdate 0.006

The plants were sprayed with aqueous nickel sulfate (NiSC .ehhO) (200 ppm) and again 3 days later. After 7 days it was observed that very few aphids were left on the stem and there was a decrease of aphids on the leaves.

Example 2: Treatment of green peach aphids (Myzus persicae) on hydroponically grown jalapenos peppers

Aphids had been observed infesting hydroponically grown jalapenos peppers. The hydroponic nutrient solution used is described in Example 1. A single leaf was sprayed with aqueous nickel sulfate (NiSC .ehhO) (200 ppm), but little reduction in infestation by aphids of the leaf or any other leaves on the plant was observed after 5 days. The whole plant was then sprayed with aqueous nickel sulfate (NiSC .ehhO) (200 ppm). After 4 days some leaves were observed to be clear of aphids, but the plant was still badly infested, especially around the terminal leaves where new leaf growth was occurring. The plant was again sprayed with the same nickel solution. After 5 days a great decrease in aphids was observed and after a further 4 days the terminal leaves were free of aphids. After an additional 13 days only 2-3 aphids were found on the whole plant, and the plant had resumed normal growth. The plant showed no signs of nickel toxicity. Example 3: Treatment of two-spotted spider mite (Tetranychus urticae) on hydroponically grown cucumber and melon

Cucumber, rock melon, honeydew melon and water melon were grown hydroponically in the presence of nickel as described above. The hydroponic nutrient solution used is described in Example 1. The plants were sprayed once with summer oil to prevent spider mites. Instead of continuing the weekly treatment of summer oil, as in previous seasons, the treatment was stopped to see if the plants now grown in the presence of nickel would be less susceptible to infestation by spider mites.

Spider mites were noted to be present in the greenhouse as fine webs across the openings in the gullies. Throughout the season no spider mites were found on the rock melon. For the honeydew melon 2-3 leaves showed signs of damage consistent with spider mites and examination under a microscope showed the presence of the two-spotted spider mites. However, throughout the rest of the growing season no spread of the spider mites to adjacent leaves or new growth was observed.

Spider mites found on the water melon after five months of growth were kept under control with 2 sprayings of aqueous nickel sulfate (N 1SO4.6H2O) (200 ppm), although towards the end of the season half of the rock melon leaves showed chlorosis due to spider mite infestation. The cucumber showed some signs of spider mite infestation, but even by the end of the season most of the leaves showed only mild damage.

Example 4: Treatment of other hydroponic growing plants.

Hydroponically grown strawberry plants, which are susceptible to spider mite and aphid infestations, showed no signs of infestation throughout the growing season. Hydroponically grown tomato plants which in previous seasons had aphid infesting the roots showed no signs of aphid infestation.

Example 5: Treatment of pear slugs (Caliroa cerasi)

During summer pear slugs were observed stripping the upper surface of the leaves on a plum tree leaving behind a skeleton of veins. The tree was sprayed twice with aqueous nickel sulfate (N1SO4.6H2O) (200 ppm) and sprayed several times with a balanced nutrient foliar spray containing nickel. The balanced spray resulted in a much improved leaf colour and the growth of new shoots. By late summer no pear slugs were observed on the tree.

In the first month of summer, a triple grafted pear tree was sprayed twice with summer oil to control codling moth, which had also been noted to be effective in controlling pear slugs. The summer oil spraying was stopped, and no nickel sprays were applied to the tree throughout summer. Only 2-3 pear slugs were found on the tree by the second month of summer. However, in late summer there was a severe infestation of pear slugs especially on the new growth which had not been exposed to nickel treatment. Some leaves showed 80 % damage. But, as noted above, not one pear slug was found on the nearby nickel -treated plum tree. Example 6: Treatment of pear for fire blight ( Erwinia amyiovora)

In mid-summer of 2015 it was noticed that a triply grafted pear tree had shrunken, blackened shoots and stems including fruit symptoms characteristic of fire blight. The tree was sprayed with aqueous nickel sulfate (NiSC .ehhO) (200 ppm) in late summer 2015. In the summer of 2016 there were no symptoms of fire blight on any part of the pear tree throughout the growing season.

Example 7: Treatment of kowhai to treat infestation of kowhai moth (Ureiphita polygonalis)

A small kowhai plant was treated with aqueous nickel sulfate (NiSC .ehhO) (50 ppm) in the spring of 2015. The plant responded favourably with good growth throughout the summer of 2016. No kowhai moth caterpillars were observed on the plant even though for the previous decade the kowhai plant had been denuded of all leaves in the spring from kowhai moth caterpillars. The plant almost tripled in size during this one season whereas for the previous seasons very little growth was observed.

Example 8: Treatment of fruit trees for increased yield of fruits

In autumn of 2015 plum, pear, fig and a New Zealand grapefruit trees were sprayed with aqueous nickel sulfate (NiSC .ehhO) (200 ppm). It was estimated that in the following summer the yield of fruit from the fig and pear trees had almost doubled and from the plum tress had quadrupled. In the spring of 2014 the grapefruit trees were treated with a foliar spray of aqueous nickel sulfate (NiSC .ehhO) (200 ppm) which resulted in a good flowering. The yield of fruit in the winter of 2015 was good considering the tree size. The g rapefruit trees were again treated with a foliar spray of nickel sulphate in the autumn of 2015 but not in the spring of 2015. No flowers were formed on any of the grapefruit but good leaf growth was observed throughout the summer of 2015/2016.

Example 9: Treatment of jalapeno pepper with nickel to increase capsaicin content

Prior to treatment with nickel, jalapeno peppers were taste-tested for pungency (spicy hot). No pungency could be detected. After treatment for aphids as described in Example 2 the peppers were again taste-tested. Small slices of the pepper were found to be very pungent.

Example 10: Analysis of leaf samples of kiwi fruit showing signs of PSA and those showing no signs of PSA

Leaf samples of two commercial kiwifruit vines (Haywards) one showing spotting due to PSA (Pseudomonas syringae actinidiae) and the other without any spotting were obtained from different areas in a single orchard in the Bay of Plenty, New Zealand. Both samples were dried at 100 °C for 45 min, dissolved in aqua regia and analysed by ICP-MS for nickel content. The leaf sample from the vine showing infection with PSA had a nickel content below detectable limits of 0.5 mg/Kg. The leaf sample from the vine that had no spotting had a nickel content of 0.9 mg/Kg.

Example 11: Treatment of two-spotted spider mite (Tetranychus urticae) on hydroponically grown cucumber previously infected with spider mite

Five Fl hybrid cucumbers (Cucumis sativus) were grown hydroponically from seed using a gully system. The nutrient solution added to the tank (40 L) was a modified Hoaglund solution as described in Example 1, which was replaced every two weeks. After 4 weeks the plants showed damage due to spider mites already present in the glasshouse. Nickel sulphate (5 mg) was then added to the tank. All the new growth showed little to no sign of spider mite damage. Example 12: Treatment of two-spotted spider mite (Tetranychus urticae) on hydroponically grown cucumber infected with spider mite using a control

Eight Fl hybrid cucumbers (Cucumis sativus) were grown from seed using a guily system and 40 L tanks. After 5 weeks all plants showed considerable damage from spider mites present in the hothouse. Four of the plants were placed in a separate gu!ly and NiS04.6H?.0 (8 mg) was added to one of the tanks. Four day later more NiS0 .6H20 (8 mg) was added to this tank. Two weeks later the leaves of the treated and untreated plants were examined for spider mite infestations. Leaves were examined for spider mite infestations 4 cm away from the growing tip. For the untreated cucumber plants, on average 2.5 leafs away from this 4 cm mark there was signs of spider mite damage. For the treated cucumbers, the average value was 3.7.

Example 13: Treatment of leaf curl plum aphids (Brachycaudus helichrysi)

Plum trees were sprayed with iS04.6H?.0 (200 ppm) in the third week of April one month before leaf fall. The leaves of the new spring leaf growth on the trees were examined for leaf curl aphids. No aphids were found. For the past 5 years serious infestation of leaf curl plum aphid had occurred leading to the eventual death of two grafted trees.

Example 14: Treatment of fireblight in pear tree (Erwinia amylowora)

Two of the grafts of the pear tree of Example 6 were sprayed with 200 ppm iSCM.ehbO in late April. By early January the untreated graft showed symptoms consistent with fireblight whereas none of the treated grafts showed any symptoms. Example 15: Treatment of an orange tree to induce off-season blossoming

Aglime (CaCGs, 1200g) and dolomite (1000 g) were stirred with pure playground sterilised sand (quartz based). A young grafted orange tree which was 60 cm tall was purchased and the roots were washed clean of the soil and compost, and the tree was planted in the sand mixture. A foliar spray was made by dissolving the following components in water:

The potassium nitrate, ammonium nitrate and potassium hydrogen phosphate were dissolved in 400 ml of water and the remaining components dissolved in 200 ml of water. The two solutions were mixed and then diluted to a quarter of their previous concentration and this was used as a foliar spray. The foliar spray was then applied to the orange tree every two weeks. The results were:

14 Feb 2016: Orange tree planted in sand mixture.

9 March 2016: New shoots appear on the orange tree.

11 April 2016: Numerous flower buds at the leaf nodes appeared on several of the old stems.

8 May 2016: First flower opens and new flower buds noticed on new shoots.

3 June 2016: Up to eight small fruit set at each node on the old growth. This was a surprising outcome since the flowering is in the opposite season to what would be expected. In contrast untreated orange trees remaining with the seller had no new growth or flowering.

Example 16; Treatment of a grapefruit tree with a nickei containing soiution

The New Zealand gra pefruit tree of Example 8 was sprayed with a foliar spray having a composition as described in Example 14 every two weeks for 2 months in April and May of 2016. This produced spectacular flowering in the spring of 2016, with many bud clusters on the tree in the spring of 2016.

Example 17; Treatment of a white sapote (Casimiroa Eduiis) grapefruit tree with a nickei containing solution

A variety of white sa pote was sprayed three times in March of 2016 with a foliar spray having composition as described in Example 14, By mid-April there were many flower clusters forming along the woody branches of the tree, where leaves were completely absent, and many on the new shoot growth as well .

Although the invention has been described by way of example, it should be appreciated that variations and modifications may be made without departing from the scope of the invention as defined in the claims. Furthermore, where known equivalents exist to specific features, such equivalents are incorporated as if specifically referred in this specification.

REFERENCES

1. I. V. Seregin and A. D. Kozhevnikova. Russian Journal of Plant Physiology 53, 257, 2006.

2. Cheng Bai, Liping Liu and Bruce W Wood . BMC Plant Biology 13, 207, 2013.

3. Bruce Wood, Charles Reilly, Andrew Nyczepir. HortScience 39, 858, 2004.

4. L.V. Kochian. Micronutrients in Agriculture. 2nd ed. Soil Sci. Soc. Amer., Inc p 229- 296.

5. D. Mishra and M. Kar. Botanical Review 40, 395, 1974.

6. H . Wang, F. Zeng, C. Cao, and J. Kang, J. Hunan Agr. Univ. 26: 119-121, 2000.

7. Florida Citrus Pest Management Guide, http://www.crec.ifas.ufl.edu/extension/pest/

8. Patrick H. Brown. CRC-Handbook of Plant Nutrition 2007 ch 14.

9. M.R. Grant and J.D.G Jones Science 324, 750, 2009.

10. C.R. Carlini and R. Ligabue-Braun. Toxicon 110, 90, 2016.

11. G. E. Vallad and R. M. Goodman. Crop Sci. 44, 1920, 2004.

12. John A. Ryals, Urs H. Neuenschwander, Michael G. Willits, Antonio Molina, Henry-York Steiner, and Michelle D. Hunt. The Plant Cell, 8, 1809, 1996. 13. Clemens Reimann, Friedrich Koller, U , Bjorn Frengstad, Galina Kashulina, Heikki Niskavaara and Peter Englmaier, The Science of the Total Environment 278, 87, 2001.

14. Sharon M. Douglas. Fire Blight. The Connecticut Agricultural Experiment Station, New Haven.

15. Joshua J. Tewksbury and Gary P. Nabhan. Nature 412, 403, 2001.

16. Dennis Hoaglund. The water-culture method for growing plants without soil (Circular (California Agricultural Experiment Station), 347. ed.). Berkeley, Calif, : University of California, College of Agriculture, Agricultural Experiment Station. Retrieved 1 October 2014.