TECHNICAL FIELD OF THE INVENTION

The present invention relates to the use of polyethylene glycol to prevent and/or combat the proliferation of plant-infesting nematodes.

PRIOR ART

Spontaneous or cultivated plants are normally exposed to adversities of plant or animal origin capable of producing severe damage to the plants themselves and to the products obtained from them.

One of the adversities of biotic origin that is least known, but of enormous importance, is represented by nematodes. These multicellular organisms, having a filiform appearance and a size such that they are mostly invisible to the human eye, generally live in the soil and feed on and reproduce at the expense of the roots of plants. These phytophages, normally endowed with a mouth apparatus known in biology as“piercing and sucking”, feed by piercing the roots and in rare cases the stems and leaves, assimilating the lymph present in them and causing more or less severe deterioration of the plant.

Various species of nematodes are also known as carriers of plant diseases of a viral, bacterial or fungal nature, propagated from plant to plant via contaminated saliva or lesions produced on the roots.

Due to their prevalently terricolous habitat and minimal size, the infestations produced by nematodes are often underestimated, resulting in blights or severe damage in plants, whose cause is often apparently inexplicable in the absence of thorough investigations.

Some genera of nematodes, such as, for example Meloidogyne spp. (gall- forming root-knot nematodes), upon piercing the plant roots produce characteristic swellings on them known in agronomy by the name galls, whose greater or lesser presence constitutes a visible and unmistakable symptom of the presence of infestation. In the present state of the art, the fight against nematodes harmful to plants is conducted with both chemical methods and physical methods.

The chemical means for controlling nematodes are represented by products known in agronomy as nematicides and are universally included in the broader category of so-called pesticides, that is, plant protection products or phytosanitary products. Though normally more effective than physical means, the nematicidal phytosanitary products used to date are not devoid of problems, sometimes very serious ones, which greatly limit their practical use, first and foremost their environmental and human toxicity, which is sometimes also extremely high. In this regard, it is noted that out of about 21 active substances officially classified as nematicides and legally usable in Europe in 1986 (M. Muccinelli, Prontuario dei fitofarmaci, Edagricole, Ed. IV), more than half have been banned on toxicological grounds, whereas the remaining ones have been made subject to strong limitations as regards dosages and sectors of application. The largest drawback of newly registered synthetic nematicides as well is that of leaving residues of chemical substances in varying amounts in the soil and on plants, thus contributing to pesticide pollution in the environment and in foodstuffs.

Moreover, the exiguous number of synthetic nematicides present on the market can also be ascribed to the high cost of the investments required to produce the documentation that is necessary for regulatory purposes. The cost in Europe can easily arrive at 100-150 million Euros over a period of 5-8 years, which has discouraged the development of new synthetic active substances.

The few new low environmental impact nematicides present on the market, respectively of natural origin (e.g. garlic extract) or microbiological origin (e.g. Bacillus firmus) show to possess scant effectiveness and thus cannot be used with any real benefits in practice.

As for the known physical methods for controlling nematodes, they are essentially based on the use of natural or artificial heat, concentrated on bare soil suitably covered by plastic sheets, through a technique known as solarisation. This method, despite having a smaller environmental impact and therefore theoretically preferable to chemical treatments, is not devoid of drawbacks, first and foremost its scant effectiveness and excessive impact on crop rotation cycles, since the treatment must be carried out on unplanted bare soil.

There is thus still a strongly felt need in the sector to have synthetic substances which associate a high nematicidal effectiveness with low human toxicity and a low environmental impact.

The Applicant has surprisingly found that polyethylene glycol can be used as an active phytosanitary substance to prevent and/or combat the proliferation of some classes of nematodes, as described below.

Tenuta M. and Ferris H. {Journal of Nematology, 36(1 ):85-94, 2004) have highlighted that persistent nematodes, i.e. nematodes having a high c-p (= colonizer-persister, Bongers, 1990) value, that is, equal to 4 or 5, are subject to osmotic stress. In contrast, the growth of the prevalently colonizer nematodes to which plant parasite groups belong (i.e. which are attributed to groups having c-p values of 1 , 2 or 3) is only slightly influenced or not at all by variations in osmotic pressure. This result was obtained by treating, in a laboratory, specimens of nematodes taken from different soils with aqueous solutions of PEG>8000 at different concentrations, starting from 260 g/l. At the tested concentrations of PEG, the viability of nematodes in the c-p 1 -3 groups, all belonging to the class Secernentea, was not reduced by the treatment any more than it was reduced by a treatment carried out with simple water.

SUMMARY OF THE INVENTION

In a first aspect thereof, the present invention relates to the use of polyethylene glycol as an effective substance to prevent and/or combat the proliferation of phytoparasitic nematodes on a plant and/or rooting substrate of the plant. In a further aspect thereof, the present invention relates to a method to prevent and/or combat the proliferation of phytoparasitic nematodes on a plant and/or in the rooting substrate of said plant, comprising at least one step (a) of distributing 3-100 Kg/ha of polyethylene glycol on the aerial part of at least one plant and/or on the rooting substrate of said at least one plant.

DETAILED DESCRIPTION OF THE INVENTION

In the context of the present invention, the percentages are understood as expressed by weight, unless specified otherwise.

In the context of the present invention,“room temperature” refers to the air temperature in the place in which the use or the method according to the invention is implemented. It is typically comprised between about 4° and about 40°C.

In a first aspect thereof, the present invention relates to the use of polyethylene glycol as an effective substance to prevent and/or combat the proliferation of phytoparasitic nematodes on the aerial part and/or in the rooting substrate of the plant, preferably in the rooting substrate of the plant.

Moreover, the present disclosure relates to the use of polyethylene glycol as a single effective substance to prevent and/or combat the proliferation of the phytoparasitic nematodes on the aerial part and/or in the rooting substrate of the plant, preferably in the rooting substrate of the plant. Polyethylene glycol has revealed to be effective in combating and/or reducing the damage caused to plants by the above-described nematodes, preferably caused to the rooting apparatus of the plants.

Furthermore, glycol has demonstrated to be capable of improving the resistance of plants to the attack of nematodes, preferably the resistance of the rooting apparatus of the plants.

Therefore, the present disclosure relates to the use of polyethylene glycol as an effective substance to combat and/or reduce the damage caused by the nematodes to plants, preferably to the rooting apparatus of plants, and/or to improve the resistance of plants, preferably of the rooting apparatus of the plants, to the attack of nematodes.

The polyethylene glycol and nematodes are as described below in detail. Polyethylene glycol (or PEG) is a polymer of ethylene oxide having the general formula

Depending on the number n of monomeric units, the molecular weight of polyethylene glycol can range from a few hundred daltons, in which case it appears as a colourless fluid liquid, to several thousand daltons, when it takes on the appearance of a more or less viscous liquid, varying in colour from transparent to amber. Polyethylene glycol is generally to be found on the market as a mixture of glycols having different molecular weights.

The polyethylene glycol for use according to the present invention can have a molecular weight of about 400-20,000 daltons, preferably comprised from about 400 to about 7,000 daltons, more preferably from about 400 to about 5,000 daltons.

Low molecular weight PEGs are frequently employed as inert excipients for drugs intended for intramuscular injections, whereas those of a higher molecular weight, in the order of several thousand daltons, constitute the basis for preparing common drugs taken orally. Finally, PEGs of an even higher molecular weight and high density have application as thickeners for syrups, tablet coatings and in the food industry.

As they are not toxic for humans or for the environment, polyethylene glycols can be used for the treatment and/or prevention of nematode infestations of plants without negative environmental impacts and without leaving toxic residues on the parts of the plant intended for human or animal consumption.

Furthermore, since their use as ingredients of pharmaceutical and/or food products is approved at a European level, the procedure for registering the active substance as a plant protection product pursuant to European Regulation EU1 107/2009, Arts. 22 and 23, will be extremely simplified and inexpensive.

The Applicant has surprisingly found that polyethylene glycol can be used as an effective substance to prevent and/or combat the proliferation of phytoparasitic nematodes belonging to the class Secernentea. Preferably, said nematodes can belong to the c-p group (Bongers, 1990) 1 , 2 or 3.

In one embodiment, the polyethylene glycol can be used as an effective substance to prevent and/or combat the proliferation of phytoparasitic nematodes belonging to at least one species of the genus Meloidogyne or to at least one species of the genus Heterodera. The phytoparasitic nematodes belonging to at least one species of the genus Meloidogyne are responsible for root knot and can infest various herbaceous and shrubby plants.

In a further aspect, therefore, the use according to the invention comprises distributing on the plant, preferably on the rooting substrate of the plant, an aqueous composition comprising polyethylene glycol as the sole effective nematicidal substance, i.e. the aqueous composition does not comprise further nematicidal active substances; it preferably does not contain further biocidal active substances, besides polyethylene glycol.

The polyethylene glycol for use as described above can be mixed with (dissolved in) water at a concentration of about 0.1 - 10 Kg every 1000 litres of water, preferably about 1 -10 Kg every 1000 litres of water, i.e. at a concentration of about 0.1 -10 g/l, preferably about 1 -10 g/l.

In one embodiment, the use comprises distributing on the plant, preferably on the rooting substrate of the plant, an aqueous composition comprising about 0.1 -10 g/l, of polyethylene glycol, preferably about 1 -10 g/l. Said aqueous composition is preferably an aqueous solution.

Said aqueous composition can further comprise at least one additive selected from colourants, anti-fermentation agents, olfactory tracers, emetics, surfactants, and mixtures thereof. The nature and amount of additives used can be those normally used in the art for the preparation of phytotherapeutic products.

In one embodiment, the aqueous composition can consist of:

- about 0.1 -10 g/l, preferably about 1 -10 g/l, of polyethylene glycol;

- at least one additive selected from colourants, anti-fermentation agents, olfactory tracers, emetics, surfactants, and mixtures thereof; and

- water.

Water with any degree of purity can be used to prepare the aqueous composition, including mains water and the water normally used for the irrigation of cultivated land. The polyethylene glycol and optionally the at least one further additive can be added to the water at ambient temperature and pressure, under stirring, using apparatus for mixing liquid compounds that are in themselves known.

The polyethylene glycol can be used to prevent the proliferation of the phytoparasitic nematodes and/or to combat the proliferation thereof on a plant and/or in the rooting substrate of a plant, and/or to increase the degree of resistance of the plant to the attacks of nematodes, said rooting substrate preferably being soil.

Phytoparasitic nematodes can affect different vegetative organs of plants, such as roots and, less frequently, stems, leaves, buds, blossoms and fruits. The phytoparasitic nematodes belonging to the group Secernentea, even more preferably belonging to at least one species of the genus Meloidogyne or to at least one species of the genus Heterodera, proliferate in the rooting substrate of the plant, preferably in the soil, and cause damage to the plant roots.

The plant can preferably be a herbaceous or shrubby plant, either a vegetable or ornamental plant, spontaneous or cultivated.

In one embodiment, the polyethylene glycol and the composition described above have demonstrated to be able to reduce the infestation by nematodes belonging to at least one species of the genus Meloidogyne or to at least one species of the genus Heterodera, and/or to combat the proliferation thereof on the plant and/or in the rooting substrate of the plant, and/or to increase the degree of resistance of the plant, in particular tomato plants {Solarium lycopersicum).

As polyethylene glycol has no phytotoxic effects, the use according to the present invention can be repeated whenever it is necessary, either before the plants are sown in the field or throughout the vegetative life of the plants themselves.

In one aspect thereof, the present invention relates to a method for preventing and/or combating the proliferation of phytoparasitic nematodes on a plant and/or in the rooting substrate of said plant and/or for combating and/or reducing the damage caused by nematodes to plants, preferably to the rooting apparatus of plants, and/or for improving the resistance of plants, preferably of the rooting apparatus of plants, to the attack of nematodes, wherein said method comprises at least one step (a) of distributing about 3-100 Kg/ha of polyethylene glycol on the aerial part of at least one plant and/or on the rooting substrate of at least one plant. The method according to the present invention can be implemented on plants in an open field or in a greenhouse, as described above. The amount of polyethylene glycol distributed refers to the portion of treated rooting substrate, on which the effective substance is evenly distributed. The rooting substrate of the at least one plant can be a solid or liquid substrate; it can preferably be a solid substrate, more preferably it can be soil.

The polyethylene glycol can preferably be as described above.

The method according to the invention can preferably be used to prevent and/or combat the proliferation of phytoparasitic nematodes belonging to the class Secernentea. Said nematodes can preferably belong to the c-p group (Bongers, 1990) 1 , 2 or 3.

In one embodiment, the method according to the invention can preferably be used to prevent and/or combat the proliferation of phytoparasitic nematodes belonging to at least one species of the genus Meloidogyne or to at least one species of the genus Heterodera.

The method according to the invention can be used to combat and/or reduce the damage caused to the plant by the nematodes described above and/or to improve the resistance of plants to the attack of nematodes.

The aerial part of the at least one plant can be selected from the stem, leaves, buds, flowers or fruits.

In a preferred embodiment, the method according to the invention can be used to prevent and/or combat the proliferation of phytoparasitic nematodes in the rooting substrate of at least one plant and/or to combat and/or to reduce the damage caused to the rooting apparatus of the plant by nematodes and/or to improve the resistance of the rooting apparatus of the plant to the attack of nematodes. According to this embodiment, the method can comprise at least one step (a) of distributing about 3-100 Kg/ha of polyethylene glycol on the rooting substrate of at least one plant, said rooting substrate being optionally, but preferably, solid, more preferably being soil.

This embodiment is particularly advantageous for preventing and/or combating the proliferation of phytoparasitic nematodes belonging preferably, but not exclusively, to at least one species of the genus Meloidogyne or to at least one species of the genus Heterodera.

In a preferred embodiment, step (a) of the method according to the invention can be implemented using an aqueous composition comprising about 0.1 -10 g/l, preferably about 1 -10 g/l, of polyethylene glycol.

In one embodiment, the aqueous composition can be characterised in that it comprises about 0.1 -10 g/l, preferably about 1 -10 g/l, of polyethylene glycol having the above-described properties as the sole effective nematocidal substance, i.e. the composition does not comprise any further nematocidal active substances, preferably it does not contain any further biocidal active substances, besides polyethylene glycol. In one embodiment, the aqueous composition of step (a) of the method can consist of:

- about 0.1 -10 g/l, preferably about 1 -10 g/l, of polyethylene glycol;

- at least one additive selected from colourants, anti-fermentation agents, olfactory tracers, emetics, surfactants, and mixtures thereof; and

- water.

The aqueous composition can have one or more of the above-described characteristics. Furthermore, said aqueous composition can be placed on the market at the concentration specified above, i.e. ready for use, or in a concentrated form, to be suitably diluted at the time of use.

The at least one step (a) of the method according to the invention can be implemented by distributing, preferably evenly, the polyethylene glycol on the aerial part of at least one plant and/or on the rooting substrate of said at least one plant using apparatus normally employed in the agricultural sector for the localised application of active substances, such as, for example, hoses, drippers, sprayers, pumps, soil injectors, etc.

Given that in the amounts envisaged by the invention there is not expected to be any phytotoxic effect on the plant, much less any effect that is toxic for humans or animals, the at least one step (a) can be repeated when necessary to keep the proliferation of phytoparasitic nematodes under control.

In particular, the at least one step (a) can be implemented:

(i) on the rooting substrate before sowing or planting new plants; and/or

(ii) on the aerial part of at least one nursery plant in its cell before it is transplanted in the field; and/or

(iii) on the aerial part of at least one plant and/or on the rooting substrate of said at least one plant at any time during the vegetative life of the plant. Depending on the severity of the infestation, and in consideration of the low level of hazardousness of the effective substance for humans or the environment, the method according to the invention can advantageously comprise a plurality of steps (a), wherein each step (a) can be independently implemented according to any one of the variants (i), (ii) and (iii).

According to one embodiment, the method according to the invention can comprise a plurality of steps (a), wherein each step (a) is implemented 1 -3 weeks after the preceding step.

The method according to the invention enables the proliferation of phytoparasitic nematodes to be effectively prevented and/or combated, it can be implemented in a simple manner using apparatus normally employed in the art and has a low impact both on humans and the environment.

Furthermore, the method according to the invention makes it possible to combat and/or to reduce the damage caused by nematodes to the aerial part and/or rooting apparatus of plants.

In addition, the method described makes it possible to improve the resistance of the aerial part and/or the rooting apparatus of a plant, in particular of the rooting apparatus of the plant, to the attack of nematodes. The examples that follow have an illustrative character, and do not limit the subject matter of the present invention.

Example 1

Polyethylene glycol with a molecular weight of 400-5000 daltons was mixed with irrigation water at a concentration of 1.0 g/l (1.0 Kg of PEG in 1000 litres of water) and the composition was distributed over soil in which tomato plants were grown, in an amount equal to about 1 kg of PEG every 1000m ² of irrigated surface area.

Fosthiazate, a synthetic nematocide considered to be extremely effective, was used as a positive control (CTR+). The frequency of the treatment with fosthiazate was established according to the provisions of European laws regulating the use of this active substance.

Pure irrigation water was used as a negative control (CTR-).

The protocols for the treatments carried out are shown in table 1 below.

Table 1

One week after the end of the last treatment, the galls produced by nematodes present on the plant roots were checked in order to verify the severity of the nematode infestation.

The treated test plots were divided into four randomised replications and the final result was subjected to statistical analysis. The results of the analysis are shown in Table 2. Table 2

The percentage of galls produced by nematodes subjected to treatment according to the method of the invention (example 1 ) was not significantly different from the percentage of galls produced by nematodes subjected to chemical treatment as per the CTR+ (P<0.05 with the Duncan test).

The number of galls, significantly lower than the number found on the untreated test plot (CTR-), is indicative of the decrease in infestation by nematodes in the test plot of example 1.

Example 2

An alternative evaluation of the number of galls present on the roots of tomato plants treated as indicated in example 1 was carried out using the so-called Bridge & Page scale (1980). The visual presence of galls produced by nematodes on the roots is assessed on this scale, the roots being assigned a sequential value ranging from zero (completely intact root) to 10 (completely necrotized root covered with galls). Conventionally, on herbaceous plants such as tomato, pepper or melon under normal vegetative conditions, it is judged that an index greater than 4.0 may lead to significant reductions in productivity. The results of this evaluation are shown in table 3 below.

Table 3

The value obtained for the roots of plants treated with the method according to the invention was significantly different from the value obtained for untreated plants (CTR-) (P[0.05%] with Duncan’s test).