Abstract:
The invention relates to the method for producing the fungus  Paecilomyces  spp., and to the uses, methods and nematicide compositions for the prevention and/or control and/or eradication of nematodes that form cysts developing in solanaceae cultures. Said invention includes a system for the application of the fungus combined with the rotation of cultures for effectively and efficiently reducing the viability of the cysts of nematodes in solanaceae cultures.

Description:
FIELD OF THE INVENTION 
       [0001]    This invention relates to a method to obtain the fungus  Paecilomyces  spp. and further relates to the uses, methods and biological compositions functioning as nematicide for the prevention and/or control and/or eradication of cyst-forming nematodes growing onto Solanaceae cultures. 
         [0002]    This invention includes a system to apply said fungus combined with crops rotation, so as to surprisingly decrease in an effective and efficient manner, never known in the state of the art, the viability of the nematode cysts highly harmful to the production of Solanaceae cultures, such as potato, tomato and eggplant, among other. This invention has the advantage of being innocuous both for the product, the soil, and the producer and end consumer, as it does not contribute in the obtainment of an organic product. 
       BACKGROUND OF THE INVENTION 
       [0003]    Nematodes are generally multi-cellular filiform microscopic plant parasites, with reduced diameter ends. Generally, nematodes are dioic (there are males and females) and the reproduction thereof implies a crossed fecundation. As per their biologic cycle this has six stages: a cyst, four juvenile stages (J1, J2, J3 and J4) and adult. Cysts remain into the soil infecting the roots of cultures during the following culture cycle. A nematode may have up to six generations during a culture time depending on the date of seeding, the temperature of the soil, the length of the growing time, the type of host and the geographic zone. Cysts can remain viable for long periods of time of up to twenty years. 
         [0004]    Generally, nematodes, upon infecting cultures, produce aerial damages, such as a reduction in the seeds germination, low production, foliar stains, decay and chlorosity, dwarfism, leaf and stem deformation, internal necrosis in stem, defoliation and lack of greenness and under earth damages such as nodulated roots, detaching and necrosis of the skin on roots, corms and tubercles, low production of seeds and buds or shoots, deformation and necrosis of roots, bulbs, tubercles and corns, excess proliferation of roots, atrophy of radical apexes or an underdeveloped radical system. 
         [0005]    An infestation by  Globodera rostochiensis  ( FIG. 2 ) is produced when larvae J2 burst under the stimuli from radicals exuded out from the plant. Said J2 enter the apical portion of the root, into the epidermal cells and the cellular walls through the stylus thereof, and start feeding from the pericycle cells. Thus, the infested plants become subject to a reduced roots system, rachitis, dwarfism and chlorosis; and due to a reduction in the water absorption the plant may die. 
         [0006]    Golden nematode or  Globodera rostochiensis  is a cyst-forming nematode infecting food and ornamental plants, with million dollars loses in a year. This nematode has affected potato-producing zones back from years 70s and from then it has spread to lukewarm and/or mountain zones where said tubercle is grown. Under a conservative appreciation, this nematode produces In Mexico loses for about 800,000 tons of potato production yearly, equivalent to about 3000 million Mexican pesos yearly. In Veracruz state the loses are esteemed in about 50,000 tons of potatoes yearly, equivalent to about 415 million Mexican pesos yearly, since a statistical lowering in the potato production has been observed in Veracruz state (OEIDRUS, 2008). 
         [0007]    During 2008 a production of 300 million tons of potato was reported in the world over, with the Republic of China being the main provider and consumer thereof. This food shows a high nutritional value, is a versatile and noble food, has a variety of uses of industrial character, such as starch, maltose, dextrose, molasses and glucose. 
         [0008]    According to United Nations&#39; Food and Agriculture Organization (FAO), the world production of potato in 2007 was 325.30 million tons, the main producers being China, Russian Federation, India, United States, Ukraine, Poland, Belarus, Netherlands and France. On the other side, the main consumers of this tubercle are European countries whilst the lower consumers are in Africa and Latin America. 
         [0009]    According to the Secretary of Agriculture, Cattle, Rural Development, Fishing and Food (SAGARPA) potato is ranked in Mexico in the 35th place for seeded surface; however, regarding the production value, this tubercle is ranked in the 9th place. More than 8700 producers are dedicated to potato farming in 12 states of the country, the farming of which generates 17,500 direct employments, 51,600 indirect ones and 6.3 million journals every year (2008). 
         [0010]    In Veracruz state the mountain zones, such as the slopes of Cofre de Perote and Pico de Orizaba, are the ideal places for growing potatoes. Since 1987 the entire producing zone (6000 Ha) of Veracruz state has been under a quarantine according to the official document “Permanent Interior Quarantine Number 17 against the Potato Golden Nematode  Globodera rostochiensis ”, as published in 1987 and in force nowadays, through the Mexican Official Standard NOM040-FITO-2002, wherein requirements and specifications are given for the production and national mobilization of commercial potato, forbidding the use of said tubercle for seeds since this can be a propagator vehicle of nematodes, specifically of  Globodera rostochiensis . This forbidding has been disrespected thus resulting in a greater dispersion of this noxious plague. These rules show that the problem has remained unsolved for more than 30 agricultural cycles. 
         [0011]    Basically, the golden nematode or  Globodera rostochiensis  has affected the Mexican potato-producing zones as back as from about 1972; since it was then when it scattered to the mountain zones where this tubercle is grown. The records of the presence of said parasite show that it reached populations 100 times greater than the recommended value in order to avoid the damage to the production (i.e., 40 cysts per soil kg), according to the European Organization for Plant Protection. 
         [0012]    The most commonly employed methods for controlling  Globodera rostochiensis  are the application of nematicides, such as carbofuran, which, further of being toxic, is not a definitive solution. There are in the state of the art methods for controlling nematodes of the  Globodera  spp. by means of the use of synthetic proteins (WO 01/94601 A2), the use of 2R,5R-dihydroxymethyl-3R, R-dihydroxypirrolidine (WO/1999/05941; a chemical agent obtained from plants of the Solanaceae family (WO/1999/059414); through the use of proteinase inhibitors (WO/1992/015690); and by the use of  Bacillus thuringiensis  (WO/1993/019604); compositions with one or more essential oils from  Tagetes erecta, Ocimum basilicium  or  Cymbopongon martini ; a carrier and an emulsifier (WO/2007/13222); compositions based on the bark of  Afrostyrax lepidephyllus  (WO/1995/010187), addition of cisterna proteinase (WO/2008/087555) and, specifically for the control of  Globodera rostochiensis , the use of  Arthrobotrys  sp. (KR20030094540), which is too much sensitive to ambient factors and is inoperable on de-structured soils; chemical agents obtained from Solanaceae (JP3190807); and a chemical agent of the condensed formula C 27 H 30 O 9  (WO/1993/002083). These control methods, jointly with the strategy of culture rotation, are generally carried out in a disorganized manner and with scarce knowledge on the nematode cycle of life. This and the doses employed, cause pollution and damages to the freatical mantles, pollution of the product and, in some instances, generate resistance on said nematodes and, consequently, making the eradication thereof more difficult. 
         [0013]    The genus  Paecilomyces  spp. has being used in the following applications:  Paecilomyces tenuipes  in the control of flying insects in plants (U.S. Pat. No. 7,033,586 B2, US 20030124098 and US 2007 0141032); a composition containing  Paecilomyces lilacinus  used in the control of soil pests (U.S. Pat. No. 7,435,411 B2, CN101418264 and US 20050008619);  Paecilomyces fumosoroseus  or  Paecilomyces javanicus  in the control of under earth thermites (U.S. Pat. No. 7,390,480);  Paecilomyces javanicus , combined with imidacloprid in the control of sucker insects (CN101502270);  Paecilomyces javanicus  combined with azadirachtin (CN101331880 and CN101331884);  Paecilomyces fumosoroseus  combined with azadirachtin (CN101273728 and WO/1991/011856); a protein obtained from  Paecilomyces farinosus  as an insecticide (WO/2001/000841); and more particularly, in the control of plants parasite nematodes among which there are:  Paecilomyces lilacinus  (U.S. Pat. No. 5,989,54, Pat No. CN101422168, Pat No. DE102005024783, Pat No. CN101081982 and Pat No. CA2059642);  Paecilomyces fumosoroseus  (U.S. Pat. No. 5,360,607);  Paecilomyces lilacinus  251,  Paecilomyces lilacinus  252,  Paecilomyces lilacinus  253 and  Paecilomyces lilacinus  254 (U.S. Pat. No. 5,989,543); and  Paecilomyces cicadae  (CN101518265), without any mention to the use of  Paecilomyces carneus  in the control of nematodes in crops. 
         [0014]    According to the above discussed, and as a response to a problem of such an important market throughout the world as the potato market, this invention has been developed to provide an optimized method of production of  Paecilomyces carneus , as well as uses, methods and compositions easy to be applied in the control and/or prevention and/or eradication of nematodes in Solanaceae, and without a negative effect neither on the soil and on the product or the final consumer. 
       OBJECTS OF THE INVENTION 
       [0015]    This novel invention relates to a fungus  Paecilomyces carneus , in a non-limitative manner, the strain IE-431 of fungus  Paecilomyces carneus ; methods for producing and spreading the same, and the use thereof as a nematicide on Solanaceae plants for the control of nematodes of the Heteroderidae family, in a non-limitative manner,  Globodera rostochiensis.    
         [0016]    Additionally, compositions and methods are described for the control and/or prevention and/or eradication of nematodes in Solanaceae in order to avoid the damage caused on the crop, characterized by the effect thereof starting from the first culture cycle (which is surprising and non-reported in the state of the art) and without affecting the end quality of the product, but improving the biological and chemical characteristics of the soil. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0017]      FIG. 1 . A comparative study of the effect of  Paecilomyces carneus  and  Paecilomyces lilacinus  on  Globodera rostochiensis.    
           [0018]      FIG. 2 . A microphotography of strain IE-431 of  Paecilomyces carneus.    
           [0019]      FIG. 3 . A microphotography of  Globodera rostochiensis.    
           [0020]      FIG. 4 .  Globodera rostochiensis  cysts. 
           [0021]      FIG. 5 . J2 of  Globodera rostochiensis  infected with  Paecilomyces carneus  strain IE-431, inside the egg. 
       
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
       [0022]    This novel invention disclosed herein below comprises uses, methods and biological compositions of  Paecilomyces carneus  for the control and/or prevention and/or eradication of plant parasites in Solanaceae cultures. 
         [0023]      Paecilomyces carneus  strain IE-431 is a fungus produced according to the methods of this invention. Said strain is presently maintained and registered in the fungal collection of the Instituto de Ecologia (INECOL), and is registered with the World Federation Culture Collection (WFCC) under No. IEWDCM782. The isolation and/or maintain of the  Paecilomyces carneus  strain IE-431 are carried out in one or more solid culture means, constituted by at least a nitrogen source and/or at least a carbon source and/or one or more mineral salts and/or at least a suitable vehicle and/or at least an antibiotic agent and/or at least a growing helper agent. Said culture is incubated within a temperature within the range of from 12° C. to 35° C. for a time enough to obtain a pertinent concentration. 
         [0024]    The massive reproduction of the fungus consists in the inoculation of living cells of  Paecilomyces carneus  in liquid media comprising one or more amino acids and/or one or more carbohydrates and/or one or more mineral salts and/or at least one suitable vehicle and/or at least an antibiotic agent and/or at least a growing helper agent. Said culture is incubated at a temperature within a range of from 12° C. to 35° C. for a time enough to obtain a pertinent concentration ( FIG. 3 ). 
         [0025]    Said fungus  Paecilomyces carneus  strain IE-431—obtained from a novel process of isolation and/or maintain and/or massive reproduction disclosed in this novel investigation—, exhibits a series of genetic characteristics that are conserved without representative structural changes, which make it different from any other species of  Paecilomyces  and never observed before with any other species belonging to genus  Paecilomyces  spp., characterized in that presents a colony diameter of from 15 to 21 mm in 10 days on agar-oats (AA) media, of from 15 to 19 mm on Agar-dextrose-potato (PDA), while on Agar-carrot-potato (PZA) it grows from 15 to 17 mm. The mycelium thereof is of white to hyaline color (1A1) in the color chart of Farver and Faver of Wanscher and Kornerup (1961); it does not stain the culture media tested both on the front and reverse of the agar plate. The texture thereof is filamentous accented on PZA; it grows very close to the surface of the media (sparse) and becomes dusty when sporulates. Said mycelium deforms the PDA with the appearance of notorious grooves at the reverse of the box. It is shaped as erect, macronematous, simple conidiophores, with no branches with catenulated equinulated conidia of subglobe shape, and some of them have ends slightly pointed at the ends, dimensioned to 2.4-4 μm×2-3.2 μm. 
         [0026]    In the solid culture medium, 86% of the conidia germinate at 16 hours, the germ tube varies between 6 and 54 micrometers in length. At 24 hrs 100% of the spores germinate and the mycelium is greater than 30 and less than 110 micrometers in length. 
         [0027]    Under the light of the above discussed technical knowledge developed for this invention, we are filing the DNA nucleotide sequence coding  Paecilomyces carneus  strain IE-431, as disclosed at the end hereof. 
         [0028]    In this novel investigation, it was surprisingly found that, when living cells of  Paecilomyces carneus  IE-431 were placed in contact with cysts of the nematode  Globodera rostochiensis , substantial damages were produced on the cysts, such that the propagation of this parasite is avoided. During our investigations, we found that the interaction between  Paecilomyces carneus  and  Globodera rostochiensis  occurs through the fungus germination which, when contacts the cysts ( FIG. 4 ) said cysts are colonized by the mycelium at the interior thereof ( FIG. 5 ) thus surprisingly affecting the whole amount of eggs and larvae J1 and J2 contained within said cyst. The cyst infection starts through an enzymatic process when  Paecilomyces carneus  permeates the cyst wall and then the mycelium grows on the surface of the eggs and the interior of juvenile J1 and J2; therefore, the development of said mycelium becomes invasive to the point of completely degrade the eggs and/or juveniles J1 and J2. 
         [0029]    During the investigation a study was carried out on the nematicide effect of  Paecilomyces carneus  strain IE-431 against the effect of  Paecilomyces lilacinus  IE-430 on the  Globodera rostochiensis  cysts, and It was surprisingly found that  Paecilomyces carneus  IE-431 acts in a more rapid and noxious manner against the cysts of said nematode; i.e., there is a significant difference between to pathogenicity of one species and the other against said nematode. This study was carried to laboratory level, evaluating the degree of infection of said cysts with a concentration of living cells of either fungus of 1×10 6  living cells/mL, and a third control group with the only addition of distilled sterile water. Said pathogenicity was evaluated for a period of 14 days, according to the macroscopic and microscopic aspects of cysts, and a significant unexpected difference was obtained relating the effect of fungus  Paecilomyces carneus  strain IE-431, compared to  Paecilomyces lilacinus . Results are shown in  FIG. 5 . 
         [0030]    The next test stage of the use of fungus  Paecilomyces carneus  strain IE-431 as a nematicide was applied singly (TP) and jointly with lima bean (HP) and peas (ChP) crops rotation, with an absolute witness (TA) where neither control nor crop were applied. Said lima bean and peas crops with no treatment with said fungus, are considered as witnesses (H) and (Ch). When the population density is above 200 cysts/100 g of soil (viability average 10-40 and up to 100 eggs/g soil) the first application is effected before seeding; a second application is made during the seeding and a third application is carried out after the harvest of lima bean and peas in order to provide for a decreasing in the golden nematode population down to practically zero (Table 1 and Table 2). 
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 1 
               
             
             
               
                   
               
               
                 Effect of  Paecilomyces carneus  on the control of 
               
               
                 the potato golden nematode, compared to the lima bean 
               
               
                 crop in rotation and non-farmed plots of land. 
               
             
          
           
               
                   
                 Treatments 
                   
               
             
          
           
               
                 parameters 
                 Monitoring 
                 H 
                 HP 
                 TP 
                 TA 
               
               
                   
               
             
          
           
               
                 Cysts 
                 1 
                 227 
                 272 
                 171 
                 180 
               
               
                 100 g 
                 2 
                 155 
                 144 
                 200 
                 189 
               
               
                   
                 3 
                 140 
                 102 
                 154 
                 183 
               
               
                   
                 4 
                 133 
                 101 
                 153 
                 211 
               
               
                 Viability 
                 1 
                 40 
                 64 
                 13 
                 15 
               
               
                  1 g 
                 2 
                 14 
                 14 
                 20 
                 13 
               
               
                   
                 3 
                 12 
                 7 
                 11 
                 14 
               
               
                   
                 4 
                 15 
                 5 
                 9 
                 15 
               
               
                 J2-Gr 
                 1 
                 4 
                 14 
                 3 
                 4 
               
               
                 100 g 
                 2 
                 7 
                 9 
                 4 
                 3 
               
               
                   
                 3 
                 21 
                 6 
                 12 
                 16 
               
               
                   
                 4 
                 17 
                 2 
                 3 
                 20 
               
               
                 Free-living 
                 1 
                 62 
                 131 
                 119 
                 124 
               
               
                 100 g 
                 2 
                 148 
                 137 
                 78 
                 70 
               
               
                   
                 3 
                 72 
                 82 
                 61 
                 52 
               
               
                   
                 4 
                 42 
                 72 
                 57 
                 43 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
             
               
               
               
             
               
               
               
               
               
               
             
               
               
               
               
               
               
             
           
               
                 TABLE 2 
               
             
             
               
                   
               
               
                 Effect of  Paecilomyces carneus  on the control 
               
               
                 of the potato golden nematode, compared to the peas 
               
               
                 crop in rotation and non-farmed plots of land. 
               
             
          
           
               
                   
                 Treatments 
                   
               
             
          
           
               
                 parameters 
                 Monitoring 
                 cH 
                 chP 
                 TP 
                 TAC 
               
               
                   
               
             
          
           
               
                 Cysts 
                 1 
                 151 
                 173 
                 250 
                 237 
               
               
                 100 g 
                 2 
                 139 
                 134 
                 139 
                 109 
               
               
                   
                 3 
                 146 
                 95 
                 126 
                 130 
               
               
                   
                 4 
                 128 
                 118 
                 124 
                 99 
               
               
                 Viability 
                 1 
                 24 
                 19 
                 17 
                 26 
               
               
                  1 g 
                 2 
                 17 
                 19 
                 15 
                 19 
               
               
                   
                 3 
                 15 
                 13 
                 14 
                 15 
               
               
                   
                 4 
                 8 
                 7 
                 9 
                 10 
               
               
                 J2-Gr 
                 1 
                 5 
                 3 
                 11 
                 10 
               
               
                 100 g 
                 2 
                 1 
                 3 
                 11 
                 8 
               
               
                   
                 3 
                 9.8 
                 13 
                 8 
                 18 
               
               
                   
                 4 
                 6 
                 8 
                 4 
                 16 
               
               
                 Free-living 
                 1 
                 73 
                 105 
                 84 
                 70 
               
               
                 100 g 
                 2 
                 73 
                 101 
                 80 
                 69 
               
               
                   
                 3 
                 61 
                 55 
                 69 
                 72 
               
               
                   
                 4 
                 62 
                 56 
                 115 
                 50 
               
               
                   
               
             
          
         
       
     
         [0031]    As can be seen from the above,  Paecilomyces carneus  strain IE-341 has a nematicide effect on the viability of the potato golden nematode cysts but no action on free life nematodes, which are to be kept on farming soils. Under this investigation, we found a synergic behavior for reducing the population density of golden nematode when the application of  Paecilomyces carneus  is combined with the rotation of cultures that are not favorable to the bursting of J2  Globodera rostochiensis.    
         [0032]    It is to be said that the use of rotation with lima bean and peas without the application of  Paecilomyces carneus  as a bionematicide is also effective, but requires of a period of time greater than 4 years. 
         [0033]    Following an exhaustive investigation, a method is provided for isolation, maintaining, massive reproduction of  Paecilomyces carneus  strain IE-431, as well as the use thereof in the control and/or prevention and/or eradication of cyst-forming nematodes or having characteristics similar to  Globodera rostochiensis  on cultures of the Solanaceae family, such as potato, chili, tomato, eggplant, among other plants where this type of parasite grows, such as  Avena sativa  and  Vicia villosa  (Winter Veza), and tobacco. 
         [0034]    Further to the products of this invention, nematicide compositions are provided comprising living cells of  Paecilomyces carneus  strain IE-431 and at least a living cells carrier and optionally a device for the application of said nematicide. The composition preferred for application to farmings is in a suspension shape; however, these compositions can be developed and applied in the shape of pellets, powder, capsules, emulsion, microemulsion, solution or any other form of application existing in the state of the art. 
       EXAMPLES 
       [0035]    Herein below, intended to be descriptive rather than restrictive, are provided compositions employed for the propagation of fungus  Paecilomyces carneus  strain IE-431 and the nematicide compositions obtained. 
       Example 1 
     Composition A for Mass Propagation of  Paecilomyces Carneus  Strain IE-431 
       [0036]      
         [0000]    
       
         
               
               
               
             
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 Carrot juice 
                 80-100 
                 mL 
               
               
                   
                 Ampicilina 
                 500 
                 mg 
               
               
                   
                 Water 
                 q.s. 1000 
                 mL 
               
               
                   
                   
               
             
          
         
       
     
       Example 2 
     Composition B for Mass Propagation of  Paecilomyces Carneus  Strain EI-431 
       [0037]      
         [0000]    
       
         
               
               
               
             
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 Oats 
                 20-50 
                 gL 
               
               
                   
                 yeast 
                 0.1-5.0 
                 g/l 
               
               
                   
                 Carrot juice 
                 50-150 
                 mL7L 
               
               
                   
                 Chloramphenicol 
                 1000 
                 mg 
               
               
                   
                 Water 
                 q.s. 1000 
                 mL 
               
               
                   
                   
               
             
          
         
       
     
       Example 3 
     Composition C for Isolation and Maintain of  Paecilomyces Carneus  Strain IE-431 
       [0038]      
         [0000]    
       
         
               
               
               
             
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 Carrot 
                 50-160 
                 g/L 
               
               
                   
                 Oats 
                 20-50 
                 g/L 
               
               
                   
                 Chloramphenicol 
                 1000 
                 mg 
               
               
                   
                 Water 
                 c.b.p. 1000 
                 mL 
               
               
                   
                   
               
             
          
         
       
     
       Example 4 
     Nematicide Composition A-1 
       [0039]      
         [0000]    
       
         
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
                 Living cells of  P. carneus   
                 0.5 × 10 7 /mL 
               
               
                   
                 Composition A 
                 1000 mL 
               
               
                   
                   
               
             
          
         
       
     
       Example 5 
     Nematicide Composition B1 
       [0040]      
         [0000]    
       
         
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
                 Living cells of  P. carneus   
                 3 × 10 7 /mL 
               
               
                   
                 Composition B 
                 1000 mL 
               
               
                   
                   
               
             
          
         
       
     
       Example 6 
     Nematicide Composition C-1 
       [0041]      
         [0000]    
       
         
               
               
               
             
           
               
                   
                   
               
               
                   
                 INGREDIENT 
                 AMOUNT 
               
               
                   
                   
               
             
             
               
                   
                 Living cells of  P. carneus   
                 0.3 × 10 7 /mL 
               
               
                   
                 Composition C 
                 1000 mL 
               
               
                   
                   
               
             
          
         
       
     
         [0042]    In an integral manner, this invention provides the following advantages: 
         [0043]    1.—A method of agricultural control, based on clean and supporting technologies. 
         [0044]    2.—Said method produces an improvement on the soil quality since allows for the recovery of soils used in Solanaceae cultures. 
         [0045]    3.—The time to eradicate nematodes is one cycle, whereby it may stop the permanent interior quarantine number 17 against the potato golden nematode  Globodera rostochiensis.    
         [0046]    4.—An immediate improvement is produced on the quality of the obtained product, enlarging the shelf life thereof. 
         [0047]    5.—The nematicide compositions obtained are innocuous for both humans and animals. 
         [0048]    6.—The population of the golden nematode is reduced from the very first application. 
         [0049]    7.—The cysts are directly attacked whereby better agricultural products are obtained from the very first application. 
         [0050]    8.—It can be combined with other control methods, such as rotation.