NZ245276A

NZ245276A - Vesicular arbuscular (va) mycorrhizae inoculant and its preparation

Info

Publication number: NZ245276A
Application number: NZ245276A
Authority: NZ
Inventors: Williams Ingrid Arias De; Jonathan Day
Original assignee: Agricultural Genetics Co
Priority date: 1991-11-28
Filing date: 1992-11-26
Publication date: 1994-04-27
Also published as: KR100239152B1; KR930010176A; TW224930B; AU2963492A; MY108466A; AU649898B2

Description

New Zealand Paient Spedficaiion for Paient Number £45276 Priority Date(s): \ \. 7; K .v.v.fU.

Complete Specific 3iicn F;io-d: Chw . £. OV • • PubHcat-on Date: .^P?.^ P.O. JNo: l?0.% r\n &;S Patents Form No. 5 NEW ZEALAND PATENTS ACT 1953 COMPLETE SPECIFICATION METHOD OF PREPARING VA MYCORRHIZAE INOCULANT WE, AGRICULTURAL GENETICS COMPANY LIMITED, a British company of Church Street, Thriplow, Royston, Herts SG8 7RE, UNITED KINGDOM hereby declare the invention, for which we pray that a patent may be granted to us, and the method by which it is to be performed, to be particularly described in and by the following statements (followed by page la) METHOD OF PREPARING VA MYCORRHIZAE INOCULANT BACKGROUND OF THE INVENTI ON The present invention relates to a method of preparing a VA mycorrhizae inoculant which is useful in the field of agriculture and horticulture and, more precisely, to a method of preparing a VA mycorrhizae inoculant substance having a high VA mycorrhizal spore density and having a well stabilized activity.

VA mycorrhizae (Vesicular Arbuscular mycorrhizae) are known to live together with plants by symbiosis to promote the growth of plants as infected therewith and to improve the disease resistance of the infected plants (N. Kobayashi. Utilization of VA Mycorrhizae to Soil Diseases, Prevention of Plant Diseases. Vol. 42, No. 5, pages 259 to 266, 1988); and cultivation of plants with utilizing such a natural power is desired.

Therefore, various methods of artificially propagating VA mycorrhizae for preparing a VA mycorrhizae inoculant substance have heretofore been proposed.

For instance, there have been proposed a method of separating and recovering spores of VA mycorrhizae as propagated in a soil, blending the spores with a carrier such as vermiculite, attapuigite or diatomaceous earth along with an adhesive substance such as carboxymethyI cellulose, and granulating the In accordance with the former method, however, spores of VA mycorrhizae resulting blend; and a method of using a mixture with charcoal. la. 2 5 JAN 1994 2 O 7 are damaged during the step of separating them or during the step of granulating the spores-containing inoculant. In addition, in the step of granulation, the inoculant is forcedly dried so that many spores would die with considerable frequency. As a result it was difficult to obtain a VA mycorrhizae inoculant of high quality by the method. The same shall apply to the latter method of using a mixture with charcoal, in which many spores would also die during the drying step.

On the other hand, other methods have also been proposed. For instance, there are mentioned a method of using a soi I ; a method of using a porous structure such as a foamed clay or pumice EP 163840, EP 15103) and a method of using a medium soil containing a porous ampho-ion exchanger (US 4,945,059). |n these methods, roots of plants and VA mycorrhizae are made to live together by symbiosis so as to propagate the VA mycorrhizae, and the VA mycorrhizae as adhered to the carrier are directly used as they are as a VA mycorrhizae inoculant.

In these methods, however, where a natural soil is used, there would have a problem of contamination with pathogenic microbes.

With respect to the method of using a porous structure such as a foamed clay or pumice, a high-density VA mycorrhizae inoculant could not be obtained.

In the method of using a medium soil containing a porous ampho-ion exchanger, host plants are limited to only potato and the like plants and the carrier of itself is an expensive one such as DEAE-cellulose. Therefore, the method is not practical.

The object of the present invention is to overcome these drawbacks in 9 r> 24 5 2 7 the above related arts and to provide a method of preparing a VA mycorrhizae inoculant having a high spore density of VA mycorrhizae and having a stabi lized activity of them.

SUMMARY OF THE INVENTION Specifically, the present invention provides a method of preparing a VA mycorrhizae inoculant substance characterized by cultivating plants as infected with VA mycorrhizae of the genus Glomus in a medium containing calcined attapuigite or calcined montomoriI lonite so as to propagate the VA mycorrhizae.

DETAILED DESCRIPTION OF THE INVENTION VA mycorrhizae are a kind of Zygomycetes living in soil, and it is known that the mycelium of them infect into roots of various plants to form mycorrhizal organs therein so that the both live together symbiotically.

VA mycorrhizae to be used in the present invention are those of the genus Glomus.

As VA mycorrhizae of the genus Glomus to be used in the present invention, more concretely, there are mentioned Glomus fasciculatum, Glomus mosseae, Glomus etunicatum, Glomus intraradicies. Glomus manifotis and Glomus caIedon i um.

These VA mycorrhizae of the genus Glomus are collected from the field(Suzuki, T., Problems on VA Mycorrhizae 5, Agriculture and Horticulture, Vol. 62, No. 3, pp. 28 to 33, 1987); or they may be artificially propagated by a nutrient thin film culture method (EP 15103) or by a method of using roots as cultivated by tissue culture (EP 100691) . ! 3 24 H 9 L tc^ra Any of them can be used in the present invention.

In the case that the medium containing calcined montomori Ilonite is usd as the medium, other VA Mycorrhizae than VA Mycorrhizae belonging to the genus Glomus may be used.

As other VA Mycorrhizae than VA Mycorrhizae belonging to the genus Glomus, there are various kinds of VA mycorrhizae.

For example, those are VA Mycorrhizae belonging to the genera Gigaspora, Acaulospora, Entrophospora, Sclerocystis and Scutellospora.

More concretely, those are Gigaspora margarita, Acaulospora laevis, Entrophospora infrequens, Sclerocystis dussii and Scutellospora gregaria.

Plants to be infected with VA mycorrhizae or host plants for propagation of VA mycorrhizae for use in the present invention are not specifically defined provided that they may grow rapidly to have well grown roots and may easily be infected with VA mycorrhizae. For instance, seeds are sowed in a field and cultivated there to plants or the seedlings from them are then transplanted in a different field and are further cultivated there; or plants are cultivated and propagated by vegetable propagation or by cutting or grafting or from bulbs.

Concretely, as host plants for propagation of VA mycorrhizae, there are mentioned true grasses of Gramineae such as corn, crabgrass, sorgo (sweet sorghum or sugar sorghum), wheat, barley and lawn grass; solanous plants of Solanaceae such as eggplant, tomato, pimento (Spanish paprika) and green pepper; leguminous plants of Leguminosae such as soy bean and tare; and liliaceous plants of Li!iaceae such as Welsh onion and common onion.

VA mycorrhizae may be appl ied to the medium before or after rooting of 4 • : ; o the above-mentioned host plants. Infection of the host plants with VA mycorrhizae may be effected by any known method. For instance, the conditions are such that the temperature is from 5 to 60° C, preferably from 10 to 45° C, and the pH of the soil is from 4 to 9.5, preferably from 4.5 to 7.5.

In accordance with the present invention, the above-mentioned host plants as infected with VA mycorrhizae of the genus Glomus are characteristicaI ly cultivated in a medium containing calcined attapuigite or calcined montomoriIlonite.

In the present invention, the host plants as infected with VA mycorrhizae of the genus Glomus may well be cultivated in at least a medium containing calcined attapuigite or calcined montomori Ilonite. If desired, any other medium may be used for infection of host plants with VA mycorrhizae of the genus Glomus.

The calcined attapuigite to be used in the present invention is one as obtained by calcining attapuigite at a temperature of from 200 to 1300° C, preferably from 300 to 1000° C. If desired, calcined attapuigite granules as obtained by granulating a powdery attapuigite along with a binder of alumina or boehmite may also be used. In the case, the calcined attapuigite granules are desired to be adjusted to have pH of from 4.5 to 7.5. The grain size of them may be from 0.25 to 10 mm, preferably from 1 to 5 mm.

On the other hand, the calcined montmoriIlonite to be used in the present invention is one as obtained by calcining montmori Ilonite at a temperature of from 200 to 1300 0 C, preferably from 300 to 1000° C. If desired, calcined montmoriIlonite granules as obtained by granulating a powdery montmoriIlonite along with a binder of alumina or boehmite may also be used. r-~- ft / ' In the case, the calcined montmoriIlonite granules are desired to be adjusted to have pH of from 4.5 to 7.5. The grain size of them may be from 0.25 to 10 mm, preferably from 1 to 5 mm.

In the present invention, the above-mentioned calcined attapuigite or calcined montmori Ilonite may be used singly as the medium, or as the case may be, it may be combined with one or more of any other medium components such as zeolite, pearlite, vermiculite, (calcined) diatomaceous earth and/or pumice.

The proportion of the calcined attapuigite to additional medium components, if any, may be from 1:0 to 1:1 (v/v), preferably from 1:0.1 to 1:1 (v/v), more preferably from 1:1/6 to 1:1/2 (v/v). As the additional medium component, preferred is pumice having a grain size of from 0.5 to 10 mm, preferably from 1 to 5 mm.

The proportion of the calcined montmoriIlonite to additional medium components, if any, may be from 1:0 to 1:1 (v/v), preferably from 1:0.1 to 1:1 (v/v), more preferably from 1:1/8 to 1:1/3 (v/v). As the additional medium component, preferred is pumice having a grain size of from 0.5 to 5 mm, preferably from 1 to 3 mm.

With growth of the host plants, VA mycorrhizae propagate. In general, after the host plants have well grown in 2 to 5 months, irrigation and others thereto is stopped and the thus grown plants are allowed to stand as they are for a while, whereupon VA mycorrhizae form spores. Cultivation of the host plants may be effected under ordinary conditions. For instance, the temperature may be generally from 5 to 60 0 C. If desired, water and fertilizer may be applied to the host plants being cultivated.

Then, the medium to which the thus formed VA mycorrhizal spores have 6 adhered (VA mycorrhizae inoculant) may be recovered. Thus, the intended VA mycorrhizae inoculant is obtained.

In accordance with the method of the present invention, a VA mycorrhizae inoculant having a high VA mycorrhizal spore density and having a stabilized VA mycorrhizal activity can be prepared at low cost.

Accordingly, the method of the present invention can be utilized effectively in various fields of agriculture, horticulture, gardening and nursery.

EXAMPLE Next, the present invention will be explained in more detail by way of the foI lowing examples.

Example 1: A plastic pot No. 8 (diameter 240 mmX height 169 mm) was filled with calcined attapuigite passing through a 3 mm-mesh sieve but not through a 1 mm-mesh sieve (grain size, 1 to 3 mm; firing temperature 540 0 C). In the center of the pot, 320 spores of VA mycorrhiza (Glomus fasciculatum) were embedded at the depth of 3 cm, they being wrapped with tissue paper so that they would not run out downwards along with water. Applicant has a mind to furnish a sample of Glomus fasciculatum to any interested party in response to request for the furnishing of the sample.

Next, two grains of corn (a variety of Golden Dent DK649, by Kaneko Nursery Co.) were put 1 cm above the spores.

Next, the calcined attapuigite montmoriIlonite, VA mycorrhizae and corn seeds in the plastic pot were well wetted with water, and was put in a glass 24 5 27 * green house.

The temperature in the inside of the glass green house in which the pots were put was kept to be from 20 to 25° C, and the plants were cultivated for one week with applying sufficient water thereto. Among the grown seedlings, others except healthy ones were removed. Then, the remaining seedlings were cultivated further in the glass green house with sprinkling water over them every day.

After one month from the beginning of the cultivation, a 1/1000 diluted solution of Peter's liquid fertilizer (N/P/K = 20/10/20) was sprinkled over the plants being cultivated once a week.

The cycle was repeated, and the plants were cultivated for further 2.5 months. Afterwards, sprinkling of water and the liquid fertilizer were stopped, and the plants were allowed to be as they were for 30 days.

Next, the medium pot was turned over a polyvinyl resin sheet and the roots of the corn plants and the calcined attapuigite containing VA mycorrhiza were spread over the sheet. The thick roots of the corn plants were removed and the others were dried as they were at 15° C in the dark.

Five samples, each weighing one gram, of the thus processed medium (calcined attapuigite containing roots of corn plants and VA mycorrhiza) were taken out at random, and the number of the adhered spores was counted in each sample. An average number was obtained from the counted data and was shown in Table 1 below as the number (/g) of spores adhered to the medium.

Example 2: The same process as in Example 1 was repeated except that a mixture comprising 6 parts of calcined attapuigite and one part of pumice was used as 8 the medium in place of calcined attapuigite only. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 1 as the number of spores adhered to the medium.

Comparative Example 1: The same process as in Example 1 was repeated except that a foamed clay (Lecadan as registered trade mark by Lecadan Deutsche I and) as pulverized to have a grain size of from 1 to 3 mm was used as the medium in place of calcined attapuigite. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 1 as the number of spores adhered to the medium.

Comparative Example 2: The same process as in Example 1 was repeated except that amber clay as sterilized with methyl bromide and well degassed to have a grain size of from 2 to 4 mm was used as the medium in place of fired attapuigite. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 1 as the number of spores adhered to the medium.

Example 3: The same process as in Example 1 was repeated except that 40 spores of Glomus caledonium were used in place of 320 spores of Glomus fasciculatum as the VA mycorrhiza. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 2 below as the number of spores adhered to the medium. Applicant has a mind to furnish a sample of Glomus caledonium to any interested party in 9 response to request for the furnishing of the sample.

Example 4: The same process as in Example 1 was repeated except that 40 spores of Glomus mosseae were used in place of 320 spores of Glomus fasciculatum as the VA mycorrhiza. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 2 as the number of spores adhered to the medium. Applicant has a mind to furnish a sample of Glomus mosseae to any interested party in response to request for the furnishing of the sample.

Comparative Example 3: The same process as in Example 1 was repeated except that 40 spores of Scutellospora gregaria were used in place of 320 spores of Glomus fasciculatum as the VA mycorrhiza. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 2 as the number of spores adhered to the medium.

Table 1 Example 1 Example 2 Comparative Example 1 Comparative Example 2 Type of the medium Calcined attapuigite Calcined attapuigite.

Pumice Foamed clay Amber clay VA mycorrhiza Glomus fasciculatum Glomus fasciculatum Glomus fasciculatum Glomus fasciculatum Number (/g) of spores adhered to the medium 87 108 36 44 Table 2 Example 3 Example 4 Comparative Example 3 Type of the medium Calcined attapuigite Calcined attapuigite Calcined attapuigite VA mycorrhiza Glomus caledonium Glomus mosseae Scutellospora gregaria Number (/g) of spores adhered to the medium 61 85 8 Example 5: A long pot No. 5 (diameter 150 mm x height 157 mm) was filled with calcined montmori Ilonite passing through a 3 mm-mesh sieve but not through a 1 mm-mesh sieve (grain size, 1 to 3 mm; calcining temperature 600° C). In the center of the pot, 250 spores of VA mycorrhiza (Glomus fasciculatum) were embedded at the depth of 3 cm, they being wrapped with tissue paper so that they would not run out downwards along with water. Next, two seeds of Sudan grass (a variety of Best Sudan, by Takii Nursery Co. ) were put 1 cm above the spores.

Next, the calcined montmoriIlonite, VA mycorrhizae and Sudan grass seeds in the long pot were well wetted with water, and was put in a glass green house.

The cycle was repeated, and the plants were cultivated for further 2.5 months. Afterwards, sprinkling of water and the liquid fertilizer were stopped, and the plants were allowed to be as they were for 20 days.

Next, the parts above the ground of the thus grown Sudan grass plants ' r 9 7B were cut and removed, and each long pot was turned over a polyvinyl resin sheet and the roots of the Sudan grass plants and the calcined montmori I loni te containing VA mycorrhiza were spread over the sheet. The thick roots of the Sudan grass plants were removed and the others were dried as they were at 15° C in the dark. Five samples, each weighing one gram, of the thus processed medium (calcined montmori I lonite) were taken out at random, and the number of the adhered spores was counted in each sample. An average number was obtained from the counted data and was shown in Table 3 below as the number (/g) of spores adhered to the medium of calcined montmori I lonite.

Example 6: The same process as in Example 5 was repeated except that a mixture comprising 6 parts of calcined montmori I lonite and one part of pumice was used as the medium in place of calcined montmori I loni te only. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 3 as the number of spores adhered to the medium.

Comparative Example 4: The same process as in Example 5 was repeated except that a foamed clay (Lecadan as registered trade mark by Lecadan Deutscheland) as pulverized to have a grain size of from 1 to 3 mm was used as the medium in place of calcined montmori I lonite. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 3 as the number of spores adhered to the medium.

Comparative Example 5: The same process as in Example 5 was repeated except that amber clay as 24 5 2 7 sterilized with methyl bromide and well degassed to have a grain size of from 2 to 4 mm was used as the medium in place of calcined montmoriIlonite. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 3 as the number of spores adhered to the medium.

Example 7: The same process as in Example 5 was repeated except that 40 spores of Glomus mosseae) were used in place of 250 spores of Glomus fasciculatum as the VA mycorrhiza. The number of the spores as adhered to the medium was counted, and an average number was obtained from the counted data and was shown in Table 3 as the number of spores adhered to the medium.

Claims

Table 3 Example 5 Example 6 Comparative Example 4 Comparative Example 5 Example 7 Type of the medium Calcined montmori-1lonite Calcined montmori-1 lonite, Pumice Foamed clay Amber clay Calcined montmori-Ilonite VA mycorrhiza Glomus fasciculatum Glomus fasciculatum Glomus fasciculatum Glomus fasciculatum Glomus mosseae Number (/g of spores adhered to the medium 76 88 41 48 61 0 7 6 WHATV/WE CLAIM IS:-W&KT IS CIS+MiO K :

1. A method of preparing a VA mycorrhizae inoculant characterized by cultivating plants as infected with VA mycorrhizae of the genus Glomus in a medium containing calcined attapuigite or calcined montmori Iloni te so as to propagate the VA mycorrhizae.

2. The method as claimed in claim 1. in which a mixture comprising calcined attapuigite and pumice in a proportion of from 1/0.1 to 1/1 (v/v) is used as the medium.

3. The method as claimed in claim 1. in which a mixture comprising calcined montmoriIlonite and pumice in a proportion of from 1/0.1 to 1/1 (v/v) is used as the medium.

4. The method as claimed in claim 1, in which the VA mycorrhiza is at least one selected from the group consisting of Glomus fasciculatum. Glomus mosseae. Glomus etunicatum. Glomus intraradicies. Glomus manifotis and Glomus caledonium.

5. A method or preparing a VA mycorrhizae inoculant, substantially as herein described with reference to the Examples.

6. Mycorrhizae inoculant when obtained by the method as claimed in any one of the preceding claims. AGRICULTURAL GENETICS COMPANY LIMITED By Their Attorneys BALDWIN SON & CAREY 2 o ;j:J2