FI79343C - FOERFARANDE FOER FRAMSTAELLNING AV MIKROBIOLOGISKA BEKAEMPNINGSMEDEL. - Google Patents

Description

1 79343

Method for the preparation of microbiological pesticides

The invention relates to a process for the preparation of biological insecticide and / or fungicide preparations which can be used for controlling plant diseases and / or pests in agriculture.

Several microbes that can be used in the biological control of plant diseases or pests are already known. However, the commercial exploitation of these microorganisms is very problematic because the efficacy of microbial preparations is based on the fact that the microbe in question remains alive in the preparation long enough. For this reason, it would be advantageous to produce spore-containing preparations because spores are known to be much more resistant than vegetative beers or vegetative mycelium. In addition, it is practically necessary for the industrial production of preparations that the products are in dried form, because when dried, the preservation of microbes, and in particular their spores, in preparations is considerably better than in products containing moisture.

Among the microorganisms useful in biological control, there are several that are capable of degrading chitin in the culture medium. Chitin is a white or colorless amorphous polysaccharide that is the major structural component of the backbone of insects, crustaceans, and some other invertebrates. Many of the chitin-degrading microbes have been found to be fungicidal, such as Streptomyces. Trichoderma and Gliocladium fungi and some bacteria. Insect pathogenic fungi are usually also degraders of chitin, due to the fact that they must be able to penetrate the chitin shell of insects. Examples of such fungi are Beauveria bassiana and Metarhizium anisop- ii »a · 2 79343

The addition of chitin and its derivative, chitosan, to plant media has been found per se to control plant disease-causing fungi and nematodes (Agric. Wastes, 4 (1982), p. 205; Phytopathol. 71 (1981), p. 569; U.S. Patent 4,536). 207). The potency is based on the fact that when the culture medium contains chitin, the number of microbes that decompose it in the vicinity of the plant's roots increases, which reduces the chances of plant disease-causing organisms growing due to competition. Chitosan has also been found to trigger certain defense mechanisms against diseases in plants (Plant. Physiol., 66 (1980), p. 205).

It is previously known from U.S. Pat. No. 4,534,965 to use chitin or chitin-containing waste as a component of a plant fungus for the control of Streotomvcea. However, the cultivation is carried out as a submerged culture, i.e. a colloidal chitinous suspension is used as the growth medium for the radial fungus, in which case no stable spores are formed but the radial fungus grows as a vegetative mycelium. The method is therefore based on the observation that when fungi are grown on a chitin-containing medium as a submerged culture, a fungicidal substance produced in the medium is obtained.

Also known from U.S. Patent 4,713,342 is the growth of plant disease or insecticidal fungi on solid media. However, the media used have been those whose sole function is to provide abundant microbial mass and have not contained chitin. Upon application, such media are available to any microbe, including pathogens.

It has now surprisingly been found that the spores of microbes for the control of plant diseases and pests can be efficiently produced on a solid medium containing chitin.

In the method of the invention, the microbial antagonists are grown on a chitin medium having a dry matter content of at least 10% by weight. In this case, the upbringing is the so-called. solidstate breeding on a solid 3 79343 platform. Chitin-degrading, mainly mycelium-like microbes grow on the surface of chitin particles in the same way as in soil and form abundant spores.

To improve growth, potassium and magnesium-containing nutrient salts can be added to the medium. Highly lime-containing soils are usually alkaline. The pH is lowered to a suitable level by adding acid. When the substrate is fully grown, it is dried using either air drying or air drying.

The advantage of the method is that it is neither necessary nor even desirable to separate the microbial spores from the chitin substrate, but chitin also acts as a selective growth medium for antagonists and thus as an active component of the product. In the case of insect control, it is furthermore advantageous that the production of the enzymes required for the penetration of the insect chitin shell is already induced during the growth of the antagonist. Microbes do not produce chitinolytic enzymes unless chitin is present in the medium.

The method according to the invention also has many advantages over known methods for the industrial production of microbiological pesticides. The water content of the product is already ‘low’, so dewatering and drying costs are low.

A suitable chitin-containing waste material 1 i, such as crab shell waste, is suitable as a raw material. Chitin is abundant as a by-product of the fishing industry and its availability is good. Chitin-degrading microbes use it as a source of both carbon and nitrogen.

The invention is described in more detail below by means of examples.

Example 1

Manufacture of Streptomyces spore fungi

The Streotomvces ariseoviridis fungus isolated from peat was grown with partially purified chitin (Sigma 3387).

48 g of saline with a composition of 4 79343 k2hpo4 * 3h2o i.o g / i

MgSO 4 * 7H 2 O> 299 was soaked in 20 g of chitin. The medium was sterilized in an autoclave o (20 min, 121 C) in a 1 liter Erlenmeyer flask. The cooled medium was mixed with 1.65 ml of a spore suspension of the radial fungus and grown at 2 ° C for 7 days. After culturing, the entire medium with spores was lyophilized. The obtained preparation had an amount of spores 6 of about 3 * 10 grams.

Example 2 Biological potency of a radish preparation

The fungal spore-chitin preparation prepared according to Example 1 was ground. A batch of cabbage seeds was contaminated with an Alternaria seedling burner and a part was pickled with a ray mushroom preparation using 15 g of preparation per seed. The seeds were sown in pots on a fertilized peat medium, and 0.5 g / l of the preparation was added to the seed layer on top of the uncovered seeds. The rearing time was 15 days. Seedlings, fresh weight of seedlings and disease were determined from the dishes. Healthy and contaminated cabbage seeds were used as controls. The results are shown in Table 1. It can be seen from the results that the preparation was effective against cabbage seedling burning. Seed pickling gave a better control result than applying the preparation to the seed sowing layer.

5 79343

table 1

Effect of the preparation produced by the method of Example 1 on the cauliflower Alternaria seedling burn.

test vegetation% fresh weight (g) disease index (0-3) 1 92.5 56.6 0.38 2 35.0 29.6 2.51 3 65.6 53.2 0.53 4 60.8 51, 0 0.68 1 * healthy control 2 = seed contaminated with seedlings 3 * contaminated and pickled seeds 4 = contaminated seeds, seedbed treated with preparation disease index: 0 = healthy seedling, 1 = mildly diseased, 2 = diseased, 3 = dead

EelmaEkki 3

Preparation of an air-dried ray spore preparation

Streptomycea ariseoviridis isolated from peat was grown with chitin from crab shell (Sigma 3132). 20 g of chitin was suspended in a saline solution having the composition as in Example 1. The pH of the suspension was adjusted to about 6 with concentrated hydrochloric acid. Excess saline was drained through a glass wool filter. The wet chitin medium was autoclaved in a 1 L Erlenmeyer flask and 1.5 ml of spore suspension of the fungus was poured into the cooled medium. The flask was incubated at room temperature for 7 days, after which the grown medium was applied in a thin layer about 1 cm thick to Petri dishes. The plates were allowed to dry at room temperature for 2 days. The resulting preparation had a fungal spore content of about 10 grams.

6 79343

Manufacture of Trichoderma sprouts

The fungicidal Trichoderma strain was grown with chitin from crab shell (Sigma 3132). 20 g of chitin was dissolved in 200 mL of 1 M hydrochloric acid in about half an hour. The acid was decanted off and the chitin was washed with 3 x 300 ml of the brine mentioned in Example 1. During the last wash, the pH of the suspension was adjusted to about 5 with 10 * KOH. Excess saline was drained through a glass wool filter, the wet chitin medium was autoclaved in a 1 L Erlenmeyer flask, and 1.5 ml of Trichoderma spore suspension was poured into the cooled medium. The flask was incubated at room temperature for 7 days, after which the medium was dried as in Example 3. The resulting preparation had a spore content of 9 to about 10 grams.

Example 5

Gliocladium it manufacture

The fungicidal G1iocladium strain was grown as in Trichoderma in Example 4. The air-dried preparation had a spore-9 content of about 10 grams.

Example 6

Manufacture of Beauveria base sprouts

The insect pathogenic Beauveria bassiana strain was grown with crab shell chitin (Sigma 3132) as well as the ray fungus in Example 3. Both the air-dried and lyophilized preparation had a B. bassiana spore content of about 10 grams.

Monitoring 7

Manufacture of Metarhizium anisopliae sprouts

The insect pathogenic strain Metarhizium anisopliae was grown, such as Trichoderma and Gliocladium in Examples 4 and 5.

9

The air-dried preparation had a spore content of about 10 grams.

Claims (6)

7 79343 A process for the preparation of microbiological insecticides and / or fungicidal preparations, in which the antagonist organism is grown on a solid medium to form spores, after which the medium is dried without separating the antagonist organism or its spores, characterized in that the antagonist organ is chitinolytic and ins. Process according to Claim 1, characterized in that the dry matter content of the chitin-containing medium is at least 10%. Process according to Claim 1, characterized in that the preparation is freeze-dried. Process according to Claim 1, characterized in that the preparation is air-dried. The method of claim 1, characterized in that the antagonist organism is a Streptomyces. Genus Trichoderma or Gliocladium. A method according to claim 1, characterized in that the antagonist organism belongs to the genus Beauveria or Metarhizium.