TWI405744B - Candida albicans Candida ethanolica ) CC-DH2011 and its formulations and uses - Google Patents

Abstract

The invention relates to a strain, which is separated, purified and cultured, that is Candida ethanolica CC-DH201. The strain is separated and sieved from fermentation disposal solution. After purifying and culturing, the strain is identified to have multifunctional activities of phosphate solubilizing, aluminum-phosphorous, and ferrite-phosphorous through the activity test of solid sieved culture medium. Further, the strain is also verified to be a safe and non-known pathogenic germ and analyzed with features of enzyme activities and carbon source utilities, with the pot experiments, and the strain can be applied in developing and utilizing microorganism fertilizer and organic waste treatment.

Description

Candida ethanolica CC-DH2011 and its preparation and use

The present invention relates to a Candida ethanolica CC-DH2011 having a multifunctional activity such as calcium phosphate, aluminum phosphorus and iron phosphorus, and a preparation and use thereof.

According to the traditional use of chemical fertilizers in farmland, it can quickly provide the nutrients needed for crops, but it will lead to problems of soil acidification, uneven fertility and groundwater pollution. It is known that soil microbes can be used as microbial fertilizers if they have nitrogen fixation, dissolution of ineffective nutrients, and decomposition of organic matter in the decomposed soil. Microbial fertilizers can replace some chemical fertilizers to solve the long-term application of chemical fertilizers. Problems such as soil degradation. Plant growth promoting rhizobacteria (PGPR) has four mechanisms according to its action mechanism: (1) biofertilization, (2) biocontrol, and (3) bioremediation. (4) phytostimulation.

It can be seen from the above that conventional microbial fertilizers can replace some chemical fertilizers, but in the application of actual microorganisms, due to the use of a plurality of single-function bacterial agents, various strains may compete with each other or cause antagonistic effects. In order to reduce the effect of microbial fertilizers, it is obvious that the incorporation of a plurality of single-function microbial agents has its inconvenience and its absence, and needs to be improved. The reason is that the creator feels that the above can be improved. He is dedicated to research and cooperate with the application of theory, and finally proposes a yeast that is reasonable and can be multifunctional.

The taxonomic status of Candida ethanolica is: phylum is called Ascomycota, Saccharomycotina, class is called Saccharomycetes, and order is yeast (Saccharomycetales) ), the family name is mitosporic Saccharomycetaceae, and the genus is genus Candida .

Although yeast has long been widely used in winemaking and baking, the function of Candida ethanolica in agricultural soil or plant growth promotion is still unclear. However, it has been pointed out that Candida valida, Rhodotorula glutinis and Trichosporon asahii have promoted fields. Vegetable crop growth and ability to inhibit the growth of pathogenic bacteria ( Rizoctonia solani AG-2-2) (El-Tarabily, 2004). Up to now, the strain of yeast C. ethanolica has not been patented in Taiwan, but there are one patents in foreign countries. As shown in Table 1, the patent only relates to biomass. The strain of the present invention was confirmed to be a multi-functional yeast strain through a multifunctional test, and a patent application was filed according to law.

The main object of the present invention is to provide a Candida ethanolica CC-DH2011 which has multifunctional activities such as calcium phosphate, aluminum phosphorus and iron phosphorus, and is now deposited in the food industry development of Hsinchu, Taiwan. Institute, registration number is BCRC920070.

A second object of the present invention is to provide a Candida ethanolica CC-DH2011 which is a multi-functional yeast strain for developing a multi-functional yeast microbial agent, and It can respond to the multiple functions of the microbial agents themselves in response to complex loops.

For the purpose of the foregoing, the present invention provides a strain which is isolated and purified and cultured, which is Candida ethanolica CC-DH2011, which is a strain isolated from the fermentation waste liquid and purified. After the culture, the activity of the solid screening medium was tested, and it was found that the bacteria had multifunctional activities such as calcium phosphate, aluminum phosphorus and iron phosphorus, and the strain was confirmed to be a non-known pathogen of safety, and the enzyme activity and carbon were analyzed. The characteristics of the source utilization and the verification effect of the pot experiment can be applied to the development and utilization of microbial fertilizers and organic waste treatment.

The above and other objects and advantages of the present invention will be readily understood from

The present invention relates to Candida ethanolica CC-DH2011 having functional activities such as dissolved iron phosphorus, calcium phosphate and aluminum phosphorus, which is stored in the food industry of Hsinchu, Taiwan, on January 10, 1998. The Development Institute, the registration number is BCRC920070, and was re-invested by the Food Industry Development Research Institute on February 20 of the same year and confirmed that the test was alive (please refer to Annex I and Annex II).

The basic characteristics of the Candida ethanolica CC-DH2011 strain of the present invention are as follows:

1. Characteristics of culture and morphology

The strain of the present invention is a yeast species which grows in a group of round or elliptical single cells, eukaryotes having a nucleus. In the nutrient ager (NA) culture, the colony morphology is similar to that of bacteria, but it is thicker and thicker, has a milky white shape, and has a rough surface and propagates in a budding manner.

2. Special characteristics of culture traits

The growth temperature of the yeast strain of the present invention is between 17 and 45 ° C, the optimum temperature is 30 to 40 ° C, the growth pH is 2 to 10, and the optimum growth pH is 3 to 5.

3. The physiological characteristics of the strain are as follows:

The yeast strain of the present invention has the following organic decomposition enzyme activities: alkaline phosphatase, butyrate esterase (C4), caprylate esterase (C8), lipase (lipase) ), leucine arylamidase, valine arylamidase, cystine arylamidase, acid phosphatase, Naphthol-AS-BI-phosphohydrolase is reactive.

The Candida ethanolica CC-DH2011 strain of the present invention includes the following characteristics:

1. The strain of the present invention is a natural strain of the self-fermented waste liquid without any artificial genetic modification.

2. The strain of the present invention has a multi-functional activity yeast species such as iron phosphate, calcium phosphate and aluminum phosphorus.

3. The genus of the invention belongs to the National Institutes of Health (NIH) Guideline biohazard classification and is not a known human pathogen.

Based on the above characteristics, the yeast can be developed as a microbial agent for use in microbial fertilizers and waste treatment.

The Candida ethanolica CC-DH2011 strain of the invention has the characteristics of being versatile such as iron-dissolving phosphorus, calcium phosphorus and aluminum phosphorus, and can be grown in organic substances of different materials to make the bacteria body. It is in full contact with the organic matter, and thus the organic substance is decomposed as a carbon source. In addition, the bacteria can also survive in the fermentation waste liquid, and can be used for the application of the bio-technical fermentation waste liquid, and has a practical function. Therefore, the multi-functional strain of the invention can be applied to agricultural biological fertilizers, which not only enhances the hydrolysis efficiency of microorganisms to different organic substances, but also overcomes the mixed different strains encountered when using various microorganisms which are conventionally mixed with a single function for use in microbial fertilizers. Since the "antagonism" between the two, the multifunctional Candida ethanolica CC-DH2011 proposed by the present invention has a higher efficiency than a conventional single function mixing a plurality of microbial fertilizers.

The present invention also encompasses the combination of the yeast C. ethanolica CC-DH2011, which is used in biotechnology waste liquids, including waste materials such as animals and plants and waste materials from food processing industries. The combined substance may further contain other microbial agents capable of decomposing organic substances and the like to enhance the function of organic decomposition. On the other hand, the composition of matter optionally may further comprise a yeast culture for C. ethanolica CC-DH2011 culture or stabilizers, present in order to facilitate growth of the yeast C. ethanolica CC-DH2011 of propagation.

According to the invention, treatment with a composition which is available for growth of the yeast C. ethanolica CC-DH2011 can be used. In a specific embodiment of the present invention, the yeast C. ethanolica CC-DH2011 is firstly cultured in a liquid medium, and the prepared microbial agent is inoculated into the root zone soil of the crop, and directly used as a microbial fertilizer of the crop.

According to a preferred embodiment of the present invention, the culture of the cultured yeast C. ethanolica CC-DH2011 has a temperature range of 17 to 47 ° C, an optimum temperature of 30 to 40 ° C, a growth pH of 2 to 10, and an optimum growth pH. It is 3 to 5.

The present invention is described in detail in the following description of the invention, and is not intended to be construed as limited.

Isolation and screening of multifunctional yeast strains

The strain isolated from the fermentation waste liquid by continuous dilution and smear method is first separated and purified by the nutrient agar medium (NA; nutrient broth), and the multi-functional characteristics are separated and screened as follows:

1. Determination of the ability to dissolve calcium and phosphorus: The yeast C. ethanolica CC-DH2011 strain was subcultured on calcium phosphate medium with the following formula: Sucrose 10.0 g/L, tricalcium phosphate (Ca ₃ (PO ₄ ) ₂ ) 4.0 g / L, ammonium nitrate (NH ₄ NO ₃ ) 0.27 g / L, potassium chloride (KCl) 0.2 g / L, magnesium sulfate (MgSO ₄ .7H ₂ O) 0.1 g / L, yeast extract (Yeast) Extract) 0.1 g/L, manganese sulfate (MnSO ₄ .6H ₂ 0) 0.001 g/L, iron sulfate (FeSO ₄ .7H ₂ O) 0.001 g/L, agar 15.0 g/L, pH 6.8. The growth was carried out at 30 ° C in an incubator, and the colony growth on the calcium phosphate medium was observed after the 5th day, and the ability to dissolve the tricalcium phosphate was 1.79 (the ratio of the transparent dissolution ring/colony diameter). As shown in Annex III, the strain of the present invention is dissolved in a medium of tricalcium phosphate, aluminum phosphate or iron phosphate.

2. Determination of the ability to dissolve aluminum and phosphorus: The yeast C. ethanolica CC-DH2011 strain was subcultured on the aluminum phosphate medium, and the formula was as follows: sucrose: 10.0 g/L, aluminum phosphate (AlPO ₄ ) 4.0 g/L, ammonium nitrate (NH ₄ NO ₃ ) 0.27 g/L, potassium chloride (KCl) 0.2 g/L, magnesium sulfate (MgSO ₄ .7H ₂ O) 0.1 g/L, yeast extract (Yeast extract) 0.1 g/L, manganese sulfate (MnSO ₄ .6H ₂ 0) 0.001 g/L, iron sulfate (FeSO ₄ .7H ₂ O) 0.001 g/L, agar 15.0 g/L, pH 6.8. The growth was carried out at 30 ° C in an incubator, and the colony growth on the iron-phosphorus medium was observed after the 13th day, and the aluminum phosphate-dissolving power was measured to be 1.22 (transparent melting circle/colony diameter ratio). As shown in Annex III, the strain of the present invention is dissolved in a medium of tricalcium phosphate, aluminum phosphate or iron phosphate.

3. Determination of the ability to dissolve iron and phosphorus: The yeast C. ethanolica CC-DH2011 strain was subcultured on iron-phosphorus medium, and its formula was: sucrose (Sucrose) 10.0 g / L, iron phosphate (FePO ₄ ) 4.0 g/L, ammonium nitrate (NH ₄ NO ₃ ) 0.27 g/L, potassium chloride (KCl) 0.2 g/L, magnesium sulfate (MgSO ₄ .7H ₂ O) 0.1 g/L, yeast extract (Yeast extract) 0.1 g/L, manganese sulfate (MnSO ₄ .6H ₂ 0) 0.001 g/L, iron sulfate (FeSO ₄ .7H ₂ O) 0.001 g/L, agar 15.0 g/L, pH 6.8. The growth was carried out at 30 ° C in an incubator, and the colony growth on the aluminum phosphate medium was observed after the 15th day, and the capacity of the dissolved iron phosphate was determined to be 1.09 (the ratio of the transparent dissolution ring/colony diameter). As shown in Annex III, the strain of the present invention is dissolved in a medium of tricalcium phosphate, aluminum phosphate or iron phosphate.

Identification of the yeast CC-DH2011 strain of the present invention by molecular biotechnology

A. Culture of the bacterial liquid: The yeast strain of the purified CC-DH2011 was cultured therein using a nutrient broth (NB) having a sufficient nutrient as a culture medium.

B. DNA extraction and PCR reaction: DNA was extracted according to the method of the laboratory, and the polymerase chain reaction (polymerase) was carried out according to the 18S rDNA sequence published by Cheng et al. (2004) and the 26S rDNA sequence published by Kurtzman and Robnett (1997). The chain reaction (PCR) technique amplifies the 18S rDNA and 26S rDNA of the yeast, and analyzes the obtained DNA by sequence analysis. The thermal cycle of the 18S rDNA gene fragment (Flexigene, PCR) was performed with specific primers NS1, NS5R and NS8 (Table 2). Thermocycler), proliferation conditions were 94 ° C for 4 minutes 1 cycle, 94 ° C 1 minute 52 ° C 2 minutes 72 ° C 2 minutes 35 cycles, 72 ° C 10 minutes 1 cycle. The thermostable reaction of the 26S rDNA gene fragment was carried out with specific primers NL1 and NL4 (Table 3) under the conditions of 94 ° C for 5 minutes, 1 cycle, 94 ° C for 1 minute, 52 ° C for 2 minutes, 72 ° C for 2 minutes, 36 cycles. The reaction product was analyzed by 1% TBE agarose colloidal electrophoresis.

C. 18S and 26S rDNA sequencing and identification: NS1, NS5R and NS8 were 18S rNDA primers (Table 2), and NL1 was 26S rNDA primers (Table 3), and were performed using ABI PRISM 310 Genetic Analyzer (Perkin-Elmer). Sequence reaction, the obtained sequences are compared with the known strain sequences in the GenBank database of the National Center for Bioinformatics (NCBI), and the results obtained are the scientific names of the strains. Based on the results of the 18S rDNA sequencing, the CC-DH2011 strain was 100% similar to Candida ethanolica , and the similarity to Pichia deserticola was 99.88% (Table 6). Using the 26S rDNA sequencing results, the CC-DH2011 strain was similar to Candida ethanolica and Pichia deserticola by 99.82% (Table 7). It can be confirmed that the CC-DH2011 strain of the present invention is the yeast Candida ethanolic . According to the scholars, Candida ethanolica and Pichia deserticola are classified into the same cluster, and the taxonomic status is very close (Kurtzman et al ., 2008). Although it is a different genus, it may be classified as the same species in the future.

Growth pattern of bacteria

The strain of the present invention is a yeast species which grows in a group of round or elliptical single cells, eukaryotes having a nucleus. After incubation at 30 ° C for 48 hours, visible (about 2 mm) milky white colonies were formed. Growth was not observed at more than 47 ° C, while small colonies were observed at 15 ° C for extended incubation, and obvious colonies were observed at 20 ° C for 72 hours. In the nutrient ager (NA) culture growth, the colony morphology is similar to that of bacteria, protruding and thicker, milky white shape, rough surface, and propagated by budding (as shown in Annex IV, the strain of the present invention The colony morphology of the growth was cultured in nutrient ager (NA).

Biological analysis of bacteria

Take 1 ml of different dilutions (100X, 500X 1000X, 2000X and 3000X) of the yeast C. ethanolica CC-DH2011 strain onto the crepe paper, and contain it in the sensitive plant lettuce seed ( Lactuca sative var. Salina). The performance of biologically active substances. Soak the lettuce seeds in water for about 30 minutes, carefully place the seeds in a Petri dish with a built-in crepe paper (70 mm Whatman No. 4) and evenly sow them on the edge of the plastic Petri dish with 12 seeds per dish. Thereafter, it was placed in the dark at about 28 ° C, and after germination for 72 hours, the growth of the radicle and the embryo stem was measured, and the culture solution of distilled water and the propagation strain was used as a control.

The results showed that the bacterial solution of the yeast C. ethanolica CC-DH2011 strain promoted the growth of radicle and embryo stems in a 1000-fold dilution (Table 8, Table 9).

Analysis of the carbon source utilization capacity of the yeast C. ethanolica CC-DH2011 strain by BIOLOG

The yeast C. ethanolica CC-DH2011 strain was visually inoculated into a BUY Agar (Biology Universal Yeast Growth Agar medium; BIOLOG USA) petri dish. Incubate at 30 ° C, pick a single colony to BUY agar medium, incubate at 30 ° C for 24 hours, pick up the colony with a sterile cotton rod, suspend and culture in GN / GP-1F inoculant, adjust the bacterial solution to a wavelength of 590 NM Transmittance 65%, take 150μl of the adjusted suspension into the YT MicroPlate, and then incubate in the growth chamber for 3 days and then use the Biolog MicroStation Reader to interpret the OD value of the non-dark color reaction in each well of the microplate. (λ1=590, λ2=750) Subtracting the value of A-1 hole (blank test, no carbon source), it can be divided into three types: strong reaction, weak reaction and no reaction.

The yeast C. ethanolica CC-DH2011 strain of the present invention was analyzed by the software analysis of the MicroStationTM system provided by BIOLOG, and the similarity of the phenotype to Pichia membranaefaciens was 0.84, and the probability of 0.99 was 0.99. The case where the strain of the strain of the present invention utilizes a carbon source is shown in Table 10, and it is shown that among the 95 kinds of carbon sources tested, there are a total of 27 kinds of positive reaction, among which (1) 10 kinds of sugars: maltotriose (maltotriose), α-D-glucose, dextrin, cellobiose, D-melibiose, D-galactose , D-psicose, L-sorbose, inulin; (2) 5 kinds of organic acids: fumaric acid, L-malic acid (L-malic acid), bromo succinic acid, acetic acid, L-proline; (3) 2 kinds of amino acids: N-acetyl-D- N-acetyl-D-glucosamine, D-glucosamine; (4) 2 kinds of aromatic compounds: Salicin, amygdalin; (5) There are three kinds of alcohols: glycerol, D-mannitol, D-arabitol; (6) two kinds of esters: dimethyl succinate Carbon sources such as methyl succinate and methyl succinate (D-xylose) are reactive.

Determination of the yeast C. ethanolica CC-DH2011 strain by API-ZYM Physiological properties

The specific enzyme activity of the yeast strain C. ethanolica CC-DH2011 of the present invention was tested by API-ZYM. First, the bacteria on the culture dish were inoculated by inoculation, and dissolved in 5 ml of sterile water. The adjusted bacterial concentration makes the transmittance close to 50%. 65 μl of the adjusted bacterial solution was added to each reaction well. The sample was placed in a reaction at 30 ° C for 4 hours. Then add one drop of each of API-ZYM A solution and B solution, and let stand for 5 minutes to observe the color change. The measurement results are shown in Table 11. Whether the strain has a reaction or no reaction activity is indicated by a + or - symbol in the table. According to API-ZYM analysis, C. ethanolica CC-DH2011 strain has alkaline phosphatase, butyrate esterase (C4), caprylate esterase (C8), and fat. Lipase, leucine arylamidase, valine arylamidase, cystine arylamidase, acid phosphatase Phosphatase, naphthol-AS-BI-phosphohydrolase is reactive, but for trypsin, tryhypeptin, and α-galactosidase (α- Galactosidase), β-galactosidase, β-glucuronidase, α-Glucosidase, β-Glucosidase, N - N- acetyl-β-glucosaminidase, α-mannosidase (α-Mannosidase), and α-fucosidase (α-Fucosidase) are not reactive.

Multifunctional test of the yeast C. ethanolica CC-DH2011 strain

The versatile performance of the strain yeast C. ethanolica CC-DH2011 was tested by the aforementioned method, and the strain had three multifunctional activities such as calcium phosphate, aluminum phosphorus and iron phosphorus. The strain is applied to the soil, and the ineffective phosphate fertilizer in the soil can be converted into the effective phosphate fertilizer by the metabolism of the yeast strain C.ethanolica CC-DH2011. The phosphates (calcium phosphate, aluminum phosphate and iron phosphate) which are ineffective in the soil can be converted into effective phosphate fertilizer, so that the application of the phosphate fertilizer can be reduced, and the application of the phosphate fertilizer can be saved for the crop growers. The cost of chemical fertilizers can increase the beneficial microbes in the soil, thereby increasing the agglomeration and soil strength of the soil.

The strain of the invention has three functions of calcium phosphate, aluminum phosphorus and iron phosphorus. Therefore, adding the composition of the strain of the present invention to produce a biological fertilizer can play a plurality of functions: (1) decomposition of organic matter to produce an acid, which can dissolve nutrients that are not easily dissolved, thereby improving the effectiveness of nutrients; and (2) releasing the product after mineralization. Nutrient for plant absorption; (3) A variety of organic carbon sources can be used as a matrix for microbial proliferation, which is conducive to colonization of microorganisms; (4) The present invention can utilize a variety of amino acids, and thus can utilize amine groups. Acid is used as a bacterial cell for reproduction. Therefore, the yeast C. ethanolica CC-DH2011 strain of the present invention has the function of calcium and phosphorus, and has the functions of dissolving aluminum phosphorus and iron phosphorus, and is applied to crop roots for the function of biofertilization of crops. It can be directly or indirectly used in related industries such as microbial fertilizer, biological control and organic matter decomposition.

The medium used in the strain of the present invention has the following characteristics: (1) Activity of amino acid utilization: analysis of the activity of the strain of the present invention having several amino acids by BIOLOG, which avoids feedback caused by amino acid of protein decomposition product The feedback inhibit-ion is beneficial to the continuous action of decomposition. (2) Amylolytic activity: The strain of the present invention showed reactivity to dextrin by BIOLOG, indicating that the strain of the present invention has amylase activity of cleaving α-1,6-glycosidic bonds. (3) Utilization of glucose: According to BIOLOG analysis, the glucose of the product of the present invention for the decomposition of starch is known, which can also facilitate the reaction by avoiding the effect of feedback inhibition.

In summary, the preliminary test for the activity of the strain in the test medium is to use the microbial characteristics of the specific substrate as a basis for a large number of screening, and compared with the commercially available test kits (such as BIOLOG, API-ZYM), the study was found. The solid screening method adopted has incomplete substitution, and has both physical and industrial applicability. Therefore, the strain of the present invention selects a suitable solid medium as a multi-functional test for the strain and a method for separation and screening.

Pot experiment results and multi-functional effects

In order to understand the effect of the bacterial agent of the present invention ( C. ethanolica ) CC-DH2011, a pot experiment was designed to carry out the following two items:

1. Qingjiang cuisine test: Bok Coy's scientific name is Brassica chinensis L.CV.Ching-Geeng. The test period was from December 5, 2007 to January 2, the next year. The test site was at the National Chung Hsing University Greenhouse. The test treatment uses a Randomized Complete Block Design (RCBD). This test uses 5 吋 plastic pots and 1 kg of soil per pot. The test treatment is as follows: (1) CK: control group, (2) the yeast CC-DH2011: application 10 ml / pot of yeast CC-DH2011 bacterial solution (10-fold dilution), (3) CF: application recommendation The total amount of Taiwan compost compound fertilizer No. 1 (0.15g / pot) and (4) 1/2 of the yeast CC-DH2011 + 1/2CF: half of the CC-DH2011 bacterial solution (5ml / pot) and semi-quantitative fertilizer ( 0.075 g / pot), each treatment was carried out in 4 replicates.

The growth situation after 4 weeks of treatment in this experiment is shown in Annex V. After 4 weeks, the fresh weight and dry weight were harvested and weighed. The Duncan's multiple range test was used to check whether there was any difference (p=0.005). Table 12, Figure 1, and Figure 2 show. The fresh weight and dry weight of Qingjiang cuisine are shown in the bacterial liquid containing the yeast strain C.ethanolica CC-DH2011 of the present invention, the semi-quantitative fertilizer containing half of the bacterial liquid (1/2 CC-DH2011+1/2CF) and the whole quantitative fertilizer The (CF) treatment group was superior to the control treatment group, and the fresh weight and dry weight of the semi-quantitative fertilizer (1/2 CC-DH2011+1/2CF) added to the semi-quantitative liquid were better than the full-quantity fertilizer (CF) treatment. It is shown that the C. ethanolica CC-DH2011 strain is beneficial to the growth of Qingjiang cuisine. If combined with chemical fertilizers and can reduce the application amount of 1/2 chemical fertilizer, it shows that the yeast has the help of Qingjiang cuisine. The effect of growth.

2. Leaf lettuce test: Lettuce has its scientific name Lactuca sativa L. The test period was from March 24, 2008 to April 22 of the same year. The test site was at the greenhouse of National Chung Hsing University. The test treatment uses a Randomized Complete Block Design (RCBD). This test uses 5 吋 plastic pots and 1 kg of soil per pot. The test treatment was as follows: (1) CK: control group, (2) the yeast CC-DH2011: application 10 ml / pot of yeast CC-DH2011 bacteria (100-fold dilution), (3) CF: application recommendation The total amount of the composting compound fertilizer No. 1 (0.15 g / pot) and (4) 1/2 of the yeast CC-DH2011 + 1/2 CF: a half amount of the yeast CC-DH2011 bacterial solution (5 ml / pot) was applied.

During the test period, except for the treatment group with the relevant fertilizer application, the other bacterial liquid and liquid fertilizer treatment groups were applied once a week. The growth after 4 weeks of the test treatment is shown in Annex VI, and was collected after 4 weeks. The fresh weight and dry weight were weighed and counted by Duncan's multiple range test (p=0.005). The results are shown in Table 13, Figure 3 and Figure 4. Display fresh weight and dry weight of leaf lettuce. Add half-quantity chemical fertilizer (1/2 CC-DH2011+1/2CF) and full amount in the bacterial liquid containing the yeast strain C.ethanolica CC-DH2011 of the present invention. The chemical fertilizer (CF) treatment group was superior to the control treatment group. The semi-quantitative fertilizer (1/2 CC-DH2011+1/2CF) containing half of the bacterial liquid was very similar to the dry weight of the fully quantified fertilizer treatment group. The results show that the C. ethanolica CC-DH2011 strain is beneficial to the growth of leaf lettuce, and can reduce the application amount of 1/2 chemical fertilizer. If combined with chemical fertilizer interaction, it shows that the yeast has assistance. The effect of leaf lettuce growth.

However, the above-described embodiments are merely illustrative of the principles of the invention and its effects, and are not intended to limit the invention. Therefore, modifications and variations of the embodiments described above will be made without departing from the spirit of the invention. The scope of the invention should be as set forth in the appended claims.

Fig. 1 shows the application of the yeast C. ethanolica CC-DH2011 strain of the present invention, chemical fertilizer (CF Taiwan compound fertilizer No. 1), 1/2 CC-DH2011 bacterial solution + 1/2 chemical fertilizer to Qingjiang vegetable Fresh weight histogram

Fig. 2 shows the application of the yeast ( C. ethanolica ) CC-DH2011 strain bacterial solution, chemical fertilizer (Taiwan compound fertilizer No. 1), 1/2 CC-DH2011 bacterial solution + 1/2 chemical fertilizer of the present invention. Histogram of dry weight of Qingjiang vegetable

Figure 3 is the fresh weight of the lettuce after applying the yeast C. ethanolica CC-DH2011 strain of the invention, the full amount of chemical fertilizer (CF compound fertilizer No. 1), 1/2CC-DH2011 bacterial solution + 1/2 chemical fertilizer. Histogram of influence

Figure 4 is a leaf lettuce applied to the yeast of the invention C. ethanolica CC-DH2011 bacterial liquid, full chemical fertilizer (CF Taiwan compound fertilizer No. 1), 1/2CC-DH2011 bacterial liquid + 1/2 chemical fertilizer, dried to the plants Histogram of heavy influence

Claims (5)

A yeast preparation which can be used for the manufacture of a multifunctional microbial fertilizer, which comprises Candida ethanolica CC-DH2011, which has been deposited at the Food Industry Development Research Institute of Hsinchu, Taiwan, under the accession number BCRC920070, The preparation can be directly inoculated into the root zone soil of the crop to make it have the action of dissolving iron phosphorus, calcium phosphorus and aluminum phosphorus, and the organic matter can be decomposed to generate acid, so as to dissolve the nutrients which are not easily dissolved, thereby improving the availability of nutrients and releasing mineralization. The latter product is a nutrient for plant absorption, and a plurality of organic carbon sources are more useful as a matrix for microbial proliferation to facilitate colonization of microorganisms. A yeast preparation for promoting plant growth, the preparation comprising Candida ethanolica CC-DH2011, which has been deposited in the Food Industry Development Research Institute of Hsinchu, Taiwan, with the accession number BCRC920070, the preparation can be Direct inoculation into the crop root zone soil, so that it has the action of dissolved iron phosphorus, calcium phosphorus and aluminum phosphorus, which can decompose organic matter to produce acid, dissolve nutrients that are not easily dissolved, improve nutrient availability, and release mineralized The product is a nutrient for plant absorption, and a plurality of organic carbon sources are more useful as a substrate for microbial proliferation to facilitate colonization of microorganisms. A yeast preparation which can be used for soil improvement, which comprises Candida ethanolica CC-DH2011, which has been deposited in the Food Industry Development Research Institute of Hsinchu, Taiwan, with the accession number BCRC920070, which can be directly Inoculated into the root zone soil of the crop to make it have the action of dissolving iron phosphorus, calcium phosphorus and aluminum phosphorus, which can decompose the organic matter to produce acid, dissolve the nutrients which are not easily dissolved, improve the availability of nutrients, and release the products after mineralization. The nutrient fraction is absorbed by the plant, and a plurality of organic carbon sources are more useful as a matrix for microbial proliferation to facilitate colonization of microorganisms. A yeast preparation for biological control, which comprises Candida ethanolica CC-DH2011, which has been deposited in the Food Industry Development Research Institute of Hsinchu, Taiwan, and has the registration number BCRC920070. The multifunctional activity of the action of dissolving iron phosphorus, calcium phosphorus and aluminum phosphorus. A yeast preparation which can be used for decomposition of organic matter, which comprises Candida ethanolica CC-DH2011, which has been deposited in the Food Industry Development Research Institute of Hsinchu, Taiwan, and has the registration number BCRC920070. The multifunctional activity of the action of dissolving iron phosphorus, calcium phosphorus and aluminum phosphorus.