TWI333941B

TWI333941B -

Info

Publication number: TWI333941B
Application number: TW94119507A
Authority: TW
Inventors: Ching Yen Huang; Yao Hsing Huang; Li Ai Huang; Hsieh Cheng Lin; Yen Yu Cheng; Kuang Jia Cheng; De Jung Huang; Ren Che Shieh; Tzen Wu Tsai; Jau Min Wu
Original assignee: Taiwan Fertilizer Co Ltd
Priority date: 2005-06-13
Filing date: 2005-06-13
Publication date: 2010-12-01
Also published as: TW200642991A

Description

1333941 九、發明說明：【發明所屬之技術領域】 ' 本發明係關於一種生物肥料，特別是關於一種可拮抗、抑制病源菌且可增進農產品品質的生物肥料。 ^ 【先前技術】二次世界大戰以後，化學肥料及農藥被大量地應用在農作物的栽種上。此種栽種方式雖然可以提高農業生產的效率，但是，長此以往，大量施用化學肥料及農藥已經對 Φ 土壤以及環境造成了相當大的傷害。長期施用無機化學肥料而忽視有機肥料的補充，使得土壤呈現酸化、固化、缺乏有機物，並且造成土壤的酸鹼失衡。再者，施灑後，無法被植物吸收利用的部分，會造成土壤的污染，而且，因、為化學肥料均為可溶性的鹽類，因此，會造成地下水源的 . 污染，同時會因為重複施灑的次數越多，使污染狀況更形嚴重。此外，為了滅絕病源菌與雜草，遂大量使用除草劑、除蟲劑等農藥，如此雖暫時抑制了病源菌的感染，但是一 ^ 但藥效降低，病源菌仍會快速繁殖，甚至出現抗藥性更強的病源菌，而使得農藥越用越多，如此的長期使用與殘留，使得土壤中的有益微生物及蚯蚓等生物均無法存活，土壤中的有機質日益減少，而污染卻日益加劇，如此不但使得作物的產量及品質降低，甚至會造成農地無法耕種的結果。為了改善使用化學肥料的缺點，有機肥料的利用逐漸 - 受到重視。一般常見用於製造有機肥料的來源包括禽畜糞便、廚餘以及農業廢棄物（如稻梗、花生殼、甘蔗渣）等等。因為這些有機物無法直接提供作物利用，因此均需經過腐 ⑧ 5 熟的處理過程。與化學肥料相比，通常有機肥料具有養分含量較低，養分釋出較慢，成本高（因需長時間的堆肥腐熟過程），成分不穩定等缺點。但是，相對上，其具有肥效持久，可改善土壌的物理、化學、生物等特性的優點。但是，製造有機肥料時，來源及製造過程均會影響最終成品的品質，若於製造過程中未去除來源物裡的毒性物質或有害成分，未去除雜草種子、蟲纟卩或幼蟲、病源菌，或是未完全腐熟，直接施用於土壤中，不但無法改善土壤，反而可能造成土壌品質的降低，或是影響作物生長。近來，另一種微生物肥料（microbial fertilizer)或稱生物肥料（biofertilizer)也相當受到重視。其是指含有某種活的微生物或酵素的粉狀或液狀製劑，將其施用在種子、幼苗或土壤上，可以加強作物對營養素的利用或增加土壤中的營養分，補充土壤中有益微生物的數量，使土壤在良好的生態環境下發揮功能。例如，固氮菌具有固氮作用，可以將空氣中的氮素固定為氨，轉變成植物可以利用的含氮化合物，如此可增加土壤的氮素來源。而某些溶解菌可以使土壤中不能被利用的結合型營養元素溶解，使植物得以利用。又如菌根菌可以增加根系吸收能力及表面積，促進養分的吸收。使用微生物肥料可以部分取代或減少化學肥料的施用，或是促進養分的吸收效率，進而獲得較佳的生長效果。本發明即提供一種兼具可拮抗抑制病源菌，且可增進多種農作物品質的生物肥料。因為此種生物肥料可以拮抗抑制多種病源菌，因此可以減少農藥的施用，同時，此種生物肥料可以促進多種農作物生長，提供高品質的農產品。 1333941 【發明内容】本發明之目的為提供一種生物肥料，其包含芽孢枯草 • 桿菌a/wj/o/Wwe/flc/ewWQSOSlW此芽孢枯草桿菌已寄存於食品工業發展研究所，寄存編號為BCRC 910284)，其可拮抗多種不同的病源菌，並可增進農產品的品質。上述生物肥料可拮抗的病源菌係選自立枯絲核菌 so/izm)所組成的群組。上述生物肥料可拮抗的病源菌係選自草皮褐斑病菌馨（似加以續α如/細· TCCH)、甘藍立枯病菌（似⑹伽論 «so/izm· Cab4-1) ’ 水稻紋枯病菌（及如/⑽^ R96)所組成的群組。上述生物肥料可增進農產品的品質，該農產品係選自西瓜、番茄、洋香瓜、小白菜，以及小胡瓜所組成 • 的群組。上述之增進農產品的品質係為增加產量、增加果膏章量或是增加果實甜度。以下將以具體的實施例進一步說明本發明。需瞭解的 • 是，下述所列舉的實施例係用以闡明本發明，並非用以限定本發=之範圍，任何熟習此技藝者，在不脫離本發明之精^和範圍内，當可做些許更動與潤飾，因此本發明之保護範圍當視後附之申請專利範圍所界定者為準。【實施方式】實施例1本發明生物肥料之製備〈製備接種菌液〉 1333941 使菌絲的病菌菌絲塊，放置於含PDA培養基的培養塊位於中央。另外，將枯草桿菌召# (下文中以代號Bstw 代表⑽#.(下文中以代號BSK代表）以及本發明之生物肥料（其製備方法如實施例1所示）均以Bs培養基（其成分為1%黃豆粉、〇.5%酵母粉、〇 5%魚溶聚，以"及蜜）製備為菌液濃度為！〇4xl〇9CFU/mL的菌 =〇.0二的水：BS培養基以及上述三種菌液，分別滴 :ί 2 ?慮紙塊。將濾紙塊置放於上述具有病源菌囷…Α、平板培養基上，使濾紙塊距離培養皿邊緣為二iUfc的環境下培養，於第二、三天後觀察么己=原=絲生長距離及拮抗菌的拮抗距離，且每症二：ϊί進行。前述之「菌絲生長距離」係指自 2 5:卜生長出的菌絲長度，而「拮抗距二7落算起，至病源菌菌絲之距離。「菌絲 :拮抗距離」越大，均表示滤紙塊上所帶有的賴物貝對雙測試的病菌有較佳的拮抗效果。士，;表1所示。不論對於何種病源菌而 a本發明之生物肥料均較其他_具有較佳的抑菌效果。作用因金對於病源菌有良好的拮抗:巧的用量。辰作物時，可減少病害，並且減少農藥本發㈣物肥料的西瓜田間肥效試驗 δ式驗瓜田位於台中縣勘斗… 為李姓農友所栽種，以鄉’西瓜品種係㈣西瓜’ 人定期派人前往進行栽培方法為基礎’由申請間知*灑作業。試驗組是於定植後之 ⑧ 1333941 15天以及23天分別施灑一次本發明之生物肥料（其製備方法如實施例1所示）以及溶填菌π.)，亦即於植株營養生長期施用本發明之生物肥料，而於開花結果期施用溶磷菌。另外，以不施用本發明之生物肥料的區域為對照組。試驗面積為二畦，施灑時，以107 CFU/ml的濃度每株施灑200mL，施灑部位為植株根圈土壤附近。試驗期間觀察並記錄西瓜生育情形，並調查果實重量與甜度。再者，於92年7月30日進行土壤採樣，並分析土壤樣品中之菌相及菌數。結果如表2所示。以果實的產量而言，試驗組之平均果重為3,730g，較對照組3,370g多360g，產量約可增加 10.7%。若以每公頃種2,200株，每株採5粒果實計，試驗組每公頃產量約41,030公斤，較對照組37,070公斤多3,960 公斤。以果實的甜度而言，試驗組的平均甜度為10.6 Brix，較對照組的10.1 Brix高0.5 Brix。以土壤的分析結果而言土壤中試驗組之菌相以所施用之菌種成為土壤中之優勢菌種，因為所施用之菌種具有拮抗病源菌、固定氮素、促進土壤中磷之釋放與促進土壤團粒構造形成等作用，故對於作物與土壤具有正面的效果。試驗組的施用菌種的殘留菌數約達106CFU/g，證實所施用的菌種可以存活於土壤中，並且具有一定的數量，因此可以幫助改善土壤中的菌相。另外，試驗組的瓜葉較對照組厚實且亮麗，生長勢亦普遍較好。由上述結果可知，本發明之生物肥料可以使西瓜的生長勢較好，果重、產量及甜度均增加，而且所施用的菌種已成為土壤中最多菌數的族群，因此可以改善土壤中的菌相。 1333941 魅Μ本發明生物肥料對番祐的肥效試驗於簡易網室中進行番茄肥致 301。所使用之培養土為台肥種花円益，番茄種苗為農友鉀總含量在1%以下。肥料ΐ土’其氣、磷、 (Ν-Ρ2〇5-Κ2〇= 5.0-2.5-2.5)。另外，''八木 $ 豐牌有機肥料、溶填菌CSflcz’Z/ws印.）、以及菌 &明之生物肥 /id⑼(/brm⑷的三個不同試驗袓，发為SH2(^^·//⑽ MC、BC、SH2表示，三種菌、、存±/:^驗組別代號分別以 107CFU/ml備用。：以…、菌水稀釋至菌數為栽培土施用東豐有機肥為基肥，於9 綱種後’挑選生長勢相近之番祐年9月Π 30曰定植’定時堯水’並依作物施肥::2年:二機肥為追肥。試驗組於92/1_第一次施用=用^ = =週施用一次。每組之樣品數均2 5 株生月If形，番加開化後約3〇天，果實不再長大皮顏色轉紅時即行採收，番#果實採收後，測量其重^及甜度。 ’、結果如表3所示。以果實的產量而言，每株平均產量 MC^且產量最高，較對照組增加239 g (增產5〇%卜bc纽產量次之，增產l22g (25.5%)。SH2也較對照組增產22g (4.6%)。以果重而言，以SH2組之果重最重。各組之間並無明顯差異。 '度由上述結果可知，本發明之生物肥料可以使番茄的產量增加。 ⑧ 1333941 堂^Li本發明生物肥料的洋香瓜田間肥效試驗 #試驗瓜田位於苗栗縣通宵鎮，為吳姓農友所栽種，以其慣行之栽培方法為基礎，由申請人定期派人前往進行田 . 間施灑作業，試驗區域分為原試驗區與A試驗區，A試驗區之試驗期間較原試驗區晚一季進行。且每試驗區中均設有試驗組及對照組’試驗組係定期施灑依照實施例1的方法所製備的生物肥料菌液，選定2()株洋香瓜作為試驗對象’以108 CFU/mL的菌液濃度’每株施灑2〇〇mL，每1〇天施灑一次，而對照組則不施用本發明之生物肥料。試驗 • 期間為92年4-8月，觀察並記錄洋香瓜的生育情形、記錄果實重ΐ ’並將採收果實分為上、中、下部，分別測其甜度。另外，分別於92年7月11日、92年7月17日以及 92年8月8日進行土壤採樣，並分析土壤樣品中之菌相及菌數。結果如表4所示。以果實重量而言，原試驗區之試驗組中，每個洋香瓜果實平均重量比對照組增加51g，而A 試驗區中的試驗組中，每個洋香瓜之果實平均重量比對照組增加19g。以果貫甜度而言，不論是原試驗區或a試驗區中的試驗組’洋香瓜果實的上、中、下部，其平均甜度均比對照組為高。至於土壤中的菌數，原試驗區的試驗組之土壤’芽孢枯草桿菌(Bacillus am少19(BCRC 910284)已成為土士穿中之優勢菌種，菌數約在l〇6-l〇7CFU/g。A試驗區中試^組之土壌’芽抱枯草桿菌(Bacillus 910284)也已成為土土穿中之優勢菌種，菌數約在i〇5-i〇6CFU/g。由上述結果可^， . 本發明之生物肥料可以使洋香瓜的果重、產量及甜度均增 ⑧ 加 ’而且所施用的菌種可以存活於土壤中，並成為優勢菌〇芽抱枯草桿菌(Bacillus amyloliquefaciens) 2Q819添加於複合肥料中之肥效試驗1333941 IX. Description of the invention: [Technical field to which the invention pertains] The present invention relates to a biological fertilizer, and more particularly to a biological fertilizer which can antagonize, inhibit, and improve the quality of agricultural products. ^ [Prior Art] After the Second World War, chemical fertilizers and pesticides were widely used in the cultivation of crops. Although this kind of planting method can improve the efficiency of agricultural production, in the long run, the large-scale application of chemical fertilizers and pesticides has caused considerable damage to Φ soil and the environment. The long-term application of inorganic chemical fertilizers and the neglect of organic fertilizer supplementation make the soil acidified, solidified, lack of organic matter, and cause acid-base imbalance in the soil. In addition, after being applied, the part that cannot be absorbed by the plant will cause soil pollution, and because the chemical fertilizer is a soluble salt, it will cause pollution of the groundwater source and will be repeated. The more times you sprinkle, the more serious the pollution situation. In addition, in order to eliminate pathogenic bacteria and weeds, a large number of pesticides such as herbicides and insecticides are used, which temporarily inhibits the infection of pathogenic bacteria, but the efficacy is reduced, the pathogenic bacteria will rapidly multiply, and even anti-antibiotics will appear. The more medicinal pathogens, the more the pesticides are used, the long-term use and residue, the beneficial microorganisms and the cockroaches in the soil can not survive, the organic matter in the soil is decreasing, and the pollution is increasing, so Not only does the yield and quality of the crops decrease, but it can even result in the inability of the farmland to cultivate. In order to improve the shortcomings of using chemical fertilizers, the use of organic fertilizers has gradually gained attention. Common sources of organic fertilizers commonly used include livestock manure, kitchen waste, and agricultural waste (such as rice stalks, peanut hulls, bagasse). Because these organics cannot directly provide crop utilization, they all need to undergo a process of rot. Compared with chemical fertilizers, organic fertilizers generally have disadvantages such as low nutrient content, slow release of nutrients, high cost (due to long-term composting and decomposing processes), and unstable composition. However, relatively speaking, it has a long-lasting effect, which can improve the physical, chemical, biological and other characteristics of the soil. However, when manufacturing organic fertilizers, the source and manufacturing process will affect the quality of the final product. If the toxic or harmful components in the source are not removed during the manufacturing process, weed seeds, worms or larvae, pathogens are not removed. Or not fully decomposed, directly applied to the soil, not only can not improve the soil, but may cause the soil quality to reduce, or affect crop growth. Recently, another microbial fertilizer or biofertilizer has also received considerable attention. It refers to a powdery or liquid preparation containing a living microorganism or enzyme, which is applied to seeds, seedlings or soil, which can enhance the utilization of nutrients by the crop or increase the nutrient content in the soil, and supplement the beneficial microorganisms in the soil. The amount of soil that makes the soil function in a good ecological environment. For example, nitrogen-fixing bacteria have nitrogen fixation, which can fix nitrogen in the air to ammonia and convert it into nitrogen-containing compounds that can be used by plants, thus increasing the nitrogen source of the soil. Some lytic bacteria can dissolve the combined nutrients in the soil that cannot be utilized, allowing plants to be utilized. Another example is mycorrhizal fungi, which can increase root absorption capacity and surface area and promote nutrient absorption. The use of microbial fertilizers can partially replace or reduce the application of chemical fertilizers, or promote the absorption efficiency of nutrients, thereby achieving better growth. The present invention provides a biological fertilizer which can antagonize the inhibition of pathogenic bacteria and can improve the quality of various crops. Because this bio-fertilizer can antagonize the inhibition of a variety of pathogenic bacteria, it can reduce the application of pesticides. At the same time, this bio-fertilizer can promote the growth of a variety of crops and provide high-quality agricultural products. 1333941 SUMMARY OF THE INVENTION The object of the present invention is to provide a biological fertilizer comprising Bacillus subtilis bacillus a/wj/o/Wwe/flc/ewWQSOSlW. This Bacillus subtilis has been deposited in the Food Industry Development Research Institute under the registration number BCRC 910284 ), it can antagonize a variety of different pathogenic bacteria and can improve the quality of agricultural products. The pathogenic bacteria which can be antagonized by the above biological fertilizer are selected from the group consisting of Rhizoctonia solani so/izm). The pathogenic bacteria that can be antagonized by the above bio-fertilizer are selected from the group consisting of turf brown spot disease (like to continue to be α/fine TCCH), and cabbage blight (like (6) gamma «so/izm· Cab4-1) A group of germs (and such as /(10)^R96). The above-mentioned bio-fertilizer can improve the quality of the agricultural product, and the agricultural product is selected from the group consisting of watermelon, tomato, cantaloupe, Chinese cabbage, and courgette. The above-mentioned promotion of the quality of agricultural products is to increase the yield, increase the amount of fruit paste or increase the sweetness of the fruit. The invention will now be further illustrated by the specific examples. It is to be understood that the following examples are set forth to illustrate the invention and are not intended to limit the scope of the present invention, and those skilled in the art can, without departing from the scope of the invention, The scope of protection of the present invention is defined by the scope of the appended claims. [Embodiment] Example 1 Preparation of Biological Fertilizer of the Present Invention <Preparation of Inoculating Bacteria> 1333941 The mycelial mass of the mycelium was placed in the center of the culture block containing the PDA medium. In addition, B. subtilis (hereinafter referred to as code Bstw representative (10) #. (hereinafter referred to as code BSK) and the biological fertilizer of the present invention (the preparation method thereof is as shown in Example 1) are all in Bs medium (the composition thereof is 1% soy flour, 〇.5% yeast powder, 〇 5% fish dissolved, prepared with "and honey as the concentration of the bacteria solution! 〇4xl〇9CFU/mL bacteria = 〇.0 2 water: BS medium And the above three kinds of bacterial liquids are respectively dropped: ί 2 ? paper pieces. The filter paper pieces are placed on the above-mentioned pathogenic bacteria Α, 平板, plate medium, so that the filter paper pieces are cultured in an environment of two iUfc from the edge of the culture dish, After the second and third days, observe the original = the original growth distance of the silk and the antagonistic distance of the antagonistic bacteria, and the second disease: the progress of the "mycelium growth distance" refers to the hyphae grown from the 25: The length, and the "antagonism distance from 2 to 7 counts, the distance to the mycelium of the pathogenic bacteria. The larger the "hyphae: antagonistic distance", means that the larvae on the filter paper block are better for the double-tested pathogens. Antagonistic effect. Shi,; Table 1. Regardless of which pathogenic bacteria and a biological fertilizer of the present invention It has better antibacterial effect than other _. The effect of gold on the pathogenic bacteria has a good antagonism: the amount of clever use. When the crop is cropped, it can reduce the disease, and reduce the pesticide field (four) fertilizer fertilizer field fertilizer efficiency test δ type The melon field is located in Taichung County. It is planted for the farmer of Li Xing, and the village 'watermelon variety (four) watermelon 'people regularly send people to the cultivation method based on 'applications to know * sprinkle operations. The test group is after planting 8 1333941 15 days and 23 days, respectively, the biological fertilizer of the present invention (the preparation method is as shown in Example 1) and the dissolved bacteria π.), that is, the biological fertilizer of the present invention is applied in the vegetative growth period of the plant, Phosphorus solubilizing bacteria are applied during the flowering period. Further, a region in which the biological fertilizer of the present invention was not applied was used as a control group. The test area was two 畦, and when spraying, 200 mL per plant was applied at a concentration of 107 CFU/ml, and the application site was near the root zone soil of the plant. During the test, the growth of watermelon was observed and recorded, and the weight and sweetness of the fruit were investigated. Furthermore, soil sampling was conducted on July 30, 1992, and the number of bacteria and bacteria in the soil samples was analyzed. The results are shown in Table 2. In terms of fruit yield, the average fruit weight of the test group was 3,730 g, which was 360 g more than the control group of 3,370 g, and the yield increased by about 10.7%. If 2,200 plants per hectare are used, and 5 fruits per plant are used, the yield per hectare of the test group is about 41,030 kg, which is 3,960 kg more than the control group of 37,070 kg. In terms of the sweetness of the fruit, the average sweetness of the test group was 10.6 Brix, which was 0.5 Brix higher than the 10.1 Brix of the control group. Based on the soil analysis results, the bacterial phase of the test group in the soil became the dominant species in the soil by the applied strain, because the applied strain had antagonistic pathogens, fixed nitrogen, and promoted the release of phosphorus from the soil. It promotes the formation of soil aggregate structure and so on, so it has a positive effect on crops and soil. The residual bacteria of the applied strain of the test group was about 106 CFU/g, which confirmed that the applied strain could survive in the soil and had a certain amount, and thus could help improve the bacterial phase in the soil. In addition, the melon leaves of the test group were thicker and brighter than the control group, and the growth potential was generally better. It can be seen from the above results that the biological fertilizer of the present invention can make watermelon grow better, the fruit weight, the yield and the sweetness increase, and the applied strain has become the most common population in the soil, so the soil can be improved. The bacterial phase. 1333941 Charming the bio-fertilizer of the invention to the effect of the fruit blessing in the simple net room to achieve tomato fertilizer 301. The culture soil used is the plant fertilizer of Taiwanese fertilizer, and the tomato seedlings are the farmers. The total potassium content is below 1%. Fertilizer bauxite 'gas, phosphorus, (Ν-Ρ2〇5-Κ2〇= 5.0-2.5-2.5). In addition, ''Yamu $ Feng brand organic fertilizer, dissolved bacteria CSflcz 'Z / ws printed.), and bacteria & Ming biological fertilizer / id (9) (/brm (4) three different test 袓, issued as SH2 (^ ^ · / /(10) MC, BC, SH2 indicates that the three strains, and the ±/:^ test group codes are reserved at 107 CFU/ml respectively. The dilution of the bacteria to the number of bacteria is used to apply Dongfeng organic fertilizer as the base fertilizer. After 9 classes, 'selecting the growth potential of the near year of the year of the year of the year Π 30曰 planting 'timed drowning' and fertilizing according to the crop: 2 years: two machine fertilizer for topdressing. The test group at 92/1_ first time Application = use ^ = = weekly application. The number of samples in each group is 2 5 plants and the shape of the month, about 3 days after the opening, the fruit no longer grows when the skin color turns red, the fruit is harvested. After harvesting, the weight and sweetness were measured. ', the results are shown in Table 3. In terms of fruit yield, the average yield per plant was MC^ and the yield was the highest, which increased by 239 g compared with the control group (increased yield by 5〇%). The production of bc New Zealand was second, with an increase of l22g (25.5%). SH2 also increased yield by 22g (4.6%) compared with the control group. In terms of fruit weight, the fruit weight of SH2 group was the heaviest. There was no significant difference between the groups. Degree As can be seen from the results, the bio-fertilizer of the present invention can increase the yield of tomato. 8 1333941 堂^Li The bio-fertilizer of the invention can be tested in the field. The test melon field is located in Tongyu Town, Miaoli County, and is planted by Wu’s farmer. Based on the customary cultivation method, the applicant regularly sends people to carry out field operations. The test area is divided into the original test area and the A test area. The test period of the A test area is carried out later than the original test area. The test group and the control group were set in the test area. The test group was regularly sprayed with the bio-fertilizer liquid prepared according to the method of Example 1. The selected 2 () strain of cantaloupe was used as the test object '108 CFU/mL bacteria The liquid concentration was sprayed 2 mL per plant and applied once every 1 day, while the control group did not apply the biological fertilizer of the present invention. The test period was from April to August of 1992, and the fertility of melon was observed and recorded. The situation, the record of the fruit is heavy, and the harvested fruit is divided into upper, middle and lower parts, and the sweetness is measured separately. In addition, on July 11, 1992, July 17, 1992 and August 8, 1992, respectively. Soil sampling and analysis of soil The bacterial phase and the number of bacteria in the sample. The results are shown in Table 4. In terms of fruit weight, the average weight of each cantaloupe fruit in the test group of the original test area increased by 51 g compared with the control group, and the test in the A test area In the group, the average weight of each melon fruit was 19g higher than that of the control group. In terms of sweetness, whether it was in the original test area or the test group in the test area, the upper, middle and lower parts of the melon fruit, The average sweetness was higher than that of the control group. As for the number of bacteria in the soil, the soil of the test group in the original test area, Bacillus subtilis (Bacillus am. 19 (BCRC 910284), has become the dominant strain of the tortoise. The number is about l〇6-l〇7 CFU/g. The B. bacillus (Bacillus 910284), a soil test in the test group of A test area, has also become the dominant strain in soil penetration, and the number of bacteria is about i〇5-i〇6 CFU/g. According to the above results, the bio-fertilizer of the invention can increase the fruit weight, yield and sweetness of the cantaloupe by 8 plus ' and the applied strain can survive in the soil, and becomes the dominant bacterium Bacillus subtilis (Bacillus amyloliquefaciens) Fertilizer effect test of 2Q819 added to compound fertilizer

試驗肥料是以台肥特1號複合肥料（20-5-10)中添加芽 ?已枯草♦干菌（如化似)920819(BCRC 料，直菌絮:A 61添加芽抱枯草桿菌之台肥特1號複合肥 , ^l〇7CFU/mL〇 ί成：為ί氮肥料為對照組。台肥㈣複合肥料 s ivv、爐供i 2〇.0%、銨態氮（N) 8.0%、硝酸態氮（N) /n。/0，:二心溶性碟酐（P2〇5) 5.0%、水溶性填酐（p2〇5) π °7’, 1及水溶性氧化鉀（K20) 10.0%。芽孢枯草桿菌 ⑽卿/〇/如e/⑽循)920819(BCRC 910284)之製備方法如實施例1所示。之作物$小白菜（品種為農友種苗「鳳山白菜」） : 加°〇種為「種苗7號」），作物施肥之規劃如表5 所不。仏，f之栽培方式係先將10L之培養土分別與3g的試 j波…、組的特1號複合肥料混合均勻。接著，將其裝 ^各培^中舖平，再將小白菜以穴播方式種植於盒祕二1〇穴’每穴播種2顆小白菜種子。發芽後留取1 Ϊ生Ϊif之幼苗’其餘拔除。韻組與對照組各為四重、二^菊固栽培倉。發芽後約二週施追肥一次，每盒3g， ί方式施用。平曰注意水分管理與病蟲害防治。二3 1月為93年9月27曰至10月20曰。栽培期間定期 π相並$錄植株生長情形。植株達採收標準時，進行全株 1333941 採收，照相、秤重、量株高，記錄根部生育情形，並將生育調查數據進行生物統計分析。 • 番茄之栽培方式係先將15L之培養土分別與5g的試驗 , 組及對照組的特1號複合肥料混合均勻。接著，將其裝入黑色塑膠盆中鋪平，將番茄幼苗定植於盆子中央，每盆一株。試驗組與對照組各10盆，共20盆。施追肥三次，每次每盆3g，分別於定植後20-25天、40-50天、60-75天，以環施覆土方式施用。平日注意水分管理與病蟲害防治。栽培期間定期照相並紀錄植株生育情形。果實達採收標準 Φ 時，秤量並紀錄採收果實之鮮重、粒數、果徑、甜度等，並統計產量。實驗結束時採收根部，記錄生育情形，並將生育調查數據進行生物統計分析。小白菜之試驗結果如表6所示。生長初期試驗組之株高、全株鮮重與根鮮重皆較對照組佳，全株鮮重增加59% ; ' 根長方面，雖然試驗組的根長較短，但根鮮重則重於對照組，顯示試驗組的根長雖然較短，但根系生長旺盛，並且植株生長旺盛。與對照組相比，試驗組可促進小白菜生長，提高小白菜賣相，增進農民收益。番茄之試驗結果如表7所示。盆栽肥效觀察試驗日期為93年9月16日至12月20日。將成熟之番茄採收、秤重並測量甜度。結果顯示試驗組較對照組的番茄果實單粒重量平均增加31克（增加27%)，在果實總產量與甜度方面有微量增加。由上述之結果可知，特1號複合肥料添加芽孢枯草桿菌後，其對試驗作物的平均果重有明顯的增加效果。 14 ⑧ 1333941 實施例7本發明生物肥料對小胡瓜的肥效試驗於簡易網室中進行小胡瓜肥效試驗。所使用之培養土為台肥種花草園藝栽培土，其氮、磷、鉀總含量在1%以下。依照實施例1製備本發明之生物肥料作為試驗組1，以施用無菌水者為對照組，施用時試驗組1之菌液以無菌水稀釋至菌數1〇7 CFU/ml使用。The test fertilizer is added with buds from Taiwan Fertilizer No. 1 compound fertilizer (20-5-10). The dried grass ♦ dry bacteria (such as chemical) 920819 (BCRC material, straight fungus: A 61 added Bacillus subtilis) Fert No. 1 compound fertilizer, ^l〇7CFU/mL〇ίcheng: for the nitrogen fertilizer as the control group. Taiwan fertilizer (four) compound fertilizer s ivv, furnace for i 2〇.0%, ammonium nitrogen (N) 8.0% Nitric acid nitrogen (N) / n. / 0,: dicentric soluble disc anhydride (P2 〇 5) 5.0%, water-soluble anhydride (p2 〇 5) π ° 7 ', 1 and water-soluble potassium oxide (K20) 10.0%. The preparation method of Bacillus subtilis (10) qing / 〇 / eg e / (10) 920819 (BCRC 910284) is shown in Example 1. Crops: Chinese cabbage (variety is the seedlings of the farmer "Fengshan cabbage"): Add the ° seedling to the "seed seedling No. 7"). The plan for crop fertilization is shown in Table 5.仏, f cultivation method is to first mix 10L of culture soil with 3g of test j wave ..., the group of special compound No. 1 compound evenly. Then, it will be flattened in each of the cultures, and then the cabbage will be planted in the box at the secret point of the 2nd hole, and 2 small cabbage seeds will be planted at each hole. After germination, leave 1 seedlings of Ϊif Ϊif and remove the rest. The rhyme group and the control group were each four-fold and two-cured solid cultivation chambers. Apply fertilizer once every two weeks after germination, 3g per box, ί. Pay attention to water management and pest control. February and January are from September 27 to October 20, 1993. During the cultivation period, the π phase is regularly recorded and the growth of the plant is recorded. When the plants reached the harvesting standard, the whole plant was harvested at 1,333,941, photographed, weighed, and planted, and the roots were recorded, and the biopsy data were analyzed. • The cultivation method of tomato is to first mix 15L of culture soil with 5g of test, group and control compound No. 1 compound fertilizer. Next, it was placed in a black plastic pot to be flattened, and tomato seedlings were planted in the center of the pot, one per pot. There were 10 pots in the test group and the control group, a total of 20 pots. Apply topdressing three times, each time 3g per pot, 20-25 days, 40-50 days, 60-75 days after planting, apply by ring application. Pay attention to water management and pest control on weekdays. Take regular photos during planting and record plant growth. When the fruit reaches the harvesting standard Φ, weigh and record the fresh weight, grain number, fruit diameter, sweetness, etc. of the harvested fruit, and count the yield. At the end of the experiment, the roots were harvested, the births were recorded, and the birth survey data was biostatistically analyzed. The test results of pakchoi are shown in Table 6. In the initial growth test group, the plant height, whole plant fresh weight and root fresh weight were better than the control group, and the whole plant fresh weight increased by 59%. 'In terms of root length, although the test group has a shorter root length, the root fresh weight is heavier than In the control group, the root length of the test group was short, but the roots grew vigorously and the plants grew vigorously. Compared with the control group, the experimental group can promote the growth of pakchoi, improve the sales of pakchoi, and increase farmers' income. The test results of tomato are shown in Table 7. The pot experiment results of the pot experiment were from September 16 to December 20, 1993. Mature tomatoes are harvested, weighed and measured for sweetness. The results showed that the average weight of tomato fruit in the experimental group was increased by 31 g (27% increase) compared with the control group, and there was a slight increase in total fruit yield and sweetness. From the above results, it was found that the addition of Bacillus subtilis to the No. 1 compound fertilizer showed a significant increase in the average fruit weight of the test crop. 14 8 1333941 Example 7 Fertilization test of bio-fertilizer of the invention on courgette The courgette efficacy test was carried out in a simple net room. The culture soil used is a planting soil of turf and horticulture, which has a total nitrogen, phosphorus and potassium content of less than 1%. The biological fertilizer of the present invention was prepared as the test group 1 according to Example 1, and the sterile water was used as the control group, and the bacterial liquid of the test group 1 was diluted with sterile water to the bacterial count of 1〇7 CFU/ml at the time of application.

小胡瓜種苗係於92年5月30日播種，發芽後挑選生長勢相近之小苗，於92年6月10日定植於栽培土，每日洗水兩次。於92年6月13日第一次以試驗組1之菌液及對照組的無菌水分別施用，每株各施用200mL，爾後每隔一週施用一次。各組之樣品數均為5株。另設一試驗組2，其施用菌液方式亦為每株各施用200 mL，菌液中則另添加適量之台肥5號即溶複肥，每隔一週施用一次。小胡瓜開花後，摘除主蔓四節内所開之雌花，主蔓五節以上開始留果。小胡瓜果實於開花後5-7天採收，以早 • 晨露水乾後果溫尚未升高時採收，測量其重量及甜度。結果如表8所示。以產量而言，試驗組1與對照組的甜度相同，但是試驗組1的產量較對照組增加134g (26%)。而試驗組2的甜度雖較對照組稍有降低，但其產量大幅增加，且較試驗組1增加375g (73%)。二試驗組之產量均較對照組大幅增加，尤以試驗組2之效果更佳，且生長勢較旺盛，亦即以本發明之生物肥料配合台肥5號的施用，對於提昇產量有更好的效果。 ⑧ 1333941 以培養土中的菌數而言，試驗結束後，試驗組1及2 中，芽孢枯草桿菌am少/o//gwe/<2c/e/w)920819(BCRC 910284)已成為培養土中之優勢菌種。由前述的說明以及各實施例中可知，本發明的生物肥料可拮抗抑制病源菌，減少農藥用量，並且對多種作物均有提高產量、增加單果重或是增加甜度的作用。對於改善土壤品質的效果而言，施用本發明生物肥料中的有益菌群可存活於土壤中，並且維持優勢菌群。若與化學肥料或有機肥料混用，對作物的生長有更佳的促進生長或增進產量 Φ 的效果。The small courgette seedlings were planted on May 30, 1992. After germination, the seedlings with similar growth potential were selected. On June 10, 1992, they were planted in cultivated soil and washed twice a day. On June 13, 1992, the bacterial liquid of the test group 1 and the sterile water of the control group were applied for the first time, and each plant was administered with 200 mL, and then applied once every other week. The number of samples in each group was 5 strains. Another test group 2 is set up, and the application of the bacterial liquid method is also 200 mL per plant, and the appropriate amount of the fertilizer is added to the bacterial liquid, and the compound fertilizer is applied once every other week. After the small courgette is flowered, the female flower opened in the four sections of the main vine is removed, and the main vine begins to leave the fruit for more than five knots. The courgette fruit is harvested 5-7 days after flowering, and harvested in the morning and morning dew. The weight and sweetness are measured when the temperature has not risen. The results are shown in Table 8. In terms of yield, the sweetness of the test group 1 was the same as that of the control group, but the yield of the test group 1 was increased by 134 g (26%) compared with the control group. While the sweetness of the test group 2 was slightly lower than that of the control group, the yield was significantly increased, and it was 375 g (73%) higher than that of the test group 1. The yield of the second test group was significantly higher than that of the control group, especially the effect of the test group 2 was better, and the growth potential was more vigorous, that is, the application of the bio-fertilizer of the invention with the application of the platform No. 5 was better for improving the yield. Effect. 8 1333941 In terms of the number of bacteria in the culture soil, after the test, in the test groups 1 and 2, the B. subtilis am/o//gwe/<2c/e/w) 920819 (BCRC 910284) has been cultured. The dominant species in the soil. As can be seen from the foregoing description and the respective examples, the biological fertilizer of the present invention can antagonize the inhibition of pathogenic bacteria, reduce the amount of pesticides, and have an effect of increasing yield, increasing single fruit weight or increasing sweetness for various crops. For the effect of improving soil quality, the beneficial flora in the application of the biological fertilizer of the present invention can survive in the soil and maintain the dominant flora. When mixed with chemical fertilizers or organic fertilizers, it has a better effect on growth of crops or increased yield Φ.

⑧ 16 1333941 【圖式簡單說明】無【主要元件符號說明】 1333941 表l'本發明生物肥料拮抗病源菌的效果菌絲生長距離(cm) 項目草皮褐斑病菌甘藍立枯病菌水稻紋枯病菌括抗距離(cm) 草皮甘藍水稻褐斑立枯紋枯拮抗能力草皮褐甘藍立水稻紋病菌二二斑病菌括病菌枯病菌8 16 1333941 [Simple description of the diagram] No [Major component symbol description] 1333941 Table l 'The effect of the bio-fertilizer of the invention against the pathogenic bacteria Mycelial growth distance (cm) Item Phytophthora brown spot pathogen Bacterial blight Anti-distance (cm) turf cabbage, brown spot, brown stalk, antagonism, turf, brown cabbage, rice smut, diphtheria sphaeroides

1.40 127 2-57 1.70 3.27 0 〇〇 --- BSTW 0-90 0.90 ---- 0.93 qZj ~~~ -- -------^67 1-67 0.63 0.60 0.43 + + + + + ++ 〇., 0.92 '—' —------—- ~~----1.10 0.60 0.63 0.37 + + + + + ++ + + 0.83 〇77 註：+++:病源菌菌 0.77 1 113 1.13 0.73 0.93 0.50 + + + + + ++ + + “：病源菌菌落一___. f菌落生長蝴受抑制^形撕制間距，S落無法再生長舰；. 洛生長初期受抑制， >成抑制間距，但菌絲仍可緩慢生長擴展；+ :病源菌絲生長正常’不受影響。制間距’但菌絲仍可繼續生長擴展一 + + :病源 :病源菌菌 13339411.40 127 2-57 1.70 3.27 0 〇〇--- BSTW 0-90 0.90 ---- 0.93 qZj ~~~ -- -------^67 1-67 0.63 0.60 0.43 + + + + + + + 〇., 0.92 '-' —-------- ~~----1.10 0.60 0.63 0.37 + + + + + ++ + + 0.83 〇77 Note: +++: pathogen bacteria 0.77 1 113 1.13 0.73 0.93 0.50 + + + + + ++ + + ": pathogen colony ___. f colony growth butterfly is inhibited ^ shape tearing spacing, S fall can not regenerate long ship;. Luo growth is inhibited at the beginning, &gt The inhibition spacing, but the hyphae can still grow slowly; +: the pathogen mycelial growth is normal 'not affected. The spacing is 'but the hyphae can continue to grow and expand one + + : pathogen: pathogenic bacteria 1333941

表2、實施例3中之西瓜平均單果重及甜度處理別平均果重 (g/顆）平均甜度 (Brix) 試驗組 3,730 10.6 對照組 3,370 10.1 表3、實施例4中之番茄果實平均產量、甜度與果重處理產量(g/株）甜度(Brix) 果重(g/顆）對照組 478 5.5 122 MC 717 5.5 121 BC 600 5.7 123 SH2 500 5.7 131 19 1333941Table 2, average fruit weight and sweetness of watermelon in Example 3 average fruit weight (g/piece) average sweetness (Brix) test group 3,730 10.6 control group 3,370 10.1 Table 3, average fruit of tomato in example 4 Yield, sweetness and fruit weight treatment yield (g/strain) sweetness (Brix) fruit weight (g/piece) control group 478 5.5 122 MC 717 5.5 121 BC 600 5.7 123 SH2 500 5.7 131 19 1333941

表4、實施例5中之洋香瓜平均單果重及甜度試驗區果實重量 (g/顆）甜度(Brix) 原试驗區對照組 1477.6 上 9.6 中 10.9 下 10.5 試驗組 1528.7 上 10.0 中 11.3 下 11.5 A 5式驗對照組 1418.0 上 8.8 中 11.5 下 10.8 試驗組 1437.3 上 10.7 中 11.8 下 11.7 註：洋香瓜採收日期為92.08.11。 1333941 表5、實施例6之作物施肥規劃處理項目特1號複合肥料(試驗組/對照組）供試驗作物小白菜番莊栽種天數 25-30 天 65天可開始採收作物栽培數量/盆 20株 1株培養土量/盆 10L 15L 基肥 3g 5g 施肥量Table 4, average single fruit weight and sweetness test area fruit weight (g/piece) sweetness (Brix) in the original test area 1477.6 upper 9.6 medium 10.9 lower 10.5 test group 1528.7 upper 10.0 medium 11.3 Lower 11.5 A 5 test control group 1418.0 upper 8.8 medium 11.5 lower 10.8 test group 1437.3 upper 10.7 medium 11.8 lower 11.7 Note: cantaloupe harvest date is 92.08.11. 1333941 Table 5, Crop Fertilization Planning and Treatment Project of Example 6 Special Compound Fertilizer No. 1 (test group/control group) For test crops, Chinese cabbage, Baizhuang planting days 25-30 days, 65 days can start harvesting crop cultivation number / pot 20 1 strain of culture soil / pot 10L 15L base fertilizer 3g 5g fertilization amount

註：1.基肥:以全層施肥方式施用；追肥：小自菜以條—-— 覆土方式施用。環施 2.培養土採用台肥公司基隆廠生產之台肥種花草園敲裁 1333941 表6、實施例6之小白菜栽培試驗結果項目對照組試驗組株高(cm) 16.71 19.00 根長(cm) 9.23 9.04 全株鮮重(g) 11.95 19.03 根鮮重(g) 0.30 0.37Note: 1. Base fertilizer: applied by full-layer fertilization; top dressing: small self-dish is applied by strip---covering method. Huanshi 2. Culture soil was planted by Taiwanese fertilizer company Kelon plant, which was produced by Taiwanese fertilizer planting grass garden 1339941 Table 6 and Example 6 cultivation results of cabbage cultivation project control group plant height (cm) 16.71 19.00 root length (cm 9.23 9.04 Fresh weight of whole plant (g) 11.95 19.03 Fresh weight (g) 0.30 0.37

表7、實施例6之番茄栽培試驗結果項目對照組試驗組平均果實重量（g/顆） 113 144 平均植株總產量（g/株） 1584 1601 平均果實甜度（Brix) 5.63 5.77Table 7. Results of tomato cultivation test in Example 6 Item Control group Test group Average fruit weight (g/piece) 113 144 Total plant total yield (g/plant) 1584 1601 Average fruit sweetness (Brix) 5.63 5.77

表8、實施例7之小胡瓜果實平均產量、甜度與試驗後培養土中之菌數項目產量 (g/株）甜度 (Brix) 土壤菌數 (CFU/mL) 對照組 517 3.8 1.5x10s 試驗組1 651 3.8 8.9xl05 試驗組2 892 3.4 1·42χ106 22 ⑧Table 8, the average yield, sweetness of the courgette fruit of Example 7 and the number of bacteria in the culture soil after the test (g/strain) sweetness (Brix) soil bacteria count (CFU/mL) control group 517 3.8 1.5x10s Test group 1 651 3.8 8.9xl05 Test group 2 892 3.4 1·42χ106 22 8

Claims

Howling 941 X. Patent Application: Announcement 1. A biological fertilizer for antagonizing Rhizoctonia solani (and /nzoc such as "/β(10)/(10)/), which contains Bacillus subtilis ^7/72 less / 〇&quot "The condition is (: chemistry) 920819 8 € 1 ^ 910284 and the endospore of Bacillus subtilis with a yield of at least 109 CFU / mL, wherein the Rhizoctonia solani is selected from the genus Bacillus licheniformis

TCCH), Brassica oleracea strain Cab4-1) and Rhizoctonia soiani R96. 2. A method for improving the quality of agricultural products, the method comprising the steps of applying the biological fertilizer according to claim 1 of the patent application to watermelon, yuyou, cantaloupe, pakchoi or courgette to enhance the fruits and vegetables Yield, fruit weight or fruit sweetness. 3. The method of claim 2, wherein the biofertilizer increases the average weight (g/piece) of the watermelon by 10.7%.

4. The method of claim 2, wherein the biofertilizer increases the average yield of watermelon (kg/ha) by 10.7%. 5. The method of claim 2, wherein the biofertilizer increases the average sweetness (Brix/particle) of the watermelon by 5%. 6. The method of claim 2, wherein the biological moon material increases the average yield (g/s) of the tomato by 50%. J. The method of claim 2, wherein the bio-fertilizer increases the average weight (grams/piece) of the melon by 35%. g. The method of claim 2, wherein the biological moon material increases the average sweetness (Brix/particle) of the melon by 21.6%. 23 1333941

9. The method of claim 2, wherein the bio-fertilizer increases the average weight (g/c) of the cabbage by 59.2%. 10. The method of claim 2, wherein the biofertilizer increases the average yield (g/s) of the courgette by 26%. twenty four