Microbacterium testaseum 1-39

Abstract

To provide a novel bacteria strain that can improve the quality of rice.SOLUTION: The present invention provides Microbacterium testaceum 1-39 (Accession number NITE AP-03803) that can improve the quality of rice.SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to Microbacterium testaceum 1-39.

Background Art

[0002] The ranking of the palatability (taste quality) of rice is roughly determined every year. The taste quality of rice is affected by genetic factors such as variety differences, and environmental factors such as cultivation management, weather, and fertilizers. However, even if cultivated under the same conditions with the same variety, the yield and taste quality will never be identical. Rice that has been cultivated since ancient times may exhibit different taste qualities even in adjacent paddy fields, despite the fact that its cultivation methods have been well established.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0004] Such differences in taste quality caused by soil preparation and terroir in adjacent paddy fields are considered to be partly attributable to differences in the soil microbiota. A wide variety of microorganisms exist in paddy field soil, and it is said that the number of such microorganisms can reach several million per 1 gram of soil.

[0005] While it is possible to improve the taste of rice through genetic analysis and the development of new varieties through crossbreeding, this is an extremely time-consuming process. Furthermore, crossbreeding is subject to influences from genetic background and other factors, making it difficult to consistently obtain the desired rice. Although the fertilizer applied during the rice cultivation period is thought to be a contributing factor, the factors involved in nutrient absorption differ depending on the cultivation environment and variety of rice. Therefore, achieving good-tasting rice requires careful consideration of the planting time and cultivation management methods, and creating superior-tasting rice requires considerable effort and time.

[0006] Methods have been developed to improve the taste and other qualities of rice after harvest. For example, Patent Document 1 discloses a rice flavor improver using a rice protein hydrolysate treated with a microbial proteolytic enzyme. Patent Document 2 also discloses a rice flavor improver using rice koji.

[0007] The present invention aims to provide a novel bacterial strain that can improve the quality of cooked rice. [Means for solving the problem]

[0008] The inventors have discovered that a specific strain of Microbacterium bacteria, collected from brown rice, has the ability to improve the quality of cooked rice when used in rice cooking.

[0009] One aspect of the present invention is Microbacterium testaseum 1-39.

[0010] One aspect of the present invention is a method for producing cooked rice, which includes adding the above-mentioned bacteria and cooking the rice.

[0011] One aspect of the present invention is a rice additive containing the above-mentioned bacteria. [Effects of the Invention]

[0012] The present invention provides a novel bacterial strain that can improve the quality of cooked rice. [Brief explanation of the drawing]

[0013] [Figure 1] This graph shows the taste scores of rice to which various bacteria have been added. [Figure 2] This graph shows the appearance score of cooked rice to which each type of bacteria has been added. [Figure 3] This graph shows the stickiness score of rice to which each type of bacteria has been added. [Modes for carrying out the invention]

[0014] Preferred embodiments of the present invention will be described in detail below. However, the present invention is not limited to the following embodiments.

[0015] (bacterial strain) One aspect of this disclosure is Microbacterium testaseum 1-39. Microbacterium testaseum 1-39 is deposited with the Patent Microorganism Depositary Center of the National Institute of Technology and Evaluation (NPMD) (2-5-8 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan), with receipt number NITE AP-03803 (deposit date: January 19, 2023).

[0016] Microbacterium testaseum 1-39 is a strain of bacteria collected from brown rice, as shown in the examples described below. When used in cooking rice, it can improve the quality of the resulting cooked rice, particularly its taste, appearance, and / or stickiness. This invention modifies the flavor components of rice using microorganisms, which differs significantly from the conventional concept of rice flavor, which is determined by a combination of genetic traits and environmental factors. This technology makes it possible to artificially modify the flavor components of rice after harvest. Furthermore, since the effect can be obtained by adding it during cooking, without depending on the genetic factors or nutritional status of the rice, the time required to obtain high-quality rice can be significantly reduced.

[0017] Culturing of Microbacterium testaceum 1-39 can be performed aerobically. The culturing can be carried out by, for example, shake culturing. The medium is not particularly limited as long as the strain can be cultured, and for example, a TSB liquid medium can be used. The culturing temperature may be, for example, 20 to 35°C, preferably 25 to 30°C, and more preferably 27 to 29°C or 28°C. The culturing time may be, for example, 10 to 72 hours, 12 to 36 hours, or 18 to 24 hours.

[0018] (Cooked rice improving agent) One aspect of the present disclosure is a cooked rice improving agent comprising Microbacterium testaceum 1-39. Since the cooked rice improving agent according to the present embodiment comprises Microbacterium testaceum 1-39, use of the agent in rice cooking can improve the quality of the resulting cooked rice, for example, the taste, appearance, and stickiness of the cooked rice.

[0019] The cooked rice improving agent may consist solely of Microbacterium testaceum 1-39, and in addition to Microbacterium testaceum 1-39, may also contain a carrier, an excipient, a surfactant, a binder, a dispersant, a thickener, a preservative, an antioxidant, a pH adjuster, an organic acid, an amino acid, common salt, a sweetener, a coloring agent, a flavoring agent, and the like, and may also contain the medium in which the bacterial cells have been cultured. The cooked rice improving agent may be formulated into dosage forms such as, for example, powders, granules, liquids, emulsions, oils, suspensions, pastes, and capsules. The cooked rice improving agent may contain viable cells or dead cells of Microbacterium testaceum 1-39, and preferably contains viable cells. The cooked rice improving agent may contain microorganisms other than Microbacterium testaceum 1-39. The cooked rice improving agent can be produced, for example, by mixing Microbacterium testaceum 1-39 with other components as necessary. As the cooked rice improving agent, the culture of Microbacterium testaceum 1-39 may be used as it is, the bacterial cells isolated after culturing may be used, or a dried product of the culture or the bacterial cells may be used.

[0020] The amount of Microbacterium testaceum 1-39 contained in the cooked rice modifier according to the present embodiment may be, relative to the total amount of the cooked rice modifier, for example, 0.001 mass% or more, 0.01 mass% or more, 0.1 mass% or more, 1 mass% or more, 5 mass% or more, 10 mass% or more, 20 mass% or more, 30 mass% or more, 50 mass% or more, 70 mass% or more, 80 mass% or more, 90 mass% or more, 95 mass% or more, 99 mass% or more, or 100 mass%, and may be 100 mass% or less, 99 mass% or less, 95 mass% or less, 90 mass% or less, 80 mass% or less, 70 mass% or less, 50 mass% or less, 30 mass% or less, 20 mass% or less, 10 mass% or less, 5 mass% or less, 1 mass% or less, or 0.1 mass% or less.

[0021] The rice to which the cooked rice modifier is applied may be, for example, white rice, partially polished rice, or brown rice, and may be a mixture thereof. The partially polished rice may be, for example, 10% polished or more, 20% polished or more, 30% polished or more, 40% polished or more, 50% polished or more, 60% polished or more, 70% polished or more, 80% polished or more, or 90% polished or more. The rice to which the cooked rice modifier is applied may be non-glutinous rice or glutinous rice. The rice may be of the Japonica variety, Indica variety, or Javanica variety, with the Japonica variety being preferred. The rice variety to which the cooked rice modifier is applied is not particularly limited.

[0022] The cooked rice modifier according to the present embodiment can be used, for example, by adding it to rice before cooking, rice soaking water, or cooking water. The cooked rice modifier may be added, for example, to the rice or the soaking water after washing the rice and before soaking the rice, or may be added to the rice or the cooking water after soaking the rice. Rice cooking is preferably carried out in a state where the cooked rice modifier is present in the cooking water during rice cooking.

[0023] The usage amount of the cooked rice modifier is, as the amount of bacterial cells relative to 100 g of rice, for example, 10 5 cells to 10 9 cells. Specifically, for example, a liquid in which the cooked rice modifier is suspended, adjusted to have an OD 600 of 1.0 in terms of the concentration of Microbacterium testaceum 1-39, may be added in an amount of 10 μL to 10 mL, 50 μL to 5 mL, or 80 μL to 1 mL relative to 100 g of rice.

[0024] When cooking rice using a rice conditioning agent, soaking the rice before cooking is not mandatory, but it is preferable to soak the rice from the viewpoint of improving the quality of the resulting cooked rice. Soaking should be done for the time and temperature normally used for soaking before cooking, and the soaking temperature may be, for example, 10-40°C, 15-35°C, or 20-30°C. The soaking time may be, for example, 5 minutes or more, 10 minutes or more, 30 minutes or more, 1 hour or more, or 1.5 hours or more, and may be 6 hours or less, 4 hours or less, 3 hours or less, 2.5 hours or less, or 2 hours or less.

[0025] The amount of water used when cooking rice may be, for example, 1.1 to 10 times, 1.1 to 7 times, 1.1 to 3 times, or 1.1 to 2 times the amount of raw rice. To more favorably obtain the modification effect of the rice modifier, it is preferable that the amount of water used when cooking rice is 1.1 to 1.4 times the amount of raw rice. Rice can be cooked by conventional methods.

[0026] The rice to which the rice modifier is applied may contain, for example, grains other than rice, chestnuts, beans, other ingredients, and seasonings.

[0027] (Method of producing cooked rice) One aspect of this disclosure is a method for cooking rice using Microbacterium testaseum 1-39, i.e., a method for producing cooked rice. The method for producing cooked rice according to this embodiment includes cooking rice with the addition of Microbacterium testaseum 1-39. In this specification, cooked rice refers to rice that has been cooked and is ready to eat.

[0028] In the method for producing cooked rice using Microbacterium testaseum 1-39, the aspects of adding the strain, rice, soaking, and cooking methods can be adapted to the aspects of the cooked rice modifier described above. Microbacterium testaseum 1-39 may be added as a cooked rice modifier or as a component of a cooked rice modifier. The method for producing cooked rice according to this embodiment may include soaking the rice. [Examples]

[0029] The present invention will be described in more detail below with reference to examples. However, the present invention is not limited to the following examples.

[0030] [Sample collection, amplicon sequencing] Soil, rhizosphere, roots, or brown rice samples were collected from nine Haenuki rice cultivation sites in Yamagata Prefecture. Genomic DNA of microorganisms was extracted and purified from each sample, and the 16S rRNA gene was sequenced using the MiSeq sequencer. The microbial community was then analyzed using QIIME2.

[0031] We conducted sensory evaluation tests on rice grown at each sampling site and analyzed the correlation between the microbial flora and the overall taste score of the rice. As a result, we found that the higher the overall taste score of the rice, the higher the proportion of Actinobacteria bacteria in the microbial flora of the brown rice.

[0032] [Network Analysis] For each item of rice composition such as amylose content, weather conditions such as temperature during growth, soil composition, microbial flora, and rice taste values ​​such as taste and aroma, we extracted items with an absolute value of 0.4 or higher for Pearson's product-moment correlation coefficient, and further extracted items with a p-value of less than 0.05 using a t-test. As a result, it was found that there was a high correlation between the presence of Actinobacteria bacteria and the aroma, taste, hardness, and overall taste value of rice.

[0033] [Rice Cooking Test] Of the bacteria collected from brown rice during the sample collection described above, Actinobacteria strains X, Y, and Z, Sphingomonas strain X, and Psudomonas strain X, which were successfully cultured, were used in the rice cooking test described below. Of these, Actinobacteria strain X was identified as Microbacterium testaseum by whole-genome analysis, and was designated strains 1-39.

[0034] 1. Culturing bacteria The bacterial strains, stored in a -80°C freezer within a clean bench, were lightly poked with a Nunc® disposable platinum loop to allow a small amount of the strain to adhere to the tip of the loop, and then streaked onto 50% TSB agar. Subsequently, to prevent contamination, the petri dish was sealed with Micropore® Surgical Tape, and streak culture was performed in an incubator (Panasonic) at 28°C for 60 hours.

[0035] A single colony was punctured using a sterile toothpick (Yanagi Products Co., Ltd.), and the toothpick and colony were placed in a 14 mL Falcon® polypropylene round tube (Corning Corporation) containing 3 mL of 50% TSB liquid medium. The mixture was then vortexed using a vortex mixer (Kenis Corporation) to break up the colony. Afterwards, the mixture was incubated in a benchtop shaking incubator (TAITEC Corporation) at 28°C for 170 min. -1 The cells were then cultured in liquid for 20 hours.

[0036] 2. Measurement and homogenization of turbidity by bacteria After culturing, 1 mL of culture medium was added to a disposable cell in a clean bench, and the absorbance at a wavelength of 600 nm was measured using a spectrophotometer (Thermo Scientific). The sample was then diluted with sterile water (Nacalai Tesque) to an OD of 1.0, and the measurement was repeated to homogenize the turbidity.

[0037] 3. Preparing the rice 20g of 90% polished rice (Nipponbare) and 1.3 times the amount of RO water were added to an aluminum pudding cup (55mm diameter at the top, 40mm diameter at the bottom, 55mm depth), and the rice was washed by rotating the stainless steel cup 2-3 times. After that, the RO water was discarded, and this process was repeated 5 times. After washing the rice, the total weight of the polished rice and RO water was adjusted to 46g using an electronic balance calibrated with an empty stainless steel cup. 20μL of bacterial solution (20μL of 50% TSB for the control) was added to each of the adjusted stainless steel cups, and the tops of the aluminum pudding cups were covered and sealed with a 200×200mm aluminum sheet.

[0038] 4.Soaking The aluminum pudding cups, sealed with aluminum foil, were placed at equal intervals in a rice cooker pressure cooker (Panasonic SR-LU181) containing 150 mL of RO water, and left to stand for 2 hours at 28°C in an incubator (TAITEC MULTI SHAKER OVEN HB).

[0039] 5. Cooking rice The pressure cooker, removed from the incubator, was placed in a rice cooker (Panasonic SR-LU181), the setting was changed to white rice / pre-washed rice, and the rice was cooked for 55 minutes. After cooking, the aluminum pudding cups were removed from the rice cooker and allowed to cool on a tabletop at 25°C for 1 hour to obtain cooked rice.

[0040] 6. Rice Cooker Taste Measurement Using a spatula, the cooked rice was transferred to a petri dish covered with Saran® plastic wrap, ensuring that the part that had been in contact with the bottom of the aluminum pudding cup was facing upwards. Then, a slit was made in the top of the cooked rice with the spatula, and the rice was spread out. Finally, the dish was sealed with plastic wrap to prevent drying until measurement.

[0041] Before measuring the cooked rice with the Rice Taste Meter (trademark, SATAKE, STA1A), 8g of cooled cooked rice was packed into the measuring metal ring of the Rice Taste Meter. The lever of the hand press attached to the Rice Taste Meter was pushed forward, and the cooked rice was compressed for 10 seconds. The ring was turned over and placed on the measuring jig, and compressed again on the press table for 10 seconds. The compressed cooked rice was inserted into the Rice Taste Meter, and its taste, appearance, and stickiness were measured. The sample cell was removed from the Rice Taste Meter, the ring was turned over, and the measurement was taken again. This was repeated three times (three times on each side, for a total of six times). The same measurement was performed in three repetitions, and the average value was calculated from the total measurement data of 18 times, and this was taken as the result for the Rice Taste Meter for the sample.

[0042] 7.Results Box plots were created using R ver4.1.2 for the taste, appearance, and stickiness results from the rice cooking taste meter, and significance testing was performed using the Kruskal-Wallis test. The results are shown in Figures 1-3. Figure 1 shows the taste score, Figure 2 shows the appearance score, and Figure 3 shows the stickiness score. In all three categories—taste, appearance, and stickiness—only Microbacterium testaseum 1-39 showed a significant difference (p<0.05) compared to the control. [Accession Number]

[0043] (1) Name of depositary institution: NPMD, National Institute of Technology and Evaluation, Patent Microbial Depository Center (2) Contact information: 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture 292-0818 (3) Receipt number: NITE AP-03803 (4) Identification markings: 1-39 (5) Entrustment date: January 19, 2023

Claims

1. Microbacterium testaseum 1-39 (receipt number NITE AP-03803).

2. A method for producing cooked rice, comprising adding the bacteria described in claim 1 and cooking the rice.

3. A rice additive containing the bacteria described in claim 1.

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