Candida utilis with high efficiency of converting ethanol and acetic acid under stress conditions and application thereof

Abstract

The application provides a Candida utilis for efficiently converting ethanol and acetic acid under stress conditions and an application thereof, and belongs to the technical field of microorganisms. The Candida utilis is Candida utilis TU389-37XHZ, which is preserved in the China General Microbiological Culture Collection Center and has a preservation number of CGMCC No.39159. The Candida utilis TU389-37XHZ is a domesticated strain capable of tolerating high temperature and low pH at the same time. The strain can be used for treating distiller's wastewater or bioethanol production wastewater and producing single-cell protein. The domesticated strain provided by the application can efficiently convert ethanol and acetic acid under stress conditions of 37 DEG C high temperature and low pH, so as to realize resource utilization of the wastewater and production of the single-cell protein.

Description

Technical Field

[0001] This invention relates to the field of microbial technology. More specifically, this invention relates to a Candida utilis strain that efficiently converts ethanol and acetic acid under stress conditions and its applications. Background Technology

[0002] Candida utilis, a microorganism suitable for single-cell protein (SCP) production and the resource utilization of organic waste, has a certain application foundation in the food, feed, fermentation, and wastewater treatment industries. In practical applications such as brewing and bioethanol production, the wastewater or byproduct systems often contain carbon sources that can be utilized by microorganisms, such as ethanol, acetic acid, or acetate. However, conditions such as temperature, pH, and substrate composition vary under different production environments. Currently, methods for utilizing microorganisms to convert organic carbon sources include methods primarily using a single carbon source or methods primarily using a combination of carbon sources. Different methods exhibit differences in the environmental adaptability of strains, carbon source utilization efficiency, fermentation stability, and the effect of microbial protein accumulation in practical applications. When the treated object has characteristics such as high temperature, strong acidity, or complex substrate composition, the growth performance and conversion efficiency of conventional strains are easily affected, thus limiting their application in relevant industrial scenarios.

[0003] Wild-grown Candida utilis exhibits weak tolerance to high temperatures, with its optimal growth temperature typically concentrated between 30-32℃. Studies have shown that around 32℃ is the suitable temperature for biomass and protein accumulation in Candida utilis. However, at temperatures of 37℃ and above, strain growth is inhibited, and biomass decreases significantly, limiting its application in high-temperature fermentation systems. Furthermore, wild-grown strains are intolerant of pH environments below 5.5; therefore, their conversion efficiency, sustainable utilization, and application adaptability remain insufficient for complex systems containing components such as ethanol, acetic acid, or acetates. Summary of the Invention

[0004] In view of the above, in order to at least partially solve at least one of the aforementioned technical problems, the present invention provides a Candida utilis strain that efficiently converts ethanol and acetic acid under stress conditions and its application, wherein the Candida utilis strain of the present invention, TU389-37XHZ, is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 39159 and deposit date of March 16, 2026.

[0005] According to one embodiment of the present invention, an application of Candida utilis TU389-37XHZ, a yeast strain that efficiently converts ethanol / acetic acid under stress conditions, is provided, comprising at least one of the following: (1) converting ethanol; (2) converting acetic acid; (3) co-converting ethanol and acetic acid; (4) producing single-cell protein; (5) treating wastewater containing ethanol, acetic acid and / or acetate; (6) treating acidic wastewater; (7) treating brewing wastewater; and (8) treating bioethanol production wastewater.

[0006] According to another aspect of the present invention, an application of Candida utilis TU389-37XHZ, which is highly efficient at converting ethanol / acetic acid under stress conditions, is provided, including at least one of the following: (1) for food processing; (2) for feed processing.

[0007] According to another embodiment of the present invention, a method for producing single-cell protein is provided, comprising the step of co-culturing Candida utilis TU389-37XHZ with a carbon source fermentation broth, wherein the carbon source fermentation broth comprises ethanol and acetic acid.

[0008] According to another embodiment of the present invention, a single-cell protein is provided, which is prepared by the method described above for producing single-cell proteins from Candida utilis TU389-37XHZ.

[0009] According to another embodiment of the present invention, a method for treating wastewater is provided, comprising the step of inoculating the above-mentioned Candida utilis TU389-37XHZ into the wastewater for cultivation.

[0010] According to another embodiment of the present invention, a composition is provided containing Candida utilis TU389-37XHZ.

[0011] Compared with the prior art, the present invention has the following beneficial technical effects:

[0012] The *Candida utilis* TU389-37XHZ strain that efficiently converts ethanol / acetic acid under stress conditions according to the present invention was obtained through continuous adaptive domestication using ethanol and acetic acid as carbon sources. This process enabled the strain to develop good high-temperature adaptability and tolerance to acidic environments, maintaining good growth and fermentation performance at 37°C and pH 3. It is suitable for industrial fermentation systems with high temperatures or strong acidity, which helps reduce the need for cooling and acid-base regulation during fermentation, thereby reducing the difficulty of process control and operating costs.

[0013] Meanwhile, *Candida utilis* TU389-37XHZ can utilize a combined carbon source of ethanol and acetic acid for metabolic transformation, providing a strain basis for the resource utilization of systems containing ethanol, acetic acid, or acetate, such as brewing wastewater and bioethanol production wastewater. This strain can maintain its ability to utilize the ethanol-acetic acid carbon source even under low pH conditions, which is beneficial for adapting to acidic industrial wastewater environments and reducing or simplifying neutralization pretreatment steps.

[0014] Furthermore, this strain exhibits good performance in biomass accumulation and cell protein formation. Under optimal conditions of 37℃ and pH 3.5, the fermentation biomass reached 35.18 g / L after 72 hours, the cell protein content reached 41.57%, and the protein yield reached 14.62 g / L, representing an increase of approximately 10.9% compared to the starting strain under pH 6.0 conditions. Therefore, this strain can simultaneously form microbial biomass containing cell protein during the conversion of organic carbon sources, which is beneficial for improving single-cell protein production efficiency and expanding the high-value utilization pathways of industrial by-products such as ethanol and acetic acid.

[0015] Furthermore, Candida utilis TU389-37XHZ has both adaptability within a temperature range of 31℃ to 37℃ and tolerance within a pH range of 3.5 to 6.0, which can reduce the impact of temperature and pH fluctuations on strain growth and carbon source conversion during industrial fermentation, and improve the stability of the fermentation process. It is especially suitable for open fermentation systems, acidic organic wastewater treatment systems, and complex industrial raw material conversion systems. Attached Figure Description

[0016] The above and other objects, features and advantages of the present invention will become more apparent from the following description of embodiments of the invention with reference to the accompanying drawings, in which:

[0017] Figure 1 The diagram shows growth curves of strains during the acclimatization stage and biomass graphs of different strains under different culture conditions in embodiments of the present invention. Specifically, A is the growth curve of *Candida utilis* TU389 under high-temperature acclimatization from generation 0 to 131; B is the growth curve of *Candida utilis* X131 under low-pH acclimatization from generation 0 to 60; C is the biomass graph of different strains fermented in shake flasks; and D is the biomass graph of different strains fermented in a 5L fermenter. W-31 / 6 represents the wild type (31℃, pH...). H6.0), W-37 / 6 represents the wild type (37℃, pH 6.0), X-31 / 6 represents the high-temperature acclimatized strain X131 (31℃, pH 6.0), X-37 / 6 represents the high-temperature acclimatized strain X131 (37℃, pH 6.0), X-37 / 3.5 represents the high-temperature acclimatized strain X131 (37℃, pH 3.5), and P-37 / 3.5 represents the final acclimatized strain P-37 / 3.5 (37℃, pH 3.5).

[0018] Figure 2 The figures show the carbon source conversion rate (g / g), protein content (%), and protein yield (g / L) of different strains fermented in a 5L fermenter under different culture conditions according to embodiments of the present invention. In the figures, A is the carbon source conversion rate, B is the protein content, and C is the protein yield.

[0019] Figure 3 The diagram shows principal component analysis (PCA) plots of different strains of the present invention at different temperatures, where A is a scatter plot of PCA for different groups of strains and B is a grouping plot of PCA for different groups of strains.

[0020] Figure 4 The figures show bar charts and Venn diagrams of the number of differentially expressed genes (DEGs) in three comparison groups (X-31 vs W-31, X-37 vs W-37, X-37 vs W-31) according to embodiments of the present invention. A is a bar chart of the number of differentially expressed genes in different comparison groups, and B is a Venn diagram of the number of differentially expressed genes in different comparison groups.

[0021] Candida utilis TU389-37XHZ is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 39159 on March 16, 2026.

[0022] Wild-type Candida utilis strain TU389 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 29510 and deposit date of January 4, 2024. Detailed Implementation

[0023] Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings. However, it should be understood that these descriptions are exemplary only and are not intended to limit the scope of the invention. In the following detailed description, numerous specific details are set forth to provide a thorough understanding of the embodiments of the invention for ease of explanation. However, it will be apparent that one or more embodiments may be practiced without these specific details. Furthermore, descriptions of well-known structures and techniques are omitted in the following description to avoid unnecessarily obscuring the concept of the invention.

[0024] The terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the invention. The term "comprising" as used herein indicates the presence of features, steps, or operations, but does not exclude the presence or addition of one or more other features.

[0025] When using expressions such as "at least one of A, B, and C," the expression should generally be interpreted in accordance with the meaning commonly understood by a person skilled in the art (e.g., "a system having at least one of A, B, and C" should include, but is not limited to, a system having A alone, a system having B alone, a system having C alone, a system having A and B, a system having A and C, a system having B and C, and / or a system having A, B, and C, etc.). When using expressions such as "at least one of A, B, or C," the expression should generally be interpreted in accordance with the meaning commonly understood by a person skilled in the art.

[0026] In realizing the concept of this invention, it was discovered that *Candida utilis*, a microorganism suitable for single-cell protein production and the resource utilization of organic waste, possesses good carbon source utilization and protein synthesis capabilities. However, in practical applications such as industrial fermentation and wastewater treatment, it is often affected by factors such as increased temperature, acidic environments, and complex carbon source conditions like ethanol and acetic acid, which limit cell growth, carbon source conversion, and protein accumulation. Compared to simply relying on external process control, obtaining strains with better environmental adaptability and carbon source conversion capabilities is more beneficial for improving the stability of the fermentation process and the efficiency of resource utilization. Therefore, screening and obtaining *Candida utilis* strains suitable for systems containing ethanol, acetic acid, or acetate, and capable of maintaining good growth and protein accumulation capabilities under stress conditions, has become the technical focus of this invention.

[0027] Specifically, according to one embodiment of the present invention, a Candidautilis TU389-37XHZ is provided.

[0028] Information on the preservation of biological material samples:

[0029] Strain name: Candida utilis;

[0030] Strain number: TU389-37XHZ;

[0031] Preservation period: March 16, 2026;

[0032] Preservation Center: China General Microbiological Culture Collection Center (CGMCC);

[0033] Address: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing;

[0034] Registration number at the Preservation Center: CGMCC No. 39159.

[0035] According to an embodiment of the present invention, a wild-type Candida utilis strain (deposited at the China General Microbiological Culture Collection Center, accession number: CGMCCNo.29510, deposit date: January 4, 2024, deposit address: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing) was continuously adapted and domesticated using ethanol and acetic acid as carbon sources. The obtained Candida utilis TU389-37XHZ exhibits good high-temperature adaptability and tolerance to acidic environments. The domesticated strain maintains good fermentation performance at 37℃ and pH 3.5, with a fermentation biomass of 35.18 g / L after 72 hours, a cell protein content of 41.57%, and a protein yield of 14.62 g / L. This strain is applicable to systems containing ethanol, acetic acid, or acetate, such as brewing wastewater and bioethanol production wastewater. It accumulates single-cell proteins while converting organic carbon sources, which is conducive to the simultaneous utilization of wastewater resources and preparation of protein products. It provides a microbial resource and biotransformation pathway with industrial application potential for the treatment of acidic organic wastewater and the production of single-cell proteins.

[0036] According to one embodiment of the present invention, an application of Candida utilis TU389-37XHZ, a yeast strain that efficiently converts ethanol / acetic acid under stress conditions, is provided, comprising at least one of the following: (1) converting ethanol; (2) converting acetic acid; (3) co-converting ethanol and acetic acid; (4) producing single-cell protein; (5) treating wastewater containing ethanol, acetic acid and / or acetate; (6) treating acidic wastewater; (7) treating brewing wastewater; and (8) treating bioethanol production wastewater.

[0037] According to an embodiment of the present invention, *Candida utilis* TU389-37XHZ can utilize organic carbon sources such as ethanol, acetic acid, or acetate for growth and metabolic transformation under stress conditions, and simultaneously accumulate cell protein. Specifically, this strain can utilize ethanol or acetic acid as a carbon source, or can achieve synergistic transformation of multiple carbon sources when both are present. Synergistic transformation refers to the strain's joint utilization and metabolic transformation of ethanol and acetic acid in the same fermentation system, enabling both types of C2 carbon sources to participate in cell growth and cell protein formation, thereby converting the system containing ethanol, acetic acid, or acetate into cell biomass and single-cell protein.

[0038] Furthermore, this strain can be used to treat wastewater containing organic carbon sources, such as acidic wastewater, brewing wastewater, and bioethanol production wastewater. Brewing wastewater refers to wastewater containing organic components such as ethanol, acetic acid, and / or acetates generated during the production, fermentation, distillation, and washing of baijiu, beer, huangjiu, wine, or other alcoholic beverages. Bioethanol production wastewater refers to wastewater containing organic components such as ethanol, acetic acid, and / or acetates generated during the production of fuel ethanol or industrial ethanol from starchy, sugary, cellulose, or other fermentable raw materials. Through the above treatment, this strain can reduce the organic carbon source load in wastewater and form microbial biomass containing bacterial protein. The obtained bacterial cells, after collection and treatment, can be used as single-cell protein products or single-cell protein raw materials for the preparation of microbial protein products, fermented protein products, protein supplement raw materials, nitrogen supplement raw materials, or organic waste resource products, providing a microbial resource basis for the high-value treatment of organic wastewater and the development of single-cell protein products.

[0039] According to another embodiment of the present invention, an application of Candida utilis TU389-37XHZ, which is highly efficient at converting ethanol / acetic acid under stress conditions, is provided, including at least one of the following: (1) for food processing; (2) for feed processing.

[0040] According to an embodiment of the present invention, the bacterial cells obtained from the fermentation culture of *Candida utilis* TU389-37XHZ contain bacterial protein. After collection, drying, pulverizing, or other suitable treatments, the bacterial cells can form single-cell protein products or their raw materials. The strain efficiently synthesizes bacterial protein under high temperature and low pH conditions, with higher protein content and yield than the wild type. Furthermore, the amino acid composition meets the nutritional requirements of animals. After drying and pulverizing, it can be directly used as a feed additive. The strain has a protein content as high as 41.57% and a balanced amino acid composition (total amino acid content 36.10 g / 100 g), making it a high-quality protein source that can replace traditional protein raw materials such as fishmeal and soybean meal, reducing feed costs and alleviating protein resource shortages.

[0041] According to another embodiment of the present invention, a method for producing single-cell protein is provided, comprising the step of co-culturing Candida utilis TU389-37XHZ with a carbon source fermentation broth, wherein the carbon source fermentation broth comprises ethanol and acetic acid in a ratio of (9:1) to (1:9), preferably, the ratio of ethanol to acetic acid is 2:1.

[0042] According to an embodiment of the present invention, the conditions for co-culturing Candida utilis TU389-37XHZ and the carbon source fermentation broth further include a carbon source fermentation broth pH of 3-9 and a carbon source fermentation broth temperature of 25℃-45℃. For example, the carbon source fermentation broth pH can be 3, 4, 6, 7, or 9; and the carbon source fermentation broth temperature can be 25℃, 27℃, 29℃, 33℃, 37℃, 41℃, or 45℃. Preferably, the carbon source fermentation broth pH is 3-6 and the carbon source fermentation broth temperature is 31℃-37℃. For example, the carbon source fermentation broth pH can be 3, 4, 5, or 6; and the carbon source fermentation broth temperature can be 31℃, 32℃, 34℃, 35℃, 36℃, or 37℃. Specifically, *Candida utilis* TU389-37XHZ was used as the production strain, with ethanol and acetic acid as carbon sources. Fermentation was carried out at 37°C and pH 3–9, and the cells were collected to obtain single-cell protein. This method requires no additional pH adjustment and consumes no energy for cooling. The fermentation biomass reached 35.18 g / L after 72 hours, with a protein content of 41.57%.

[0043] According to embodiments of the present invention, the pH and temperature of the carbon source fermentation broth can affect the growth status, carbon source conversion capacity, and cell protein accumulation of *Candida utilis* TU389-37XHZ. Maintaining the pH and temperature of the carbon source fermentation broth within a suitable range helps maintain the metabolic activity of the cell cells, promotes their utilization of ethanol / acetic acid, and improves the microbial biomass and cell protein formation efficiency during fermentation. Specifically, when the pH of the carbon source fermentation broth is 3–6 and the temperature is 31°C–37°C, this strain can still maintain good environmental adaptability and fermentation stability, thus making it suitable for single-cell protein production using ethanol and acetic acid as carbon sources.

[0044] According to another embodiment of the present invention, a single-cell protein produced by the above-described single-cell protein production method is provided. The bacterial protein obtained by the above-described method has a high content, reaching up to 41.57%, and possesses a relatively balanced amino acid composition, making it suitable for use as a microbial protein product or protein raw material. The single-cell protein originates from *Candida utilis* cells and, in addition to bacterial protein, may also contain amino acids, nucleic acids, vitamins, and other nutrients formed by the microbial cells themselves, thus possessing a good nutritional composition. Therefore, single-cell protein can be used as a protein supplement raw material, a fermented protein product, a microbial protein raw material, or a nitrogen source supplement raw material, providing a stable product foundation for the development of related protein products.

[0045] According to another embodiment of the present invention, a method for treating wastewater is provided, comprising the step of inoculating and culturing Candida utilis TU389-37XHZ into the wastewater.

[0046] According to an embodiment of the present invention, the wastewater includes at least one of ethanol, acetic acid, and acetate. After inoculating the wastewater with Candida utilis TU389-37XHZ, the strain can utilize organic components such as ethanol, acetic acid, or acetate in the wastewater for growth and metabolic transformation, thereby reducing the organic carbon source content in the wastewater and obtaining treated wastewater.

[0047] According to an embodiment of the present invention, cultivation can be carried out at a temperature of 37°C and a pH of 3.5–6.0. ​​Because *Candida utilis* TU389-37XHZ exhibits good high-temperature adaptability and tolerance to acidic environments, it can still maintain its ability to utilize ethanol, acetic acid, or acetate in acidic or high-temperature wastewater environments. Therefore, this method can be used for the biological treatment of wastewater containing ethanol, acetic acid, or acetate, and is particularly suitable for the treatment of organic wastewater such as brewing wastewater and bioethanol production wastewater. It is beneficial for improving the conversion efficiency of organic components in wastewater and reducing the adverse effects of changes in wastewater acidity or temperature on the treatment effect.

[0048] According to another embodiment of the present invention, a composition is provided containing Candida utilis TU389-37XHZ.

[0049] According to embodiments of the present invention, the composition can use *Candida utilis* TU389-37XHZ as the active ingredient or main functional ingredient, enabling the composition to utilize organic carbon sources such as ethanol, acetic acid, and / or acetate for growth and metabolic transformation. The composition can be in the form of a microbial agent, and can be used in fermentation, wastewater treatment, or single-cell protein preparation scenarios according to actual application needs. This improves the convenience of the strain during transportation, preservation, inoculation, and use, and enhances its adaptability for engineering applications.

[0050] The *Candida utilis* TU389-37XHZ strain obtained in this invention possesses the ability to grow and metabolize using organic carbon sources such as ethanol, acetic acid, and / or acetate under stress conditions. This strain is suitable for fermentation systems using ethanol and acetic acid as carbon sources and maintains good fermentation performance under relatively high temperatures and acidic conditions. According to an embodiment of the present invention, when fermented at 37°C and pH 3.5 for 72 hours, the biomass of this strain can reach 35.18 g / L, the cell protein content can reach 41.57%, and the protein yield can reach 14.62 g / L, representing an increase of approximately 10.9% in protein yield compared to the starting strain under pH 6.0 conditions.

[0051] Furthermore, the *Candida utilis* TU389-37XHZ strain provided by this invention can be used to treat wastewater containing ethanol, acetic acid, and / or acetate, and is particularly suitable for organic wastewater systems such as brewing wastewater and bioethanol production wastewater. This strain can utilize organic carbon sources in the wastewater for metabolic transformation, which helps to reduce the content of related organic components in the wastewater and improves the adaptability and stability of wastewater biological treatment.

[0052] The present invention will be further explained below through specific embodiments. Unless otherwise specified, all reagents used are commercially available reagents and all experimental methods used are conventional experimental methods in the art.

[0053] Example 1

[0054] 1.1 Strain origin and isolation identification

[0055] The wild-type strain used in this invention was isolated from barley samples. Under aseptic conditions, 10g of barley sample was taken, washed thoroughly with sterile physiological saline, and the supernatant was collected after standing. 1mL of the sample solution was inoculated into 100mL of enrichment medium (malt extract medium with added lactic acid to inhibit bacterial growth) and cultured at 28–30℃ and 150–200 rpm for 24–48 hours with shaking. These enrichment conditions allow acid-tolerant yeasts such as *Candida utilis* to proliferate rapidly, while inhibiting the growth of most bacteria and molds.

[0056] The enriched culture was streaked onto malt extract agar plates or yeast extract peptone dextrose agar plates (YPD agar plates) and incubated upside down at 28–30°C for 48–72 hours. Typical yeast colonies that are milky white, smooth, moist, and have regular edges (sometimes appearing as hyphae) were selected for purification culture.

[0057] The purified strain was analyzed for 26S rDNA D1 / D2 region sequence and identified as *Candida utilis*, named *Candida utilis TU389*. It is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 29510, deposited on January 4, 2024, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0058] 1.2 Culture medium

[0059] Seed culture medium (g / L): glucose 10g / L, yeast extract 3g / L, malt extract 3g / L, peptone 5g / L, pH natural (6.0~6.5), autoclaved at 121℃ for 20min.

[0060] Shake-flask fermentation medium (g / L): (NH4)2SO4 5g / L, MgSO4 1g / L, KH2PO4 1.5g / L, CaCl2 0.5g / L, ZnSO4·7H2O 100mg / L, FeSO4·7H2O 30mg / L, anhydrous ethanol 6.7g / L, sodium acetate 3.3g / L, pH adjusted to 7; autoclave at 121℃ for 20min (ethanol and acetic acid are sterilized separately by filtration and added aseptically after the medium has cooled to below 50℃).

[0061] Fermentation medium for fermenter (g / L): The composite carbon source is ethanol-acetic acid (mass ratio 2:1), and the other components are the same as those for shake flask fermentation medium. The pH is adjusted to 6 with ammonia water. Ethanol and acetic acid are filtered and sterilized separately, and added aseptically after the medium has been sterilized and cooled.

[0062] Slant culture medium (g / L): glucose 20.0, yeast extract 3.0, peptone 5.0, agar 20.0, pH natural, autoclaved at 121℃ for 20 min.

[0063] Example 2: Temperature and pH acclimatization of Candida utilis

[0064] 2.1 High-temperature acclimatization

[0065] Domestication initiation: A single colony was picked from the slant culture strain *Candida utilis* TU389 and inoculated into 50 mL of seed culture medium. The culture was incubated at 31°C and 200 r / min for 18 h to obtain the seed culture. 10 mL of the seed culture was then inoculated into 100 mL of shake-flask fermentation medium and incubated at 31°C and 200 r / min for 48 h, serving as the first generation of domesticated strain. The strain after 131 generations of domestication was named *Candida utilis* X131.

[0066] Domestication process: The domestication temperature was 37℃; generations 1-27 used a 48-hour fermentation cycle for transfer, while generations 28-131 had a shortened 24-hour fermentation cycle to enhance screening pressure. The transfer ratio was 10% (v / v), i.e., 10 mL of the previous generation fermentation broth was inoculated into 100 mL of fresh fermentation medium and cultured at the corresponding temperature, 200 r / min, and pH 6.0.

[0067] Domestication index monitoring: After each generation of fermentation, the OD 600 value of the fermentation broth was measured using a UV-Vis spectrophotometer. Three biological replicates were set up for each sample, the average value was calculated, and a domestication curve was plotted to track changes in biomass.

[0068] 2.2 Low pH Acclimation

[0069] Based on the above-mentioned domesticated Candida utilis X131 that can withstand high temperatures, pH domestication was carried out.

[0070] Domestication initiation: A single colony of *Candida utilis* X131 was picked from the slant culture and inoculated into 50 mL of seed culture medium. The culture was incubated at 37°C and 200 r / min for 18 h to obtain the seed culture. 10 mL of the seed culture was inoculated into 100 mL of shake-flask fermentation medium and incubated at 37°C, 200 r / min, and pH 6.0 for 24 h, serving as the first generation of domesticated strain. After 60 rounds of domestication, the domesticated strain was obtained and named *Candida utilis* P-37 / 3.5 (i.e., strain TU389-37XHZ with accession number CGMCC No. 39159).

[0071] Domestication process: The initial pH of domestication was 6.0. Multiple rounds of gradual pH reduction were used, with each round lasting 24 hours. 10% (v / v) was transferred at a temperature of 37℃. The pH was eventually stabilized at 3.5. A total of 60 generations of continuous transfer domestication were carried out.

[0072] Domestication index monitoring: After each generation of fermentation, the OD600 value of the fermentation broth was measured using a UV-Vis spectrophotometer. Three biological replicates were set up for each sample, the average value was calculated, and a domestication curve was plotted to track changes in biomass.

[0073] 2.3 Evaluation of High-Temperature Acclimation and Shake-Flavor Fermentation of High-Temperature Acclimated Strains and Wild-Type Strains

[0074] Seed culture: Single colonies of the domesticated strain X131 (X) and the wild-type strain Candida utilis TU389 (W) after 131 generations of high-temperature domestication were selected and inoculated into 50 mL of seed culture medium. The culture was carried out at 31 °C and 200 r / min for 18 h. The OD of the bacterial solution was adjusted to 1.0 to obtain standardized seed culture.

[0075] Shake flask fermentation: Take 5 mL of standardized seed liquid and inoculate it into 50 mL of shake flask fermentation medium (250 mL Erlenmeyer flask). Set up two temperature groups of 31℃ and 37℃. Set up 3 replicates for each strain-temperature combination. Incubate at 200 r / min for 72 h.

[0076] 2.4 Fermentation Tank Scale-up Experiment

[0077] Seed culture: The seed culture method is the same as that for shake flask fermentation in 2.3, and the culture is expanded to 200 mL of standardized seed solution (OD600=1.0).

[0078] Fermentation in a 5L fermenter with a liquid volume of 3L and an inoculum size of 10% (v / v). Two temperature groups were set up: 31℃ and 37℃. The aeration rate was 1.0~1.5 vvm, and the stirring speed was 300~500 r / min. The pH was maintained at 6.0±0.2 by automatically adding 2mol / L HCl or ammonia. A dissolved oxygen cascade fed-batch fermentation mode was used. Carbon source consumption was monitored in real time by HPLC, and ethanol-acetic acid mother liquor (mass ratio 2:1) was added to maintain the total carbon source concentration of the fermentation broth at 5~8 g / L. Three replicates were set up for each strain-temperature combination.

[0079] Fermentation process monitoring: Samples were taken at 0, 12, 24, 30, 36, 48, 51, 58, and 72 h of fermentation to determine the OD600 value and biomass dry weight; the consumption of ethanol and acetic acid was measured and calculated, and the carbon source conversion rate (biomass dry weight / total carbon source consumed × 100%) was calculated.

[0080] Sample collection: After 72 hours of fermentation, cell samples of each strain-temperature combination were collected, centrifuged (8000 r / min, 10 min) to collect the precipitate, washed three times with pre-cooled deionized water, some samples were used for protein content determination, and the remaining samples were quickly frozen at -80℃ for transcriptome analysis.

[0081] 2.5 Evaluation of low-pH acclimatization shake-flask fermentation of low-pH acclimatized strains

[0082] Seed culture: Select single colonies of low pH acclimatized strain P-37 / 3.5 after 60 generations of low pH acclimatization, inoculate them into 50 mL of seed culture medium, and incubate at 31℃ and 200 r / min for 18 h. Adjust the bacterial OD of the solution to 1.0 to obtain standardized seed culture.

[0083] Shake flask fermentation: Take 5 mL of standardized seed culture and inoculate it into 50 mL of shake flask fermentation medium (250 mL Erlenmeyer flask). Set the culture temperature to 37℃, pH 3.5, set 3 replicates, and culture at 200 r / min for 72 h.

[0084] 2.6 Fermentation Tank Scale-up Experiment

[0085] Seed culture: The seed culture method is the same as that of shake flask fermentation in 2.5, and the culture is expanded to 200 mL of standardized seed liquid (OD600=1.0).

[0086] Fermentation in a 5L fermenter with a liquid volume of 3L and an inoculum size of 10% (v / v). During fermentation, the temperature was maintained at 37℃, the aeration rate at 1.0~1.5 vvm, and the stirring speed at 300~500 r / min. The pH was maintained at 3.5±0.2 by automatic addition of 2mol / L HCl or ammonia. A dissolved oxygen cascade fed-batch fermentation mode was used, and carbon source consumption was monitored in real time by HPLC. Ethanol-acetic acid mother liquor (mass ratio 2:1) was added to maintain a total carbon source concentration of 5~8 g / L in the fermentation broth. Three replicates were set up for each strain-pH combination.

[0087] Fermentation process monitoring: Samples were taken at 0, 12, 24, 30, 36, 48, 51, 58, and 72 h of fermentation to determine the OD600 value and biomass dry weight; the consumption of ethanol and acetic acid was measured and calculated, and the carbon source conversion rate (biomass dry weight / total carbon source consumed × 100%) was calculated.

[0088] Sample collection: After 72 hours of fermentation, cell samples of each strain-pH combination were collected, centrifuged (8000 r / min, 10 min) to collect the precipitate, washed three times with pre-cooled deionized water, some samples were used for protein content determination, and the remaining samples were quickly frozen at -80℃ for transcriptome analysis.

[0089] Determination methods for ethanol and acetic acid

[0090] Ethanol and acetic acid were determined by gas chromatography using a gas chromatograph (Tianmei Technology Co., Ltd., Shanghai, China) equipped with an HP InnoWax capillary column (Agilent Technologies, California, USA).

[0091] Place the sample to be tested in a centrifuge tube and centrifuge at a suitable speed (e.g., 8000 rpm, 4℃) for 10 min to remove solid impurities such as bacterial residue. Take the supernatant after centrifugation, draw it up with a disposable syringe, filter it through a 0.22 μm organic phase filter membrane into a clean sample bottle, and obtain cell-free supernatant for testing.

[0092] Gas chromatograph injection port temperature: 200℃, detector temperature: 250℃, column temperature program: initial temperature: 60℃, hold for 1 min; increase to 100℃ at a rate of 10℃ / min, hold for 2 min; then increase to 200℃ at a rate of 30℃ / min, hold for 6 min; internal standard: isobutanol, used for quantitative correction.

[0093] A series of ethanol and acetic acid standard solutions of known concentrations were precisely prepared, and an equal volume of isobutanol was added as an internal standard. The standard solutions were analyzed under the chromatographic conditions described above, and the peak areas of ethanol, acetic acid, and isobutanol were recorded. A standard curve was plotted with the concentration of ethanol or acetic acid on the x-axis and the ratio of the ethanol peak area to the isobutanol peak area on the y-axis, and the regression equation was obtained. The cell-free supernatant was taken, and an equal volume of isobutanol was added as used in the preparation of the standard curve. After mixing, GC detection was performed. The peak areas of ethanol and isobutanol in the sample were recorded, substituted into the standard curve regression equation, and the ethanol content (g / L) in the sample was calculated.

[0094] Biomass determination: Take 10 mL of fermentation broth into a centrifuge tube, centrifuge at 8000 r / min for 10 min, collect the cell precipitate, wash 3 times with deionized water, dry at 60℃ to constant weight, weigh the dry weight, and calculate the biomass dry weight (g / L).

[0095] Take an appropriate amount of freeze-dried bacterial cells, grind them into powder, pass them through a 220-mesh sieve, and take a uniform sample. Weigh 3.2-3.8 mg of the powder and determine the nitrogen content using a Flashsmart elemental analyzer. The protein content and yield are calculated using the following formulas: Protein content (%) = Nitrogen content × 6.25, Protein yield (g / L) = Biomass × Protein content.

[0096] Results Analysis

[0097] Temperature acclimatization of Candida utilis

[0098] Using ethanol and acetic acid as the sole carbon sources, wild-grown *Candida utilis* TU389, with an optimal growth temperature of 31℃, underwent 131 generations of high-temperature transfer acclimatization (fermentation cycles of 48 hours for generations 1-27, and 24 hours for generations 28-131). The OD600 value of the fermentation broth was continuously monitored at each generation to characterize biomass changes. Specific results are shown below. Figure 1 and Figure 2 As shown.

[0099] Depend on Figure 1As shown in Figure A, the average OD600 of the strain during the domestication process exhibits the characteristics of initial rapid increase, mid-term fluctuation and optimization, and high-level stability in the later stage. The initial average OD600 of the first-generation strain was only 1.54, which increased to 2.39 after rapid adaptation in the first 10 generations, an increase of 55.6% compared to the initial value. The biomass of generations 11-26 fluctuated between 1.74 and 2.24. After reaching stability, considering the possibility of carbon source concentration limitations (carbon source was exhausted in 48 hours), the OD measurement time was changed to 24 hours starting from the 27th transfer. The OD value of the 27th transfer reached 1.9, and then gradually increased, reaching the peak value of the entire cycle (2.424) in the 64th generation, reflecting the metabolic optimization process of the strain under the dual selection pressure of high temperature and complex carbon source. The average OD600 of generations 65-131 was stable in the high range of 1.905-2.389, with the 131st generation reaching 2.195, an increase of 43.0% compared to the initial value, and there was no significant downward trend. This indicates that the domesticated strain has formed a stable high-temperature adaptive phenotype, and short-cycle transfer did not lead to performance degradation. The domestication effect has genetic stability.

[0100] Depend on Figure 1 As shown in Figure C, using ethanol-sodium acetate (2:1, total carbon source 10 g / L) as substrate, the biomass dry weight of domesticated strain (X) and wild type (W) was compared at 31℃ and 37℃. The average biomass dry weight of wild type at 31℃ (W-31 / 6) was 4.87 g / L, which decreased to 4.06 g / L at 37℃ (W-37 / 6), a decrease of 16.6%, indicating a significant high-temperature inhibitory effect. In contrast, the biomass dry weight of domesticated strain at 31℃ (X-31 / 6) and 37℃ (X-37 / 6) was 4.944 g / L and 4.478 g / L, respectively, which were 1.6% and 10.4% higher than that of wild type at the same temperature. Moreover, the decrease in biomass at 37℃ (9.4%) was lower than that of wild type, indicating that domesticated strain has a wider temperature adaptation range.

[0101] Depend on Figure 1 As shown in Figure D, fed-batch fermentation with ethanol-acetic acid (2:1) as the carbon source for 72 hours resulted in an average dry biomass of 29.87 g / L for the wild-type strain at 31℃ (W-31 / 6) and a decrease to 23.27 g / L at 37℃ (W-37 / 6), a decrease of 22.1%. The biomass of the domesticated strain reached 35.41 g / L at 31℃ (X-31 / 6), an increase of 18.5% compared to W-31 / 6, and 33.39 g / L at 37℃ (X-37 / 6), an increase of 43.4% compared to W-37 / 6. Moreover, the biomass of X-37 / 6 was not significantly different from that of W-31 / 6. The significance analysis showed that the biomass of the domesticated strain was not significantly different at the two temperatures, indicating that the domesticated strain could tolerate the high temperature of 37℃. Furthermore, the domesticated strain X131 showed tolerance to temperature fluctuations, suggesting that the strain exhibited a certain degree of robustness to temperature under industrial conditions.

[0102] Depend on Figure 2 As shown in Figure A, the carbon source conversion rates of the domesticated strains at 31℃ and 37℃ were higher than those of the wild type at the same temperature, with the carbon source conversion rate of X-37 / 6 being significantly higher than that of W-37 / 6.

[0103] Depend on Figure 2 As shown in B, the protein content of the domesticated strain was slightly higher than that of the wild type at both temperatures.

[0104] Depend on Figure 2 As shown in C, the protein yields of X-31 / 6 and X-37 / 6 increased by 19.2% and 45.1% compared to W-31 / 6 and W-37 / 6, respectively, and the protein yield of X-37 / 6 has exceeded that of W-31 / 6.

[0105] This indicates that after high-temperature domestication, *Candida utilis* exhibits more efficient production performance.

[0106] As shown in Table 1, there is a difference in the total content of 18 amino acids between the domesticated strain X-37 / 6 and the wild strain W-31 / 6: the total amino acid content of X-37 / 6 is 36.10 g / 100 g, which is 10.3% lower than that of W-31 / 6 (40.24 g / 100 g). This difference indicates that high temperature affects microbial protein synthesis. Although the high temperature environment of 37℃ did not cause the amino acid synthesis system to collapse, it resulted in a slightly lower accumulation of total protein (total amino acids) than the wild strain at the optimum temperature (31℃). However, the domesticated strain did not show a complete absence of any type of amino acid, indicating that through 131 generations of continuous domestication, the domesticated strain has achieved stable adaptation of the amino acid synthesis system under high temperature, avoiding the risk of protein synthesis collapse under extreme high temperature.

[0107] Table 1

[0108]

[0109] pH acclimatization of Candida utilis

[0110] The results of pH acclimatization of Candida utilis are as follows: Figure 1 and Figure 2 As shown.

[0111] like Figure 1 As shown in B, Candida utilis X131, obtained by high-temperature domestication at 37℃, was used as the starting strain. Under the stress condition of pH 3.5, it underwent 60 generations of adaptive evolution. Its cell growth density (OD600) showed a typical adaptive improvement-stabilization process with the change of domestication generations.

[0112] In the early stages of acclimatization (generations 0-15), the strain exhibited significant growth inhibition under low pH stress, with OD600 remaining between 1.0 and 1.4, lower than its growth level under pH 6.0 conditions. This reflects the initial inhibitory effect of the acidic environment on cell membrane integrity, enzyme activity, and central metabolism. As acclimatization progressed (generations 16-25), the strain gradually adapted to low pH stress, with OD600 rapidly increasing to above 2.0. This indicates that through multiple generations of selection, the cells had accumulated adaptive mutations, effectively alleviating the effects of H+. + Efflux pressure and the toxicity of acetic acid dissociation restored core metabolic activity.

[0113] During the later stages of domestication (generations 26-60), the cell growth density plateaued and continued to optimize, with OD600 stabilizing between 2.0 and 2.4. In some generations (such as generation 32 and 52), it reached its peak, even exceeding the growth level of the original strain at the optimal pH of 6.0. This indicates that long-term low-pH domestication not only enabled the strain to acquire stable acid tolerance, but also that adaptive evolution successfully screened out a superior mutant strain with both high tolerance and high yield potential under pH 3.5 conditions, laying the foundation for subsequent low-pH high-density fermentation.

[0114] like Figure 1 As shown in Figure C, under shake flask culture conditions, the dry biomass of strain P-37 / 3.5 was 4.56±0.05, which was higher than that of strain X-37 / 6 (4.478±0.01). This indicates that low pH acclimatization not only enabled the strain to acquire tolerance to pH 3.5, but also improved its growth performance.

[0115] like Figure 1 As shown in Figure D, at the 5L fermenter level, the dry biomass of strain P-37 / 3.5 reached 35.18±0.27 g / L, which was also superior to the 33.39±1.00 g / L of strain X-37 / 6, verifying the growth advantage of domesticated strains in large-scale fermentation.

[0116] like Figure 2 As shown in Figure A, the carbon source conversion rate of strain P-37 / 3.5 was 0.51±0.01 g / g (based on consumed ethanol and acetic acid), which was slightly lower than that of strain X-37 / 6 (0.56±0.02 g / g), but the difference was not significant, indicating that low pH acclimatization did not significantly weaken the conversion efficiency of the strain for C2 carbon source.

[0117] like Figure 2 As shown in B, the cell protein content of strain P-37 / 3.5 was 41.57±0.33%, which was higher than that of strain X-37 / 6 (39.49±0.13%).

[0118] like Figure 2As shown in C, its final protein yield reached 14.62±0.01g / L, which is about 10.9% higher than that of strain X-37 / 6 (13.18±0.35g / L), demonstrating superior protein production capacity.

[0119] In summary, the P-37 / 3.5 strain, obtained through 60 generations of low-pH adaptive evolution, achieved simultaneous improvement in growth performance, protein content, and protein yield under pH 3.5 stress conditions, while maintaining high C2 carbon source conversion efficiency. This provides an excellent strain foundation for single-cell protein production of Candida utilis in low-pH industrial fermentation scenarios.

[0120] Example 3: Effect of Candida utilis strain P-37 / 3.5 on the conversion of ethanol, acetic acid, and wastewater.

[0121] Using wild-type strain TU389 as a control, and pure ethanol (10 g / L) as the carbon source, *Candida utilis* strain P-37 / 3.5 was fermented at pH 3.5 to convert and utilize ethanol. The fermentation results are shown in Table 2. The results indicate that *Candida utilis* P-37 / 3.5 has a good ability to convert and utilize ethanol.

[0122] Table 2

[0123]

[0124] Using wild-type strain TU389 as a control, and pure acetic acid (10 g / L) as the carbon source, the fermentation of *Candida utilis* strain P-37 / 3.5 at pH 3.5 was studied. The fermentation results are shown in Table 3. The results indicate that *Candida utilis* strain P-37 / 3.5 has a better ability to convert and utilize acetic acid under low pH conditions than the wild-type strain.

[0125] Table 3

[0126]

[0127] Using wild-type strain TU389 as a control, acidic wastewater rich in ethanol with a low pH (4-5) was selected as the culture medium to study the growth of *Candida utilis* strain P-37 / 3.5. The fermentation results are shown in Table 4. The results indicate that *Candida utilis* P-37 / 3.5 can effectively convert and utilize industrial wastewater.

[0128] Table 4

[0129]

[0130] Test Example 1

[0131] 1.1 Transcriptome Analysis

[0132] Transcriptome sequencing technology, as a core tool for elucidating the molecular mechanisms of microbial environmental adaptation, can comprehensively screen differentially expressed genes (DEGs) under stress conditions, thereby revealing the regulatory patterns of strains in carbon source utilization, energy metabolism, genome stability, and stress response.

[0133] Total RNA extraction and quality control: Bacterial samples frozen at -80℃ (wild-type TU389 cultured at 31℃ and 37℃, and domesticated strain X131 cultured at 31℃ and 37℃ in a 5L fermenter were named W-31, W-37, X-31, and X-37, respectively, with 3 biological replicates for each) were used. Total RNA was extracted using the Trizol method. RNA integrity was detected by 1% agarose gel electrophoresis, RNA purity was determined by Nanodrop (A260 / A280 ratio 1.8~2.0), and RNA concentration was quantified using Qubit to ensure that the RNA quality met the sequencing requirements.

[0134] cDNA library construction: Qualified total RNA was collected, and mRNA was enriched using an mRNA enrichment kit. After fragmentation, cDNA first and second strands were synthesized using the mRNA as templates. End repair, A-tailing, and adapter ligation were performed. The cDNA library was obtained by PCR amplification. After passing quality testing (Agilent 2100 for fragment size detection, Qubit quantification), the library was used for sequencing.

[0135] High-throughput sequencing: The constructed cDNA library was subjected to paired-end sequencing on the Illumina NovaSeq 6000 platform to obtain raw sequencing data (Raw Reads).

[0136] Data filtering and assembly: The raw sequencing data was quality controlled to remove reads containing adapters, low-quality reads (Q value <20), and reads with an excessively high N ratio, to obtain clean reads. The clean reads were aligned to the Candida utilis reference genome (NCBI GenBank database), and transcript assembly and quantitative analysis were performed using software to calculate the transcript abundance of each gene (expressed as TPM value).

[0137] All experimental data were set up with 3 biological replicates, and the results are expressed as mean ± standard deviation (mean ± SD). Statistical analysis was performed using GraphPad Prism10 software. One-way ANOVA was used to compare differences between groups, and P < 0.05 was considered statistically significant. Domestication curves, fermentation curves, and index comparison charts were plotted. Transcriptome data analysis was performed using relevant R language packages (DESeq2, clusterProfiler, etc.) for visualization and enrichment analysis.

[0138] 1.2 Transcriptomics Data Analysis

[0139] This invention conducted rigorous quality control (QC) assessments on the transcriptome sequencing data of all samples (W-31, W-37, X-31, and X-37) from the fermenter after 72 hours. The sequencing data quality statistics are shown in Table 2. The abbreviations of the strains are as follows: W-31-1, W-31-2, and W-31-3 represent wild-type Candida utilis TU389 (31℃, pH 6.0); W-37-1, W-37-2, and W-37-3 represent wild-type Candida utilis TU389 (37℃, pH 6.0); X-31-1, X-31-2, and X-31-3 represent high-temperature acclimatized strain X131 (31℃, pH 6.0); and X-37-1, X-37-2, and X-37-3 represent high-temperature acclimatized strain X131 (37℃, pH 6.0). The results show that the overall data quality is excellent and meets the requirements for subsequent analysis. The number of raw sequencing reads for each sample was 3.90 × 10⁻⁶. 7 Up to 5.05×10 7 After quality control filtering, the number of effective clean reads was highly consistent with the number of original reads, indicating a high data retention rate. The sequencing error rate for all samples was below 0.013%, far below the quality threshold (≤0.1%) for conventional transcriptome sequencing. Regarding base quality, the proportion of Q20 (base recognition accuracy ≥99%) was above 99.5%, and the proportion of Q30 (base recognition accuracy ≥99.9%) exceeded 97.0%, indicating extremely high accuracy and reliability of the sequencing data. GC content remained stable between 46.69% and 46.99%, showing no obvious GC bias or contamination. Furthermore, the rRNA contamination rate was controlled below 2.6%, indicating good mRNA enrichment in the samples. Overall, the transcriptome sequencing data of this invention are of reliable quality. Specific sequencing results are shown in Table 5.

[0140] Table 5

[0141]

[0142] like Figure 3 As shown, the first two principal components (PC1 and PC2) explained a total of 75.57% of the total variance, with PC1 explaining 43.67% and PC2 explaining 31.90%, indicating that these two dimensions are sufficient to reflect the main differences between samples. Figure 3As shown in Figure A, the PCA scatter plot reveals a clear grouping trend: W-series (wild-type) samples are concentrated in the negative PC1 region, while X-series (domesticated strains) samples are distributed in the positive PC1 region, indicating that strain background is the primary factor driving sample differences. Simultaneously, within the same strain, samples cultured at 31℃ and 37℃ show significant separation along the PC2 axis, indicating that culture temperature is a secondary but significant factor affecting the transcriptome expression profiles of the samples. The close clustering of biological replicates within each group indicates good experimental reproducibility and reliable data. Figure 3 The PCA grouping diagram shown in Figure B, with the addition of confidence ellipses, makes the clustering trends of each group more intuitive. The confidence ellipses of the four treatment groups W-31, W-37, X-31, and X-37 do not overlap, further confirming that strain type and culture temperature are two key variables affecting transcriptome expression profiles, and that their effects are independent. This result provides a solid foundation for subsequent differential expression analysis and functional enrichment studies.

[0143] In summary, this invention, using ethanol and acetic acid as the sole carbon sources, successfully increased the fermentation tolerance temperature of *Candida utilis* from 31℃ to 37℃ through 131 generations of continuous high-temperature transfer and domestication, obtaining a domesticated strain with stable high-temperature adaptability. Based on this, further domesticated strain P-37 / 3.5, which is simultaneously tolerant to both 37℃ high temperature and pH 3.5 acidic environment, was screened out through 60 generations of low-pH adaptive evolution. This domesticated strain exhibits higher biomass, carbon source conversion rate, protein content, and protein yield than the wild type at the same temperature at both 31℃ and 37℃. Specifically, at 37℃, biomass increased by 43.4% and protein yield increased by 45.1% compared to the wild type. Under low pH conditions, the P-37 / 3.5 strain achieved a dry biomass weight of 35.18 g / L, a protein content of 41.57%, and a protein yield of 14.62 g / L, representing a 10.9% increase in protein yield compared to the starting strain at the optimal pH 6.0. Furthermore, it demonstrated good phenotypic stability during fermenter scale-up culture.

[0144] This invention provides excellent strain resources for the high-temperature and low-pH fermentation applications of Candida utilis. The excellent performance of the low-pH acclimated strain P-37 / 3.5 under the dual stress of high temperature and low pH has been verified by fermentation experiments. This invention provides new ideas and experimental basis for the stress resistance acclimation of microorganisms that utilize complex carbon sources, and is of great significance for promoting the green and low-carbon transformation of industrial microorganisms.

[0145] The specific embodiments described above further illustrate the purpose, technical solution, and beneficial effects of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A type of Candida utilis TU389-37XHZ, characterized in that, It is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 39159.

2. The application of the Candida utilis TU389-37XHZ as described in claim 1, characterized in that, It is at least one of the following: (1) Conversion to ethanol; (2) Conversion of acetic acid; (3) Co-conversion of ethanol and acetic acid; (4) Production of single-cell proteins; (5) Treating wastewater containing ethanol, acetic acid and / or acetate; (6) Treatment of brewing wastewater; (7) Treat wastewater from bioethanol production.

3. The application of Candida utilis TU389-37XHZ as described in claim 1, characterized in that, It is at least one of the following: (1) Used in food processing; (2) Used in feed processing.

4. A method for producing single-cell protein, characterized in that, The method includes the step of co-culturing the Candida utilis TU389-37XHZ as described in claim 1 with a carbon source fermentation broth, wherein the carbon source fermentation broth comprises ethanol and acetic acid.

5. The method according to claim 4, characterized in that, The carbon source fermentation broth meets at least one of the following conditions: (1) The pH value of the carbon source fermentation broth is 3~9; (2) The temperature of the carbon source fermentation broth is 25℃~45℃.

6. A method for treating wastewater, characterized in that, Includes the step of inoculating the Candida utilis TU389-37XHZ as described in claim 1 into wastewater for cultivation; The wastewater contains at least one of ethanol, acetic acid, and acetate.

7. A composition, characterized in that, The composition contains Candidautilis TU389-37XHZ as described in claim 1.

8. The composition according to claim 7, characterized in that, The composition is a microbial agent.

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