Method for determining salinity of soil or water body by using microorganisms and application

Abstract

The invention relates to the technical field of environmental monitoring, in particular to a method for measuring soil or water salinity by using microorganisms and application. In a salt environment, the growth rate of the selected microorganisms is related to the salinity; when the salinity is enhanced, the growth of microorganisms is inhibited; according to the method, the salinity of the environment is judged by monitoring microorganisms OD600 in real time; the method comprises the following steps: selecting Nocardia listeria BFY2-1 as a strain to be tested as a microorganism, and selecting Nocardia listeria BFY2-1 as a strain to be tested; a to-be-tested water body or soil 1: 5 suspension; adjusting the pH value to 6-9, and taking 20 ML; adding 10 ML of a three-time culture medium, inoculating a test strain, and comparing with sterile water; carrying out shake cultivation at 28 DEG C, wherein the rotating speed is 200 rpm; after 48 hours, measuring the absorbance at the wavelength of 600 nm by using a spectrophotometer; calculating the salt content according to the absorbance, and converting the initial salt content according to the dilution.

Description

Technical Field

[0001] This invention relates to the field of environmental monitoring technology, and in particular to methods and uses for determining soil or water salinity using microorganisms. Background Technology

[0002] Pollution from saline soils and highly saline water bodies is becoming increasingly serious globally, particularly in arid, semi-arid, and coastal regions. These areas experience high evaporation rates, low rainfall, and seawater intrusion, leading to the continuous accumulation of salt in soil and water, resulting in widespread salinization. High salinity not only directly hinders crop growth, hinders root water absorption, and causes nutrient imbalances, leading to reduced yields and quality degradation, but also disrupts soil structure, causing compaction and reduced aeration, further exacerbating land degradation. Simultaneously, salt stress affects microbial community structure and vegetation distribution, resulting in ecosystem dysfunction, a significant reduction in biodiversity, and a serious threat to sustainable agricultural development and regional ecological security.

[0003] Currently, commonly used salinity determination methods mainly rely on traditional chemical analysis techniques, such as conductivity methods, ion chromatography, or titration. While these methods offer high accuracy and reliable results, they generally suffer from drawbacks such as complex operating procedures, stringent sample pretreatment requirements, reliance on specialized equipment, long detection times, and high operating costs. For example, ion chromatography requires precision instruments and standard solutions, while titration demands skilled operators and stringent experimental conditions. These limitations make it difficult to meet the practical needs of rapid on-site detection, large-scale dynamic monitoring, and real-time data feedback. Furthermore, chemical analysis methods typically consume large amounts of auxiliary reagents, and improper disposal of waste liquids can cause secondary pollution, further exacerbating the environmental burden.

[0004] Therefore, developing a simple, rapid, economical, and highly reliable method for salinity measurement has become an urgent need in the fields of environmental monitoring and agricultural management. Such a method should enable real-time, non-destructive testing in fields or at pollution sites, reducing the professional requirements for equipment and personnel while maintaining high measurement accuracy and repeatability. It will provide key technical support for the precise management of salinized areas, real-time monitoring of irrigation water quality, rational allocation of water resources, and scientific evaluation of ecological restoration projects, possessing significant practical value and potential for widespread application.

[0005] On December 26, 2025, a search was conducted in the China Patent Publication Database using "microorganisms and salts and absorbance and content" as the abstract keywords, with the option to allow synonym expansion. CN115478018A relates to a reagent-grade yeast extract, its preparation method, and its applications. The reagent-grade yeast extract has a total nitrogen content of 10-13%, an amino acid nitrogen content ≥5%, a magnesium ion content ≤500ppm, a calcium ion content ≤500ppm, a manganese ion content ≤10ppm, a copper ion content ≤5ppm, and an absorbance of ≤0.2 at 600nm for an aqueous solution of the reagent-grade yeast extract with a weight percentage concentration of 30-40%. The reagent-grade yeast extract prepared by this invention exhibits significantly improved clarity, a substantial reduction in divalent metal ion content, and does not produce precipitation when combined with phosphates, reducing cell aggregation and enhancing the product's microbial growth-promoting ability, thus meeting the purity requirements of high-end biopharmaceutical products.

[0006] CN107561025A discloses a rapid quantitative method for detecting the iron oxidation capacity of iron-oxidizing bacteria. The steps are as follows: (1) sampling and extraction of enrichment solution; (2) solution preparation; (3) determination of standard curve; (4) measurement of Fe; (5) measurement of total active iron (Fe and Fe); (6) calculation of the iron oxidation capacity of iron-oxidizing bacteria under anaerobic conditions. This method has high efficiency, relatively low cost, and good experimental repeatability. The linear relationship between the iron concentration extracted by hydrochloric acid and absorbance is good in the range of 0-6 mg / L. This method is reliable and has a wide range of applications. It can quickly detect the content of ferrous and ferric iron in various environmental systems such as soil, sediment, microbial culture medium, and groundwater.

[0007] CN104312937A belongs to the field of wastewater biological treatment technology, specifically relating to a method for cultivating polyphosphate-containing bacteria (B8) using polyphosphate staining. The cultivated strain is named B8 and is currently deposited at the Microbial Culture Collection Center of the Institute of Microbiology, Chinese Academy of Sciences, with accession number CGMCC NO.9168. It has been identified as *Pseudomonas putida* sp. The polyphosphate staining cultivation method using the above-mentioned strain is as follows: B8 strain is inoculated into CCZU culture medium and cultured in a shaking incubator to obtain a bacterial culture. Then, 2% polyphosphate (Poly-P) staining solution is added to the bacterial culture and the culture is incubated at 30℃ for 24 hours. A blank is prepared using the same procedure. The seed culture is centrifuged at 8000 r / min for 15 min, and the absorbance of the supernatant is measured at 625 nm to characterize the content of polyphosphate particles in the seed culture.

[0008] On December 26, 2025, an abstract search was conducted on CNKI (China National Knowledge Infrastructure) using the keywords "microorganisms, salt content, absorbance, and content". The following articles were found: "Optimization of Cassava Residue Composting Parameters and the Influence of Surfactant Addition on the Composting Process" (published June 1, 2023); "Screening of Oil-Degrading Strains and Their Application in Food Waste Degradation" (published May 1, 2022); "Desalination Process of Pickled Vegetables, Analysis of Spoilage Bacteria Community, and Preservative Study Based on ε-Polylysine" (published January 1, 2019); and "Preparation Process and Quality Study of Seasoning from Litopenaeus vannamei By-products" (published April 1, 2016). These articles are unrelated to this invention.

[0009] On December 26, 2025, a search was conducted on the website of the United States Patent and Trademark Office for the term "microorganisms with salt with absorbance with content," but no relevant literature was found; the search URL is https: / / ppubs.uspto.gov / pubwebapp / .

[0010] On December 26, 2025, a search was conducted on WIPO's website https: / / patentscope2.wipo.int / for the term "microorganisms and salt and absorbance and content", but no relevant literature was found.

[0011] On December 26, 2025, a search was conducted on the website of the Japan Patent Office (https: / / www.j-platpat.inpit.go.jp / ) for the term "microorganisms and salt and absorbance and content," but no relevant literature was found.

[0012] It is completely different from the concept of this patent. Summary of the Invention

[0013] Purpose of the invention: To provide a more effective method and application for measuring soil or water salinity using microorganisms, the specific purpose of which is described in the detailed implementation section for several substantial technical effects.

[0014] To achieve the above objectives, the present invention adopts the following technical solution: A method for determining soil or water salinity using microorganisms, characterized in that... In a saline environment, the growth rate of the selected microorganisms showed a certain correlation with salinity; when salinity increased, the growth of the microorganisms was inhibited. The method determined the salinity of the environment by real-time monitoring of the microbial OD600. Listeria monocytogenes was selected as the microorganism.Nocardiopsis listeri BFY2-1 was used as the test strain; The test solution is a 1:5 suspension of water or soil. Adjust the pH to 6-9 and take 20 ml. Add 10 mL of 3 times the amount of culture medium and inoculate with the test strain; the control is sterile water. Incubate at 28℃ on a shaker at 200 rpm; 48 hours later, the absorbance was measured at 600 nm using a spectrophotometer. The salt content is calculated based on the absorbance, and the initial salt content is converted based on the dilution.

[0015] A further technical solution of the present invention is that the culture medium provides the nutrients required for microbial growth; data acquisition: a handheld spectrophotometer is used to measure the values, the absorbance is measured at 600 nm, and the measured values ​​are converted into readable data.

[0016] A further technical solution of the present invention is that by substituting the formula y = -0.107x + 1.143, where x is the initial salt content and y is the absorbance, the initial salt content can be calculated.

[0017] A further technical solution of the present invention is that the culture medium is a 3x culture medium formula: 9.0g beef extract, 30.0g peptone, 15.0g NaCl, 1000ml water, and the pH value is adjusted to 6.5-8.5.

[0018] A further technical solution of the present invention is Listeria monocytogenes. Nocardiopsis listeri BFY2-1 is deposited at the China Center for Type Culture Collection (CCTCC), located at No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province, with accession number CCTCC M 20211324 and deposit date of October 26, 2021.

[0019] Listeria monocytogenes Nocardiopsis listeri BFY2-1 is used for determining the salinity of soil or water.

[0020] The present invention, employing the above technical solution, offers the following advantages over existing technologies: It enables the determination of salinity in soil or water. This method utilizes specific salt-tolerant microorganisms, whose growth and metabolic products reflect salinity, thus achieving rapid and accurate environmental monitoring. It eliminates the need for complex chemical reagents, making it environmentally friendly and setting a precedent in this field. High sensitivity: It possesses extremely high detection sensitivity, responding rapidly to minute changes in salinity, ensuring the accuracy and timeliness of measurement results, and is suitable for various environmental conditions.

[0021] Environmentally friendly: Using natural microorganisms as sensing elements completely avoids the use of chemical reagents, reducing environmental pollution and ecological risks, and conforms to the concept of sustainable development.

[0022] Low cost: Compared with traditional chemical analysis methods, the manufacturing process is simple, the material cost is low, and the maintenance requirements are minimal, which significantly reduces the overall cost and makes it more economical and accessible.

[0023] Real-time monitoring: It can continuously and in real time monitor the salinity of soil and water, and provide instant information through an efficient data transmission system to support rapid decision-making and precise management. Attached Figure Description

[0024] To further illustrate the present invention, the following description is provided in conjunction with the accompanying drawings: Figure 1 The OD600 absorbance of the bacterial culture after 48 hours of incubation under different initial pH conditions; Figure 2 The OD600 absorbance of the bacterial culture after 48 hours of incubation under different NaCl contents; Figure 3 This is a schematic diagram illustrating the working principle of the method. Figure 4 Proof of the survival of the microorganisms used in the invention Select Listeria monocytogenes Nocardiopsis listeri BFY2-1, the test strain, is deposited at the China Center for Type Culture Collection (CCTCCM), located at No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province, with accession number CCTCCM 20211324 and deposit date of October 26, 2021. Detailed Implementation

[0025] The present invention will be further illustrated below with reference to the accompanying drawings and specific embodiments. It should be understood that the following specific embodiments are for illustrative purposes only and are not intended to limit the scope of the present invention.

[0026] Select Listeria monocytogenes Nocardiopsis listeri BFY2-1, the test strain, is deposited at the China Center for Type Culture Collection (CCTCCM), located at No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province, with accession number CCTCCM 20211324 and deposit date of October 26, 2021.

[0027] method Prepare one 3x culture medium, one bacterial suspension, and one control. The culture medium used is beef extract peptone medium. The original formula is: 3.0g beef extract, 10.0g peptone, 5.0g NaCl, and 1000ml water, with the pH adjusted to 6.5-8.5. The 3x culture medium formula is: 9.0g beef extract, 30.0g peptone, 15.0g NaCl, and 1000ml water, with the pH adjusted to 6.5-8.5.

[0028] The method includes the following components: Culture medium: Provides the nutrients required for microbial growth. The formulation of the culture medium is optimized according to the characteristics of the target microorganism.

[0029] Data acquisition: A handheld spectrophotometer was used to measure the absorbance at 600 nm, and the measured values ​​were converted into readable data.

[0030] Working principle: In saline environments, the growth rate of the selected microorganisms showed a certain correlation with salinity. Microbial growth was inhibited as salinity increased. The method determined the environmental salinity by real-time monitoring of the microbial OD600.

[0031] Calibration and Data Analysis: The method needs to be calibrated regularly to ensure accuracy. Data analysis and result prediction are achieved by establishing a mathematical model between salinity and changes in microbial metabolites.

[0032] in conclusion This invention provides a novel microbial method for determining salinity through microbial metabolic activity, which has broad application prospects and can effectively promote research and application in fields such as agriculture and environmental protection.

[0033] Experimental data and results Water body 1: After filtration with filter paper, the initial pH was 6.5. 20 mL of water was added to 10 mL of a 3x culture medium, and the test bacterial strain was inoculated. This was repeated three times. The control was prepared by adding 10 mL of sterile water. After 48 hours, the absorbance values ​​were 0.357, 0.426, and 0.398, with an average of 0.394. Substituting into the formula y = -0.107x + 1.143, where x is the initial salinity and y is the absorbance, the initial salinity was calculated to be 7.0%.

[0034] Soil 1: Soil was prepared as a 1:5 soil-to-water suspension. After filtration with filter paper, the initial pH was 8.5. 20 mL of this suspension was added to 10 mL of a 3x culture medium, and the test bacterial strain was inoculated. This was repeated three times. The control was prepared by adding 10 mL of sterile water. After 48 hours, the absorbance values ​​were 0.932, 1.247, and 0.987, with an average of 1.056. Substituting into the formula y = -0.107x + 1.143, where x is the initial salinity and y is the absorbance, the initial salinity was calculated to be 0.82%.

[0035] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the claims.

Claims

1. A method for determining soil or water salinity using microorganisms, characterized in that, In a saline environment, the growth rate of the selected microorganisms showed a certain correlation with salinity; when salinity increased, the growth of the microorganisms was inhibited. The method determined the salinity of the environment by real-time monitoring of the microbial OD600. Listeria monocytogenes was selected as the microorganism. Nocardiopsis listeri BFY2-1 was used as the test strain; The test solution is a 1:5 suspension of water or soil. Adjust the pH to 6-9 and take 20 ml. Add 10 mL of 3 times the amount of culture medium and inoculate with the test strain; the control is sterile water. Incubate at 28℃ on a shaker at 200 rpm; 48 hours later, the absorbance was measured at 600 nm using a spectrophotometer. The salt content is calculated based on the absorbance, and the initial salt content is converted based on the dilution.

2. The method for determining soil or water salinity using microorganisms as described in claim 1, characterized in that, The culture medium provides the nutrients required for microbial growth; Data acquisition: A handheld spectrophotometer was used to measure the absorbance at 600 nm, and the measured values ​​were converted into readable data.

3. The method for determining soil or water salinity using microorganisms as described in claim 1, characterized in that, Substituting the formula y = -0.107x + 1.143, where x is the initial salt content and y is the absorbance, the initial salt content can be calculated.

4. The method for determining soil or water salinity using microorganisms as described in claim 1, characterized in that, The culture medium is a 3x culture medium formula: 9.0g beef extract, 30.0g peptone, 15.0g NaCl, 1000ml water, and the pH value is adjusted to 6.5-8.

5.

5. The method for determining soil or water salinity using microorganisms as described in claim 1, characterized in that, Listeria monocytogenes Nocardiopsis listeri BFY2-1 is deposited at the China Center for Type Culture Collection (CCTCC), located at No. 299 Bayi Road, Wuchang District, Wuhan City, Hubei Province, with accession number CCTCC M 20211324 and deposit date of October 26, 2021.

6. Listeria monocytogenes Nocardiopsis listeri BFY2-1 is used for determining the salinity of soil or water.

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