A method of quantifying the ability of a microbial mat to capture and bind particles

Abstract

The present application relates to a method for quantifying the ability of microbial mat to capture and bind particles, the method comprises the following steps: cultivating microbial mat according to the physical and chemical conditions of the water body at the sampling point; taking several portions of the microbial mat, and carrying out multiple groups of capture experiments on mixed particles of different particle sizes, wherein for each group of capture experiments, the weight of the particles captured by the microbial mat is measured, and the microbial mat after capturing the particles is further cultivated, and the weight of the particles bound by the microbial mat is measured after the capture experiment ends; taking several portions of the microbial mat, and carrying out multiple groups of blank experiments on mixed particles of different particle sizes after sterilization and cleaning, wherein for each group of blank experiments, the weight of the particles captured by the microbial mat is measured, and the microbial mat after capturing the particles is further cultivated, and the weight of the particles bound by the microbial mat is measured after the blank experiment ends; determining the weight proportion of particles of different particle sizes by comparing the measurement results of the capture experiments and the blank experiments, and analyzing the ability of the microbial mat to capture and bind particles based on the weight proportion.

Description

Technical Field

[0001] This invention relates to the field of microbial and microbial rock research technology, and in particular to a method for quantifying the ability of microbial mats to capture and bind particles. Background Technology

[0002] Microbial rocks refer to "in-situ mineral-deposited organic sediments formed by benthic microbial communities through the capture and binding of clastic sediments, or through inorganic / organic induction by microorganisms," encompassing two important geological processes: capture and binding, and induced precipitation. The metabolic processes of these microorganisms (such as cyanobacteria and diatoms) can produce large amounts of electronegative extracellular polymeric substances (EPS), which overcome energy barriers, absorb cations from surrounding water, and thus induce in-situ precipitation of carbonate rocks. The ability of microorganisms to induce precipitation has been confirmed by numerous ancient examples and microbial culture experiments, and is widely used in crack filling, dam repair, and material protection.

[0003] The capture and adhesion process specifically refers to the blocking and protection of particles by uncalcified filamentous microorganisms and the EPS (extracellular polymeric substances) slime matrix produced by metabolism within the microbial mat, causing water-borne particles to settle and form sediments. The particle composition is often composed of intraclastic particles of varying sizes and roundness, forming fine- to coarse-grained aggregated stromatolites or tuffaceous rocks under the influence of the microbial mat. This process is considered crucial in the formation and development of stromatolites and tuffaceous rocks in ancient and modern environments such as the Bahamas and Shark Bay. Previous research indicates that, in both ancient and modern environments, the capture and adhesion processes of microbial mats are closely related to microbial community ecology, hydrodynamics, water saturation, and particle size.

[0004] Current experimental techniques largely focus on inducing the formation of carbonate minerals such as calcite and dolomite through microbial mat cultivation, providing valuable experimental guidance for the century-old mystery of "dolomite." However, research on the ability of microorganisms to capture and bind particles of different sizes during metabolism is relatively limited, especially in terms of quantitative studies. This significantly restricts our understanding of the paleoatmospheric and marine / lake environments during the formation of ancient agglomerated stromatolites and tuffaceous rocks, as well as the ecological evolution of microorganisms (such as cyanobacteria). Therefore, there is an urgent need for an experimental method to quantitatively study the ability of microbial mats to bind and capture particles of different sizes. This would help to quantitatively assess microbial communities and their contribution to atmospheric oxygen content, and further lay an important foundation for understanding the formation mechanism of ancient microbial rocks and reconstructing sedimentary processes.

[0005] After searching patents and literature, the Derwent database shows that: Patent CN112308936A discloses a method for determining the influence of microbial activity on the development of microbial carbonate reservoirs, bridging the relationship between the strength of microbial activity and structure and the reservoir. This method quantitatively characterizes the ability of microbial mats to capture and bind particles, addressing the formation mechanism of microbial rocks from a mechanistic perspective, and can provide a reference for the formation of microbial rocks in this patent. Therefore, there are differences in the technical field and application scenarios. Patent CN111411127A discloses a method for inducing calcium carbonate precipitation using microorganisms, by adding sodium montmorillonite... Soil removal accelerates calcium carbonate precipitation, which is one of the processes in microbial rock formation (induced precipitation). However, this method differs in that it addresses another capture and bonding process in microbial rock formation through microbial culture experiments, thus differing from existing technologies in terms of method, purpose, and application. Patent CN110628624B discloses a magnetic microbial capture material and method, which greatly enhances the capture efficiency of magnetic beads through the introduction of magnetic materials. In contrast, this method involves the capture and bonding of microbial particles, rather than microbial particles themselves, belonging to the fields of microbiology and geological sedimentology, and significantly differing from existing technologies in terms of method and purpose. Summary of the Invention

[0006] To address the aforementioned technical problems, at least one embodiment of the present invention provides a method for quantifying the ability of microbial mats to capture and bind particles through microbial culture experiments. This method...

[0007] In some embodiments of the present invention, the method mainly includes the following steps:

[0008] Microbial mats that meet the experimental conditions are cultivated based on the physicochemical conditions of the water at the sampling point;

[0009] Take several microbial mats that meet the experimental conditions and conduct multiple capture experiments on mixed particles of different sizes. For each capture experiment, measure the weight of the particles captured by the microbial mat and continue to cultivate the microbial mat after capturing the particles. After the capture experiment is completed, measure the weight of the particles adhered to the microbial mat.

[0010] Take several microbial mats that meet the experimental conditions, and after sterilization and cleaning, conduct multiple sets of blank experiments on mixed particles of different sizes. For each set of blank experiments, measure the weight of the particles captured by the microbial mat, and continue to cultivate the microbial mat after capturing the particles. After the blank experiments are completed, measure the weight of the particles adhered to the microbial mat.

[0011] The weight ratio of particles of different sizes was determined by comparing the measurement results of the capture experiment and the blank experiment, and the ability of the microbial mat to capture and bind particles was analyzed based on the weight ratio.

[0012] In some embodiments of the present invention, the physicochemical conditions of the water body at the sampling point include one or more of the following: seawater illumination, temperature, salinity, pH, anion and cation composition, and nutrients.

[0013] In some embodiments of the present invention, the cultivation of a microbial mat that meets experimental conditions based on the physicochemical conditions of the water at the sampling point includes:

[0014] The types and components of the microbial mat were determined by rRNA gene sequence analysis.

[0015] Determine whether the microbial mat meets the experimental conditions based on its type and composition.

[0016] In some embodiments of the present invention, determining whether the microbial mat meets the experimental conditions based on its type and composition includes:

[0017] Calculate the percentage of principal components in the microbial mat;

[0018] When the proportion of the main component in the microbial mat exceeds a preset threshold, the microbial mat is determined to meet the experimental conditions.

[0019] In some embodiments of the present invention, the method further includes, prior to performing the capture experiment:

[0020] Based on the physicochemical conditions of the water at the sampling point, the relevant conditions of the microbial mat experimental environment were set up. Preliminary experimental analysis was conducted on particles of the same weight but different sizes to determine the maximum particle size that the microbial mat could capture and the stabilization time of the bound particles. In some embodiments of the present invention, the sterilization cleaning includes:

[0021] The microbial mat is sterilized by high pressure and cleaned with ethanol.

[0022] In some embodiments of the present invention, determining the weight ratio of particles of different sizes by comparing the measurement results of the capture experiment with those of the blank experiment, and analyzing the ability of the microbial mat to capture and bind particles based on the weight ratio, includes:

[0023] Statistical analysis was performed on the weight ratio of particles of different sizes in each group of microbial mats;

[0024] A quantitative indicator of the ability of the microbial mat to capture and bind different particles was calculated based on the weight ratio.

[0025] At least one embodiment of the present invention also provides an electronic device, characterized in that it comprises:

[0026] At least one processor; and,

[0027] A memory communicatively connected to the at least one processor; wherein,

[0028] The memory stores instructions that can be executed by the at least one processor to enable the at least one processor to perform the method described above for quantifying the ability of microbial mats to capture and bind particles.

[0029] At least one embodiment of the present invention also provides a computer-readable storage medium storing a computer program that, when executed by a processor, implements the method described above for quantifying the ability of microbial mats to capture and bind particles.

[0030] At least one embodiment of the present invention also provides a computer program product, including a computer program that, when executed by a processor, implements the steps of the method for quantifying the ability of microbial mats to capture and bind particles as described above.

[0031] This invention provides a method for quantifying the ability of microbial mats to capture and bind particles. This method is simple, highly feasible, and yields significant results, effectively filling a gap in existing microbial experimental techniques. By quantitatively calibrating particle capture and binding capacity indicators, and combining this with the particle size, relative content, and distribution location of ancient microbial rocks, the method effectively quantifies the scale of ancient microbial communities, their growth and metabolic environment, and their contribution to pO2 in the ancient atmosphere and ocean. Furthermore, this lays a solid experimental foundation for establishing the formation mechanism of ancient microbial rocks and reconstructing sedimentary processes. Attached Figure Description

[0032] One or more embodiments are illustrated by way of example with reference to the accompanying drawings, and these illustrative descriptions do not constitute a limitation on the embodiments.

[0033] Figure 1 This is a flowchart of the steps of the method for quantifying the ability of microbial mats to capture and bind particles, as used in Embodiment 1 of the present invention. Detailed Implementation

[0034] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the various embodiments of the present invention will be described in detail below with reference to the accompanying drawings. However, those skilled in the art will understand that many technical details are presented in the embodiments of the present invention to facilitate a better understanding of the invention. However, the technical solutions claimed in the present invention can be implemented even without these technical details and various variations and modifications based on the following embodiments. The division of the following embodiments is for ease of description and should not constitute any limitation on the specific implementation of the present invention. The various embodiments can be combined with and referenced by each other without contradiction.

[0035] As mentioned above, this invention aims to provide a method for quantifying the particle-capturing and binding ability of microbial mats, filling the gap in existing experiments on particle capture and binding by microbial mats. This method allows for the quantitative calibration of the particle capture and binding ability of modern microbial mats. Combined with studies on particle size, relative abundance, and distribution in ancient microbial rocks, it can quantitatively infer the size of microbial communities and their metabolic contributions to atmospheric-oceanic pO2. Furthermore, it lays an important foundation for understanding the formation mechanism of ancient microbial rocks and reconstructing sedimentary processes.

[0036] Example 1

[0037] like Figure 1 As shown, this method mainly includes the following steps:

[0038] S01, Microbial culture: The physical and chemical conditions of the water body at the indoor sampling point are simulated. The collected microbial mats are placed in a large water tank for a certain period of time for culture. The growth of the microbial community is observed to determine its evolution to the mature stage.

[0039] S02, Microbial species identification: The mature microbial mat is divided into several equal-sized portions, and several portions are taken for rRNA gene sequence analysis to identify the main microbial species and proportions, and to check whether they meet the experimental standards (>90%).

[0040] S03, Preliminary capture and adhesion experiment: Several microbial mats were placed in petri dishes and then fixed in the same large water tank at a 20° inclination angle. Further, particles of the same weight but different sizes were weighed and placed on the microbial mats for a preliminary capture and adhesion experiment. After the experiment, uncaptured and unadhere- ...

[0041] S04, Capture Experiment: Take several samples of microbial mats and conduct multiple parallel capture experiments. Unlike step S03, the particles are a mixture of particles of different sizes in the same weight ratio. Specifically, the experimental conditions and steps are the same as in step S03 above. After a short period of settling, collect the uncaptured particles, dry them, weigh them, and record the weight data of the captured particles for each group.

[0042] S05, Adhesion Experiment: The experimental sample in step S04 is cultured for a certain period of time. After the experiment is over, the unadheded particles are collected by inverting the sample, dried, weighed, and the weight data of each group of adhered particles are recorded.

[0043] S06, Blank Experiment Comparison: Several microbial mats were taken for blank parallel experiments for comparison. Similarly, the experimental conditions and procedures were the same as in steps S04 and S05, except that the microbial mats underwent autoclaving and ethanol cleaning. After the experiment, uncaptured and unbonded particles were collected, dried, weighed, and the weight data of captured and bonded particles in each group were recorded.

[0044] S07, Evaluate the ability of the microbial mat to capture and bind particles: Specifically, calculate the weight ratio of particles bound and captured by the microbial mat, statistically analyze the weight ratio of particles of different sizes in each group, and further, quantitatively calculate the index δ value of the particle capture and binding ability of each microbial mat.

[0045] Example 2

[0046] Another embodiment of the present invention uses cyanobacterial microbial mats as an example to quantitatively study the capture and binding ability of particles of different sizes. The specific implementation steps are as follows:

[0047] S01, Microbial Culture: Simulating the seawater conditions of region A in the South China Sea, including light, temperature, salinity, pH, anion and cation composition, and nutrients, the collected cyanobacterial mats were placed in a large 1m×1m×1m water tank for cultivation. The tank was connected to a bubbler to prevent excessive oxygen accumulation. Further, the growth of the cyanobacteria was observed. When the cyanobacteria stopped growing significantly, showed no signs of decay, and continued to produce bubbles, it indicated that the cyanobacterial mat had evolved to a mature and stable stage, capable of producing the maximum amount of EPS (excessive sludge matrix).

[0048] S02, Microbial species identification: The mature cyanobacterial mat from step S01 was divided into 36 equal parts, each measuring 15cm × 15cm × 15cm. Three parts from different regions were selected for rRNA gene sequence analysis to identify the main microbial components, including cyanobacteria, diatoms, and sulfate-reducing bacteria. The cyanobacteria content, dominated by *Coleofasciculus*, averaged 90%, meeting the experimental standards.

[0049] S03, Preliminary experiment on capture and adhesion: Five cyanobacterial mats were placed in 20cm×20cm×20cm petri dishes and then fixed in the same large water tank (1m×1m×1m). The physical and chemical conditions inside and outside the water tank were the same as in step S01 to ensure normal growth of the microbial mats. Further, based on the estimated capture capacity of 5g particles per cyanobacterial mat (maximum particle size 4mm), 5g particles with different diameters (0.125–0.625mm, 0.625–1.0mm, 1.0–3.0mm, 3.0–5.0mm, 5.0–8.0mm) were weighed and gently placed on the five cyanobacterial mats. After standing for half an hour, the particles that fell from each petri dish were collected, dried, and weighed. The weight ratio of particles captured by each cyanobacterial mat was recorded. Similarly, five more cyanobacterial mats were used for adhesion experiments. Five 5g samples of particles with a diameter of 0.625–1.0 mm were placed on the cyanobacterial mats. The experiments were completed at 6h, 12h, 18h, 24h, and 30h, respectively. Unadhesive particles were collected, dried, and weighed. The proportion of adherent particles in each group was calculated. The results showed that the cyanobacterial mats of this size had a capture capacity of less than 10% for particles of 5.0–8.0 mm, and their adhesion capacity remained basically stable after 18h of cultivation.

[0050] S04, Capture Experiment: Six cyanobacterial mats were used to conduct multiple parallel experiments to test the capture capacity. Unlike step S03, six 5g portions of mixed particles with different particle sizes (<5.0mm) in the same weight ratio were weighed to better reflect actual geological conditions. Specifically, the experimental conditions and procedures were the same as in step S03. After standing for half an hour, the uncaptured particles in each petri dish were collected, dried, and weighed. The weight data a1-6 of the captured particles for each group were recorded.

[0051] S05, Adhesion Experiment: In step S04, the 6 cyanobacterial mat experimental samples were cultured for another 18 hours. After the experiment, the unadhesive particles in each culture dish were collected by inverting the dish, dried, and weighed. The weight data b1 to b6 of the adhesive particles in each group were recorded.

[0052] S06, Blank Experiment Comparison: Six cyanobacterial mats were used for a blank parallel experiment comparison. The experimental conditions and procedures were the same as in steps S04 and S05. The difference was that the cyanobacterial mats were autoclaved and cleaned with ethanol. After the experiment, uncaptured and unattached particles were collected from each petri dish, dried, and weighed. The weight data c1-6 and d1-6 of the captured and attached particles for each group were recorded.

[0053] S07, Evaluate the ability of cyanobacterial mats to capture and bind particles: Specifically, calculate the weight ratio of captured and bound particles in 6 groups of capture and binding experiments and 6 groups of blank experiments (Formulas 1 and 2), and statistically analyze the weight ratio of particles of different sizes in each group of cyanobacterial mats. Furthermore, quantitatively calculate the δ value (Formulas 3 and 4) of the particle capture and binding ability of the cyanobacterial mat with an area of ​​0.0225 m². Specifically, the calculation formulas are as follows:

[0054]

[0055] Where m represents the total weight of particles before the experiment (g), x and y represent the weight of uncaptured and unbonded particles after each experiment (g), S represents the total area of ​​each cyanobacterial mat, Pcapture represents the ratio of captured particles to the weight before the experiment (%), Bbonding represents the ratio of bonded particles to captured particles (%), and δcapture and δbonding represent quantitative indicators (g / m2) of the cyanobacterial mat's ability to capture and bond different particles. In the example capture and bonding experiment, Pcapture and Bbonding were 78.17% and 81.87%, respectively, and δcapture and δbonding were 173.71 g / m2 and 142.22 g / m2, respectively. In the example blank control experiment, Pcapture and Bbonding were 43.47% and 39.99%, respectively, and δcapture and δbonding were 96.38 g / m2 and 38.54 g / m2, respectively.

[0056] This embodiment uses cyanobacterial microbial mats as a specific example to quantitatively calibrate the ability of cyanobacterial mats to capture and bind particles, effectively filling the gap in existing technologies in this experimental field. It lays a solid experimental foundation for quantitative research on the atmospheric-oceanic pO2 contribution of ancient microbial mats and the mechanisms of microbial rock formation. Cyanobacteria are among the most common types of ancient and modern microorganisms in this embodiment. This method remains effective for red algae, green algae, diatoms, sulfate-reducing bacteria, etc. Therefore, any modifications that do not deviate from the essence of this method still fall within the scope of this invention. The method of this invention is simple in process and highly feasible in operation, effectively filling the gap in microbial experimental technology, and can be promoted in paleoenvironmental analysis, mineralization precipitation, CCUS, and oil and gas reservoir prediction.

[0057] Example 3

[0058] Another embodiment of the present invention relates to an electronic device, comprising: at least one processor; and a memory communicatively connected to the at least one processor; wherein the memory stores instructions executable by the at least one processor to enable the at least one processor to perform the methods for quantifying the ability of microbial mats to capture and bind particles as described in the above embodiments.

[0059] The memory and processor are connected via a bus, which can include any number of interconnecting buses and bridges, connecting various circuits of one or more processors and memories. The bus can also connect various other circuits, such as peripheral devices, voltage regulators, and power management circuits, which are well known in the art and will not be described further herein. The bus interface provides an interface between the bus and the transceiver. The transceiver can be a single element or multiple elements, such as multiple receivers and transmitters, providing a unit for communicating with various other devices over a transmission medium. Data processed by the processor is transmitted over the wireless medium via an antenna, which further receives data and transmits it to the processor.

[0060] The processor manages the bus and general processing, and also provides various functions, including timing, peripheral interfaces, voltage regulation, power management, and other control functions. Memory is used to store data used by the processor during operation.

[0061] Example 4

[0062] Another embodiment of the present invention relates to a computer-readable storage medium storing a computer program. When executed by a processor, the computer program implements the method described above for quantifying the ability of microbial mats to capture and bind particles.

[0063] That is, those skilled in the art will understand that all or part of the steps in the methods of the above embodiments can be implemented by a program instructing related hardware. This program is stored in a storage medium and includes several instructions to cause a device (which may be a microcontroller, chip, etc.) or processor to execute all or part of the steps of the methods described in the various embodiments of the present invention. The aforementioned storage medium includes various media capable of storing program code, such as USB flash drives, portable hard drives, read-only memory (ROM), random access memory (RAM), magnetic disks, or optical disks.

[0064] Example 5

[0065] Another embodiment of the present invention relates to a computer program product, including a computer program that, when executed by a processor, implements the steps of the method for quantifying the ability of microbial mats to capture and bind particles as described in the above embodiments.

[0066] Those skilled in the art will understand that the above embodiments are specific embodiments for implementing the present invention, and in practical applications, various changes in form and detail may be made without departing from the spirit and scope of the present invention.

Claims

1. A method for quantifying the ability of microbial mats to capture and bind particles, characterized in that, Includes the following steps: Microbial mats that meet the experimental conditions are cultivated based on the physicochemical conditions of the water at the sampling point; Take several microbial mats that meet the experimental conditions and conduct multiple capture experiments on mixed particles of different sizes. For each capture experiment, measure the weight of the particles captured by the microbial mat and continue to cultivate the microbial mat after capturing the particles. After the capture experiment is completed, measure the weight of the particles adhered to the microbial mat. Take several microbial mats that meet the experimental conditions, and after sterilization and cleaning, conduct multiple sets of blank experiments on mixed particles of different sizes. For each set of blank experiments, measure the weight of the particles captured by the microbial mat, and continue to cultivate the microbial mat after capturing the particles. After the blank experiments are completed, measure the weight of the particles adhered to the microbial mat. The weight ratio of particles of different sizes was determined by comparing the measurement results of the capture experiment and the blank experiment, and the ability of the microbial mat to capture and bind particles was analyzed based on the weight ratio.

2. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 1, characterized in that, The physicochemical conditions of the water at the sampling point include one or more of the following: seawater illumination, temperature, salinity, pH, anion and cation composition, and nutrients.

3. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 2, characterized in that, The cultivation of microbial mats that meet experimental conditions based on the physicochemical conditions of the water at the sampling point includes: The types and components of the microbial mat were determined by rRNA gene sequence analysis. Determine whether the microbial mat meets the experimental conditions based on its type and composition.

4. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 3, characterized in that, The step of determining whether the microbial mat meets the experimental conditions based on its type and composition includes: Calculate the percentage of principal components in the microbial mat; When the proportion of the main component in the microbial mat exceeds a preset threshold, the microbial mat is determined to meet the experimental conditions.

5. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 1, characterized in that, Prior to conducting the capture experiment, the method further includes: Based on the physicochemical conditions of the water at the sampling point, the relevant conditions of the microbial mat experimental environment were set up. Preliminary experimental analysis was conducted on particles of the same weight but different sizes to determine the maximum particle size that the microbial mat could capture and the stabilization time of the bound particles.

6. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 1, characterized in that, The sterilization cleaning includes: The microbial mat is sterilized by high pressure and cleaned with ethanol.

7. The method for quantifying the hydrodynamic influence on the ability of microbial mats to capture and bind particles according to claim 1, characterized in that, The step of determining the weight ratio of particles of different sizes by comparing the measurement results of the capture experiment and the blank experiment, and analyzing the ability of the microbial mat to capture and bind particles based on the weight ratio, includes: Statistical analysis was performed on the weight ratio of particles of different sizes in each group of microbial mats; A quantitative indicator of the ability of the microbial mat to capture and bind different particles was calculated based on the weight ratio.

8. An electronic device, characterized in that, include: At least one processor; as well as, A memory communicatively connected to the at least one processor; wherein, The memory stores instructions executable by the at least one processor to enable the at least one processor to perform the method for quantifying the ability of microbial mats to capture and bind particles as described in any one of claims 1 to 7.

9. A computer-readable storage medium storing a computer program, characterized in that, When the computer program is executed by a processor, it implements the method for quantifying the ability of microbial mats to capture and bind particles as described in any one of claims 1 to 7.

10. A computer program product, comprising a computer program, characterized in that, When executed by a processor, the computer program implements the steps of the method for quantifying the ability of microbial mats to capture and bind particles as described in any one of claims 1 to 7.

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