A ship ballast water biological risk identification and evaluation method, device, medium and product

Abstract

This application discloses a method, equipment, medium, and product for identifying and assessing biological risks in ship ballast water, relating to the fields of marine engineering and environmental safety technology. The method includes: obtaining ship ballast water samples; screening and identifying hazardous substances based on the ballast water samples; determining the concentration values ​​of the identified hazardous substances using qPCR; constructing a risk assessment model; using the risk assessment model to perform risk substance analysis simulation based on the concentration values ​​of the hazardous substances, obtaining simulation results of ballast water biological risk substances; and obtaining ballast water biological risk assessment results based on the simulation results. This application can construct a risk management system for identifying and assessing ballast water quality safety risks to address new challenges in biosafety.

Description

Technical Field

[0001] This application relates to the fields of marine engineering and environmental safety technology, and in particular to a method, equipment, medium and product for identifying and assessing biological risks in ship ballast water. Background Technology

[0002] Ballast water is water and suspended matter added to a ship to control its list, trim, draft, stability, or stress. Its main functions are to adjust the ship's displacement, balance the hull, improve seaworthiness, and reduce hull vibration by injecting into or removing it from ballast tanks. Due to the vast differences in water quality and marine life across the globe, no ballast water treatment technology or device has yet been found that is suitable for the vast majority of sea areas and ships. Therefore, while ballast water ensures navigational safety, it also brings global environmental problems—biological invasion.

[0003] Every year, over 10 billion tons of ballast water are transferred globally, with more than 7,000 species of organisms migrating to new environments daily, including bacteria, planktonic plants and animals, pathogens, and dormant spores. If these organisms become invasive species, they can trigger catastrophic consequences such as ecological collapse, public health crises, and the spread of toxic algal blooms.

[0004] Furthermore, current biological risk control measures in ship ballast water are limited to the existing Convention on the Control and Management of Ship Ballast Water and Sediments, which specifies requirements for the content of live organisms and some microorganisms larger than or equal to 10 μm. The risks associated with live organisms smaller than 10 μm or other emerging pollutants have consistently been underestimated. One study identified 710 subtypes of 26 antibiotic resistance genes (ARGs) in ballast water samples from 13 ships from 11 countries and regions, including multidrug resistance genes. Existing ballast water treatment processes cannot remove all pollutants; toxic and harmful organic matter, pathogens, ARGs, and other pollutants may enter the ecosystem of the port area during ballast water reception and treatment, thus posing biological risks. Therefore, how to construct a risk management system for ballast water quality safety risk identification and assessment through a multi-dimensional linkage of screening and identification, technical assessment, and policy management to address new biosafety challenges has become an urgent technical problem to be solved in this field. Summary of the Invention

[0005] The purpose of this application is to provide a method, equipment, medium, and product for identifying and assessing biological risks in ship ballast water, which can build a risk management system for identifying and assessing ballast water quality safety risks in order to address new challenges in biosafety.

[0006] To achieve the above objectives, this application provides the following solution: Firstly, this application provides a method for identifying and assessing biological risks in ship ballast water, including: Obtain ballast water samples from the ship; Hazardous substances were screened and identified based on the ship's ballast water samples. The concentration values ​​of the risk substances were determined by qPCR quantitative analysis based on the screened and identified risk substances. Develop a risk assessment model; Using the aforementioned risk assessment model, risk substance analysis simulation was performed based on the qPCR quantitative analysis of risk substance concentration values ​​to obtain the simulation results of ballast water biological risk substances. The ballast water biological risk assessment results are obtained based on the simulation results of the ballast water biological risk substances.

[0007] Secondly, this application provides a computer device, including: a memory, a processor, and a computer program stored in the memory and executable on the processor, wherein the processor executes the computer program to implement the steps of the above-described method for identifying and assessing biological risks in ship ballast water.

[0008] Thirdly, this application provides a computer-readable storage medium having a computer program stored thereon, which, when executed by a processor, implements the steps of the above-described method for identifying and assessing biological risks in ship ballast water.

[0009] Fourthly, this application provides a computer program product, including a computer program that, when executed by a processor, implements the steps of the above-described method for identifying and assessing biological risks in ship ballast water.

[0010] According to the specific embodiments provided in this application, this application has the following technical effects: This application provides a method, equipment, medium, and product for identifying and assessing biological risks in ship ballast water. It involves screening and identifying hazardous substances based on ballast water samples, and then determining the concentration of these substances using qPCR. This concentration is used as the initial input concentration, and combined with a constructed risk assessment model, simulation results of ballast water biological risks can be obtained, thereby achieving risk assessment. Based on this, this application constructs a risk management system for ballast water quality safety risk identification and assessment through a multi-dimensional linkage of processes including screening and identification, risk assessment, and ballast water zoning management, thus addressing new challenges to biosafety. Attached Figure Description

[0011] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0012] Figure 1 A flowchart illustrating a method for identifying and assessing biological risks in ship ballast water, provided as an embodiment of this application; Figure 2 This is a schematic diagram of a risk factor screening and identification process provided in an embodiment of this application; Figure 3 A schematic diagram illustrating the results of an indoor simulation experiment of bacterial cells and eDNA provided in an embodiment of this application; Figure 4 A schematic diagram of the technical route for a method for identifying and assessing biological risks in ship ballast water provided in an embodiment of this application; Figure 5 This is a schematic diagram of the structure of a computer device provided in an embodiment of this application. Detailed Implementation

[0013] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0014] To make the above-mentioned objectives, features and advantages of this application more apparent and understandable, the application will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0015] In one exemplary embodiment, this application provides a method for identifying and assessing biological risks in ship ballast water. This method is executed by computer equipment, specifically by a terminal or server, or by both. In this embodiment, the method is described using a server as an example. Figure 1 As shown, the method includes: Step 100: Obtain a sample of the ship's ballast water.

[0016] Step 101: Screening and identification of hazardous substances based on ship ballast water samples.

[0017] Step 102: Based on the screened and identified risk substances, determine the concentration value of the risk substances by qPCR.

[0018] Step 103: Construct a risk assessment model.

[0019] Step 104: Using a risk assessment model, conduct risk substance analysis and simulation based on the concentration values ​​of the risk substances to obtain the simulation results of risk substances for ballast water organisms. This analysis and simulation can be achieved by performing migration simulations in a terrestrial environment.

[0020] Step 105: Obtain the ballast water biological risk assessment results based on the simulation results of ballast water biological risk substances.

[0021] By implementing steps 100-105 above, this application can significantly improve the efficiency and accuracy of ballast water biological risk assessment, providing key technical support for effectively preventing and controlling invasive alien species and protecting the marine ecological environment.

[0022] In one exemplary embodiment of this application, in order to improve the accuracy of screening and identification of hazardous substances, this application may, based on considerations of biological invasiveness, ecological hazard, detection frequency, and the environmental tolerance and cumulative response of biological hazardous substances, and on a large number of documents (such as... Figure 2 Based on research and analysis of the "Discharge Standard for Municipal Wastewater Treatment Plants" and its amendments, as well as literature reports, and the requirements of the "2024 International Convention for the Control and Management of Ballast Water and Sediments," a risk control pollutant identification method for ballast water reception and treatment was established by comparing it with relevant urban wastewater treatment standards (such as the "Urban Wastewater Reuse" series of standards) to screen and identify risky substances. The screened and identified risky substances include conventional pollutants (primary controlled biological risk substances) and emerging pollutants (key controlled biological risk substances). While conventional pollutants are not specified in the D-2 discharge standard, they are restricted by the "Discharge Standard for Municipal Wastewater Treatment Plants" and its amendments, and the "Urban Wastewater Reuse" series of standards; these mainly include fecal coliform count and total coliform count. Emerging pollutants refer to pollutants reported in at least one publication, such as ARGs and toxic microalgae, but not covered by the D-2 discharge standard. Based on this, the primary controlled biological pollutants for ship ballast water reception designed in this application are shown in Table 1.

[0023] Table 1. List of Primary Controlled Biological Pollutants in Ship Ballast Water Reception

[0024] Based on the above description, during the identification of hazardous substances, marine organisms (fish, invertebrates, mammals, microorganisms, etc.) release DNA fragments (i.e., eDNA) into the environment through skin cells, mucus, excrement, gametes, etc. eDNA can be detected in extremely small amounts from ship ballast water samples, allowing for the detection of difficult-to-observe biological and other genetic information. Therefore, by extracting and detecting eDNA from ship ballast water samples, the content of primary controlled biological hazardous substances in ballast water can be analyzed, thereby avoiding the underestimation of biological hazardous substances in ship ballast water samples due to detection methods, and using this as the initial quantity for the risk assessment system. In this application, ballast water refers to ballast water samples that have been received and treated at ports and meet the D-2 discharge standards of international conventions. After ballast water treatment, eDNA is extracted from it. Therefore, in practical applications, the implementation process of step 101 of this application can be described as follows: Step 101-1: Extracting eDNA from ship ballast water samples. This involves filtering the ship ballast water samples using a nitrocellulose membrane of a set density (0.22 μm) to remove intracellular DNA from the ballast water, obtaining a filtrate. Magnetic beads are then used to extract eDNA from the filtrate. For example, the steps are as follows: (1) Add 400 μL of isopropanol and 300 μL of C1 buffer (proteinase K dissolved at 20 g / L in 30 mM Tris-Cl, Tiangen Biotech, China) to 200 μL of ballast water sample (hereinafter referred to as the sample) filtrate, mix thoroughly to remove impurities (such as proteins), and then add 6.5 μL of magnetic bead stock solution (30 g / L) (1 μm nucleic acid extraction silanol magnetic beads, vortex for 3-4 min). Use a magnet to adsorb the nucleic acids carried by the magnetic beads in the mixture, and discard the supernatant without magnetic beads.

[0025] (2) The magnetic beads were washed with 0.5 mL of CW1 buffer (7 mol / L guanidine hydrochloride dissolved in 50% isopropanol), and the supernatant was removed again by magnetic adsorption. Then, the magnetic beads were washed twice with 0.5 mL of CW2 buffer (75% ethanol), and then placed at room temperature for 5 min to evaporate the remaining ethanol. After that, 30 μL of elution buffer (10 mM Tris-HCl, pH 8.5, preheated to 55 °C) was added to the magnetic beads and incubated for 5 min. During incubation, the mixture was vortexed for 20 s per minute. Finally, the mixture was placed on a magnetic rack, and the supernatant was collected for subsequent qualitative or quantitative analysis.

[0026] Step 101-2: Quantitative analysis of the extracted eDNA is performed using qPCR (multi-gene marker) to screen and identify risk substances based on the quantitative analysis results. During this process, qPCR-high-throughput sequencing can be used simultaneously to obtain the levels of fecal coliforms, total coliforms, and ARGs in the sample for quantitative analysis. The primer sequences used for qPCR are shown in Table 2.

[0027] Table 2 Primer sequence listing

[0028] Furthermore, based on the above description, the process of analyzing the content of primary biohazard substances in ballast water can be described as follows: First, based on different water pollutant discharge standards, the Ballast Water Convention D-2 discharge standard, and literature review, biohazard pollutants that need to be controlled in ballast water are identified, as shown in Table 1 above. Second, according to different pollutants, extracellular DNA (eDNA) is extracted from ballast water samples and amplified using conventional PCR (using the same primer sequences as qPCR). The amplified products are then subjected to 1.5% agarose gel electrophoresis for 40 min, and the amplified strips are observed using a gel imaging analyzer to qualitatively determine the presence of the target gene in the sample. Third, based on the presence of the gene, qPCR is used for quantification to analyze the copy number of different target genes.

[0029] In one exemplary embodiment of this application, after ballast water is received and treated onshore, it is discharged into port waters and may also enter soil and groundwater systems via the shoreline, thereby entering various ecosystems and posing biological risks. Therefore, it is necessary to establish an effective risk assessment model.

[0030] Because the hydrogeology of different ports varies, in order to unify the assessment standards, the model construction process in this embodiment can be based on a homogeneous, anisotropic, two-dimensional, unstable groundwater flow system, using a hydrogeological conceptual model. Referring to existing research on biocolloids such as E. coli and soil column experimental models simulating ARGs migration, a convection-diffusion equation considering a colloidal filtration model is used to simulate the migration process of bacteria and eDNA in the aquifer, thus constructing a mathematical model that considers the leaching effect during bacterial colloidal migration. The migration of hazardous substances in the porous medium is mainly affected by convection and diffusion, without considering biodegradation and other complex processes. Based on this, the implementation process of step 103 provided above can be described as follows: Based on a biocolloid and soil column experimental model simulating ARGs migration, a convection-diffusion equation considering a colloidal filtration model is used to simulate the migration process of bacteria and eDNA in the aquifer, thereby constructing a mathematical model that considers the leaching effect during bacterial colloidal migration. This mathematical model is used as a risk assessment model. The constructed risk assessment model is expressed as follows: .

[0031] In the formula, For pollutant concentration, The porosity of the medium. The density is the medium. The concentration of colloids adsorbed onto the surface of soil particles by reversible adsorption is expressed in copies / g. The concentration of colloids adsorbed onto the surface of soil particles by irreversible adsorption, expressed in copies / g. The direction of pollutant movement The diffusion coefficient is... For fluid velocity, For time. The corresponding term for adsorption is: , .

[0032] The corresponding item for leaching is: .

[0033] In the formula, The reversible adsorption rate parameter ( sul 1, 2.25 × 10 -6 [1 / s]), The first-order dynamic blocking coefficient, . The time-dependent dimensionless retention coefficient. The maximum adsorption capacity on the medium is expressed in copies / g. It is a dimensionless retention coefficient that depends on distance. Average diameter of medium particles, in cm. Empirical values ​​for controlling the morphological distribution state. For the resolution rate coefficient, This represents the amount of adsorption on the medium.

[0034] Through such Figure 3 The resistant bacteria and antibiotic resistance genes shown sul The indoor simulation results of 1 demonstrate that the simulation results of the risk assessment model constructed in this application are good.

[0035] In one exemplary embodiment of this application, based on the conclusions of different literature studies, the stable effluent concentration of bacteria and eDNA (the ratio of effluent pollutant concentration C to influent pollutant concentration C0) is used as one of the risk assessment criteria for the sample. Based on this, a risk assessment model can be adopted according to the abundance of different types of eDNA in the sample (i.e., the concentration values ​​of risk substances analyzed by qPCR). Through indoor soil column experiments, soil columns are filled according to the soil properties of the site, and pollutants are reinjected into the soil column to simulate the diffusion patterns and stable effluent concentrations of different eDNAs, thereby obtaining the simulation results of ballast water biological risk substances.

[0036] Based on the simulation results of ballast water biohazard substances, risk assessments were conducted on treated samples from different regions. Then, based on the assessment results, the risk levels of the ballast water source ports were classified (through continuous data accumulation, risk classification of ocean-going vessels entering or ports of entry was performed using ballast water biohazard substance data), and a ballast water risk level map was drawn. For example, when the concentration of fecal coliform bacteria in the effluent is greater than 1000 per liter, and the cDNA C / C0 ≥ 30% and absolute abundance is 10, risk levels are considered high. 4 The above indicates that this sample is classified as high-risk. When the concentration of fecal coliforms in the effluent is 1000 per liter, the cDNA content is 10% ≤ C / CO < 30%, and the absolute abundance is 10. 3 The above indicates that this ballast water sample is classified as medium risk. When the concentration of fecal coliform bacteria in the effluent is less than 1000 per liter, and the C / C0 ratio of eDNA is less than 10% and the absolute abundance is 10, it is considered a medium risk. 3 The sample is classified as low risk.

[0037] In one exemplary embodiment of this application, the method for identifying and assessing biological risks in ship ballast water provided by this application can generate risk response measures based on the ballast water biological risk assessment results. For example, for ship ballast water in high-risk areas, receiving and processing work is strictly prohibited. Ballast water must be replaced by sailing to a designated open sea area (usually requiring at least 200 nautical miles from the shore and a water depth of more than 2,000 meters), or the ballast water must be sealed in the tank and carried by the ship to the next designated high-risk water area where discharge is permitted or returned to the source for processing.

[0038] For ballast water from ships in medium-risk areas, a mandatory regulatory approach of "onboard treatment" + "enhanced monitoring" is required. If the onboard treatment system experiences a temporary malfunction, a backup plan must be activated or port-approved mobile receiving and treatment facilities must be used for treatment at a designated location. Direct discharge of treated ballast water is strictly prohibited; it is primarily intended for ship ballast. In addition to routine compliance inspections, random checks for indicator pathogens and key ARGs (Argumentative Genetic Spectrum Organisations) markers will be conducted. The standards for these random checks can be determined based on actual needs.

[0039] For ballast water from ships in low-risk areas, onshore reception and processing can be provided where conditions permit. After ships provide complete electronic evidence such as the ballast water loading location and replacement coordinates and file it, they can be given priority for berthing and operations, thereby improving port efficiency.

[0040] For ballast water from ships in risk-free areas, it can be discharged directly in designated compliant areas within the port, with a green channel for customs clearance opened to achieve zero-wait berthing and operation, and priority allocation of berth and loading / unloading resources.

[0041] In summary, the technical approach for implementing the ship ballast water biological risk identification and assessment method provided in this application is as follows: Figure 4 As shown, it can be widely used by port state control authorities for rapid screening of ballast water from international vessels, for ship operators for self-assessment and decision support of ballast water management, for ballast water treatment system manufacturers for verification and optimization of equipment performance, and for ecological protection zones for precise management of ship ballast water discharge.

[0042] In one exemplary embodiment, a computer device is provided, which may be a server or a terminal, and its internal structure diagram may be as follows. Figure 5 As shown, the computer device includes a processor, memory, input / output (I / O) interfaces, and a communication interface. The processor, memory, and I / O interfaces are connected via a system bus, and the communication interface is also connected to the system bus via the I / O interfaces. The processor provides computational and control capabilities. The memory includes non-volatile storage media and internal memory. The non-volatile storage media stores the operating system, computer programs, and a database. The internal memory provides the environment for the operation of the operating system and computer programs stored in the non-volatile storage media. The database stores data on the identification and assessment of biological risks in ship ballast water. The I / O interfaces are used for exchanging information between the processor and external devices. The communication interface is used for communication with external terminals via a network connection. When the computer program is executed by the processor, it implements a method for identifying and assessing biological risks in ship ballast water.

[0043] Those skilled in the art will understand that Figure 5 The structure shown is merely a block diagram of a portion of the structure related to the present application and does not constitute a limitation on the computer equipment to which the present application is applied. Specific computer equipment may include, for example, [the following is a list of possible additional structures]. Figure 5 The diagram shows more or fewer components, or combinations of certain components, or different component arrangements.

[0044] In one exemplary embodiment, a computer device is provided, including a memory and a processor, wherein the memory stores a computer program, and the processor executes the computer program to implement the steps in the above-described method embodiments.

[0045] In one exemplary embodiment, a computer-readable storage medium is provided storing a computer program that, when executed by a processor, implements the steps in the above-described method embodiments.

[0046] In one exemplary embodiment, a computer program product is provided, including a computer program that, when executed by a processor, implements the steps in the above-described method embodiments.

[0047] It should be noted that the user information (including but not limited to user device information, user personal information, etc.) and data (including but not limited to data used for analysis, data stored, data displayed, etc.) involved in this application are all information and data authorized by the user or fully authorized by all parties, and the collection, use and processing of the relevant data must comply with relevant regulations.

[0048] Those skilled in the art will understand that all or part of the processes in the above embodiments can be implemented by a computer program instructing related hardware. The computer program can be stored in a non-volatile computer-readable storage medium, and when executed, it can include the processes of the embodiments described above. Any references to memory, databases, or other media used in the embodiments provided in this application can include at least one of non-volatile and volatile memory. Non-volatile memory can include read-only memory (ROM), magnetic tape, floppy disk, flash memory, optical memory, high-density embedded non-volatile memory, resistive random access memory (RRAM), magnetic random access memory (MRAM), ferroelectric random access memory (FRAM), phase change memory (PCM), graphene memory, etc. Volatile memory can include random access memory (RAM) or external cache memory, etc. By way of illustration and not limitation, RAM can take many forms, such as Static Random Access Memory (SRAM) or Dynamic Random Access Memory (DRAM).

[0049] The databases involved in the embodiments provided in this application may include at least one type of relational database and non-relational database. Non-relational databases may include, but are not limited to, blockchain-based distributed databases. The processors involved in the embodiments provided in this application may be general-purpose processors, central processing units, graphics processing units, digital signal processors, programmable logic devices, quantum computing-based data processing logic devices, etc., and are not limited to these.

[0050] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0051] This document uses specific examples to illustrate the principles and implementation methods of this application. The descriptions of the above embodiments are only for the purpose of helping to understand the methods and core ideas of this application. Furthermore, those skilled in the art will recognize that, based on the ideas of this application, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of this application.

Claims

1. A method for identifying and assessing biological risks in ship ballast water, characterized in that, include: Obtain ship ballast water samples; Hazardous substances were screened and identified based on the ship's ballast water samples. The concentration of the risk substances identified through screening and identification was determined by qPCR. Develop a risk assessment model; Using the aforementioned risk assessment model, risk substance analysis and simulation were performed based on the concentration values ​​of the risk substances to obtain simulation results of ballast water biological risk substances. The ballast water biological risk assessment results are obtained based on the simulation results of the ballast water biological risk substances.

2. The method for identifying and assessing biological risks in ship ballast water according to claim 1, characterized in that, Hazardous substance screening and identification based on the aforementioned ship ballast water samples includes: eDNA was extracted from the ship's ballast water sample; Qualitative analysis of the extracted eDNA was performed using PCR amplification to screen and identify the risk substances.

3. The method for identifying and assessing biological risks in ship ballast water according to claim 2, characterized in that, Extracting eDNA from the ship's ballast water samples includes: The ship's ballast water sample was filtered using a nitrocellulose filter membrane of a set density to obtain filtrate; The eDNA in the filtrate was extracted using magnetic beads.

4. The method for identifying and assessing biological risks in ship ballast water according to claim 1, characterized in that, Construct a risk assessment model, including: Based on the convection-diffusion theory and coupled with the colloidal filtration mechanism, a mathematical model for the migration of bacterial cells and eDNA in the terrestrial environment considering leaching is constructed. The mathematical model is used as the risk assessment model.

5. The method for identifying and assessing biological risks in ship ballast water according to claim 4, characterized in that, The risk assessment model is expressed as follows: ； In the formula, For pollutant concentration, The porosity of the medium. For the density of the medium, This represents the concentration of colloids adsorbed onto the surface of soil particles through reversible adsorption. This represents the concentration of colloids adsorbed onto the surface of soil particles through irreversible adsorption. The direction of pollutant movement Where is the diffusion coefficient. For fluid velocity, For time.

6. The method for identifying and assessing biological risks in ship ballast water according to claim 2, characterized in that, Using the aforementioned risk assessment model, risk substance analysis simulations were performed based on the qPCR quantitative analysis of risk substance concentrations to obtain simulation results of ballast water biological risk substances, including: Based on the concentration values ​​of risk substances obtained through qPCR quantitative analysis after screening and identification, the diffusion patterns and stable effluent concentrations of different eDNAs are simulated using the risk assessment model to obtain the simulation results of the risk substances in ballast water organisms.

7. The method for identifying and assessing biological risks in ship ballast water according to claim 1, characterized in that, Also includes: Risk response measures are generated based on the results of the ballast water biological risk assessment.

8. A computer device, comprising: A memory, a processor, and a computer program stored in the memory and capable of running on the processor, characterized in that the processor executes the computer program to implement the method for identifying and assessing biological risks in ship ballast water as described in any one of claims 1-7.

9. A computer-readable storage medium having a computer program stored thereon, characterized in that, When executed by a processor, the computer program implements the method for identifying and assessing biological risks in ship ballast water as described in any one of claims 1-7.

10. A computer program product, comprising a computer program, characterized in that, When executed by a processor, the computer program implements the method for identifying and assessing biological risks in ship ballast water as described in any one of claims 1-7.

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