CN-122022179-A - Ecological health evaluation method for reclaimed water river based on environment DNA and co-occurrence network

CN122022179ACN 122022179 ACN122022179 ACN 122022179ACN-122022179-A

Abstract

The application relates to the technical field of environmental monitoring and ecological environment protection, and discloses a regenerated water river ecological health evaluation method based on an environment DNA and co-occurrence network, which comprises the steps of setting sampling points and collecting water samples in a monitoring area; extracting environmental DNA, carrying out PCR amplification and high-throughput sequencing, carrying out species annotation on three large biological groups, calculating a water quality comprehensive pollution index, determining a reference point, constructing a co-occurrence network of various group organisms in each point location, calculating network topology parameters, establishing a candidate index library of four dimensions of community structure, diversity, functional groups and network topology parameters for the three large biological groups, calculating an integrity index after optimization of each group and an integrity index of multiple species after optimization, and defining river health grades according to the score of an integrity evaluation system. Therefore, the topology parameters of the ecological co-occurrence networks of various groups are incorporated into the integrity index calculation, and objective weights are determined through training of a machine learning model, so that the structure and the functional state of the river ecological system can be scientifically and comprehensively reflected.

Inventors

ZHAO FEI
CHENG JUNTAO
WANG LIYAN
CHEN TAO
CHEN DONG
ZHAO XIA
Wei Penghao
SHI XUEQING

Assignees

青岛理工大学

Dates

Publication Date: 20260512
Application Date: 20260204

Claims (10)

1. The ecological health evaluation method of the regenerated water river based on the environment DNA and the co-occurrence network is characterized by comprising the following steps of: S1, selecting a sampling area, and distributing sampling points in the sampling area; s2, extracting the environment DNA of the samples collected by the sampling points; S3, carrying out PCR amplification and high-throughput sequencing on the environmental DNA, and carrying out species annotation on three major biotics of phytoplankton, invertebrate and fish based on the operation classification unit information corresponding to the sample; s4, calculating the water pollution index of the sampling area, and determining a reference point and a damaged point; s5, constructing a co-occurrence network of various groups of organisms in each point location, and calculating network topology parameters; S6, establishing a candidate index library with four dimensions of community structure, diversity, functional group and network topology parameters for each biological group, carrying out correlation analysis on candidate indexes of each group and each dimension, eliminating indexes with strong collinearity, carrying out PCA analysis on the screened indexes, and calculating indexes of each dimension of each group; S7, calculating weights of community structure, diversity, functional groups and network topology parameters under the single group level, and calculating an integrity index of each group after optimization by a weighted summation method; And S8, calculating the weight of each biological group, calculating to obtain an optimized multi-species integrity index by a weighted summation method, and defining the river health grade according to the score of the integrity evaluation system.
2. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein in step S5, the calculating network topology parameters includes: Integrating the relative abundance of each OTU within each class group into the overall network dataset; carrying out correlation analysis on all the points, and reserving the corresponding OTU for subsequent network construction when the absolute value of the correlation coefficient is larger than a first preset value and the P value is smaller than a second preset value; and calculating a plurality of network topology parameters including network density, average degree, clustering coefficient, modularity, natural connectivity, network damage resistance index, community condensation degree and average variation degree.
3. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S6 comprises: Selecting the relative abundance of gates, classes, orders, families, genera and species with the abundance of various groups being larger than the preset abundance and corresponding classification units as indexes of community structure dimensions, adopting Shannon, chao1, pielou and Simpson indexes as indexes of diversity dimensions, selecting proper functional group categories for different groups as indexes of functional group dimensions, and adopting Density, MEAN DEGREE, clustering Coefficient, modularity, natural Connectivity, invulnerability, cohesion and AVD as indexes of network topology parameter dimensions; Carrying out correlation analysis on candidate indexes of various groups and dimensions, screening candidate indexes with relative independence larger than an independence threshold value and biological information redundancy smaller than a redundancy threshold value, and screening candidate indexes with relative independence larger than the independence threshold value or biological information redundancy smaller than the redundancy threshold value if the correlation coefficient is larger than or equal to a preset coefficient; And carrying out PCA analysis on the indexes reserved after screening, carrying out Z-score standardization on candidate indexes of each dimension of each group, then carrying out PCA analysis, screening principal components with reserved characteristic values larger than or equal to 1, calculating variance contribution rate weights of the principal components, and obtaining indexes of each dimension of each group through weighted summation of principal component scores.
4. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S7 comprises: under the level of a single group, taking four-dimensional indexes as explanatory variables and taking states of different point positions as dependent variables to perform random forest model training; Calculating feature importance based on a random forest, adopting out-of-band OOB as a test set, constructing a plurality of data sets by utilizing a resampling technology, and respectively carrying out feature importance measurement e kj on each data set, wherein k is the kth resampling; Invoking importance a function to calculate a feature importance measure e kj , and averaging the multiple resampling results to obtain the average importance of the j-th dimension ; Average importance to all dimensions And carrying out normalization processing to obtain the importance weight of the comprehensive evaluation feature, and calculating the integrity indexes of different groups according to the weight.
5. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S8 comprises: taking the integrity indexes of various groups as explanatory variables and the states of different point positions as dependent variables to perform random forest model training; Calculating feature importance based on a random forest, adopting out-of-band OOB as a test set, constructing a plurality of data sets by utilizing a resampling technology, and respectively carrying out feature importance measurement e kj on each data set, wherein k is the kth resampling; Invoking importance a function to calculate a feature importance measure e kj , and averaging the multiple resampling results to obtain the average importance of the j-th class group ; Average importance to all clusters Normalizing to obtain the importance weight of the comprehensive evaluation characteristics, and calculating the optimized multi-species integrity index according to a weighted summation method; Dividing the distribution range of the integrity index to obtain multiple kinds of evaluation grades based on optimized multi-species integrity evaluation, and corresponding the score of each sampling point to each grade to obtain a final evaluation result.
6. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S4 comprises: five water quality factors capable of reflecting the levels of nutrient salts, organic matters and pH value of the reclaimed water moisturizing river channel are selected for comprehensive pollution index calculation; The formula for calculating the single factor pollution index of the unsuitable pH is: ; Wherein, the Is the concentration index of the contaminant; is the measured concentration of the contaminant; is a standard limit for contaminants; the calculation formula of the pH pollution index is as follows: ; ; Wherein, the Pollution index for pH; Is the actual measurement value; is standard to Lower limit value of (2); Is the upper limit value of the standard pH; The calculation formula of the water quality comprehensive pollution index P is as follows: ; Wherein P is the comprehensive pollution index of water quality; P is the arithmetic average value of the single factor pollution indexes of n indexes; and selecting the point with the lowest preset number as the reference point and the rest point as the damaged point according to the calculated water quality comprehensive pollution index result.
7. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S3 comprises: The selected universal primers comprise CITSF-CITSR phytoplankton, 16S1634MF_16S1634MR fish, 16SF1F_16SR1R invertebrate; constructing a library and performing high-throughput sequencing; OTU clustering is carried out on the sequences according to the preset similarity, and the sequences annotated to chloroplasts and mitochondria in all samples are removed; All sample sequences were leveled, OTU species taxonomic annotation was performed using RDP CLASSIFIER alignment NT gene databases, and the community composition of each sample was counted at different species taxonomic levels.
8. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S1 comprises: selecting a region with wide river surface, large water volume and adequate water plant coverage and biomass as the sampling region; And according to the geographical position and hydrologic condition of the regenerated water supply point, distributing and controlling the sampling points along the river according to the principle of uniform distribution.
9. The method for evaluating ecological health of a regenerated water river based on an environmental DNA and co-occurrence network according to claim 1, wherein step S2 comprises: In a preset time period, mixing water samples collected at each point in an equal ratio, and filtering by using a sterile filter membrane; and (3) extracting DNA from the filter membrane of the filtered water sample by using a genome DNA extraction kit, and preserving the extracted DNA at a preset temperature.
10. The method for ecological health assessment of a reclaimed water river based on an environmental DNA and co-occurrence network as recited in any one of claims 1 to 9, further comprising, after step S8: S9, calculating an original multi-species integrity index, wherein a candidate index library of each group comprises three dimensions of a community structure, diversity and functional groups, and carrying out statistical consistency analysis on the original multi-species integrity index and the optimized multi-species integrity index by adopting a multi-level verification mode of correlation analysis, regression analysis and Bland-Altman difference analysis.

Description

Ecological health evaluation method for reclaimed water river based on environment DNA and co-occurrence network Technical Field The application relates to the technical field of environmental monitoring and ecological environment protection, in particular to a regenerated water river ecological health evaluation method based on an environment DNA and co-occurrence network. Background At present, reclaimed water in a sewage treatment plant is used as a second water source and is widely used for river channel water replenishing and the like so as to realize the dual purposes of sewage reclamation and water ecological restoration. Scientific evaluation of influence of the reclaimed water on river channel health is necessary for pertinently improving the quality of the reclaimed water and promoting the safe utilization of the reclaimed water. The biological integrity index is a core method for evaluating the influence of reclaimed water supplementing water on the ecological health of a receiving river by integrating multidimensional information such as community structure, diversity, functional groups and the like. The existing assessment method depends on the traditional morphological identification method, and species counting and abundance statistics are carried out only on single nutrition level (such as phytoplankton or fish), so that information dimension is limited, accuracy of ecological health assessment of the regenerated water moisturizing type river is reduced, and structural integrity and functional stability of a river ecological system cannot be comprehensively reflected. It should be noted that the information disclosed in the above background section is only for enhancing understanding of the background of the application and thus may include information that does not form the prior art that is already known to those of ordinary skill in the art. Disclosure of Invention The following presents a simplified summary in order to provide a basic understanding of some aspects of the disclosed embodiments. This summary is not an extensive overview, and is intended to neither identify key/critical elements nor delineate the scope of such embodiments, but is intended as a prelude to the more detailed description that follows. The embodiment of the disclosure provides a regenerated water river ecological health evaluation method based on an environment DNA and co-occurrence network so as to comprehensively reflect the structural integrity and the functional stability of an ecological system. In some embodiments, the method for evaluating ecological health of a regenerated water river based on environmental DNA and co-occurrence networks comprises the steps of S1, selecting a sampling area, distributing sampling points in the sampling area, S2, conducting environmental DNA extraction on samples collected by the sampling points, S3, conducting PCR amplification and high-throughput sequencing on the environmental DNA, conducting species annotation on three biological groups of phytoplankton, invertebrates and fishes based on operation classification unit information corresponding to the samples, S4, calculating water pollution indexes of the sampling area, determining reference points and damaged points, S5, constructing co-occurrence networks of all biological groups in all the points, calculating network topology parameters, S6, establishing candidate index libraries of four dimensions of community structures, diversity, functional groups and network topology parameters for all biological groups, conducting correlation analysis on candidate indexes of all the groups and all the dimensions, removing the indexes of the co-linearity, conducting PCA analysis on the indexes after screening, calculating indexes of all the dimensions of all the groups, S7, calculating the water pollution indexes of all the groups at the single group level, calculating the structural weights, the diversity weights and the functional groups, calculating the total biological group health index of all the groups and the biological groups by the method after calculating the total health of the systems, and optimizing the total health of all the biological groups by the methods, and calculating the total health index of all the systems after the calculation of all the biological groups. The regenerated water river ecological health evaluation method based on the environment DNA and co-occurrence network provided by the embodiment of the disclosure can realize the following technical effects: The network topology parameters of various groups of ecological co-occurrence networks are incorporated into the calculation of the integrity index, the information dimension is increased, the information quantity of the multi-species integrity index is improved, and the optimized integrity evaluation system can reflect the change of river health more sensitively, comprehensively and objectively. The method fills the gap of the integrity evaluati