Table S1 List of 345 specimens of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 used in this study.

Table S2 Nucleotide sequences of microsatellite primers and fluorescence dyes used for labelling used in this study.

Table S3 Pairwise comparison of linkage disequilibrium of 15 microsatellite loci in bighead catfish (Clarias macrocephalus) collected from SNK18NH (Ban Don Sao Thong, Sakon Nakhon Province) population in 2018. Numbers indicate p-values with 110 permutations.

Table S4 Pairwise comparison of linkage disequilibrium of 15 microsatellite loci in bighead catfish (Clarias macrocephalus) collected from SNK23SDKJ (Susan Don Kajo, Sakon Nakhon Province) population in 2023. Numbers indicate p-values with 110 permutations.

Table S5 Pairwise comparison of linkage disequilibrium of 15 microsatellite loci in bighead catfish (Clarias macrocephalus) collected from SNK23BTW (Baan Tha Wat, Sakon Nakhon Province) population in 2023. Numbers indicate p-values with 110 permutations.

Table S6 Pairwise comparison of linkage disequilibrium of 15 microsatellite loci in bighead catfish (Clarias macrocephalus) collected from SNK22NH (The hatchery of Sakon Nakhon Provincial Fisheries Office, the Department of Fisheries, Ministry of Agriculture and Cooperatives, Sakon Nakhon Province) population in 2022. Numbers indicate p-values with 110 permutations.

Table S7 Genetic diversity of each microsatellite locus in four populations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023. Detailed information of all individuals is presented in Table S1.

Table S8 Observed (H_o) and expected (H_e) heterozygosity in four populations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 estimated by the Welch’s t-test using 15 microsatellite loci.

Table S9 Comparison of observed (H_o) and expected (H_e) heterozygosity between four populations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 based on the genotyping data of 15 microsatellite loci.

Table S10 Distributions of genetic relatedness values (r) and inbreeding coefficients (F_IS) in bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023.

Table S11 Analysis of molecular variance for bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023based on the results of AMOVA analysis with15 microsatellite loci.

Table S12 Nei’s genetic distance (D) between four populations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023.

Table S13 Wilcoxon sign rank test to evaluate the mutation–drift equilibrium in 345 bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 under different models (TPM and SMM).

Table S14 The mean migration rates between populations and 95% confidence intervals for standard deviation determined via BayesAss version 3.0.5 using microsatellite data for bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023.

Table S15 The median scaled population size (Θ) in four populations of bighead catfish and asymmetric migration rates (M) between bighead catfish populations estimated with 15 microsatellite loci.

Table S16 The effective number of immigrants (N_m) from population i into population j per generation between four populations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 estimated with 15 microsatellite loci.

Table S17 Genetic diversity of tentative subpopulation of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 estimated using 15 microsatellite loci.

Table S18 Observed (H_o) and expected (H_e) heterozygosity in six tentative subpopulations of bighead catfish (Clarias macrocephalus) from Sakon Nakhon Province, Thailand, in 2018, 2022, and 2023 estimated by Welch’s t-test using 15 microsatellite loci.

Table S19 Comparison of observed (H_o) and expected (H_e) heterozygosity between six tentative subpopulations of bighead catfish (Clarias macrocephalus) based on genotyping data of 15 microsatellite loci.

Table S20 Pairwise comparison of genetic differentiation (F_ST) between tentative subpopulations of bighead catfish (Clarias macrocephalus).

Table S21 The result of analysis of molecular variance (AMOVA) for tentative subpopulations of bighead catfish (Clarias macrocephalus) based on 15 microsatellite loci.

Table S22 The effective population size and the ratio of effective population size to census population size (N_e/N) in tentative subpopulations of bighead catfish (Clarias macrocephalus).

Table S23 The effective number of immigrants (N_m) from population i into population j per generation in tentative subpopulation of bighead catfish (Clarias macrocephalus) estimated with 15 microsatellite loci.

Table S24 Changes of area in water and flooded vegetation at Nong Han Lake, Sakon Nakhon Province, Thailand from 2017 to 2023.

Table S25 The number and origin of bighead catfish (Clarias macrocephalus) released in Nong Han Lake, Sakon Nakhon over years.

Figure S1 Distribution of (a) inbreeding coefficient (F_IS) and relatedness (r) among population of bighead catfish (Clarias macrocephalus).

Figure S2 Discriminant analysis of principal components (DAPC) of bighead catfish (Clarias macrocephalus) populations based on genotyping data of 15 microsatellite loci. Each population is represented with a different color, and each point represents an individual.

Figure S3 Factorial correspondence analysis (FCA) of bighead catfish (Clarias macrocephalus) populations based on genotyping data of 15 microsatellite loci. Each population is represented with a different color, and each point represents an individual.

Figure S4 Principal component analysis (PCoA) of bighead catfish (Clarias macrocephalus) populations based on genotyping data of 15 microsatellite loci. Each population is represented with a different color, and each point represents an individual.

Figure S5 Multidimensional scaling analysis (MDS) of bighead catfish (Clarias macrocephalus) populations based on genotyping data of 15 microsatellite loci. Each population is represented with a different color, and each point represents an individual.

Figure S6 Different patterns of population structures of bighead catfish (Clarias macrocephalus) populations based on genotyping data of 15 microsatellite loci generated by the model-based Bayesian clustering algorithms implemented in STRUCTURE. (a) Plot based on ln P(K). (b) Plot based on Evano’s ΔK.

Figure S7 Estimation of the optimal number of migration edges in bighead catfish (Clarias macrocephalus) using the R package OptM with the default Evanno method. (a) value of variance explained, (b) the number of migration edges.

Figure S8 Plot of correlation between genetic distance and geographical distance among bighead catfish (Clarias macrocephalus) populations.

Figure S9 Genetic structures of six tentative subpopulation of bighead catfish (Clarias macrocephalus) revealed by Bayesian structural analysis. The x-axis represents the proportion of membership in each genetic cluster, and each vertical bar on the y-axis represents an individual. All individuals from six subpopulations are superimposed on the plot. Black horizontal lines indicate the boundaries.

Figure S10 Principal component analysis (PCoA) of bighead catfish (Clarias macrocephalus) tentative subpopulations based on the genotyping data of 15 microsatellite loci. Each subpopulation is represented with a different color, and each point represents an individual.

Figure S11 Discriminant analysis of principal components (DAPC) of bighead catfish (Clarias macrocephalus) tentative subpopulations based on genotyping data of 15 microsatellite loci. Each subpopulation is represented with a different color, and each point represents an individual.

Figure S12 Multidimensional scaling analysis (MDS) of bighead catfish (Clarias macrocephalus) tentative subpopulations based on genotyping data of 15 microsatellite loci. Each subpopulation is represented with a different color, and each point represents an individual.

Figure S13 Factorial correspondence analysis (FCA) of bighead catfish (Clarias macrocephalus) tentative subpopulations based on genotyping data of 15 microsatellite loci. Each subpopulation is represented with a different color, and each point represents an individual.