Objective To detect the mutations in the knockdown resistance (kdr) gene of the Aedes albopictus populations resistant and susceptible to permethrin, deltamethrin, and α-cypermethrin, and to elucidate the association between the resistance phenotypes and the kdr gene mutations. Methods From June to September, 2016, the Ae. albopictus populations in Ruili, Yunnan province, China, resistant to permethrin, deltamethrin, and α-cypermethrin and the Ae. albopictus populations, and the susceptible lab population of Jiangsu Ae. albopictus strain were collected, respectively, and their resistance phenotypes were detected by a bioassay. Individual genomic DNA was extracted. Three pairs of primers were sgnthetized for PCR amplification of partial gene fragments of voltage-gated sodium channel on the nerve cell membrane to detect the kdr gene mutations. The genotypes and their frequencies of the kdr gene of the resistant and susceptible populations were statistically analyzed. The chi-square test was used to analyze the association between the kdr gene mutations and the resistance phenotypes. Results A total of 500 kdr gene fragments were obtained, including 440 kdr gene fragments of Ae. albopictus resistant populations in Ruili and 60 kdr gene fragments of Ae. albopictus susceptible populations of Jiangsu strain. There was no mutation detected in the gene fragments from the susceptible populations. In Ae. albopictus resistant populations in Ruili, mutations were detected at the 1532, 1534, and 1763 sites. Both F1534S and I1532T mutations were detected in one sample. There were two alleles at the 1532 site, i.e., wild-type ATC/I (isoleucine) and mutant ACC/T (threonine), and their frequencies were 99.32% (292/294) and 0.68% (2/294), respectively; the frequencies of three genotypes-wild-type homozygous I/I, wild/mutant heterozygous I/T, and mutant homozygous T/T were 98.64% (145/147), 1.36% (2/147), and 0 (0/147), respectively. There were five alleles at the 1 534 site, i.e., wild-type TTC/F (phenylalanine), mutant TCC/S (serine), TCG/S (serine), TTG/L (leucine), and TGC/C (cysteine), and their frequencies were 59.53% (175/294), 29.93% (88/294), 0.68% (2/294), 2.72% (8/294), and 7.14% (21/294), respectively; the frequencies of six genotypes-wild-type homozygous F/F, wild-type/mutant heterozygous F/C, F/S, and F/L, mutant homozygous S/S, and mutant heterozygous C/S were 40.14% (59/147), 6.80% (10/147), 26.53% (39/147), 5.44% (8/147), 13.61% (20/147), and 7.48% (11/147), respectively. There were two alleles at the 1 763 site, i.e., wild-type GAC/D (aspartic acid) and mutant TAC/Y (tyrosine), their frequencies were 99.32% (292/294) and 0.68% (2/294), respectively; the frequencies of three genotypes-wild-type homozygous D/D, wild-type/mutant heterozygous D/Y, and mutant homozygous Y/Y were 98.64% (145/147), 1.36% (2/147), and 0 (0/147), respectively. Conclusion In the kdr gene of Ae. albopictus population resistant to pyrethroid insecticides in Ruili, mutations are detected at the 1532, 1534, and 1763 sites and no mutations are detected at the 989 and 1 014 sites. The mutations at the 1534 site are dominant and the mutations at the 1763 site are first discovered. Single mutation is dominant and only one sample contains both the F1534S and I1532T mutations. This study confirms that the kdr mechanism is one of the resistance mechanisms of Ae. albopictus populations against pyrethroid insecticides in Ruili.